Characterization of Two Virulent Phages of Lactobacillus plantarum

We characterized two Lactobacillus plantarum virulent siphophages, ATCC 8014-B1 (B1) and ATCC 8014-B2 (B2), previously isolated from corn silage and anaerobic sewage sludge, respectively. Phage B2 infected two of the eight L. plantarum strains tested, while phage B1 infected three. Phage adsorption was highly variable depending on the strain used. Phage defense systems were found in at least two L. plantarum strains, LMG9211 and WCSF1. The linear double-stranded DNA genome of the pac-type phage B1 had 38,002 bp, a G+C content of 47.6%, and 60 open reading frames (ORFs). Surprisingly, the phage B1 genome has 97% identity with that of Pediococcus damnosus phage clP1 and 77% identity with that of L. plantarum phage JL-1; these phages were isolated from sewage and cucumber fermentation, respectively. The double-stranded DNA (dsDNA) genome of the cos-type phage B2 had 80,618 bp, a G+C content of 36.9%, and 127 ORFs with similarities to those of Bacillus and Lactobacillus strains as well as phages. Some phage B2 genes were similar to ORFs from L. plantarum phage LP65 of the Myoviridae family. Additionally, 6 tRNAs were found in the phage B2 genome. Protein analysis revealed 13 (phage B1) and 9 (phage B2) structural proteins. To our knowledge, this is the first report describing such high identity between phage genomes infecting different genera of lactic acid bacteria.     

A Virulent Phage Infecting Lactococcus garvieae,with Homology to Lactococcus lactis Phages

A new virulent phage belonging to the Siphoviridae family and able to infect Lactococcus garvieae strains was isolated from compost soil. Phage GE1 has a prolate capsid (56 by 38 nm) and a long noncontractile tail (123 nm). It had a burst size of 139 and a latent period of 31 min. Its host range was limited to only two L. garvieae strains out of 73 tested. Phage GE1 has a double-stranded DNA genome of 24,847 bp containing 48 predicted open reading frames (ORFs). Putative functions could be assigned to only 14 ORFs, and significant matches in public databases were found for only 17 ORFs, indicating that GE1 is a novel phage and its genome contains several new viral genes and encodes several new viral proteins. Of these 17 ORFs, 16 were homologous to deduced proteins of virulent phages infecting the dairy bacterium Lactococcus lactis, including previously characterized prolate-headed phages. Comparative genome analysis confirmed the relatedness of L. garvieae phage GE1 to L. lactis phages c2 (22,172 bp) and Q54 (26,537 bp), although its genome organization was closer to that of phage c2. Phage GE1 did not infect any of the 58 L. lactis strains tested. This study suggests that phages infecting different lactococcal species may have a common ancestor.     

Lysogeny in Bradyrhizobium japonicum and Its Effect on Soybean Nodulation

Rhizobiophage V, isolated from soil in the vicinity of soybean roots, was strongly lytic on Bradyrhizobium japonicum 123B (USDA 123) but only mildly lytic on strain L4-4, a chemically induced small-colony mutant of 123. Numerous bacteriophage-resistant variants were isolated from L4-4 infected with phage V; two were studied in detail and shown to be lysogenic. The two, L4-4 (V5) and L4-4 (V12), are the first reported examples of temperate-phage infection in B. japonicum. Phage V and its derivative phages V5 and V12 were closely related on the basis of common sensitivity to 0.01 M sodium citrate and phage V antiserum, phage immunity tests, and apparently identical morphology when examined by electron microscopy. However, the three phages differed in host range and in virulence. Lysogens L4-4 (V5) and L4-4 (V12) were immune to infection by phages V and V5 but not to infection by V12. Southern hybridization analysis confirmed the incorporation of phage V into the genomes of strains L4-4(V5) and L4-4(V12) and also demonstrated the incorporation of phage V into the genome of a phage V-resistant derivative of USDA 123 designated 123 (V2). None of the three lysogens, L4-4(V5), L4-4(V12), or 123B(V2), was able to nodulate soybean plants. However, Southern hybridization profile data indicated that phage V had not incorporated into any of the known B. japonicum nodulation genes.     

The P-SSP7 cyanophage has a linear genome with direct terminal repeats

P-SSP7 is a T7-like phage that infects the cyanobacterium Prochlorococcus MED4. MED4 is a member of the high-light-adapted Prochlorococcus ecotypes that are abundant in the surface oceans and contribute significantly to primary production. P-SSP7 has become a model system for the investigation of T7-like phages that infect Prochlorococcus. It was classified as T7-like based on genome content and organization. However, because its genome assembled as a circular molecule, it was thought to be circularly permuted and to lack the direct terminal repeats found in other T7-like phages. Here we sequenced the ends of the P-SSP7 genome and found that the genome map is linear and contains a 206 bp repeat at both genome ends. Furthermore, we found that a 728 bp region of the genome originally placed downstream of the last ORF is actually located upstream of the first ORF on the genome map. These findings suggest that P-SSP7 is likely to use the direct terminal repeats for genome replication and packaging in a similar manner to other T7-like phages. Moreover, these results highlight the importance of experimentally verifying the ends of phage genomes, and will facilitate the use of P-SSP7 as a model for the correct assembly and end determination of the many T7-like phages isolated from the marine environment that are currently being sequenced.     

Genomic analysis of Pseudomonas aeruginosa phages LKD16 and LKA1: establishment of the phiKMV subgroup within the T7 supergroup

Lytic Pseudomonas aeruginosa phages LKD16 and LKA1 were locally isolated and morphologically classified as Podoviridae. While LKD16 adsorbs weakly to its host, LKA1 shows efficient adsorption (ka = 3.9 x 10(-9) ml min(-1)). LKA1, however, displays a narrow host range on clinical P. aeruginosa strains compared to LKD16. Genome analysis of LKD16 (43,200 bp) and LKA1 (41,593 bp) revealed that both phages have linear double-stranded DNA genomes with direct terminal repeats of 428 and 298 bp and encode 54 and 56 genes, respectively. The majority of the predicted structural proteins were experimentally confirmed as part of the phage particle using mass spectrometry. Phage LKD16 is closely related to bacteriophage phiKMV (83% overall DNA homology), allowing a more thoughtful gene annotation of both genomes. In contrast, LKA1 is more distantly related, lacking significant DNA homology and showing protein similarity to phiKMV in 48% of its gene products. The early region of the LKA1 genome has diverged strongly from phiKMV and LKD16, and intriguing differences in tail fiber genes of LKD16 and LKA1 likely reflect the observed discrepancy in infection-related properties. Nonetheless, general genome organization is clearly conserved among phiKMV, LKD16, and LKA1. The three phages carry a single-subunit RNA polymerase gene adjacent to the structural genome region, a feature which distinguishes them from other members of the T7 supergroup. Therefore, we propose that phiKMV represents an independent and widespread group of lytic P. aeruginosa phages within the T7 supergroup.     

Improving the Safety of Staphylococcus aureus Polyvalent Phages by Their Production on a Staphylococcus xylosus Strain

Team1 (vB_SauM_Team1) is a polyvalent staphylococcal phage belonging to the Myoviridae family. Phage Team1 was propagated on a Staphylococcus aureus strain and a non-pathogenic Staphylococcus xylosus strain used in industrial meat fermentation. The two Team1 preparations were compared with respect to their microbiological and genomic properties. The burst sizes, latent periods, and host ranges of the two derivatives were identical as were their genome sequences. Phage Team1 has 140,903 bp of double stranded DNA encoding for 217 open reading frames and 4 tRNAs. Comparative genomic analysis revealed similarities to staphylococcal phages ISP (97%) and G1 (97%). The host range of Team1 was compared to the well-known polyvalent staphylococcal phages phi812 and K using a panel of 57 S. aureus strains collected from various sources. These bacterial strains were found to represent 18 sequence types (MLST) and 14 clonal complexes (eBURST). Altogether, the three phages propagated on S. xylosus lysed 52 out of 57 distinct strains of S. aureus. The identification of phage-insensitive strains underlines the importance of designing phage cocktails with broadly varying and overlapping host ranges. Taken altogether, our study suggests that some staphylococcal phages can be propagated on food-grade bacteria for biocontrol and safety purposes.     

Two Phages,phiIPLA-RODI and phiIPLA-C1C,Lyse Mono- and Dual-Species Staphylococcal Biofilms

Phage therapy is a promising option for fighting against staphylococcal infections. Two lytic phages, vB_SauM_phiIPLA-RODI (phiIPLA-RODI) and vB_SepM_phiIPLA-C1C (phiIPLA-C1C), belonging to the Myoviridae family and exhibiting wide host ranges, were characterized in this study. The complete genome sequences comprised 142,348 bp and 140,961 bp and contained 213 and 203 open reading frames, respectively. The gene organization was typical of Spounavirinae members, with long direct terminal repeats (LTRs), genes grouped into modules not clearly separated from each other, and several group I introns. In addition, four genes encoding tRNAs were identified in phiIPLA-RODI. Comparative DNA sequence analysis showed high similarities with two phages, GH15 and 676Z, belonging to the Twort-like virus genus (nucleotide identities of >84%); for phiIPLA-C1C, a high similarity with phage phiIBB-SEP1 was observed (identity of 80%). Challenge assays of phages phiIPLA-RODI and phiIPLA-C1C against planktonic staphylococcal cells confirmed their lytic ability, as they were able to remove 5 log units in 8 h. Exposure of biofilms to phages phiIPLA-RODI and phiIPLA-C1C reduced the amount of adhered bacteria to about 2 log units in both monospecies and dual-species biofilms, but phiIPLA-RODI turned out to be as effective as the mixture of both phages. Moreover, the frequencies of bacteriophage-insensitive mutants (BIMs) of Staphylococcus aureus and S. epidermidis with resistance to phiIPLA-RODI and phiIPLA-C1C were low, at 4.05 × 10⁻⁷ ± 2.34 × 10⁻⁹ and 1.1 × 10⁻⁷ ± 2.08 × 10⁻⁹, respectively. Overall, a generally reduced fitness in the absence of phages was observed for BIMs, which showed a restored phage-sensitive phenotype in a few generations. These results confirm that lytic bacteriophages can be efficient biofilm-disrupting agents, supporting their potential as antimicrobials against staphylococcal infections.     

Genomic analysis of bacteriophages SP6 and K1-5, an estranged subgroup of the T7 supergroup

We have determined the genome sequences of two closely related lytic bacteriophages, SP6 and K1-5, which infect Salmonella typhimurium LT2 and Escherichia coli serotypes K1 and K5, respectively. The genome organization of these phages is almost identical with the notable exception of the tail fiber genes that confer the different host specificities. The two phages have diverged extensively at the nucleotide level but they are still more closely related to each other than either is to any other phage currently characterized. The SP6 and K1-5 genomes contain, respectively, 43,769 bp and 44,385 bp, with 174 bp and 234 bp direct terminal repeats. About half of the 105 putative open reading frames in the two genomes combined show no significant similarity to database proteins with a known or predicted function that is obviously beneficial for growth of a bacteriophage. The overall genome organization of SP6 and K1-5 is comparable to that of the T7 group of phages, although the specific order of genes coding for DNA metabolism functions has not been conserved. Low levels of nucleotide similarity between genomes in the T7 and SP6 groups suggest that they diverged a long time ago but, on the basis of this conservation of genome organization, they are expected to have retained similar developmental strategies.     

Characterization of the Genome of the Dairy Lactobacillus helveticus Bacteriophage ΦAQ113

The complete genomic sequence of the dairy Lactobacillus helveticus bacteriophage ΦAQ113 was determined. Phage ΦAQ113 is a Myoviridae bacteriophage with an isometric capsid and a contractile tail. The final assembled consensus sequence revealed a linear, circularly permuted, double-stranded DNA genome with a size of 36,566 bp and a G+C content of 37%. Fifty-six open reading frames (ORFs) were predicted, and a putative function was assigned to approximately 90% of them. The ΦAQ113 genome shows functionally related genes clustered together in a genome structure composed of modules for DNA replication/regulation, DNA packaging, head and tail morphogenesis, cell lysis, and lysogeny. The identification of genes involved in the establishment of lysogeny indicates that it may have originated as a temperate phage, even if it was isolated from natural cheese whey starters as a virulent phage, because it is able to propagate in a sensitive host strain. Additionally, we discovered that the ΦAQ113 phage genome is closely related to Lactobacillus gasseri phage KC5a and Lactobacillus johnsonii phage Lj771 genomes. The phylogenetic similarities between L. helveticus phage ΦAQ113 and two phages that belong to gut species confirm a possible common ancestral origin and support the increasing consideration of L. helveticus as a health-promoting organism.     

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 M11S (Siphoviridae) did not correspond to any other described Rhizobium phages and represents a new species.     

Comparative whole genome analysis of six diagnostic brucellaphages

Whole genome sequencing of six diagnostic brucellaphages, Tbilisi (Tb), Firenze (Fz), Weybridge (Wb), S708, Berkeley (Bk) and R/C, was followed with genomic comparisons including recently described genomes of the Tb phage from Mexico (Tb_M) and Pr phage to elucidate genomic diversity and candidate host range determinants. Comparative whole genome analysis revealed high sequence homogeneity among these brucellaphage genomes and resolved three genetic groups consistent with defined host range phenotypes. Group I was composed of Tb and Fz phages that are predominantly lytic for Brucella abortus and Brucella neotomae; Group II included Bk, R/C, and Pr phages that are lytic mainly for B. abortus, Brucella melitensis and Brucella suis; Group III was composed of Wb and S708 phages that are lytic for B. suis, B. abortus and B. neotomae. We found that the putative phage collar protein is a variable locus with features that may be contributing to the host specificities exhibited by different brucellaphage groups. The presence of several candidate host range determinants is illustrated herein for future dissection of the differential host specificity observed among these phages.     

Isolation of new Pseudomonas tolaasii bacteriophages and genomic investigation of the lytic phage BF7

Sixteen lytic bacteriophages that infect Pseudomonas tolaasii LMG 2342(T) were isolated from smashed sporocarps of oyster mushroom (Pleurotus ostreatus) showing necrotic symptoms. On the basis of the host range investigation of the phages, they have wide infection abilities against the genus Pseudomonas, mainly in the case of phages Bf3, Bf7, Bf10, and Bf15. Molecular investigations have revealed that they all have dsDNA genomes about 40?kbp in size. Identical restriction patterns resulting from restriction enzyme analysis suggest that the isolates probably belong to the same phage species. However, there was a difference between these phage isolates in their infecting abilities. Phage isolate Bf7 was investigated and characterized more deeply. Morphological characterization of Bf7 by transmission electron microscopy (TEM) has shown that it has a short, noncontractile tail, an icosahedral phage head, and the size is about 60?nm in diameter, suggesting that it belongs to the Podoviridae family. Complete genome sequence analysis of the Bf7 phage isolate revealed a 40?058?bp genome, 58.4% G+C content, 46 open reading frames encoding different proteins showing homology to proteins of the bacteriophage Caulobacter crescentus φCd1 from the Podoviridae family. On the basis of these results and comparative genomic studies, we classified the Bf7 phage to the subfamily of Autographivirinae, φKMV-like phages.     

Genomic Diversity of Phages Infecting Probiotic Strains of Lactobacillus paracasei

Strains of the Lactobacillus casei group have been extensively studied because some are used as probiotics in foods. Conversely, their phages have received much less attention. We analyzed the complete genome sequences of five L. paracasei temperate phages: C_L1, C_L2, iLp84, iLp1308, and iA2. Only phage iA2 could not replicate in an indicator strain. The genome lengths ranged from 34,155 bp (iA2) to 39,474 bp (C_L1). Phages iA2 and iLp1308 (34,176 bp) possess the smallest genomes reported, thus far, for phages of the L. casei group. The GC contents of the five phage genomes ranged from 44.8 to 45.6%. As observed with many other phages, their genomes were organized as follows: genes coding for DNA packaging, morphogenesis, lysis, lysogeny, and replication. Phages C_L1, C_L2, and iLp1308 are highly related to each other. Phage iLp84 was also related to these three phages, but the similarities were limited to gene products involved in DNA packaging and structural proteins. Genomic fragments of phages C_L1, C_L2, iLp1308, and iLp84 were found in several genomes of L. casei strains. Prophage iA2 is unrelated to these four phages, but almost all of its genome was found in at least four L. casei strains. Overall, these phages are distinct from previously characterized Lactobacillus phages. Our results highlight the diversity of L. casei phages and indicate frequent DNA exchanges between phages and their hosts.     

A New Microviridae Phage Isolated from a Failed Biotechnological Process Driven by Escherichia coli

Bacteriophages are present in every environment that supports bacterial growth, including manmade ecological niches. Virulent phages may even slow or, in more severe cases, interrupt bioprocesses driven by bacteria. Escherichia coli is one of the most widely used bacteria for large-scale bioprocesses; however, literature describing phage-host interactions in this industrial context is sparse. Here, we describe phage MED1 isolated from a failed industrial process. Phage MED1 (Microviridae family, with a single-stranded DNA [ssDNA] genome) is highly similar to the archetypal phage phiX174, sharing >95% identity between their genomic sequences. Whole-genome phylogenetic analysis of 52 microvirus genomes from public databases revealed three genotypes (alpha3, G4, and phiX174). Phage MED1 belongs to the phiX174 group. We analyzed the distribution of single nucleotide variants in MED1 and 18 other phiX174-like genomes and found that there are more missense mutations in genes G, B, and E than in the other genes of these genomes. Gene G encodes the spike protein, involved in host attachment. The evolution of this protein likely results from the selective pressure on phages to rapidly adapt to the molecular diversity found at the surface of their hosts.     

Isolation and Characterization of Bacteriophages Infecting Staphylococcus epidermidis

Bacteriophages infecting Staphylococcus epidermidis were isolated by mitomycin C induction. Three distinct phages (vB_SepiS-phiIPLA5, vB_SepiS-phiIPLA6, and vB_SepiS-phiIPLA7)—defined by plaque morphology, structure, virion proteins pattern, DNA restriction bands, and host range—were obtained. One-step growth curves of bacteriophages under optimal growth conditions for S. epidermidis F12 revealed eclipse and latent periods of 5–10 and 10–15 min, respectively, with burst sizes of about 5 to 30 PFU per infected cell. Transmission electron microscopy revealed that the phages were of similar size and belonged to the Siphoviridae family. Phage phi-IPLA7 had the broadest host range infecting 21 out of 65 S. epidermidis isolates. Phage phi-IPLA5 seemed to be a virulent phage probably derived from phi-IPLA6. Phages phi-IPLA5 and phi-IPLA7 exhibited increasing plaques surrounded by a halo that could be indicative of a polysaccharide depolymerase activity. Viable counts, determined during the infection of S. epidermidis F12, confirmed that phi-IPLA5 had a potent lytic capability and reduced S. epidermidis population by 5.67 log units in 8 h of incubation; in the presence of the mixture of phi-IPLA6 and phi-IPLA7, however, a reduction of 2.27 log units was detected     

Properties of the new D3-like Pseudomonas aeruginosa bacteriophage phiPMG1: Genome structure and prospects for the use in phage therapy

Results of studying the novel virulent phage phiPMG1 active on Pseudomonas aeruginosa are presented. It is shown that phiPMG1 exhibits significant homology and the similarity in the overall structure with the genome of a temperate phage converts D3. Phage phiPMG1 differs from D3 in that it fails to stably lysogenize bacteria and can grow on strains carrying plasmids that cause growth inhibition of phage D3 and some other phages. This significantly diminishes the probability of horizontal gene transfer with phage phiPMG1 and suggests the possible employment of this phage in phage therapy. A comparison of phages phiPMG1 and D3 structures of genomes in demonstrated not only high homology of 65 genes, but also the presence of 16 genes in the phiPMG1 genome that were not included in the in NCBI database. Apparently, the evolution of genomes in phages of this species is mostly associated with migrations into other species of bacteria, and recombinations with phages of other species (for example, F116). A detailed analysis of structure of one region genomes, which significant nonhomology for the three D3-like phages (D3, phiPMG1 and PAJU2), revealed that the phiPMG1 genome possible closest to a hypothetical genome of ancestral phage of this species.     

The terminally redundant, nonpermuted genome of Listeria bacteriophage A511: a model for the SPO1-like myoviruses of gram-positive bacteria

Only little information on a particular class of myoviruses, the SPO1-like bacteriophages infecting low-G+C-content, gram-positive host bacteria (Firmicutes), is available. We present the genome analysis and molecular characterization of the large, virulent, broad-host-range Listeria phage A511. A511 contains a unit (informational) genome of 134,494 bp, encompassing 190 putative open reading frames (ORFs) and 16 tRNA genes, organized in a modular fashion common among the Caudovirales. Electron microscopy, enzymatic fragmentation analyses, and sequencing revealed that the A511 DNA molecule contains linear terminal repeats of a total of 3,125 bp, encompassing nine small putative ORFs. This particular genome structure explains why A511 is unable to perform general transduction. A511 features significant sequence homologies to Listeria phage P100 and other morphologically related phages infecting Firmicutes such as Staphylococcus phage K and Lactobacillus phage LP65. Equivalent but more-extensive terminal repeats also exist in phages P100 (approximately 6 kb) and K (approximately 20 kb). High-resolution electron microscopy revealed, for the first time, the presence of long tail fibers organized in a sixfold symmetry in these viruses. Mass spectrometry-based peptide fingerprinting permitted assignment of individual proteins to A511 structural components. On the basis of the data available for A511 and relatives, we propose that SPO1-like myoviruses are characterized by (i) their infection of gram-positive, low-G+C-content bacteria; (ii) a wide host range within the host bacterial genus and a strictly virulent lifestyle; (iii) similar morphology, sequence relatedness, and collinearity of the phage genome organization; and (iv) large double-stranded DNA genomes featuring nonpermuted terminal repeats of various sizes.     

A new group of cosmopolitan bacteriophages induce a carrier state in the pandemic strain of Vibrio parahaemolyticus

A clonal population of pathogenic Vibrio parahaemolyticus O3 : K6 serovar has spread in coastal waters, causing outbreaks worldwide since 1996. Bacteriophage infection is one of the main factors affecting bacterial strain concentration in the ocean. We studied the occurrence and properties of phages infecting this V. parahaemolyticus pandemic strain in coastal waters. Analysing 143 samples, phages were found in 13. All isolates clustered in a closely related group of podophages with at least 90% nucleotide sequence identity in three essential genes, despite distant geographical origins. These bacteriophages were able to multiply on the V. parahaemolyticus pandemic strain, but the impact on host concentration and subsequent growth was negligible. Infected bacteria continued producing the phage but were not lysogenized. The phage genome of prototype strain VP93 is 43 931 nucleotides and contains 337 bp direct terminal repeats at both ends. VP93 is the first non‐Pseudomonas phage related to the ΦKMV‐like subgroup of the T7 supergroup. The lack of a major effect on host growth suggests that these phages exert little control on the propagation of the pandemic strain in the environment. This form of phage growth can be modelled if phage‐sensitive and ‐resistant cells that convert to each other with a high frequency are present in clonal cultures of pandemic V. parahaemolyticus.     

Biology and genome sequence of Streptococcus mutans phage M102AD

M102AD is the new designation for a Streptococcus mutans phage described in 1993 as phage M102. This change was necessitated by the genome analysis of another S. mutans phage named M102, which revealed differences from the genome sequence reported here. Additional host range analyses confirmed that S. mutans phage M102AD infects only a few serotype c strains. Phage M102AD adsorbed very slowly to its host, and it cannot adsorb to serotype e and f strains of S. mutans. M102AD adsorption was blocked by c-specific antiserum. Phage M102AD also adsorbed equally well to heat-treated and trypsin-treated cells, suggesting carbohydrate receptors. Saliva and polysaccharide production did not inhibit plaque formation. The genome of this siphophage consisted of a linear, double-stranded, 30,664-bp DNA molecule, with a GC content of 39.6%. Analysis of the genome extremities indicated the presence of a 3'-overhang cos site that was 11 nucleotides long. Bioinformatic analyses identified 40 open reading frames, all in the same orientation. No lysogeny-related genes were found, indicating that phage M102AD is strictly virulent. No obvious virulence factor gene candidates were found. Twelve proteins were identified in the virion structure by mass spectrometry. Comparative genomic analysis revealed a close relationship between S. mutans phages M102AD and M102 as well as with Streptococcus thermophilus phages. This study also highlights the importance of conducting research with biological materials obtained from recognized microbial collections.