Abstract
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.
Highlights
Avian colibacillosis, which is caused by strains of avian pathogenic Escherichia coli (APEC), is a severe bird disease that affects the poultry industry and is responsible for enormous economic losses worldwide [1, 2]
Escherichia coli is typically a commensal microorganism in the intestinal tract of chickens, but APEC strains are extraintestinal pathogenic E. coli (ExPEC) whose infection is characterized by various symptoms, such as airsacculitis, polyserositis, enteritis, cellulitis, salpingitis, omphilitis, coligranuloma, egg peritonitis and septicemia [1,2,3]
Phage genomics have been advancing as a result of the remarkable development in phage genome sequencing, which contributes to the progress of genetic, biotechnological and clinical tools [12, 13]
Summary
Avian colibacillosis, which is caused by strains of avian pathogenic Escherichia coli (APEC), is a severe bird disease that affects the poultry industry and is responsible for enormous economic losses worldwide [1, 2]. Escherichia coli is typically a commensal microorganism in the intestinal tract of chickens, but APEC strains are extraintestinal pathogenic E. coli (ExPEC) whose infection is characterized by various symptoms, such as airsacculitis, polyserositis, enteritis, cellulitis, salpingitis, omphilitis, coligranuloma, egg peritonitis and septicemia [1,2,3]. Because the process of phage evolution is being affected by frequent horizontal gene transfer (HGT), phage genomes show pervasive mosaicism and it is difficult to characterize them by classical taxonomic study based on hierarchical classification [10, 11, 13]. To overcome the limitation of the classic approach, Lima-Mendez et al [11] proposed a reticulated network model representing the phage-phage relationship in consideration of shared gene (protein) contents, including the evolutionary reconstructions of individual gene phylogeny; this model gives a clear understanding of the true evolutionary history of phage genomes [10, 14]
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