Escherichia coli bacteriophage Gostya9, representing a new specieswithin the genus T5virus

AbstractEscherichia coli bacteriophage Gostya9 (genus T5virus) was isolated from horse feces collected in Moscow, Russia, in 2013.This phage was associated in a single plaque with the previously reported phage 9g and was subsequently purified. Analysisof the complete genomic sequence of Gostya9 revealed that it is closely related to the T5-like bacteriophage DT57C, whichhad been isolated at the same location in 2007. These two viruses share 79.5% nucleotide sequence identity, which is belowthe 95% threshold applied currently to demarcate bacteriophage species. The most significant features distinguishing Gostya9from DT57C include 1) the presence of one long tail fiber protein gene, 122c (ltf), instead of the two genes, ltfA and ltfB,that are present in DT57C; 2) the absence of the gene for the receptor-blocking lytic conversion lipoprotein precursor llp;and 3) the divergence of the receptor-recognition protein, pb5, which is only distantly related at the amino acid sequencelevel. The observed features of the Gostya9 adsorption apparatus are suggestive of a possible novel specificity for the finalreceptor and make this phage interesting for possible direct application in phage therapy of E. coli infections or as a sourceof receptor-recognition protein for engineering new phage specificities

RB49-like Bacteriophages Recognize O Antigens as One of the Alternative Primary Receptors

The power of most of the enterobacterial O antigen types to provide robust protection against direct recognition of the cell surface by bacteriophage receptor-recognition proteins (RBP) has been recently recognized. The bacteriophages infecting O antigen producing strains of E. coli employ various strategies to tackle this nonspecific protection. T-even related phages, including RB49-like viruses, often have wide host ranges, being considered good candidates for use in phage therapy. However, the mechanisms by which these phages overcome the O antigen barrier remain unknown. We demonstrate here that RB49 and related phages Cognac49 and Whisky49 directly use certain types of O antigen as their primary receptors recognized by the virus long tail fibers (LTF) RBP gp38, so the O antigen becomes an attractant instead of an obstacle. Simultaneously to recognize multiple O antigen types, LTFs of each of these phages can bind to additional receptors, such as OmpA protein, enabling them to infect some rough strains of E. coli. We speculate that the mechanical force of the deployment of the short tail fibers (STF) triggered by the LTF binding to the O antigen or underneath of it, allows the receptor binding domains of STF to break through the O polysaccharide layer

Complete genome sequences of T5-related Escherichia coli bacteriophages DT57C and DT571/2 isolated from horse feces

We report the complete genome sequencing of two Escherichia coli T5-related bacteriophages, DT57C and DT571/2, isolated from the same specimen of horse feces. These two isolates share 96 % nucleotide sequence identity and can thus be considered representatives of the same novel species within the genus T5likevirus. The observed variation in the ltfA gene of these phages, resulting from a recent recombination event, may explain the observed host-range differences, suggesting that a modular mechanism makes a significant contribution to the short-term evolution (or adaptation) of T5-like phage genomes in the intestinal ecosystem. Comparison of our isolates to their closest relative, coliphage T5, revealed high overall synteny of the genomes and high conservation of the sequences of almost all structural proteins as well as of the other proteins with identified functions. At the same time, numerous alterations and non-orthologous replacements of non-structural protein genes (mostly of those with unknown functions) as well as substantial differences in tail fiber locus organization support the conclusion that DT57C and DT571/2 form a species-level group clearly distinct from bacteriophage T5

Genomic Sequencing and Biological Characteristics of a Novel Escherichia Coli Bacteriophage 9g, a Putative Representative of a New Siphoviridae Genus

Bacteriophage 9g was isolated from horse feces using Escherichia coli C600 as a host strain. Phage 9g has a slightly elongated capsid 62 × 76 nm in diameter and a non-contractile tail about 185 nm long. The complete genome sequence of this bacteriophage consists of 56,703 bp encoding 70 predicted open reading frames. The closest relative of phage 9g is phage PhiJL001 infecting marine alpha-proteobacterium associated with Ircinia strobilina sponge, sharing with phage 9g 51% of amino acid identity in the main capsid protein sequence. The DNA of 9g is resistant to most restriction endonucleases tested, indicating the presence of hypermodified bases. The gene cluster encoding a biosynthesis pathway similar to biosynthesis of the unusual nucleoside queuosine was detected in the phage 9g genome. The genomic map organization is somewhat similar to the typical temperate phage gene layout but no integrase gene was detected. Phage 9g efficiently forms stable associations with its host that continues to produce the phage over multiple passages, but the phage can be easily eliminated via viricide treatment indicating that no true lysogens are formed. Since the sequence, genomic organization and biological properties of bacteriophage 9g are clearly distinct from other known Enterobacteriaceae phages, we propose to consider it as the representative of a novel genus of the Siphoviridae family

Branched Lateral Tail Fiber Organization in T5-Like Bacteriophages DT57C and DT571/2 is Revealed by Genetic and Functional Analysis

The T5-like siphoviruses DT57C and DT571/2, isolated from horse feces, are very closely related to each other, and most of their structural proteins are also nearly identical to T5 phage. Their LTFs (L-shaped tail fibers), however, are composed of two proteins, LtfA and LtfB, instead of the single Ltf of bacteriophage T5. In silico and mutant analysis suggests a possible branched structure of DT57C and DT571/2 LTFs, where the LtfB protein is connected to the phage tail via the LtfA protein and with both proteins carrying receptor recognition domains. Such adhesin arrangement has not been previously recognized in siphoviruses. The LtfA proteins of our phages are found to recognize different host O-antigen types: E. coli O22-like for DT57C phage and E. coli O87 for DT571/2. LtfB proteins are identical in both phages and recognize another host receptor, most probably lipopolysaccharide (LPS) of E. coli O81 type. In these two bacteriophages, LTF function is essential to penetrate the shield of the host's O-antigens. We also demonstrate that LTF-mediated adsorption becomes superfluous when the non-specific cell protection by O-antigen is missing, allowing the phages to bind directly to their common secondary receptor, the outer membrane protein BtuB. The LTF independent adsorption was also demonstrated on an O22-like host mutant missing O-antigen O-acetylation, thus showing the biological value of this O-antigen modification for cell protection against phages