Abstract / Summary
Mycobacteriophages are being developed as therapeutics for mycobacterial infections, but their highly variable host ranges limit this application and the bacterial factors that determine whether infection succeeds remain poorly defined. The determinants characterized so far are dominated by cell envelope lipids and glycolipids. Here, we performed a genome-wide CRISPRi screen in Mycobacterium smegmatis to identify host functions associated with susceptibility to BPsΔ33HTH HRM10. After prolonged phage challenge, the outer membrane porin genes mspA and mspC were among the most strongly enriched hits, whereas the closely related paralogs mspB and mspD were not enriched despite sharing high amino-acid identity across the family. Deletion of mspA or mspC reduced BPsΔ33HTH HRM10 plaque formation and produced smaller, more turbid plaques, with the stronger phenotype following loss of mspA, the major outer membrane porin of M. smegmatis. Across a panel of 25 mycobacteriophages, 11 showed either a more than tenfold reduction in titer or loss of discrete plaque formation on ΔmspA, while the remainder were largely unaffected. These findings identify outer membrane porins as phage-dependent determinants of mycobacterial susceptibility, revealing an additional layer of specificity in phage infection beyond previously characterized cell envelope lipids and glycolipids.