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Bacterial diseases caused by Vibrio spp. are prevalent in aquaculture and can lead to high mortality rates among aquatic species and significant economic losses. With the increasing emergence of multidrug-resistant Vibrio strains, phage therapy is being explored as a potential alternative to antibiotics for biocontrol of infectious diseases. Here, a new lytic phage named vB_VhaS_R21Y (R21Y) was isolated against Vibrio harveyi BVH1 obtained from seawater from a scallop-farming area in Rongcheng, China. Its morphology, infection cycle, lytic profile, phage stability, and genetic features were characterized. Transmission electronic microscopy indicated that R21Y is siphovirus-like, comprising an icosahedral head (diameter 73.31 ± 2.09 nm) and long noncontractile tail (205.55 ± 0.75 nm). In a one-step growth experiment, R21Y had a 40-min latent period and a burst size of 35 phage particles per infected cell. R21Y was highly species-specific in the host range test and was relatively stable at pH 4–10 and 4–55 °C. Genomic analysis showed that R21Y is a double-stranded DNA virus with a genome size of 82,795 bp and GC content of 47.48%. Its high tolerance and lytic activity indicated that R21Y may be a candidate for phage therapy in controlling vibriosis in aquacultural systems.

A high temperature-adapted bacteriophage, vB_PtoS_NIIg3.2 (NIIg3.2), was isolated in Lithuania from compost heaps using Parageobacillus toebii strain NIIg-3 as a host for phage propagation. Furthermore, NIIg3.2 was active against four strains of Geobacillus thermodenitrificans, and it infected the host cells from 50 to 80 °C. Transmission electron microscopy analysis revealed siphovirus morphology characterized by an isometric head (~59 nm in diameter) and a noncontractile tail (~226 nm in length). The double-stranded DNA genome of NIIg3.2 (38,970 bp) contained 71 probable protein-encoding genes and no genes for tRNA. In total, 29 NIIg3.2 ORFs were given a putative functional annotation, including those coding for the proteins responsible for DNA packaging, virion structure/morphogenesis, phage–host interactions, lysis/lysogeny, replication/regulation, and nucleotide metabolism. Based on comparative phylogenetic and bioinformatic analysis, NIIg3.2 cannot be assigned to any genus currently recognized by ICTV and potentially represents a new one within siphoviruses. The results of this study not only extend our knowledge about poorly explored thermophilic bacteriophages but also provide new insights for further investigation and understanding the evolution of Bacilllus-group bacteria-infecting viruses.

In the human gut, temperate bacteriophages interact with bacteria through predation and horizontal gene transfer. Relying on taxonomic data, metagenomic studies have associated shifts in phage abundance with a number of human diseases. The temperate bacteriophage VEsP-1 with siphovirus morphology was isolated from a sample of river water using Enterococcus faecalis as a host. Starting from the whole genome sequence of VEsP-1, we retrieved related phage genomes in blastp searches of the tail protein and large terminase sequences, and blastn searches of the whole genome sequences, with matches compiled from several different databases, and visualized a part of viral dark matter sequence space. The genome network and phylogenomic analyses resulted in the proposal of a novel genus “Vespunovirus”, consisting of temperate, mainly metagenomic phages infecting Enterococcus spp.

Phage vB_YenS_P400 isolated from deer, is a virulent siphovirus of Y. enterocolitica, whose circularly permutated genome (46,585 bp) is not substantially related to any other phage deposited in public nucleotide databases. vB_YenS_P400 showed a very narrow host range and exclusively lysed two Y. enterocolitica B4/O:3 strains. Moreover, lytic activity by this phage was only discernible at room temperature. Together with the finding that vB_YenS_P400 revealed a long latent period (90 to 100 min) and low burst size (five to ten), it is not suitable for applications but provides insight into the diversity of Yersinia phages.

Bacillus anthracis is a potent biowarfare agent, able to be highly lethal. The bacteria dwell in the soil of certain regions, as natural flora. Bacteriophages or their lytic enzymes, endolysins, may be an alternative for antibiotics and other antibacterials to fight this pathogen in infections and to minimize environmental contamination with anthrax endospores. Upon screening environmental samples from various regions in Poland, we isolated three new siphophages, J5a, F16Ba, and z1a, specific for B. anthracis. They represent new species related to historical anthrax phages Gamma, Cherry, and Fah, and to phage Wbeta of Wbetavirus genus. We show that the new phages and their closest relatives, phages Tavor_SA, Negev_SA, and Carmel_SA, form a separate clade of the Wbetavirus genus, designated as J5a clade. The most distinctive feature of J5a clade phages is their cell lysis module. While in the historical phages it encodes a canonical endolysin and a class III holin, in J5a clade phages it encodes an endolysin with a signal peptide and two putative holins. We present the basic characteristic of the isolated phages. Their comparative genomic analysis indicates that they encode two receptor-binding proteins, of which one may bind a sugar moiety of B. anthracis cell surface.

A novel siphovirus, vB_PagS_MED16 (MED16) was isolated in Lithuania using Pantoea agglomerans strain BSL for the phage propagation. The double-stranded DNA genome of MED16 (46,103 bp) contains 73 predicted open reading frames (ORFs) encoding proteins, but no tRNA. Our comparative sequence analysis revealed that 26 of these ORFs code for unique proteins that have no reliable identity when compared to database entries. Based on phylogenetic analysis, MED16 represents a new genus with siphovirus morphology. In total, 35 MED16 ORFs were given a putative functional annotation, including those coding for the proteins responsible for virion morphogenesis, phage–host interactions, and DNA metabolism. In addition, a gene encoding a preQ0 DNA deoxyribosyltransferase (DpdA) is present in the genome of MED16 and the LC–MS/MS analysis indicates 2′-deoxy-7-amido-7-deazaguanosine (dADG)-modified phage DNA, which, to our knowledge, has never been experimentally validated in genomes of Pantoea phages. Thus, the data presented in this study provide new information on Pantoea-infecting viruses and offer novel insights into the diversity of DNA modifications in bacteriophages.

A cold-adapted siphovirus, vB_PagS_AAS23 (AAS23) was isolated in Lithuania using the Pantoea agglomerans strain AUR for the phage propagation. The double-stranded DNA genome of AAS23 (51,170 bp) contains 92 probable protein encoding genes, and no genes for tRNA. A comparative sequence analysis revealed that 25 of all AAS23 open reading frames (ORFs) code for unique proteins that have no reliable identity to database entries. Based on the phylogenetic analysis, AAS23 has no close relationship to other viruses publicly available to date and represents a single species of the genus Sauletekiovirus within the family Drexlerviridae. The phage is able to form plaques in bacterial lawns even at 4 °C and demonstrates a depolymerase activity. Thus, the data presented in this study not only provides the information on Pantoea-infecting bacteriophages, but also offers novel insights into the diversity of cold-adapted viruses and their potential to be used as biocontrol agents.

Despite a wealth of knowledge on Salmonella phages worldwide, little is known about poultry-associated Salmonella phages from Thailand. Here, we isolated 108 phages from Thai poultry farms that infect Salmonella enterica serovar Typhimurium. Phages STm101 and STm118 were identified as temperate Siphoviridae phages. Genome sequencing and analyses revealed these phages share approximately 96% nucleotide sequence similarity to phage SPN19, a member of the Chi-like virus genus. PCR amplification of the gene encoding capsid protein E of the Chi-like phage was positive for 50% of phage isolates, suggesting a predominance of this phage type among the sampled poultry farms. In addition to the flagella, two phages required the lipopolysaccharide to infect and lyse Salmonella. Furthermore, phylogenomic analysis demonstrated that phages STm101 and STm118 formed a monophyletic clade with phages isolated from Western countries, but not from closer isolated phages from Korea. However, further investigation and more phage isolates are required to investigate possible causes for this geographic distribution.

Nineteen Vibrio anguillarum-specific temperate bacteriophages isolated across Europe and Chile from aquaculture and environmental sites were genome sequenced and analyzed for host range, morphology and life cycle characteristics. The phages were classified as Siphoviridae with genome sizes between 46,006 and 54,201 bp. All 19 phages showed high genetic similarity, and 13 phages were genetically identical. Apart from sporadically distributed single nucleotide polymorphisms (SNPs), genetic diversifications were located in three variable regions (VR1, VR2 and VR3) in six of the phage genomes. Identification of specific genes, such as N6-adenine methyltransferase and lambda like repressor, as well as the presence of a tRNAArg, suggested a both mutualistic and parasitic interaction between phages and hosts. During short term phage exposure experiments, 28% of a V. anguillarum host population was lysogenized by the temperate phages and a genomic analysis of a collection of 31 virulent V. anguillarum showed that the isolated phages were present as prophages in >50% of the strains covering large geographical distances. Further, phage sequences were widely distributed among CRISPR-Cas arrays of publicly available sequenced Vibrios. The observed distribution of these specific temperate Vibriophages across large geographical scales may be explained by efficient dispersal of phages and bacteria in the marine environment combined with a mutualistic interaction between temperate phages and their hosts which selects for co-existence rather than arms race dynamics.

Phage vB_YenS_P400 isolated from deer, is a virulent siphovirus of Y. enterocolitica , whose circularly permutated genome (46,585 bp) is not substantially related to any other phage deposited in public nucleotide databases. vB_YenS_P400 showed a very narrow host range and exclusively lysed two Y. enterocolitica B4/O:3 strains. Moreover, lytic activity by this phage was only discernible at room temperature. Together with the finding that vB_YenS_P400 revealed a long latent period (90 to 100 min) and low burst size (five to ten), it is not suitable for applications but provides insight into the diversity of Yersinia phages.

A cold-adapted siphovirus, vB_PagS_AAS23 (AAS23) was isolated in Lithuania using the Pantoea agglomerans strain AUR for the phage propagation. The double-stranded DNA genome of AAS23 (51,170 bp) contains 92 probable protein encoding genes, and no genes for tRNA. A comparative sequence analysis revealed that 25 of all AAS23 open reading frames (ORFs) code for unique proteins that have no reliable identity to database entries. Based on the phylogenetic analysis, AAS23 has no close relationship to other viruses publicly available to date and represents a single species of the genus Sauletekiovirus within the family Drexlerviridae . The phage is able to form plaques in bacterial lawns even at 4 °C and demonstrates a depolymerase activity. Thus, the data presented in this study not only provides the information on Pantoea -infecting bacteriophages, but also offers novel insights into the diversity of cold-adapted viruses and their potential to be used as biocontrol agents.