Experimental evolution of Vibrio cholerae identifies hypervesiculation as a way to increase motility in the presence of polymyxin B

Abstract

<i>Vibrio cholerae</i> includes strains responsible for the cholera disease and is a natural inhabitant of aquatic environments. <i>V. cholerae</i> possesses a unique polar flagellum essential for motility, adhesion, and biofilm formation. In a previous study, we showed that motility and biofilm formation are altered in the presence of subinhibitory concentrations of polymyxin B in <i>V. cholerae</i> O1 and O139. In this study, we performed an experimental evolution to identify the genes restoring the motility in the presence of a subinhibitory concentration of polymyxin B. Mutations in five genes have been identified in three variants derived from two different parental strains A1552 and MO10: <i>ihfA</i> that encodes a subunit of the integration host factor (IHF), <i>vacJ</i> (<i>mlaA</i>) and <i>mlaF</i>, two genes belonging to the maintenance of the lipid asymmetry (Mla) pathway, <i>dacB</i> that encodes a penicillin-binding protein (PBP4) and involved in cell wall synthesis, and <i>ccmH</i> that encodes a c-type cytochrome maturation protein. We further demonstrated that the variants derived from MO10 containing mutations in <i>vacJ</i>, <i>mlaF</i>, and <i>dacB</i> secrete more and larger membrane vesicles that titer the polymyxin B, which increases the bacterial survival and is expected to limit its impact on the bacterial envelope and participate in the flagellum's retention and motility.