F. tularensis LVS FTL_0699::Tn5 and FTL_0724::Tn5 transposon mutants modulated Francisella-induced IL-1β and IL-18 secretion and cytotoxicity. (A to C) LPS-primed BMMϕ were infected with wild-type F. tularensis LVS or the indicated transposon mutant at an MOI of 50:1. Supernatants and cell lysates were collected at the indicated times postchallenge. Secretion of IL-1β (A) and IL-18 (B) into the supernatants was assessed by ELISA. (C) Cytotoxicity was assessed by measuring LDH release into the supernatant and expressed as a percentage of LDH release by Triton X-100 detergent. Determinations were performed in triplicate, and data are expressed as means ± standard deviations (SD). Data are representative of the results of three independent experiments. *, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001 (for FTL_0724::Tn5 compared to LVS); #, P ≤ 0.05; ##, P ≤ 0.01; ###, P ≤ 0.001 (for FTL_0699::Tn5 compared to LVS).