FIG 1.

Contribution of Ca²⁺ to virulence gene expression in V. cholerae. (A) tcpA expression in the presence of taurocholate (TC) and different mental ions. V. cholerae containing P_tcpA-luxCDABE plasmids were grown in LB medium containing 10 mM ion without or with 0.1 mM TC and grown microaerobically until reaching mid-log phase (OD₆₀₀ ∼0.2). The luminescence was measured and normalized for growth against the OD₆₀₀. (B) Bile salt effects. The tcpA expression was measured for cultures grown in 0.1 mM TC, 0.1 mM glycocholate (GC), and 0.1 mM deoxycholate (DC) in the presence of indicated calcium concentration. (C) TcpA production. Wild-type strains were grown in the absence or in the presence of 0.1 mM TC and 10 mM CaCl₂ microaerobically at 37°C until reaching mid-log phase. The lysed cells were normalized by protein concentration (200 μg/sample) and subjected to SDS-PAGE and immunoblotting with anti-TcpA antiserum. (D) Intestinal extract. tcpA expression was measured without intestinal extract, with intestinal extract, or with intestinal extract previously incubated with EGTA. Intestinal extracts were incubated for 12 h with or without EGTA and added to cultures at a final EGTA concentration of 0.25 mM. The data shown are from three independent experiments. *, P < 0.05 (Student t test).