Growth characteristics of the FTL_0699::Tn5 and FTL_0724::Tn5 mutants. (A) LPS-primed BMMϕ were infected with LVS, the FTL_0699::Tn5 mutant, or the FTL_0724::Tn5 mutant (50:1 MOI); 1 h later, BMMϕ monolayers were washed to remove extracellular bacteria. At the indicated times postinfection, BMMϕ were lysed with 0.5% saponin and plated on bacterial media to determine numbers of CFU/well. (B and C) F. tularensis LVS and the FTL_0699::Tn5 and FTL_0724::Tn5 mutants were grown in Chamberlain's defined media (CDM; pH 6.8) (B) or modified Mueller-Hinton (MMH) broth at pH 4.5, 5.2, or 6.8 (C) at 37°C; at the indicated times, growth was determined by measuring absorbance at 600 nm. Data are representative of the results of three (A) or two (B and C) independent experiments. Determinations were performed in triplicate, and the results are expressed as means ± SD. *, P ≤ 0.05 for FTL_0724::Tn5 compared to LVS; #, P ≤ 0.05 for FTL_0699::Tn5 compared to LVS.