Genomic characterization of phages infecting Stenotrophomonas maltophilia reveals novel lineages

Stenotrophomonas maltophilia is an emerging opportunistic pathogen with high intrinsic antimicrobial resistance, motivating renewed interest in bacteriophages as ecological components of microbial communities, with potential applications in phage therapy. Here, we isolated four lytic phages infecting clinical S. maltophilia strains, recovered from hospital wastewater and generated complete genomes using long-read sequencing. Using comparative genomics, and phylogenomic analyses, we examined the genome architecture, functional gene content, and evolutionary relationships within the 146 currently available Stenotrophomonas phage genomes. The double-stranded DNA phages ranged in size from 56.3 to 74.8 kb and encoded 79–94 proteins, with no detectable virulence or antimicrobial resistance genes. Comparative analyses showed that all isolated phages represent novel species and genera. Three phages clustered within a divergent clade of the family Schitoviridae , whereas one phage was more closely related to the Casjensviridae family members. Host range assays revealed variable infectivity patterns across clinical isolates, indicating host-dependent infection dynamics. Additionally, the phages encoded diverse anti-defense systems that were unevenly distributed among lineages and reflected their evolutionary history. These results expand the genomic and taxonomic landscape of S. maltophilia bacteriophages and highlight lineage-specific evolutionary strategies shaped by host immunity and environmental pressure.