Analysis of a Novel T1-like Phage KanT1 Reveals a Standalone SH3 Domain as a Widespread Component of Drexlerviridae Cell Lysis Module.

Bacteriophages are ubiquitous biological entities that profoundly influence microbiology research and biotechnology. Among coliphages, T1-like viruses (family Drexlerviridae) are notoriously known for their environmental stability and propensity to contaminate laboratory cultures and equipment. Despite this, the genomic features that may underlie their persistence and recurrent detection as laboratory contaminants remain insufficiently characterized. Here, we describe a novel T1-like bacteriophage, KanT1, identified as a recurrent contaminant emerging from environmental samples. Comparative genomics and phylogenetic analyses position KanT1 within the Tunavirus lineage, confirming its close relationship to canonical T1-like phages. Structure-informed annotation enabled the functional characterization of previously unannotated proteins, highlighting the importance of integrating structural predictions into phage genome analysis. Notably, we provide novel details regarding the distribution of superinfection exclusion cassette cor and identify an SH3 domain-containing protein associated with the lysis cassette. We show that SH3 is widespread, though non-universal, across Drexlerviridae genomes. Given the established role of SH3 domains as determinants of cell-wall binding specificity for endolysins of phages infecting Gram-positive bacteria, we propose that this protein represents an auxiliary component of the T1-like lysis module. Together, these findings expand the current understanding of T1-like phage genome organization and provide new insights into molecular features that may contribute to their broad host range and persistence in laboratory environments.