The Multidrug Efflux System AcrABZ-TolC Is Essential for Infection of Salmonella Typhimurium by the Flagellum-Dependent Bacteriophage Chi

Bacteriophages are the most abundant biological entities in the biosphere. Due to their host specificity and ability to kill bacteria rapidly, bacteriophages have many potential health care applications, including therapy against antibiotic-resistant bacteria. Infection by flagellotropic bacteriophages requires a properly rotating bacterial flagellar filament. The flagella-dependent phage chi (chi) infects serovars of the pathogenic enterobacterium Salmonella enterica. However, cell surface receptors and proteins involved in other stages of chi infection have not been discovered to date. We screened a multigene deletion library of S. enterica serovar Typhimurium by spotting mutants on soft agar plates seeded with bacteriophage chi and monitoring their ability to grow and form a swim ring, a characteristic of bacteriophage-resistant motile mutants. Those multigene deletion regions identified to be important for chi infectivity were further investigated by characterizing the phenotypes of corresponding single-gene deletion mutants. In this way, we identified motile mutants with various degrees of resistance to chi. Deletions in individual genes encoding the AcrABZ-TolC multidrug efflux system drastically reduced infection by bacteriophage chi. Furthermore, an acrABtolC triple deletion strain was fully resistant to chi. Infection was severely reduced but not entirely blocked by the deletion of the gene tig, encoding the molecular chaperone trigger factor. Finally, deletion in genes encoding enzymes involved in the synthesis of the antioxidants glutathione (GSH) and uric acid resulted in reduced infectivity. Our findings begin to elucidate poorly understood processes involved in later stages of flagellotropic bacteriophage infection and inform research aimed at the use of bacteriophages to combat antibiotic-resistant bacterial infections. IMPORTANCE Antimicrobial resistance is a large concern in the health care field. With more multidrug-resistant bacterial pathogens emerging, other techniques for eliminating bacterial infections are being explored. Among these is phage therapy, where combinations of specific phages are used to treat infections. Generally, phages utilize cell appendages and surface receptors for the initial attachment to their host. Phages that are flagellotropic are of particular interest because flagella are often important in bacterial virulence, making resistance to attachment of these phages harder to achieve without reducing virulence. This study discovered the importance of a multidrug efflux pump for the infection of Salmonella enterica by a flagellotropic phage. In theory, if a bacterial pathogen develops phage resistance by altering expression of the efflux pump, then the pathogen would simultaneously become more susceptible to the antibiotic substrates of the pump. Thus, coadministering antibiotics and flagellotropic phage may be a particularly potent antibacterial therapy.

Automated summary

The study identified the impact of different gene deletion regions on the infectivity of flagellotropic phage chi, showing that multi-gene deletions affecting the efflux system reduced infection, deletion of the tig gene reduced infection, and deletion of genes involved in antioxidant synthesis reduced infectivity.