Characterization and complete genome sequence of vB_EcoP-Bp4, a novel polyvalent N4-like bacteriophage that infects chicken pathogenic Escherichia coli.
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ABSTRACT:
SUBMITTER: Zhang C
PROVIDER: S-EPMC8193434 | biostudies-literature | 2016 Aug
REPOSITORIES: biostudies-literature
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Characterization and complete genome sequence of vB_EcoP-Bp4, a novel polyvalent N4-like bacteriophage that infects chicken pathogenic Escherichia coli.
Zhang Can C Ma Yanxiang Y Wang Ting T Sun Huzhi H Lu Guomin G Ren Huiying H
Virologica Sinica 20160801 4
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Project description:A novel virulent bacteriophage vB_EcoM_swi3 (swi3), isolated from swine feces, lyzed 9% (6/65) of Escherichia coli and isolates 54% (39/72) of Salmonella enteritidis isolates, which were all clinically pathogenic multidrug-resistant strains. Morphological observation showed that phage swi3 belonged to the Myoviridae family with an icosahedral head (80 nm in diameter) and a contractile sheathed tail (120 nm in length). At the optimal multiplicity of infection of 1, the one-step growth analysis of swi3 showed a 25-min latent period with a burst size of 25-plaque-forming units (PFU)/infected cell. Phage swi3 remained stable both at pH 6.0-8.0 and at less than 50°C for at least 1 h. Genomic sequencing and bioinformatics analysis based on genomic sequences and the terminase large subunit showed that phage swi3 was a novel member that was most closely related to Salmonella phages and belonged to the Rosemountvirus genus. Phage swi3 harbored a 52-kb double-stranded DNA genome with 46.02% GC content. Seventy-two potential open reading frames were identified and annotated, only 15 of which had been assigned to functional genes. No gene associated with pathogenicity and virulence was identified. The effects of phage swi3 in treating pathologic E. coli infections in vivo were evaluated using a mouse model. The administration of a single intraperitoneal injection of swi3 (106 PFU) at 2 h after challenge with the E. coli strain (serotype K88) (108 colony-forming units) sufficiently protected all mice without toxic side effects. This finding highlighted that phage swi3 might be used as an effective antibacterial agent to prevent E. coli and S. enteritidis infection.
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Project description:BACKGROUND: Bacteriophage EC1-UPM is an N4-like bacteriophage which specifically infects Escherichia coli O78:K80, an avian pathogenic strain that causes colibacillosis in poultry. The complete genome sequence of bacteriophage EC1-UPM was analysed and compared with other closely related N4-like phage groups to assess their genetic similarities and differences. RESULTS: Bacteriophage EC1-UPM displays a very similar codon usage profile with its host and does not contain any tRNA gene. Comparative genomics analysis reveals close resemblance of bacteriophage EC1-UPM to three N4-like bacteriophages namely vB_EcoP_G7C, IME11 and KBNP21 with a total of 44 protein coding genes shared at 70% identity threshold. The genomic region coding for the tail fiber protein was found to be unique in bacteriophage EC1-UPM. Further annotation of the tail fiber protein using HHpred, a highly sensitive homology detection tool, reveals the presence of protein structure homologous to various polysaccharide processing proteins in its C-terminus. Leveraging on the availability of multiple N4-like bacteriophage genome sequences, the core genes of N4-like bacteriophages were identified and used to perform a multilocus phylogenetic analysis which enabled the construction of a phylogenetic tree with higher confidence than phylogenetic trees based on single genes. CONCLUSION: We report for the first time the complete genome sequence of a N4-like bacteriophage which is lytic against avian pathogenic Escherichia coli O78:K80. A novel 928 amino acid residues tail fiber protein was identified in EC1-UPM which may be useful to further the understanding of phage-host specificity. Multilocus phylogenetic analysis using core genes of sequenced N4-like phages showed that the evolutionary relationship correlated well with the pattern of host specificity.
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Project description:Avian pathogenic Escherichia coli (APEC) is a major pathogen that causes avian colibacillosis and is associated with severe economic losses in the chicken-farming industry. Here, bacteriophage KBNP1315, infecting APEC strain KBP1315, was genomically and functionally characterized. The evolutionary relationships of KBNP1315 were analyzed at the genomic level using gene (protein)-sharing networks, the Markov clustering (MCL) algorithm, and comparative genomics. Our network analysis showed that KBNP1315 was connected to 30 members of the Autographivirinae subfamily, which comprises the SP6-, T7-, P60-, phiKMV-, GAP227- and KP34-related groups. Network decomposition suggested that KBNP1315 belongs to the SP6-like phages, but our comparison of putative encoded proteins revealed that key proteins of KBNP1315, including the tail spike protein and endolysin, had relative low levels of amino acid sequence similarity with other members of the SP6-like phages. Thus KBNP1315 may only be distantly related to the SP6-like phages, and (based on the difference in endolysin) its lysis mechanism may differ from theirs. To characterize the lytic functions of the holin and endolysin proteins from KBNP1315, we expressed these proteins individually or simultaneously in E. coli BL21 (DE3) competent cell. Interestingly, the expressed endolysin was secreted into the periplasm and caused a high degree of host cell lysis that was dose-dependently delayed/blocked by NaN3-mediated inhibition of the SecA pathway. The expressed holin triggered only a moderate inhibition of cell growth, whereas coexpression of holin and endolysin enhanced the lytic effect of endolysin. Together, these results revealed that KBNP1315 appears to use a pin-holin/signal-arrest-release (SAR) endolysin pathway to trigger host cell lysis.
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Project description:The characterization of a recently isolated bacteriophage, vB_Eco4M-7, which effectively infects many, though not all, Escherichia coli O157 strains, is presented. The genome of this phage comprises double-stranded DNA, 68,084 bp in length, with a GC content of 46.2%. It contains 96 putative open reading frames (ORFs). Among them, the putative functions of only 35 ORFs were predicted (36.5%), whereas 61 ORFs (63.5%) were classified as hypothetical proteins. The genome of phage vB_Eco4M-7 does not contain genes coding for integrase, recombinase, repressors or excisionase, which are the main markers of temperate viruses. Therefore, we conclude that phage vB_Eco4M-7 should be considered a lytic virus. This was confirmed by monitoring phage lytic development by a one-step growth experiment. Moreover, the phage forms relatively small uniform plaques (1 mm diameter) with no properties of lysogenization. Electron microscopic analyses indicated that vB_Eco4M-7 belongs to the Myoviridae family. Based on mass spectrometric analyses, including the fragmentation pattern of unique peptides, 33 phage vB_Eco4M-7 proteins were assigned to annotated open reading frames. Importantly, genome analysis suggested that this E. coli phage is free of toxins and other virulence factors. In addition, a similar, previously reported but uncharacterized bacteriophage, ECML-117, was also investigated, and this phage exhibited properties similar to vB_Eco4M-7. Our results indicate that both studied phages are potential candidates for phage therapy and/or food protection against Shiga toxin-producing E. coli, as the majority of these strains belong to the O157 serotype.
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