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PubMed1999

Similarly organized lysogeny modules in temperate Siphoviridae from low GC content gram-positive bacteria.

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)/cI-like repressor/cro-like repressor/antirepressor (optional)] differ clearly from that of lambda-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 gram-negative bacteria, but also in insect viruses.

Amino Acid Sequence↗

PubMed1998

The structural gene module in Streptococcus thermophilus bacteriophage phi Sfi11 shows a hierarchy of relatedness to Siphoviridae from a wide range of bacterial hosts.

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

Biological Evolution↗

PubMed2001

Comparative phage genomics and the evolution of Siphoviridae: insights from dairy phages.

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.

Dairying↗

PubMed2002

The dilemma of phage taxonomy illustrated by comparative genomics of Sfi21-like Siphoviridae in lactic acid bacteria.

The complete genome sequences of two dairy phages, Streptococcus thermophilus phage 7201 and Lactobacillus casei phage A2, are reported. Comparative genomics reveals that both phages are members of the recently proposed Sfi21-like genus of Siphoviridae, a widely distributed phage type in low-GC-content gram-positive bacteria. Graded relatedness, the hallmark of evolving biological systems, was observed when different Sfi21-like phages were compared. Across the structural module, the graded relatedness was represented by a high level of DNA sequence similarity or protein sequence similarity, or a shared gene map in the absence of sequence relatedness. This varying range of relatedness was found within Sfi21-like phages from a single species as demonstrated by the different prophages harbored by Lactococcus lactis strain IL1403. A systematic dot plot analysis with 11 complete L. lactis phage genome sequences revealed a clear separation of all temperate phages from two classes of virulent phages. The temperate lactococcal phages share DNA sequence homology in a patchwise fashion over the nonstructural gene cluster. With respect to structural genes, four DNA homology groups could be defined within temperate L. lactis phages. Closely related structural modules for all four DNA homology groups were detected in phages from Streptococcus or Listeria, suggesting that they represent distinct evolutionary lineages that have not uniquely evolved in L. lactis. It seems reasonable to base phage taxonomy on data from comparative genomics. However, the peculiar modular nature of phage evolution creates ambiguities in the definition of phage taxa by comparative genomics. For example, depending on the module on which the classification is based, temperate lactococcal phages can be classified as a single phage species, as four distinct phage species, or as two if not three different phage genera. We propose to base phage taxonomy on comparative genomics of a single structural gene module (head or tail genes). This partially phylogeny-based taxonomical system still mirrors some aspects of the current International Committee on Taxonomy in Virology classification system. In this system the currently sequenced lactococcal phages would be grouped into five genera: c2-, sk1, Sfi11-, r1t-, and Sfi21-like phages.

Base Sequence↗

PubMed2000

Comparative genomics of the late gene cluster from Lactobacillus phages.

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 Sfi11/Lactococcus lactis phage TP901-1 on one hand and Lactobacillus delbrueckii phage LL-H/Lactobacillus plantarum phage phig1e/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 Gram-positive bacteria is defined by temperate cos-site phages including Lactobacillus gasseri 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.

Amino Acid Sequence↗

PubMed2001

Comparative genomics of lactococcal phages: insight from the complete genome sequence of Lactococcus lactis phage BK5-T.

Lactococcus lactis phage BK5-T and Streptococcus thermophilus phage Sfi21, two cos-site temperate Siphoviridae with 40-kb genomes, share an identical genome organization, sequence similarity at the amino acid level over about half of their genomes, and nucleotide sequence identity of 60% over the DNA packaging and head morphogenesis modules. Siphoviridae with similarly organized genomes and substantial protein sequence similarity were identified in several genera of low-GC-content Gram-positive bacteria. These phages demonstrated a gradient of relatedness ranging from nucleotide sequence similarity to protein sequence similarity to gene map similarity over the DNA packaging and head morphogenesis modules. Interestingly, the degree of relatedness was correlated with the evolutionary distance separating their bacterial hosts. These observations suggest elements of vertical evolution in phages. The structural genes from BK5-T shared no sequence relationships with corresponding genes/proteins from lactococcal phages belonging to distinct lactococcal phage species, including phage sk1 (phage species 936) that showed a closely related gene map. Despite a clearly distinct genome organization, lactococcal phages sk1 and c2 showed nine sequence-related proteins. Over the early gene cluster phage BK5-T shared nine regions of high nucleotide sequence similarity, covering at most two adjacent genes, with lactococcal phage r1t (phage species P335). Over the structural genes, the closest relatives of phage r1t were not lactococcal phages belonging to other phage species, but Siphoviridae from Mycobacteria (high-GC-content Gram-positive bacteria). Evidence for recent horizontal gene transfer between distinct phage species was obtained for dairy phages, but these transfers were limited to phages infecting the same bacterial host species.

Computational Biology↗

PubMed2006

Anatomy of a lactococcal phage tail.

Bacteriophages of the Siphoviridae family utilize a long noncontractile tail to recognize, adsorb to, and inject DNA into their bacterial host. The tail anatomy of the archetypal Siphoviridae lambda has been well studied, in contrast to phages infecting gram-positive bacteria. This report outlines a detailed anatomical description of a typical member of the Siphoviridae infecting a gram-positive bacterium. The tail superstructure of the lactococcal phage Tuc2009 was investigated using N-terminal protein sequencing, Western blotting, and immunogold transmission electron microscopy, allowing a tangible path to be followed from gene sequence through encoded protein to specific architectural structures on the Tuc2009 virion. This phage displays a striking parity with lambda with respect to tail structure, which reenforced a model proposed for Tuc2009 tail architecture. Furthermore, comparisons with lambda and other lactococcal phages allowed the specification of a number of genetic submodules likely to encode specific tail structures.

Electrophoresis, Polyacrylamide Gel↗

PubMed1988

A Study of 33 Bacteriophages of Rhizobium meliloti.

A total of 33 Rhizobium meliloti bacteriophages were studied. Of those, 21 were isolated in northern France from field soil in which Medicago sativa L. was grown. The other 12 phages were obtained by UV light and mitomycin C induction from 46 R. meliloti strains. Rhizobiophages were characterized by their morphology, host range, serological properties, restriction endonuclease patterns, DNA-DNA homologies, and DNA molecular weights. Five morphotypes were observed showing tailed phages with icosahedral heads. The categories of morphotypes included the Myoviridae (11 phages), Siphoviridae (3 morphotypes and 20 phages), and Podoviridae (2 phages). Type NM1 phage (Siphoviridae) is highly unusual because of the presence of transverse bars on the phage tail. Soil phages had broad host ranges, whereas phages isolated from bacterial cultures showed more or less narrow host ranges. Restriction endonuclease patterns and DNA-DNA hybridization experiments showed that the five phage type genomes were unrelated. Molecular weights of phage type DNAs were estimated, and they corresponded to values expected for capsid sizes, except for phage NM8. Type phiM11S (Siphoviridae) did not correspond to any other described Rhizobium phages and represents a new species.

Journal Article↗

PubMed1994

Differential Effect of Tetrazolium Dyes upon Bacteriophage Plaque Assay Titers.

This study examined whether the practice of incorporating either tetrazolium red or tetrazolium violet dye into plaque assay medium deleteriously influences plaque assay titers. Representative members of six different virus families were studied: Cystoviridae (varphi6), Leviviridae (MS2), Microviridae (varphiX174), Myoviridae (T2), Podoviridae (P22), and Siphoviridae (Denver, T1, and VD13). Each of the members of the Podoviridae and Siphoviridae families appeared to be suppressed by either one or both dyes at a 300-mug/ml concentration. The chosen representatives of the other bacteriophage families were not suppressed by either dye at a 300-mug/ml concentration. Subsequent trials revealed no suppression of Podoviridae or Siphoviridae plaque assay titers when members of these virus families were tested with the same two dyes at the lower concentrations of 150 and 50 mug/ml. Interestingly, the bacteriophage families whose members were affected by the dyes have additional commonality in that they are the two bacteriophage families whose members possess both double-stranded DNA genomes and noncontractile tails.

Journal Article↗

PubMed2002

Comparative genomics of phages and prophages in lactic acid bacteria.

Comparative phage genomics has become possible due to the availability of more than 100 complete phage genome sequences and the development of powerful bioinformatics tools. This technology, profiting from classical molecular-biology knowledge, has opened avenues of research for topics, which were difficult to address in the past. Now, it is possible to retrace part of the evolutionary history of phage modules by comparative genomics. The diagnosis of relatedness is hereby not uniquely based on sequence similarity alone, but includes topological considerations of genome organization. Detailed transcription maps have allowed in silico predictions of genome organization to be verified and refined. This comparative knowledge is providing the basis for a new taxonomic classification concept for bacteriophages infecting low G + C-content Gram-positive bacteria based on the genetic organization of the structural gene module. An Sfi21-like and an Sfi11-like genus of Siphoviridae is proposed. The gene maps of many phages show remarkable synteny in their structural genes defining a lambda super-group within Siphoviridae. A hierarchy of relatedness within the lambda super-group suggests elements of vertical evolution in Siphoviridae. Tailed phages are the result of both vertical and horizontal evolution and are thus fascinating objects for the study of molecular evolution. Prophage sequences integrated into the genomes of their bacterial host present theoretical challenges for evolutionary biologists. Prophages represent up to 10% of the genome in some LAB. In pathogenic streptococci prophages confer genes of selective value for the lysogenic cell. The lysogenic conversion genes are located between the lysin gene and the right phage attachment site. Non-attributed genes were found at the same genome position of prophages from lactic streptococci. These genes belong to the few prophage genes transcribed in the lysogen. Prophages from dairy bacteria might therefore also contribute to the evolutionary fitness of non-pathogenic LAB.

Bacteriophages↗

PubMed2003

Homogeneity of the morphological groups of bacteriophages infecting Bacteroides fragilis strain HSP40 and strain RYC2056.

Bacteriophages infecting Bacteroides fragilis strains RYC2056 and HSP40 have been proposed as indicators of water quality. To accomplish this function, homogeneity of the group of phages detected by these strains is necessary to ensure that the final results are not due to the different kinetics of inactivation of the phages. To evaluate homogeneity, we observed by electron microscopy bacteriophages isolated from sewage with two Bacteroides fragilis strains (HSP40 and RYC2056). A predominant group of phages was observed, Siphoviridae with slightly curved tails. Detection of other minority groups, which could be present in the sample, was done with neutralization experiments by using antiserum against the majority group and with host mutants resistant to infection with the predominant phage. Although two other minority groups were observed, results showed that bacteriophages infecting B. fragilis strain HSP40 and strain RYC2056 form a homogeneous group, Siphoviridae with slightly curved tails being the most predominant in sewage.

Animals↗

PubMed1999

Study of the potential relationship between the morphology of infectious somatic coliphages and their persistence in the environment.

The proportions of different morphological types of infectious somatic coliphages were determined in faecally polluted freshwaters. Myoviridae, followed by Siphoviridae, were the most frequently isolated morphological types in raw sewage, treated sewage and river water collected a few metres downstream from a sewage outfall. However, in river water collected further downstream from the pollution point, in river water after 'in situ' inactivation experiments and in chlorinated raw and treated sewage significant changes in the proportions of the different somatic coliphage morphological types occurred. In all cases, Siphoviridae, especially those with flexible and curled tails, became more abundant to the detriment of Myoviridae.

Caudovirales↗

PubMed1994

Structural proteins and DNA characteristics of 14 Listeria typing bacteriophages.

The major structural proteins of 13 temperate and one virulent Listeria typing bacteriophages were analysed and compared using isoelectric focusing in immobilized pH gradients (IPG), ultrathin-layer two-dimensional electrophoresis, amino acid analysis and N-terminal amino acid sequences of selected proteins. Isoelectric points for major capsid and tail proteins of the 12 members of the siphoviridae family included in this study ranged from 4.70 to 5.92, whereas one of the two myoviridae investigated (B054) showed structural proteins in the 6.1 to 6.3 range. In comparison to protein profiles from one-dimensional SDS gels, the IPG technique gave better resolution and improved discrimination of phage proteins. Combination of this technique and SDS gel electrophoresis made it possible to correlate M(r) and isoelectric points of major structural proteins. Tail polypeptides of all siphoviridae are smaller and, with one exception, more acidic than their corresponding capsid counterparts. We also determined the amino acid composition of capsid and tail proteins. When compared with an average protein, they were found to be fairly rich in acidic and short-chain hydrophobic amino acids, as well as in lysine. In addition, the N-terminal amino acid sequences of major capsid and tail proteins of four representative listeriaphages were compared. The base composition of listeriaphage DNAs was between 37% and 39% G + C, reflecting that of their bacterial hosts. Each phage had a distinct restriction endonuclease pattern, and genome sizes ranged from 35 to 116 kb. DNA-DNA hybridization permitted the identification of five DNA homology groups. The two myoviruses studied (A511 and B054) showed no DNA homology to other phages, confirming their unique nature. The 12 siphoviruses were classified into three DNA homology groups with little cross-homology. Furthermore, phage A006 was found to share little DNA homology with the other investigated members of species 2671. Therefore, a new species (A006) is proposed. With respect to phage classification and taxonomy, a good correlation between the various approaches was observed, mostly corresponding to particle morphology.

Amino Acid Sequence↗

PubMed2002

Isolation and characterization of bacteriophages from fermenting sauerkraut.

This paper presents the first report of bacteriophage isolated from commercial vegetable fermentations. Nine phages were isolated from two 90-ton commercial sauerkraut fermentations. These phages were active against fermentation isolates and selected Leuconostoc mesenteroides and Lactobacillus plantarum strains, including a starter culture. Phages were characterized as members of the Siphoviridae and Myoviridae families. All Leuconostoc phages reported previously, primarily of dairy origin, belonged to the Siphoviridae family.

Bacteriophages↗

PubMed2025

CRISPRi-mediated repression of three cI repressors induces the expression of three related Neisseria gonorrhoeae bacteriophages.

The Neisseria gonorrhoeae FA1090 isolate encodes nine prophage islands (Ngoɸ1-9). Ngoɸ1-3 contain genes consistent with a Siphoviridae-dsDNA bacteriophage (phage). Saturating transposon-sequencing screens using two different N. gonorrhoeae isolates predicted that multiple prophage genes were essential, including three putative transcriptional repressors: ngo0479 (present in Ngoɸ1), ngo1116 (present in Ngoɸ2), and ngo1630 (present in Ngoɸ3). All three genes display homology to the Lambda phage cI, a regulator important for maintaining the lysogenic state and inhibiting lytic induction, but these proteins are not close paralogs. Using a Neisseria lactamica-derived Type I-C CRISPR-interference system, we show that these cI orthologs are essential, as the knockdown of each gene results in bacterial death. We determined that the repression of the three cI orthologs resulted in the significant induction of phage gene expression. Finally, we detected Siphoviridae-like phage particles released from N. gonorrhoeae following repression of ngo0479, ngo1116, or ngo1630. We hypothesize that these cI orthologs are critical for preventing phage lytic infection and cell death and allow N. gonorrhoeae to benefit from the carriage and expression of prophage genes.IMPORTANCEBacteriophage, or phage, are bacteria-infecting viruses and are the most abundant natural entities in the world. Here, we report that Neisseria gonorrhoeae's three most complete double-stranded DNA prophage islands each encode essential and related transcriptional repressors. CRISPRi-mediated repression of these transcriptional repressors leads to a significant increase in prophage gene expression and phage induction. This study marks an important initial step in studying the interaction between N. gonorrhoeae and its resident phage.

Neisseria gonorrhoeae↗

PubMed2005

Sequence analysis of the Lactobacillus plantarum bacteriophage PhiJL-1.

The complete genomic sequence of a Lactobacillus plantarum virulent phage PhiJL-1 was determined. The phage possesses a linear, double-stranded, DNA genome consisting of 36,677 bp with a G+C content of 39.36%. A total of 52 possible open reading frames (ORFs) were identified. According to N-terminal amino acid sequencing and bioinformatic analyses, proven or putative functions were assigned to 21 ORFs (41%), including 5 structural protein genes. The PhiJL-1 genome shows functionally related genes clustered together in a genome structure composed of modules for DNA replication, DNA packaging, head and tail morphogenesis, and lysis. This type of modular genomic organization was similar to several other phages infecting lactic acid bacteria. The structural gene maps revealed that the order of the head and tail genes is highly conserved among the genomes of several Siphoviridae phages, allowing the assignment of probable functions to certain uncharacterized ORFs from phage PhiJL-1 and other Siphoviridae phages.

Amino Acid Sequence↗

PubMed2003

The prophage sequences of Lactobacillus plantarum strain WCFS1.

The Lactobacillus plantarum commensal WCFS1 contains four prophage elements in its genome. Lp1 and Lp2 are two about 40-kb-long uninducible prophages that share closely related DNA packaging, head and tail genes defining a second lineage of pac-site Siphoviridae in L. plantarum, distinct from L. plantarum phage phig1e, but related to Bacillus phage SPP1 and Lactococcus phage TP901-1. Northern analysis revealed transcribed prophage genes exclusively near both attachment sites. Comparative genomics identified candidate lysogenic conversion genes (LCG) downstream of the lysis cassette and within the lysogeny module. Notable are genes with sequence similarities to putative LCG from Streptococcus pyogenes prophages and to a Bacillus plasmid. Both prophages harbored tRNA genes. R-Lp3 and R-Lp4 represent short prophage remnants; R-Lp3 abuts Lp2 and displays sequence links to cos-site Siphoviridae.

Base Sequence↗

PubMed2005

Partial characterization of a novel bacteriophage of Vibrio harveyi isolated from shrimp culture ponds in Thailand.

A bacteriophage was isolated together with Vibrio harveyi (VH) 1114 a from a black tiger shrimp-rearing pond in Thailand. By negative staining transmission electron microscopy (TEM), the phage had an icosahedral head (diameter 60-62 nm), a rigid, non-contractile tail (9-10 nm x 100-120 nm) without a collar or terminal fibers and a genome of double stranded DNA of approximately 80 kb as determined by analysis of restriction enzyme digestion fragments. Since these features would place it in the family Siphoviridae, it was tentatively named V. harveyi siphoviridae-like phage or VHS1. VHS1 could also infect two VH reference strains LMD 22.30 and LMD 80.33 (=ATCC 14126) but yielded smaller plaques than with VH1114. The phage tolerated temperatures as high as 60 degrees C for up to 2h and overnight exposure to a broad range of pH. VHS1 lysogens of VH1114 were unstable, contained unaltered VHS1 DNA, were immune to VHS1 lysis and spontaneously released VHS1 in liquid cultures. Approximately 20 kb of the genome has been sequenced and deposited at GenBank but it mostly showed no significant homology with existing sequences in the database.

Animals↗