FIG 2 .

ΔctpA and ΔlbcA mutants share common phenotypes. (A) T3SS function. Wild-type or ΔctpA or ΔlbcA mutant strains contained the tac promoter expression plasmid pVLT35 (vector) or derivatives encoding CtpA, CtpA-S302A, or LbcA as indicated. TSB growth medium contained 5 mM EGTA to induce T3SS activity and 10 µM IPTG to induce the tac promoter of the complementing plasmids. Cell-free supernatants derived from equivalent amounts of cells were separated on an SDS-PAGE gel, which was stained with silver. The region with the abundant T3SS effectors ExoT and ExoS is shown. (B) Surface attachment. The same strains as in panel A were incubated in glass test tubes at 30°C without agitation for 8 h. The growth medium contained 10 µM IPTG to induce the tac promoter of the complementing plasmids. Bacterial cells attached to the glass at the air-liquid interface were stained with crystal violet (purple rings). The stain was dissolved in ethanol and quantified by measuring absorbance at 595 nm. All strains were tested in triplicate, and error bars represent the standard deviation from the mean.