Bacteriophage Ecology Group Members' Publications
	Dedicated to the ecology and evolutionary biology of the parasites of unicellular organisms (UOPs)
	© Stephen T. Abedon
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© Phage et al.	last updated on Wednesday, December 26, 2001

1. Bacteriophage therapy rescues mice bacteremic from a clinical isolate of vancomycin-resistant Enterococcus faecium. Biswas, B., Adhya, S., Washart, P., Paul, B., Trostel, A.N., Powell, B., Carlton, R., Merril, C.R. (2002). Infect. Immun. 70:204-210. Colonization of the gastrointestinal tract with vancomycin-resistant Enterococcus faecium (VRE) has become endemic in many hospitals and nursing homes in the United States. Such colonization predisposes the individual to VRE bacteremia and/or endocarditis, and immunocompromised patients are at particular risk for these conditions. The emergence of antibiotic-resistant bacterial strains requires the exploration of alternative antibacterial therapies, which led our group to study the ability of bacterial viruses (bacteriophages, or phages) to rescue mice with VRE bacteremia. The phage strain used in this study has lytic activity against a wide range of clinical isolates of VRE. One of these VRE strains was used to induce bacteremia in mice by intraperitoneal (i.p.) injection of 10 9 CFU. The resulting bacteremia was fatal within 48 h. A single i.p. injection of 3 _ 10 8 PFU of the phage strain, administered 45 min after the bacterial challenge, was sufficient to rescue 100% of the animals. Even when treatment was delayed to the point where all animals were moribund, approximately 50% of them were rescued by a single injection of this phage preparation. The ability of this phage to rescue bacteremic mice was demonstrated to be due to the functional capabilities of the phage and not to a nonspecific immune effect. The rescue of bacteremic mice could be effected only by phage strains able to grow in vitro on the bacterial host used to infect the animals, and when such strains are heat inactivated they lose their ability to rescue the infected mice. [TOP OF PAGE]

2. Bacteriophage latent-period evolution as a response to resource availability. Abedon, S.T., Herschler, T.D., Stopar, D. (2001). Appl. Environ. Microbiol. 67:4233-4241. Bacteriophages (phages) modify microbial communities by lysing hosts, transferring genetic material, and effecting lysogenic conversion. To understand how natural communities are affected it is important to develop predictive models. Here we consider how variation between models in eclipse period, latent period, adsorption constant, burst size, the handling of differences in host quantity and host quality, and in modeling strategy can affect predictions. First we compare two published models of phage growth, which differ primarily in terms of how they model the kinetics of phage adsorption; one is a computer simulation and the other is an explicit calculation. At higher host quantities (~10⁸ cells/ml), both models closely predict experimentally determined phage population growth rates. At lower host quantities (10⁷ cells/ml), the computer simulation continues to closely predict phage growth rates, but the explicit model does not. Next we concentrate on predictions of latent-period optima. A latent-period optimum is the latent period that maximizes the population growth of a specific phage growing in the presence of a specific quantity and quality of host cells. Both models predict similar latent-period optima at higher host densities (e.g., 17 min at 10⁸ cells/ml). At lower host densities, however, the computer simulation predicts latent-period optima that are much shorter than those suggested by explicit calculations (e.g., 90 versus 1,250 min at 10⁵ cells/ml). Finally, we consider the impact of host quality on phage latent-period evolution. By taking care to differentiate latent-period phenotypic plasticity from latent-period evolution, we argue that the impact of host quality on phage latent-period evolution may be relatively small. [TOP OF PAGE]

3. Le matin des bactériophages. Ackermann, H.-W. (2001). Virologie 5:35-43. With about 5 150 electron microscopic observations, phages contstitute the larges of all viral groups. The International Committee on Taxonomy of Viruses (ICTV) persently recognized one order, 13 families, and 30 genera. The order Caudovirales includes three families of tailed phages and about 5 000 members (96.4%). The 10 families of icosahedral, filamentous, or pleomorphic phages totalize 186 viruses. Phages are found in all the bacterial world and, with up to 10¹⁰ particles/ml in seawater, seem to be the most frequent microbes of Earth. Phages are polyphyletic in origin. Tailed phages appear to be the msot ancient viruses and recombination with exchange of genes or gene blocs (modular evolution) seems to be their preferred way of evolution. Tailed phages and herpesviruses present multiple analogies. Harmful phages can create havoc in bacterial fermentations, especially in the dairy industry. By contrast, phages are very useful in general bacteriology, therapy of infectious diseases is making a come-back and phages are likely to have a brilliant future in research. [TOP OF PAGE]

4. Frequency of morphological phage descriptions in the year 2000. Brief Review. Ackermann, H.-W. (2001). Archives of Virology 146:843-857. Over 5100 bacteria viruses have been examined in the electron microscope since 1959. About 4950 phages (96%) are tailed and only 186 phages (3.6%), are cubic, filamentous, or pleomorphic. Phages belong to 13 virus families and occur in over 140 bacterial genera. Phages are listed by morphotypes and host genera. Siphoviridae or phages with long, noncontractile tails compromise 61% of tailed phages. The distribution of phages in different bacterial phylogenetic divisions is shown. [TOP OF PAGE]

5. The Vibrio cholerae VPI?/CTX?/TCP: Interactions of PHAGE-PHAGE-bacterium. Ai, Y.-C., Meng, F. (2001). Acta Microbiologica Sinica 41:[TOP OF PAGE]

6. The genome of the archael virus SIRV1 has features in common with genomes of eukaryal viruses. Blum, H., Zillig, W., Mallock, S., Domdey, H., Prangishvili, D. (2001). Virology 281:6-9. The virus SIRV1 of the extremely thermophilic archaeon Sulfolobus has a double-stranded DNA genome similar in architecture to the genomes of eukaryal viruses of the families Poxviridae, Pycodnaviridae, and Asfarviridae: the two strands of the 32,301 bp long linear genome are covalently connected forming a continuous polynucleotide chain and 2029 kb long inverted repeats are present at the termini. Very likely it also shares with these viruses mechanisms of initiation of replication and resolution of replicative intermediates. [TOP OF PAGE]

7. Phages of Lactococcus lactis: an ecological and economical equilibrium. Boucher, I., Moineau, S. (2001). Recent Research Developments in Virology 3:243-256. Lactic acid bacteria (LAB) are a group of organisms widely used in food fermentation. Interests in these microorganisms have increased sharply in the last decade; these organisms have even been dubbed the bugs of the new millennium. One distinctive fact about LAB-fermented foods is that they are produced in non-sterile conditions. Thus, LAB are susceptible to infection by lytic bacteriophages naturally present in these environments. Recent developments (by our group and others) in the field of bacteriophages of Lactococcus, the most studied LAB, are investigated and presented in the review. [TOP OF PAGE]

8. Comparative phage genomics and the evolution of Siphoviridae: Insights from dairy phages. Brussow, H., Desiere, F. (2001). Molecular Microbiology 39:213-222. Comparative phage genomics can retrace part of the evolutionary history of phage modules encoding phage-specific functions such as capsid building or establishment of the lysogenic state. The diagnosis of relatedness is not based exclusively on sequence similarity, but includes topological considerations of genome organization. The gene maps from the lambda-, psiM2-, L5-, Sfi21-, Sfi11-, phiC31-, sk1- and TM4-like phages showed a remarkable synteny of their structural genes defining a lambda supergroup within Siphoviridae (Caudovirales with long non-contractile tails). A hierarchy of relatedness within the lambda supergroup suggested elements of vertical evolution in the capsid module of Siphoviridae. Links to P22-like Podoviridae and P2-like Myoviridae were also detected. Numerous cases of horizontal gene transfer were observed, but recent transfers were limited to interbreeding phage populations. We suggest that tailed phages are the result of both vertical and horizontal evolution and are thus a good model system for web-like phylogenies. [TOP OF PAGE]

9. Phages and their application against drug-resistant bacteria. Chanishvili, N., Chanishvili, T., Tediashvili, M., Barrow, P.A. (2001). Journal of Chemical Technology and Biotechnology. 76:689-699. At the beginning of the 20th century the phenomenon of spontaneous bacterial lysis was discovered independently by Twort and d'Herelle. Despite the suggestion at that time by d'Herelle that these agents might be applied to the control of bacterial diseases in the west this idea was explored in a desultory fashion only and was eventually discarded largely due to the advent of extensive antibiotic usage. However, interest was maintained in countries of the former Soviet Union where bacteriophage therapy has been applied extensively since that time. Central to this work was the Eliava Institute of Bacteriophage, Microbiology and Virology in Tbilisi, Georgia, which was founded in 1923 through the joint efforts of d'Herelle and the Georgian George Eliava. Ironically, given his contributions to public health in the Soviet Union, Eliava was branded as an enemy of the people in 1937 and executed. d'Herelle never again returned to Georgia. In spite of these tragic events this institute remained the focus for phage therapy in the world and despite being continuously active in this field for 75 years, now struggles for its financial life. In the Eliava Institute, phages were sought for bacterial pathogens implicated in disease outbreaks in different parts of the Soviet Union and were dispatched for use in hospitals throughout the country. Although infections caused by a wide variety of bacterial pathogens have been treated, much of this has been published in Russian and is not readily available in the west. Work has also been carried out in Poland over many years and this has only recently been published in English. By contrast, interest in the west has been limited to a small number of enthusiasts and academics and until very recently little interest has been shown. The main reason that the medical and scientific communities are now beginning to take notice, is the continuing world-wide rise in the incidence of multiply-antibiotic-resistant bacterial pathogens and the absence of effective means for their control. Recent publicity over the work of the Eliava Institute has concentrated the minds of the western world on the potential for infectious disease control that bacteriophage offer, a procedure that is biologically more acceptable than antibiotic use and which has been in use for several decades already. [TOP OF PAGE]

10. A conserved genetic module that encodes the major virion components in both the coliphage T4 and the marine cyanophage S-PM2. Hambly, E., Tétart, F., Desplats, C., Wilson, H., Krisch, H.M., Mann, N.H. (2001). Proc. Natl. Acad. Sci. USA 98:11411-11416. Sequence analysis of a 10-kb region of the genome of the marine cyanomyovirus S-PM2 reveals a homology to coliphage T4 that extends as a contiguous block from gene (g)18 to g23. The order of the S-PM2 genes in this region is similar to that of T4, but there are insertions and deletions of small ORFs of unknown function. In T4, g18 codes for the tail sheath, g19, the tail tube, g20, the head portal protein, g21, the prohead core protein, g22, a scaffolding protein, and g23, the major capsid protein. Thus, the entire module that determines the structural components of the phage head and contractile tail is conserved between T4 and this cyanophage. The significant differences in the morphology of these phages must reflect the considerable divergence of the amino acid sequence of their homologous virion proteins, which uniformly exceeds 50%. We suggest that their enormous diversity in the sea could be a result of genetic shuffling between disparate phages mediated by such commonly shared modules. These conserved sequences could facilitate genetic exchange by providing partially homologous substrates for recombination between otherwise divergent phage genomes. Such a mechanism would thus expand the pool of phage genes accessible by recombination to all those phages that share common modules. [TOP OF PAGE]

11. A novel virus (HaNIV) causes lysis of the toxic bloom-forming alga Heterosigma akashiwo (Raphidophyceae). Lawrence, J.E., Chan, A.M., Suttle, C.A. (2001). Journal of Phycology 37:216-222. We describe a previously unknown virus that causes lysis of the toxic blopm-forming alga Heterosigma akashiwo (Hada) Hara et Chihara (Raphidophyceae). Heterosigma akashiwo nuclear inclusion virus (HaNIV) does not resemble other algal viruses described to date. HaNIV is small (ca. 30 nm diameter), is assembled in the nucleus, and forms crystalline arrays. We estimate that approximately 10⁵ HaNIV particles are released during lysis of a cell. During a time-course experiment, TEM revealed the first signs of HaNIV infection 24 h after viral addition, and by 74 h 98% of observed cells were visibly infected. The onset of cell lysis, as indicated by a decrease in the relative fluorescence of the cultures, was apparent by 42 h postinfection. The heterochromatin of infected cells is frequently found at the margin of the nucleoplasm, which is consistent with virus-mediated programmed cell death, or apoptosis. HaNIV is clearly different from other described viruses that infect alg ae, including other viral pathogens of H, akashiwo. These results indicate that viruses other than Phycodnaviridae are pathogens and cause mortality of microalgae in marine systems. It is Likely that HaNIV plays an integral role in the population dynamics of H. akashiwo. [TOP OF PAGE]

12. Viruses in the plankton of freshwater and saline Antarctic lakes. Laybourn-Parry, J., Hofer, J.S., Sommaruga, R. (2001). Freshwater Biology 46:1279-1287. 1. Virus-like particle (VLP) abundances in nine freshwater to saline lakes in the Vestfold Hills, Eastern Antarctica (68degree S) were determined in December 1999. In the ultra-oligotrophic to oligotrophic freshwater lakes, VLP abundances ranged from 1.01 to 3.28 X 106 mL-1 in the top 6 m of the water column. In the saline lakes the range was between 6.76 and 36.5 X 10⁶ mL-1. The lowest value was found in meromictic Ace Lake and the highest value in hypersaline Lake Williams. Virus to bacteria ratios (VBR) were lowest in the freshwater lakes and highest in the saline lakes, with a maximum of 23.4 in the former and 50.3 in the latter. 2. A range of morphologies among VLP was observed, including phages with short (Podoviridae) and long tails, icosahedric viruses of up to 300 nm and star-like particles of about 80 nm diameter. 3. In these microbially dominated ecosystems there was no correlation between VLP and either bacterial numbers of chlorophyll a. There was a significant correlation between VLP abundances and dissolved organic carbon concentration (r = 0.845, P < 0.01). 4. The data suggested that viruses probably attack a spectrum of bacteria and protozoan species. Virus-like particle numbers in the freshwater lakes were lower than values reported for lower latitude systems. Those in the saline lakes were comparable with abundances reported from other Antarctic lakes, and were higher than most values published for lower latitude lakes and many marine systems. Across the salinity spectrum from freshwater through brackish to hypersaline, VLP concentrations increased roughly in relation to increasing trophy. 5. Given that Antarctic lakes have a plankton almost entirely made up of bacteria and protists, and that VLP abundances are high, it is likely that viruses play a pivotal role in carbon cycling in these extreme ecosystems. [TOP OF PAGE]

13. Prevention and elimination of upper respiratory colonization of mice by group A streptococci by using a bacteriophage lytic enzyme. Nelson, D., Loomis, L., Fischetti, V.A. (2001). Proc. Natl. Acad. Sci. USA 98:4107-4112. Bacteriophage lytic enzymes quickly destroy the cell wall of the host bacterium to release progeny phage. Because such lytic enzymes specifically kill the species in which they were produced, they may represent an effective way to control pathogenic bacteria without disturbing normal microflora. In this report, we studied a murein hydrolase from the streptococcal bacteriophage C1 termed lysin. This enzyme is specific for groups A, C, and E streptococci, with little or no activity toward several oral streptococci or other commensal organisms tested. Using purified lysin in vitro, we show that 1,000 units (10 ng) of enzyme is sufficient to sterilize a culture of apprxeq10⁷ group A streptococci within 5 seconds. When a single dose of lysin (250 units) is first added to the oral cavity of mice, followed by 10⁷ live group A streptococci, it provides protection from colonization (28.5% infected, n = 21) compared with controls without lysin (70.5% infected, n = 17) (P < 0.03). Furthermore, when lysin (500 units) was given orally to 9 heavily colonized mice, no detectable streptococci were observed 2 h after lysin treatment. In all, these studies show that lysin represents a unique murein hydrolase that has a rapid lethal effect both in vitro and in vivo on group A streptococci, without affecting other indigenous microorganisms analyzed. This general approach may be used to either eliminate or reduce streptococci from the upper respiratory mucosal epithelium of either carriers or infected individuals, thus reducing associated disease. [TOP OF PAGE]

14. Three-component-mediated serotype conversion in Pseudomonas aeruginosa by bacteriophage D3. Newton, G.J., Daniels, C., Burrows, L.L., Kropinski, A.M., Clarke, A.J., Lam, J.S. (2001). Molecular Microbiology 39:1237-1247. Bacteriophage D3 is capable of lysogenizing Pseudomonas aeruginosa PAO1 (serotype O5), converting the O-antigen from O5 to O16 and O-acetylating the N-acetylfucosamine moiety. To investigate the mechanism of lysogenic conversion, a 3.6 kb fragment from the D3 genome was isolated capable of mediating serotypic conversion identical to the D3 lysogen strain (AK1380). The PAO1 transformants containing this 3.6 kb of D3 DNA exhibited identical lipopolysaccharide (LPS) banding patterns to serotype O16 in silver-stained SDS-PAGE gels and displayed reactivity to an antibody specific for O-acetyl groups. Further analysis led to the identification of three open reading frames (ORFs) required for serotype conversion: an alpha-polymerase inhibitor (iap); an O-acetylase (oac); and a beta-polymerase (wzybeta). The alpha-polymerase inhibitor (lap) is capable of inhibiting the assembly of the serotype-specific O5 B-band LPS and allows the phage-encoded beta-polymerase (Wzybeta) to form new beta-linked B-band LPS. The D3 phage also alters the LPS by the addition of O-acetyl groups to the FucNAc residue in the O-antigen repeat unit by the action of the D3 O-acetylase (Oac). These three components form a simple yet elegant system by which bacteriophage D3 is capable of altering the surface of P. aeruginosa PAO1. [TOP OF PAGE]

15. Naturally occurring lactococcal plasmid pAH90 links bacteriophage resistance and mobility functions to a food-grade selectable marker. O' Sullivan, D., Ross, R.P., Twomey, D.P., Fitzgerald, G.F., Hill, C., Coffey, A. (2001). Appl. Environ. Microbiol. 67:929-937. The bacteriophage resistance plasmid pAH90 (26,490 bp) is a natural cointegrate plasmid formed via homologous recombination between the type I restriction-modification specificity determinants (hsdS) of two smaller lactococcal plasmids, pAH33 (6,159 bp) and pAH82 (20,331 bp), giving rise to a bacteriophage-insensitive mutant following phage challenge (D. O'Sullivan, D. P. Twomey, A. Coffey, C. Hill, G. F. Fitzgerald, and R. P. Ross, Mol. Microbiol. 36:866-876; 2000). In this communication we provide evidence that the recombination event is favored by phage infection. The entire nucleotide sequence of plasmid pAH90 was determined and found to contain 24 open reading frames (ORFs) responsible for phenotypes which include restriction-modification, phage adsorption inhibition, plasmid replication, cadmium resistance, cobalt transport, and conjugative mobilization. The cadmium resistance property, encoded by the cadA gene, which has an associated regulatory gene (cadC), is of particular interest, as it facilitated the selection of pAH90 in other phage-sensitive lactococci after electroporation. In addition, we report the identification of a group II self-splicing intron bounded by two exons which have the capacity to encode a relaxase implicated in conjugation in gram-positive bacteria. The functionality of this intron was evident by demonstrating splicing in vivo. Given that pAH90 encodes potent phage defense systems which act at different stages in the phage lytic cycle, the linkage of these with a food-grade selectable marker on a replicon that can be mobilized among lactococci has significant potential for natural strain improvement for industrial dairy fermentations which are susceptible to phage inhibition. [TOP OF PAGE]

16. Viruses of the extremely therophilic archaeon Sulfolobus. Prangishvili, D., Stedman, K., Zillig, W. (2001). Trends Microbiol. 9:39-42. Viruses of Sulfolobus are highly unusual in their morphology, and genome structure and sequence. Certain characteristics of the replication strategies of these viruses and the virus-host interactions suggest relationships with eukaryal and bacterial viruses. Moreover, studying these viruses led to the discovery of archaeal promoters and has provided tools for the development of the molecular genetics of these organisms. The Sulfolobus viruses contain unique regulatory features and structures that undoubtedly hold surprises for researchers in the future. [TOP OF PAGE]

17. Isolation and characterization of five Erwinia amylovora bacteriophages and assessment of phage resistance in strains of Erwinia amylovora. Schnabel, E.L., Jones, A.L. (2001). Appl. Environ. Microbiol. 67:59-64. Phages able to infect the fire blight pathogen Erwinia amylovora were isolated from apple, pear, and raspberry tissues and from soil samples collected at sites displaying fire blight symptoms. Among a collection of 50 phage isolates, 5 distinct phages, including relatives of the previously described phages variant phiEa1 and variant phiEa7 and 3 novel phages named variant phiEa100, variant phiEa125, and variant phiEa116C, were identified based on differences in genome size and restriction fragment pattern. variant phiEa1, the phage distributed most widely, had an approximately 46-kb genome which exhibited some restriction site variability between isolates. Phages variant phiEa100, variant phiEa7, and variant phiEa125 each had genomes of approximately 35 kb and could be distinguished by their EcoRI restriction fragment patterns. variant phiEa116C contained an approximately 75-kb genome. variant phiEa1, variant phiEa7, variant phiEa100, variant phiEa125, and variant phiEa116C were able to infect 39, 36, 16, 20, and 40, respectively, of 40 E. amylovora strains isolated from apple orchards in Michigan and 8, 12, 10, 10, and 12, respectively, of 12 E. amylovora strains isolated from raspberry fields (Rubus spp.) in Michigan. Only 22 of 52 strains were sensitive to all five phages, and 23 strains exhibited resistance to more than one phage. variant phiEa116C was more effective than the other phages at lysing E. amylovora strain Ea110 in liquid culture, reducing the final titer of Ea110 by >95% when added at a ratio of 1 PFU per 10 CFU and by 58 to 90% at 1 PFU per 105 CFU. [TOP OF PAGE]

18. Bacteriophage K1-5 encodes two different tail fiber proteins, allowing it to infect and replicate on both K1 and K5 strains of Escherichia coli. Scholl, D., Rogers, S., Adhya, S., Merril, C.R. (2001). J. Virol. 75:2509-2515. A virulent double-stranded DNA bacteriophage, PHIK1-5, has been isolated and found to be capable of infecting Escherichia coli strains that possess either the K1 or the K5 polysaccharide capsule. Electron micrographs show that the virion consists of a small icosohedral head with short tail spikes, similar to members of the Podoviridae family. DNA sequence analysis of the region encoding the tail fiber protein showed two open reading frames encoding previously characterized hydrolytic phage tail fiber proteins. The first is the K5 lyase protein gene of PHIK5, which allows this phage to specifically infect K5 E. coli strains. A second open reading frame encodes a protein almost identical in amino acid sequence to the N-acetylneuraminidase (endosialidase) protein of PHIK1E, which allows this phage to specifically infect K1 strains of E. coli. We provide experimental evidence that mature phage particles contain both tail fiber proteins, and mutational analysis indicates that each protein can be independently inactivated. A comparison of the tail gene regions of PHIK5, PHIK1E, and PHIK1-5 shows that the genes are arranged in a modular or cassette configuration and suggests that this family of phages can broaden host range by horizontal gene transfer. [TOP OF PAGE]

19. Changes in bacterial community composition and dynamics and viral mortality rates associated with enhanced flagellate grazing in a mesoeutrophic reservoir. Simek, K., Weinbauer, M.G., Hornak, K., Dolan, J.R., Nedoma, J., Masin, M., Amann, R. (2001). Appl. Environ. Microbiol. 67:2723-2733. Bacterioplankton from a meso-eutrophic dam reservoir was size fractionated to reduce (<0.8-mum treatment) or enhance (<5-mum treatment) protistan grazing and then incubated in situ for 96 h in dialysis bags. Time course samples were taken from the bags and the reservoir to estimate bacterial abundance, mean cell volume, production, protistan grazing, viral abundance, and frequency of visibly infected cells. Shifts in bacterial community composition (BCC) were examined by denaturing gradient gel electrophoresis (DGGE), cloning and sequencing of 16S rDNA genes from the different treatments, and fluorescence in situ hybridization (PISH) with previously employed and newly designed oligonucleotide probes, Changes in bacterioplankton characteristics were clearly linked to changes in mortality rates. In the reservoir, where bacterial production about equaled protist grazing and viral mortality, community characteristics were nearly invariant, In the "grazer-free" (0.8-mum-filtered) treatment, subject only to a relatively low mortality rate (similar to 17% day(-1)) from viral lysis, bacteria increased markedly in concentration. While the mean bacterial cell volume was invariant, DGGE indicated a shift in BCC and FISH revealed an increase in the proportion of one lineage within the beta proteobacteria, In the grazing-enhanced treatment (5-mum filtrate), grazing mortality was similar to 200% and viral lysis resulted in mortality of 30% of daily production. Cell concentrations declined, and grazing-resistant flocs and filaments eventually dominated the biomass, together accounting for > 80% of the total bacteria by the end of the experiment. Once again, BCC changed strongly and a significant fraction of the large filaments was detected using a FISH probe targeted to members of the Flectobacillus lineage, Shifts of BCC were also reflected in DGGE patterns and in the increases in the relative importance of both beta proteobacteria and members of the Cytophaga-Flavobacterium cluster, which consistently formed different parts of the bacterial flocs. Viral concentrations and frequencies of infected cells were highly significantly correlated with grazing rates, suggesting that protistan grazing may stimulate viral activity. [TOP OF PAGE]

20. Application of Streptococcus thermophilus DPC1842 as an adjunct to counteract bacteriophage disruption in a predominantly lactococcal Cheddar cheese starter: use in bulk starter culture systems. Stokes, D., Ross, R.P., Fitzgerald, G.F., Coffey, A. (2001). Lait 81:327-334. A significant amount of Cheddar cheese manufactured world-wide relies on bulk starter cultures instead of direct vat set (DVS) cultures. While the inclusion of S. thermophilus is sometimes used to counteract failure due to lactococcal phage in the latter system, it is considered difficult to implement in bulk starter systems and is normally avoided. This stems from the problem in controlling the ratio of S. thermophilus to lactococci during the bulk starter preparation such that suitable acidification rates can be achieved. The current study demonstrates how S. thermophilus numbers can be controlled during growth in the bulk starter medium prior to inoculation of a culture, based on three lactococcal strains and S. thermophilus DPC1842, into the cheese vat. The concentration of inorganic phosphate necessary to inhibit the growth of strain DPC1842 in a whey-based bulk starter medium was found to be 0.18 mol.L^-1. Since higher levels of phosphate exist in different commercial bulk starter media used for mesophilic cultures, a number of these media can be used for propagation of this starter blend without domination of strain DPC1842 over its lactococcal counterparts. Strain DPC1842 is highly phage resistant and is particularly acid-fast in the cheese milk and reduces the pH efficiently at very low inocula. Cheddar cheese was manufactured in a commercial plant with this system and the resulting cheese had good flavour characteristics. This study demonstrates the effectiveness of the S. thermophilus DPC1842 component in rescuing the fermentation in the event of severe lactococcal phage attack. [TOP OF PAGE]

21. Community Structure: Viruses. Suttle, C.A. (2001). p. ???-??? In Hurst, C.J., Knudson, G.R., McInerney, M.J., Stezenbach, L.D., and Walter, M.V. (eds.), Manual of Environmental Microbiology (2nd Edition). ASM Press, Washington, DC. [TOP OF PAGE]

22. Phylogeny of the major head and tail genes of the wide-ranging T4-type bacteriophages. Tétart, F., Desplats, C., Kutateladze, M., Monod, C., Ackermann, H.-W., Krisch, H.M. (2001). J. Bacteriol. 183:358-366. We examined a number of bacteriophages with M-type morphology that propagate in different genera of enterobacteria, Aeromonas, Burkholderia, and Vibrio. Most of these phages had a prolate icosahedral head, a contractile tail, and a genome size that was similar to that of T4. A few of them had more elongated heads and larger genomes. All these phages are phylogenetically related, since they each had sequences homologous to the capsid gene (gene 23), tail sheath gene (gene 18), and tail tube gene (gene 19) of T4. On the basis of the sequence comparison of their virion genes, the T4-type phages can be classified into three subgroups with increasing divergence from T4: the T-evens, pseudoT-evens, and schizoT-evens. In general, the phages that infect closely related host species have virion genes that are phylogenetically closer to each other than those of phages that infect distantly related hosts. However, some of the phages appear to be chimeras, indicating that, at least occasionally, some genetic shuffling has occurred between the different T4-type subgroups. The compilation of a number of gene 23 sequences reveals a pattern of conserved motifs separated by sequences that differ in the M-type subgroups. Such variable patches in the gene 23 sequences may determine the size of the virion head and consequently the viral genome length. This sequence analysis provides molecular evidence that phages related to T4 are widespread in the biosphere and diverged from a common ancestor in acquiring the ability to infect different host bacteria and to occupy new ecological niches. [TOP OF PAGE]

23. Interaction of the PhiHSIC virus with its host: lysogeny or pseudolysogeny? Williamson, S.J., McLaughlin, M.R., Paul, J.H. (2001). Appl. Environ. Microbiol. 67:1682-1688. The marine phage PhiHSIC has been previously reported to enter into a lysogenic relationship with its host, HSIC, identified as Listonella pelagia. This phage produces a variety of plaques on its host, including turbid and haloed plaques, from which lysogens were previously isolated. These lysogens were unstable during long-term storage at -80° C and were lost. When HSIC was reinfected with phage PhiHSIC, pseudolysogen-like interactions between the phage and its host were observed. The cells (termed HSIC-2 or HSIC-2e) produced high viral titers (10¹¹ ml^-1) in the absence of inoculating phage and yet reached culture densities of nearly 10⁹ ml^-1. Prophages were not induced by mitomycin C or the polyaromatic hydrocarbon naphthalene in cells harboring such infections. However, such cells were homoimmune to superinfection. Colonies hybridized strongly with a gene probe from a 100-bp fragment of the PhiHSIC genome, while the host did not. Analysis of chromosomal DNA preparations suggested the presence of a chromosomally integrated prophage. Phage adsorption experiments suggested that HSIC-2 was adsorption impaired. Because of the chromosomal prophage integration and homoimmunity, we interpret these results to indicate that PhiHSIC establishes a lysogenic relationship with its host that involves an extremely high level of spontaneous induction. This could be caused by a weak repressor of phage production. Additionally, poor phage adsorption of HSIC-2 compared to the wild type probably helped maintain this pseudolysogen-like relationship. In many ways, pseudolysogenic phage-host interactions may provide a paradigm for phage-host interactions in the marine environment. [TOP OF PAGE]

24. Quantifying viral propagation in vitro: toward a method for characterization of complex phenotypes. Yin, J., Duca, K., Lam, V., Keren, I., Endler, E.E., Letchworth, G.J., Novella, I.S. (2001). Biotechnology Progress [TOP OF PAGE]

25. Characterization of a lytic Lactobacillus plantarum bacteriophage and molecular cloning of a lysin gene in Escherichia coli. Yoon, S.S., Kim, J.W., Breidt, F., Fleming, H.P. (2001). Int. J. Food Microbiol. 65:63-74. Bacteriophage SC921, which can infect Lactobacillus plantarum specifically, was isolated from a fermented vegetable source, Kimchi. This phage is active against six of 11 strains of L. plantarum tested as hosts. Morphologically, it has an isometric head (60 nm in diameter) and a non-contractile tail (260 nm long and 9-11 nm wide), indicating that it belongs to Bradley's group B or the Siphoviridae family according to the International Committee on Taxonomy of Viruses (ICTV). The bouyant density was 1.58 g/cm3. SDS-PAGE experimentation indicated that the phage particle contains two major structural proteins and several minor proteins. The genome was a double stranded linear DNA molecule with cohesive ends and 66.5 kb long by mapping genomic DNA digested with the restriction endonucleases: KpnI, SmaI, and XbaI. The (G + C) content of the phage DNA is 39.4%. For this lysin gene study, 9.4 kb of KpnI-digested DNA fragment was cloned into pUC19 and expressed in Escherichia coli. The KpnI fragment was considered as the genetic element responsible for the lysis gene of L. plantarum bacteriophage. The cloned fragment in pUC19 was hybridized to a 9.4-kb fragment generated by KpnI digestion of SC 921 as a probe. This confirmed that the fragment in pUC19 originated from phage DNA. The lysin gene was near the middle of the phage genome. [TOP OF PAGE]

26. Simulating the growth of viruses. You, L., Yin, J. (2001). Pacific Symposium on Biocomputing 532-543. To explore how the genome of an organism defines its growth, we have developed a computer simulation for the intracellular growth of phage T7 on its E. coli host. Our simulation, which incorporates 30 years of genetic, biochemical, physiological, and biophysical data, is used here to study how the intracellular resources of the host, determined by the specific growth rate of the host, contribute toward phage development. It is also used to probe how changes in the linear organization of genetic elements on the T7 genome can affect T7 development. Further, we show how time-series trajectories of T7 mRNA and protein levels generated by the simulation may be used as raw data to test data-mining strategies, specifically, to identify partners in protein-protein interactions. Finally, we suggest how generalization of this work can lead to a knowledge-driven simulation for the growth of any virus. [TOP OF PAGE]

27. Purification of Piscirickettsia salmonis and associated phage particles. Yuksel, S.A., Thompson, K.D., Ellis, A.E., Adams, A. (2001). Diseases of Aquatic Organisms 44:231-235. Piscirickettsia salmonis was isolated from cell culture using differential centrifugation and purified on a 30% Percol gradient. The purity of the preparation was assessed by transmission electron microscopy and phage-like particles were found to be associated with some of the P. salmonis isolates examined. This is believed to be the first report of a phage associated with rickettsia from fish. [TOP OF PAGE]

28. The murky origin of Snow White and her T-even dwarfs. Abedon, S.T. (2000). Genetics 155:481-486. The T-even bacteriophages—T2, T4, and T6—represent facile experimental systems that are both relatively complex and meticulously well defined. They played essential roles in the birth and early nurturing of the field of molecular genetics, and could serve similarly as model organisms for ecology. Identification of the source habitat from which these phages were isolated would be satisfying from an ecological as well as historical perspective. Here I infer, mostly from published materials, the habitats from which these three phages were isolated, plus I delve into the history of their host, Eschcerichia coli B. [TOP OF PAGE]

29. The evolution of pathogen-host interactions mediated by bacteriophages. Ai, Y.-C., Meng, F., Zeng, Y. (2000). Acta Microbiologica Sinica 40:657-661. [TOP OF PAGE]

30. A proposal for the reclassification of Bdellovibrio stolpii and Bdellovibrio starrii into a new genus, Bacteriovorax gen. nov. as Bacteriovorax stolpii comb. nov. and Bacteriovorax starrii comb. nov., respectively. Baer, M.L., Ravel, J., Chun, J., Hill, R.T., Williams, H.N. (2000). Int J Syst Evol Microbiol 50 Pt 1:219-224. Bdellovibrios are unique bacteria with the ability to prey upon a wide variety of susceptible Gram-negative bacteria. Micro-organisms exhibiting this trait have been included in the genus Bdellovibrio despite their isolation from diverse habitats and relatively unstudied taxonomic relatedness. In this study, 16S rDNA sequences were compared from known terrestrial Bdellovibrio species, Bdellovibrio bacteriovorus 100T, Bdellovibrio stolpii Uki2T and Bdellovibrio starrii A3.12T in order to study their phylogenetic relationship. The two sequences from B. stolpii Uki2T and B. starrii A3.12T were 90.0% similar to each other but exhibited only 81.7% and 81.2% similarity, respectively to B. bacteriovorus 100T. Phylogenetic analysis indicated that B. bacteriovorus 100T clustered in a separate clade from B. starrii A3.12T and B. stolpii Uki2T, demonstrating only a distant relationship between B. bacteriovorus 100T and the other two recognized type species. DNA-DNA hybridization experiments also demonstrated <4% hybridization between these three species. On the basis of the results obtained from the phylogenetic analysis and DNA-DNA hybridization studies, it is proposed that B. stolpii Uki2T and B. starrii A3.12T should be transferred to a new genus, Bacteriovorax gen. nov. as Bacteriovorax stolpii comb. nov. and Bacteriovorax starrii comb. nov., respectively. It is also proposed that the type species for the new genus Bacteriovorax should be Bacteriovorax stolpii comb. nov. [TOP OF PAGE]

31. Characterization of mesophilic mixed starter cultures used for the manufacture of aged cheddar cheese. Bissonnette, F., Labrie, S., Deveau, H., Lamoureux, M., Moineau, S. (2000). Journal of Dairy Science 83:620-627. Seventy-one different Lactococcus lactis subsp. cremoris strains were isolated from seven mesophilic mixed starters used in the manufacture of aged Cheddar cheese in Canada. Based on plasmid profiles and growth in milk (with or without glucose, Casamino Acids or both), two mixed starters were highly heterogeneous, containing at least 18 to 24 distinct L. lactis strains. Three mixed starters were comprised of seven to nine strains, whereas two starters were relatively homogeneous, containing two or three strains. Many strains with similar plasmid profiles behaved differently during growth in milk, indicating variability in the phenotypes. Only 20% of the strains could grow in plain milk, whereas 30% could not grow in milk supplemented with glucose and Casamino Acids. Twenty-five lactococcal bacteriophages were also isolated from whey samples with single strains as hosts. Eighteen phages belonged to the 936 species and seven to the c2 species. Thirteen strains were insensitive to all 25 phages. Almost all sensitive strains were phage species-specific. The 936-like phages had a broader host range. [TOP OF PAGE]

32. Linking genetic change to community evolution: insights from studies of bacteria and bacteriophage. Bohannan, B.J.M., Lenski, R.E. (2000). Ecological Letters 3:362-377. A major goal of community ecology is to link biological processes at lower scales with community patterns. Microbial communities are especially powerful model systems for making these links. In this article, we review recent studies of laboratory communities of bacteria and bacteriophage (viruses that infect bacteria). We focus on the ecology and evolution of bacteriophage-resistance as a case study demonstrating the relationship between specific genes, individual interactions, population dynamics, community structure, and evolutionary change. In laboratory communities of bacteria and bacteriophage, bacteria rapidly evolve resistance to bacteriophage infection. Different resistance mutations produce distinct resistance phenotypes, differing, for example, in whether resistance is partial or complete, in the magnitude of the physiological cost associated with resistance, and in whether the mutation can be countered by a host-range mutation in the bacteriophage. These differences determine whether a mutant can invade, the effect its invasion has on the population dynamics of sensitive bacteria and phage, and the resulting structure of the community. All of these effects, in turn, govern the community's response to environmental change and its subsequent evolution. [TOP OF PAGE]

33. The relative importance of competition and predation varies with productivity in a model community. Bohannan, B.J.M., Lenski, R.E. (2000). Am. Nat. 156:329-340. Recent theory predicts that productivity can influence the relative importance of predation and competition in determining patterns in abundance, diversity, and community structure. In low-productivity systems, competition is predicted to be the major influence on community patterns, while at high productivity, the major influence is predicted to be predation. We directly tested this theory using a laboratory model community. Our model community consisted of the bacteriophage T2 (a virus that feeds on Escherichia coli) and two populations of E. coli, in glucose-limited chemostats. One E. coli population consisted of individuals that were sensitive to predation by T2 ("vulnerable" E. coli), and the other population consisted of individuals that were partially resistant to predation by T2 ("less vulnerable" E. coli). We manipulated productivity in this experiment by running replicate chemostats with different input concentrations of glucose. Our observations were consistent with theoretical predictions. We observed the decline of the more vulnerable prey population at higher productivity but not at lower productivity, and the decline of the less vulnerable prey population at lower productivity but not at higher productivity. However, the rate of decline in some replicates was slower than predicted, and extinctions were not observed during the experiments, contrary to theoretical predictions. We present some testable hypotheses that might explain the slow rate of decline observed. [TOP OF PAGE]

34. Big-benefit mutations in a bacteriophage inhibited with heat. Bull, J.J., Badgett, M.R., Wichman, H.A. (2000). Molecular Biology and Evolution 17:942-950. High temperature inhibits the growth of the wild-type bacteriophage phiX174. Three different point mutations were identified that each recovered growth at high temperature. Two affected the major capsid protein (residues F188 and F242), and one affected the internal scaffolding protein (B114). One of the major capsid mutations (F242) is located in a beta strand that contacts B114 in the procapsid during viral maturation, whereas the other capsid mutation (F188) is involved in subunit interactions at the threefold axis of symmetry. Selective coefficients of these mutations ranged from 13.9 to 3.8 in the inhibitory, hot environment, but all mutations reduced fitness at normal temperature. The selective effect of one of the mutations (F242) was evaluated at high temperature in four different genetic backgrounds and exhibited epistasis of diminishing returns: as log fitness of the background genotype increased from -0.1 to 4.1, the fitness boost provided by the F242 mutation decreased from 3.9 to 0. 8. These results support a model in which viral fitness is bounded by an upper limit and the benefit of a mutation is scaled according to the remaining opportunity for fitness improvement in the genome. [TOP OF PAGE]

35. Evolvability of an RNA virus is determined by its mutational neighbourhood. Burch, C.L., Chao, L. (2000). Nature 406:625-628. The ubiquity of mechanisms that generate genetic variation has spurred arguments that evolvability, the ability to generate adaptive variation, has itself evolved in response to natural selection. The high mutation rate of RNA viruses is postulated to be an adaptation for evolvability, but the paradox is that whereas some RNA viruses evolve at high rates, others are highly stable. Here we show that evolvability in the RNA bacteriophage phi6 is also determined by the accessibility of advantageous genotypes within the mutational neighbourhood (the set of mutants one or a few mutational steps away). We found that two phi6 populations that were derived from a single ancestral phage repeatedly evolved at different rates and toward different fitness maxima. Fitness measurements of individual phages showed that the fitness distribution of mutants differed between the two populations. Whereas population A, which evolved toward a higher maximum, had a distribution that contained many advantageous mutants, population B, which evolved toward a lower maximum, had a distribution that contained only deleterious mutants. We interpret these distributions to measure the fitness effects of genotypes that are mutationally available to the two populations. Thus, the evolvability of phi6 is constrained by the distribution of its mutational neighbours, despite the fact that this phage has the characteristic high mutation rate of RNA viruses. [TOP OF PAGE]

36. Evolution of virulence in parasites: making hard and soft choices. Chao, L., Hanley, K.A., Burch, C.L., Dahlberg, C., Turner, P.E. (2000). Q. Rev. Biol. 75:261-275. [TOP OF PAGE]

37. Transfer of conjugative plasmids and bacteriophage lambda occurs in the presence of antibiotics that prevent de novo gene expression. Cooper, T.F., Heinemann, J.A. (2000). Plasmid ???:???-??? [TOP OF PAGE]

38. Evolutionary Reversals During Viral Adaptation to Alternating Hosts. Crill, W.D., Wichman, H.A., Bull, J.J. (2000). Genetics 154:27-37. Experimental adaptation of the bacteriophage fX174 to a Salmonella host depressed its ability to grow on the traditional Escherichia host, whereas adaptation to Escherichia did not appreciably affect growth on Salmonella. Continued host switching consistently exhibited this pattern. Growth inhibition on Escherichia resulted from two to three substitutions in the major capsid gene. When these phages were forced to grow again on Escherichia, fitness recovery occurred predominantly by reversions at these same sites, rather than by second-site compensatory changes, the more frequently observed mechanism in most microbial systems. The affected residues lie on the virion surface and they alter attachment efficiency, yet they occur in a region distinct from a putative binding region previously identified from X-ray crystallography. These residues not only experienced high rates of evolution in our experiments, but also exhibited high levels of radical amino acid variation among fX174 and its known relatives, consistent with a history of adaptation involving these sites. [TOP OF PAGE]

39. Comparative genomics of the late gene cluster from Lactobacillus phages. Desiere, F., Pridmore, R.D., Brossow, H. (2000). Virology 275:294-305. Three prophage sequences were identified in the Lactobacillus johnsoni strain NCC533. Prophage Lj965 predicted a gene map very similar to those of pac-site Streptococcus thermophilus phages over its DNA packaging and head and tail morphogenesis modules. Sequence similarity linked the putative DNA packaging and head morphogenesis genes at the protein level. Prophage Lj965/S. thermophilus phage Sfi 11/Lactococcus lactis phage TP901-1 on one band and Lactobacillus delbrueckii phage LL-H/Lactbacillus plantarum phage phig 1e/Listeria monocytogenes phage A118 on the other hand defined two sublines of structural gene clusters in pac-site Siphoviridae from low-GC Gram-positive bacteria. Bacillus subtilis phage SPP1 linked both sublines. The putative major head and tail proteins from Lj965 shared weak sequence similarity with phages from Gram-negative bacteria. A clearly independent line of structural genes in Siphoviridae from low-GC Grampositive bacteria is defined by temperate cos-site phages including Lactobacillus gasserl phage adh, which also shared sequence similarity with phage D3 infecting a Gram-negative bacterium. A phylogenetic tree analysis demonstrated that the ClpP-like protein identified in four cos-site Siphoviridae from Lactobacillus, Lactococcus, Streptococcus, and Pseudomonas showed graded sequence relationships The tree suggested that the ClpP-like proteins from the phages were not acquired by horizontal gene transfer from their corresponding bacterial hosts. [TOP OF PAGE]

40. Bacteriophages of spirochetes. Eggers, C.H., Casjens, S., Hayes, S.F., Garon, C.F., Damman, C.J., Oliver, D.B., Samuels, D.S. (2000). J Mol Microbiol Biotechnol 2:365-373. Historically, a number of bacteriophage-like particles have been observed in association with members of the bacterial order Spirochetales, the spirochetes. In the last decade, several spirochete bacteriophages have been isolated and characterized at the molecular level. We have recently characterized a bacteriophage of the Lyme disease agent, Borrelia burgdorferi, which we have designated phiBB-1. Here we review the history of the association between the spirochetes and their bacteriophages, with a particular emphasis on phiBB-1 and its prophage, the 32-kb circular plasmid family of B. burgdorferi. [TOP OF PAGE]

41. The two faces of mutation: Extinction and adaptation in RNA viruses. Elena, S.F., Miralles, R., Cuevas, J.M., Turner, P.E., Moya, A. (2000). IUBMB Life 49:5-9. From a population standpoint, two main features characterize the replication of RNA viruses and viruses that use RNA as a replicative intermediate: high genetic variability, and enormous fluctuations in population size. Their genetic variability mainly reflects a lack of the proof-reading and post-replicative error correction mechanisms that operate during cellular DNA replication, but recombination and segment exchange can also play an important role, Viral population size can change tremendously as a consequence of transmission between hosts or between different tissues within an infected host. A new infection can be initiated with very few particles that subsequently expand many trillion-fold. Repeated bottleneck events can lead to drastic fitness losses or even to viral extinction, whereas continuously large population sizes result in fitness gains and adaptation. Here we review experimental evidence for the effects of mutation, selection, and genetic drift on the adaptation and extinction of RNA viruses. [TOP OF PAGE]

42. Computation, prediction, and experimental tests of fitness for bacteriophage T7 mutants with permuted genomes. Endy, D., You, L., Yin, J., Molineux, I.J. (2000). Proc. Natl. Acad. Sci. USA 97:5375-5380. We created a simulation based on experimental data from bacteriophage T7 that computes the developmental cycle of the wild-type phage and also of mutants that have an altered genome order. We used the simulation to compute the fitness of more than 10⁵ mutants. We tested these computations by constructing and experimentally characterizing T7 mutants in which we repositioned gene 1, coding for T7 RNA polymerase. Computed protein synthesis rates for ectopic gene 1 strains were in moderate agreement with observed rates. Computed phage-doubling rates were close to observations for two of four strains, but significantly overestimated those of the other two. Computations indicate that the genome organization of wild-type T7 is nearly optimal for growth: only 2.8% of random genome permutations were computed to grow faster, the highest 31% faster, than wild type. Specific discrepancies between computations and observations suggest that a better understanding of the translation efficiency of individual mRNAs and the functions of qualitatively "nonessential" genes will be needed to improve the T7 simulation. In silico representations of biological systems can serve to assess and advance our understanding of the underlying biology. Iteration between computation, prediction, and observation should increase the rate at which biological hypotheses are formulated and tested. [TOP OF PAGE]

43. Toward antiviral strategies that resist viral escape. Endy, D., Yin, J. (2000). Antimicrobial Agents and Chemotherapy 44:1097-1099. We studied the effect on viral growth of drugs targeting different virus functions using a computer simulation for the intracellular growth of bacteriophage T7. We found that drugs targeting components of negative-feedback loops gain effectiveness against mutant viruses that attenuate the drug-target interaction. The greater inhibition of such mutants than of the wild type suggests a drug design strategy that would hinder the development of drug resistance. [TOP OF PAGE]

44. Bacterial indicator occurrence and the use of an F+ specific RNA coliphage assay to identify fecal sources in Homosassa Springs, Florida. Griffin, D.W., Stokes, R., Rose, J.B., Paul, J.H., III (2000). Microb. Ecol. 39:56-64. A microbiological water quality study of Homosassa Springs State Wildlife Park (HSSWP) and surrounding areas was undertaken. Samples were collected in November of 1997 (seven sites) and again in November of 1998 (nine sites). Fecal bacterial concentrations (total and fecal coliforms, Clostridium perfringens, and enterococci) were measured as relative indicators of fecal contamination. F⁺-specific coliphage genotyping was performed to determine the source of fecal contamination at the study sites. Bacterial levels were considerably higher at most sites in the 1997 sampling compared to the 1998 sampling, probably because of the greater rainfall that year. In November of 1997, 2 of the 7 sites were in violation of all indicator standards and guidance levels. In November of 1998, 1 of 9 sites was in violation of all indicator standard and guidance levels. The highest concentrations of all fecal indicators were found at a station downstream of the animal holding pens in HSSWP. The lowest levels of indicators were found at the Homosassa Main Spring vent. Levels of fecal indicators downstream of HSSWP (near the point of confluence with the river) were equivalent to those found in the Southeastern Fork and areas upstream of the park influences. F⁺ specific RNA coliphage analysis indicated that fecal contamination at all sites that tested positive was from animal sources (mammals and birds). These results suggest that animal (indigenous and those in HSSWP) and not human sources influenced microbial water quality in the area of Homosassa River covered by this study. [TOP OF PAGE]

45. Characterization of natural isolates of Lactobacillus strains to be used as starter cultures in dairy fermentation. Hebert, E.M., Raya, R.R., Tailliez, P., de, G.G. (2000). Int. J. Food Microbiol. 59:19-27. The technological relevant characteristics of five homofermentative lactobacilli strains, isolated from natural fermented hard cheeses, were studied. Isolates CRL 581 and CRL 654, from Argentinian artesanal hard cheeses, and isolates CRL 1177, CRL 1178, and CRL 1179, from Italian Grana cheeses, were identified as Lactobacillus delbrueckii subsp. lactis and Lactobacillus helveticus, respectively, by physiological and biochemical tests, SDS-PAGE of whole-cell proteins and sequencing of the variable (V1) region of the 16S ribosomal DNA. All strains showed high levels of beta-galactosidase activity. However, proteolytic activity varied widely among isolates. Strains CRL 581, CRL 654, and CRL 1177 hydrolyzed alpha- and beta-caseins and were able to coagulate reconstituted skim milk in less than 16 h at 42 degrees C. According to the substrate specificity, these proteinases have a caseinolytic activity comparable to that of the P(III)-type of lactococcal proteinases. No strains produced inhibitor substances (bacteriocin) and all were insensitive to attack by 14 L. helveticus- and L. delbrueckii subsp. lactis-specific bacteriophages. [TOP OF PAGE]

46. Ultraviolet radiation effects on bacterioplankton and viruses in marine ecosystems. Jeffrey, W.H., Kase, J.P., Wilhelm, S.W. (2000). pp. 206-236. In In De Mora, S.J. and et al. (eds.), Effects Of UV Radiation On Marine Ecosystems. Cambridge University Press, Cambridge. [TOP OF PAGE]

47. Sequence of the genome of the temperate, serotype-converting, Pseudomonas aeruginosa bacteriophage D3. Kropinski, A.M. (2000). J. Bacteriol. 182:6066-6074. Temperate bacteriophage D3, a member of the virus family Siphoviridae, is responsible for serotype conversion in its host, Pseudomonas aeruginosa. The complete sequence of the double-stranded DNA genome has been determined. The 56,426 bp contains 90 putative open reading frames (ORFs) and four genes specifying tRNAs. The latter are specific for methionine (AUG), glycine (GGA), asparagine (AAC), and threonine (ACA). The tRNAs may function in the translation of certain highly expressed proteins from this relatively AT-rich genome. D3 proteins which exhibited a high degree of sequence similarity to previously characterized phage proteins included the portal, major head, tail, and tail tape measure proteins, endolysin, integrase, helicase, and NinG. The layout of genes was reminiscent of lambdoid phages, with the exception of the placement of the endolysin gene, which parenthetically also lacked a cognate holin. The greatest sequence similarity was found in the morphogenesis genes to coliphages HK022 and HK97. Among the ORFs was discovered the gene encoding the fucosamine O-acetylase, which is in part responsible for the serotype conversion events. [TOP OF PAGE]

48. Multiplex PCR for detection and identification of lactococcal bacteriophages. Labrie, S., Moineau, S. (2000). Appl. Environ. Microbiol. 66:987-994. Three genetically distinct groups of Lactococcus lactis phages are encountered in dairy plants worldwide, namely, the 936, c2, and P335 species. The multiplex PCR method was adapted to detect, in a single reaction, the presence of these species in whey samples or in phage lysates. Three sets of primers, one for each species, were designed based on conserved regions of their genomes. The c2-specific primers were constructed using the major capsid protein gene (mcp) as the target. The mcp sequences for three phages (eb1, Q38, and Q44) were determined and compared with the two available in the databases, those for phages c2 and bIL67. An 86.4% identity was found over the five mcp genes. The gene of the only major structural protein (msp) was selected as a target for the detection of 936-related phages. The msp sequences for three phages (p2, Q7, and Q11) were also established and matched with the available data on phages sk1, bIL170, and F4-1. The comparison of the six msp genes revealed an 82. 2% identity. A high genomic diversity was observed among structural proteins of the P335-like phages suggesting that the classification of lactococcal phages within this species should be revised. Nevertheless, we have identified a common genomic region in 10 P335-like phages isolated from six countries. This region corresponded to orfF17-orf18 of phage r1t and orf20-orf21 of Tuc2009 and was sequenced for three additional P335 phages (Q30, P270, and ul40). An identity of 93.4% within a 739-bp region of the five phages was found. The detection limit of the multiplex PCR method in whey was 10(4) to 10(7) PFU/ml and was 10(3) to 10(5) PFU/ml with an additional phage concentration step. The method can also be used to detect phage DNA in whey powders and may also detect prophage or defective phage in the bacterial genome. [TOP OF PAGE]

49. Properties of natural interspecific hybrids of transposable phages of Pseudomonas aeruginosa: specific characteristics of phage PL24 transposition. Mit'kina, L.N., Krylov, V.N. (2000). Genetika 36:1330-1339. Properties of natural hybrid transposable phages (TP) of Pseudomonas aeruginosa, including phage PL24 and lysogens for this phage, were studied. PL24 possesses the properties of TP from two previously described groups, B3 and D3112. Its genome, unlike the genome of D3112, contains many sites susceptible to the SalGI restriction endonuclease and possesses no more than 100 nucleotides of bacterial origin located at the left genome end. However, unlike B3, phage PL24 failed to induce auxotrophic mutants upon integration in the bacterial genome. This phage differed from both B3 and D3112 in sensitivity to chloroform treatment. A more detailed examination of a group containing 25 randomly isolated lysogens for phage PL24 revealed previously unknown processes occurring at early stages of bacterial lysogenization. There are at least two different modes of cell lysogenization with phage PL24. In the first case, the emerging lysogens contained a single prophage genome located (in each lysogen) at individual sites. In the second case, polylysogenic bacteria appeared, and, after primary integration of a phage genome, replicative transposition occurred at new sites (often accompanied by the appearance of prophage clusters at these sites). The choice of the mode of lysogenization can be determined both by differences in the physiological state of bacteria and by specific features of phage PL24, which possibly affect the time of repressor accumulation to the concentration sufficient for blocking phage growth or the stability of the lysogenic state. [TOP OF PAGE]

50. Investigation of the relationship between lysogeny and lysis of Lactococcus lactis in cheese using prophage-targeted PCR. O'Sullivan, D., Ross, R.P., Fitzgerald, G.F., Coffey, A. (2000). Appl. Environ. Microbiol. 66:2192-2198. The ability of lactococcal strains to lyse (and release intracellular enzymes) during cheese manufacture can be a very desirable trait and has been associated with improvement of flavor and acceleration of cheese ripening. Using a laboratory-scale cheese manufacturing assay, the autocatalytic behavior of 31 strains of Lactococcus lactis was assessed. In general, marked variation was observed between strains with a 20-fold difference between best and worst lysing strains based on the release of intracellular enzyme lactate dehydrogenase. In a parallel experiment, the genomes of these strains were examined for the presence of prophage integrase (int) sequences by using concerved primer sequences from known lysogenic phage. Results demonstrated that the lytic behavior of lactococcal starter strains significantly correlates with the presence of prophage sequences. These results highlight not only the contribution of prophage to starter cell lysis but also the potential of PCR as a useful screen to assess strains for this important industrial trait. [TOP OF PAGE]

51. Genotypic variations of Shiga toxin-converting phages from enterohaemorrhagic Escherichia coli O157: H7 isolates. Osawa, R., Wada, A., Iyoda, S., Nakayama, S.I., Yamai, S., Watanabe, H. (2000). J. Med. Microbiol. 49:565-574. Pulsed-field gel electrophoresis (PFGE) analysis revealed that enterohaemorrhagic Escherichia coli (EHEC) O157:H7 strains had considerable variations in their genomes. This study investigated whether or not the molecular profile of Shiga toxin (Stx) 1- and Stx2-converting phages isolated from EHEC O157:H7 strains, derived from various sources in the USA and Japan, corresponded to the variations of host strains' genotypes as determined by PFGE. A total of 51 Stx-converting phages including 12 Stx1-converting phages and 37 Stx2-converting phages was isolated from seven USA isolates and 20 Japanese isolates. The average Dice coefficient values showed 44% similarity between phage DNAs in Stx2-converting phages digested with SmaI and 55% in Stx1-converting phages digested with HindIII, indicating considerable variation among phage DNA. In particular, restriction fragment length polymorphism (RFLP) patterns of Stx2-converting phage DNA varied according to the PFGE type of their host strain, which suggests that the phage genomes have altered their genotypic characteristics with those of host genomes. However, there are several exceptions: the RFLP patterns of some Stx2-converting phages were quite similar irrespective of the different genotypes of the host strains, indicating that horizontal transfer of Stx2-converting phage may also occur under some circumstances. [TOP OF PAGE]

52. Bacteriophage phiYeO3-12, specific for Yersinia enterocolitica serotype O:3, is related to coliphages T3 and T7. Pajunen, M., Kiljunen, S., Skurnik, M. (2000). J. Bacteriol. 182:5114-5120. Bacteriophage phiYeO3-12 is a lytic phage of Yersinia enterocolitica serotype O:3. The phage receptor is the lipopolysaccharide O chain of this serotype that consists of the rare sugar 6-deoxy-L-altropyranose. A one-step growth curve of phiYeO3-12 revealed eclipse and latent periods of 15 and 25 min, respectively, with a burst size of about 120 PFU per infected cell. In electron microscopy phiYeO3-12 virions showed pentagonal outlines, indicating their icosahedral nature. The phage capsid was shown to be composed of at least 10 structural proteins, of which a protein of 43 kDa was predominant. N-terminal sequences of three structural proteins were determined, two of them showing strong homology to structural proteins of coliphages T3 and T7. The phage genome was found to consist of a double-stranded DNA molecule of 40 kb without cohesive ends. A physical map of the phage DNA was constructed using five restriction enzymes. The phage infection could be effectively neutralized using serum from a rabbit immunized with whole phiYeO3-12 particles. The antiserum also neutralized T3 infection, although not as efficiently as that of phiYeO3-12. phiYeO3-12 was found to share, in addition to the N-terminal sequence homology, several common features with T3, including morphology and nonsubjectibility to F exclusion. The evidence conclusively indicated that phiYeO3-12 is the first close relative of phage T3 to be described. [TOP OF PAGE]

53. Rapid movement of wastewater from on-site disposal systems into surface waters in the Lower Florida Keys. Paul, J.H., McLaughlin, M.R., Griffin, D.W., Lipp, E.K., Stokes, R., Rose, J.B. (2000). Estuaries 23:662-668. Viral tracer studies have been used previously to study the potential for wastewater contamination of surface marine waters in the Upper and Middle Florida Keys. Two bacteriophages, the marine bacteriophage phiHSIC and the Salmonella phage PRD1, were used as tracers in injection well and septic tank studies in Saddlebunch Keys of the Lower Florida Keys and in septic tank studies in Boot Key Harbor, Marathon, of the Middle Keys. In Boot Key Harbor, both phages were detected in a canal adjacent to the seeded septic tank within 3 h 15 min of the end of the seed period. The tracer was then detected at all sampling sites in Boot Key Harbor, including one on the opposite side of U.S. Highway 1 in Florida Bay, and at an Atlantic Ocean beach outside Boot Key Harbor. Rates of migration based on first appearance of the phage ranged from 1.7 to 57.5 m h-1. In Saddlebunch Keys, phiHSIC and PRD1 were used to seed a residential septic tank and a commercial injection well. The septic tank tracer was not found in any surface water samples. The injection well tracer was first detected at a site most distant from the seed site, a channel that connected Sugarloaf Sound with the Atlantic Ocean. The rate of tracer migration from the injection well to this channel ranged from 66.8 to 141 m h-1. Both tracer studies showed a rapid movement of wastewater from on-site sewage treatment and disposal systems in a southeasterly direction toward the reef tract and Atlantic Ocean, with preferential movement through tidal channels. These studies indicate that wastewater disposal systems currently in widespread use in the Florida Keys can rapidly contaminate the marine environment. [TOP OF PAGE]

54. Ecology of bacteriophages in nature. Paul, J.H., Kellogg, C.A. (2000). pp. 211-246. In In Hurst, C.J. (ed.), Viral Ecology. Academic Press, San Diego. [no abstract?]. [TOP OF PAGE]

55. Dynamics of bacterial community composition and activity during mesocosm diatom blooms. Riemann, L., Steward, G.F., Azam, F. (2000). Appl. Environ. Microbiol. 66:578-587. Bacterial community composition, enzymatic activities, and carbon dynamics were examined during diatom blooms in four, 200 liter laboratory seawater mesocosms. The objective was to determine whether the dramatic shifts in growth rates and ectoenzyme activities, which are commonly observed during the course of phytoplankton blooms and their subsequent demise, could result from shifts in bacterial community composition. Nutrient enrichment of metazoan-free seawater resulted in diatom blooms dominated by Thalassiosira sp. which peaked nine days after enrichment ( 24 g chl a l^-1). At this time bacterial abundance abruptly decreased from 2.8 to 0.75 x 10⁶ ml^-1 and analysis of bacterial community composition, by denaturing gradient gel electrophoresis (DGGE) of PCR-amplified, 16S rRNA gene fragments, revealed a disappearance of three dominant phylotypes. Increased viral and flagellate abundance suggested that both lysis and grazing could have played a role in the observed phylotype-specific mortality. Subsequently, new phylotypes appeared and bacterial production, abundance and enzyme activities shifted from being predominantly associated with the <1.0 m size-fraction towards the >1.0 m size-fraction indicating a pronounced microbial colonization of particles. Sequencing of DGGE bands suggested that the observed rapid and extensive colonization of particulate matter was mainly by specialized ??Proteobacteria and Cytophagales-related phylotypes. These particle-associated bacteria had high growth rates as well as high cell specific aminopeptidase, ??glucosidase and lipase activities. Rate measurements as well as bacterial population dynamics were almost identical among the mesocosms indicating that the observed bacterial community dynamics were systematic and repeatable responses to the manipulated conditions. [TOP OF PAGE]

56. The complete genomic sequence of the marine phage Roseophage SIO1 shares homology with nonmarine phages. Rohwer, F., Segall, A., Steward, G., Seguritan, V., Breitbart, M., Wolven, F., Azam, F. (2000). Limnol. Oceanogr. 45:408-418. Viruses are ubiquitous components of the marine environment, frequently reaching concentrations of 10⁷-10⁸ viruses per milliliter of surface seawater The majority of these viral particles are bacteriophages (phages). Although the oceans are probably the largest pool of bacteriophages on the planet, the evolutionary relationships of marine phages to phages from other environments are unknown. To address this issue, we have completely sequenced the genome of the lytic marine phage, Roseophage SIO1, that infects the heterotrophic marine bacterium Roseobacter SIO67. This phage has an isometric capsid with a diameter of approximately 43 nm, a short tail, a buoyant density of 1.49 g cm^-3 in CsCl, and a 39,906-bp dsDNA genome. Sequence similarities and relative positions within the genome suggest that three of the open reading frames (ORFs) are homologous to the primase, DNA polymerase, and endodeoxyribonuclease I proteins of coliphages T3 and T7. The results are consistent with the mosaic theory of phage evolution and indicate a genetic link between marine and nonmarine phages. Additionally, basic life histories of marine phages can be elucidated by comparison of complete genomes to those of other extensively studied phages (e.g., lambda, T4, T7). The DNA replication machinery of Roseophage SIO1 shows a clear homology with that of coliphages T3 and T7, suggesting that the process of DNA replication may be similar among these phages. The Roseophage SIO1 genome also encodes four predicted proteins involved in phosphate metabolism (RP PhoH, RP ribonucleotide reductase, RP Thy1, and RP endodeoxyribonuclease I) suggesting that phosphate recycling is important to Roseophage SIO1's life cycle. Other interesting clues about Roseophage SIO1's life history come from the absence of certain expected protein regions. For example, we have not been able to identify the Roseophage SIO1 structural proteins (e.g., capsid proteins) by homology to other phages. It is also conspicuous that the Roseophage SIO1 genome lacks a recognizable RNA polymerase, an essential component of T3 and T7 life cycles. Analysis of the Roseophage SIO1 genome shows that marine and nonmarine phages are genetically related but basic life histories may be significantly different. [TOP OF PAGE]

57. [Isolation and comparative study of a group of temperate bacteriophages of rhizospheric pseudomonads Pseudomonas putida]. Shaburova, O.V., Burkal'tseva, M.V., Pleteneva, E.A., Krylov, V.N. (2000). Genetika 36:915-919. We have isolated several new temperate bacteriophages for rhizosphere pseudomonads Pseudomonas putida. Examination of these phages, along with two previously isolated temperate phages PP56 and PP71 of P. putida PpG1 (biovar A), allowed us to classify them into four species on the basis of DNA cross-homology; relative genomic size; and, to a certain extent, the morphology of phage particles. Two of these species are represented by nonidentical variants. No transposable phages were found among these two new species. Three phage species cause various-types of lysogenic conversion manifested in growth suppression of other phage species. This seems to account for the fact that the temperate phage of rhizosphere pseudomonads are seldom encountered. The new phages described can be used for selection of phage-resistant bacterial forms exhibiting antifungal activity that are commercially produced and used for treatment of seeds of cultivated plants. [TOP OF PAGE]

58. Denaturing gradient gel electrophoresis resolves virus sequences amplified with degenerate primers. Short, S.M., Suttle, C.A. (2000). BioTechniques 28:20 [TOP OF PAGE]

59. Genome size distributions indicate variability and similarities among marine viral assemblages from diverse environments. Steward, G.F., Montiel, J.L., Azam, F. (2000). Limnol. Oceanogr. 45:1697-1706. Pulsed field gel electrophoresis (PFGE) was used to determine the size distributions of virus-like DNA in seawater from diverse environments (Arctic Ocean, Ross Sea, Coastal Pacific Ocean, and Northern Adriatic Sea). Changes in DNA banding patterns indicated that shifts in the viral assemblage composition occurred on the order of = 2 d during an intense dinoflagellate bloom in coastal Pacific waters. Different DNA banding patterns from diverse locations also indicated spatial variability in composition, but all of the samples analyzed had similar features. Size frequency distributions for virus-like genomes (VLGs) were multi-modal with major peaks occurring around 31-36 kilobases (kb) and 58-63 kb. The smallest discrete band resolved was 26 kb and the largest was >200 kb and the overall mean virus-like genome size was 50 ± 4 kb (mean ± sd, n = 30). On average, in surface seawater, > 90% of the VLGs occurred in the 26-69 kb size range and at least half were between 28 to 45 kb. This first extensive survey of viral genome sizes in seawater indicates that most marine viruses have physical properties similar to other known viruses. The distributions revealed that the vast majority of the detected VLGs have sizes typical of bacteriophages while only a few percent were in the size range of known algal viruses. [TOP OF PAGE]

60. Analysis of marine viral assemblages. Steward, G.F., Azam, F. (2000). pp. 159-165. In In Bell, C.R., Brylinski, M., and Johnson-Green, P. (eds.), Microbial Biosystems: New Frontiers. Atlantic Canada Society for Microbial Ecology, ??? Viruses are the numerically dominant microbes in every oceanic environment from the surface into the sediments. A liter of surface seawater from a typical mesotrophic area contains 10¹⁰ of them, about ten times more than bacteria. While total counts of viruses are becoming easier to make, we still know very little about the viruses that comprise a given assemblage. Infectivity assays are extremely useful and still the best way to assay for infectious viruses for any particular host. However, this approach requires that each potential host organism be cultured, making it impractical if not impossible to completely characterize natural assemblages. Morphological studies have been enlightening, but are time consuming and difficult to do quantitatively. Here we report a fingerprinting approach to characterize natural viral assemblages. In this approach, viruses are concentrated and intact viral genomes are separated based on their size via pulsed-field gel electrophoresis. The number of distinguishable bands provides a minimum estimate of the number of different viruses, while band position and staining intensity reveal the genome size distribution within the assemblage. With this technique we have detected spatial and temporal differences, as well as many similarities, in viral assemblages among a variety of marine habitats. Current efforts are directed toward combining this technique with other methods of fractionation and sequence analysis to allow both morphological and genetic description of uncultivated marine viruses. Direct investigation of dominant or particularly widespread viruses may ultimately provide clues as to which marine organisms contribute most to the viral pool, and which organisms are likely to be significantly influenced by viral mortality. [TOP OF PAGE]

61. Cyanophages and their role in the ecology of cyanobacteria. Suttle, C.A. (2000). pp. 563-589. In In Whitton, B.A. and Potts, M. (eds.), The Ecology of Cyanobacteria: Their Diversity in Time and Space. Kluwer Academic Publishers, Boston. [TOP OF PAGE]

62. The ecology, evolutionary and geochemical consequences of viral infection of cyanobacteria and eukaryotic algae. Suttle, C.A. (2000). pp. 248-286. In In Hurst, C.J. (ed.), Viral Ecology. Academic Press, New York. [TOP OF PAGE]

63. Cost of host radiation in an RNA virus. Turner, P.E., Elena, S.F. (2000). Genetics 156:1465-1470. Although host radiation allows a parasite to expand its ecological niche, traits governing the infection of multiple host types can decrease fitness in the original or alternate host environments. Reasons for this reduction in fitness include slower replication due to added genetic material or modifications, fitness trade-offs across host environments, and weaker selection resulting from simultaneous adaptation to multiple habitats. We examined the consequences of host radiation using vesicular stomatitis virus (VSV) and mammalian host cells in tissue culture. Replicate populations of VSV were allowed to evolve for 100 generations on the original host (BHK cells), on either of two novel hosts (HeLa and MDCK cells), or in environments where the availability of novel hosts fluctuated in a predictable or random way. As expected, each experimental population showed a substantial fitness gain in its own environment, but those evolved on new hosts (constant or fluctuating) suffered reduced competitiveness on the original host, However, whereas evolution on one novel host negatively correlated with performance on the unselected novel host, adaptation in fluctuating environments led to fitness improvements in both novel habitats. [TOP OF PAGE]

64. Experimental evolution recapitulates natural evolution. Wichman, H.A., Scott, L.A., Yarber, C.D., Bull, J.J. (2000). Philosophical Transactions of the Royal Society of London B Biological Sciences 355:1677-1684. Genomes of the closely related bacteriophages phiX174 and S13 are 5386 bases long and differ at 114 nucleotides, affecting 28 amino acids. Both parental phages were adapted to laboratory culture conditions in replicate lineages and analysed for nucleotide changes that accumulated experimentally. Of the 126 experimental substitutions, 90% encoded amino-acid changes, and 62% of the substitutions occurred in parallel in more than one experimental line. Furthermore, missense changes at 12 of the experimental sites were at residues differing between the parental phages; in ten cases the phiX174 experimental lineages were convergent with the S13 parent, or vice versa, at both the nucleotide and amino-acid levels. Convergence at a site was even obtained in both directions in three cases. These results point to a limited number of pathways taken during evolution in these viruses, and also raise the possibility that much of the amino-acid variation in the natural evolution of these viruses has been selected. [TOP OF PAGE]

65. Quantification of algal viruses in marine samples. Wilhelm, S.W., Poorvin, L. (2000). Methods in Microbiology 30:???-??? [TOP OF PAGE]

66. Bacterial carbon production in Lake Erie is influenced by viruses and solar radiation. Wilhelm, S.W., Smith, R.E.H. (2000). Canadian Journal of Fisheries and Aquatic Sciences 57:317-326. Bacterial production is an integral recycling mechanism that facilitates carbon flow through aquatic food webs. Factors influencing bacterial activity therefore impact carbon flow. Although ecologists consider grazing and dissolved organic carbon flux to be the major regulators of bacterial activity, we explored two other important pressures. Virus-like particle abundance ranged from 3.7 to 37.9 x 10¹⁰ L-1 in samples collected during August 1997 and July 1998. Bacterial abundance during these periods ranged from 1.8 to 4.6 x 10⁹ L-1. Based on electron microscopic analysis, viruses in Lake Erie would have been responsible for 12.1 to 23.4 % of bacterial mortality and, in quasi-steady state conditions, a comparable loss of bacterial productivity. In the central basin, solar radiation was also demonstrated to regulate bacterial productivity. Ultraviolet radiation (UVR, 295-400 nm) was shown to inhibit bacterial productivity according to a cumulative exposure kinetic model, and biological weighting functions were derived to enable calculation of time- and depth-integrated photoinhibition. The daytime photoinhibitory loss of bacterial carbon production was estimated to be 14 to 30% over the upper 5 m, primarily due to UVR > 320 nm. Viruses and sunlight are therefore of comparable importance as regulators of bacterial activity in this system. [TOP OF PAGE]

67. Virioplankton: viruses in aquatic ecosystems. Wommack, K.E., Colwell, R.R. (2000). Microbiology and Molecular Biology Reviews 64:69-114. [no abstract]. [TOP OF PAGE]

68. An exfoliative toxin A-converting phage isolated from Staphylococcus aureus strain ZM. Yoshizawa, Y., Sakurada, J., Sakurai, S., Machida, K., Kondo, I., Masuda, S. (2000). Microbiology and Immunology 44:189-191. Exfoliative toxin A (ETA) causes staphylococcal scalded-skin syndrome in children. The gene for ETA was believed to be coded by the chromosomal DNA. We isolated temperate phages from an ETA-producing strain, ZM, using a restriction minus strain, 1039, as an indicator. One of the prophages, designated phi-ZM-1 mediated lysogenic conversion of ETA. The polymerase chain reaction assay of the eta gene revealed that phage phi-ZM-1 carries the structural gene for ETA. [TOP OF PAGE]

69. Phages will out: strategies of host cell lysis. Young, R., Wang, I.-N., Roof, W.D. (2000). Trends in Microbiology 8:120-128. Most phages accomplish host lysis using a muralytic enzyme, or endolysin, and a holin, which permeabilizes the membrane at a programmed time and thus controls the length of the vegetative cycle. By contrast, lytic single-stranded RNA and DNA phages accomplish lysis by producing a single lysis protein without muralytic activity. [TOP OF PAGE]

70. Bacteriophage T4 resistance to lysis-inhibition collapse. Abedon, S.T. (1999). Genet. Res. 74:1-11. Lysis-inhibition is a mechanism of latent-period extension and burst-size increase that is induced by the T4 bacteriophage adsorption of T4-infected cells. Mutants of T4 genes imm, sp, and 5 (specifically the ts1 mutant of the latter) display some lysis inhibition. However, these mutants experience lysis-inhibition collapse, the lysis of lysis-inhibited cells, earlier than wild type-infected cells (i.e., their collapse occurs prematurely). Lysis from without is a lysis induced by excessive T4 adsorption. Gp5 is an inducer of lysis from without while gpimm and gpsp effect resistance to lysis from without. This paper shows that interfering with the adsorption of phages to imm-, sp-, or 5ts1-mutant-infected cells, in a variety of contexts, inhibits premature lysis-inhibition collapse. From these data, it is inferred that wild-type T4-infected cells display resistance to lysis-inhibition collapse by a mechanism resembling resistance to lysis from without. [TOP OF PAGE]

71. The genetic element pSSVx of the extremely thermophilic crenarchaeon Sulfolobus is a hybrid between a plasmid and a virus. Arnold, H.P., She, Q., Phan, H., Stedman, K., Prangishvili, D., Holz, I., Kristjansson, J.K., Garrett, R., Zillig, W. (1999). Molecular Microbiology 34:217-226. A new Sulfolobus islandicus strain, REY15/4, harboured both a novel fusellovirus, SSV2, and a small plasmid, pSSVx. The plasmid spread in S. solfataricus P1 together with the virus after infection with either the supernatant of a culture of REY15/4 or purified virus. Spreading of the plasmid required co-transfection with either SSV2 or the related SSV1 as helpers. Virus purified from REY15/4 constituted a mixture of two sizes of particles, one with the dimensions of a normal fusellovirus and the other smaller. Cloned SSV2 produced only the larger particles and only SSV2 DNA, indicating that the smaller particles contained pSSVx packaged into capsids made up of SSV2 components. The 5.7 kb genome of pSSVx revealed regions of high sequence similarity to the cryptic Sulfolobales plasmids pRN1, pRN2 and pDL10. Thus, pSSVx belongs to the family of pRN plasmids that share a highly conserved region, which probably constitutes the minimal replicon. They also contain a variable region showing no sequence similarity. In pSSVx, this region contains three open reading frames (ORFs), two of which are juxtapositioned and show high sequence similarity to a tandem of ORFs in fusellovirus genomes. Neither pRN1 nor pRN2, which lack this tandem, spread in the presence of the fuselloviruses, which implies that the sequences of these ORFs enable pSSVx to use the packaging system of the viral helpers for spreading. [TOP OF PAGE]

72. Epistatic interactions can lower the cost of resistance to multiple consumers. Bohannan, B.J.M., Travisano, M., Lenski, R.E. (1999). Evolution 53:292-295. It is widely assumed that resistance to consumers (e.g., predators or pathogens) comes at a "cost"; i.e., that when the consumer is absent the resident organisms are less fit than their susceptible counterparts. It is unclear what factors determine this cost. We demonstrate that epistasis between genes that confer resistance to two different consumers can alter the cost of resistance. We used as a model system the bacterium Escherichia coli and two different viruses (bacteriophage), T4 and l, that prey upon E. coli. Epistasis tended to reduce the costs of multiple resistance in this system. However, the extent of cost savings and its statistical significance depended on the environment in which fitness was measured, whether the null hypothesis for gene interaction was additive or multiplicative, and subtle differences among mutations that conferred the same resistance phenotype. [TOP OF PAGE]

73. Effect of resource supply rate on host-pathogen dynamics. Bohannan, B.J.M. (1999). AnonymousProceedings of the 8th International Symposium on Microbial Ecology. The dynamics of model host cell (E. coli) and model pathogen (bacteriophage) populations were studied in chemostats with different resource supply rates. Resource supply rate was manipulated by altering the concentration of the limiting resource (glucose) in the incoming media. Population responses to increased resourse supply rate were influenced by the vulnerability of the host cells to infection. When the host cell population consisted entirely of cells equally vulnerable to infection, both pathogen and host cells responded to increased resource supply rate with an increase in their average densities. In contrast, when the host cell contained some cells that were less vulnerable to infection (i.e., partially phage-resistant E. coli), only the pathogen population responded to increased supply rate with a signficant increase in average density. Furthermore, when the host cell population contained some cells completely invulnerable to infection (i.e., phage-resistant E. coli) only the host cell population responded to increased supply rate with an increase in average density. These responses were in general agreement with the predictions of mechanistic models of resource-consumer interactions. [TOP OF PAGE]

74. Effect of prey heterogeneity on the response of a food chain to resource enrichment. Bohannan, B.J.M., Lenski, R.E. (1999). Am. Nat. 153:73-82. We demonstrated that the presence of invulnerable prey can result in a shift in the balance between top-down and bottom-up control of a model food chain. Our model food chain consisted of the bacterium Escherichia coli and the bacteriophage T4 (a virus that feeds on E. coli) in chemostats supplied with different concentrations of glucose. The E. coli population consisted of individuals that were susceptible to predation by T4 ("edible" E. coli) and individuals that were resistant to predation by T4 ("inedible" E. coli). The equilibrium density of a hetergeneous prey population (consisting of edible and inedible E. coli) increased strongly in response to an enrichment of its resources. This response consisted of an increase in the inedible fraction of the prey population but no change in the edible fraction. In contrast, a homogeneous prey population (edible E. coli only) increased only marginally. The equilibrium density of the predator population (bacteriophage T4) did not significantly increase in response to enrichment when its prey were heterogeneous, but it increased when its prey were homogeneous. [TOP OF PAGE]

75. Evolution by small steps and rugged landscapes in the RNA virus phi6. Burch, C.L., Chao, L. (1999). Genetics 151:921-927. Fisher's geometric model of adaptive evolution argues that adaptive evolution should generally result from the substitution of many mutations of small effect because advantageous mutations of small effect should be more common than those of large effect. However, evidence for both evolution by small steps and for Fisher's model has been mixed. Here we report supporting results from a new experimental test of the model. We subjected the bacteriophage phi6 to intensified genetic drift in small populations and caused viral fitness to decline through the accumulation of a deleterious mutation. We then propagated the mutated virus at a range of larger population sizes and allowed fitness to recover by natural selection. Although fitness declined in one large step, it was usually recovered in smaller steps. More importantly, step size during recovery was smaller with decreasing size of the recovery population. These results confirm Fisher's main prediction that advantageous mutations of small effect should be more common. We also show that the advantageous mutations of small effect are compensatory mutations whose advantage is conditional (epistatic) on the presence of the deleterious mutation, in which case the adaptive landscape of phi6 is likely to be very rugged. [TOP OF PAGE]

76. Phage therapy: past history and future prospects. Carlton, R.M. (1999). Archivum Immunologii et Therapiae Experimentalis 47:267-274. Bacterial viruses (bacteriophages, also called "phages") can be robust antibacterial agents in vitro. However, their use as therapeutic agents, during a number of trials from the 1920s to the 1950s, was greatly handicapped by a number of factors. In part, there were certain limitations inherent in phage physiology (e. g. narrow host range, and rapid clearance from the body); in part there were technological limitations in the era (e.g. lysogeny not yet discovered); but the greatest limitation was the highly inadequate scientific methodologies used by practitioners at the time (e.g., their failure to conduct placebo-controlled studies, to remove endotoxins from the preparations, and to re-confirm phage viability after adding sterilizing agents to the preparations). In recent years, well-controlled animal models have demonstrated that phages can rescue animals from a variety of fatal infections, while non-controlled clinical reports published in Eastern Europe have shown that phages can be effective in treating drug-resistant infections in humans. This encouraging data, combined with the fact that drug-resistant bacteria have become a global crisis, have created a window of opportunity for phage therapy to be tested anew, this time using modem technologies and placebo-controlled designs. If successful, it can be used as a stand-alone therapy when bacteria are fully resistant to antibiotics, and as a valuable adjunct to antibiotics when the bacteria are still susceptible. [TOP OF PAGE]

77. Comparative sequence analysis of the DNA packaging, head, and tail morphogenesis modules in the temperate cos-site Streptococcus thermophilus bacteriophage Sfi21. Desiere, F., Lucchini S, Brussow, H. (1999). Virology 260:244-253. The temperate Streptococcus thermophilus bacteriophage Sfi21 possesses 15-nucleotide-long cohesive ends with a 3' overhang that reconstitutes a cos-site with twofold hyphenated rotational symmetry, Over the DNA packaging, head and tail morphogenesis modules, the Sfi21 sequence predicts a gene map that is strikingly similar to that of lambdoid coliphages in the absence of any sequence similarity. A nearly one to one gene correlation was found with the phage lambda genes Nul to H, except for gene B-to-E complex, where the Sfi21 map resembled that of coliphage HK97. The similarity between Sfi21 and HK97 was striking: both major head proteins showed an N-terminal coiled-coil structure, the mature major head proteins started at amino acid positions 105 and 104, respectively, and both major head genes were preceded by genes encoding a possible protease and portal protein. The purported Sfi21 protease is the first viral member of the CipP protease family The prediction of Sfi21 gene functions by reference to the gene map of intensively investigated coliphages was experimentally confirmed for the major head and tail gene. Phage Sfi21 shows nucleotide sequence similarity with Lactococcus phage BK5-T and a lactococcal prophage and amino acid sequence similarity with the Lactobacillus phage A2 and the Staphylococcus phage PVL. PVL is a missing link that connects the portal proteins from Sfi21 and HK97 with respect to sequence similarity. These observations and database searches, which demonstrate sequence similarity between proteins of phage from gram-positive bacteria, proteobacteria, and Archaea, constrain models of phage evolution. [TOP OF PAGE]

78. Causative agents of bacterial mortality and the consequences to marine food webs. Fuhrman, J.A., Noble, R.T. (1999). p. ???-??? In Bell, C.R., Brylinsky, M., and Johnson-Green, P. (eds.), Microbial Biosystems: New Frontiers. Proc 8th Int Symp Microb. Ecol. Atlantic Canada Society for Microbial Ecology, Halifax, Canada. [TOP OF PAGE]

79. Marine viruses and their biogeochemical and ecological effects. Fuhrman, J.A. (1999). Nature 399:541-548. Viruses are the most common biological agents in the sea, typically numbering ten billion per litre. They probably infect all organisms, can undergo rapid decay and replenishment, and influence many biogeochemical and ecological processes, including nutrient cycling, system respiration, particle size-distributions and sinking rates, bacterial and algal biodiversity and species distributions, algal bloom control, dimethyl sulphide formation and genetic transfer. Newly developed fluorescence and molecular techniques leave the field poised to make significant advances towards evaluating and quantifying such efforts. [TOP OF PAGE]

80. Impact of viruses on planktonic bacteria. Fuhrman, J.A. (1999). p. ???-??? In Kirchman, D.L. (ed.), Microbial Ecology of the Oceans. Wiley & Sons, ??? [TOP OF PAGE]

81. Biocontrol of Erwinia amylovora using bacteriophage. Gill, J.J., Svircev, A.M., Myers, A.L., Castle, A.J. (1999). Phytopathology 89:S27 [TOP OF PAGE]

82. Development and characterization of a fluorescent-bacteriophage assay for detection of Escherichia coli O157:H7. Goodridge, L., Chen, J., Griffiths, M. (1999). Appl. Environ. Microbiol. 65:1397-1404. In this paper we describe evaluation and characterization of a novel assay that combines immunomagnetic separation and a fluorescently stained bacteriophage for detection of Escherichia coli O157:H7 in broth. When it was combined with flow cytometry, the fluorescent-bacteriophage assay (FBA) was capable of detecting 10(4) cells/ml. A modified direct epifluorescent-filter technique (DEFT) was employed in an attempt to estimate bacterial concentrations. Using regression analysis, we calculated that the lower detection limit was between 10(2) and 10(3) cells/ml; however, the modified DEFT was found to be an unreliable method for determining bacterial concentrations. The results of this study show that the FBA, when combined with flow cytometry, is a sensitive technique for presumptive detection of E. coli O157:H7 in broth cultures. [TOP OF PAGE]

83. The use of a fluorescent bacteriophage assay for detection of Escherichia coli O157:H7 in inoculated ground beef and raw milk. Goodridge, L., Chen, J., Griffiths, M. (1999). Int. J. Food Microbiol. 47:43-50. The objective of this study was to develop a fluorescent bacteriophage assay (FBA) for the detection of E. coli O157:H7 in ground beef and raw milk. The FBA is a two step assay that combines immunomagnetic separation, to separate the target organism from mixed culture, with a highly specific fluorescently stained bacteriophage to label the E. coli O157:H7 cells. When used in conjunction with flow cytometry, the FBA was able to detect 2.2 CFU/g of artificially contaminated ground beef following a 6 h enrichment. Between 10(1) and 10(2) CFU/ml of artificially contaminated raw milk were detectable after a 10 h enrichment step. The results show that the FBA is potentially useful as a rapid technique for the preliminary detection of E. coli O157:H7 in food. [TOP OF PAGE]

84. Comparative genomics of Streptococcus thermophilus phage species supports a modular evolution theory. Lucchini S., Desiere, F., Brussow, H. (1999). J. Virol. 73:8647-8656. The comparative analysis of five completely sequenced Streptococcus thermophilus bacteriophage genomes demonstrated that their diversification was achieved by a combination of DNA recombination events and an accumulation of point mutations. The five phages included lytic and temperate phages, both pac site and cos site, from three distinct geographical areas. The units of genetic exchange were either large, comprising the entire morphogenesis gene cluster, excluding the putative tail fiber genes, or small, consisting of one or maximally two genes or even segments of a gene. Many indels were flanked by DNA repeats. Differences in a single putative tail fiber gene correlated with the host ranges of the phages. The predicted tail fiber protein consisted of highly conserved domains containing conspicuous glycine repeats interspersed with highly variable domains. As in the T-even coliphage adhesins, the glycine-containing domains were recombinational hot spots. Downstream of a highly conserved DNA replication region, all lytic phages showed a short duplication; in three isolates the origin of replication was repeated. The lytic phages could conceivably be derived from the temperate phages by deletion and multiple rearrangement events in the lysogeny module, giving rise to occasional selfish phages that defy the superinfection control systems of the corresponding temperate phages. [TOP OF PAGE]

85. Similarly organized lysogeny modules in temperate Siphoviridae from low GC content gram-positive bacteria. Lucchini, S., Desiere, F., Brussow, H. (1999). Virology 263:427-435. Temperate Siphoviridae from an evolutionarily related branch of low GC content gram-positive bacteria share a common genetic organization of lysogeny-related genes and the predicted proteins are linked by many sequence similarities. Their compact lysogeny modules [integrase/1-2 orfs (phage exclusion? and metalloproteinase motif proteins)/cl-like repressor/cro-like repressor/antirepressor (optional)] differ clearly from that of h-like and L5-like viruses, the two currently established genera of temperate Siphoviridae, while they resemble those of the P2-like genus of Myoviridae. In all known temperate Siphoviridae from low GC content gram-positive bacteria the lysogeny module is flanked by the lysis module and the DNA replication module. This modular organization is again distinct from that of the known genera of temperate Siphoviridae. On the basis of comparative sequence analysis we propose a new genus of Siphoviridae: "Sfi21-like" phages. With a larger database of phage sequences it might be possible to establish a genomics-based phage taxonomy and to retrace the evolutionary history of selected phage modules or individual phage genes. The antirepressor of Sfi21-like phages has an unusual widespread distribution since proteins with high aa similarity (40%) were found not only in phages from gramnegative bacteria, but also in insect viruses. [TOP OF PAGE]

86. The genetic relationship between virulent and temperate Streptococcus thermophilus bacteriophages: Whole genome comparison of cos-site phages Sfi19 and Sfi21. Lucchini, S., Desiere, F., Brussow, H. (1999). Virology 260:232-243. The virulent cos-site Streptococcus thermophilus bacteriophage Sfi19 has a 37,392-bp-long genome consisting of 44 open reading frames all encoded on the same DNA strand. The genome of the temperate cos-site S. thermophilus phage Sfi21 is 3.3 kb longer (40,740 bp, 53 orfs). Both genomes are very similarly organized and differed mainly by gene deletion and DNA rearrangement events in the lysogeny module; gene replacement, duplication, and deletion events in the DNA replication module, and numerous point mutations. The level of point mutations varied from 15% (DNA packaging and head morphogenesis modules). A dotplot analysis showed nearly a straight line over the left 25 kb of their genomes. Over the right genome half, a more variable dotplot pattern was observed. The entire lysogeny module from Sfi21 comprising 12 genes was replaced by 7 orfs in Sfi19, six showed similarity with genes from temperate pac-site S. thermophilus phages. Noneof the genes implicated in the establishment of the lysogenic state (integrase, superinfection immunity, repressor) or remnants of it were conserved in Sfi19, while a Cro-like repressor was detected. Downstream of the highly conserved DNA replication module 11 and 13 orfs were found in Sfi19 and phiSfi21, respectively: Two orfs from Sfi21 were replaced by a different gene and a duplication of the phage origin of replication in Sfi19; a further orf was only found in Sfi21. All other orfs from this region, which included a second putative phage repressor, were closely related between both phages. Two noncoding regions of Sfi19 showed sequence similarity to pST1, a small cryptic plasmid of S. thermophilus. [TOP OF PAGE]

87. Virulence evolution in a virus obeys a trade-off. Messenger, S.L., Molineux, I.J., Bull, J.J. (1999). Proc. R. Soc. Lond. B 266:297-404. The evolution of virulence was studied in a virus subjected to alternating episodes of vertical and horizontal transmission. Bacteriophage f1 was used as the parasite because it establishes a debilitating but non-fatal infection that can be transmitted vertically (from a host to its progeny) as well as horizontally (infection of new hosts). Horizontal transmission was required of all phage at specific intervals, but was prevented otherwise. Each episode of horizontal transmission was followed by an interval of obligate vertical transmission, followed by an interval of obligate horizontal transmission etc. The duration of vertical transmission was eight times longer per episode in one treatment than in the other, thus varying the relative intensity of selection against virulence while maintaining selection for some level of virus production. Viral lines with the higher enforced rate of infectious transmission evolved higher virulence and higher rates of virus production. These results support the trade-off model for the evolution of virulence. [TOP OF PAGE]

88. [Natural interspecific hybrids of transposable phages of Pseudomonas aeruginosa]. Mit'kina, L.N., Krylov, V.N. (1999). Genetika 35:1182-1190. Bacterial viruses of Pseudomonas aeruginosa assigned to two groups, D3112 and B3, recombine with very low frequencies. Previous study of the genome structure of intergroup hybrids suggested the incompatibility of some genetic modules of these bacteriophages. In this work, several natural hybrid transposable phages that had the genomes largely consisting of modules of phages from group D3112 and B3, were described. The discovery of these phages suggests the continuous genetic exchange in nature of these viruses belonging to different species. This model is considered as promising from the viewpoint of monitoring virus evolution. [TOP OF PAGE]

89. Applications of phage resistance in lactic acid bacteria. Moineau, S. (1999). Antonie van Leeuwenhoek 76:377-382. [TOP OF PAGE]

90. Breakdown and microbial uptake of marine viruses and other lysis products. Noble, R.T., Fuhrman, J.A. (1999). Aquat. Microb. Ecol. 20:1-11. To understand the roles of marine viruses in marine microbial food webs, it is important to determine rates and mechanisms of virus degradation and subsequent uptake of degraded virus material and other cell lysis products by heterotrophic marine bacteria. We radiolabeled and concentrated viruses and viral lysis products from either pure cultures (³H) or natural communities (³H and ³³P) and added them to seawater samples of differing trophic status from coastal (mesotrophic) and offshore (oligotrophic) California waters and French Mediterranean waters (oligotrophic). Rates of degradation were determined by the loss of high molecular weight radiolabel over time and the fate of the degraded material (microbial uptake or accumulation in low molecular weight pools) was followed by size fractionation and/or acid extraction. Preliminary experiments with ³H-labeled, single-stranded RNA phage MS2 and marine phage H11/1 demonstrated that MS2 degraded significantly faster in coastal Santa Monica Bay seawater (2.5 ± 0.6% h^-1), than the marine phage, H11/1 (0.99 ± 0.1% h^-1). For labeled virus material from natural populations, rates of degradation were slower in oligotrophic waters (ranges from 1.0 to 3.3% h^-1) than in mesotrophic waters (ranges from 4.9 to 6.0% h^-1), corresponding to turnover rates of 1 to 4 d for this material. Degradation rates of labeled virus material are likely underestimates, because during preparation, degradation and uptake are continually occurring, resulting in accumulation of the less reactive products. The proportion of radiolabeled material taken up by microbes was greatest in oligotrophic waters, especially in the phosphate-limited Villefranche Bay, France, where most of the ³³PO₄-labeled material was taken up in less than 7 h. In contrast, the majority of degraded ³H-labeled material was not accumulated into biomass, and in 3 of 4 samples, accumulation was hardly detectable. The results suggest that viruses and lysis products are labile and turn over relatively rapidly, but often may not be efficiently incorporated into bacterial biomass. [TOP OF PAGE]

91. The effects of viral enrichment on the mortality and growth of heterotrophic bacterioplankton. Noble, R.T., Middelboe, M., Fuhrman, J.A. (1999). Aquat. Microb. Ecol. 18:1-13. [TOP OF PAGE]

92. Microbial gene transfer: an ecological perspective. Paul, J.H. (1999). J. Mol. Microbiol. Biotechnol. 1:45-50. Microbial gene transfer or microbial sex is a means of exchanging loci amongst prokaryotes and certain eukaryotes. Historically viewed as a laboratory artifact, recent evidence from natural populations as well as genome research has indicated that this process may be a major driving force in microbial evolution. Studies with natural populations have taken two approaches-either adding a defined donor with a traceable gene to an indigenous community, and detecting the target gene in the indigenous bacteria, or by adding a model recipient to capture genes being transferred from the ambient microbial flora. However, both approaches usually require some cultivation of the recipient, which may result in a dramatic underestimation of the ambient transfer frequency. Novel methods are just evolving to study in situ gene transfer processes, including the use of green fluorescent protein (GFP)-marked plasmids, which enable detection of transferrants by epifluorescence microscopy. A transduction-like mechanism of transfer from viral-like particles produced by marine bacteria and thermal spring bacteria to Escherichia coli has been documented recently, indicating that broad host range transduction may be occurring in aquatic environments. The sequencing of complete microbial genomes has shown that they are a mosaic of ancestral chromosomal genes interspersed with recently transferred operons that encode peripheral functions. Archaeal genomes indicate that the genes for replication, transcription, and translation are all eukaryotic in complexity, while the genes for intermediary metabolism are purely bacterial. And in eukaryotes, many ancestral eukaryotic genes have been replaced by bacterial genes believed derived from food sources. Collectively these results indicate that microbial sex can result in the dispersal of loci in contemporary microbial populations as well as having shaped the phylogenies of microbes from multiple, very early gene transfer events. [see http://www.jmmb.net/v1n1/07/07.html for full-text entry]. [TOP OF PAGE]

93. A novel virus family, the Rudiviridae: Structure, virus-host interactions and genome variability of the sulfolobus viruses SIRV1 and SIRV2. Prangishvili, D., Arnold, H.P., Gotz, D., Ziese, U., Holz, I., Kristjansson, J.K., Zillig, W. (1999). Genetics 152:1387-1396. The unenveloped, stiff-rod-shaped, linear double-stranded DNA viruses SIRV1 and SIRV2 from Icelandic Sulfolobus isolates form a novel virus family, the Rudiviridae. The sizes of the genomes are 32. 3 kbp for SIRV1 and 35.8 kbp for SIRV2. The virions consist of a tube-like superhelix formed by the DNA and a single basic 15.8-kD DNA-binding protein. The tube carries a plug and three tail fibers at each end. One turn of the DNA-protein superhelix measures 4.3 nm and comprises 16.5 turns of B DNA. The linear DNA molecules appear to have covalently closed hairpin ends. The viruses are not lytic and are present in their original hosts in carrier states. Both viruses are quite stable in these carrier states. In several laboratory hosts SIRV2 was invariant, but SIRV1 formed many different variants that completely replaced the wild-type virus. Some of these variants were still variable, whereas others were stable. Up to 10% nucleotide substitution was found between corresponding genome fragments of three variants. Some variants showed deletions. Wild-type SIRV1, but not SIRV2, induces an SOS-like response in Sulfolobus. We propose that wild-type SIRV1 is unable to propagate in some hosts but surmounts this host range barrier by inducing a host response effecting extensive variation of the viral genome. [TOP OF PAGE]

94. A high incidence of prophage carriage among natural isolates of Streptococcus pneumoniae. Ramirez, M., Severina, E., Tomasz, A. (1999). J. Bacteriol. 181:3618-3625. The majority (591 of 791, or 76%) of Streptococcus pneumoniae clinical isolates examined showed the presence of two or more chromosomal SmaI fragments that hybridized with the lytA-specific DNA probe. Only one of these fragments, frequently having an approximate molecular size of 90 kb, was shown to carry the genetic determinant of the pneumococcal autolysin (N-acetylmuramic acid-L-alanine amidase). Strains carrying multiple copies of lytA homologues included both antibiotic-susceptible and -resistant isolates as well as a number of different serotypes and strains recovered from geographic sites on three continents. Mitomycin C treatment of strains carrying several lytA-hybridizing fragments caused the appearance of extrachromosomal DNA hybridizing to the lytA gene, followed by lysis of the bacteria. Such lysates contained phage particles detectable by electron microscopy. The findings suggest that the lytA-hybridizing fragments in excess of the host lytA represent components of pneumococcal bacteriophages. The high proportion of clinical isolates carrying multiple copies of lytA indicates the widespread occurrence of lysogeny, which may contribute to genetic variation in natural populations of pneumococci. [TOP OF PAGE]

95. Prophage carriage as a molecular epidemiological marker in Streptococcus pneumoniae. Severina, E., Ramirez, M., Tomasz, A. (1999). Journal of Clinical Microbiology 37:3308-3315. The great majority of clinical isolates of Streptococcus pneumoniae carry prophages that may be identified through their hybridization with a DNA probe specific for the pneumococcal lytA gene (M. Ramirez, E. Severina, and A. Tomasz, J. Bacteriol. 181:3618-3625, 1999). We now show that the lytA hybridization pattern of chromosomal SmaI digests is stable for a given strain during extensive serial culturing in the laboratory; the pattern is specific for the strain's clonal type, as defined by pulsed-field gel electrophoretic (PFGE) pattern, and variations in PFGE subtypes may be explained by changes in the number and chromosomal localization of this prophage(s). These observations indicate that the lytA hybridization pattern may be used as a molecular epidemiological marker that offers additional resolution of the genetic background of S. pneumoniae strains. [TOP OF PAGE]

96. Use of the polymerase chain reaction and denaturing gradient gel electrophoresis to study diversity in natural virus communities. Short, S.M., Suttle, C.A. (1999). Hydrobiologia 401:19-32. Viruses are abundant members of marine and freshwater microbial communities, and are important players in aquatic ecology and geochemical cycles. Recent methodological developments have allowed the use of the polymerase chain reaction (PCR) to examine the diversity of natural communities of viruses without the need for culture. DNA polymerase genes are highly conserved and are, therefore, suitable targets for PCR analysis of microbes that do not encode rRNA. As natural virus communities are largely made up of dsDNA viruses, and as many dsDNA algal viruses encode their own DNA polymerase, PCR primers can be designed to amplify fragments of these genes. This approach has been used to examine the genetic diversity in natural communities of viruses that infect phytoplankton. Algal-virus-specific primers were used to amplify polymerase fragments from natural virus samples, demonstrating the presence of a diverse community of viruses closely related to those that are known to infect phytoplankton. We have modified this approach by using denaturing gradient gel electrophoresis (DGGE) to rapidly analyze PCR products. DGGE will permit rapid and efficient fingerprinting of natural marine viral communities, and allow spatial and temporal differences in viral community structure to be examined. This paper provides a brief overview of how PCR and DGGE can be used to examine diversity in natural viral communities drawing on viruses that infect phytoplankton as an example. [TOP OF PAGE]

97. An in situ enclosure experiment to test the solar UVB impact on plankton in a high altitude mountain lake: II) effects on the microbial food web. Sommaruga, R., Sattller, B., Oberleiter, A., Wille, A., Sommaruga-Wögrath, S., Psenner, R., Felip, M., Camarero, L., Pina, S., Gironés, R., Catalán, J. (1999). Journal of Plankton Research 21:859-879. We studied the impact of ambient levels of solar UVB radiation on the planktonic microbial food web (viruses, heterotrophic bacteria, heterotrophic flagellates and ciliates) of a high-mountain lake (2417 m above sea level) under in situ conditions for 16 days. Enclosures of 1 m3 receiving either the full sunlight spectrum or sunlight without UVB radiation were suspended at the lake surface. We found that the abundance of heterotrophic flagellates was always lower in the +UVB treatment than in the -UVB one. In addition, bacterial consumption, measured by the disappearance of fluorescently labelled bacteria, was significantly (p<0.05) reduced in the +UVB treatment. The abundance of non-filamentous bacteria (<10 µm long) was also lower in the +UVB treatment, suggesting a direct effect of UVB on their growth. This was supported by the significantly (p<0.05) lower cell-specific activity ([3H]-thymidine incorporation) found on the fifth day of the experiment. In contrast, UVB radiation had no effect on filamentous bacteria (>10 µm long) that represented only a small fraction of the total abundance (<4%) but up to ~70 % of the total bacterial biovolume. Ciliates, mainly Urotricha pelagica and U. furcata, were less impacted by UVB radiation although the net growth rate during the first week of the experiment was lower in the +UVB treatment than in the -UVB one (0.22 and 0.39 d-1, respectively). The abundance of virus-like particles during the first week of the experiment was higher in the -UVB treatment. After reaching the maximum value for the interaction viruses x bacteria, their number decreased dramatically (by ~85%) in both treatments with a decay rate of ~0.017 h-1. This study illustrates the complexity in assessing the impact of UVB radiation when more than one trophic level is considered and indicates the existence of different sensitivity to UVB radiation among components of the microbial food web. [TOP OF PAGE]

98. Do viruses control the oceans? Suttle, C.A. (1999). Natural History 108:48-51. [TOP OF PAGE]

99. Cyanophage. Suttle, C.A. (1999). p. ???-??? In Whitton, B. and Potts, M. (eds.), Ecology of Cyanobacteria: Their Diversity in Time and Spac. ???, ??? [TOP OF PAGE]

100. Hybrid frequencies confirm limit to coinfection in the RNA bacteriophage phi-6. Turner, P.E., Burch, C.L., Hanley, K., Chao, L. (1999). J. Virol. 73:2420-2424. Coinfection of the same host cell by multiple viruses may lead to increased competition for limited cellular resources, thus reducing the fitness of an individual virus. Selection should favor viruses that can limit or prevent coinfection, and it is not surprising that many viruses have evolved mechanisms to do so. Here we explore whether coinfection is limited in the RNA bacteriophage phi6 that infects Pseudomonas phaseolicola. We estimated the limit to coinfection in phi6 by comparing the frequency of hybrids produced by two marked phage strains to that predicted by a mathematical model based on differing limits to coinfection. Our results provide an alternative method for estimating the limit to coinfection and confirm a previous estimate between two to three phages per host cell. In addition, our data reveal that the rate of coinfection at low phage densities may exceed that expected through random Poisson sampling. We discuss whether phage phi6 has evolved an optimal limit that balances the costly and beneficial fitness effects associated with multiple infections. [TOP OF PAGE]

101. Prisoner's dilemma in an RNA virus. Turner, P.E., Chao, L. (1999). Nature 398:441-443. The evolution of competitive interactions among viruses was studied in the RNA phage phi6 at high and low multiplicities of infection (that is, at high and low ratios of infecting phage to host cells). At high multiplicities, many phage infect and reproduce in the same host cell, whereas at low multiplicities the viruses reproduce mainly as clones. An unexpected result of this study was that phage grown at high rates of co-infection increased in fitness initially, but then evolved lowered fitness. Here we show that the fitness of the high-multiplicity phage relative to their ancestors generates a pay-off matrix conforming to the prisoner's dilemma strategy of game theory. In this strategy, defection (selfishness) evolves, despite the greater fitness pay-off that would result if all players were to cooperate. Viral cooperation and defection can be defined as, respectably, the manufacturing and sequestering of diffusible (shared) intracellular products. Because the low-multiplicity phage did not evolve lowered fitness, we attribute the evolution of selfishness to the lack of clonal structure and the mixing of unrelated genotypes at high multiplicity. [TOP OF PAGE]

102. Sunlight-induced DNA damage and resistance in natural viral communities. Weinbauer, M.G., Wilhelm, S.W., Suttle, C.A., Pledger, R.J., Mitchell, D.L. (1999). Aquat. Microb. Ecol. 17:111-120. Using a highly specific radioimmunoassay, the sunlight-induced formation of cyclobutane pyrimidine dimers (CPDs) and pyrimidine (6-4) pyrimidone photoproducts ([6-4] PPs) in viral DNA was investigated for natural virus communities in offshore and coastal waters of the western Gulf of Mexico as well as for clonal viral isolates. Concentrations of (6-4) PPs were consistently lower than CPD concentrations, and ranged from 1.5 to 17.0% of total measured photodamage. The accumulation of photoproducts varied among the natural viral community, the marine Vibrio phage PWH3a-P1 and the Synechococcus sp. DC2 (WH7803) cyanophage SYN-M3, which were deployed in situ from dawn until dark. Natural viral communities were more resistant to DNA damage than the cyanophage isolate SYN-M3, which was more resistant to damage than bacteriophage PWH3a-P1. Moreover, depth profiles revealed that photodamage in viral isolates deployed in the water column accumulated more rapidly at offshore stations than at coastal stations. In natural virus communities collected from offshore surface waters, photodamage accumulated during the solar day with maximum damage occurring between 15:00 and 18:00 h. Depth profiles obtained during calm seas showed that photodamage concentrations were high in surface waters at the offshore stations and at 1 coastal station. Results at other coastal stations undergoing significant mixing demonstrated no photoproduct accumulations. Results demonstrate that natural virus communities were more tolerant to DNA damaging radiation than the laboratory isolates used in this study. Consequently, laboratory isolates can be poor proxies for UV impacts on natural viral communities. [TOP OF PAGE]

103. Lysogeny and prophage induction in coastal and offshore bacterial communities. Weinbauer, M.G., Suttle, C.A. (1999). Aquat. Microb. Ecol. 18:217-225. The influence of solar radiation and hydrogen peroxide on induction of lysogens, and the resulting effect on bacteriophage production and bacterial mortality was investigated for coastal and oceanic marine bacterial communities at 6 stations in the western Gulf of Mexico. The percentage of lysogenic cells induced by mitomycin C was also determined. Solar radiation and hydrogen peroxide were not as effective as mitomycin C at inducing phage production. The burst size of cells induced by mitomycin C was estimated by transmission electron microscopy, assuming that cells completely filled with viral particles were on the verge of bursting. The smallest estimates of burst size were associated with oligotrophic oceanic stations and ranged from 15 to 28 viruses produced per lytic event, while in more productive coastal waters the estimated burst sizes ranged from 33 to 64. The mitomycin C-induced phage production and burst size were used to estimate the number of lysogenic bacterial cells. On average, the percentage of inducible lysogens was higher at offshore (1.5 to 11.4%) than at coastal (0.8 to 2.2%) stations. However, with the exception of 1 station, less than 5% of the bacteria could be induced to produce phage, suggesting that lysogens only occasionally comprised a significant component of these bacterial communities. The proportion of lysogens that could be induced by sunlight, relative to those that could be induced by mitomycin C, was lower at oceanic than coastal stations. This implies that prophages in optically transparent offshore waters were more resistant to induction by solar radiation, or that most lysogens that could be triggered by sunlight were already induced. Based on a steady-state model, induction of lysogenic bacteria by solar radiation or hydrogen peroxide could result in between 0 and 3.5% or 0.9 and 3.4% of the total bacterial mortality, respectively. Our results imply that solar radiation and hydrogen peroxide induced lysogenic phage production were not an important source of phage production or bacterial mortality in offshore or coastal waters of the western Gulf of Mexico. [TOP OF PAGE]

104. Resistance to DNA damage in natural viral communities from the Gulf of Mexico. Weinbauer, M.G., Wilhelm, S.W., Pledger, R.J., Mitchell, D.L., Suttle, C.A. (1999). Aquat. Microb. Ecol. 17:111-120. Using a highly specific radioimmunoassay, the sunlight-induced formation of cyclobutane pyrimidine dimers (CPDs) and pyrimidine (6-4) pyrimidone photoproducts ([6-4] PPs) in viral DNA was investigated for natural virus communities in offshore and coastal waters of the western Gulf of Mexico as well as for clonal viral isolates. Concentrations of (6-4) PPs were consistently lower than CPD concentrations, and ranged from 1.5 to 17.0% of total measured photodamage. The accumulation of photoproducts varied among the natural viral community, the marine Vibrio phage PWH3a-P1 and the Synechococcus sp. DC2 (WH7803) cyanophage SYN-M3, which were deployed in situ from dawn until dark. Natural viral communities were more resistant to DNA damage than the cyanophage isolate SYN-M3, which was more resistant to damage than bacteriophage PWH3a-P1. Moreover, depth profiles revealed that photodamage in viral isolates deployed in the water column accumulated more rapidly at offshore stations than at coastal stations. In natural virus communities collected from offshore surface waters, photodamage accumulated during the solar day with maximum damage occurring between 15:00 and 18:00 h. Depth profiles obtained during calm seas showed that photodamageconcentrations were high in surface waters at the offshore stations and at 1 coastal station. Results at other coastal stations undergoing significant mixing demonstrated no photoproduct accumulations. Results demonstrate that natural virus communities were more tolerant to DNA damaging radiation than the laboratory isolates used in this study. Consequently, laboratory isolates can be poor proxies for UV impacts on natural viral communities. [TOP OF PAGE]

105. Different trajectories of parallel evolution during viral adaptation. Wichman, H.A., Badgett, M.R., Scott, L.A., Boulianne, C.M., Bull, J.J. (1999). Science 285:422-424. [TOP OF PAGE]

106. Viruses and nutrient cycles in the sea. Wilhelm, S.W., Suttle, C.A. (1999). BioScience 49:781-788. Viruses play critical roles in the structure and function of aquatic food webs. [TOP OF PAGE]

107. Analysis of cyanophage diversity and population structure in a south-north transect of the Atlantic ocean. Wilson, W.H., Fuller, N.J., Joint, I.R., Mann, N.H. (1999). Bulletin de l'Institut Oceanographique (Monaco). 0:209-216. Cyanophages (viruses which infect cyanobacteria) are abundant in the marine environment and are thought to be a significant factor in determining the dynamics of Synechococcus spp. populations. In an effort to use molecular techniques to characterise cyanophage populations, we designed cyanophage-specific (CPS) PCR primers based on a gene found in three genetically distinct marine cyanophages (Fuller et al., 1998). CPS primers were used to amplify cyanophage DNA extracted from viral communities concentrated from sea-water samples obtained during a cruise transect between the Falkland Islands, in the south Atlantic ocean, to the UK. Following phylogenetic analysis of cloned and sequenced PCR products, it was revealed that genetic diversity of marine cyanophage clones within a single water sample was as great as clones and cyanophage isolates collected between different oceans. Denaturing gradient gel electrophoresis (DGGE) analysis confirmed this high diversity. DGGE analysis also revealed changes in cyanophage population structure in surface seawater over the south-north transect and throughout depth profiles in the water column. Maximum Synechococcus spp. concentrations, in a stratified water column, correlated with maximum cyanophage diversity. [TOP OF PAGE]

108. Hybridization analysis of Chesapeake Bay Virioplankton. Wommack, K.E., Ravel, J., Hill, R.T., Colwell, R.R. (1999). Appl. Environ. Microbiol. 65:241-250. It has been hypothesized that, by specifically lysing numerically dominant host strains, the virioplankton may play a role in maintaining clonal diversity of heterotrophic bacteria and phytoplankton populations. If viruses selectively lyse only those host species that are numerically dominant, then the number of a specific virus within the virioplankton would be expected to change dramatically over time and space, in coordination with changes in abundance of the host. In this study, the abundances of specific viruses in Chesapeake Bay water samples were monitored, using nucleic acid probes and hybridization analysis. Total virioplankton in a water sample was separated by pulsed-field gel electrophoresis and hybridized with nucleic acid probes specific to either single viral strains or a group of viruses with similar genome sizes. The abundances of specific viruses were inferred from the intensity of the hybridization signal. By using this technique, a virus comprising 1/1,000 of the total virioplankton abundance (ca. 10(4) PFU/ml) could be detected. Titers of either a single virus species or a group of viruses changed over time, increasing to peak abundance and then declining to low or undetectable levels, and were geographically localized in the bay. Peak signal intensities, i.e., peak abundances of virus strains, were 10-fold greater than the low background level. Furthermore, virus species were found to be restricted to a particular depth, since probes specific to viruses from bottom water did not hybridize with virus genomes from surface water at the same geographical location. Overall, changes in abundances of specific viruses within the virioplankton were episodic, supporting the hypothesis that viral infection influences, if not controls, clonal diversity within heterotrophic bacteria and phytoplankton communities. [TOP OF PAGE]

109. Population dynamics of Chesapeake Bay virioplankton: total-community analysis by pulsed-field gel electrophoresis. Wommack, K.E., Ravel, J., Hill, R.T., Colwell, R.R. (1999). Appl. Environ. Microbiol. 65:231-240. It has been hypothesized that, by specifically lysing numerically dominant host strains, the virioplankton may play a role in maintaining clonal diversity of heterotrophic bacteria and phytoplankton populations. If viruses selectively lyse only those host species that are numerically dominant, then the number of a specific virus within the virioplankton would be expected to change dramatically over time and space, in coordination with changes in abundance of the host. In this study, the abundances of specific viruses in Chesapeake Bay water samples were monitored, using nucleic acid probes and hybridization analysis. Total virioplankton in a water sample was separated by pulsed-field gel electrophoresis and hybridized with nucleic acid probes specific to either single viral strains or a group of viruses with similar genome sizes. The abundances of specific viruses were inferred from the intensity of the hybridization signal. By using this technique, a virus comprising 1/1,000 of the total virioplankton abundance (ca. 10(4) PFU/ml) could be detected. Titers of either a single virus species or a group of viruses changed over time, increasing to peak abundance and then declining to low or undetectable levels, and were geographically localized in the bay. Peak signal intensities, i.e., peak abundances of virus strains, were 10-fold greater than the low background level. Furthermore, virus species were found to be restricted to a particular depth, since probes specific to viruses from bottom water did not hybridize with virus genomes from surface water at the same geographical location. Overall, changes in abundances of specific viruses within the virioplankton were episodic, supporting the hypothesis that viral infection influences, if not controls, clonal diversity within heterotrophic bacteria and phytoplankton communities. [TOP OF PAGE]

110. Amplification and spread of viruses in a growing plaque. You, L., Yin, J. (1999). J. Theor. Biol. 200:365-373. The two-dimensional propagation of viruses through a "lawn" of receptive hosts, commonly called plaque growth, reflects the dynamics of interactions between viruses and host cells. Here we treat the amplification of viruses during plaque growth as a reaction-diffusion system, where interactions among the virus, uninfected host cells, and virus-producing host-virus complexes are accounted for using rates of viral adsorption to and desorption from the host-cell surface, rates of reproduction and release of progeny viruses by lysis of the host, and by the coupling of these reactions with diffusion of free virus within the agar support. Numerical solution of the system shows the development of a traveling wave of reproducing viruses, where the velocity of the wave is governed by the kinetic and diffusion parameters. The model has been applied to predict the propagation velocity of a bacteriophage plaque. Different mechanisms may account for the dependence of this velocity on the host density during early stages of a growing plaque. The model provides a means to explore how changes in the virus-host interactions may be manifest in a growing plaque. [TOP OF PAGE]

111. Tailed bacteriophages: the order caudovirales. Ackermann, H.-W. (1998). Advances in Virus Research 51:135-201. Tailed bacteriophages have a common origin and constitute an order with three families, named Caudovirales. Their structured tail is unique. Tailed phages share a series of high-level taxonomic properties and show many facultative features that are unique or rare in viruses, for example, tail appendages and unusual bases. They share with other viruses, especially herpesviruses, elements of morphogenesis and life-style that are attributed to convergent evolution. Tailed phages present three types of lysogeny, exemplified by phages lambda, Mu, and P1. Lysogeny appears as a secondary property acquired by horizontal gene transfer. Amino acid sequence alignments (notably of DNA polymerases, integrases, and peptidoglycan hydrolases) indicate frequent events of horizontal gene transfer in tailed phages. Common capsid and tail proteins have not been detected. Tailed phages possibly evolved from small protein shells with a few genes sufficient for some basal level of productive infection. This early stage can no longer be traced. At one point, this precursor phage became perfected. Some of its features were perfect enough to be transmitted until today. It is tempting to list major present-day properties of tailed phages in the past tense to construct a tentative history of these viruses: 1. Tailed phages originated in the early Precambrian, long before eukaryotes and their viruses. 2. The ur-tailed phage, already a quite evolved virus, had an icosahedral head of about 60 nm in diameter and a long non-contractile tail with sixfold symmetry. The capsid contained a single molecule of dsDNA of about 50 kb, and the tail was probably provided with a fixation apparatus. Head and tail were held together by a connector. a. The particle contained no lipids, was heavier than most viruses to come, and had a high DNA content proportional to its capsid size (about 50%). b. Most of its DNA coded for structural proteins. Morphopoietic genes clustered at one end of the genome, with head genes preceding tail genes. Lytic enzymes were probably coded for. A part of the phage genome was nonessential and possibly bacterial. Were tailed phages general transductants since the beginning? 3. The virus infected its host from the outside, injecting its DNA. Replication involved transcription in several waves and formation of DNA concatemers. Novel phages were released by burst of the infected cell after lysis of host membranes by a peptidoglycan hydrolase (and a holin?). a. Capsids were assembled from a starting point, the connector, and around a scaffold. They underwent an elaborate maturation process involving protein cleavage and capsid expansion. Heads and tails were assembled separately and joined later. b. The DNA was cut to size and entered preformed capsids by a headful mechanism. 4. Subsequently, tailed phages diversified by: a. Evolving contractile or short tails and elongated heads. b. Exchanging genes or gene fragments with other phages. c. Becoming temperate by acquiring an integrase-excisionase complex, plasmid parts, or transposons. d. Acquiring DNA and RNA polymerases and other replication enzymes. e. Exchanging lysin genes with their hosts. f. Losing the ability to form concatemers as a consequence of acquiring transposons (Mu) or proteinprimed DNA polymerases (phi 29). Present-day tailed phages appear as chimeras, but their monophyletic origin is still inscribed in their morphology, genome structure, and replication strategy. It may also be evident in the three-dimensional structure of capsid and tail proteins. It is unlikely to be found in amino acid sequences because constitutive proteins must be so old that relationships were obliterated and most or all replication-, lysogeny-, and lysis-related proteins appear to have been borrowed. However, the sum of tailed phage properties and behavior is so characteristic that tailed phages cannot be confused with other viruses. [TOP OF PAGE]

112. Vyiavlenie roli divergentsii v evoliutsii fagov-transpozonov gruppy B3 Pseudomonas aeruginosa [Role of divergence in evolution of group B3 Pseudomonas aeruginosa transposable phage evolution]. Akhverdian, V.Z., Khrenova, E.A., Lobanov, A.O., Krylov, V.N. (1998). Genetika 34:846-849. A heteroduplex analysis was performed to identify and map divergent DNA sequences in the genomes of the P. aeruginosa transposable phages (TPs) of group B3 using different formamide concentrations (30, 50, and 70%). Six PTs were classified into three related species--B3, PM681, and PM57. The role of DNA divergence in the evolution of TPs within one species is insignificant: the genomes of phages pM105 and PM681 (species PM681) and phages Hw12 and pM57 (species pM57) were shown to contain either homologous (98%) or nonhomologous DNA (2%). Homologous, divergent, and nonhomologous DNA regions (modules) were identified in the genomes of the TP of different species. Homologous modules with a level of DNA homology higher than 86% constitute approximately 30% of the phage genome; they are located at the left (1-5 kb) and right (29-38 kb) ends of the phage genome. Divergent modules with a DNA homology level between 50 and 67% and nonhomologous modules represent 30 to 35% and 25 to 30% of the phage genome, respectively. These regions form a mosaic structure in a 5-29-kb region. Thus, the key role of DNA divergence in the evolution of the natural TPs of three related species of group B3 was shown. A single region containing a 5-11-kb divergent DNA sequence was detected in the pM62 phage genome (species pM57). As shown by our previous data, this region was integrated into phage pM62 via interspecific recombination with a phage of species B3. [TOP OF PAGE]

113. Rekominatsionnoe proiskhozhdenie prirodnykh fagov-transpozonov rodstvennykh vidov gruppy B3, aktivnykh na bakteriiakh vida Pseudomonas aeruginosa [Recombinational origin of natural transposable phages of related species belonging to group B3, active in Pseudomonas aeruginosa species]. Akhverdian, V.Z., Lobanov, A.O., Khrenova, E.A., Krylov, V.N. (1998). Genetika 34:697-700. A heteroduplex analysis of four related transposable phages--B3, PM57, PM62, and Hw12--of the Pseudomonas aeruginosa B3 group was performed. Heteroduplex structures, restriction maps, and data on DNA-DNA hybridization obtained upon hybridization of phage DNA restriction fragments with labeled probes representing different regions of the phage genomes are in good agreement. The data obtained strongly confirmed the recombinational origin of the analyzed phages. Thus, all natural transposable phages of P. aeruginosa, including phages from both group B3 and species D3112, were shown to have a recombinational origin. [TOP OF PAGE]

114. His1, and archaeal virus of the Fuselloviridae family that infects Haloarcula hispanica. Bath, C., Dyall-Smith, M.L. (1998). J. Virol. 72:9392-9395. A novel archaeal virus, His1, was isolated from hypersaline waters in south-eastern Australia. It was lytic, grew only on Ha. hispanica (up to titres of 1011p.f.u./ml), and displayed a "lemon-shaped" morphology (74nm x 44nm) previously reported only for a virus of the extreme thermophiles (SSV1). The density of His1 was approximately 1.28g/ml - similar to that of SSV1 (1.24g/ml). Purified particles were resistant to low salt. The genome was linear, dsDNA and 14.9kb in size, which was similar in size to the genome of the SSV1 (ie. 15.5kb). Morphologically, this isolate clearly belongs to the recently proposed Fuselloviridae family of archaeal viruses. It represents the first member from the extremely halophilic archaea, and its host, Ha. hispanica, is one that can be readily manipulated genetically. [TOP OF PAGE]

115. Bioluminescence-based assays for detection and characterization of bacteria and chemicals in clinical laboratories. Billard, P., DuBow, M.S. (1998). Clinical Biochemistryy 31:1-14. OBJECTIVES: To survey recent advances in the application of bioluminescence to public health problems. The usefulness of bacterial (lux) and eucaryotic (luc) luciferase genes is presented, along with several examples that demonstrate their value as "reporters" of many endpoints of clinical concern. CONCLUSIONS: The development of new technologies for monitoring biological and chemical contaminants is in continuous progress. Recent excitement in this area has come from the use of genes encoding enzymes for bioluminescence as reporter systems. Applications of the recombinant luciferase reporter phage concept now provide a sensitive approach for bacterial detection, their viability, and sensitivity to antimicrobial agents. Moreover, a number of fusions of the lux and luc genes to stress inducible genes in different bacteria can allow a real-time measurement of gene expression and determination of cellular viability, and also constitute a new tool to detect toxic chemicals and their bioavailability. [TOP OF PAGE]

116. Response of model microbial communities to increased productivity. Bohannan, B.J.M. (1998). Michigan State Univeristy. [TOP OF PAGE]

117. Molecular ecology and evolution of Streptococcus thermophilus bacteriophages--a review. Brussow, H., Bruttin, A., Desiere, F., Lucchini, S., Foley, S. (1998). Virus Genes 16:95-109. Bacteriophages attacking Streptococcus thermophilus, a lactic acid bacterium used in milk fermentation, are a threat to the dairy industry. These small isometric-headed phages possess double-stranded DNA genomes of 31 to 45 kb. Yoghurt-derived phages exhibit a limited degree of variability, as defined by restriction pattern and host range, while a large diversity of phage types have been isolated from cheese factories. Despite this diversity all S. thermophilus phages, virulent and temperate, belong to a single DNA homology group. Several mechanisms appear to create genetic variability in this phage group. Site-specific deletions, one type possibly mediated by a viral recombinase/integrase, which transformed a temperate into a virulent phage, were observed. Recombination as a result of superinfection of a lysogenic host has been reported. Comparative DNA sequencing identified up to 10% sequence diversity due to point mutations. Genome sequencing of the prototype temperate phage f Sfi21 revealed many predicted proteins which showed homology with phages from Lactococcus lactis suggesting horizontal gene transfer. Homology with phages from evolutionary unrelated bacteria like E. coli (e.g. lambdoid phage 434 and P1) and Mycobacterium f L5 was also found. Due to their industrial importance, the existence of large phage collections, and the whole phage genome sequencing projects which are currently underway, the S. thermophilus phages may present an interesting experimental system to study bacteriophage evolution. [References: 48]. [TOP OF PAGE]

118. Viral escape from antisense RNA. Bull, J.J., Jacoboson, A., Badgett, M.R., Molineux, I.J. (1998). Molecular Microbiology 28:835-846. RNA coliphage SP was propagated for several generations on a host expressing an inhibitory antisense RNA complementary to bases 31-270 of the positive-stranded genome. Phages evolved that escaped inhibition. Typically, these escape mutants contained 3-4 base substitutions, but different sequences were observed among different isolates. The mutations were located within three different types of structural features within the predicted secondary structure of SP genomic RNA: (i) hairpin loops; (ii) hairpin stems; and (iii) the 5' region of the phage genome complementary to the antisense molecule. Computer modelling of the mutant genomic RNAs showed that all of the substitutions within hairpin stems improved the Watson-Crick pairing of the stem. No major structural rearrangements were predicted for any of the mutant genomes, and most substitutions in coding regions did not alter the amino acid sequence. Although the evolved phage populations were polymorphic for substitutions, many substitutions appeared independently in two selected lines. The creation of a new, perfect, antisense RNA against an escape mutant resulted in the inhibition of that mutant but not of other escape mutants nor of the ancestral, unevolved phage. Thus, at least in this system, a population of viruses that evolved to escape from a single antisense RNA would require a cocktail of several antisense RNAs for inhibition. [TOP OF PAGE]

119. The bacteriophages. Champagne, C.P., Moineau, S. (1998). pp. 89-116. In In Champagne, C.P. (ed.), Production of Dairy Starter Cultures (in French). [TOP OF PAGE]

120. Seasonal abundance of lysogenic bacteria in a subtropical estuary. Cochran, P.K., Paul, J.H. (1998). Appl. Environ. Microbiol. 64:2308-2312. Seasonal changes in the abundance of inducible lysogenic bacteria in a eutrophic estuarine environment were investigated over a 13-month period. Biweekly water samples were collected from Tampa Bay, Fla., and examined for prophage induction by mitomycin C treatment. At the conclusion of the study, we determined that 52.2% of the samples displayed prophage induction, as indicated by significant increases in viral direct counts compared with uninduced controls. Samples that displayed prophage induction occurred during the warmer months (February through October), when surface water temperatures were above 19 degree C, and no induction was observed in November, December, or January. This study presents clear evidence that there is seasonal variation in the number of inducible lysogenic bacteria in an estuarine environment. [TOP OF PAGE]

121. Prophage induction of indigenous marine lysogenic bacteria by environmental pollutants. Cochran, P.K., Kellogg, C.A., Paul, J.H. (1998). Mar. Ecol. Prog. Ser. 164:124-133. Lysogenic bacteria may be abundant components of bacterial assemblages in marine waters. The tremendous number of viruses found in estuarine and other eutrophic environments may be the result in part of induction of prophages. Mitomycin C is the inducing agent of choice for prophage induction; however this is not naturally found in the marine environment. We determined the capability of environmentally important pollutants to effect prophage induction in natural populations of marine bacteria. We investigated Aroclor 1248, a PCB mixture, bunker C fuel oil No. 6, and a pesticide mixture as inducing agents for natural bacterial communities from the Gulf of Mexico. Mitomycin C was also employed as a positive control for induction. Induction was determined as a significant increase in viral direct counts compared to control and ranged from 149 to 1336% of the controls. Two-thirds of the environments sampled showed prophage induction by one of the methods utilized, with the PCB mixture and Aroclor 1248 giving the highest percent efficiency (75%) of induction. This study shows that many environmentally important pollutants may be inducing agents for natural lysogenic viral production in the marine environment. [TOP OF PAGE]

122. Increasing phage resistance of cheese starters: A case study using Lactococcus lactis DPC4268. Coffey, A., Coakley, M., McGarry, A., Fitzgerald, G.F., Ross, R.P. (1998). Letters in Applied Microbiology 26:51-55. This study serves as an example of strategies used to increase the phage resistance of an important Irish Cheddar cheese starter, Lactococcus lactis DPC4268. It describes the emergence and persistence of a lytic bacteriophage, 4268, that has a relatively large burst size and exhibits no homology to the most common phage types encountered in Irish cheese plants. Inherent difficulties were encountered that prevented the effective introduction of conjugative phage-resistance plasmids pNP40 and pMRCO1 to strain DPC4268. In fact, pNP40-associated Abi systems were naturally present in six of 19 starters. Control of phage 4268 was eventually achieved by generating a mutant of DPC4268, which was subsequently used for cheese manufacture. [TOP OF PAGE]

123. Increasing the phage resistance of cheese starter Lactococcus lactis DPC4268 in response to the emergence of a novel highly virulent phage in industry. Coffey, A., Coakley, M., McGarry, A., Fitzgerald, G.F., Ross, R.P. (1998). pp. 460-481. In In ??? (ed.), Actes du Colloque LACTIC-97 Lactic Acid Bacteria: Which strains? For which products? Villers Bocage, Cedex, France. [TOP OF PAGE]

124. Phages infecting Vibrio vulnificus are abundant and diverse in oysters (Crassostrea virginica) collected from the Gulf of Mexico. Depaola, A., Motes, M.L., Chan, A.M., Suttle, C.A. (1998). Appl. Environ. Microbiol. 64:346-351. Phages infecting Vibrio vulnificus were abundant (10^-4 phages g of oyster tissue^-1) throughout the year in oysters (Crassostrea virginica) collected from estuaries adjacent to the Gulf of Mexico (Apalachicola Bay, Fla.; Mobile Bay, Ala.; and Black Bay, La.). Estimates of abundance ranged from 10^-1 to 10^-5 phages g of oyster tissue^-1 and were dependent on the bacterial strain used to assay the sample. V. vulnificus was near or below detection limits ( lt 0.3 cell g^-1) from January through March and was most abundant (10^-3 to 10^-4 cells g^-1) during the summer and fall, when phage abundances also tended to be greatest. The phages isolated were specific to strains of V. vulnificus, except for one isolate that caused lysis in a few strains of V. parahaemolyticus. Based on morphological evidence obtained by transmission electron microscopy, the isolates belonged to the Podoviridae, Styloviridae, and Myoviridae, three families of double-stranded DNA phages. One newly described morphotype belonging to the Podoviridae appears to be ubiquitous in Gulf Coast oysters. Isolates of this morphotype have an elongated capsid (mean, 258 nm; standard deviation, 4 nm; n = 35), with some isolates having a relatively broad host range among strains of V. vulnificus. Results from this study indicate that a morphologically diverse group of phages which infect V. vulnificus is abundant and widely distributed in oysters from estuaries bordering the northeastern Gulf of Mexico. [TOP OF PAGE]

125. Evolution of Streptococcus thermophilus bacteriophage genomes by modular exchanges followed by point mutations and small deletions and insertions. Desiere, F., Lucchini, S., Brussow, H. (1998). Virology 241:345-356. Comparative sequence analysis of 40 % of the genomes from two prototype Streptococcus thermophilus bacteriophages (lytic group I phage fSfi19 and the cos-site containing temperate phage fSfi21) suggested two processes in the evolution of their genomes. In a first evolutionary distant phase the basic genome structure was apparently constituted by modular exchanges. Over the 17 kb long DNA segment analyzed in the present report we observed clusters of genes with similarity to genes from Leuconostoc oenos phage L10, Lactococcus lactis phage BK5-T and Streptococcus pneumoniae phage Dp-1. A chimeric protein was predicted for orf 1291 which showed similarity both to phage BK5-T and phage Dp-1. The very large orf 1626 gene product showed similarity to two adjacent genes from the Lactobacillus delbrueckii phage LL-H and further phage proteins (Lactococcus lactis, Bacillus subtilis). The similarities were localized to distinct parts of this apparently multifunctional protein. The putative fSfi19 lysin showed similarity both to lysins of phages and cellular enzymes. In a second evolutionary more recent phase the S. thermophilus phage genomes apparently diversified by point mutations and small deletions/insertions. Over the investigated 17 kb DNA region fSfi19 differed from fSfi21 by 10 % base pair changes, the majority of which were point mutations (mainly at the third codon position), while a third of the bp differences were contributed by small deletions/insertions. The bp changes were unevenly distributed: Over the Leuconostoc phage-related DNA the change rate was high, while over the Lactococcus and S. pneumoniae phage-related DNA the change rate was low. We speculate that the degree of bp changes could provide relative time scales for the modular exchange reactions observed in S. thermophilus phages. [TOP OF PAGE]

126. Occurrence of a sequence in marine cyanophages similar to that of T4 gp20 and its application to PCR-based detection and quantification techniques. Fuller, N.J., Wilson, W.H., Joint, I.R., Mann, N.H. (1998). Appl. Environ. Microbiol. 64:2051-2060. Viruses are ubiquitous components of marine ecosystems and are known to infect unicellular phycoerythrin-containing cyanobacteria belonging to the genus Synechococcus. A conserved region from cyanophage genome was identified in three genetically distinct cyanomyoviruses, and a sequence analysis revealed that this region exhibited significant similarity to a gene encoding a capsid assembly protein (gp20) from the enteric coliphage T4. The results of a comparison of gene 20 sequences from three cyanomyoviruses and T4 allowed us to design two degenerate PCR primers, CPS1 and CPS2, which specifically amplified a 165-bp region from the majority of cyanomyoviruses tested. A competitive PCR (cPCR) analysis revealed that cyanomyovirus strains should be accurately enumerated, and it was demonstrated that quantification was log-linear over ca. 3 orders of magnitude. Different calibration curves were obtained for each of the three cyanomyovirus strains tested; consequently, cPCR performed with primers CPS1 and CPS2 could lead to substantial inaccuracies in estimates of phage abundance in natural assemblages. Further sequence analysis of cyanomyovirus gene 20 homologs would be necessary in order to design primers which do not exhibit phage-to-phage variability in priming efficiency. It was demonstrated that PCR products of the correct size could be amplified from seawater samples following 100x concentration and even directly without any prior concentration. Hence, the use of degenerate primers in PCR analysis of cyanophage populations should provide valuable data on the diversity of cyanophages in natural assemblages. Further optimization of procedures may ultimately lead to a sensitive assay which can be used to analyze natural cyanophage populations both quantitatively (by cPCR) and qualitatively following phylogenetic analysis of amplified products. [TOP OF PAGE]

127. The effect of cyanophages on the morality of Synechococcus spp. and selection for UV resistant viral communities. Garza, D.R., Suttle, C.A. (1998). Microb. Ecol. 36:281-??? [TOP OF PAGE]

128. Gene transfer by transduction in the marine environment. Jiang, S.C., Paul, J.H. (1998). Appl. Environ. Microbiol. 64:2780-2787. To determine the potential for bacteriophage-mediated gene transfer in the marine environment, we established transduction systems by using marine phage host isolates. Plasmid pQSR50, which contains transposon Tn5 and encodes kanamycin and streptomycin resistance, was used in plasmid transduction assays. Both marine bacterial isolates and concentrated natural bacterial communities were used as recipients in transduction studies. Transductants were detected by a gene probe complementary to the neomycin phosphotransferase (nptII) gene in Tn5. The transduction frequencies ranged from 1.33 X 10^-7 to 5.13 X 10^-9 transductants/PFU in studies performed with the bacterial isolates. With the mixed bacterial communities, putative transductants were detected in two of the six experiments performed. These putative transductants were confirmed and separated from indigenous antibiotic-resistant bacteria by colony hybridization probed with the nptII probe and by PCR amplification performed with two sets of primers specific for pQSR50. The frequencies of plasmid transduction in the mixed bacterial communities ranged from 1.58 X 10^-8 to 3.7 X 10^-8 transductants/PFU. Estimates of the transduction rate obtained by using a numerical model suggested that up to 1.3 X 10¹⁴ transduction events per year could occur in the Tampa Bay Estuary. The results of this study suggest that transduction could be an important mechanism for horizontal gene transfer in the marine environment. [TOP OF PAGE]

129. Characterization of marine temperate phage-host systems isolated from Mamala Bay, Oahu, Hawaii. Jiang, S.C., Kellogg, C.A., Paul, J.H. (1998). Appl. Environ. Microbiol. 64:535-542. To understand the ecological and genetic role of viruses in the marine environment, it is critical to know the infectivity of viruses and the types of interactions that occur between marine viruses and their hosts. We isolated four marine phages from turbid plaques by using four indigenous bacterial hosts obtained from concentrated water samples from Mamala Bay, Oahu, Hawaii. Two of the rod-shaped bacterial hosts were identified as Sphingomonas paucimobilis and Flavobacterium sp. All of the phage isolates were tailed phages and contained double-stranded DNA. Two of the phage isolates had morphologies typical of the family Siphoviridae, while the other two belonged to the families Myoviridae and Podoviridae. The head diameters of these viruses ranged from 47 to 70.7 nm, and the tail lengths ranged from 12 to 146 nm. The burst sizes ranged from 7.8 to 240 phage/bacterial cell, and the genome sizes, as determined by restriction digestion, ranged from 36 to 112 kb. The members of the Siphoviridae, T-f HSIC, and T-f D0, and the member of the Myoviridae, T-f D1B, were found to form lysogenic associations with their bacterial hosts, which were isolated from the same water samples. Hybridization of phage T-f HSIC probe with lysogenic host genomic DNA was observed in dot blot hybridization experiments, indicating that prophage T-f HSIC was integrated within the host genome. These phage-host systems are available for use in studies of marine lysogeny and transduction. [TOP OF PAGE]

130. Significance of lysogeny in the marine environment: studies with isolates and a model of lysogenic phage production. Jiang, S.C., Paul, J.H. (1998). Microb. Ecol. 35:235-243. The importance of lysogeny in marine microbial populations is just beginning to be understood. To determine the abundance of lysogens in bacterial populations, we studied the occurrence of lysogenic bacteria among bacterial isolates from a variety of marine environments. More than 116 bacteria isolated on artificial seawater nutrient agar plates were tested for the presence of inducible prophage by mitomycin C and UV radiation. Induction was determined as a decrease in culture absorbance at 600 nm, after treatment with inducing agents. Samples in which optical density decreased or remained the same after induction were further examined by transmission electron microscopy, for the presence of virus-like particles. More than 40% of the bacterial isolates contained inducible prophage, as determined by mitomycin C induction. A higher percentage of lysogenic bacteria was found in isolates from oligotrophic environments, compared to coastal or estuarine environments. These studies suggest that lysogenic bacteria are important components in marine microbial populations. However, a mathematical model based on viral and bacterial abundance and production rates suggests that, under normal conditions, lysogenic viral production contributes less than 0.02% of total viral production. Therefore, lysogens in the marine environment may serve as a source of viruses and only contribute significantly to viral production during natural induction events. [TOP OF PAGE]

131. Viruses in Antarctic lakes. Kepner, R.L., Wharton, R.A.Jr., Suttle, C.A. (1998). Limnol. Oceanogr. 43:1754-1761. Water samples collected from four perennially ice-covered Antarctic lakes during the austral summer of 1996-1997 contained high densities of extracellular viruses. Many of these viruses were found to be morphologically similar to double-stranded DNA viruses that are known to infect algae and protozoa. These constitute the first observations of viruses in perennially ice-covered polar lakes. The abundance of planktonic viruses and data suggesting substantial production potential (relative to bacterial secondary and photosynthetic primary production) indicate that viral lysis may be a major factor in the regulation of microbial populations in these extreme environments. Furthermore, we suggest that Antarctic lakes may be a reservoir of previously undescribed viruses that possess novel biological and biochemical characteristics. [TOP OF PAGE]

132. Phage Therapy: Bacteriophages as Antibiotics. Kutter, E. (1998). [TOP OF PAGE]

133. The structural gene module in Streptococcus thermophilus bacteriophage f Sfi11 shows a hierarchy of relatedness to Siphoviridae from a wide range of bacterial hosts. Lucchini, S., Desiere, F., Brussow, H. (1998). Virology 246:63-73. The structural gene cluster and the lysis module from lytic group II Streptococcus thermophilus bacteriophage f Sfi11 was compared to the corresponding region from other Siphoviridae. The analysis revealed a hierarchy of relatedness. f Sfi11 differed from the temperate S. thermophilus bacteriophage f O1205 by about 10% at the nucleotide level. The majority of the changes were point mutations, mainly at the third base position. Only a single gene (orf 695) differed substantially between the two phages. Over the putative minor tail and lysis genes, f Sfi11 and the lytic group 1 S. thermophilus f Sfi19 shared regions with variable degrees of similarity. Orf 1291 from f Sfi19 was replaced by four genes in f Sfi11, two of which (orf 1000 and orf 695) showed a complicated pattern of similarity and nonsimilarity compared with f Sfi19. The predicted orf 695 gp resembles the receptor-recognizing protein of T-even coliphages in its organization, but not its sequence. No sequence similarity was detected between f Sfi11 and f Sfi19 in the region covering the major head and tail genes. Comparison of the structural gene map of f Sfi11 with that of Siphoviridae from gram-positive and -negative bacterial hosts revealed a common genomic organization. Sequence similarity was only found between f Sfi11 and Siphoviridae from gram-positive hosts and correlated with the evolutionary distance between the bacterial hosts. Our data are compatible with the hypothesis that the structural gene operon from Siphoviridae of the low G + C group of gram-positive bacteria is derived from a common ancestor. [TOP OF PAGE]

134. Elevated production of dimethylsulfide resulting from viral infection of cultures of Phaeocystis pouchetii. Malin, G., Wilson, W.H., Bratbak, G., Liss, P.S., Mann, N.H. (1998). Limnol. Oceanogr. 43:1389-1393. [TOP OF PAGE]

135. Taxonomy of bacterial viruses: establishment of tailed virus genera and the order Caudovirales [news]. Maniloff, J., Ackermann, H.-W. (1998). Archives in Virology 143:2051-2063. [TOP OF PAGE]

136. Evidence of pseudolysogeny in a marine phage host system. McLaughlin, M.R., Paul, J.H. (1998). Abstracts of the General Meeting of the American Society for Microbiology 98:387-??? [TOP OF PAGE]

137. Comparison of the lysogeny modules from the temperate Streptococcus thermophilus bacteriophages TP-J34 and Sfi21: implications for the modular theory of phage evolution. Neve, H., Zenz, K.I., Desiere, F., Koch, A., Heller, K.J., Brussow, H. (1998). Virology 241:61-72. A 7.6-kb DNA segment covering the putative lysogeny module of the pac-site-containing temperate Streptococcus thermophilus bacteriophage TP-J34 was sequenced. Sequence alignment with the lysogeny module from the cos-site-containing S. thermophilus bacteriophage fSfi21 revealed areas of high sequence conservation (e.g., over the int gene), interspersed with regions of low or no sequence similarity (e.g., over the cro gene). Four of the six sharp transition zones from high to low sequence conservation were found within open reading frames coding for the CI repressor, the Anti-repressor, the Immunity protein, and a protein of unknown function. The transition points in the cI and ant genes appear to separate gene segments coding for distinct functional domains of these proteins. In addition, these two transition points were located at or near the deletion sites observed in spontaneous phage fSfi21 deletion mutants, thus suggesting these transition points as recombinational hotspots. Furthermore, the sequence at the transition point in the cI gene resembles the attachment site of the phage, suggesting the involvement of the phage integrase in at least some of the exchange reactions. Contrary to the initial formulation of the modular theory of phage evolution the unit of the evolutionary exchange in streptococcal phages is not a group of functional genes, but can be as small as a single gene. Exchange reactions can also occur within genes, possibly between gene segments encoding distinct protein domains. [TOP OF PAGE]

138. Use of SYBR Green I for rapid epifluorescence counts of marine viruses and bacteria. Noble, R.T., Fuhrman, J.A. (1998). Aquat. Microb. Ecol. 14:113-118. A new nucleic acid stain, SYBR Green I, can be used for the rapid and accurate determination of viral and bacterial abundances in diverse marine samples. We tested this stain with formalin-preserved samples of coastal water and also from depth profiles (to 800 m) from sites 19 and 190 km offshore, by filtering a few ml onto 0.02 mu m pore-size filters and staining for 15 min. Comparison of bacterial counts to those made with acridine orange (AO) and virus counts with those made by transmission electron microscopy (TEM) showed very strong correlations. Bacterial counts with AO and SYBR Green I were indistinguishable and almost perfectly correlated (r super(2) = 0.99). Virus counts ranged widely, from 0.03 to 15 x 10 super(7) virus ml super(-1). Virus counts by SYBR Green I were on the average higher than those made by TEM, and a SYBR Green I versus TEM plot yielded a regression slope of 1.28. The correlation between the two was very high with an r super(2) value of 0.98. The precision of the SYBR Green I method was the same as that for TEM, with coefficients of variation of 2.9%. SYBR Green I stained viruses and bacteria are intensely stained and easy to distinguish from other particles with both older and newer generation epifluorescence microscopes. Detritus is generally not stained, unlike when the alternative dye YoPro I is used, so this approach may be suitable for sediments. SYBR Green I stained samples need no desalting or heating, can be fixed with formalin prior to filtration, the optimal staining time is 15 min (resulting in a total preparation time of less than 25 min), and counts can be easily performed at sea immediately after sampling. This method may facilitate incorporation of viral research into most aquatic microbiology laboratories. [TOP OF PAGE]

139. Design of a phage-insensitive lactococcal dairy starter via sequential transfer of naturally occurring conjugative plasmids. O'Sullivan, D., Coffey, A., Fitzgerald, G.F., Hill, C., Ross, R.P. (1998). Appl. Environ. Microbiol. 64:4618-4622. [TOP OF PAGE]

140. Abundance, morphology and distribution of planktonic virus-like particles in two high-mountain lakes. Pina, S., Creus, A., Ganzález, N., Gironés, R., Felip, M., Sommaruga, R. (1998). Journal of Plankton Research 20:2413-2421. Direct counts of virus-like particles (VLP) by transmission electron microscopy revealed abundances of up to 3 x 10⁷ ml^-1 in the plankton of two remote high-mountain lakes in the Alps and in the Pyrenees. Most VLP were icosahedric without tail and with diameters between 40 and 90 nm, but also very large ones with diameter of up to 325 nm were observed. VLP outnumbered bacteria by a factor of 4.2 to 42.8 and bacterial cells were infected with large numbers (>50) of viral particles. This study constitutes the first report on aquatic viruses for alpine lakes and it suggests that they may be an important additional source of bacterial mortality in these systems. [TOP OF PAGE]

141. Biochemical and phylogenetic characterization of the dUTPase from the archaeal virus SIRV. Prangishvili, D., Klenk, H.P., Jakobs, G., Schmiechen, A., Hanselmann, C., Holz, I., Zillig, W. (1998). Journal of Biological Chemistry 273:6024-6029. The derived amino acid sequence from a 474-base pair open reading frame in the genome of the Sulfolobus islandicus rod-shaped virus SIRV shows striking similarity to bacterial dCTP deaminases and to dUTPases from eukaryotes, bacteria, Poxviridae, and Retroviridae. The putative gene was expressed in Escherichia coli, and dUTPase activity of the recombinant enzyme was demonstrated by hydrolysis of dUTP to dUMP. Deamination of dCTP by the enzyme was not detected. Phylogenetic analysis based on amino acid sequences of the characterized enzyme and its homologues showed that the dUTPase-encoding dut genes and the dCTP deaminase-encoding dcd genes constitute a paralogous gene family. This report is the first identification and functional characterization of an archaeal dUTPase and the first phylogeny derived for the dcd-dut gene family. [TOP OF PAGE]

142. Detection, quantification and morphological characterization of Vibrio cholerae indicator bacteriophages [Spanish]. Talledo, M.A., Gutiérrez, S., Merino, F., Rojas, N. (1998). Rev. Peru. Biol. 5:90-97. [TOP OF PAGE]

143. Sex and the evolution of intrahost competition in RNA virus f6. Turner, P.E., Chao, L. (1998). Genetics 150:523-532. Sex allows beneficial mutations that occur in separate lineages to be fixed in the same genome. For this reason, the Fisher-Muller model predicts that adaptation to the environment is more rapid in a large sexual population than in an equally large asexual population. Sexual reproduction occurs in populations of the RNA virus phi6 when multiple bacteriophages coinfect the same host cell. Here, we tested the model's predictions by determining whether sex favors more rapid adaptation of phi6 to a bacterial host, Pseudomonas phaseolicola. Replicate populations of phi6 were allowed to evolve in either the presence or absence of sex for 250 generations. All experimental populations showed a significant increase in fitness relative to the ancestor, but sex did not increase the rate of adaptation. Rather, we found that the sexual and asexual treatments also differ because intense intrahost competition between viruses occurs during coinfection. Results showed that the derived sexual viruses were selectively favored only when coinfection is common, indicating that within-host competition detracts from the ability of viruses to exploit the host. Thus, sex was not advantageous because the cost created by intrahost competition was too strong. Our findings indicate that high levels of coinfection exceed an optimum where sex may be beneficial to populations of phi6, and suggest that genetic conflicts can evolve in RNA viruses. [TOP OF PAGE]

144. Significance of viral lysis and flagellate grazing as factors controlling bacterioplankton production in a eutrophic lake. Weinbauer, M.G., Hofle, M.G. (1998). Appl. Environ. Microbiol. 64:431-438. The effects of viral lysis and heterotrophic nanoflagellate (HNF) grazing on bacterial mortality were estimated in a eutrophic lake (Lake Plussee in northern Germany) which was separated by a steep temperature and oxygen gradient into a warm and oxic epilimnion and a cold and anoxic hypolimnion. Two transmission electron microscopy-based methods (whole-cell examination and thin sections) were used to determine the frequency of visibly infected cells, and a model was used to estimate bacterial mortality due to viral lysis. Examination of thin sections also showed that between 20.2 and 29.2% (average, 26.1%) of the bacterial cells were empty (ghosts) and thus could not contribute to viral production. The most important finding was that the mechanism for regulating bacterial production shifted with depth from grazing control in the epilimnion to control due to viral lysis in the hypolimnion. We estimated that in the epilimnion viral lysis accounted on average for 8.4 to 41.8% of the summed mortality (calculated by determining the sum of the mortalities due to lysis and grazing), compared to 51.3 to 91.0% of the summed mortality in the metalimninon and 88.5 to 94.2% of the summed mortality in the hypolimnion. Estimates of summed mortality values indicated that bacterial production was controlled completely or almost completely in the epilimnion (summed mortality, 66.6 to 128.5%) and the hypolimnion (summed mortality, 43.4 to 103.3%), whereas in the metalimnion viral lysis and HNF grazing were not sufficient to control bacterial production (summed mortality, 22.4 to 56.7%). The estimated contribution of organic matter released by viral lysis of cells into the pool of dissolved organic matter (DOM) was low; however, since cell lysis products are very likely labile compared to the bulk DOM, they might stimulate bacterial production. The high mortality of bacterioplankton due to viral lysis in anoxic water indicates that a significant portion of bacterial production in the metalimnion and hypolimnion is cycled in the bacterium-virus-DOM loop. This finding has major implications for the fate and cycling of organic nutrients in lakes. [TOP OF PAGE]

145. Size-specific mortality of lake bacterioplankton by natural virus communities. Weinbauer, M.G., Hoefle, M.G. (1998). Aquat. Microb. Ecol. 15:103-113. The potential effect that viral lysis has on the cell size distribution of bacterioplankton was investigated during late summer stratification in Lake Plusssee, Germany. Size-specific bacterial mortality due to viral lysis was estimated from in situ samples by a transmission electron microscopy based examination of visibly infected cells (VIC) and in an experiment with varying concentrations of the natural virus community. In all depth layers the highest percentage of cells was found in a cell length class that was smaller for the entire bacterial community (0.3-0.6 mu m) than for VIC (0.6-0.9 mu m). For cells <2.4 mu m the highest frequency of VIC (FVIC) was detected in the size classes 0.6-0.9 and 0.9-1.2 mu m, and the FVIC was high in the size classes 1.2-1.5 (all depth layers) and 1.5-1.8 mu m (meta- and hypolimnion). The estimated mortality due to viral lysis in these size classes was significant with maxima of 29 to 55% in the epilimnion, 30 to 59% in the metalimnion and 56 to 107% in the hypolimnion. In all depth layers the FVIC of bacteria <0.3 mu m in length was ca 30% of that averaged for the entire bacterial community, and in the experiment the percentage of cells <0.3 mu m was highest in enclosures with high viral activity. [TOP OF PAGE]

146. Cell size-specific lysis of lake bacterioplankton by natural virus communities. Weinbauer, M.G., Höfle, M.G. (1998). Aquat. Microb. Ecol. 156:103-113. [TOP OF PAGE]

147. The role of sunlight in the removal and repair of viruses in the sea. Wilhelm, S.W., Weinbauer, M.G., Suttle, C.A., Jeffrey, W.H. (1998). Limnol. Oceanogr. 43:586-592. We investigated the in situ destruction rates of marine viral particles as well as the decay rates of infectivity for viral isolates along an similar to 400-km transect from oligotrophic offshore waters to productive coastal waters in the Gulf of Mexico. Light-mediated decay rates of viral infectivity averaged over the solar day ranged from 0.7 to 0.85 h super(-1) in surface waters at all stations and decreased with depth in proportion to the attenuation of UVB (305 nm). The destruction rates of viral particles also decreased with depth, although the rates of particle destruction were only 22-61% of infectivity when integrated over the mixed layer. The rates of viral particle destruction indicated that at three of four stations 6-12% of the daily bacterial production would have to be lysed in order to maintain ambient viral concentrations. At the fourth station, where there was a dense bloom of Synechococcus spp. and the mixed layer was shallower, 34-52% of the daily bacterial production would have to be lysed. A comparison of the difference between destruction rates of viral particles and infectivity integrated over the depth of the mixed layer implies that host-mediated repair must have restored infectivity to 39-78% of the sunlight-damaged viruses daily. The calculated frequency of contacts between viral particles and bacterial cells that resulted in infection (contact success) ranged from similar to 18 to 34% in offshore waters, where the frequency of contacts between viruses and bacteria was much lower, to similar to 1.0% at the most inshore station, where contact rates are much higher. This suggests that in offshore waters bacterial communities are less diverse, and that there is less selection to be resistant to viral infection. This paper provides a framework for balancing viral production, destruction, and light-dependent repair in aquatic viral communities. [TOP OF PAGE]

148. Measurements of DNA damage and photoreactivation imply that most viruses in marine surface waters are inefective. Wilhelm, S.W., Weinbauer, M.G., Suttle, C.A., Pledger, R.J., Mitchell, D.L. (1998). Aquat. Microb. Ecol. 14:215-222. The proportion of viruses in natural marine communities that are potentially infectious was inferred from the relationship between DNA damage and the loss of infectivity in marine viral isolates and measurements of the DNA damage in natural viral communities. Several viral isolates which infect marine Vibrio spp. were exposed to UV-C radiation and the concentration of cyclobutane pyrimidine dimers in the viral DNA was measured with a highly sensitive radioimmunoassay. The loss of infectivity in the UV-exposed isolates was also determined under conditions which either activated or repressed the blue light dependent photolyase enzyme in host cells in order to examine the damage-dependent response of this bacterial repair system. In addition, the accumulation of DNA photodamage during the solar day was measured in DNA isolated from natural viral communities collected along a transect in the western Gulf of Mexico. Using the correlation between DNA damage and infectivity for one of the viral isolates, we estimated the proportion of the natural viral community which was infective. The results imply that, due to light-mediated repair of damaged viral DNA by host-cell mechanisms (photoreactivation), greater than 50% of the viruses in natural communities are infective despite high rates of DNA damage. Furthermore, the accumulation of cyclobutane pyrimidine dimers was highest at the station where the surface mixed layer was shallowest, emphasizing the importance of mixing depth in relation to the accumulation of DNA damage. These experiments demonstrate that physical parameters such as mixing depth are critically interwoven with light penetration in influencing the infectivity of marine viral communities. [TOP OF PAGE]

149. Estimating the infectivity of marine viral communities from measurements of DNA damage and photoreactivation. Wilhelm, S.W., Weinbauer, M.G., Jeffrey, W.H., Suttle, C.A. (1998). Aquat. Microb. Ecol. 14:215-222. [TOP OF PAGE]

150. Population dynamics of phytoplankton and viruses in a phosphate-limited mesocosm and their effect on DMSP and DMS production. Wilson, W.H., Turner, S., Mann, N.H. (1998). Estuarine, Coastal and Shelf Science 46 (Supplement a):49-59. The effect of phosphate limitation on viral abundance, phytoplankton bloom dynamics and production of dimethylsulphoniopropionate (DMSP) and dimethyl sulphide (DMS) was investigated in seawater mesocosm enclosures, in a Norwegian fjord, during June 1995. Daily estimates of viral concentrations, based on transmission electron microscope (TEM) counts, varied on an apparently random basis in each of the enclosures. A large Synechococcus spp. bloom developed in an enclosure which was maintained at a high N:P ratio, simulating phosphate-deplete growth conditions. Following phosphate addition to this enclosure, there was a large increase in estimated virus numbers shortly before an apparent collapse of the Synechococcus bloom. It is tentatively suggested that lysogenic viruses were induced following phosphate addition to the phosphate-limited enclosures, and that these observations add to a growing body of evidence which supports the hypothesis that nutrient availability may be responsible for the switch between lysogeny and lytic production. High DMS concentrations and viral numbers were observed on the demise of the flagellate (predominantly Emiliania huxleyi) and diatom blooms, but overall there was no significant correlation. Highest concentrations of DMSP were associated with blooms of E. huxleyi, for which an intracellular concentration of 0.5 pg cell^-1 (SD, 0.06) was calculated. Good correlation of DMSP with Synechococcus spp. cell numbers was observed, suggesting that these species of picoplankton may be significant producers of DMSP. No effects of phosphate limitation on DMS and/or DMSP production were evident from the data. [TOP OF PAGE]

151. A catalogue of T4-type bacteriophages. Ackermann, H.-W., Krisch, H.M. (1997). Archives of Virology 142:2329-2345. The T4-type of bacteriophages is broadly defined on the basis of particle morphology. It occurs in enterobacteria (125 representatives), acinetobacters, aeromonads, pseudomonads, and vibrios (16 isolates). In addition, 18 apparently unrelated phages with prolate heads and contractile tails are found in a wide range of bacteria. A descriptive catalogue of these phages is presented. The T4-type probably originated in precursors of enterobacteria. [TOP OF PAGE]

152. Bacteriophage ecology. Ackermann, H.-W. (1997). pp. 335-339. In In Martins, M.T., Sato, M.I.Z., Tiedje, J.M., Hagler, L.C.N., Döbereiner, J., and Sanchez, P.S. (eds.), Progress in Microbial Ecology (Proceedings of Seventh International Symposium on Microbial Ecology). Brazilian Society for Microbiology, Bacteriophage are classified into 12 families. Phages are tailed or cubic, filamentous or pleomorphic. Phages occur in all parts of the bacteria world and in every possible habitat. Phage titers may attain 8 log/ml in seawater and 9-10 log/ml in rumen fluid. Somatic coliphages, F-RNA and Bacteriodes fragilis phages indicate sewage and/or fecal contamination. Industrial microbiology provides particular environments in which phages proliferate. [TOP OF PAGE]

153. Taxonomic changes in tailed phages of enterobacteria. Ackermann, H.W., DuBow, M.S., Gershman, M., Karska-Wysocki, B., Kasatiya, S.S., Loessner, M.J., Mamet-Bratley, M.D., Regue, M. (1997). Archives of Virology 142:1381-1390. Out of 136 new phages, 80 (59%) are classified into 23 species according to morphology and physicochemical properties. Six new species are described and species beta 4, from a previous classification scheme, is renamed T1. The morphology of 36 phage species is schematically represented. [TOP OF PAGE]

154. Isolation and Characterization of a New Lactobacillus delbrueckii ssp.bulgaricus Temperate Bacteriophage. Auad, L., de Ruiz Holgado, A.A.P., Forsman, P., Alatossava, T., Raya, R.R. (1997). J. Dairy Sci. 80:2706-2712. Lactobacillus delbrueckii ssp. bulgaricus strain CRL 539 was shown to be lysogenic and inducible with mitomycin C. The conditions were determined for an optimal induction of temperate bacteriophage lb539 with mitomycin C as well as the sensitivity of lb539 to physical and chemical agents. Electron microscopy of lysates revealed bacteriophage particles with an isometric head of 47 nm and a noncontractile tail of 159 nm. Phage lb539 was classified within Bradley's B1 phage group and the Siphoviridae family. The host range of lb539 encompassed mainly Lactobacillus delbrueckii ssp. lactis strains; strain LKT (CNRZ 700) was the most sensitive for detection of lb539 lysates induced by mitomycin C. The lb539 genome is a linear, double-stranded DNA molecule of approximately 35 kbp. The presence of submolar fragments in restriction enzyme digests suggests that lb539 DNA may contain a pac site. Dot-blot experiments showed that the lb539 genome hybridized with the genomes of phages mv4 and LL-H, which are type phages of group a of L. delbrueckii ssp. phages. Restriction enzyme patterns and morphological features showed lb539 to be distinct from mv4 and LL-H. [TOP OF PAGE]

155. Effect of resource enrichment on a chemostat community of bacteria and bacteriophage. Bohannan, B.J.M., Lenski, R.E. (1997). Ecology 78:2303-2315. We determined the responses of a model laboratory community to resource enrichment and compared these responses to the predictions of prey-dependent and ratio-dependent food chain models. Our model community consisted of Escherichia coli B and bacteriophage T4 in chemostats supplied with different concentrations of glucose. We observed the following responses to enrichment: (1) a large and highly significant increase in the equilibrium population density of the predator, bacteriophage T4, (2) a small but significant increase in the equilibrium population density of the prey, E. coli, and (3) a large and highly significant decrease in the stability of both the predator and prey populations. These responses were better predicted by a prey-dependent model (altered to include a time delay between consumption and reproduction by predators) than by a ratio-dependent model. Enrichment had a large effect on evolutionary change in our system. Enrichment significantly decreased the amount of time required for mutants of E. coli that were resistant to predation by bacteriophage to appear in the chemostats. Enrichment also significantly increased the rate at which these bacteriophage-resistant mutants invaded the chemostats. These results were also better predicted by the prey-dependent model. Invasion by bacteriophage-resistant mutants had a large effect on the subsequent population dynamics of both predator and prey. Both the equilibrium density and stability of the E. coli population increased following invasion, and the population shifted from being primarily limited by predators to being primarily limited by resources. After invasion by the mutants, the T4 population decreased in equilibrium density, and.the population cycled with an increased period. These results were compared to the predictions of a ratio-dependent model and a prey-dependent model altered to include T4-resistant mutants. The dynamics of this community were better predicted by the modified prey-dependent model; however, this model was more complex mathematically than the simpler ratio-dependent model. [TOP OF PAGE]

156. Characterization of the lysogeny DNA module from the temperate Streptococcus thermophilus bacteriophage f Sfi21. Bruttin, A., Desiere, F., Lucchini S, Foley S, Brussow, H. (1997). Virology 233:136-148. Phage f Sfi21, the only temperate Streptococcus thermophilus phage from our phage collection, showed extensive DNA homology with virulent phages from lytic group I. Southern blot hybridizations demonstrated that the f Sfi21- specific DNA was clustered in an approximately 6.6- kb-long region, the putative lysogeny module. Sequence analysis and database research identified an integrase within this module; orf 203 with homology to an anonymous orf 258 from the temperate lactococcal phage BK5-T; orf 127 and orf 122 with weak homology to the N- and C- terminal parts, respectively, of the cl-like repressor from lactococcal phages Tuc2009 and BK5-T; orf 75 with homology to a repressor protein from lambdoid phage 434 and an anti-repressor ant with homology to phage P1. The molecular arrangement of the predicted orfs in phage phi Sfi21 was very similar to that of the lactococcal phage BK5-T. The transition from f Sfi21-specific DNA into DNA shared with virulent phages was abrupt and flanked at one side by notable DNA repeats. Sequence analysis identified a holin protein to the left of the lysogeny module. A site-specific deletion of 2.4 kb, which reproducibly transformed f Sfi21 into a lytic phage, was localized in the lysogeny module. It was flanked at both sides by conspicuous DNA repeats. One repeat region reflected the DNA around the attP site, while the other reflected the putative genetic switch region between repressor and anti- repressor genes. S. thermophilus host Sfi1 transformed with a plasmid containing int and orf 203 showed resistance to superinfection by heterologous phages, but not by the homologous f Sfi21. Part of the int gene could be deleted without loss of this activity, while a deletion in orf 203 resulted in loss of the phage resistance. We speculate on the possibility of a bipartite immunity system for the control of lysogeny in f Sfi21. [TOP OF PAGE]

157. Molecular ecology of Streptococcus thermophilus bacteriophage infections in a cheese factory. Bruttin, A., Desiere, F., d'Amico N, Guerin JP, Sidoti J, Huni B, Lucchini S, Brussow, H. (1997). Appl. Environ. Microbiol. 63:3144-3150. A mozzarella cheese factory using an undefined, milk-derived Streptococcus thermophilus starter system was monitored longitudinally for 2 years to determine whether the diversity of the resident bacteriophage population arose from environmental sources or from genetic changes in the resident phage in the factory. The two hypotheses led to different predictions about the genetic diversity of the phages. With respect to host range, 12 distinct phage types were observed. With two exceptions, phages belonging to different lytic groups showed clearly distinct restriction patterns and multiple isolates of phages showing the same host range exhibited identical or highly related restriction patterns. Sequencing studies in a conserved region of the phage genome revealed no point mutations in multiple isolates of the same phage type, while up to 12% nucleotide sequence diversity was observed between the different phage types. This diversity is as large as that between the most different sequences from phages in our collection. These observations make unlikely a model that postulates a single phage invasion event and diversification of the phage during its residence in the factory. In the second stage of our factory study, a defined starter system was introduced that could not propagate the resident factory phage population. Within a week, three new phage types were observed in the factory while the resident phage population was decreased but not eliminated. Raw milk was the most likely source of these new phages, as phages with identical host ranges and restriction patterns were isolated from raw milk delivered to the factory during the intervention trial. Apparrently, all of the genetic diversity observed in the S. thermophilus phages isolated during our survey was already created in their natural environment. A better understanding of the raw-milk ecology of S. thermophilus phages is thus essential for successful practical phage control. [TOP OF PAGE]

158. Parallel molecular evolution of deletions and nonsense mutations in bacteriophage T7 [letter]. Cunningham, C.W., Jeng, K., Husti, J., Badgett, M.R., Molineux, I.J., Hillis, D.M., Bull, J.J. (1997). Molecular Biology and Evolution 14:113-116. [TOP OF PAGE]

159. A highly conserved DNA replication module from Streptococcus thermophilus phages is similar in sequence and topology to a module from Lactococcus lactis phages. Desiere, F., Lucchini S, Bruttin, A., Zwahlen MC, Brussow, H. (1997). Virology 234:372-382. A highly conserved DNA region extending over 5 kb was observed in Streptococcus thermophilus bacteriophages. Comparative sequencing of one temperate and 26 virulent phages demonstrated in the most extreme case an 18% aa difference for a predicted protein, while the majority of the phages showed fewer, if any aa changes. The relative degree of aa conservation was not homogeneous over the DNA segment investigated. Sequence analysis of the conserved segment revealed genes possibly involved in DNA transactions. Three predicted proteins (orf 233, 443, and 382 gene product (gp)) showed nucleoside triphosphate binding motifs. Orf 443 gp showed in addition a DEAH box motif, characteristically found in a subgroup of helicases, and a variant zinc finger motif known from a phage T7 helicase/primase. Tree analysis classified orf 443 gp as a distant member of the helicase superfamily. Orf 382 gp showed similarity to putative plasmid DNA primases. Downstream of orf 382 a noncoding repeat region was identified that showed similarity to a putative minus origin from a cryptic S. thermophilus plasmid. Four predicted proteins showed not only high degrees of aa identity (34 to 63%) with proteins from Lactococcus lactis phages, but their genes showed a similar topological organization. We interpret this as evidence for a horizontal gene transfer event between phages of the two bacterial genera in the distant past. [TOP OF PAGE]

160. Intracellular kinetics of a growing virus: A genetically-structured simulation for bacteriophage T7. Endy, D., Kong, D., Yin, J. (1997). Biotechnology and Bioengineering 55:375-389. Viruses have evolved to efficiently direct the resources of their hosts toward their own reproduction. A quantitative understanding of viral growth will help researchers develop antiviral strategies, design metabolic pathways, construct vectors for gene therapy, and engineer molecular systems that self-assemble. As a model system we examine here the growth of bacteriophage T7 in Escherichia coli using a chemical-kinetic framework. Data published over the last three decades on the genetics, physiology, and biophysics of phage T7 are incorporated into a genetically structures simulation that accounts for entry of hte T7 genome into its host, expression of T7 genes, replication of T7 DNA, assembly of T7 procapsids, and packaging of T7 DNA to finally produce intact T7 progeny. Good agreement is found between the simulated behavior and experimental observations for the shift in transcription capacity from the host to the phage, the initation times of phage protein synthesis, and the intracellular assembly of both wild-type phage and a fast-growing deletion mutant. The simulation is utilized to predict the effect of antisense molecules targetted to different T7 mRNA. Further, a postulated mechanism for the down regulation of T7 transcription in vivo is quantitatively examined and shown to agree with available data. The simulation is found to be a useful tool for exploring and understanding the dyanamics of virus growth at the molecular level. [TOP OF PAGE]

161. Polymorphism of bacteriophage T7. Gabashvili, I.S., Khan, S.A., Hayes, S.J., Serwer, P. (1997). J. Mol. Biol. 273:658-667. For viruses made of nucleic acid and protein, the structure of the protein outer shell has, in the past, been found to be uniquely determined by the viral genome. However, here, non-denaturing agarose gel electrophoresis of bacteriophage T7 reveals two states of the mature T7 capsid; the conditions of growth are found to alter the population by T7 of these two electrophoretically defined states. Both states have been previously observed for a genetically altered T7 and they are observed here for wild-type T7. The average electrical surface charge density of a bacteriophage particle (delta) determines its state; the delta of particles in both states is negative. For a given condition of growth, the population of these two states is influenced by the extent to which the major T7 outer shell protein, p10A, is accompanied by its minor readthrough variant, p10B. Comparison of the two electrophoretic states reveals the following. (1) No difference in radius is present in the outer shell (+/-2%). (2) As the pH of electrophoresis is either increased or decreased from neutrality, the state becomes more highly populated for which delta is greater in magnitude (state 1). By changing the pH, some T7 particles are made to change state. (3) Particles in state 1 adsorb less quickly to host cells than do the particles in the alternative state (state 2). This latter observation suggests the hypothesis that state 1 evolved to reduce the probability of re-initiating an infection when conditions are not favorable for growth. This hypothesis is supported by the observation that, as conditions of growth become apparently more unfavorable, progeny increasingly populate state 1. [TOP OF PAGE]

162. The abundance of planktonic viruses in antarctic lakes. Kepner, R., Galchenko, V., Wharton, R. (1997). pp. 241-252. In In Lyons, W.B., Howard-Williams, C., and Hawes, I. (eds.), Ecosystem Processes in Antarctic Ice-Free Landscapes. Balkema Press, Rotterdam. [TOP OF PAGE]

163. Two groups of bacteriophages infecting Streptococcus thermophilus can be distinguished on the basis of mode of packaging and genetic determinants for major structural proteins. Le Marrec, C., van Sinderen, D., Walsh, L., Stanley, E., Vlegels, E., Moineau, S., Heinze, P., Fitzgerald, G., Fayard, B. (1997). Appl. Environ. Microbiol. 63:3246-3253. A comparative study of 30 phages of Streptococcus thermophilus was performed based on DNA restriction profiles, DNA homology, structural proteins, packaging mechanisms, and host range data. All phages exhibited distinct DNA restriction profiles, with some phages displaying similarly sized restriction fragments. DNA homology was shown to be present among all 30 phages. The phages could be divided into two groups on the basis of their packaging mechanism as was derived from the appearance of submolar DNA fragments in restriction enzyme digests and the presence (cos-containing phages) or absence (pac-containing phages) of cohesive genomic extremities. Interestingly, the 19 identified cos-containing phages possessed two major structural proteins (32 and 26 kDa) in contrast to the remaining 11 pac-containing phages, which possessed three major structural proteins (41, 25, and 13 kDa). Southern hybridization demonstrated that all pac-containing phages tested contain homologs of the genes encoding the three major structural proteins of the pac-containing phage O1205, whereas all cos-containing phages tested exhibit homology to the gene specifying one of the structural components of the cos-containing phage phi 7201. Fifty-seven percent of the phages (both cos and pac containing) possessed the previously identified 2.2-kb EcoRI fragment of the temperate S. thermophilus phage Sfi18 (H. Brussow, A. Probst, M. Fremont, and J. Sidoti, Virology 200:854-857, 1994). No obvious correlation was detected between grouping based on packaging mechanism and host range data obtained with 39 industrial S. thermophilus strains. [TOP OF PAGE]

164. Antiserum inhibition of propagating viruses. Lee, Y., Eisner, S.D., Yin, J. (1997). Biotechnology & Bioengineering 55:542-546. The design and implementation of controlled environments to continuously culture and evolve viruses provides a means to track how their populations respond to natural and designed anti-viral agents. We have previously demonstrated how the growth of viruses in spreading plaques enables detection and characterization of their evolutionary dynamics. Using plaques of phage T7 growing on E. coli as a model system, we observe here that velocities of propagation can be readily controlled by the level of anti-viral antiserum incorporated into the propagation medium. Further, we develop a simple analytic expression for the radial velocity of propagation in terms of the microscopic rates of viral amplification, Fickian diffusion of the virions and their neutralization by antiserum. Our analysis captures the essential dependence of propagation velocity on antiserum concentration. This study provides an ex vivo foundation for exploring how medically relevant viruses escape suppression by the immune system. [TOP OF PAGE]

165. Genetic diversity in temperate bacteriophages of Streptococcus pyogenes: Identification of a second attachment site for phages carrying the erythrogenic toxin A gene. McShan, W.M., Ferretti, J.J. (1997). J. Bacteriol. 179:6509-6511. Bacteriophage T12, the prototypic bacteriophage of Streptococcus pyogenes carrying the erythrogenic toxin A gene (speA), integrates into the bacterial chromosome at a gene for a serine tRNA (W. M. McShan, Y.-F. Tang, and J. J. Ferretti, Mol. Microbiol. 23:719-728, 1997). This phage is a member of a group of related temperate phages, and we show here that not all speA-carrying phages in this group use the same attachment site for integration into the bacterial chromosome. Additionally, other phages in the group use the same serine tRNA gene attachment site as phage T12 and yet do not carry speA. The evidence suggests that recombination between phage genomes has been an important means of generating diversity and disseminating virulence-associated genes like speA. [TOP OF PAGE]

166. Genetic studies of erythrogenic toxin carrying temperate bacteriophages of Streptococcus pyogenes. McShan, W.M., Ferretti, J.J. (1997). Advances in Experimental Medicine & Biology 418:971-973. [TOP OF PAGE]

167. A novel mechanism of virus-virus interactions: bacteriophage P2 Tin protein inhibits phage T4 DNA synthesis by poisoning the T4 single-stranded DNA binding protein, gp32. Mosig, G., Yu, S., Myung, H., Haggard-Ljungquist, E., Davenport, L., Carlson, K., Calendar, R. (1997). Virology 230:72-81. P2 prophages have been known to inhibit DNA replication and growth of T-even phages. We show here that this inhibition is due to poisoning of the T-even single-stranded DNA binding protein gp32 by the product of the nonessential P2 tin gene. Synthesis of Tin protein from a gene cloned in a multicopy plasmid is necessary and sufficient to completely prevent de novo DNA replication and growth of wild-type T2 or T4 phage. We isolated more than 20 independent mutants that render T-even phages resistant to poisoning by the P2 Tin protein. In all of these mutants, which we call asp, Asp codon 163 of gene 32 is changed to a Gly or Asn codon. The mutant alleles are recessive; i.e., when wild-type and asp mutants coinfect the same host cells, most DNA replication is poisoned by P2 Tin protein. To explain our results, we propose that the P2 Tin protein interacts with T-even gp32 at position 163 and distorts the helical filament of gene 32 protein on single-stranded DNA. Thereby Tin protein inhibits either assembly or function, or both, of the T4 replisome. The inhibition of late gene expression by P2 Tin protein may be an indirect consequence of inhibition of DNA replication. [TOP OF PAGE]

168. Virus decay and its cause in coastal waters. Noble, R.T., Fuhrman, J.A. (1997). Appl. Environ. Microbiol. 63:77-83. Mesocosms filled with 80 liters of coastal seawater from Santa Monica, California, were used twice (June and November) to budget bacterial production and loss, as well as to assess the relative significance of viral lysis and protist grazing in bacterial mortality. Bacterial abundance was similar to 6 x 10 super(9) cells/liter in June and 2 x 10 super(9) in November, with viral abundances similar to 2 x 10 super(10) particles/liter in June and 1.5 x 10 super(10) in November. Incorporation of [ super(3)H]thymidine and leucine yielded essentially identical production estimates and allowed calculation on total bacterial mortality in these closed systems. Bacterial growth rates were 1-2/d in June and 1-3/d in November. Three independent lines of evidence indicated that bacterial mortality attributed to grazing by protists was about equal to that attributed to viruses: size fractionation of disappearance of labeled DNA, with a 50% reduction after protists were removed; comparison of protist grazing rates estimated with fluorescently labeled bacteria and virus production-based bacterial lysis rates, with 40-50% of the total ascribed to viruses; and model-based interpretation of the 3.3-4.6% of bacteria visibly infected with assembled intracellular viruses, suggesting that 24-66% of loss is due to infection. Redundant production and loss measurements as well as the independent loss process estimates agreed within similar to 30%, yielding a reasonably balanced budget. We believe the loss of bacteria to viruses reflects a significant dissipation of energy in this ecosystem and that viruses and protists contribute similarly to bacterial mortality. [TOP OF PAGE]

169. Coliphage and indigenous phage in Mamala Bay, Oahu, Hawaii. Paul, J.H., Rose, J.B., Jiang, S.C., London, P., Xhou, X., Kellogg, C. (1997). Appl. Environ. Microbiol. 63:133-138. Public concern over the discharge of primarily treated sewage by two offshore outfalls in Mamala Bay, Oahu, prompted a multidisciplinary study to determine the impact of such activities on the water quality in the bay and at adjacent recreational beaches. As part of this study, we determined the abundance of coliphage as an indicator of fecal pollution along with total viral direct counts and phages infective for Vibrio parahaemoltyicus 16 at stations in Mamala Bay in four quarterly samplings over 13 months. Coliphage ( lt 1 to 1.2 times 10^-3/liter) were found during each quarterly sampling along an offshore transect to the Sand Island waste treatment facility outfall. The nonpoint coastal stations (Pearl Harbor, Ala Wai Canal, and Ke'ehi Lagoon) had high levels of coliphage during the storm event sampling in February 1994 but much lower levels or none when sampled during dry weather. Coliphage were absent at all samplings at Waikiki Beach and at the control station off Diamond Head. Viral direct counts in eutrophic coastal stations (Pearl Harbor, Ke'ehi Lagoon, Ala Moana Beach, and Ala Wai canal) averaged 10^-9/liter, while counts at offshore stations ranged from 9 times 10^-7 to 1 times 10^-9 viruses/liter, values similar to those for other marine environments. Vibriophage were found mainly in eutrophic coastal environments (Ala Wai Canal, Pearl Harbor, and Ke'ehi Lagoon) and at the Sand Island Transect stations D1 and D2. The greatest abundance was found during the storm event (February 1994) sampling. These results suggest that the Sand Island outfall influenced the water quality of the immediate surrounding waters but had little effect on the quality of the recreational beaches. Nonpoint discharge sources appeared to be more important in the distribution of fecal indicators in the coastal zone. [TOP OF PAGE]

170. Evidence for groundwater and surface marine water contamination by waste disposal wells in the Florida Keys. Paul, J.H., Rose, J.B., Jiang, S.C., Xhou, X., Cochran, P., Kellogg, C., Kang, J.B., Griffin, D., Farrah, S., Lukasik, J. (1997). Water Res. 31:1448-1454. [TOP OF PAGE]

171. Comparison of PCR and plaque assay for detection and enumeration of coliphage in polluted marine waters. Rose, J.B., Zhou, X., Griffin, D.W., Paul, J.H. (1997). Appl. Environ. Microbiol. 63:4564-4566. [TOP OF PAGE]

172. Photoreactivation compensates for UV damage and restores infectivity to natural marine virus communities. Weinbauer, M.G., Wilehelm, S.W., Suttle, C.A., Garza, D.R. (1997). Appl. Environ. Microbiol. 63:2200-2205. We investigated the potential for photoreactivation to restore infectivity to sunlight-damaged natural viral communities in offshore (chlorophyll a, lt 0.1 mu-g liter-1), coastal (chlorophyll a, ca. 0.2 mu-g liter-1), and estuarine (chlorophyll a, ca. 1 to 5 mu-g liter-1) waters of the Gulf of Mexico. In 67% of samples, the light-dependent repair mechanisms of the bacterium Vibrio natriegens restored infectivity to natural viral communities which could not be repaired by light-independent mechanisms. Similarly, exposure of sunlight-damaged natural viral communities to gt 312-nm-wavelength sunlight in the presence of the natural bacterial communities restored infectivity to 21 to 26% of sunlight-damaged viruses in oceanic waters and 41 to 52% of the damaged viruses in coastal and estuarine waters. Wavelengths between 370 and 550 nm were responsible for restoring infectivity to the damaged viruses. These results indicate that light-dependent repair, probably photoreactivation, compensated for a large fraction of sunlight-induced DNA damage in natural viral communities and is potentially essential for the maintenance of high concentrations of viruses in surface waters. [TOP OF PAGE]

173. Comparison of epifluorescence and transmission electron microscopy for counting viruses in natural marine waters. Weinbauer, M.G., Suttle, C.A. (1997). Aquat. Microb. Ecol. 13:225-232. [TOP OF PAGE]

174. Lysogenic and lytic viral production in marine microbial communities. Wilson, W.H., Mann, N.H. (1997). Aquat. Microb. Ecol. 13:95-100. It is now well established that viruses are an abundant component of marine ecosystems and they are being increasingly recognised and accepted as important contributors to element cycling within the microbial loop. However, some of the key questions regarding the ecological significance of viruses in the marine environment still remain largely unanswered. Thus, particular interest is currently focused on the extent to which lytic production or lysogeny predominates and the nature of factors in the marine environment, particularly nutrient availability and multiplicity of infection (MOI), which might influence the lysis/lysogeny 'decision'. The present evidence is still insufficient to unambiguously assess the relative ecological significance of lysogeny versus lysis and progress in this area will rely on the development and application of new techniques. This review attempts to collect recent information relating to this central question, focusing particularly on those viruses which infect the bacterioplankton and nano- and picophytoplankton. [TOP OF PAGE]

175. Frequency of morphological phage descriptions in 1995. Ackermann, H.-W. (1996). Archives of Virology 141:209-218. At least 4500 bacterial viruses have been examined in the electron microscope since 1959. About 4400 phages (96%) are tailed and only 162 phages (4%) are cubic, filamentous, or pleomorphic. Phages belong to 12 virus families and occur in about 130 bacterial genera. Phages are listed by morphotypes and host genera. Siphoviridae or phages with long, noncontractile tails include about 60% of tailed phages. [References: 21]. [TOP OF PAGE]

176. Selection of the phage-resistant mutant strains of E.coli M 17 and study of their main biological properties. Chanishvili, N., Tediashvili, M., Eliashvili, T., Zviadadze, N., Porchkhidze, K., Alavidze, Z., Giorkhelidze, D., Kvatadze, N., Natroshvili, G., Giorkhelidze, R., Adamia, R., Chanishvili, T. (1996). Proceedings of the Georgian Academy of Sciences 22:203-209. [TOP OF PAGE]

177. Genetic diversity in marine algal virus communities as revealed by sequence analysis of DNA polymerase genes. Chen, F., Suttle, C.A., Short, S.M. (1996). Appl. Environ. Microbiol. 62:2869-2874. Algal-virus-specific PCR primers were used to amplify DNA polymerase gene (pol) fragments (683 to 689 bp) from the virus-sized fraction (0.02 to 0.2 mu m) concentrated from inshore and offshore water samples collected from the Gulf of Mexico. Algal-virus-like DNA pol genes were detected in five samples collected from the surface and deep chlorophyll maximum. PCR products from an offshore station were cloned, and the genetic diversity of 33 fragments was examined by restriction fragment length polymorphism and sequence analysis. The five different genotypes or operational taxonomic units (OTUs) that were identified on the basis of restriction fragment length polymorphism banding patterns were present in different relative abundances (9 to 34%). One clone from each OTU was sequenced, and phylogenetic analysis showed that all of the OTUs fell within the family Phycodnaviridae. Four of the OTUs fell within a group of viruses (MpV) which infect the photosynthetic picoplankter Micromonas pusilla. The genetic diversity among these genotypes was as large as that previously found for MpV isolates from different oceans. The remaining genotype formed its own clade between viruses which infect M. pusilla and Chrysochromulina brevifilum. These results imply that marine Virus communities contain a diverse assemblage of MpV-like viruses, as well as other unknown members of the Phycodnaviridae. [TOP OF PAGE]

178. Evolutionary relationships among large double-stranded DNA viruses that infect microalgae and other organisms as inferred from DNA polymerase genes. Chen, F., Suttle, C.A. (1996). Virology 219:170-178. In order to examine genetic relatedness among viruses that infect microalgae, DNA polymerase gene (DNA pol) fragments were amplified and sequenced from 13 virus clones that infect three genera of distantly related microalgae (Chlorella strains NC64A and Pbi, Micromonas pusillia and Chrysochromulina spp.). Phylogenetic trees based on DNA pol sequences and hybridization of total genomic DNA showed similar branching patterns. Genetic relatedness calculated from the hybridization and sequence data showed good concordance (r = 0.90), indicating that DNA pol sequences can be used to determine genetic relatedness and infer phylogenetic relationships among these viruses. The phylogenetic tree inferred from the deduced amino acid sequences of DNA pol from 24 dsDNA viruses, including phycodnaviruses, herpesviruses, poxviruses, baculoviruses, and African swine fever virus corresponded well with groupings based on the International Committee on Taxonomy of Viruses. Microalgal viruses are more closely related to each other than to the other dsDNA viruses and form a distinct phyletic group, suggesting that they share a common ancestor and belong to the Phycodnaviridae. Moreover, the Phycodnaviridae are more closely related to the Herpesviridae than to other virus families for which DNA pol sequences are available. [TOP OF PAGE]

179. Study of biological properties of CB phages isolated from the cells of the industrial strain E. coli M17. Eliashvili, T., Tediashvili, M., Chanishvili, N., Zviadadze, N., Kvachadze, L., Adamia, R., Chanishvili, T. (1996). Selected articles, G. Eliava Institute of BMV 9:152-154. [TOP OF PAGE]

180. Influence of the contaminating phages on the propagation process of the industrial strain E-coli M17, held in industrial conditions. Goderdzishvili, M., Kvatadze, N., Eliashvili, T., Tediashvili, M., Chanishvili, N., Zviadadze, N., Chanishvili, T. (1996). Selected articles, G. Eliava Institute of BMV 9:56-59. [TOP OF PAGE]

181. A moderately halophilic Vibrio from a Spanish saltern and its lytic bacteriophage. Goel, U., Kauri, T., Ackermann, H.W., Kushner, D.J. (1996). Canadian Journal of Microbiology 42:1015-1023. A number of bacteria and their phages were isolated from a saltern near Alicante, Spain. One isolate, Vibrio B1, a moderate halophile that is probably a strain of Vibrio costicola, was host to a lytic phage, UTAK. Studies of the host bacterium included the effects of salt concentrations on the action of a number of inhibitory agents. Phage UTAK has a head, a tail, and a baseplate. It contains 80 kbp of double-stranded DNA with no unusual bases. It was stable for long periods in the absence of high salt concentrations and even in distilled water. Salt concentrations had little effect on adsorption of UTAK to its host but resulted in considerable changes in burst size. It appears that phages of halophilic and salt-tolerant eubacteria, and also of some marine bacteria, have much lower salt requirements for stability than the phages of halophilic archaebacteria. Our results suggest that ionic controls of phage replication in these eubacteria may differ from those of growth. [TOP OF PAGE]

182. Occurence of lysogenic bacteria in marine microbial communities as determined by prophage induction. Jiang, S.C., Paul, J.H. (1996). Mar. Ecol. Prog. Ser. 142:27-38. [TOP OF PAGE]

183. Selection and properties of totally phage-resistant mutant Pseudomonas putida PpG1. Krylov, V.N., Rasskazchikova, S.A., Al'nikin, A.F. (1996). Genetika 32:348-353. The efficiency of using bacteria in open systems to degrade different anthropogenic toxic pollutants can depend strongly on the interaction between these bacteria and natural bacteriophages. The possibility of selecting bacterial Pseudomonas putida mutants resistant to all bacteriophages of this species known so far was tested (in our work, these mutants were designated totally phage-resistant mutants). In a model experiment, changes in the composition of a population upon prolonged growth of bacteria in the presence of one of the virulent phages were examined. On the basis of the results obtained, it is postulated that: (1) Mutants differing in resistance to various phages accumulate in a population; relative numbers of different mutants can undergo alterations over the course of time; mutants selected in the presence of a given virulent phage do not often manifest complete resistance to this phage. (2) It is possible to isolate totally phage-resistant mutants of P. putida PpG1. These mutants carry up to three different mutations simultaneously; however, these mutants regain sensitivity to many phages upon pseudoreversion occurrence. [TOP OF PAGE]

184. Evolution of T4-related phages. Kutter, E., Gachechiladze, K., Poglazov, A., Marusich, E., Shneider, M., Aronsson, P., Napuli, A., Porter, D., Mesyanzhinov, V. (1996). Virus Genes 11:285-297. Much progress has been made in understanding T-even phage biology in the last 50 years. We now know the entire sequence of T4, encoding nearly 300 genes, only 69 of which have been shown to be essential under standard laboratory conditions; no specific function is yet known for about 140 of them. The origin of most phage genes is unclear, and only 42 genes in T4 have significant similarity to anything currently included in GenBank. Comparative analysis of related phages is now being used to gain insight into both the evolutionary origins and interrelationships of these phage genes, and the functions of their protein products. The genomes of phages isolated from Tbilisi hospitals, Long Island sewage plants, the Denver zoo, and Khabarovsk show basic similarity. However, these phages show substantial insertions and deletions in a number of regions relative to each other, and closer investigation of specific sequences often reveals much more complex relationships. There are only a few cases in T4- related phages in which there is evidence for evolution through DNA duplication. These include the fibrous products of genes 12, 34, and 37; head proteins gp23 and gp24; and the Alt enzyme and its downstream neighbors. T4 also contains 13 apparent relatives of group I and group II intron homing endonucleases. Distal portions of the tail fibers of various T-even phages contain segments closely related total-fiber regions of other DNA coliphages, such as Mu, P1, P2, and lambda. Horizontal gene transfer clearly emerges as a major factor in the evolution of at least the tail-fiber regions, where site-specific recombination probably is involved in the exchange of host-range determinants. [TOP OF PAGE]

185. Detection of evolving viruses. Lee, Y., Yin, J. (1996). Nature Biotechnology 14:491-493. The spread of viruses on a homogeneous lawn of receptive hosts provides an opportunity to detect the dynamics of their evolution. We have previously found that when repeated virus passages are confined to the expanding perimeter of a growing plaque, the appearance and outgrowth of genetically diverse strains (all descended from the same parent strain) can be traced along different radii of the plaque. As a plaque grows, the random mutation and selection of new fast-growing strains reduce the roundness or circularity of the growing plaque. Here we have quantified such changes in growing plaques of bacteriophage T7 using a digital imaging system. We find that T7 populations not adapted for fast growth exhibit a broader diversity of growth rates than populations adapted for fast growth. These results provide a foundation for understanding how viruses exploit mutation and selection processes to persist in nature. [TOP OF PAGE]

186. Imaging the propagation of viruses. Lee, Y., Yin, J. (1996). Biotechnology and Bioengineering 52:438-442. The propagation of viruses in a growing plaque has been measured using a digital image acquisition and analysis system. Plaques of phage T7 incubated at 37°C and illuminated against a dark field emerged as dark growing spots against a background of host bacteria. Images of the growth were acquired using a charge-coupled device (CCD) camera at 1-h intervals over 24 h. The first 10 h of plaque development coincided with rapid growth of the agar-immobilized Escherichia coli host, measured as a reduction in gray value. Following this period, the average radial velocity of plaque growth remained constant at 0.059 mm/h while the standard deviation about this velocity increased. These results suggest the suitability of the system for spatially resolving the dynamics of viral evolution during plaque growth. [TOP OF PAGE]

187. Phage therapy revisited: the population biology of a bacterial infection and its treatment with bacteriophage and antibiotics. Levin, B.R., Bull, J.J. (1996). Am. Nat. 147:881-898. Phage therapy is the use of bacterial viruses (bacteriophage) to treat bacterial infections. It has been practiced sporadically on humans and domestic animals for nearly 75 yr. Nevertheless, phage therapy has remained outside the mainstream of modern medicine, presumably because of doubts about its efficacy, and possibly because it was eclipsed by antibiotics and other chemotherapeutic agents. In this report, we develop the study of phage therapy and antibiotic therapy as a population biological phenomenon-the dynamic interaction of bacteria with a predator (phage) or a toxic chemical (antibiotic) inside a host whose immune and other defenses also affect the interaction. Our goal is to identify the conditions under which phage and antibiotics can successfully control a bacterial infection and when they cannot. We review data published in the 1980s by H. Williams Smith and J. B. Huggins on the use of phage and antibiotics to control lethal, systemic infections of Escherichia coli in experimentally inoculated mice. We show that some of their observations can be accommodated by a quantitative model that invokes known or plausible assumptions about host defenses and the interactions of bacteria with phage and antibiotics; some observations remain unexplained by the model. Our analysis identifies several hypotheses about the population dynamics of phage and antibiotic therapy that can be tested experimentally. Included among these are hypotheses that account for variation in the efficacy of the different phages employed by Smith and Huggins and why, in their study, phages were more effective than antibiotics. [TOP OF PAGE]

188. Long-circulating bacteriophage as antibacterial agents. Merril, C.R., Biswas, B., Carlton, R., Jensen, N.C., Creed, G.J., Zullo, S., Adhya, S. (1996). Proc. Natl. Acad. Sci. USA 93:3188-3192. The increased prevalence of multidrug-resistant bacterial pathogens motivated us to attempt to enhance the therapeutic efficacy of bacteriophages. The therapeutic application of phages as antibacterial agents was impeded by several factors: (i) the failure to recognize the relatively narrow host range of phages; (ii) the presence of toxins in crude phage lysates; and (iii) a lack of appreciation for the capacity of mammalian host defense systems, particularly the organs of the reticuloendothelial system, to remove phage particles from the circulatory system. In our studies involving bacteremic mice, the problem of the narrow host range of phage was dealt with by using selected bacterial strains and virulent phage specific for them. Toxin levels were diminished by purifying phage preparations. To reduce phage elimination by the host defense system, we developed a serial- passage technique in mice to select for phage mutants able to remain in the circulatory system for longer periods of time. By this approach we isolated long-circulating mutants of Escherichia coli phage lambda and of Salmonella typhimurium phage P22. We demonstrated that the long- circulating lambda mutants also have greater capability as antibacterial agents than the corresponding parental strain in animals infected with lethal doses of bacteria. Comparison of the parental and mutant lambda capsid proteins revealed that the relevant mutation altered the major phage head protein E. The use of toxin-free, bacteria-specific phage strains, combined with the serial-passage technique, may provide insights for developing phage into therapeutically effective antibacterial agents. [TOP OF PAGE]

189. Evolution of the genetic switch in temperate bacteriophage. I. Basic theory. Mittler, J.E. (1996). J. Theor. Biol. 179:161-172. While the molecular mechanisms underlying lysogeny and induction in bacteriophage have been intensely studied, relatively little has been done to relate these findings to their presumed selective functions. To explore the ecological basis for these traits, I have used a resource-based model for competition between bacteriophage with different probabilities of lysogeny and different spontaneous induction rates. In any given habitat the fitness of a phage will depend on the inputs of sensitive cells and nutrient resources. In equable environments (modeled here using chemostats with constant inputs of nutrients and sensitive cells), bacteriophage with low probabilities of lysogeny and low induction rates can always invade when rare and will generally be good competitors. In variable environments (chemostats with seasonal inputs), bacteriophage with higher probabilities of lysogeny and higher induction rates are favored. In both equable and variable environments, the ability of a phage to invade when rare will depend on the properties of the resident phage, and it is possible for phages with divergent parameter values to coexist. The modeling suggests that bacteriophage that have evolved moderately low induction and lysogeny rates will be able to "hedge their bets" against environmental change without sacrificing the ability to compete well in a constant environment. Implications of this theory for understanding the molecular basis of gene regulation in temperate bacteriophage and other viruses are discussed. [TOP OF PAGE]

190. Isolation and characterization of lactococcal bacteriophages from cultured buttermilk plants in the United States. Moineau, S., Borkaev, M., Holler, B.J., Walker, S.A., Kondo, J.K., Vedamuthu, E.R., Vandenbergh, P.A. (1996). Journal of Dairy Science 79:2104-2111. From July 1993 and June 1994, 27 different lactococcal bacteriophages were isolated from 27 US cultured buttermilk plants located in 23 states. Phages were characterized by DNA homology, electron microscopy, restriction patterns, genome size, host range, and serology. Over 80% (22 of 27) of the phages were classified into the 936 species, and the remaining phages were divided almost equally between the P335 species (3 of 27) and the c2 species (2 of 27). The 936 and c2-type phages had the same basic morphological and genetic characteristics as other phages from the same species isolated in other countries. Very closely related 936 phages were isolated from widely separated areas in the US. The P335 phages had a very narrow host range and showed noticeable genetic and immunological diversity. None of the phages could propagate on the two exopolysaccharide-producing Lactococcus lactis strains tested. Novel mechanisms for phage resistance should be tested for efficiency against members of the lactococcal phage species 936, c2, and P335. To our knowledge, this study is the first thorough examination of industrial lactococcal phages isolated from buttermilk plants. [TOP OF PAGE]

191. AbiG, a genotypically novel abortive infection mechanism encoded by plasmid pCI750 of Lactococcus lactis subsp. cremoris UC653. O'Connor, L., Coffey, A., Daly, C., Fitzgerald, G. (1996). Appl. Environ. Microbiol. 62:3075-3082. AbiG is an abortive infection (Abi) mechanism encoded by the conjugative plasmid pCI750 originally isolated from Lactococcus lactis subsp. cremoris UC653. Insensitivity conferred by this Abi manifested itself as complete resistance to vphi-712 (936 phage species) with only partial resistance to vphi-c2 (c2 species). The mechanism did not inhibit phage DNA replication. The smallest subclone of pCI750 which expressed the Abi phenotype contained a 3.5-kb insert which encoded two potential open reading frames. abiGi (750 bp) and abiGii (1,194 bp) were separated by 2 bp and appeared to share a single promoter upstream of abiGi. These open reading frames showed no significant homology to sequences of either the DNA or protein databases; however, they did exhibit the typical low G+C content (29 and 27%, respectively) characteristic of lactococcal abi genes. In fact, the G+C content of a 7.0-kb fragment incorporating the abiG locus was 30%, which may suggest horizontal gene transfer from a species of low G+C content. In this context, it is notable that remnants of IS elements were observed throughout this 7.0-kb region. [TOP OF PAGE]

192. Viruses and DNA in marine environments. Paul, J.H., Kellogg, C.A., Jiang, S.C. (1996). pp. 115-124. In In Colwell, R.R., Simidu, U., and Ohwada, K. (eds.), Microbial Diversity in Time and Space. Plenum Press, New York. [TOP OF PAGE]

193. Host-parasite persistence: the role of spatial refuges in stabilizing bacteria-phage interactions. Schrag, S., Mittler, J.E. (1996). Am. Nat. 148:348-347. We manipulated a bacteria-phage model system to investigate empirical and theoretical conditions allowing for the coexistence of an Escherichia coli host with each of two virulent phage species, a T1-like phage (T1X) and lambda-vir. In minimal medium in the laboratory, bacteria coexisted with each phage species in continuous (chemostat) culture; however, in serial culture, T1X rapidly became extinct in all cases and lambda-vir became extinct in most cases. When we refined a previously developed mechanistic model of bacteria-phage interactions in continuous culture, this model failed to predict our laboratory observations of long-term stability of bacteria and phage in chemostats. A serial transfer version of this model, however, came much closer to predicting bacteria-phage dynamics. To investigate why models of continuous culture failed, we tested hypotheses for phage persistence by manipulating experimental culture conditions. We found that wall populations of bacteria strongly influenced the stability of both phage species. When wall populations were not allowed to develop in chemostats, both phage species became extinct rapidly; conversely, when wall populations were allowed to develop in serial transfer, both phage species showed evidence of long-term persistence. The final percentage of sensitive bacteria was significantly higher in walls than in liquid populations, suggesting that glass surfaces did indeed act as a spatial refuge for sensitive bacteria. In addition, serial transfer of bacteria and phage on agar surfaces produced more stable interactions than in liquid. We discuss these results in light of previous observations of bacteria-phage interactions and in the broader context of parasite-host and predator-prey coexistence conditions. [TOP OF PAGE]

194. Abundance and production of bacteria and viruses in the Bering and Chukchi sea. Steward, G.F., Smith, D.C., Azam, F. (1996). Mar. Ecol. Prog. Ser. 131:287-300. The distribution, abundance, and production of viruses and bacteria were investigated during an August to September 1992 cruise aboard the RV 'Alpha Helix' in the Bering and Chukchi Seas. Viruses were abundant in seawater samples at all stations (10⁹ to 10¹⁰ l^-1) and exceeded the bacteria concentration by an order of magnitude on average. Virus-like particles and bacteria were also observed in the pore water of a sediment sample at 27 and 2.1 ´ 10⁹ l^-1, respectively. The concentrations of viruses and bacteria in pelagic samples were correlated (r = 0.83, n = 43). In a detailed depth profile from the deepest and northern most station (72° N), bacteria and viruses displayed subsurface maxima in the upper 100 m. Below 100 m, the concentrations declined, but were detectable even in the deepest-collected samples (402 m). Integrated bacterial biomass estimates were similar to results from a previous study in this area, but bacterial production measurements ranging from 0.3 to 0.45 g C m^-2 d^-1 were an order of magnitude higher. Production rates of bacterial viruses (also known as bacteriophages or simply phages) measured by radiolabeling ranged from 0.5 to 4.2 ´ 10⁹ viruses l^-1 d^-1, which are similar to previous estimates for temperate coastal waters. The production measurements indicated turnover times ranging from 0.4 to 17 d for bacteria and maximum estimates of 1.2 to 15 d for bacterial viruses. Viral mortality of bacteria was estimated from the frequency of visibly infected cells (FVIC) and flagellate grazing was calculated from flagellate and bacterial abundances together with an assumed flagellate clearance rate. Overall, estimated viral lysis was roughly comparable to estimated grazing by flagellates as a source of bacterial mortality. Averaged over the water column, viral mortality of bacteria in the Chukchi Sea was estimated to be 23% of the bacterial production at two southern stations and approximately 10% at two northern stations. FVIC was correlated with bacterial production (r = 0.75, n=18) and specific growth rate (r = 0.74, n=18), but not with bacterial abundance (r = 0.22, n = 27). These data show viruses to be an ubiquitous and dynamic feature and a significant source of bacterial mortality in Arctic marine microbial communities. The implications of bacterial and viral production for C and N cycling in the Chukchi Sea are discussed. [TOP OF PAGE]

195. The effect of cyanophages on Synechococcus spp. during a bloom in the western Gulf of Mexico. Suttle, C.A., Chan, A.M., Rodda, K.M., Short, S.M., Weinbauer, M.G., Garza, D.R., Wilhelm, S.W. (1996). EOS 76 (suppl.):OS207-OS208 [TOP OF PAGE]

196. Viruses as biological control agents for blooms of marine phytoplankton. Suttle, C.A. (1996). pp. 71-76. In AnonymousProceedings of the Brown Tide Summit, 20-21 October, 1995. New York Sea Grant Institute???, [TOP OF PAGE]

197. Community structure: Viruses. Suttle, C.A. (1996). pp. 272-277. In In Hurst, C.J., Knudson, G.R., McInerney, M.J., Stezenbach, L.D., and Walter, M.V. (eds.), Manual of Environmental Microbiology. ASM Press, Washington DC. [TOP OF PAGE]

198. The evolution of phage lysis timing. Wang, I.-N., Dykhuizen, D.E., Slobodkin, L.B. (1996). Evol. Ecol. 10:545-558. The effect of host quantity and host quality on the evolution of phage lysis timing is analyzed using marginal value theorem of optimal foraging theory. Both factors have been shown to strongly influence the latent period. A high host density selects for short latent period, which is the same result as previous investigators have found. A good host quality also promotes short latent period. However, elasticity analysis shows that these two factors extert their influences under different sets of conditions. When host density is low, the host density is more important in determining the length of latent period, whereas when host density is high, the host quality is more important. [TOP OF PAGE]

199. Potential significance of lysogeny to bacteriophage production and bacterial mortality in coastal waters of the Gulf of Mexico. Weinbauer, M.G., Suttle, C.A. (1996). Appl. Environ. Microbiol. 62:4374-4380. The potential effect that induction of lysogenic bacteria has on bacteriophage production and bacterial mortality in coastal waters was investigated, and we present estimates for the percentage of lysogenic cells in a natural aquatic bacterial community. Various concentrations of mitomycin C and exposure times to UV C radiation (UV-C) (wavelength of 254 nm) were used to induce the lytic cycle in lysogenic cells of natural communities of marine bacteria. UV-C treatment occasionally resulted in phage production, but phage production induced by UV- C was always less than that caused by the addition of mitomycin C. There was no evidence that high growth rates of bacteria resulted in lysogenic phage production. The burst size of cells induced by mitomycin C was determined by transmission electron microscopy and ranged from 11 to 45. Dividing the induced phage production by the burst size provided an estimate of the number of lysogenic bacterial cells, which ranged from 0.07 to 4.4% (average, 1.5%) of the total bacterial population. The percentages of lysogenic bacteria that were induced by mitomycin C were similar for samples collected nearshore from the pier of the Marine Science Institute (chlorophyll a, 1.6 to 2.9 mu-g liter-1) and in relatively oligotrophic water (chlorophyll a, 0.2 to 0.9 mu-g liter-1) collected 25 to 100 km offshore. By using a steady-state model, if all lysogenic bacteria were induced simultaneously, 0.14 to 8.8% (average, 3.0%) of the total bacterial mortality would result from induction of lysogenic cells. If mitomycin C induces all or the majority of lysogenized cells, our results imply that lysogenic phage production is generally not an important source of phage production or bacterial mortality in the coastal waters of the western Gulf of Mexico. [TOP OF PAGE]

200. The effects of nutrient limitation on the kinetics of cyanophage infection of the oceanic picoplankter Synechococcus sp. WH7803. Wilson, S.H., Carr, N.G., Mann, N.H. (1996). Journal of Phycology 32:506-516. [TOP OF PAGE]

201. The effect of phosphate status on the kinetics of cyanophage infection in the oceanic cyanobacterium Synechococcus sp. WH7803. Wilson, W.H., Carr, N.G., Mann, N.H. (1996). Journal of Phycology 32:506-516. Phycoerythrin-containing Synechococcus species are considered to be major primary producers in nutrient-limited gyres of subtropical and tropical oceanic provinces, and the cyanophages that infect them are thought to influence marine biogeochemical cycles. This study begins an examination of the effects of nutrient limitation on the dynamics of cyanophage/Synechococcus interactions in oligotrophic environments by analyzing the infection kinetics of cyanophage strain S-PM2 (Cyanomyoviridae isolated from coastal water off Plymouth, UK) propagated on Synechococcus sp. WH7803 grown in either phosphate-deplete or phosphate-replete conditions. When the growth of Synechococcus sp. WH7803 in phosphate-deplete medium was followed after infection with cyanophage, an 18-h delay in cell lysis was observed when compared to a phosphate-replete control. Synechococcus sp. WH7803 cultures grown at two different rates (in the same nutritional conditions) both lysed 24 h postinfection, ruling out growth rate itself as a factor in the delay of cell lysis. One- step growth kinetics of S-PM2 propagated on host Synechococcus sp. WH7803, grown in phosphate-deplete and- replete media, revealed an apparent 80% decrease in burst size in phosphate-deplete growth conditions, but phage adsorption kinetics of S-PM2 under these conditions showed no differences. These results suggested that the cyanophages established lysogeny in response to phosphate-deplete growth of host cells. This suggestion was supported by comparison of the proportion of infected cells that lysed under phosphate-replete and-deplete conditions, which revealed that only 9.3% of phosphate-deplete infected cells lysed in contrast to 100% of infected phosphate replete cells. Further studies with two independent cyanophage strains also revealed that only approximately 10% of infected phosphate-deplete host cells released progeny cyanophages. These data strongly support the concept that the phosphate status of the Synechococcus cell will have a profound effect on the eventual outcome of phage-host interactions and will therefore exert a similarly extensive effect on the dynamics of carbon flow in the marine environment. [TOP OF PAGE]

202. Effects of sunlight on bacteriophage viability and structure. Wommack, K.E., Hill, R.T., Muller, T.A., Colwell, R.R. (1996). Appl. Environ. Microbiol. 62:1336-1341. Current estimates of viral abundance in natural waters rely on direct counts of virus-like particles (VLPs), using either transmission or epifluorescence microscopy. Direct counts of VLPs, while useful in studies of viral ecology, do not indicate whether the observed VLPs are capable of infection and/or replication. Rapid decay in bacteriophage viability under environmental conditions has been observed. However, it has not been firmly established whether there is a corresponding degradation of the virus particles. To address this question, viable and direct counts were carried out employing two Chesapeake Bay bacteriophages in experimental microcosms incubated for 56 h at two depths in the York River estuary. Viruses incubated in situ in microcosms at the surface yielded decay rates in full sunlight of 0.11 and 0.06 h^-1 for CB 38-PHI and CB 7-PHI, respectively. The number of infective particles in microcosms in the dark and at a depth of 1 m was not significantly different from laboratory controls, with decay rates averaging 0.052 h^-1 for CB 38-PHI and 0.037 h^-1 for CB 7-PHI). Direct counts of bacteriophages decreased in the estuarine microcosms, albeit only at a rate of 0.028 h^-1, and were independent of treatment. Destruction of virus particles is concluded to be a process separate from loss of infectivity. It is also concluded that strong sunlight affects the viability of bacteriophages in surface waters, with the result that direct counts of VLPs overestimate the number of bacteriophage capable of both infection and replication. However, in deeper waters, where solar radiation is not a significant factor, direct counts should more accurately estimate numbers of viable bacteriophage. [TOP OF PAGE]

203. Viruses, plasmids and other genetic elements of thermophilic and hyperthermophilic Archaea. [Review] [28 refs]. Zillig, W., Prangishvili, D., Schleper, C., Elferink, M., Holz, I., Albers, S., Janekovic, D., Gotz, D. (1996). FEMS Microbiol. Rev. 18:225-236. We review and update the work on genetic elements, e.g., viruses and plasmids (exluding IS elements and transposons) in the kingdom Crenarchaeota (Thermoproteales and Sulfolobales) and the orders Thermococcales and Thermoplasmales in the kingdom Euryarchaeota of the archael domain, including unpublished data from our laboratory. The viruses of Crenarchaeota represent four novel virus families. The Fuselloviridae represented by SSVI of S. shibatae and relatives in other Sulfolobus strains have the form of a tailed spindle. The envelope is highly hydrophobic. The DNA is double-stranded and circular. Members of this group have also been found in Methanococcus and Haloarcula. The Lipothrivciridae (e.g., T TV1 to 3) have the form of flexible filaments. They have a core containing linear double-stranded DNA and DNA-binding proteins which is wrapped into a lipid membrane. The "Bacilloviridae" (e.g., TTV4 and SIRV) are stiff rods lacking this membrane, but also featuring linear double-stranded DNA and DNA-binding proteins. Both virus types carry on both ends structures involved in the attachment to receptors. Both types are represented in Thermoproteus and Sulfolobus. The droplet-formed novel Sulfolobus virus SNDV represents the "Guttaviridae" containing circular double-stranded DNA. Though head and tail viruses distantly resembling T phages or lambdoid phages were seen electronmicroscopically in solfataric water samples, no such virus has so far been isolated. SSV1 is temperate, TTV1 causes lysis after induction, the other viruses found so far exist in carrier states. The hosts of all but TTV1 survive virus production. We discuss the implications of the nature of these viruses for understanding virus evolution. The plasmids found so far range in size from 4.5 kb to about 40 kb. Most of them occur in high copy number, probably due to the way of their detection. Most are cryptic, pNOB8 is conjugative, the widespread pDL10 alleviates in an unknown way autotrophic growth of its host Desulfurolobus by sulfur reduction. The plasmid pTIK4 appears to encode a killer function. pNOB8 has been used as a vector for the transfer of the lac S (beta-galactosidase) gene into a mutant of S. solfataricus. [References: 28]. [TOP OF PAGE]

204. Relationships of tailed phages: a survey of protein sequence identity. Ackermann, H.-W., Elzanowski, A., Fobo, G., Stewart, G. (1995). Archives of Virology 140:1871-1884. [TOP OF PAGE]

205. A Bacillus phage that is a living fossil. Ackermann, H.-W., Yoshino, S., Ogata, S. (1995). Canadian Journal of Microbiology 41:294-297. Bacillus cereus phage Bace-11 has an isometric head and a contractile tail of about 500 nm in length, and is one of the two largest bacterial viruses known. Its tail, characterized by three long, whiplike fibers, is identical to giant phage tails produced by a Clostridium strain. These particles may represent phages of ancient origin that arose before the divergence of clostridia and bacilli. [TOP OF PAGE]

206. Bacillus cereus phage typing as an epidemiological tool in outbreaks of food poisoning. Ahmed, R., Sankar-Mistry, P., Jackson, S., Ackermann, H.-W., Kasatiya, S.S. (1995). Journal of Clinical Microbiology 33:636-640. Bacillus cereus is responsible for an increasing number of food poisoning cases. By using 12 bacteriophages isolated from sewage, a typing scheme for B. cereus isolates from outbreaks or sporadic cases of food poisoning was developed. The phages belonged to three morphotypes. Ten phages with contractile tails and icosahedral heads were members of the Myoviridae family, and two phages with noncontractile tails belonged to the Siphoviridae family. Phage II represented a new species. It had an isometric head and a very long contractile tail with long wavy tail fibers and was one of the largest viruses known. The vast majority of 166 B. cereus strains (161, or 97%) isolated from food poisoning cases were typeable. Of 146 strains isolated from 18 outbreaks, 142 (97%) could be divided into 17 phage types. A good correlation, on the order of 80 to 100%, between phage types of strains isolated from suspected foods and those of strains isolated from stools of symptomatic patients was observed. Most Bacillus thuringiensis strains were also typeable, providing further evidence of the close relatedness of B. cereus and B. thuringiensis. This phage typing scheme can be a valuable epidemiological tool in tracing the origins of food poisoning caused by B. cereus. [TOP OF PAGE]

207. Nested PCR with three highly degenerate primers for amplification and identification of DNA from related organisms. Chen, F., Suttle, C.A. (1995). BioTechniques 18:609-611. This paper reports that three highly degenerate primers can be used in nested PCR, and that the second amplification can be done directly on DNA fragments excised from low melting temperature agarose. The approach provides rapid confirmation that the correct targets have been amplified. It is useful when only one internal probe sequence (or gene-specific primer) is available to confirm the identity of a PCR product, and a degenerate primer sequence must be derived from an amino acid sequence. This can occur, for example, when DNA must be amplified from a group of related organisms. [TOP OF PAGE]

208. Amplification of DNA polymerase gene fragments from viruses infecting microalgae. Chen, F., Suttle, C.A. (1995). Appl. Environ. Microbiol. 61:1274-1278. Nested PCR using three highly degenerate primers was used for amplification and identification of the DNA polymerase (pol) genes from viruses which infect three genera of microalgae. Group-specific primers (AVS1, AVS2) were designed based on inferred amino acid sequences that were unique to the DNA pol genes of viruses (PBCV-1 and NY-2A) infecting an endosymbiotic Chlorella-like algae (Chlorophyceae), and a virus (MpV-SP1) which infects the photosynthetic flagellate Micromonas pusilla (Prasinophyceae). As well, a nested primer (POL) was designed based on the highly conserved amino acid sequence YGDTDS found in most B-family (*-like) DNA pol genes. These primers were used to amplify DNA from the three viruses PBCV-1, NY-2A and MpV-SP1 for which the primers were designed, as well as eight other clonal isolates of genetically distinct viruses which infect M. pusilla and viruses which infect Chrysochromulina spp., (Prymnesiophyceae) suggesting that these are a group of related viruses. The primers could also be used to amplify DNA from natural virus communities. In contrast, a product was not produced when DNA from viruses which infect the marine brown algae Ectocarpus siliculosis and Feldmannia sp. (Phaeophyceae) was used, suggesting that these viruses may not be closely related to those infecting microalgae. The primers could also be used to amplify DNA from natural virus communities. Our results indicate that nested PCR, even under low stringency conditions, can be used as a rapid method to verify the presence in seawater of a group of related viruses which infect microalgae. Sequence analysis of these fragments should provide information on the genetic diversity and possibly the phyletic relationships among these viruses. This is the first example of a PCR-based technique designed to detect viruses which infect eukaryotic algae. [TOP OF PAGE]

209. Genetic diversity of algal viruses which lyse the photosynthetic picoflagellate Micromonas pusilla (Prasinophyceae). Cottrell, M.T., Suttle, C.A. (1995). Appl. Environ. Microbiol. 61:3088-3091. The genetic similarity among eight clones of Micromonas pusilla virus (MpV) isolated from five geographic locations was measured by DNA hybridization. Our objective was to explore the existence of genetically distinct populations of MpV by comparing the similarity among MpVs isolated from a single water sample to the similarity among viruses isolated from geographically distant locations. The highest and lowest similarities we observed were 70% plus or minus 1.1% (mean plus or minus standard error [SE], n = 3) for virus strains SP1 and SP2 isolated from a California coastal water sample and 13% plus or minus 1.9% for strains SP2 and PB6; the latter was isolated from New York estuarine water. However, the similarity between MpV isolated from a single water sample was not always greater than the similarity between viruses isolated from different locations. Viruses PB7 and PB8 were isolated from a single New York estuarine sample but were only 16% plus or minus 0.5% similar, whereas PB7 was quite similar (43% plus or minus 2.9%) to PL1, a virus from Texas coastal water. Overall, the similarity among MpVs isolated from a single geographic location, 34% plus or minus 12.6% (mean plus or minus SE, n = 4), was not significantly different from the similarity among MpVs isolated from geographically distant locations, 26.6% plus or minus 2.7% (mean plus or minus SE, n = 24) (P = 0.92, Mann-Whitney U test). Clones of MpV were more similar to each other than they were to the related algal virus PBCV-1, and three groups of MpVs consisting of (i) PL1, SG1, PB6, and PB7, (ii) PB8, and (iii) GM1, SP1, and SP2 were resolved. The genetic variation among MpVs isolated from a single water sample was as large as the variation between viruses isolated from different oceans. If MpVs within a geographic location share genetic characteristics not shared with MpVs from geographically distant locations, this was not reflected in the overall similarity of their genomes. [TOP OF PAGE]

210. Dynamics of a lytic virus infecting the photosynthetic marine picoflagellate Micromonas pusilla. Cottrell, M.T., Suttle, C.A. (1995). Limnol. Oceanogr. 40:730-739. The impact of Micromonas pusilla virus (MpV) on Micromonas pusilla was inferred from measurements of the abundance of MpV, the kinetics of MpV adsorption to host cells, and the estimated in situ decay rate of MpV infectivity. The viral production rate was calculated to balance the estimated in situ decay rate of MpV infectivity. In inshore water of the Texas coast, the abundance of infective MpV was high and decreased from 130,000/ml in January 1993 to 2100/ml, at the end of April 1993. Decay rates of MpV infectivity in seawater incubated in the dark ranged from 0.06/d at 4 degree C to 0.09/d at 25 degree C. In unattenuated sunlight, decay rates of infectivity were much higher, ranging from 6.9 to 7.1/d. Sunlight-mediated decay rate of viral infectivity was depths-dependent, with an attenuation coefficient estimated to equal 0.73/m. The MpV production rate was 0.79/d, equal to a turnover time of 1.3 d. MpV abundance changed slowly relative to its turnover time, suggesting a stable coexistence of M. pusilla and the lytic virus. The adsorption coefficient for MpV-SP1 and host strain Plymouth 27 was 1.40 x 10^-9 ml/min. Using this coefficient, we calculated that from 2 to 10% of the M. pusilla population was lysed per day (avg, 4.4%/d). These results suggest that lysis of phytoplankton by viruses is a process that needs to be incorporated into models of nutrient and energy cycling in aquatic food webs. [TOP OF PAGE]

211. Relationships among Actinomyces naeslundii (A. viscosus) bacteriophages isolated from sewage and the oral cavity. Delisle, A.L., Donkersloot, J.A. (1995). Microbial Ecology in Health & Disease 8:121-127. Several lytic phages of Actinomyces naeslundii genospecies 2 (formerly A. viscosus) have been isolated from sewage and from dental plaque. To define the relationships between these phages and ultimately to assess their role in the ecology of the human oral cavity, 13 phages isolated from these two environments were purified and their biochemical properties compared. Five small, short-tailed phages, isolated from sewage over the course of several years (Av-1, Av-2, Av-3, 1281, and BF307) were morphologically indistinguishable from each other and from five phages recovered more recently from human dental plaque (CT1, CT2, CT3, CT6 and CT7). The small phages (all morphotype C1) contained double-stranded linear DNA, 18 kb in size. In contrast, three phages from dental plaque (CT4, CT5 and CT8) possessed longer tails and much larger head structures (morphotype B1). Two of the larger phages (CT4 and CT5) contained DNA genomes estimated to be 80 kb in size, whereas large phage CT8 contained DNA of approximately 50 kb. Restriction endonuclease analysis revealed extensive differences between the large and small phages but the latter group showed similar, and in several cases identical, fragment patterns. These results indicate the existence of at least three distinct types of lytic bacteriophage active against oral Actinomyces spp. The similarities between the sewage and small dental plaque isolates indicate a high degree of relatedness and suggest that the sewage phages probably originated from the oral cavity. [TOP OF PAGE]

212. Phylogeny of Aureococcus anophagefferens and a morphologically similar bloom-forming alga from Texas as determined by 18S rDNA sequence analsysis. DeYoe, H.R., Chan, A.M., Suttle, C.A. (1995). Journal of Phycology 31:413-418. [TOP OF PAGE]

213. Bacteriophages of Erwinia carotovora and Erwinia ananas isolated from freshwater lakes. Eayre, C.G., Bartz, J.A., Concelmo, D.E. (1995). Plant Disease 79:801-804. Bacteriophages for Erwinia carotovora subsp. carotovora and for E. ananas were readily isolated from freshwater lakes in Florida and Texas. Approximately 15% of enrichment cultures with 48 strains of E. carotovora yielded phage. Nineteen of 22 phages had distinct host ranges among the 62 strains and up to 10 strains were susceptible to a single phage. Among strains representing 24 serotypes, 12 of 15 phages caused plaques in lawns of 16 serotypes. Each of 13 enrichment cultures with strains of E. ananas yielded at least one phage and five distinct host range patterns emerged among host strains and phage isolates. The number of susceptible hosts for each phage ranged from one to six. [TOP OF PAGE]

214. Partial characterization of Streptomyces phages isolated from the soils of jarrah forest in Western Australia. El-Tarabily, K.A., Kurtboke, D.I., Hardy, G.E.S. (1995). Actinomycetes 6:7-15. [TOP OF PAGE]

215. Can phage defence maintain colicin plasmids in Escherichia coli? Feldgarden, M., Golden, S., Wilson, H., Riley, M.A. (1995). Microbiology 141:2977-2984. We examined the role of plasmid-based phage defence in maintaining plasmids, using colicin plasmids in Escherichia coli as a model system. Experimental data indicated that the possession of a colicin plasmid can confer limited protection against bacteriophages. A continuous culture model, using these experimental values, indicated that the observed limited protection alone could selectively maintain colicin plasmids, without requiring a competitive advantage due to colicinogeny. Phage defence might explain the current maintenance of colicin plasmids, given the naturally occurring high levels of resistance to colicins. This model also suggests that many plasmids might be maintained in natural populations, in part, by phage resistance, including 'cryptic' plasmids for which no phenotype is known. [TOP OF PAGE]

216. Bacteriophage resistance in Lactococcus: characterization of the ScrFI restriction/modification system from Lactococcus lactis subsp. cremoris UC503. Fitzgerald, G.F., Twomey, D.P., Daly, C., Coffey, A. (1995). pp. 581-590. In In Ferretti, J., Gilmore, M., Klaenhammer, T., and Brown, F. (eds.), Genetics of streptococci, enterococci and lactococci. Karger, Basel. [TOP OF PAGE]

217. Viruses and protists cause similar bacterial mortality in coastal seawater. Fuhrman, J.A., Noble, R.T. (1995). Limnol. Oceanogr. 40:1236-1242. Mesocosms filled with 80 liters of coastal seawater from Santa Monica, California, were used twice (June and November) to budget bacterial production and loss, as well as to assess the relative significance of viral lysis and protist grazing in bacterial mortality. Bacterial abundance was apprx 6 times 10-9 cells liter-1 in June and 2 times 10-9 in November, with viral abundances apprx 2 times 10-10 particles liter-1 in June and 1.5 times 10-10 in November. Incorporation of (3H)thymidine and leucine yielded essentially identical production estimates and allowed calculation of total bacterial mortality in these closed systems. Bacterial growth rates were 1-2 d-1 in June and 1-3 d-1 in November. Three independent lines of evidence indicated that bacterial mortality attributed to grazing by protists was about equal to that attributed to viruses: size fractionation of disappearance of labeled DNA, with a 50% reduction after protists were removed; comparison of protist grazing rates estimated with fluorescently labeled bacteria and virus production-based bacterial lysis rates, with 40-50% of the total ascribed to viruses; and model-based interpretation of the 3.3-4.6% of bacteria visibly infected with assembled intracellular viruses, suggesting that 24-66% of loss is due to infection. Redundant production and loss measurements as well as the independent loss process estimates agreed within apprx 30%, yielding a reasonably balanced budget. We believe the loss of bacteria to viruses reflects a significant dissipation of energy in this ecosystem and that viruses and protists contribute similarly to bacterial mortality. [TOP OF PAGE]

218. Large double-stranded DNA viruses which cause the lysis of marine heterotrophic nanoflagellates (Bodo sp.) occur in natural marine virus communities. Garza, D.R., Suttle, C.A. (1995). Aquat. Microb. Ecol. 9:203-210. A virus (BV-PW1) which causes lysis of two strains of a marine heterotrophic nanoflagellate belonging to the genus Bodo (strains E1 and E4) was isolated from the coastal waters of Texas. Transmission electron microscopy of ultrathin sections revealed the presence of intracellular virus-like particles 48 h following infection, concomitant with a decline in flagellate numbers. The virus contains double-stranded DNA, is hexagonal in cross-section, ca. 230-300 nm in diameter and contains an electron dense core. It is morphologically similar to virus-like particles which have been observed in other heterotrophic nanoflagellates, and to viruses which have been isolated which infect eukaryotic phytoplankton. Addition of the viruses to cultures of Pseudobodo parvulus (ATCC 50091, formerly Bodo parvulus) or Paraphysomonas imperforata (strain VS1) did not result in lysis. To our knowledge this is the first virus infecting heterotrophic nanoflagellates which has been isolated and maintained in culture. The presence of viruses in seawater which cause lysis of phagotrophic nanoflagellates implies that viruses infect microzooplankton populations in the sea and suggests another important role for viruses in aquatic microbial communities. [TOP OF PAGE]

219. Extremo-phage: In vitro selection of tolerance to a hostile environment. Gupta, K., Lee, Y., Yin, J. (1995). J. Mol. Evol. 41:113-114. Extremophiles are microorganisms that thrive under extreme conditions such as temperatures above 65 degree C, pHs below 4 or above 10, salt concentrations above 0.5 M, or pressures of 600 atm. While studies of enzymes either isolated from extremophiles, or generated using site-specific mutagenesis, or adapted by in vivo or in vitro selection have established a precedent for the engineering and application of proteins at extreme conditions, generalization of the approaches to more complex multimolecular or multitask systems has remained elusive. Here we demonstrate that a significantly more complex system-a bacteriophage-can over a number of generations be adapted to tolerate a hostile and unnatural environment. An in vitro selection strategy was used to adapt phage to urea, a protein denaturing agent. As the concentration of urea employed in selections over 20 generations was gradually increased from 5 to 9 m, the surviving phages steadily improved their tolerance, finally achieving a greater than 350-fold stability enhancement over the original population. [TOP OF PAGE]

220. Fluorescently labeled virus probes show that natural virus populations can control the structure of marine microbial communities. Hennes, K.P., Suttle, C.A., Chan, A.M. (1995). Appl. Environ. Microbiol. 61:3623-3627. Fluorescently stained viruses were used as probes to label, identify and enumerate specific strains of bacteria and cyanobacteria in mixed microbial assemblages. Several marine virus isolates were fluorescently stained with YOYO-1 or POPO-1 (Molecular Probes, Inc.) and added to seawater samples that contained natural microbial communities. Cells to which the stained viruses adsorbed were easily distinguished from non-host cells; typically, there was undetectable binding of stained viruses to natural microbial assemblages containing &gt;10⁶ bacteria ml^-1, but to which host cells were not added. Host cells that were added to natural seawater were quantified with 99 &plusmn; 2 % efficiency using fluorescently labeled virus probes (FLVPs). A marine bacterial isolate (strain PWH3a) was introduced into natural microbial communities that were either supplemented with nutrients or untreated, and changes in the abundance of the isolate were followed using FLVPs. Simultaneously, the concentration of viruses that infected strain PWH3a was monitored by plaque assay. Following the addition of PWH3a, viruses infecting this strain increased from undetectable levels (&lt;1 ml^-1 to 2.9 x 10⁷ ml^-1 and 8.3 x 10⁸ ml^-1 for the untreated and nutrient-enriched samples, respectively. The increase in viruses was associated with a collapse in populations of strain PWH3a from ca. 30% to 2% and 43% to 0.01% of the microbial communities in untreated and nutrient-enriched samples, respectively. These results clearly demonstrate that FLVPs can be used to identify and quantify specific groups of bacteria in mixed microbial communities. As well, the data show that viruses which are present at low abundances in natural aquatic viral communities can control microbial community structure. [TOP OF PAGE]

221. Direct counts of viruses in natural waters and laboratory cultures by epifluorescence microscopy. Hennes, K.P., Suttle, C.A. (1995). Limnol. Oceanogr. 40:1050-1055. Epifluorescent microscopy was used to determine the abundance of viruses in samples from marine and freshwater environments and in laboratory cultures that were filtered onto 0.02-&micro;m pore-size filters and stained with a cyanine-based dye (Yo-Pro-1). Estimates of viral abundance based on Yo-Pro stained samples were 1.2 to 7.1 times greater than estimates obtained using transmission electron microscopy (TEM). Moreover, the precision of the Yo-Pro based method was much greater than that for TEM (coefficient of variation 7 % versus 20 %, respectively). DNase treatment of samples did not result in lower numbers of particles that could be stained by Yo-Pro, suggesting that the fluorescence was not the result of nucleic acids associated with the surface of particles. These results indicate that the concentration of viruses in natural waters may be higher than previously recognized and imply that the TEM-based method significantly underestimates virus abundance. Virus abundances ranged from 10⁷ to &gt; 10⁸ ml^-1 in surface waters along a transect in the western Gulf of Mexico to 10⁹ ml^-1 in water overlying a submerged cyanobacterial mat. High counting efficiency, ease of preparation, modest equipment requirements and the possibility of preparing specimens for long-term storage, make the Yo-Pro based method ideal for routine environmental analysis. [TOP OF PAGE]

222. Viral contribution to dissolved DNA in the marine environment as determined by differential centrifugation and kingdom probing. Jiang, S.C., Paul, J.H. (1995). Appl. Environ. Microbiol. 61:317-325. Dissolved or filterable (<0.2- mu m-pore-size filter) DNA is a ubiquitous component of the dissolved organic matter in the surface waters of this planet. In an effort to understand the composition and possible sources, we subjected dissolved DNA concentrated by vortex flow filtration from offshore and coastal environments to differential centrifugation and probing with 16S rRNA-targeted kingdom oligonucleotide probes. Initial studies with calf thymus soluble DNA and T2 phage particles indicated that high-speed ultracentrifugation (201,000 x g for 90 min), a method to separate viral particles from soluble DNA used by other investigators, resulted in pelleting of nearly all the DNA and virus particles. Lower-speed centrifugation (11,200 to 25,800 x g for 90 min) resulted in >99% of the virus particles being collected in the pellet and similar to 65% of the calf thymus DNA remaining in the supernatant. Employing this approach, we estimate that approximately 50% of the filterable DNA from marine environments is truly soluble or free DNA and that the other half is composed of bound forms (viral particles and, potentially, colloids). Of the bound form, 17 to 30% could be accounted for by viral particles, by calculating the amount of viral DNA on the basis of viral abundance, leaving a portion of the bound form uncharacterized. Kingdom probing with universal, eubacterial, and eucaryotic probes indicated that dissolved DNA hybridized with all of these probes, while purified standard viral DNAs did not, or hybridized only slightly with the universal probe (tailed oligonucleotide only). Collectively, these data indicate that DNA in viral particles is a small component of the dissolved DNA, the majority being of eubacterial and eucaryotic origin. [TOP OF PAGE]

223. Genetic diversity of related vibriophages isolated from marine environments around Florida and Hawaii, USA. Kellogg, C.A., Rose, J.B., Jiang, S.C., Thurmond, J.M., Paul, J.H. (1995). Mar. Ecol. Prog. Ser. 120:89-98. Although viruses from the marine environment have been enumerated, isolated, and characterized, there is little information on the abundance or global distribution of specific phage types. To this end, we studied the abundance and distribution of phages which infect a marine bacterium isolated from Tampa Bay (Florida, USA), tentatively identified (Microbial ID, Inc., Newark, Delaware, USA) as Vibrio parahaemolyticus. Using this host, we have isolated over 60 phages from the Gulf of Mexico, Tampa Bay, Florida Keys, and Oahu, Hawaii (USA). These isolates are all Myoviridae, with head sizes ranging from 50 +- 0.0 to 65 +- 4.2 nm and tail lengths of 60 +- 3.6 to 100 +- 5.0 rim. The type phage (PHI-16 from Tampa Bay) has a double-stranded DNA genome of 51 to 58 kb. A 1.5 kb EcoRI fragment of this genome has been cloned and used as a gene probe. All of the DNA from the phage isolates hybridized to this probe under stringent conditions, but not to DNA from other marine vibriophages and bacteriophages, suggesting genetic relatedness. Agarose gel electrophoresis of EcoRI digests of the DNAs, followed by Southern transfer and probing with the 1.5 kb gene probe, yielded 6 groups based upon banding patterns. These groups were not segregated geographically within the Florida isolates; however, all of the Hawaiian phages had a common restriction pattern. These data indicate that populations of genetically related phages are widely distributed over large geographic distances in the oceans. [TOP OF PAGE]

224. Whole-virus vaccine development by continuous culture on a complementing host. Kong, D., Yin, J. (1995). Bio/Technology 13:583-586. We have evaluated an adaptive strategy for generating whole-virus vaccines using a bacteriophage model. Wildtype phage T7 was cultivated in a two-stage continuous stirred-tank reactor (CSTR) utilizing a recombinant E. coli host that constitutively expressed T7 RNA polymerase, an essential enzyme of the early viral metabolism. Over the course of 180 generations a diversity of phage variants emerged, outgrew the wildtype, and were subsequently eclipsed by yet fitter variants, based on host-ranges, restriction patterns, and one-step growth responses of isolated clones. The fittest variant, which required complementation by the recombinant host in order to grow, deleted at least 12 percent of its genome and replicated twice as fast as the wildtype. Moreover, this variant was immunogenically indistinguishable from the wildtype, based on cross-reactivities of antisera raised against both. These results suggest the feasibility of the proposed strategy for the development of safe whole-virus vaccines. [TOP OF PAGE]

225. Cryptic transposable phages of Pseudomonas aeruginosa. Krylov, V.N., Mit'kina, L.N., Pleteneva, E.A., Aleshin, V.V. (1995). Genetika 31:1507-1511. Frequencies of nucleotide sequences homologous to phage transposons (PT) of two species, D3112 and B3, were assessed in genomes of natural Pseudomonas aeruginosa strains by the dot-blot hybridization method. These strains were incapable of liberating viable phages on a lawn of the PA01 standard indicator strain of P. aeruginosa. It was shown that the homologies detected belong to two groups, high and intermediate, with respect to homology level. Homology patterns were classified as high when they provided signals comparable to those for hybridization in a positive control; they were classified as intermediate when the hybridization level higher than the background level, but lower than in the positive control. Homologous PIT sequences were designated as cryptic PT. Intact cryptic PT prophages were shown to exist in genomes of particular natural strains manifesting a high level of hybridization. However, the growth of these phages was limited by the restriction system of strain PA01. It is possible to isolate strains maintaining the growth of a portion of cryptic PT. These strains differed from P. aeruginosa with respect to the specificity of the restriction and modification system. Nevertheless, in most cases, the attempt to identify a novel host capable of maintaining growth of a cryptic PT failed. Natural strains often carry cryptic PT related to both known PT species, D3112 and B3. The frequency of cryptic PT is extremely high, reaching 30% in only strains with a high level of homology and up to 50% in all strains exhibiting homology. This high PT frequency is assumed to be associated with the considerable variation of P. aeruginosa. [TOP OF PAGE]

226. Mucoid clones of Pseudomonas aeruginosa PAO1 surviving after induction of transposable prophages. Krylov, V.N., Solov'eva, T.I., Burkal'tseva, M.V. (1995). Genetika 31:1375-1379. The origin and properties of mucoid clones were studied. The clones were selected with high frequency after thermoinduction of Pseudomonas aeruginosa lysogenic for phage-transposons (PT). The production of alginate does not promote the survival of lysogenic bacteria at 42degreeC. Mucoid clones were shown to appear before thermoinduction; the frequency of their formation does not depend on the specificity of the mutator effect intrinsic to different PT. Phenotypic differences typical of mucoid clones can be mediated by different mutations promoting clone survival at 42degreeC and by simultaneously arising additional mutations. The SL21 mucoid clone selected among clones of P. aeruginosa PAO1 resistant to PT of B3 possesses an additional trait of phage resistance at 42degreeC. The presence of D3112 cts 15 prophage has no significant effect on the frequency of SL21 reversion to nonmucoidness. This means that the mutator effect of PT has made a slight contribution to this process. The appearance of mutations promoting the survival of the thermoinducible lysogen SL21 (D3112 cts 15) does not affect the frequency of the loss of mucoidness. Nonmucoid derivatives of SL21 were shown to differ in phage resistance at 42degreeC and in the extent of the residual mucoidness manifested under specific conditions. Consequently, nonmucoid clones appear as a result of pseudoreversions. Because some of these pseudorevertants cannot again be convened to the mucoid form, it is concluded that they carry mutations in genes whose functions are obligatory for the production of alginate. [TOP OF PAGE]

227. Inducible bacteriophages of Actinobacillus actinomycetemcomitans. Loftus, A., Delisle, A.L. (1995). Current Microbiology 30:317-321. Doses of 0.1 to 1.0 mu-g/ml of mitomycin C induced cell lysis of six of eight strains of Actinobacillus actinomycetemcomitans tested. Infectious phages were induced from ATCC strains 43717, 29524, 33384, and 43719; non-plaque- forming, possibly defective phages were induced from ATCC strains 29522 and 29523. No phages were detected in strain FDC 651 or ATCC strain 43718. No correlation between lysogeny and leukotoxin production or serotype of the strains could be established. Gel electrophoresis of phage DNAs indicated that the induced phages were of three types, based on size. By electron microscopy, the phages were found to belong to either morphotype A1 or morphotype B1; no other morphotypes were observed. Curing experiments led to the isolation of nonlysogenic derivatives of two strains, which supported plaque formation by the phages they originally carried. On the basis of our results, lysogeny appears to be widespread in A. actinomycetemcomitans. [TOP OF PAGE]

228. Cloning and sequencing of LlaDCHI [corrected] restriction/modification genes from Lactococcus lactis and relatedness of this system to the Streptococcus pneumoniae DpnII system [published erratum appears in Appl Environ Microbiol 1995 Sep;61(9):3514]. Moineau, S., Walker, S.A., Vedamuthu, E.R., Vandenbergh, P.A. (1995). Appl. Environ. Microbiol. 61:2193-2202. The natural 7.8-kb plasmid pSRQ700 was isolated from Lactococcus lactis subsp. cremoris DCH-4. It encodes a restriction/modification system named LlaDCHI [corrected]. When introduced into a phage-sensitive L. lactis strain, pSRQ700 confers strong phage resistance against the three most common lactococcal phage species, namely, 936, c2, and P335. The LlaDCHI [corrected] endonuclease was purified and found to cleave the palindromic sequence 5'-GATC-3'. It is an isoschizomer of Streptococcus pneumoniae DpnII. The plasmid pSRQ700 was mapped, and the genetic organization of LlaDCHI [corrected] was localized. Cloning and sequencing of the entire LlaDCHI [corrected] system allowed the identification of three open reading frames. The three genes (llaIIA, llaIIB, and llaIIC) overlapped and are under one putative promoter. A putative terminator was found at the end of llaIIC. The genes llaIIA and llaIIB coded for m6A methyltransferases, and llaIIC coded for an endonuclease. The LlaDCHI [corrected] system shares strong genetic similarities with the DpnII system. The deduced amino acid sequence of M.LlaIIA was 75% identical with that of M.DpnII, whereas M.LlaIIB was 88% identical with M.DpnA. However, R.LlalII shared only 31% identity with R.DpnII. [TOP OF PAGE]

229. Halophage HF2: genome organisation and replication strategy. Nuttall, S.D., Dyall-Smith, M.L. (1995). J. Virol. 69:2322-2327. Halophage HF2 is a lytic, broad-host-range bacteriophage of the extremely halophilic domain Archaea. It has a 79.7-kb double-stranded DNA genome which is linear, contains no modified nucleotides, and is not susceptible to cleavage by many type II restriction endonucleases. This insensitivity is attributed to selection against palindromic restriction sites, a commonly observed feature of broad-host-range phages. Interestingly, enzymes that did cut the genome recognized AT-rich sites, and five such enzymes, DraI, AseI, HpaI, HindIII,
     and SspI, were used to construct a physical map of the genome. Southern hybridization experiments used to order fragments on the map indicated homologies between the phage termini, and subsequent sequence analysis showed that HF2 possessed 306-bp direct terminal repeats. The presence of such repeats suggested replication through concatameric intermediates, and this was confirmed by analysis of the state of the phage genome in infected cells. This is a replication strategy adopted by many well-studied bacterial phages, for example T3 and T7. Other similarities between the terminal repeats of T3 or T7 and HF2 include a putative nick site at the repeat border and a series of short imperfect repeats. These observations suggest a long evolutionary history for concatamer-based strategies of phage replication, possibly predating the divergence of Archaea/Eucarya and Bacteria, or alternatively, indicate possible lateral transfer of phage genes or modules between the domains Archaea and Bacteria. [TOP OF PAGE]

230. Isolation of Dermatophilus congolensis phage from the 'lumpy wool' of sheep in Western Australia. Patten, K.M., Kurtboke, D.I., Lindsay, D.R. (1995). Letters in Applied Microbiology 20:199-203. A lytic phage with species-specific activity was isolated from wool samples infected with the actinomycete Dermatophilus congolensis, the agent of 'lumpy wool', collected from properties in Western Australia. The physiochemical properties, plaque morphology, host range and particle morphology of the phage isolated were characterized. The isolated phage reduced the cell numbers of Dermatophilus congolensis on infected wool samples in vitro. It may therefore have potential as a biocontrol agent of dermatophilosis. [TOP OF PAGE]

231. Viral tracer studies indicate contamination of marine waters by sewage disposal practices in Key Largo, Florida. Paul, J.H., Rose, J.B., Brown, J., Shinn, E.A., Miller, S., Farrah, S.R. (1995). Appl. Environ. Microbiol. 61:2230-2234. Domestic wastewater disposal practices in the Florida Keys are primarily limited to on-site disposal systems such as septic tanks, injection wells, and illegal cesspits. Poorly treated sewage is thus released into the highly porous subsurface Key Largo limestone matrix. To investigate the fate and transport of sewage in the subsurface environment and the potential for contamination of marine surface waters, we employed bacteriophages as tracers in a domestic septic system and a simulated injection well in Key Largo, Florida. Transport of bacteriophage PHI-HSIC-1 from the septic tank to adjacent surface canal waters and outstanding marine waters occurred in as little as 11 and 23 h, respectively. Transport of the Salmonella phage PRD1 from the simulated injection well to a canal adjacent to the injection site occurred in 11.2 h. Estimated rates of migration of viral tracers ranged from 0.57 to 24.2 m/h, over 500- fold greater than flow rates measured previously by subsurface flow meters in similar environments. These results suggest that current on-site disposal practices can lead to contamination of the subsurface and surface marine waters in the Keys. [TOP OF PAGE]

232. Occurrence of fecal indicator bacteria in surface waters and the subsurface aquifer in Key Largo, Florida. Paul, J.H., Rose, J.B., Kellogg, C., Shinn, E.A. (1995). Appl. Environ. Microbiol. 61:2235-2241. Sewage waste disposal facilities in the Florida Keys include septic tanks and individual package plants in place of municipal collection facilities in most locations. In Key Largo, both facilities discharge into the extremely porous Key Largo limestone. To determine whether there was potential contamination of the subsurface aquifer and nearby coastal surface waters by such waste disposal practices, we examined the presence of microbial indicators commonly found in sewage (fecal coliforms, Clostridium perfringens, and enterococci) and aquatic microbial parameters (viral direct counts, bacterial direct counts, chlorophyll a, and marine vibriophage) in injection well effluent, monitoring wells that followed a transect from onshore to offshore, and surface waters above these wells in two separate locations in Key Largo in August 1993 and March 1994. Effluent and waters from onshore shallow monitoring wells (1.8- to 3.7-m depth) contained two or all three of the fecal indicators in all three samples taken, whereas deeper wells (10.7- to 12.2-m depth) at these same sites contained few or none. The presence of fecal indicators was found in two of five nearshore wells (i.e., those that were < or = 1.8 miles [< or = 2.9 km] from shore), whereas offshore wells (> or = 2.1 to 5.7 miles [< or = 3.4 to 9.2 km] from shore) showed little sign of contamination. Indicators were also found in surface waters in a canal in Key Largo and in offshore surface waters in March but not in August. Collectively, these results suggest that fecal contamination of the shallow onshore aquifer, parts of the nearshore aquifer, and certain surface waters has occurred.(ABSTRACT TRUNCATED AT 250 WORDS). [TOP OF PAGE]

233. Increasing the phage resistance of starter cultures. Ross, R.P., Fitzgerald, G.F., Coffey, A. (1995). pp. 110-118. In In Cogan, T., Ross, R.P., and Fox, P. (eds.), National Dairy Products Research Centre 4th Cheese Symposium. TEAGASC, Fermoy, Co. Cork. [TOP OF PAGE]

234. Pseudomonas aeruginosa bacteriophage in treatment of P. aeruginosa infetion in cystic fibrosis patients. Shabalova, I.A., Karpanov, N.I., Krylov, V.N., Sharibjanova, T.O., Akhverdijan, V.Z. (1995). 443. Zurich, Switzerland, International Cystic Fibrosis Association. Proceedings of IX International Cystic Fibrosis Congress. [TOP OF PAGE]

235. Presence of large virus-like particles in a eutrophic reservoir. Sommaruga, R., Krössbacher, M., Salvenmoser, W., Catalan, J., Psenner, R. (1995). Aquat. Microb. Ecol. 9:305-330. The presence of large virus-like particles without tail was observed in the water of a severely eutrophied freshwater reservoir. We used transmission electron microscopy (TEM) coupled to a semiautomatic image analysing system to study the size distribution of aquatic viruses. The LVLP had maximum head diameters between 195 and 210 nm. Although the affiliation and host of the LVLP are unknown similarities in size and shape were found with the African swine fever virus. The diffuse organic contamination from piggeries in the catchement area may explain the presence of these LVLP. The overlap in size of the LVLP with the smallest bacteria observed imply serious methodological problems to distinguish these communities under the epifluorescence microscope. Therefore, although at present we do not know how common are LVLP in other aquatic systems we discourage the use of epifluorescence microscopy for viral abundance estimations during routine work. [TOP OF PAGE]

236. Viruses infecting the marine prymnesiophyte Chrysochromulina spp.: Isolation, preliminary characterization and natural abundance. Suttle, C.A., Chan, A.M. (1995). Mar. Ecol. Prog. Ser. 118:275-282. Sixty-four natural virus communities were concentrated from seawater collected from three locations in Texas' coastal waters (Gulf of Mexico, 27°; 31' N, 96°; 18' W; Aransas Pass 27°; 50' N, 97°; 02' W; Laguna Madre, 27°; 30' N, 97°; 18' W) and screened for the presence of lytic pathogens which infect the marine Prymnesiophyte (Haptophyte) Chrysochromulina brevifilum. Viruses were detected in 16 of the samples and ranged in abundance from 2 to 688 infectious units per liter. The pathogens were detected at the 3 locations, but not on all dates, from December through June when water temperatures were less than 28°;C. A clonal isolate of the virus (CbV-PW1) was obtained by determining the concentration of the infectious agent by a most-probable-number assay and adding 0.2 of an infective unit into each of 20 exponentially growing cultures, removing an aliquot from a culture which lysed and repeating the procedure. The isolate also caused lysis of Chrysochromulina strobilus, but did not lyse 8 other isolates of Chrysochromulina or 5 other genera of Prymnesiophytes that were screened. The double-stranded DNA virus is a polyhedron of about 145-170 nm in diameter with a heavily staining central region that is distinct from the capsid. The appearance of the virions is associated with a granular region in the cytoplasm that does not appear within uninfected cells. Ultimately, viral production results in disruption of the organelles, lysis of the cell and release of the virus particles. Although the number of viruses produced per lytic event is presently unknown we have counted more than 320 virus particles in a single ultrathin section of an infected cell. These results suggest that viruses are likely important in regulating Chrysochromulina populations in the sea and may be the reason that blooms of the genus are relatively rare and ephemeral. [TOP OF PAGE]

237. Biological properties of bacteriophages revealed throughout the manufacturing process of an industrial strain E. coli M 17. Tediashvili, M., Chanishvili, N., Eliashvili, T., Zviadadze, N., Porchkhidze, K., Natroshvili, G., Cholokashvili, N., Giorkhelidze, D., Adamia, R., Chanishvili, T. (1995). Proceedings of the Georgian Academy of Sciences 21:175-184. [TOP OF PAGE]

238. Effect of virus-rich high molecular weight concentrates of seawater on the dynamics of dissolved amino acids and carbohydrates. Weinbauer, M.G., Peduzzi, P. (1995). Mar. Ecol. Prog. Ser. 127:245-253. [TOP OF PAGE]

239. Temporal and spatial distribution of viruses and dissolved DNA in the northern Adriatic Sea. Weinbauer, M.G., Fusk, D., Puskaric, S., Peduzzi, P. (1995). Microb. Ecol. 30:25-41. [TOP OF PAGE]

240. Diel, seasonal, and depth-related variability of viruses and dissolved DNA in the Northern Adriatic Sea. Weinbauer, M.G., Fuks, D., Puskaric, S., Peduzzi, P. (1995). Microb. Ecol. 30:25-41. [TOP OF PAGE]

241. Significance of viruses versus heterotrophic nanoflagellates for controlling bacterial abundance in the northern Adriatic Sea. Weinbauer, M.G., Peduzzi, P. (1995). J. Plank. Res. 17:1851-1856. [TOP OF PAGE]

242. A simple method for the concentration of viruses from natural water samples. Wommack, K.E., Hill, R.T., Colwell, R.R. (1995). J. Microbiol. Meth. 22:57-67. A double filtration method for the concentration of viruses from water was developed. The procedure involves prefiltering a water sample through 0.22 mm and 0.1 &#181m filters, concentrating viruses in the 0.1 &#181m filtrate using a 10,000 molecular weight cut-off (mwco) filter. In tests utilizing bacteriophage host systems (PHS) isolated from the Chesapeake Bay, this method was shown to have an average recovery efficiency of 76.5% for viable bacteriophage. It was also possible to recover 100% of the virus-like particles observed in natural water samples. The recovery of bacteriophages and virus-like particles using our method compared well with other reported virus concentration methods employing adsorption/elution and ultrafiltration. Furthermore, the procedure is inexpensive and well suited to field applications. [TOP OF PAGE]

243. Lysis and the interaction between free phages and infected cells. Abedon, S.T. (1994). pp. 397-405. In In Karam, J.D. (ed.), The Molecular Biology of Bacteriophage T4. ASM Press, Washington, DC. The envelope of T4-infected cells undergoes a complex set of changes that affect its stability, its susceptibility to lysozyme-induced degradation, and the fate of other incoming phage. Particularly, superinfection exclusion, resistance to lysis from without, lysis inhibition, and lysis inhibition collapse are all mechanisms involving the interaction between T4-infected cells and subsequently adsorbing T4 phage. This chapter reviews the normal T4 lysis process and, for the first time, what is known of the complex molecular biology of free phage-infected cell interactions. [TOP OF PAGE]

244. PID, a new member of the P1 bacteriophage group. Ackermann, H.-W., Reynaud, A., Gayerie, M.C. (1994). Can. J. Microbiol. 40:63-67. Phage PID produces particles of essentially uniform head size and differs from P1 in its range and tail length. The dimensions of phage P1 are reassessed. The P1 phage group shows signs of morphological evolution. [TOP OF PAGE]

245. Classification of Acinetobacter phages. Ackermann, H.-W., Brochu, G., Emadi Konjin, H.P. (1994). Arch. Virol. 135:345-354. Eight phage species and type viruses are proposed. They below to the Myoviridae, Siphoviridae, and Podoviridae families of tailed phages and are characterized by a combination of morphological and physiochemical properties. An unusual siphovirus species has an elongated head and transverse tail disks. [TOP OF PAGE]

246. New Bacillus bacteriophage species. Ackermann, H.-W., Azizbekyan, R.R., Emadi Konjin, H.P., Lecadet, M.-M., Seldin, L., Yu, M.X. (1994). Arch. Virol. 135:333-344. Nine new species of tailed Bacillus phages, based on morphological and physiochemical properties, are defined. Phage P10 is one of the largest viruses known. The total number of tailed Bacillus phage species is presently 33. [TOP OF PAGE]

247. Bacteriophages from Bombyx mori. Ackermann, H.-W., Auclair, P., Basavarajappa, S., Emadi Konjin, H.P., Savanurmath, C. (1994). Arch. Virol. 137:185-190. Preparations of silkworm larvae contained two large phages with contractile tails (Myoviridae). One phage was active on Pseudomonas paucimobilis. The other, not cultured, was one of the largest viruses known. [TOP OF PAGE]

248. Stand der Forschung bei der Untersuchung der Phagen Problematik in biotechnologischen Prozessen der Lebensmittelfermentation. Arendt, E.K., Fitzgerald, G.F., Coffey, A., Hammes, W.P. (1994). Lebensmitteltechnik 6:40-44. [TOP OF PAGE]

249. General procedures. Carlson, K., Miller, E.S. (1994). pp. 427-437. In In Karam, J.D. (ed.), Molecular Biology of Bacteriophage T4. ASM Press, Washington, DC. [TOP OF PAGE]

250. Enumerating phage. Carlson, K., Miller, E.S. (1994). pp. 427-429. In In Karam, J.D. (ed.), Molecular Biology of Bacteriophage T4. ASM Press, Washington, DC. The plaque assay is based on the fact that when a single phage particle encounters a permissive bacterium it will infect it and a while later lyse it, with the concomitant release of newly formed phage particles. When about 100 to 500 phage are mixed with about 10⁸ sensitive cells and poured in a layer of soft agar on the surface of a nutrient agar plate supporting bacterial growth, the cells will resume growth. Each phage soon comes into contact with a bacterium, which it infects; in areas where no phage are present the bacteria will grow to stationary phase, forming a smoot opaque layer or lawn in the overlay. The progeny phge from each infected bacterium will infect neighboring bacteria, resulting in a growing zone of lysis, full of liberated phage, which eventually becomes visible to the naked eye as a "plaque" in the otherwise smooth lawn. Growth of plaque is limited by slow diffusion of phage in the semi-solid soft agar and by the fact that the host cells support phage growth only as long as they metabolize actively; when cell growth stops, phage growth also ceases. A production of about 10 to 15 phage per infected cell is sufficient for plaque formation; each phage giving rise to a plaque can be assayed as a plaque-forming unit. [TOP OF PAGE]

251. Working with T4. Carlson, K., Miller, E.S. (1994). pp. 421-426. In In Karam, J.D. (ed.), Molecular Biology of Bacteriophage T4. ASM Press, Washington, DC. [TOP OF PAGE]

252. ??? Cirillo, M.R., Trigg, M.E., Merril, C.R. (1994). Infect. Immun. 62:3254-3261. [this is cited in Lederberg, 1996 (PNAS 93:3167-3168): ". . . perhaps in the soil amoebae that harbor Legionella and other bacteria (ref)"]. [TOP OF PAGE]

253. In situ replication studies of somatic and male-specific coliphages in a tropical pristine river. Hernandez-Delgado, E.A., Toranzos, G.A. (1994). Water Science and Technology 247-250. The present study was carried out in order to evaluate coliphage survival and their ability to replicate in tropical aquatic environments. Results showed that coliphages survived for extended periods of time, however, neither sewage isolates nor laboratory phage strains replicated in the environment. Neither one of the bacterial strains tested were permissive to phage replication of "fecal coliphages" does not occur in the tropical aquatic environment and suggest that somatic or male-specific coliphages may be considered as potential indicators of fecal contamination in tropical aquatic environments. [TOP OF PAGE]

254. Seasonal and diel abundance of viruses and occurrence of lysogeny/bacteriocinogeny in the marine environment. Jiang, S.C., Paul, J.H. (1994). Mar. Ecol. Prog. Ser. 104:163-172. To understand the role of viruses in the marine environment, it is important to know the factors affecting their temporal distribution and the abundance of lysogens. We therefore performed a seasonal and a diel study on viral distribution in Tampa Bay, Florida, USA, and detected the abundance of lysogens and bacteriocinogens amongst marine bacterial isolates from diverse marine environments. We investigated the distribution of viruses, bacterial direct counts, chlorophyll a (chl a), salinity and temperature during a 13 mo period in the Tampa Bay estuary. The results indicated that the viral population had a strong seasonal pattern with the highest concentrations (2.0 +- 0.8 times 10-7) in the summer and lowest (4.8 +- 1.4 times 10-6) in the winter. Viral abundance was negatively correlated with salinity (r = -0.803), and positively correlated with chl a concentration (r = 0.725). A diel study in a seawater mesocosm indicated that viral abundance did not vary on a diel rhythm, but rather peaked after a maximum in bacterial abundance and chl a. Dissolved DNA concentrations displayed diel rhythmicity, suggesting that viruses were not the main source of dissolved DNA. An estimation of the percentage of the bacterial standing stock lysed by viruses based on 4 h intervals ranged from 3.0 to 53.3% per day. Screening bacterial isolates for the presence of inducible prophages indicated that 43% were lysogens or bacteriocinogens, suggesting that lysogeny and bacteriocinogeny are common in the marine environment. [TOP OF PAGE]

255. Rapid detection of Clavibacter toxicus and of its bacteriophage responsible for annual ryegrass toxicity in Australia and the effect of selected herbicides on toxin production. Kurtboke, D.I. (1994). Actinomycetes 5:31-39. [TOP OF PAGE]

256. Effects of bacterial growth conditions and physiology on T4 infection. Kutter, E., Kellenberger, E., Carlson, K., Eddy, S., Neitzel, J., Messinger, L., North, J., Guttman, B. (1994). pp. 406-418. In In Karam, J.D. (ed.), The Molecular Biology of Bacteriophage T4. ASM Press, Washington, DC. Most studies of T4 development have been carried out under conditions that appear to be optimal, at least for the commonly used laboratory strains: rich media, highly aerated, at 37&#176C. However, there is a growing interest in understanding the ability of phage to grow under conditions more like those they meet in nature. This interest parallels the growing number of studies of Escherichia coli in more natural environments, along with an increased understanding of how bacteria respond to altered growth conditions, such as lack of oxygen, nutrient deprivation, and environmental stress (Freter, 1983, 1992; Matin et al., 1989; Roszak and Colwell, 1987; Bohannon et al., 1991; Spiro and Guest, 1991). The interest is further stimulated by the finding that, in addition to approximately 150 genes of known function, T4 has nearly 150 apparent open reading frames (ORFs) whose functions are not yet known (see the Appendix of this volume). Most of them appear likely to endode proteins and are in regions that are deletable under standard laboratory conditions. Many may have functions that are relevant only in natural environmental conditions, while others are clearly involved in the transition from host to phage metabolism. Efforts to understand the infection process and evolutionary pressures in the natural habitat(s) of T-even phages need to take into account bacterial metabolism and intracellular environments under such conditions. Thus, in this chapter we explore the evidence regarding the probable origins of T4 and examine the effects on the composition of E. coli of various environmental factors that may affect T4 growth. We review the limited number of studies of phage growth under conditions approaching those in its native habitats: low oxygen tension, nutrient limitation, exposure to nongrowing ("stationary") host cells, and temperature variations. None of these topics has been reviewed previously. [TOP OF PAGE]

257. Evolution of a lytic bacteriophage via DNA acquisition from the Lactococcus lactis chromosome. Moineau, S., Pandian, S., Klaenhammer, T.R. (1994). Appl. Environ. Microbiol. 60:1832-1841. We discovered a phage-host interaction in which the lytic phage ul36, in response to pressure exerted by an abortive phage resistance mechanism, acquired a large DNA fragment from the chromosome of Lactococcus lactis NCK203 to form a new phage, ul37. Phage ul37 was characterized at morphological, phenotypic, and genotypic levels and was found to be a member of the P335 species. Although it exhibits a high level of DNA homology with ul36, phage ul37 is resistant to the abortive mechanism and has a longer tail, a different base plate, and apparently a different origin of replication. The chromosomal DNA implicated in the formation of new phage ul37 was disrupted by site-specific integration in NCK203. This strategy prevented the appearance of ul37 during subsequent infections with ul36. [TOP OF PAGE]

258. Virsuses and DNA in marine environments. Paul, J.H., Kellogg, C.A., Jiang, S.C. (1994). pp. 119-128. In In Colwell, R.R., Simidu, U., and Ohwada, K. (eds.), Microbial Diversity in Space and Time. Plenum Press, New York, N.Y. [TOP OF PAGE]

259. Significance of the virus-rich 2-200 nm size fraction of seawater for heterotrophic flagellates: I. Impact on growth. Pesan, B.F., Weinbauer, M.G., Peduzzi, P. (1994). P. S. Z. N. I. : Mar. Ecol. 15:281-290. [TOP OF PAGE]

260. Transduction of a freshwater microbial community by a new Pseudomonas aeruginosa generalized transducing phage, UT1. Ripp, S., Ogunseitan, O., Miller, R.V. (1994). Moleculear Ecology 3:121-126. [TOP OF PAGE]

261. The significance of viruses to mortality in aquatic microbial communities. Suttle, C.A. (1994). Microb. Ecol. 28:237-243. A variety of approaches including enumeration of visibly infected microbes, removal of viral particles, decay of viral infectivity and measurements of viral production rates have been used to infer the impact of viruses on microbial mortality. The results are surpisingly consistent and suggest that, on average, about 20 % of marine heterotrophic bacteria are infected by viruses and 10-20 % of the bacterial community is lysed daily, by viruses. The effect of viruses on phytoplankton is less certain, but ca. 3 % of Synechococcus biomass may be lysed daily. The fraction of primary productivity this represents depends upon the relative biomass and growth rate of Synechococcus. Virus enrichment experiments suggest that the productivity of eukaryotic phytoplankton would be ca. 2 % higher in the absence of viruses. Overall, probably about 2-3 % of primary productivity is lost to viral lysis. There is considerable variation about these estimates; however, they represent a starting point for incorporating viral-mediated processes into aquatic ecosystem models. [TOP OF PAGE]

262. Dynamics and distribution of cyanophages and their effect on marine Synechococcus spp. Suttle, C.A., Chan, A.M. (1994). Appl. Environ. Microbiol. 60:3167-3174. Cyanophages infecting marine Synechococcus were frequently very abundant and were found in every seawater sample along a transect in the western Gulf of Mexico, and during a 28 month period in Aransas Pass, Texas. In Aransas Pass their abundance varied seasonally with the lowest concentrations coincident with cooler water and lower salinity. Along the transect, viruses infecting Synechococcus strains DC2 and SYN48 ranged in concentration from a few hundred ml^-1 at 97 m depth and 83 km offshore, to ca. 4 x 10⁵ ml^-1 near the surface at stations within 18 km of the coast. The highest concentrations occurred at the surface where salinity decreased from ca. 35.5 to 34 ppt and concentrations of Synechococcus were greatest. Viruses infecting strains SNC1, SNC2 and 838BG were distributed in a similar manner, but were much less abundant (&lt; 10 to &gt; 5 x 10³ ml^-1). When Synechococcus exceeded ca. 10³ ml^-1, cyanophage concentrations increased markedly (ca. 10² to &gt; 10⁵ ml^-1), suggesting that there was a minimum host density required for efficient viral propagation. Data on the decay rate of viral infectivity (d; d-1) as a function of solar radiation (I; mmol quanta m^-2 s^-1) was used to develop a relationship (d = 0.2610 I - 0.00718; r² = 0.69) for conservatively estimating the destruction of infectious viruses in the mixed layer of two offshore stations. Assuming that virus production balances losses, and burst size is 250, ca. 5-7 % of Synechococcus would be infected daily by viruses. Calculations based on contact rates between Synechococcus and infectious viruses produce similar results (5-14 %). Moreover, balancing estimates of viral production with contact rates for the most offshore station required that most Synechococcus be susceptible to infection, that most contacts result in infection and that the burst size be about 324 viruses per lytic event. In contrast, in nearshore waters where ca. 80 % of Synechococcus would be contacted daily by infectious cyanophages, only ca. 1 % of the contacts would have to result in infection, in order to balance the estimated virus removal rates. These results indicate that cyanophages are an abundant and dynamic component of marine planktonic communities and are likely responsible for lysing a small but significant portion of the Synechococcus population on a daily basis. [TOP OF PAGE]

263. An alternative for the detection of Vibrio cholerae in waters [Spanish]. Talledo, M.A., Gutiérrez, S., Merino, F. (1994). Revista Peruana de Epidemiología 7:30-34. [TOP OF PAGE]

264. Frequency, size and distribution of bacteriophages in different marine bacterial morphotypes. Weinbauer, M.G., Peduzzi, P. (1994). Mar. Ecol. Prog. Ser. 108:11-20. The frequencies of cells containing mature phages, the burst sizes, the phage head sizes and the distribution of phages inside cells of different bacterial morphotypes were investigated in the northern Adriatic Sea. Coccoid bacteria more frequently (2.5 %) contained mature phages than rodshaped bacteria (1.2 %) and spirillae [1.4 %). Including an estimation of non-visible infection we found that up to 27 % of rods were infected with viruses, up to 79 % of cocci and up to 100 % of spirillae. The highest overall infection frequency of the entire bacterial community was 30 %. The percentage of rods with mature phages was significantly correlated to increasing rod densities. It is suggested that a threshold density of about 2 x 10 super(5) rods/m exists that is necessary for infection with phages. No threshold densities could be determined for cocci and spirillae. Burst sizes varied strongly between different host morphotypes. The burst sizes of rods increased significantly with the frequency of rods containing mature phages, probably as a result of superinfection of bacteria with phages. [TOP OF PAGE]

265. Bacterial viruses in coastal seawater: Lytic rather than lysogenic production. Wilcox, R.M., Fuhrman, J.A. (1994). Mar. Ecol. Prog. Ser. 114:35-45. It has recently been suggested that the majority of virus production in seawater is through lysogen induction. Here, we examine the mechanism of virus production in coastal surface seawater. Bacteria and viruses were removed from seawater by ultrafiltration (0.2 or 0.02 mu-m) and introduced back to the water in 0.6 mu-m filtered inocula. Viral and bacterial abundance was followed during sample incubations using transmission electron microscopy and acridine orange direct counts. Bacteria increased in abundance in all cases. When initial abundances were less than 1.3 times 10-6 viruses ml-1, (with extremely low initial bacterial densities), viruses disappeared at a rate of 20 to 50% d-1 for the first 2 d and did not increase in abundance over the 7 d incubation period. Continuous sunlight exposure or a pulse of sunlight did not lead to detectable virus production, indicating that natural UV light did not induce lysogens. In samples with initial abundances above 1.3 times 10-6 viruses ml-1, significant increases as well as fluctuations in virus numbers were observed after the second day. This suggests that lytic infection, rather than the induction of lysogenic bacteria, is responsible for the majority of bacteriophage production in these experiments. Since lytic infection is dependent on encounter frequency, which is controlled by densities of both viruses and bacteria, the product of these 2 concentrations determines if infection may take place. We observed lytic infection only when the product of virus and bacteria numbers equalled 10-12 or more. Reported abundances of viruses and bacteria in natural seawater samples frequently fall above this level in surface waters and below it in mid and deep waters, suggesting that lytic infection may occur in all but deeper waters. However, due to the specificity of host/virus interactions, infection is also dependent on the quantitative bacterial species composition and diversity, which are presently unknown. [TOP OF PAGE]

266. Spatially resolved evolution of viruses. Yin, J. (1994). Annals of the New York Academy of Science 745:399-408. [TOP OF PAGE]

267. The isolation of T-even phages (feces -?-> P.C./PC --> gamma --> T2; Sewage -?-> T4/T6). Abedon, S.T. (1993). T4 News (Feb 17). It may never prove possible to identify, with high confidence, the time, place, and source material for the original isolation of phages T2, T4, and T6. Nevertheless, a better understanding of where T-even phages came from has historical significance and is even of scientific consequence to those of us interested in the ecology of these organisms. Gathering clues for the discovery of a likely candidate for the original isolation of phage T2 was the more difficult task. For phages T4 and T6 this search was much less problematic and is presented first. I conclude with a conjecture on the history of free tryptophan (trp) adsorption cofactor dependence in these phages. <a href="sta0003.htm">Full text is also available</A>. [TOP OF PAGE]

268. Experimental molecular evolution of bacteriophage T7. Bull, J.J., Cunningham, C.W., Molineux, I.J., Badgett, M.R., Hills, D.M. (1993). Evolution 47:993-1007. We present an analysis of molecular evolution in a laboratory- generated phylogeny of the bacteriophage T7, a virus of 40 kilo- base pairs of double-stranded DNA. The known biology of T7 is used in concert with observed changes in restriction sites and in DNA sequences to produce a model of restriction-site convergence and divergence in the experimental lineages. During laboratory propagation in the presence of a mutagen, the phage lineages changed an estimated 0.5% 1.5% in base pairs; most change appears to have been G fwdarw A or C fwdarw T, presumably because of the mutagen employed. Some classes of restriction-site losses can be explained adequately as simple outcomes of random processes, given the mutation rate and the bias in mutation spectrum. However, some other classes of sites appear to have undergone accelerated rates of loss, as though the losses were selectively favored. Overall, the wealth of knowledge available for T7 biology contributes only modestly to these explanations of restriction-site evolution, but rates of restriction-site gains remain poorly explained, perhaps requiring an even deeper understanding of T7 genetics than was employed here. Having measured these properties of molecular evolution, we programmed computer simulations with the parameter estimates and pseudo- replicated the empirical study, thereby providing a data base for statistical evaluation of phylogeny reconstruction methods. By these criteria, replicates of the experimental phylogeny would be correctly reconstructed over 97% of the time for the three methods tested, but the methods differed significantly both in their ability to recover the correct topology and in their ability to predict branch lengths. More generally, the study illustrates how analyses of experimental evolution in bacteriophage can be exploited to reveal relationships between the basics of molecular evolution and abstract models of evolutionary processes. [TOP OF PAGE]

269. Spatial distribution of viruses, bacteria and chlorophyll a in neritic, oceanic and estuarine environments. Cochlan, W.P., Wikner, J., Steward, G.F., Smith, D.C., Azam, F. (1993). Mar. Ecol. Prog. Ser. 92:77-87. The spatial distribution of viruses was investigated in the coastal and oceanic waters of the Southern California Bight, USA, and the brackish waters of the Gulf of Bothnia, Sweden, using the direct harvesting technique and transmission electron microscopy. The vertical and horizontal distributions of viruses were examined in relation to bacterial abundance and chlorophyll a. Total virus abundances ranged from 0.3 to 52 times 10-9 l-1; higher concentrations of viruses were found in the upper 50 m of the water column and in coastal environments. Viruses with capsid diameters less than 60 nm dominated the virus community, were morphologically characterized as bacteriophages and were responsible for most of the observed spatial variability. Bacteria abundance alone explained 67% of the spatial variability in virus numbers, thereby suggesting that bacteria constituted the major host organisms for viruses in these physically diverse habitats. [TOP OF PAGE]

270. Lytic bacteriophages of Streptococcus mutans. Delisle, A.L., Rostkowski, C.A. (1993). Current Microbiology 27:163-167. Three phages of Streptococcus mutans were obtained and partially characterized. The three phages, designated M102, e10, and f1, were found to be strictly lytic, with host ranges restricted to only serotype c, e, and f strains of this species, respectively. Phage sensitivity was not correlated with the presence of plasmids, at least in host strains of serotypes c and e. Each phage produced clear plaques in a number of standard media, even in the presence of sucrose, indicating that the extracellular glucan polysaccharides (mutan) produced by the hosts from this substrate do not prevent phage adsorption and growth. The phages were similar in size and morphology, having icosahedral heads and long (283-287 nm), flexible, noncontractile tails. The genome of each phage was found to consist of linear, double-stranded DNA, 31-35 kb in length, with a base composition of 37-38% G + C. Restricting phage DNAs with four enzymes produced fragment patterns unique to each phage, but common bands between M102 and e10 and between e10 and f1 were produced by BamHI. Labeled e10 and M102 DNAs hybridized strongly with all three phage DNAs, indicating that they share some common sequences. The three phages appear to be more similar than expected and probably evolved from a common ancestor. [TOP OF PAGE]

271. Characterization of bacteriophages of Erwinia ananas. Eayre, C.G., Robertson, N.L. (1993). Phytopathology 83:1354 [TOP OF PAGE]

272. Viruses in marine planktonic systems. Fuhrman, J.A., Suttle, C.A. (1993). Oceanography 6:50-62. The last 10-15 years have seen major changes in our views of marine planktonic food webs, primarily from the realization that prokaryotic microorganisms and small eukaryotes are responsible for a significant fraction, often 50% or more, of the primary production and heterotrophic consumption of organic matter in these systems. However, it has only been in the past few years that marine scientists have investigated the roles of viruses in ecological processes. Although this research area is only in its infancy, early results suggest that viruses may be important agents in the mortality or marine microorganisms and in controlling their genetic compositions. This paper will summarize the experiments and measurements that have led to this suggestion as well as the conceptual frame-work within which they are interpreted. [TOP OF PAGE]

273. Viruses in marine food webs. Fuhrman, J.A., Wilcox, R.M., Noble, R.T., Law, N.C. (1993). pp. 295-298. In In Guerrero, R. and Pedros-Alio, C. (eds.), Trends in microbial ecology. Spanish Society for Microbiology, Barcelona. [TOP OF PAGE]

274. Grazing by marine nanoflagellates on viruses and virus-sized particles: Ingestion and digestion. González, J.M., Suttle, C.A. (1993). Mar. Ecol. Prog. Ser. 94:1-10. We examined grazing of marine viruses and bacteria by natural assemblages and cultures of phagotrophic nanoflagellates. Ingestion rates were determined using fluorescently-labelled viruses (FLVs) and bacteria (FLB), and 50 or 500 nm-diameter fluorescent microspheres (FMs). Calculated clearance rates of viruses by natural nanoflagellate assemblages were about 4 % of those for bacteria when the bacteria and viruses were present at natural concentrations. Different viruses were ingested at different rates with the smallest virus being ingested at the slowest rate. As well, we found differences in digestion times for the same flagellates grazing on different viruses and for different flagellate assemblages grazing on the same viruses. 50 nm FMs were used as a control for egestion of undigested particles. As rates of digestion were greater than those for ingestion both processes would occur simultaneously; hence, our estimates of grazing rate are likely conservative. Ingestion rates were positively correlated with the concentration of 50 nm FMs. Discrimination against 50 nm FMs in favor of FLVs was also observed. Our calculations suggest that viruses may be of nutritional significance for phagotrophic flagellates. When there are 10⁶ bacteria ml^-1 and 10⁷ to 10⁸ viruses ml^-1, viruses may represent 0.2-9 % of the carbon, 0.3-14 % of the nitrogen and 0.6-28 % of the phosphorus that the flagellates obtain from ingestion of bacteria. This study demonstrates that both natural assemblages and cultures of phagotrophic nanoflagellates consume and digest a variety of marine viruses, thereby deriving nutritional benefit and serving as a natural sink for marine viral particles. In addition, these results imply that some nanoflagellates are likely capable of consuming a wide spectrum of organic particles in the colloidal size range. [TOP OF PAGE]

275. A study of five bacteriophages of the Myoviridae family which replicate on different gram-positive bacteria. Jarvis, A.W., Collins, L.J., Ackermann, H.W. (1993). Archives of Virology 133:75-84. A comparative study is reported on five phages of the Myoviridae family which propagate on Bacillus subtilis, B. thuringiensis, Enterococcus sp., Lactobacillus plantarum, or Staphylococcus aureus. The phages are morphologically identical and characterized by isometric heads with conspicuous capsomers and by contractile tails with complex base plates. The phages show similar protein profiles, but vary considerably in burst size. Phage DNAs are about 95-166 kb in size and are unrelated by DNA-DNA hybridization and restriction endonuclease analysis. Therefore the phages are unrelated at species level. Implications of these data for our understanding of the development of phage species are discussed. [TOP OF PAGE]

276. DNA homology in species of bacteriophages active on Pseudomonas aeruginosa. Krylov, V.N., Tolmachova, T.O., Akhverdian, V.Z. (1993). Archives of Virology 131:141-151. [TOP OF PAGE]

277. Taxonomic implications of the reactions of representative Bacillus strains to Thermoactinomyces-phage. Kurtboke, D.I., Sivasithamparam, K. (1993). Actinomycetes 4:1-7. [TOP OF PAGE]

278. Occurrence of Actinomadura phage in organic mulches used for avocado plantations in Western Australia. Kurtboke, D.I., Wilson, C.R., Sivasithamparam, K. (1993). Canadian Journal of Microbiology 39:389-394. Phage infecting Actinomadura verrucosospora, Actinomadura pusilla, and Nocardiopsis flava were isolated from mulches applied to avocado trees at the University of Western Australia experimental plots. The physiochemical properties, plaque morphology, host range, and particle morphology of the phage isolated are described. Taxonomic implications of the reactions of some cell wall chemotype III actinomycetes to Actinomadura phage are also discussed. [TOP OF PAGE]

279. Use of bacteriophage for the selective isolation of thermophilic actinomycetes from composted eucalyptus bark. Kurtboke, D.I., Murphy, N.E., Sivasithamparam, K. (1993). Canadian Journal of Microbiology 39:46-51. A method was developed to reduce the numbers of thermophilic bacteria on isolation plates, which in turn facilitated the detection and isolation of thermophilic actinomycetes. The method involves exposing the test material to bacteriophage suspensions prior to inoculation on isolation plates. This method was applied to composted eucalyptus bark samples, which were then inoculated on R8 and 1/2 TSA + 0.2% casein hydrolysate agar plates. The phage susceptibility of thermophilic bacteria provided a selective means of reducing their numbers on isolation plates and hence increased the numbers of Thermomonospora, Saccharopolyspora rectivirgula, and thermophilic Streptomyces spp. on these media in comparison with the numbers recorded from control plates. [TOP OF PAGE]

280. Ch2, a novel halophilic archaeon from an Australian solar saltern. Nuttall, S.D., Dyall-Smith, M.L. (1993). International Journal of Systematic Bacteriology [INT. J. SYST. BACTERIOL. ] 43:729-734. A novel halophilic archaeon, strain Ch2, was isolated from a marine solar saltern in Geelong, Australia. The fact that this organism had a dam-methylated genome suggested that it is closely related to the taxon that includes Halobacterium saccharovorum, Halobacterium sodomense, and Holobacterium trapanicum. A sequence analysis of the 16S rRNA gene (Ch2 has three copies of this gene) showed that Ch2 is phylogenetically equidistant from the genera Haloarcula and Haloferax and closely related to H. saccharovorum. The susceptibility of both Ch2 and H. saccharovorum to the recently isolated halophage HF2 supported the hypothesis that these two organisms are closely related. [TOP OF PAGE]

281. HF1 and HF2: Novel bacteriophages of halophilic archaea. Nuttall, S.D., Dyall-Smith, M.L. (1993). Virology 197:678-684. Two novel halophilic archaebacterial bacteriophages, HF1 and HF2, were isolated from an Australian solar saltern. They were morphologically identical with icosahedral-shaped heads (diameter 58 nm) and contractile tails (length 94 nm). Other similarities included sensitivity to reduced ionic conditions, similar protein profiles by SDS-PAGE, and dsDNA genomes of identical size (73.5 kbp) with analogous restriction patterns. DNA-DNA hybridization data showed the two phages to be closely related. HF1 has a broad host-range, infecting members of three halobacterial genera including Halobacterium salinarium and the genetically well-characterized strain Haloferax volcanii WFD11. Mutants showing increased plating efficiency on alternative hosts were readily selectable. By contrast, HF2 showed a limited host range, confined to the closely related dam-methylated strains Ch2 and H. saccharovorum. [TOP OF PAGE]

282. Distribution of viral abundance in the reef environment of Key Largo, Florida. Paul, J.H., Rose, J.B., Jiang, S.C., Kellogg, C.A., Dickson, L. (1993). Appl. Environ. Microbiol. 59:718-724. The distribution of viral and microbial abundance in the Key Largo, Fla., reef environment was measured. Viral abundance was measured by transmission electron microscope direct counts and plaque titer on specific bacterial hosts in water and sediment samples from Florida Bay (Blackwater Sound) and along a transect from Key Largo to the outer edge of the reef tract in Key Largo Sanctuary. Water column viral direct counts were highest in Blackwater Sound of Florida Bay (1.2 times 10-7 viruses per ml), decreased to the shelf break (1.7 times 10-6 viruses per ml), and were inversely correlated with salinity (r = -0.97). Viral direct counts in sediment samples ranged from 1.35 times 10-8 to 5.3 times 10-8/cm-3 of sediment and average nearly 2 orders of magnitude greater than counts in the water column. Viral direct counts (both sediment and water column measurements) exceeded plaque titers on marine bacterial hosts (Vibrio natriegens and others) by 7 to 8 orders of magnitude. Water column viral abundance did not correlate with bacterial direct counts or chlorophyll a measurements, and sediment viral parameters did not correlate with water column microbial, viral, or salinity data. Coliphage, which are indicators of fecal pollution, were detected in two water column samples and most sediment samples, yet their concentrations were relatively low (2 to 15/liter for water column samples, and lt 2 to 108/cm-3 of sediment). Our findings indicate that viruses are abundant in the Key Largo environment, particularly on the Florida Bay side of Key Largo, and that processes governing their distribution in the water column (i.e., salinity and freshwater input) are independent of those governing their distribution in the sediment environment. [TOP OF PAGE]

283. Effect of the submicron size fraction including viruses on the formation of algal flocs (marine snow) in the sea. Peduzzi, P., Weinbauer, M.G. (1993). Limnol. Oceanogr. 38:1562-1565. [TOP OF PAGE]

284. Effect of concentrating the virus-rich 2-200 nm size fraction of seawater on the formation of algal flocs (marine snow). Peduzzi, P., Weinbauer, M.G. (1993). Limnol. Oceanogr. 87:105-112. [TOP OF PAGE]

285. The submicron size fraction of seawater containing high numbers of virus particles as bioactive agent in unicellular plankton community successions. Peduzzi, P., Weinbauer, M.G. (1993). Jour. Plankt. Res. 15:1375-1386. [TOP OF PAGE]

286. Calibrating estimates of phage-induced mortality in marine bacteria ultrastructural studies of marine bacteriophage development from one-step growth experimetns. Proctor, L.M., Okubo, A., Fuhrman, J.A. (1993). Microb. Ecol. 25:161-182. The timing of lytic phase development and the relationship between host generation times and latent periods were investigated by electron microscopy of one-step growth experiments in two strains of marine Vibrio species. Results were used in a correction factor developed to interpret field studies of phage-infected marine bacteria. Both the number of mature phase per average cell section and the percentage of cells with mature phase increased exponentially by 73-86% into the latent periods. Assuming that bacterial infection and lysis take place continually in the ocean, conversion factors for relating the percentage of visibly infected bacteria to the total percentage of the bacterial community that are phage-infected were calculated as 3.70-7.14. When this range of factors was applied to previously-collected field data Proctor LM, Fuhrman JA (1990) Nature (Lond) 343:60-2; Proctor LM, Fuhrman JA (1991) Mar Ecol Prog Ser 69:133-142 from 3 to 31% of the free-living bacteria and 3 to 26% of particulate-associated bacteria appeared to be phage-infected at any given time. Based upon a steady-state model in which half the daughter cells survive to divide again, the percent of total mortality would be twice the total percentage of phage-infected cells. From 6 to 62% and from 6 to 52% of mortality for the free-living and particulate-associated bacterial community, respectively, may be due to viruses. [TOP OF PAGE]

287. Biological inactivation of adhering Listeria monocytogenes by listeriaphages and a quaternary ammonium compound. Roy, B., Ackermann, H.W., Pandian, S., Picard, G., Goulet, J. (1993). Appl. Environ. Microbiol. 59:2914-2917. The use of listeriaphages as a means of disinfecting contaminated stainless-steel and polypropylene surfaces was investigated. Surfaces artificially contaminated with L. monocytogenes 10401 and 8427 were sanitized with suspensions of listeriaphages (H387, H387-A, and 2671), all belonging to the Siphoviridae family. Phage suspensions at concentrations of up to 3.5 x 10(8) PFU/ml were at least as efficient as a 20 ppm solution of a quaternary ammonium compound (QUATAL) in reducing L. monocytogenes populations. A synergistic activity was observed when two or more phages were used in combination and when phages were suspended in QUATAL. The biological activity of the three phages was not affected by QUATAL concentrations of 50 ppm and a contact time of 4 h. [TOP OF PAGE]

288. Cyanophages and sunlight: A paradox. Suttle, C.A., Chan, A.M., Chen, F., Garza, D.R. (1993). pp. 303-307. In In Guerrero, R. and Pedros-Alio, C. (eds.), Trends in Microbial Ecology. Spanish Society Microbiology, Barcelona. [TOP OF PAGE]

289. Marine cyanophages infecting oceanic and coastal strains of Synechococcus: Abundance, morphology, cross-infectivity and growth characteristics. Suttle, C.A., Chan, A.M. (1993). Mar. Ecol. Prog. Ser. 92:99-109. Eight different phycoerythrin- and phycocyanin-containing strains of Synechococcus and one strain of Anacystis were screened against 29 natural virus communities taken from 3 locations in south Texas coastal waters, at different times of the year. In addition, one sample was screened from Peconic Bay, New York. Cyanophages were detected in all samples, but the frequency with which they were detected and their abundance depended upon the strain of Synechococcus that was screened. Viruses that infected red Synechococcus strains were particularly common. In some instances, concentrations infecting a single Synechococcus strain were in excess of 10⁵ ml^-1. The abundances of cyanophages were weakly correlated with temperature (r² = 0.53 to 0.70), although they occurred at all of the temperatures (12-30.4^oC) and salinities (18-70 ppt) that were screened. The seven cyanophages that were cloned belonged to the same three families of viruses that have been observed to infect freshwater cyanobacteria, namely the Siphoviridae (formerly Styloviridae), Myoviridae and Podoviridae. The cyanophage clones varied in host-specificity. For example, one clone infected a single Synechococcus strain of 12 that were tested, whereas, another infected 4 of 9 strains tested. Growth characteristics of one of the virus clones was determined for a single host. Photosynthesis in Synechococcus was not affected until near the onset of cell lysis and the virus burst cycle was complete about 17 h post-infection. The burst size was approximately 250 infective particles. The high abundance of cyanophages in the natural environment provides further evidence that viruses are probably important regulators of phytoplankton dynamics in marine systems. [TOP OF PAGE]

290. Enumeration and isolation of viruses. Suttle, C.A. (1993). pp. 121-134. In In Kemp, P.F., Sherr, B.F., Sherr, E.F., and Cole, J.J. (eds.), Current Methods in Aquatic Microbial Ecology. Lewis Publ., Boca Raton. [TOP OF PAGE]

291. Distribution of viruses and dissolved DNA along a coastal trophic gradient in the northern Adriatic Sea. Weinbauer, M.G., Fuks, D., Peduzzi, P. (1993). Appl. Environ. Microbiol. 59:4074-4082. The distribution of viral and other microbial abundances as well as the concentrations of dissolved DNA (D-DNA) along a trophic gradient in the northern Adriatic Sea were determined. Virus abundances, covering a range of 1.2 times 10-9 to 8.7 times 10-10 liter-1 were on average 2.5-fold higher in eutrophic than in mesotrophic stations. A 2.5-fold enrichment was also measured for chlorophyll a concentrations, whereas the densities of bacteria and heterotrophic nanoflagellates were only approximately 1.5-fold higher. The frequency of bacteria containing mature phage increased linearly with bacterial abundance. Assuming that mature phage is only visible during the last 14 to 27% of the latent period (L. M. Proctor, A. Okubo, and J. A. Fuhrman, Microb. Ecol. 25:161-152, 1993), we estimated that between 3.5 and 7.3% of the bacterial population was infected at mesotrophic stations versus between 7.0 and 19.5% at eutrophic stations, indicating that the bacterial mortality due to viral lysis might increase with the degree of eutrophication. The frequency of bacteria with mature phage and the burst size varied significantly with the bacterial morphotype; rod-shape cells, the most abundant morphotype, showed low infection rates but a high burst size. Concentrations of D-DNA varied significantly with season but not with trophic conditions. The estimated percentage of viral DNA on total D-DNA concentrations averaged 17.1% (range, 0.7 to 88.3%). Some kind of interaction between heterotrophic nanoflagellates and viruses is proposed. We conclude (i) that the significance of viruses varies with changing trophic conditions and (ii) that viral activity may play a significant role in food web structure under changing trophic conditions. [TOP OF PAGE]

292. Nucleic acids from the host bacterium as a major source of nucleotides for three marine bacteriophages. Wikner, J., Vallino, J.J., Steward, G.F., Smith, D.C., Azam, F. (1993). FEMS Microbiol. Ecol. 12:237-248. The incorporation of ³²P-phosphorus into marine bacteriophage nucleic acid was studied in culture experiments to investigate the source of nucleotides used by the phage. We consistently found that the ³²P-specific activity in the phage genome increased during the 11 h incubation and was low relative to the specific activity in the medium, averaging 21% (.+-. SD 5.9) for the three phage isolates. This was in accordance with a mathematical model where most of the nucleotides for phage DNA synthesis were derived from the host cell nucleic acid rather than de novo synthesis. We propose that this metabolic strategy may be common among marine phages, as an adaptation to a nutrient poor environment. Consequently, the contribution of free DNA to the dissolved fraction through phage lysis of bacteria, may be less than previously thought. Also during radiolabelling of bacteriophages in natural water samples, isotope dilution may be dependent on the specific growth rate of the bacterial host. [TOP OF PAGE]

293. Isolation and molecular characterization of five marine cyanophages propagated on Synechococcus sp. strain WH7803. Wilson, W.H., Joint, I.R., Carr, N.G., Mann, N.H. (1993). Appl. Environ. Microbiol. 59:3736-3743. Five marine cyanophages propagated on Synechococcus sp. strain WH7803 were isolated from three different oceanographic provinces during the months of August and September 1992: coastal water from the Sargasso Sea, Bermuda; Woods Hole harbor, Woods Hole, Mass.; and coastal water from the English Channel, off Plymouth Sound, United Kingdom. The five cyanophage isolates were found to belong to two families, Myoviridae and Styloviridae, on the basis of their morphology observed in the transmission electron microscope. DNA purified from each of the cyanophage isolates was restricted with a selection of restriction endonucleases, and three distinguishably different patterns were observed. DNA isolated from Myoviridae isolates from Bermuda and the English Channel had highly related restriction patterns, as did DNA isolated from Styloviridae isolates from Bermuda and the English Channel. DNA isolated from the Myoviridae isolate from Woods Hole had a unique restriction pattern. The genome size for each of the Myoviridae isolates was ca. 80 to 85 kb, and it was ca. 90 to 100 kb for each of the Styloviridae isolates. Southern blotting analysis revealed that there was a limited degree of homology among all cyanophage DNAs probed, but clear differences were observed between cyanophage DNA from the Myoviridae and that from the Styloviridae isolates. Polypeptide analysis revealed a clear difference between Myoviridae and Styloviridae polypeptide profiles, although the major, presumably structural, protein in each case was ca. 53 to 54 kDa. [TOP OF PAGE]

294. Evolution of bacteriophage T7 in a growing plaque. Yin, J. (1993). J. Bacteriol. 175:1272-1277. The emergence of mutants during the 10⁹-fold amplification of a bacteriophage was spatially resolved in a growing plaque. When wild-type phage T7 was grown on an Escherichia coli host which expressed an essential early enzyme of the phage infection cycle, the T7 RNA polymerase, mutant phage relying on this enzyme appeared in 10⁸ phage replications and outgrow the wild type. Spatial resolution of the selection process was achieved by analyzing stab samples taken along a plaque radius. Different mutants were selected at different rates along different radii of the plaque, based on host range and restriction patterns of the isolates. The mutants deleted up to 11% of their genomes, including the gene for their own RNA polymerase. They gained an advantage over the wild type by replicating more efficiently, as determined by one-step growth cultures. [TOP OF PAGE]

295. Lysis of lysis inhibited bacteriophage T4 infected cells. Abedon, S.T. (1992). J. Bacteriol. 174:8073-8080. T4 bacteriophage (phage)-infected cells show a marked increase in latent-period length, called lysis inhibition, upon adsorption of additional T4 phages (secondary adsorption). Lysis inhibition is a complex phenotype requiring the activity of at least six T4 genes. Two basic mysteries surround our understanding of the expression of lysis inhibition: (i) the mechanism of initiation (i.e., how secondary adsorption leads to the expression of lysis inhibition) and (ii) the mechanism of lysis (i.e., how this signal not to lyse is reversed). This study first covers the basic biology of the expression of lysis inhibition and lysis of T4-infected cells at high culture densities. Then evidence is presented which implies that, as with the initiation of lysis inhibition, sudden, lysis-associated clearing of these cultures is likely caused by T4 secondary adsorption. For example, such clearing is often observed for lysis-inhibited T4-infected cells grown in batch culture during T4 stock preparation. The significance of this secondary adsorption-induced lysis to wild T4 populations is discussed. The study concludes with a logical argument suggesting that the lytic nature of the T4 phage particle evolved as a novel mechanism of phage-induced lysis. [TOP OF PAGE]

296. Molecular genetics of adaptation in an experimental model of cooperation. Bull, J.J., Molineux, I.J. (1992). Evolution 46:882-895. The evolution of cooperation was studied in an empirical system utilizing a parasitic bacteriophage (fl) and a bacterial host. Infected cells were propagated by serial passage so that a phage could increase its representation among infected hosts only by enhancing the rate of growth of its host. Loss of infectivity was therefore without selective penalty, and phage benevolence could potentially evolve through a variety of genetic changes. The infected hosts evolved to grow faster over the course of the study, but the genetic bases of this phenotypic change were more difficult to anticipate. Two fundamentally different types of genetic changes in the phage were revealed. One involved the loss of some phage genes, resulting in a noninfectious plasmid that continued to replicate via the parental phage replicon. The second change involved integration of the phage to genome into host DNA by a process that, at low frequency, could be reversed to produce infectious phage particles. Integration is a previously known property of wild-type fl, and in the system studied, may have resulted from the use of a phage bearing an insert containing nonfunctional DNA. The evolution of this novel function apparently depended only on the presence of a small region in the phage genome that provided some homology to the host DNA, with the host providing all necessary functions. Although fl is one of the simplest phages known, these observations suggest that host-parasite interactions of the filamentous phages are more complicated than previously thought. More generally, the fl system offers a useful model for many problems concerning the genetic basis of adaptation. [TOP OF PAGE]

297. Phages and phage resistance in lactococci. Actes du Colloque LACTIC 91. Daly, C., Coffey, A., Casey, C.N., Fitzgerald, G.F. (1992). In none (ed.), Les Bacteries Lactiques, Recherche et Applications Industrielles en Agro-Alimentaire. Adria Normandie, Villers Bocage, Cedex, France. [TOP OF PAGE]

298. Bacterioplankton roles in cycling of organic matter: the microbial food web. Fuhrman, J.A. (1992). pp. 361-383. In In Falkowski, P.G. and Woodhead, A.D. (eds.), Primary Productivity and Biogeochemical Cycles in the Sea. Plenum, New York. [TOP OF PAGE]

299. Experimental phylogenetics: generation of a known phylogeny. Hillis, D.M., Bull, J.J., White, M.E., Badgett, M.R., Molineux, I.J. (1992). Science 255:589-592. Although methods of phylogenetic estimation are used routinely in comparative biology, direct tests of these methods are hampered by the lack of known phylogenies. Here a system based on serial propagation of bacteriophage T7 in the presence of a mutagen was used to create the first completely known phylogeny. Restriction-site maps of the terminal lineages were used to infer the evolutionary history of the experimental lines for comparison to the known history and actual ancestors. The five methods used to reconstruct branching pattern all predicted the correct topology but varied in their predictions of branch lengths; one method also predicts ancestral restriction maps and was found to be greater than 98 percent accurate. [TOP OF PAGE]

300. Use of polyvalent phage for reduction of Streptomycetes on soil dilution plates. Kurtboke, D.I., Chen, C.F., Williams, S.T. (1992). J. Appl. Bacteriol. 72:103-111. An isolation method was developed in which prior to inoculation soil suspensions were exposed to suspensions of polyvalent phage isolated to Streptomyces spp. The phage susceptibility of streptomycetes provided a selective means of reducing streptomycetes on isolation plates subsequent to inoculation, and this reduction was persistent after long incubation periods. The efficiency and applicability of the method developed were checked with different samples from a range of sources. The increased chances of development of other genera after the reduction of streptomycetes on soil dilution plates were assessed. [TOP OF PAGE]

301. Origin of phage particles in culture fluids during the industrial production of fodder protein with the use of methanotrophic bacteria: The role of the bacteria in the discontinuing the fermentation process. Lobanov, A.O., Turin, V.S., Krylov, V.N. (1992). Biotekhnologiya 4-8. There is a direct confirmation of the possibility that the phage lysis of the attending cultures may cause the death of the main culture in the article. Besides reviewed are the properties as follows: data received during electronic-microscope observations of bacteria during continous fermentation proving the existing phages, specific for bacteria, which are in the special physiological state caused probable by the continuous character of fermentation. [TOP OF PAGE]

302. Characterization of lactococcal bacteriophages from Quebec cheese plants. Moineau, S., Fortier, J., Ackermann, H.-W., Pandian, S. (1992). Canadian Journal of Microbiology 38:875-882. This is the first study on the characterization of lactococcal phages isolated in Canada. Thirty lactococcal phages were isolated from Quebec cheese plants reporting partial loss of starter activity. Phages were characterized by electron microscopy, DNA homology, protein profile, and host range. All phages belonged to the Siphoviridae family. Seventeen phages (57%) has prolate heads (60 times 40 nm) and 100 nm long, noncontractile tails (morphotype B2, species c2). They showed strong DNA homology with other prolate-headed phages isolated from other countries (Australia, United States). In addition to normal prolate phages, most lysates contained pairs of empty heads (no DNA) connected by a small bridge. Thirteen phages (43%) had small isometric heads (55 nm in diameter) and long, noncontractile tails (morphotype B1). Based on DNA homology, 11 of these phages were found related to phage species 936 despite differences in tail length (140 to 200 nm). The two other small isometric phages, UL36 and UL39, hybridized with phage P335 DNA, and therefore belong to this species. No DNA homology was observed between prolate and small isometric phages. Phages with prolate heads showed a broader host range than small isometric-headed phages. The DNA of phage UL36, which has a relatively narrow host range, has more restriction endonuclease sites than other lactococcal bacteriophages. [TOP OF PAGE]

303. Mortality of marine bacteria in response to enrichments of the virus size fraction from seawater. Proctor, L.M., Fuhrman, J.A. (1992). Mar. Ecol. Prog. Ser. 87:283-293. The potential for viral lysis of marine bacteria in seawater enriched with the virus size fraction from seawater was investigated in seawater samples from Long Island Sound, USA, the eastern Pacific Ocean and the Caribbean Sea. Ultrafiltration was used to concentrate material from seawater in the > 0.05 .mu.m to <0.22 .mu.m size fraction. Electron microscopy counts of virus-like particles in the high molecular weight concentrate (HMWC) correlated closely to epifluorescence microscopy counts of <0.22 .mu.m DAPI-positive, DNase-resistant particles of the HMWC. Acridine orange direct counts of bacterial abundances significantly declined (often to 1/2 or less of controls) when seawater was incubated with 4- to 16-fold enrichnments of HMWC. Microwave treatment of the HMWC before addition to seawater virtually eliminated the declines in bacterial abundance. The combined evidence of the size range of particles, the heat lability of the HMWC and the presence of <0.22 .mu.m DAPI-fluorescent, DNase-resistant particles by epifluorescence microscopy and abundant virus particles by electron microscopy suggests that a dominant bacterial mortality agent in the seawater concentrate was bacteriophage, although we could not rule out an effect of high molecular weight proteins. [TOP OF PAGE]

304. Characteristics and diffusion in the rabbit of a phage for Escherichia coli 0103. Attempts to use this phage for therapy. Reynaud, A., Cloastre, L., Bernard, J., Laveran, H., Ackermann, H.W., Licois, D., Joly, B. (1992). Veterinary Microbiology 30:203-212. A bacteriophage for Escherichia coli 0103 was isolated during a study on E. coli diarrhoea in intensive breeding units of rabbits. The phage had an isometric head and a short tail and resembled coliphage N4 (Podoviridae). It had a very narrow host range and seemed to be specific for serogroup 0103, suggesting that it might be used for preliminary identification of E. coli strains of this serogroup instead of the usual slide agglutination. In view of its possible use as a therapeutic phage, we investigated its dissemination in rabbit organs after oral administration. The phage persisted in the spleen for at least 12 days. However, in vivo studies showed that this phage and a mixture of more virulent phages for E. coli 0103 were ineffective in preventing disease in rabbits inoculated with an enteropathogenic strain of E. coli 0103. [TOP OF PAGE]

305. Virus and bacteria abundances in the Drake Passage during January and August 1991. Smith, D.C., Steward, G.F., Azam, F., Hollibaugh, J.T. (1992). Antarct. J. U. S. 27:125-126. no abstract. [TOP OF PAGE]

306. Estimation of virus production in the sea: II. Field results. Steward, G.F., Wikner, J., Cochlan, W.P., Smith, D.C., Azam, F. (1992). Mar. Microb. Food Webs. 6:79-90. Virus production was estimated in samples from diverse marine environments by incorporation of super(32)P-orthophosphate ( super(32)P sub(i)) into viral DNA. Rates of virus production in the field were found to vary over three orders of magnitude (from < 1 x 10 super(8) to 2.3 x 10 super(11) viruses/l/d). Due to the possibility of intracellular isotope dilution and filtration losses, these rates should be considered minimum estimates. Virus production displayed a strong onshore-offshore gradient which covaried with bacteria abundance and chlorophyll a. Minimum estimates of mortality in two coastal samples (12 and 25% of bacteria production) suggest that viruses were a significant source of bacteria mortality in these environments. The apparent dependence of virus production on host density suggests that significant phage attack is spatially and temporally episodic in the open ocean (e.g. during phytoplankton blooms), but more persistent in the highly productive coastal waters. [TOP OF PAGE]

307. Estimation of virus production in the sea. I. Method development. Steward, G.F., Wikner, J., Smith, D.C., Cochlan, W.P., Azam, F. (1992). Marine Microbial Food Webs 6:57-78. A method for estimating virus production in seawater was developed and tested in the field. The rates of virus production were determined on the basis of the rate of ³²P-orthophosphate (³²Pi) or ³H-thymidine (³H-TdR) incorporation specifically into the nucleic acid of the viruses released during the incubation period. The ³H-TdR-incorporation method was designed to specifically detect the production of DNA-containing bacterial viruses. The ³²Pi-incorporation method has the potential to measure the production of DNA and RNA viruses of bacteria, algae and protozoa. The ³²Pi-incorporation protocols developed in this study, however, measure only the production of DNA viruses. The method provides sensitive detection of virus production with high precision and low blanks. Conversion factors relating label incorporated to viruses produced were derived empirically from culture and field measurements. Due to variable and potentially biased losses of viruses, the accuracy of the method could not be determined in the present study. Virus production measured by this method must be considered minimum estimates at present. This method should be of value in studies of the ecology of viruses in the sea and the roles of viruses in the ocean's biogeochemical dynamics. [TOP OF PAGE]

308. Marine viruses: Decay rates, diversity and ecological implications. Suttle, C.A., Chen, F., Chan, A.M. (1992). pp. 153-163. In In Nash, C.C., II (ed.), International Marine Biotechnology Conferences "IMBC-91": Short Communications of Invited Lectures. W. Brown Co., Dubuque. [TOP OF PAGE]

309. Inhibition of photosynthesis by the submicron size fraction concentrated from seawater. Suttle, C.A. (1992). Mar. Ecol. Prog. Ser. 87:105-112. Ultrafiltration was used to concentrate the 2-200 nm size fraction from seawater samples 100-1000 fold. Electron microscopy indicated that these concentrates were heavily enriched with virus-like particles (VLPs). When aliquots from these concentrates were added to natural seawater samples, primary productivity (¹⁴C-bicarbonate uptake) was either little affected or suppressed within minutes by up to 78 %. In some instances as little as a 20 % increase in the concentration of the 2-200 nm size fraction in seawater reduced relative carbon fixation rates by nearly 50 %. Autoclaving the concentrates before addition reduced or eliminated the bioactive effect. Larger additions caused greater inhibition, although a point was reached where further additions of concentrate did not result in greater photosynthetic suppression, relative to samples to which autoclaved concentrate was added. Microautoradiography indicated that there were fewer photosynthetically-active cells in seawater samples to which untreated concentrates were added, when compared to samples receiving autoclaved concentrate. Phytoplankton &gt;3 &micro;m were affected the most. This indicated that specific organisms were affected as opposed to photosynthesis being reduced throughout the community. In vivo chlorophyll fluorescence paralleled changes in carbon incorporation, and increased most slowly when the greatest additions of concentrate were made. Nonetheless, growth of phytoplankton recovered several days after the concentrates were added, suggesting either that some cells were unaffected by the addition, or that resistance to the bioactive agent(s) developed over time. The data provide further evidence for the presence of a strongly bioactive component that inhibits primary productivity, which is associated with the virus-rich, 2 to 200 nm size fraction of seawater. [TOP OF PAGE]

310. Mechanisms and rates of decay of marine viruses in seawater. Suttle, C.A., Chen, F. (1992). Appl. Environ. Microbiol. 58:3721-3729. Loss rates and loss processes for viruses in coastal seawater from the Gulf of Mexico were estimated using three different marine bacteriophages. Decay rates in the absence of sunlight ranged from 0.009 to 0.028 h^-1, with different viruses decaying at different rates. In part, decay was attributed to adsorption by heat-labile particles, as viruses did not decay or decayed very slowly in 0.2-&micro;m-filtered, autoclaved or ultracentrifuged seawater, but continued to decay in cyanide-treated seawater. Cyanide did cause decay rates to decrease, however, indicating that biological processes were also involved. As decay rates were often greatly reduced in 0.8- or 1.0 &micro;m-filtered seawater, whereas bacteria numbers were not, it suggested that most bacteria were not responsible for the decay. Decay rates were also reduced in 3-&micro;m-filtered or cycloheximide-treated seawater, but not in 8-&micro;m-filtered water, implying that flagellates consumed viruses. Viruses added to flagellate cultures decayed at 0.15 h^-1, corresponding to 3.3 viruses ingested flagellate^-1 h^-1. Infectivity was very sensitive to solar radiation and in full sunlight decay rates were 0.4-0.8 ^-1. Even when UV-B was blocked, rates were as high as 0.17 h^-1. Calculations suggest that in clear oceanic waters exposed to full sunlight that most of the viral decay, averaged over a depth of 200 m, would be attributable to solar radiation. In coastal waters, when decay rates were averaged over 24 h and 10 m depth, loss rates of infectivity attributable to sunlight were similar to those resulting from all other processes combined. Consequently, there should be a strong diel signal in the concentration of infectious viruses. In addition, as sunlight destroys infectivity but does not remove virus particles, a large proportion of the viruses in seawater are probably not infective. [TOP OF PAGE]

311. Distribution of viruses in the Chesapeake Bay. Wommack, K.E., Hill, R.T., Kessel, M., Russek-Cohen, E., Colwell, R.R. (1992). Appl. Environ. Microbiol. 58:2965-2970. High virus counts were found in water samples collected from the Chesapeake Bay. Viruses were enumerated by ultracentrifugation of water samples onto grids which were visualized by transmission electron microscopy. Virus counts in September 1990, April 1991, August 1991, and October 1991 ranged between 2.6 x 10⁶ and 1.4 x 10⁸ viruses ml^-1 with a mean of 2.5 x 10⁷ viruses ml^-1. Virus counts were usually at least three times higher than direct bacterial counts in corresponding samples. Virus counts in August and October were significantly higher than at the other sampling times, whereas bacterial counts were significantly lower at that time, yielding mean virus-to-bacterium ratios of 12.6 and 25.6, respectively. From analysis of morphology of the virus particles, it is concluded that a large proportion of the viruses are bacteriophages. The high virus counts obtained in this study suggest that viruses may be an important factor affecting bacterial populations in the Chesapeake Bay, with implications for gene transfer in natural aquatic bacterial popluations and release of genetically engineered microorganisms to estuarine and coastal environments. [TOP OF PAGE]

312. Replication of viruses in a growing plaque: A reaction-diffusion model. Yin, J., McCaskill, J.S. (1992). Biophys. J. 61:1540-1549. An understanding of viral replication process commonly referred to as "plaque growth" is developed in the context of a reaction-diffusion model. the interactions among three components: the virus, the healthy host, and the infected host are represented using rates of viral adsorption and desorption to the cell surface, replication and release by host lysis, and difdusion. The solution to the full model reveals a maximum in the dependency of the velocity of viral propagation on its equilibrium adsorption constant, suggesting that conditions can be chosen where viruses which adsorb poorly to their hosts will replicate faster in plaques than those which adsorb well. ¶ Analytic expressions for the propagation velocity as a function of the kinetic and diffusion parameters are presented for the limiting cases of equilibrated adsorption, slow adsorption, fast adsorption, and large virus yields. Hindered diffusion at high host concentrations must be included for quantitative agreement with experimental data. [TOP OF PAGE]

313. Selection for benevolence in a host-parasite system. Bull, J.J., Molineux, I.J., Rice, W.R. (1991). Evolution 45:875-882. A paradigm for the evolution of cooperation between parasites and their hosts argues that the mode of parasite transmission is critical to the long-term maintenance of cooperation. Cooperation is not expected to be maintained whenever the chief mode of transmission is horizontal: a parasite's progeny infect hosts unrelated to their parent's host. Cooperation is expected to be maintained if the chief mode of transmission is vertical: a parasite's progeny infect only the parent's host or descendants of that host. This paradigm was tested using bacteria and filamentous bacteriophage (fl). When cells harboring different variants of these phage were cultured so that no infectious spread was allowed, ensuring that all parasite transmission was vertical, selection favored the variants that were most benevolent to the host - those that least harmed host growth rate. By changing the culture conditions so that horizontal spread of the phage was allowed, the selective advantage of the benevolent forms was lost. These experiments thus support the theoretical argument that mode of transmission is a major determinant in the evolution of cooperation between a parasite and its host. [TOP OF PAGE]

314. Distinguishing mechanisms for the evolution of co-operation. Bull, J.J., Rice, W.R. (1991). J. Theor. Biol. 149:63-74. [TOP OF PAGE]

315. Plasmid-encoded bacteriophage insensitivity in members of the genus Lactococcus with special reference to pCI829. Coffey, A., Costello, V., Fitzgerald, G., Daly, C. (1991). pp. 131-135. In In Dunny, G., Cleary, P., and McKay, L. (eds.), Genetics and Molecular Biology of Streptococci, Lactococci and Enterococci. ASM, Washington, DC. [TOP OF PAGE]

316. Wide-spread occurrence and clonal variation in viruses which cause lysis of a cosmopolitan, eukaryotic marine phytoplankter Micromonas pusilla. Cottrell, M.T., Suttle, C.A. (1991). Mar. Ecol. Prog. Ser. 78:1-9. Seven clonal isolates of viruses which cause lysis of the eukaryotic, naked, photosynthetic flagellate Micromonas pusilla were isolated from the coastal waters of New York, Texas, California and British Columbia, as well as the oligotrophic waters of the central Gulf of Mexico. The viruses are large polyhedrons (ca 115 nm dia.) lacking tails, and are morphologically similar to a previously described virus (MPV) which infected M. pusilla . Restriction fragment analysis of the DNA from these clones using EcoRI revealed unique banding patterns, demonstrating that each of the clones (including 3 that were isolated from the same water sample) were genetically different. Summation of the 17 to 26 visible fragments from the restriction digests, for each of the clones, yielded estimated genomes sizes of 77 to 110 kilobase pairs. In contrast, only 4 different types of viruses could be recognized based on the molecular weights of the major proteins. In field samples the concentrations of viruses causing lysis of M. pusilla were found to be spatially and temporally variable, ranging from &lt; 20 to 4.6 x 10 super(6) infective units/l. [TOP OF PAGE]

317. HOST RANGES OF FLORIDA ISOLATES OF BACTERIOPHAGES OF ERWINIA-CAROTOVORA. Eayre, C.G., Concelmo, D.E., Bartz, J.A. (1991). Phytopathology 81:1179 [TOP OF PAGE]

318. Use of modified diatomaceous earth for removal and recovery of viruses in water. Farrah, S.R., Preston, D.R., Toranzos, G.A., GIRARD, M., ERDOS, G.A., VASUHDIVAN, V. (1991). Appl. Environ. Microbiol. 57:2502-2506. Use of modified diatomaceous earth for removal and recovery of viruses in water.Diatomaceous earth was modified by in situ precipitation of metallic hydroxides. Modification decreased the negative charge on the diatomaceous earth and increased its ability to adsorb viruses in water. Electrostatic interactions were more important than hydrophobic interactions in virus adsorption to modified diatomaceous earth. Filters containing diatomaceous earth modified by in situ precipitation of a combination of ferric chloride and aluminum chloride adsorbed greater than 80% of enteroviruses (poliovirus 1, echovirus 5, and coxsachievirus B5) and coliphage MS2 present in tap water at ambient pH (7.8 to 8.3), even after filtration of 100 liters of tap water. Viruses adsorbed to the filters could be recovered by mixing the modified diatomaceous earth with 3% beef extract plus 1 M NaCl (pH 9). [TOP OF PAGE]

319. COLIPHAGES AS ALTERNATE INDICATORS OF FECAL CONTAMINATION IN TROPICAL WATERS. Hernandez-Delgado, E.A., SERRA, M.L., Toranzos, G.A. (1991). Environmental Toxicology and Water Quality 6:131-144. Coliphages as alternate indicators of fecal contamination in tropical waters.Strong evidence has recently been found against the use of the fecal coliforms as indicator organisms of fecal contamination in tropical waters due to their indigenous nature in pristine waters. There is a great need for the development of more rapid, accurate, and low-cost techniques for determining bacteriological water quality. Coliphages seem to be an excellent alternative. We have developed a method to analyze large volumes of pristine water, which we compared to a commonly used direct method. It involves the filtration of water through positively charge filters and a virus-elution step. The eluent is then mixed with culture medium, a host bacterium, and incubated. We sampled pristine tropical rivers, water collected from bromeliads (epiphytic vegetation), sewage-contaminated waters, and marine waters. Concentrations of indicator bacteria were higher than recommended levels for recreational waters, including bromeliad waters. Indicators levels were higher in bromeliad waters than in sewage-contaminated rivers. Phages were isolated only from waters being used for recreational purposes and from sites known to be contaminated with domestic sewage, but not from pristine or bromeliad waters. These results suggest that there is a correlation between the presence of coliphages and fecal contamination. This further suggests that coliphages may be reliable indicators of fecal contamination in the environment. [TOP OF PAGE]

320. Concentration of viruses and dissolved DNA from aquatic environments by vortex flow filtration. Paul, J.H., Jiang, S.C., Rose, J.B. (1991). Appl. Environ. Microbiol. 57:2197-2204. Vortex flow filtration (VFF) was used to concentrate viruses and dissolved DNA from freshwater and seawater samples taken in Florida, the Gulf of Mexico, and the Bahamas Bank. Recoveries of T2 phage and calf thymus DNA added to artifical seawater and concentrated by VFF were 72.8 and 80%, respectively. Virus concentrations determined by transmission electron microscopy of VFF-concentrated samples ranged from 3.4 .times. 107/ml for a eutrophic Tampa Bay sample to 2.4 .times. 105 for an oligotrophic oceanic surface sample from the southeastern Gulf of Mexico. Viruslike particles were also observed in a sample taken from a depth of 1,500 m in the subtropical North Atlantic Ocean. Filtration of samples through Nucleopore or Durapore filters (pore size, 0.2 .mu.m) prior to VFF reduced phage counts by an average of two-thirds. Measurements of dissolved-DNA content by Hoechst 33258 fluorescence in environmental samples concentrated by VFF yielded values only ca. 35% of those obtained for samples concentrated by ethanol precipitation (the standard dissolved-DNA method). However, ethanol precipitation of VFF-concentrated extracts resulted in an increase in measurable DNA, reaching 80% of the value obtained by the standard method. These results indicate that a portion of the naturally occurring dissolved DNA is in a form inaccessible to nucleases and Hoechst strain, perhaps bound to protein or other polymeric material, and is released upon ethanol precipitation. Viral DNA contents estimated from viral counts averaged only 3.7% (range, 0.9 to 12.3%) of the total dissolved DNA for samples from freshwater, estuarine, and offshore oligotrophic environments. These results suggest that viruses are not a major component of the dissolved DNA, although they may be involved in its production by lysis of bacterial and phytoplankton cells. [TOP OF PAGE]

321. Genetic significance of viruses in the marine environment. Paul, J.H., Jiang, S.C., Rose, J.B. (1991). Int. Marine Biotechnology Conf. (IMBC '91), Baltimore, MD (USA), 13-16 Oct 1991; PROGRAM AND ABSTRACTS. SECOND INTERNATIONAL MARINE BIOTECHNOLOGY CONFERENCE. Viruses are now known to be abundant in aquatic environments, where they are believed to play a role in controlling bacterial and algal populations by infection and lysis. It is not known if viruses are a significant component of the dissolved (< 0.2 mu m) DNA pool. We have developed a method to concentrate both viruses and dissolved DNA from aquatic environments by vortex flow filtration (VFF). Recoveries of T2 phage and calf thymus DNA added to artificial seawater averaged 72.8 and 80%, respectively. Viral abundance in samples concentrated by VFF and determined by TEM ranged from < 10 super(6) for a sample taken at a 1500 m depth from the Gulf of Mexico to 3.4 x 10 super(7)/ml for a Tampa Bay sample. [TOP OF PAGE]

322. Roles of viral infection in organic particle flux. Proctor, L.M., Fuhrman, J.A. (1991). Mar. Ecol. Prog. Ser. 69:133-142. Lack of information on the fate of particulate-associated microorganisms prompted this investigation of viruses (including bacteriophage or phage) and phage-infected cells in sinking particles from sediment traps. Sediment trap material from 30 to 400 m collected from the north Pacific Ocean during the 'VERTEX' cruises in 1980 to 1982 was examined by transmission electron microscopy. Viruses were present in all of the sinking particles examined except for those from one sample, of highly degraded algal cells or small fecal pellets, from 400 m. Viruses in the sinking particles often appeared aggregated. From 0.7 to 3.7% of the bacteria in sinking particles contained mature phage; from these data and limited information from pure cultures, we estimate that 2 to 37% of the particulate-associated bacteria may be killed by viral lysis. Many eukaryotic cells were also apparently infected with viruses, but none (.ltoreq. 50 cells observed) of the cyanobacteria or 'Chlorella-like' cells appeared infected. Viral lysis of bacteria associated with sinking particles and free-living bacteria may be causally linked and may play a role in dissolved organic carbon production and the dynamics of sinking particles. Viral lysis may have major implications for understanding cycling of material and energy in the ocean. [TOP OF PAGE]

323. Use of ultrafiltration to isolate viruses from seawater which are pathogens to marine phytoplankton. Suttle, C.A., Chan, A.M., Cottrell, M.T. (1991). Appl. Environ. Microbiol. 57:721-726. Viruses may be major structuring elements of phytoplankton communities, and hence important regulators of nutrient and energy flux in aquatic environments. In order to ascertain if viruses are potentially important in dictating phytoplankton community structure, it is essential to determine the extent to which representative phytoplankton taxa are susceptible to viral infection. We used a spiral ultrafiltration cartridge (30,000 MW cutoff) to concentrate viruses from seawater at efficiencies approaching 100 %. Natural virus communities were concentrated from stations in the Gulf of Mexico, a barrier island pass and a hypersaline lagoon (Laguna Madre), and added to cultures of potential phytoplankton hosts. By following changes in in-vivo fluorescence over time it was possible to isolate several viruses that were pathogens to a variety of marine phytoplankton, including a prasinophyte (Micromonas pusilla), a pennate diatom (likely Navicula sp.), a centric diatom (of unknown taxa), and a chroococcoid cyanobacterium (Synechococcus sp.). As well, we observed changes in fluorescence in cultures of a cryptophyte (Rhodomonas sp.) and a chlorophyte (Nannochloropsis oculata) which were consistent with the presence of viral pathogens. Although pathogens were isolated from all stations, all the pathogens were not isolated from every station. Filterability studies on the viruses infecting Micromonas and Navicula showed that the viruses were consistently infective after filtration through polycarbonate and glass-fiber filters, but were affected by most other filter types. Establishment of phytoplankton/pathogen systems will be important in elucidating the affect that viruses have on primary producers in aquatic systems. [TOP OF PAGE]

324. Virus ecology - reply. Suttle, C.A., Chan, A.M., Cottrell, M.T. (1991). Nature 351:612-613. [TOP OF PAGE]

325. CURRENT AND POSSIBLE ALTERNATE INDICATORS OF FECAL CONTAMINATION IN TROPICAL WATERS A SHORT REVIEW. Toranzos, G.A. (1991). Environmental Toxicology and Water Quality 6:121-130. Current and possible alternate indicators of fecal contamination in tropical waters: A short review.Bacteria that belong to the total and fecal coliform groups have been used for several decades as indicators of fecal pollution of waters and other systems. The prevailing idea behind this is that these two groups of bacteria cannot replicate outside the gastrointestinal tract of warm-blooded animals. Several studies conducted by this and other laboratories have indicated that these bacteria may be naturally present in certain areas of the world. Naturally occurring fecal coliforms (the genera Escherichia and Klebsiella) have been isolated from pristine sites and from sites where there was no apparent fecal contamination. All these results lead us to the conclusion that these groups of bacteria are not adequate indicators of fecal contamination in tropical areas of the world. Alternate indicators are being tested. This laboratory and others around the world are testing the adequacy of bacteriophages (more specifically, coliphages) for this purpose. In Puerto Rico coliphages have been detected only in areas known to be contaminated with domestic sewage, but not in pristine areas, although high levels of fecal coliforms can be detected at the latter sites. All these results suggest that coliphages may be used as indicators that more realistically demonstrate fecal contamination of waters in tropical areas of the world. In addition, newer techniques, although still in the trial stages, can be used to directly detect pathogens in the environment. Of all the techniques currently available, the polymerase chain reaction is the most promising for the direct detection of pathogens and therefore for the unequivocal evaluation of the biological quality of waters. [TOP OF PAGE]

326. A quantifiable phenotype of viral propagation. Yin, J. (1991). Biochim. Biophys. Acta 174:1009-1014. A system has been identified where a virus, replicating continuously on its host, displays a distinct and quantifiable phenotype, and thereby continuously reports on the state of the virus-host relationship. When bacteriophage T7 is plated out with its host, Escherichia coli, it establishes a constant-velocity infection wave, which is driven by an autocatalytic reaction- diffusion mechanism. The velocity—which is easily measured—continuously reflects the infection environment. The simplicity of the system extends to the investigator an unprecedented ability to monitor and control a viral infection process. [TOP OF PAGE]

327. The Ecology of Bacteriophage T4. Abedon, S.T. (1990). University of Arizona. In this dissertation I explore the ecology of bacteriophage T4, a virus of Escherichia coli. In particular, I argue that the life history of bacteriophage T4 can be divided into the growth and survival of T4 phages in three distinct environments. I argue that these environments are distinguished by at least two T4 phage sensory systems. These include (i) the sensing of secondary adsorption by infecting phages and (ii) the determination of the concentration of monovalent cations and free tryptophan in solution about free T4 phage particles. The first environment consists of high concentrations of uninfected, logarithmic phase E. coli cells. These concentrations are approximately 10⁶ E. coli cells/ml and greater. This environment occurs in the prefecal colonic lumen of animals. Here T4 phages exhibit unimpeded logarithmic growth. The second environment contains high concentrations of infected E. coli cells, low concentrations of uninfected E. coli cells, and high concentrations of free T4 phage particles. This second environment also occurs in the prefecal colonic lumen of animals and represents the maturation of environments supporting logarithmic T4 phage population growth. Such phage phenotypes as secondary exclusion and lysis inhibition characterize T4 phage growth in this environment. The third environment consists of extra-colonic waters. Here T4 phages avoid infecting E. coli cells and exhibit strategies that maximize their stability. These strategies in extra-colonic waters increase the potential of T4 phages to disseminate successfully from colon to colon. I employ this enhanced understanding of T4 phage ecology, outlined above, in an exploration of the ecology of the repair of DNA damage by T4 phages. [TOP OF PAGE]

328. Selection for lysis inhibition in bacteriophage. Abedon, S.T. (1990). J. Theor. Biol. 146:501-511. For Escherichia coli cells that have been infected by T-even bacteriophages (phages T2, T4, and T6), the adsorption of a second T-even phage results in an increase in the length of the original phage infection and an associated increase in the number of phages produced by the same infected cell. This is a phage encoded response called lysis inhibition. In this study the ecological significance of lysis inhibition is explored. In particular it is argued that lysis inhibition is an adaptive response to environments containing high concentrations of infected cells and low concentrations of uninfected cells. [TOP OF PAGE]

329. CONTROL OF SOFT ROT ERWINIAS WITH BACTERIOPHAGES. Eayre, C.G., Concelmo, D.E., Bartz, J.A. (1990). Phytopathology 80:994 [TOP OF PAGE]

330. Viral mortality of marine bacteria and cyanobacteria. Proctor, L.M., Fuhrman, J.A. (1990). Nature 343:60-62. [TOP OF PAGE]

331. Infection of phytoplankton by viruses and reduction of primary productivity. Suttle, C.A., Chan, A.M., Cottrell, M.T. (1990). Nature 347:467-469. Natural marine waters contain roughly 10⁶ to 10⁹ virus particles per ml, yet their role in aquatic ecosystems and the organisms that they infect remain largely unknown. Electron microscopy has been used to study interactions between viruses and their hosts, focusing mainly on pathogens to prokaryotic organisms. The authors demonstrate that viral pathogens infect a variety of important marine primary producers, including diatoms, cryptophytes, prasinophytes and chroococcoid cyanobacteria. Also, addition to sea water of particles in the 0.002-0.2 &micro;m size range, concentrated from sea water by ultrafiltration, reduced primary productivity (¹⁴C-bicarbonate incorporation) by as much as 78%. Results indicate that in addition to grazing and nutrient limitation, infection by viruses could be a factor regulating phytoplankton community structure and primary productivity in the oceans. [TOP OF PAGE]

332. Selection for bacteriophage latent period length by bacterial density: A theoretical examination. Abedon, S.T. (1989). Microb. Ecol. 18:79-88. In bacteriophage (phage), rapid and efficient intracellular progeny production is of obvious benefit. A short latent period is not. All else being equal, a longer latent period utilizes host cell resources more completely. Using established parameters of phage growth, a simulation of three successive phage lysis cycles is presented. I have found that high, but not low, host cell densities can select for short latent periods. This results from phage with short latent periods more rapidly establishing multiple parallel infections at high host cell concentrations, whereas phage with long latent periods are restricted to growth within a single cell over the same period. This implies that phage with short latent periods habitually grow in environments that are rich in host cells. [TOP OF PAGE]

333. Isolation and characterization of a Pseudomonas aeruginosa bacteriophage with a very limited host range. Bigby, D., Kropinski, A.M.B. (1989). Can. J. Micribiol. 35:630-635. [TOP OF PAGE]

334. Identification and characterization of a plasmid encoding phage abortive infection from Lactococcus lactis ssp. lactis UC811. Coffey, A., Fitzgerald, G.F., Daly, C. (1989). Netherlands Milk and Dairy Journal 43:229-244. Identification and characterization of a plasmid encoding abortive infection from Lactococcus lactis ssp. lactis UC811.Bacteriophage infection of starter cultures used in milk fermentations is a major problem in commercial practise. A highlight of recent genetic studies has been the demonstration of a linkage between phage insensitivity mechanisms and plasmid DNA. The conjugative plasmid pCI1829 encodes abortive infection in Lactococcus lactis ssp. lactis. This was manifested by complete insensitivity (no plaques) to small isometric-headed phage and a large reduction in plaque size for prolate-headed phage. The introduction of a derivative of a similar plasmid, pCI1750, into a strain containing pCI1829 resulted in complete insensitivity against a prolate-headed phage showing that these plasmids have an additive effect against phage. The abortive infection system described here was not inhibited by temperatures as high as 40.degree. C. [TOP OF PAGE]

335. The effect of IncP-2 plasmids on propagation of Pseudomonas aeruginosa bacteriophages. Freizon, E.V., Kopylova, Yu.I., Cheremukhina, L.V., Krylov, V.N. (1989). Genetika 25:1168-1178. [TOP OF PAGE]

336. The Bacteriophages. Volume 2. Ackermann, H.-W., DuBow, M.S. (1988). Plenum Press, New York.[TOP OF PAGE]

337. The Bacteriophages. Volume 1. Ackermann, H.-W., DuBow, M.S. (1988). Plenum, New York.[TOP OF PAGE]

338. Study of transducing properties of therapeutic and prophylactic dysentery bacteriophages. Chanishvili, N. (1988). Materials of the Y Congress of the Georgian Society of Genestists and Selectioners, Tbilisi 1988:34-36. [TOP OF PAGE]

339. Improvement of the method of transduction applicable for a new transducing dysenteric phage BFl-1. Chanishvili, N. (1988). Materials of the Y Congress of the Georgian Society of Genestists and Selectioners, Tbilisi 1988:33-34. [TOP OF PAGE]

340. Marine bacteriophages and bacterial mortality. Proctor, L.M., Fuhrman, J.A., Ledbetter, M.C. (1988). EOS 69:1111-1112. [TOP OF PAGE]

341. Enteric Viruses and Coliphages in Latin America. Toranzos, G.A., Gerba, C.P., Hanssen, H. (1988). Toxicity Assessment 3:491-510. [TOP OF PAGE]

342. A study of 33 bacteriophages of Rhizobium meliloti. Werquin, M., Ackermann, H.-W., Levesque, R.C. (1988). Appl. Environ. Microbiol. 54:188-196. [TOP OF PAGE]

343. Practical applications of bacteriophages. Ackermann, H.-W., DuBow, M.S. (1987). pp. 143-172. In AnonymousViruses of Prokaryotes. Volume I. General properties of bacteriophages. CRC Press, Boca Raton, FL. [TOP OF PAGE]

344. Origin and evolution of bacteriophages. Ackermann, H.-W., DuBow, M.S. (1987). pp. 173-192. In AnonymousViruses of Prokaryotes. Volume I. General properties of bacteriophages. CRC Press, Boca Raton, FL. [TOP OF PAGE]

345. Viruses of Prokaryotes, Volume 1, General Properties of Bacteriophages. Ackermann, H.-W., DuBow, M.S. (1987). CRC Press, Boca Raton, Florida.[TOP OF PAGE]

346. Introduction: General properties of bacteriophages. Ackermann, H.-W., DuBow, M.S. (1987). pp. 1-11. In AnonymousViruses of Prokaryotes. Volume I: General Properties of Bacteriophages. CRC Press, Inc., Boca Raton, Florida. [TOP OF PAGE]

347. Viruses of Prokaryotes, Volume 2, Natural Groups of Bacteriophages. Ackermann, H.-W., DuBow, M.S. (1987). CRC Press, Boca Raton, Florida.[TOP OF PAGE]

348. Description and identification of new phages. Ackermann, H.-W., DuBow, M.S. (1987). pp. 103-142. In AnonymousViruses of Prokaryotes. Volume I. General properties of bacteriophages. CRC Press, Boca Raton, FL. [TOP OF PAGE]

349. Lysogeny. Ackermann, H.-W., DuBow, M.S. (1987). pp. 87-101. In AnonymousViruses of Prokaryotes. Volume I: General Properties of Bacteriophages. CRC Press, Inc., Boca Raton, Florida. [TOP OF PAGE]

350. Bacteriophage Taxonomy. Ackermann, H.-W., DuBow, M.S. (1987). pp. 13-28. In In Ackermann, H.-W. and DuBow, M.S. (eds.), Viruses of Prokaryotes. Volume I. General properties of bacteriophages. CRC Press, Boca Raton, FL. Taxonomy, the science of classification, is as old as science itself. In essence, it compares and summarizes facts in order to draw conclusions: it may be viewed as an abstract activity aimed at interpretation, understanding, and simplification. In itself, taxonomy is completely independent from nomenclature. Perhaps the best example of the aims and consequences of taxonomy is the Mendeleyev system of elements; once established, it conveyed information through its very arrangement and predicted the properties of elements not yet discovered. Virus data are accumulating so fast that virus research would long ago have come to a standstill without repeated efforts toward generalization which is visible in the growing number of review papers. Some familiarity with virus taxonomy, which is essentially comparative virology, appears to be necessary for teachers, students, and all but the most specialized investigators. With respect to phages, the purpose of classification is (1) to organize approximately 2800 known phages into a managable system for teaching purposes and a better understanding of phage biology; (2) to facilitate phage identification, notably in research on phage ecology and industrial microbiology; and (3) to allow predictions and control of experimental results; e.g., any DNA molecular weight that differs notably from that of simlar phages may be incorrect. [TOP OF PAGE]

351. Occurrence and frequency of bacteriophages. Ackermann, H.-W., DuBow, M.S. (1987). pp. 29-47. In AnonymousViruses of Prokaryotes. Volume I. General properties of bacteriophages. CRC Press, Boca Raton, FL. [TOP OF PAGE]

352. Phage multiplication. Ackermann, H.-W., DuBow, M.S. (1987). pp. 49-85. In AnonymousViruses of Prokaryotes. Volume I: General Properties of Bacteriophages. CRC Press, Inc., Boca Raton, Florida. [TOP OF PAGE]

353. COMPARATIVE STUDIES OF BACTERIOPHAGES SPECIFIC FOR BREVIBACTERIUM-FLAVUM. ARUTYUNYAN, S.Z., KAZHOYAN, S., V, KARABEKOV, B.P., REULETS, M.A., Khrenova, E.A., AKHVERDYAN, V.Z., Krylov, V.N. (1987). Biotekhnologiya 21-27. Comparative studies of bacteriophages specific for Brevibacterium flavum.A comparative study of some biological and physico-chemical characteristics of 10 bacteriophages active on Brevibacterium flavum cells was carried out. All bacteriophages belong to the morphological group 4 (Tikhonenko's classification) with phage particles having similar structure and size (the head diameter 60 .+-. 1 nm, the tail length 310 .+-. 8 nm). Molecular sizes of phage DNAs are 44-47 kb. There are no identical isolates among all of the 10 phages studied according to results of DNA restriction with endonucleases. It was suggested that all 10 phages belong to the same group of related phages. Nevertheless according to levels of manifested DNA/DNA homology the phages were distributed into 3 subgroups: subgroup .vphi. B (phages .vphi. B, BB1, BB4, BB8, BB10, BB12 and BB14); subgroup E.PHI.. (phages E.PHI.-Y and E.PHI.-B), subgroup H.PHI. (with the only representative). Genomes of phages belong to the same subgroup have more than 90% DNA homology. The level of DNA homology measured between phages belonging to subgroup E.PHI. and .vphi. B varies (for different pairs of phages) from practically 0 to 5% and for subgroups E.PHI. and H.PHI. is near 1-2%. There is no DNA homology between phages of subgroup .vphi. B and phage H.PHI. [TOP OF PAGE]

354. A STUDY OF PHAGE RESISTANT MUTANTS OF BREVIBACTERIUM-FLAVUM. ARUTYUNYAN, S.Z., KAZHOYAN, S., V, CHITCHYAN, M.B., CHAKHMAKHCHYAN, A.G., OGANEZOVA, G.G., KARABEKOV, B.P., AKHVERDYAN, V.Z., Krylov, V.N. (1987). Biotekhnologiya 9-13. A study of phage resistant mutants of Brevibacterium flavum.Independent phage resistant and glutacin resistant mutants were isolated using three subgroups of Brevibacterium bacteriophages and glutacin PBP-26 (a bacteriocin produced by the strain Corynebacterium glutamicum PBP-26). According to their resistance to these phages and glutacin in the mutants may be distributed into 10 groups. For these phages and glutacin a formal receptor scheme of B. flavum XX was elaborated. At least two independent mutations were shown necessary for mutants resistant to all phages and glutacin to appear. [TOP OF PAGE]

355. Mutagenic and lethal effects of near-ultraviolet radiation (290-400 nm) on bacteria and phage. Eisenstark, A. (1987). Environ. Molec. Mutagen. 10:317-337. [TOP OF PAGE]

356. Potential for transduction of plasmids in a natural freshwater environment: Effect of plasmid donor concentration and a natural microbial community on transduction in Pseudomonas aeruginosa. Saye, D.J., Ogunseitan, O., Sayler, G.S., Miller, R.V. (1987). Appl. Environ. Microbiol. 53:987-995. [TOP OF PAGE]

357. PARTIAL CHARACTERIZATION OF BACTERIOPHAGES FOR CLAVIBACTER-MICHIGANENSE-SSP-NEBRASKENSE. SHIRAKO, Y., Vidaver, A.K., Ackermann, H.-W. (1987). Annals of the Phytopathological Society of Japan 52:793-800. Partial characterization of bacteriophages for Clavibacter michiganense ssp. nebraskense.Bacteriophages Clmll, ClmX and ClmXC for Clavibacter (Cl.) michiganense subsp. nebraskense were characterized with respect to biological and physical properties. All phages produced clear plaques on strain CN18-5, the original propagating host. On strain CN76-2, ClmX produced predominantly turbid plaques with a few clear plaques, from which ClmXC was derived. The host range of the three phages was restricted to strains of Cl. michiganense subsp. nebraskense. Adsorption rate constants and burst sizes of the three phages were in the range of 2.9 to 9.4 .times. 10-9 ml/min and 6 to 19 virions/cell, respectively. All phages were morphologically similar with a hexagonal head about 60 nm in width and a flexuous tail approximately 235 nm long. The type of nucleic acid was double-stranded DNA. Agarose gel electrophoresis of restriction endonuclease digests of the phage DNAs indicated that the three phages are similar, but distinguishable in DNA structure. ClmXC was probably a deletion mutant of ClmX. [TOP OF PAGE]

358. Inactiviation of phage by near-ultraviolet radiation and hydrogen peroxide. Eisenstark, A., Buzard, R.L., Hartman, S. (1986). Photochem. Photobiol. 44:603-606. [TOP OF PAGE]

359. EXPRESSION OF PSEUDOMONAS-AERUGINOSA TRANSPOSABLE BACTERIOPHAGES IN THE CELLS OF PSEUDOMONAS-PUTIDA I. ESTABLISHMENT OF THE LYSOGENIC STATE AND EFFECTIVENESS OF LYTIC GROWTH. GORBUNOVA, S.A., YANENKO, A.S., AKHVERDYAN, V.Z., REULETS, M.A., Krylov, V.N. (1986). Genetika 21:1455-1463. Expression of Pseudomonas aeruginosa transposable bacteriophages in the cells of Pseudomonas putida: I. Establishment of the lysogenic state and effectiveness of lytic growth.Expression of transposable phages (TP) of Pseudomonas aeruginosa in the cells of P. putida was studied. The high efficiency of phage lytic development was shown both as a consequence of zygotic induction after transfer of the RP4::TPc+ plasmid into nonlysogenic recipients, and as a result of heat induction of lysogens PpG1(D3112cts15). The high phage yield (20-25 particles of D3112cts phage per one cell of P. putida) is an evidence for a high level of transposition in the cells of this bacterial species. Plasmids RP4::TP are transferred into cells of PpG1 and PAO1 with similar frequency. However, the efficiency of establishment of the lysogenic state is lower in PpG1. Transposable phages of P. aeruginosa can integrate into the chromosome of PpG1 producing stable inducible lysogens. The presence of RP4 in the P. putida cells is not necessary for expression of transposable phages. The transposable phage D3112cts15 can be used in experiments of interspecies transduction of plasmids and chromosomal genes. [TOP OF PAGE]

360. Morphology of Yersinia enterocolitica phages. Kasatiya, S., Ackermann, H.-W. (1986). Annales de l Institut Pasteur - Virology 137E:59-69. [TOP OF PAGE]

361. Aeromonas bacteriophages: Reexamination and classification. Ackermann, H.-W., Dauguet, C., Paterson, W.D., Popoff, M., Rouf, M.A., Vieu, J.-F. (1985). Ann. Inst. Pasteur Virol. 136E:175-??? [TOP OF PAGE]

362. New actinophage species. Ackermann, H.-W., Berthiaume, L., Jones, L.A. (1985). Intervirology 23:121-??? [TOP OF PAGE]

363. Les virus des bactéries. Ackermann, H.-W. (1985). p. 196-??? In Maurin, J. (ed.), Virologie Médicale. Flammarion Médecine-Sciences, Paris. [TOP OF PAGE]

364. SIMULTANEOUS LOSS OF BACTERIOPHAGE RECEPTOR AND COAGGREGATION MEDIATOR ACTIVITIES IN ACTINOMYCES-VISCOSUS MG-1. Tylenda, C.A., ENRIQUEZ, E., Kolenbrander, P.E., Delisle, A.L. (1985). Infect. Immun. 48:228-233. Simultaneous loss of bacteriophage receptor and coaggregation mediator activities in Actinomyces viscosus MG-1.Actinomyces bacteriophages were used as tools to study coaggregation between actinomyces and streptococci. Four bacteriophage isolates, phages AV-1, AV-2, AV-3,and 1281, bound to coaggregation group A A. viscosus and to group E A. naeslundii. No binding to groups B, C, D, or F was observed. Only A. viscosus MG-1 was capable of supporting a productive infection by these phages. Spontaneously occurring bacteriophage-resistant mutants of A. viscosus MG-1 were isolated and were shown to fall into 2 classes. Class I mutants were resistant to all 4 phages, whereas class II mutants were resistant only to phage AV-3. In each case, strains resistant to a particular phage were unable to bind that phage, suggesting that a loss or alteration of the cell surface phage receptor had occurred. Both classes of mutants were unable to coaggregate with streptococci representing coaggregation group 1 and had also lost the ability to mediate 1 type of coaggregation with group 4 streptococci. Class II mutants also were unable to coaggregate with group 2 streptococci. Lactose-inhibitable interactions with other streptococci (groups 3 and 4) were unchanged in the mutants. The simultaneous loss of sensitivity to phage AV-3 and the ability to coaggregate with coaggregation group1 streptococci suggests the possibility of a relationship between these 2 cell surface structures. [TOP OF PAGE]

365. New species definitions in phages of gram-positive cocci. Ackermann, H.-W., Cantor, E.D., Jarvis, A.W., Lembke, J., Mayo, J.A. (1984). Interviriology 22:181-190. [TOP OF PAGE]

366. Classification of Vibrio bacteriophages. Ackermann, H.-W., Kasatiya, S.S., Kawata, T., Koga, T., Lee, J.V., Mbiguino, A., Newman, F.S., Vieu, J.-F., Zachary, A. (1984). Intervirology 22:61-71. [TOP OF PAGE]

367. THE GROUP OF GENETICALLY RELATED PHAGES OF CORYNEBACTERIUM-GLUTAMICUM. ARUTYUNYAN, S.Z., AKHVERDYAN, V.Z., Khrenova, E.A., KARABEKOV, B.P., Krylov, V.N. (1984). Genetika 20:730-737. The group of genetically related phages of Corynebacterium glutamicum.The bacteriophages MC-1, MC-2, MC-3 and MC-4 specific for C. glutamicum were studied. The phages represent a group of genetically related phages with common physical, chemical and biological properties. All these phages belong to the morphological group IV, according to Tikhonenko's classification. Phage genomes consist of double-stranded DNA with a MW of .apprx. 37.3 megadaltons. DNA have single-stranded cohesive ends. [TOP OF PAGE]

368. ISOLATION AND MORPHOLOGY OF BACTERIO PHAGES OF KURTHIA-ZOPFII. Rocourt, J., Ackermann, H.-W., Brault, J. (1984). Annales de Virologie (Paris) 134:557-568. Isolation and morphology of bacteriophages of Kurthia zopfii.Seven bacteriophages from K. zopfit were isolated from meat samples. The phages were morphologically identical, belonged to the pedoviridae family and resembled Bacillus phage GA-1. [TOP OF PAGE]

369. Sewage coliphages studied by electron microscopy. Ackermann, H.-W., Nguyen, T.M. (1983). Appl. Environ. Microbiol. 45:1049-1059. Sewage was enriched with 35 Escherichia coli strains, and sediments of enrichment cultures were studied in the electron microscope. They contained up to 10 varieties of morphologically different particles. T-even-type phages predominated in 14 samples. Thirteen phages were enriched, representing the families Myoviridae (seven), Styloviridae (two), Podoviridae (three), and Microviridae (one). Twelve of these corresponded to known enterobacterial phage species, namely, 121, K19, FC3-9, O1, 9266, T2, 16-19, kappa, beta 4, N4, T7, and phi X174. Cubic RNA phages and filamentous phages were not detected. Types 121 and 9266 have previously been observed only in Romania and South Africa. Identification by morphology is usually simple. Our investigative technique is qualitative and will not detect all phages present. Most enrichment strains are polyvalent, and electron microscopy is always required for phage identification. In a general way, electron microscopy seems to be the method of choice for investigation of phage geography and ecology. [TOP OF PAGE]

370. Current problems in bacterial virus taxonomy. Ackermann, H.-W. (1983). pp. 105-121. In In Matthews, R.E.F. (ed.), A Critical Appraisal of Viral Taxonomy. CRC Press, Boca Raton, FL. [TOP OF PAGE]

371. Construction of the systems for the study of transducing ability of therapeutic and prophylactic dysenteric phages. Chanishvili, N., Ilina, T. (1983). pp. 64-78. In AnonymousBacterio-phages - Theoretical and Practical Issues. Moscow. [TOP OF PAGE]

372. Bacteriophage in the Ixodes dammini spirochete, etiological agent of Lyme disease. Hayes, S.F., Burgdorfer, W., Barbour, A.G. (1983). J. Bacteriol. 154:1436-1439. [TOP OF PAGE]

373. Bacteriophage T4. Mathews, C.K., Kutter, E.M., Mosig, G., Berget, P.B. (1983). American Society for Microbiology, Washington, DC.[TOP OF PAGE]

374. Long tail fibers: Genes, proteins, assembly, and structure. Wood, W.B., Crowther, R.A. (1983). pp. 259-269. In In Mathews, C.K., Kutter, E.M., Mosig, G., and Berget, P.B. (eds.), Bacteriophage T4. American Society for Microbiology, Washington, DC. "Factors that promote retraction (of long tail fibers) and therefore lower infectivity are low pH, low temperature, low ionic strength, and in some strains absence of tryptophan (Conley and Wood, 1975; Kellenberber et al., 1965). These effects probably are biologically significant in that they inhibit infection under environmental conditions that are unfavorable for phage multiplication." [p. 259]. [TOP OF PAGE]

375. Félix d'Herelle: His life and work and the foundation of a bacteriophage reference center. Ackermann, H.-W., Martin, M., Vieu, J.-F., Nicolle, P. (1982). ASM News 48:346-??? [TOP OF PAGE]

376. Alteration of bacteriophage attachment capacity by near-UV irradiation. Hartman, P.S., Eisenstark, A. (1982). J. Virol. 43:529-532. [TOP OF PAGE]

377. Comparative biology and evolution of bacteriophages. Reanney, D.C., Ackermann, H.-W. (1982). Adv. Virus Res. 27:205-280. [TOP OF PAGE]

378. A survey of Brucella phages and morphology of new isolates. Ackermann, H.-W., Simon, F., Verger, J.-M. (1981). Intervirology 16:1-7. [TOP OF PAGE]

379. PSEUDOMONAS-AERUGINOSA BACTERIO PHAGES HAVING DNA STRUCTURE SIMILAR TO THAT OF MU-1 PHAGE 3. ISOLATION AND ANALYSIS OF HYBRID PHAGE D-3112 AND PHAGE B-39 LOCALIZATION OF THE IMMUNITY REGION AND SOME GENETIC FACTORS. BOGUSH, V.G., Plotnikova, T.G., KIRSANOV, N.B., Rebentish, B.A., PERMOGOROV, V., I, Krylov, V.N. (1981). Genetika 17:967-976. Pseudomonas aeruginosa bacteriophages having DNA structure similar to that of Mu1 phage: 3. Isolation and analysis of hybrid phage D3112 and phage B39: Localization of the immunity region and some genetic factors.Several different hybrids were isolated in a cross between heteroimmune Mu-like P. aeruginosa bacteriophages D3112 and B39. Analysis of hybrid phage DNA treated with specific endonucleases and heteroduplex studies permitted mapping of the immunity region of the phages to the left part of the genome. A ts temperature-sensitive mutation in the wild-type B39 and loci influencing the growth of D3112 in P. aeruginosa strain PA0(B39) and in bacteria harboring plasmid RPL11 were localized. All recombinants studied arose by double crossing-over and can be considered as substitutions of different continuous parts of the D3112 genome by fragments of the B39 genome. A comparison of distribution of non-homologous regions in B39 and D3112 genomes as well as locations of endonuclease-sensitive sites gave some unexpected results. In contrast to the B39-D3112 pair, an other pair of related Mu-like phages, B3-D3112, have no visible homology within most of their genomes. [TOP OF PAGE]

380. Alteration of bacteriophage attachment capacity by near-UV irradiation. Hartman, P.S., Eisenstark, A. (1981). J. Virol. 43:529-532. [TOP OF PAGE]

381. ISOLATION AND CHARACTERIZATION OF PSEUDOMONAS-PUTIDA PHAGE RESISTANT MUTANTS BY USE OF NEW BACTERIO PHAGES. Krylov, V.N., Kulakov, L.A., KIRSANOV, N.B., Khrenova, E.A. (1981). Genetika 17:239-245. Isolation and characterization of Pseudomonas putida phage-resistant mutants by use of new bacteriophages.Three new bacteriophages of P. putida PpG1 were isolated. The bacteriophages differed in many characteristics and were unrelated. Phage-resistant mutants of P. putida PpG1 were selected using these phages and bacteriophage pf16. No lysogenic variants were detected among these mutants. The mutants were divided into 7 groups according to their resistance to bacteriophages. Resistance to phage PMW was caused by at least 2 different mutations. Some of the phage-resistant mutations appear to be incompatible with survival. [TOP OF PAGE]

382. Tailed phages of Pseudomonas and related bacteria. Liss, A., Ackermann, H.-W., Mayer, L.W., Zierdt, C.H. (1981). Intervirology 15:71-81. [TOP OF PAGE]

383. An updated survey of Bacillus phages. Reanney, D.C., Ackermann, H.-W. (1981). Intervirology 15:190-197. [TOP OF PAGE]

384. PSEUDOMONAS-AERUGINOSA PHAGES WHOSE DNA IS SIMILAR IN STRUCTURE TO MU-1 PHAGE DNA 2. EVIDENCE OF RELATIONSHIP OF D-3112 B-3 AND B-39 PHAGES ANALYSIS OF DNA SPLITTING BY RESTRICTION ENDO NUCLEASES ISOLATION OF A RECOMBINANT BETWEEN D-3112 AND B-3 PHAGES. Krylov, V.N., BOGUSH, V.G., YANENKO, A.S., KIRSANOV, N.B. (1980). Genetika 16:975-984. Pseudomonas aeruginosa phages whose DNA is similar in structure to mu1 phage DNA: 2. Evidence of relationship of D3112, B3 and B39 phages: Analysis of DNA splitting by restriction endonucleases, isolation of a recombinant between D3112 and B3 phages.It is found that bacteriophages B3 and B39 specific for P. aeruginosa have the same genome structure as previously described phage D3112. On the right (S) end of their genomes, a variable non-phage DNA is located (approximately 0.9-2.5 kilobases for different phages). This variable DNA probably has its origin from different regions of bacterial chromosome. In genome of 1 of the phages, B3' phage, such variable DNA (not more than 150 base pairs) was found on the left end of DNA molecule. Isolation of a viable B3 .times. D3112 recombinant phage and analysis of its genome with restriction technique and with studies of homo- and heteroduplex molecules confirmed the genetical relationship of B3 and D3112. Some essential non-homology of B3 and D3112 DNA were found on the right ends of genomes of the phages. [TOP OF PAGE]

385. Host Specificity of Glycine max genotypes with antibiotic-resistant mutants and phage-typed strains of Rhizobium japonicum. Schröder, E.C. (1980). North Carolina State University. Rhizobium japonicum Bacteriophages: Isolation and Host Range. Bacteriophages able to lyse several different Rhizobium japonicum strains were isolated from soils of North Carolina where soybeans had been grown previously. For isolation, an enrichment technique was followed. Soil-broth mixtures were incubated, centrifuged and the liquid supernatant filtered. This bacterial-free supernatant was spotted on different strains. When lysis occurred, the supernatant was diluted and single plaques were purified by several passages. For some strains, several phages differing in plaque size and morphology were obtained. Most R. japonicum bacteriophages produce larger plaques than Rhizobium trifolii or Rhizobium meliloti phages; this is probably associated with the longer generation time of the host cells. ¶ A set of different rhizobiophages was used to study their host range on a large number of different Rhizobium strains. The study shows that some phages have a very specific host range while others are able to lyse a wide range of R. japonicum strains. ¶ Strains were classified according to their phage lysis pattern in 25 different groups. With one exception, strains belonging to a certain phage group belong also to the same serogroup. However, strains in the same serogroup were subdivided into different phage groups. Results indicate there is a close relation between serotype and phage receptor sites. The use of a set of phages is suggested to help in the classification of Rhizobium japonicum. Electron microscopy pictures show that phage Rh110/1 has a head and long, contractile tails and falls into the general morphological group A. Bacteriophage Typing of Antibiotic-resistant mutants of Rhizobium japonicum. Wild type strains of Rhizobium japonicum are naturally resistant to higher concentrations of antibiotics than fast growing rhizobia. Antibiotic marked strains of Rhizobium are commonly used for ecological studies. R. japonicum mutants resistant to streptomycin, kanamycin, erythromycin, and rifampicin were isolated from a range of strains belonging to different serogroups. Mutant clones were characterized by their rhizobiophage sensitivity pattern and their ability to nodulate soybeans and reduce acetylene. Several antibiotic-resistant mutants showed a lysotype pattern different from the wild type or were unable to nodulate Lee 74 soybeans. Presumptive R. trifolii mutants were shown to be contaminated or cultures of R. japonicum USDA110. Genetic marked strains provide a useful tool, but should be carefully checked before being used for ecological studies. Influence of Glycine max Genotypes on strain selection by a mixed
     Rhizobium japonicum inoculum. Thirty-seven soybean genotypes of different origins were inoculated with a mixture of three strains of Rhizobium japonicum. Each strain could be differentiated serologically, differed in antibiotic resistance and was lysed by specific rhizobiophages. Plants were grown in the greenhouse in vermiculite for 5 weeks and then the root nodules were harvested. The strains were identified by their differential markers. The percentage of nodules produced by each strain was calculated. Strain USDA58Ery was more dominant than USDA110SN and both were more competitive than USDA123Kan. Three soybean genotypes of SE Asia origin (TGm51, PI323.275 and PI376.844) were not nodulated by strain USDA123Kan. This result can be attributed to host plant genes conferring resistance to the specific bacterial strain or suppression of nodulation by other strains. The distribution of strains in each plant genotype was compared to that of Lee 74 by chi-square analysis. There were significant differences among genotypes in their ability to select a specific strain from the mixed inoculum. Most genotypes had nodules containing more than one Rhizobium strain: Lee (Non-nod) had the highest percentage. Spontaneous resistant mutants resistant to kanamycin were less competitive than Rhizobium strains resistant to streptomycin, rifampicin, or erythromycin. It is suggested that one appropriate combination of host genotype and R. japonicum strain can be used to establish the desired strain and obtain inoculation responses in soils with an indigenous population of root-nodule bacteria. [TOP OF PAGE]

386. Bacteriophage-like particles associated with a spirochete. Berthiaume, L., Elazhary, Y., Alain, R., Ackermann, H.-W. (1979). Can. J. Microbiol. 25:114-??? [TOP OF PAGE]

387. DNA BASE COMPOSITION NATURE OF INTRA CELLULAR DNA MORPHOLOGY AND CLASSIFICATION OF BACTERIO PHAGES INFECTING MICROCOCCUS-LUTEUS. COMPTON, S.W., Mayo, J.A., Ehrlich, M., Ackermann, H.W., TREMBLAY, L., CORDS, C.E., SCALETTI, J., V (1979). Canadian Journal of Microbiology 25:1027-1035. DNA base composition, nature of intracellular DNA, morphology and classification of bacteriophages infecting Micrococcus luteus.Ten bacteriophages infecting M. luteus were characterized. All phages contain double-stranded DNA, of 64.3-73.5 mol% guanine plus cytosine (GC). The DNA of phage N7 has the highest GC content reported for any bacterial virus. No unusual bases were found. The intracellular replicating DNA of 6 phages are covalently closed circular molecules. All 10 phages have isometric, probably icosahedral, heads and long, flexible, noncontractile tails and can be sorted into 2 morphological groups based on size and presence or absence of a collar. Host-range studies indicate 6 host-range groups. [TOP OF PAGE]

388. Characterization of two Salmonella newport bacteriophages. Moazamie, N., Ackermann, H.-W., Murthy, M.R.V. (1979). Canadian Journal of Microbiology 25:1063-1072. [TOP OF PAGE]

389. La classification des phages d'Agrobacterium et Rhizobium. Ackermann, H.-W. (1978). Pathol. Biol. 26:507-??? [TOP OF PAGE]

390. Guidelines for bacteriophage characterization. Ackermann, H.-W., Audurier, A., Berthiaume, L., Jones, L.A., Mayo, J.A., Vidaver, A.K. (1978). Adv. Virus Res. 23:1-24. [TOP OF PAGE]

391. Phage-like bacteriocins. Ackermann, H.-W., Brochu, G. (1978). p. 691-??? In Laskin, A.I. and Lechevaliar, H.A. (eds.), CRC Handbook of Microbiology. CRC Press, Boca Raton, Florida. [TOP OF PAGE]

392. Natural groups of bacteriophages. Ackermann, H.-W. (1978). p. 639-??? In Laskin, A.I. and Lechevaliar, H.A. (eds.), CRC Handbook of Microbiology. CRC Press, Boca Raton, FL. [TOP OF PAGE]

393. Partial characterization of a cubic Bacillus phage. Ackermann, H.-W., Roy, R., Murthy, M.R.V., Smirnoff, W.A. (1978). Can. J. Micribiol. 24:986-??? [TOP OF PAGE]

394. Evolutionary aspects of bacteriophage development. Eisenstark, A. (1978). p. 616-??? AnonymousProceedings of the Fourth International Congress on Virology. Centre for Agricultural Publishing and Documentation, Wageningen, The Netherlands. [TOP OF PAGE]

395. Susceptibility of lipopolysaccharide-defective mutants of Pseudomonas aeruginosa strain PAO to dyes, detergents, and antibiotics. Kropinski, A.M., Chan, L., Milazzo, F.H. (1978). Antimicrobial Agents and Chemotherapy 13:494-499. Lipopolysaccharide-defective mutants of Pseudomonas aeruginosa strain PAO have been isolated on the basis of their resistance to lipopolysaccharide-specific bacteriophages. These mutants have been differentiated by their agglutination in NaCl and acriflavine, phage sensitivity, and chemical analysis of the lipopolysaccharides. The susceptibility of the wild-type strain and four mutants to a series of twenty-six agents, including dyes, detergents, antibiotics, and lysozyme, was examined. The roughest mutant (AK-43) exhibited increased susceptibility to sodium deoxycholate, hexadecylpyridinium chloride, benzalkonium chloride, ampicillin, penicillin G, erythromycin, colymycin, and polymyxin B. The role of cell envelope fractions in antibiotic resistance in P. aeruginosa is discussed. [TOP OF PAGE]

396. La classification des actinophages. Berthiaume, L., Ackermann, H.-W. (1977). Pathol. Biol. 25:195-??? [TOP OF PAGE]

397. Interactions of bacterial viruses and bacterial genes with animal systems. Merril, C.R. (1977). pp. 83-96. In In Rubenstein, I., Phillips, R.L., Green, C.E., and Desnick, R.J. (eds.), Molecular Genetic Modification of Eucaryotes. [TOP OF PAGE]

398. La classification des phages caudés des entérobactéries. Ackermann, H.-W. (1976). Pathol. Biol. X 24:359-??? [TOP OF PAGE]

399. Properties of some new Brucella phage isolates; evidence for lysogeny within the genus. Corbel, M.J., Thomas, E.L. (1976). Developm 31:38-45. A number of phages were isolated from Brucella abortus strains showing atypical phage sensitivity patterns. The phages, designated the Firenze group, were all similar in morphology, host range general properties. They were lytic for smooth Brucella abortus, Br. neotomae and Br. suis, but not for Br. melitensis strains. They were serologically distinct from phages of the Tbilisi and M51-S708-Weybridge groups and also differed from these in plaque morphology and physical and chemical stability. The parent brucella strains although smooth were resistant to lysis by their own phages or by the other available brucella phages. Their general properties suggested a lysogenic state. [TOP OF PAGE]

400. [New lambdoid Escherichia coli phages. I. Isolation, group immunity and recombination with lambda phage]. Krylov, V.N., Tsygankov, I. (1976). Genetika 12:104-111. 550 bacterial strains were isolated from sewage. 69 of them were lysogenic by phages active on Escherichia coli. The phages were divided into two groups on the basis of UV-inducibility, the ability to form plaques on Rep-E. coli mutants and particle morphology: lambdoid (23 phages) and related to P2 (46 phages). Hybrid phages isolated from the crosses of lambdoid phages with phage lambda harboured the region imm lambda and the gene of adsorption specificity from other parent. Ten groups of heteroimmune phages were found in the collection of new temperate phages are homoimmune with known phages: lambda, phi 80, phi 81, 434. Another 7 phages involved in 6 groups of immunity which were heteroimmune to known phages. Diversity of lambdoid phage genes determining the structure of repressor is discussed. [TOP OF PAGE]

401. La classification des bacteriophages des cocci gram-positifs: Micrococcus, Staphylococcus et Streptococcus. Ackermann, H.-W. (1975). Path. -Biol. 23:247-253. [TOP OF PAGE]

402. Characterization of Escherichia coli bacterial viruses in commercial sera. Geier, M.R., Attallah, A.F., Merril, C.R. (1975). In Vitro 11:55-58. [TOP OF PAGE]

403. Structure of two phages of Bacillus thuringiensis and B. cereus. Ackermann, H.-W., Smirnoff, W.A., Bilsky, A.Z. (1974). Can. J. Micribiol. 20:29-??? [TOP OF PAGE]

404. The present state of phage taxonomy. Ackermann, H.-W., Eisenstark, A. (1974). Interviriology 3:201-219. [TOP OF PAGE]

405. La classification des bactériophage de Bacillus et Clostridium. Ackermann, H.-W. (1974). Pathol. Biol. 22:909 [TOP OF PAGE]

406. Bacteriophage interactions with higher organisms. Merril, C.R. (1974). Trans. N. Y. Acad. Sci. 36:265-272. [TOP OF PAGE]

407. A phage typing scheme for Salmonella newport. Petrow, S., Kasatiya, S.S., Pelletier, J., Ackermann, H.-W., Peloquin, J. (1974). Ann. Mikcrobiol. 125A:433-445. [TOP OF PAGE]

408. Studies of immunological reactivity following syngeneic or allogeneic marrow grafts in man. Fass, L., Ochs, H.D., Thomas, E.D., Mickelson, E., Storb, R., Fefer, A. (1973). TRANSPLANTATION 16:630-640. [TOP OF PAGE]

409. The fate of bacteriophage lambda in non-immune germ-free mice. Geier, M.R., Trigg, M.E., Merril, C.R. (1973). Nature 246:221-223. [this is cited in Merril et al., 1996 (PNAS 93:3188-3192): ". . . even in the absence of an antibody response, bacteriophage tend to be rapidly eliminated from the circulation by the reticuloendothelium system (ref)"]. [TOP OF PAGE]

410. Phages defectifs chez Chromobacterium. Ackermann, H.-W., Gauvreau, L. (1972). Zentrabl. Bakteriol. Parasitenkd. Infektionskr. Hyg. Abt. 1 Orig. Reihe A 221:196-??? [TOP OF PAGE]

411. Characteristics of three phages infectious for psychrophilic fishery isolates of Pseudomonas putrefaciens. Delisle, A.L., Levin, R.E. (1972). Antonie van Leeuwenhoek J. Microbiol. 38:1-8. [TOP OF PAGE]

412. Bacteriophages of psychrophilic pseudomonads. II. Host range of phage active against Pseudomonas putrefaciens. Delisle, A.L., Levin, R.E. (1972). Antonie van Leeuwenhoek J. Microbiol. 38:1-8. [TOP OF PAGE]

413. Effect of temperature on an obligately psychrophilic phage-host system of Pseudomonas putrefaciens. Delisle, A.L., Levin, R.E. (1972). Antonie van Leeuwenhoek J. Microbiol. 38:9-15. [TOP OF PAGE]

414. lambda phage transcription in human fibroblasts. Geier, M.R., Merril, C.R. (1972). Virology 47:638-643. [TOP OF PAGE]

415. ??? Merril, C.R., Friedman, T.B., Attallah, A., Geier, M.R., Krell, K., Yarkin, R. (1972). In Vitro 8:91-93. [this is cited in Lederberg, 1996 (PNAS 93:3167-3168): "Phages are commonly found in bovine sera used for cell culture (ref)"]. [TOP OF PAGE]

416. Bacterial virus gene expression in human cells. Merril, C.R., Geier, M.R., Petricciani, J.C. (1971). Nature 233:398-??? [TOP OF PAGE]

417. Bactériophages--propriétes et premières étapes d'une classification. Ackermann, H.-W. (1969). Path. -Biol. 17:1003-1024. [TOP OF PAGE]

418. Bacteriophages of psychophilic pseudomonads. I. Host range of phage pools active against fish spoilage and fish-pathogenic pseudomonads. Delisle, A.L., Levin, R.E. (1969). Antonie van Leeuwenhoek J. Microbiol. 35:307-317. [TOP OF PAGE]

419. Detection of Salmonella carriers and Vi-phage typing of the isolates. Ha, T.-Y. (1969). Journal of the Korean Medical Association 12:836-??? [TOP OF PAGE]

420. Biological effects of substituting cytosine for 5-hydroxymethylcytosine in the deoxyribonucleic acid of bacteriophage T4. Kutter, E.M., Wiberg, J.S. (1969). J. Virol. 4:439-453. [TOP OF PAGE]

421. Histological changes of various organs of mouse by the introduction of staphylococcal phages. Ha, T.-Y. (1968). Chonnam Medical Journal 4:151-??? [TOP OF PAGE]

422. Studies on protection of Klebsiella pneumonia -infected mouse with phage. Ha, T.-Y. (1968). Journal of Korean Modern Medicine 8:395-??? [TOP OF PAGE]

423. Isolation of viral particles from large fluid volumes. Anderson, N.G., Cline, G.B., Harris, W.W., Green, J.G. (1967). pp. 75-88. In In Ber, G. (ed.), Transmission of Viruses by the Water Route. Interscience Publishers, [TOP OF PAGE]

424. Bacteriophage techniques. Eisenstark, A. (1967). Methods in Virology 1:449-524. [TOP OF PAGE]

425. Geographical distribution of Vi-phage types of Salmonella typhi isolated in East Asia. A review made mainly on a basis of results in Korea. Aoki, Y., Chun, D., Suh, J.S., Ha, T.-Y., Lee, J.S. (1965). Endemic Dis. Bulletin of Nagasaki Univ. (Japan) 7:71 [TOP OF PAGE]

426. Epidemiology of typhoid fever with special reference to phage typing of Salmonella. Ha, T.-Y. (1965). Korean Army Medical Journal 9:139-??? [TOP OF PAGE]

427. Studies on the movement of staphyloccocal phage introduced into rabbit body by various routes. Ha, T.-Y. (1964). Korean Central Journal 7:595-??? [TOP OF PAGE]

428. Phage isolated from lysogenic Bacillus anthracis. Buck, C.A., Anacker, R.L., Newman, F.S., Eisenstark, A. (1963). J. Bacteriol. 85:1423-1430. [TOP OF PAGE]

429. The influence of sulfadiazine on the intracellular multiplication of dysentery phage. Ha, T.-Y. (1961). Chonnam University. [TOP OF PAGE]

430. Erfahrungen über die Isolierung von Typhus-Vi-und Salmonellaphagen aus Fluss- und Abwasser. Lenk, V., Ackermann, H.-W. (1961). Zbl. Bakteriol. Hyg. Parasitenkd. Infektionskr. Abt. 1 Orig. Reihe A 181:224-??? [TOP OF PAGE]

431. Lysogenicity in Xanthomonas pruni. Eisenstark, A., Goldberg, S.S., Bernstein, L.B. (1955). Journal of General Microbiology 12:402-405. [TOP OF PAGE]

432. Specificity of bacteriophages of Xanthomonas pruni. Eisenstark, A., Bernstein, L.B. (1955). Phytopathology 45:596-598. [TOP OF PAGE]

433. Plaque formation by xanthomonas pruni bacteriophage. Eisenstark, A., Thornberry, H.H. (1950). Phytopathology XL:876-877. [TOP OF PAGE]

434. Bacterial viruses and hosts: Influence of culturable state. Robb, F.T., Hill, R.T. (???). p. ???-??? In Colwell, R.R. and Grimes, D.J. (eds.), Nonculturable Microorganisms in the Environment. ASM Press, Washington,D.C. [TOP OF PAGE]

435. Estimating viral proliferation in aquatic samples. Steward, G.F. (???). p. ???-??? In Paul, J.H. (ed.), Marine Microbiology. Academic Press, London. It is only within the last decade that marine viruses were determined to be consistently the most abundant biological entities in the sea (Fuhrman, 1999). Since then, many advances have been made in understanding viral ecology (Fuhrman, 1999, Wilhelm and Suttle, 1999). Initial discoveries showed that viruses are both abundant in the ocean and that many bacteria are infected with viruses (Bergh et al., 1989; Proctor and Fuhrman, 1990). These data led researchers to believe that viruses are an important source of mortality in marine microbial food webs, but only provided a static picture. Subsequent studies have shown that virus populations are extremely dynamic, and can change quickly over short time scales (Bratbak et al 1990, 1996). Estimates of viral production and decay rates provided the valuable confirmation that viruses are active members of the marine community (Heldal and Bratbak, 1991; Steward et al., 1992b). The production of viruses implies the lysis of host cells and the release of cellular material as dissolved and colloidal organic carbon. Therefore, measurements of viral replication rates are also useful for assessing the contribution of viruses to bacterial mortality and organic matter cycling in the ocean. By assuming a burst size, viral productivity can be used to estimate rates of bacterial lysis. This approach provides an additional means to assess bacterial mortality along with the original electron microscopy-based method of Proctor and Fuhrman (1990). Accurate measurements of viral productivity and turnover are required if we are to properly model their dynamics and impact within the microbial food web. So far, however, there is no standard method for measuring viral productivity. A wide variety of different approaches have been used each with associated advantages and disadvantages. These methods include:
     1. Quantifying net increases in viral abundance over time (Bratbak et al., 1990)
     2. Measuring rates of viral decay (Heldal and Bratbak, 1991)
     3. Estimating viral DNA synthesis rates by radiolabeling (Steward et al., 1992a,b)
     4. Calculating expected viral release rates from estimated rates of bacterial lysis and an assumed burst size (Weinbauer et al., 1993)
     5. Measuring tracer dilution rates using fluorescently labeled viruses (FLV) as tracers (Noble and Fuhrman, submitted)
     The first approach, observing net increases in viruses over time, is the simplest means of demonstrating viral proliferation. However, use of this method is restricted to times when viral abundance is increasing and only provides a minimum estimate of productivity unless the viral decay rate is also known. The productivity estimates obtained are also dramatically influenced by the time scale of sampling (Bratbak et al., 1994, 1996). In the second approach, virus production is prevented by poisoning or removing host organisms and the rate at which viruses disappear (or decay) is observed. If the system was initially in steady state, the decay rate is assumed equal to the original rate of production. These two approaches have been used in a number of studies and variations on them are possible. Due to space limitations, however, this chapter focuses only on the last three methods listed above. [TOP OF PAGE]

436. Fingerprinting viral assemblages by pulsed field gel electrophoresis. Steward, G.F. (???). p. ???-??? In Paul, J.H. (ed.), Marine Microbiology. Academic Press, London. Viruses are the most abundant microorganisms in marine and freshwater environments and perhaps the most genetically diverse (Fuhrman and Suttle, 1993). Counting viruses in aquatic samples is now a routine matter, but assessing the diversity and dynamics within complex assemblages is still a challenge. DNA-based fingerprinting approaches which rely on amplification of rRNA gene fragments by PCR have facilitated analyses of bacterial community composition. These approaches have more restricted application when analyzing viral assemblages, because of the extreme genetic diversity among viruses. Unlike in bacteria, there are no gene sequences conserved in all viruses which can serve as universal primer sites for PCR amplification. PCR-based analyses of viral assemblages must therefore target specific subsets of the total viral assemblage. For example, PCR amplification of specific genes has recently been used to examine the genetic diversity among cyanophages (Fuller et al., 1998) and among phycodnaviridae (Chen et al., 1996; Short and Suttle, 1999). A more general fingerprinting approach, which encompasses the total viral assemblage, is a valuable complement to these more specific, higher resolution analyses. The approach described here uses variation in genome size as the basis for obtaining a fingerprint of a viral assemblage (Klieve and Swain, 1993). A whole genome fingerprinting approach is possible, because viral genomes can vary greatly in length (a few thousand to hundreds of thousands of base pairs) yet they fall within a range that is easily resolved using pulsed field gel electrophoresis (PFGE). The PFGE fingerprinting technique provides a quick and relatively simple means of visualizing differences in the composition of viral assemblages (Swain et al., 1996; Wommack et al., 1999a; Steward and Azam, 2000). As a supplement to the more specific treatment of PFGE provided in this chapter, the reader is encouraged to consult the excellent introductory text to PFGE by Birren and Lai (1993). [TOP OF PAGE]

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