Heterogeneous Flagellar Expression in Single Salmonella Cells Promotes Diversity in Antibiotic Tolerance

Phenotypic heterogeneity among single cells in a genetically identical population leads to diverse environmental adaptation. The human and animal pathogen Salmonella enterica serovar Typhimurium exhibits heterogeneous expression of virulence genes, including flagellar and Salmonella pathogenicity island (SPI) genes. Little is known about how the differential expression of flagellar genes among single cells affects bacterial adaptation to stresses. Here, we have developed a triple-fluorescence reporter to simultaneously monitor the expression of flagellar and SPI-1 pathways. We show that the two pathways cross talk at the single-cell level. Intriguingly, cells expressing flagella (fliC-ON) exhibit decreased tolerance to antibiotics compared to fliC-OFF cells. Such variation depends on TolC-dependent efflux pumps. We further show that fliC-ON cells contain higher intracellular proton concentrations. This suggests that the assembly and rotation of flagella consume the proton motive force and decrease the efflux activity, resulting in antibiotic sensitivity. Such a trade-off between motility and efflux highlights a novel mechanism of antibiotic tolerance.