Dps-dependent in vivo mutation enhances long-term host adaptation in Vibrio cholerae
Fig 1
V. cholerae produces spontaneous nonmotility-related mutations during long-term colonization in adult mice.
(A) Different morphological V. cholerae were found in the intestine of adult mice. We collected the fecal pellets of adult mice at the fifth day post-infection and plated on plates. Unexpectedly found the small, rugose, opacity and dense colony (red arrow), which was different morphology from wild-type V. cholerae large, smooth and transparent colony (black arrow). (B) Adult mice competition assay of small colony variants. We pick several small colony variants from different individual mouse and mixed as a sample, 108 cells of wild-type and small colony variants were mixed in a 1:1 ratio and intragastrically administered to CD-1 adult mice. Fecal pellets were collected from each mouse at the fifth day after infection and plated on selective plates. The competitive index (CI) was calculated as the ratio of small colony variants to wild-type colonies normalized to the input ratio. Horizontal line: median CI. (C) Electron micrographs of small colony variants. Bacteria were harvested in mid-logarithmic growth and prepared for electron microscopy. Bars represent 1 μm, C6706, flagellum positive control. ΔflaA, flagellum negative control. Mutant-1-4, small colony variants. (D) Motility phenotype of small colony variants. Bacteria were inoculated into 0.3% agar LB plates and incubated at 37°C for 8 h. C6706, motility positive control. ΔflaA, motility negative control. Mutant-1-4, small colony variants. (E) Rate of nonmotile V. cholerae in vitro and in vivo culture. Wild-type C6706 were inoculated in 5 mL LB anaerobic test tubes and incubated anaerobically at 37°C for 5 days, and plated onto selective plates (in vitro culture). Bacteria were intragastrically administered to CD-1 adult mice, fecal pellets were collected from each mouse at the fifth day after infection and plated onto selective plates (in vivo culture). At the indicated time points, one hundred V. cholerae colonies per sample were picked and the motility was detected in 0.3% agar LB plates. Rate of nonmotile colonies were calculated as the ratio of nonmotile mutant colonies to all colonies per sample. Horizontal line: median. The illustration was created with BioRender.com.