Fig. 2. E. coli expression library contains clones that rescue ts and HOCl-sensitive phenotypes of O395 ΔhslO mutant.

A. E. coli gene expression library from MG1655 ΔhslO mutant strain was constructed in either pET11A or pBR322 plasmids and transformed into the V. cholerae O395 ΔhslO mutant strain. Transformants were cultivated on MacConkey plates for 24 h at 43°C. Two independently identified transformants, clone 5 from the pBR322 library and clone 12 from the pET11a library, were selected for further analysis. Growth of these two strains was tested on MacConkey plates for 24 h at 43°C and compared to O395 wild type, O395 ΔhslO, and O395 ΔhslO expressing the respective empty plasmids.

B. To test the bleach sensitivity of these strains, wild-type O395, O395 ΔhslO, or O395 ΔhslO expressing either clone 5 or clone 12 were cultivated in LB medium until mid-log phase was reached. Cells were washed, resuspended in phosphate buffer, and treated with 10 μM HOCl for 20 min. Cell viability was analyzed by preparing serial dilutions of the cultures and spotting them onto LB plates.

C. Schematic presentation of E. coli genomic sequences with indicated chromosomal positions that were inserted into clone 5 or clone 12.