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 r