Culturing and Characterizing Burkholderia mallei

Strains and plasmids are described in Table 1. Burkholderia mallei was cultured at 37°C using Brucella medium (BD) supplemented with 5% (vol/vol) glycerol. When indicated, antibiotics were added at the following concentrations: 7.5 μg/mL Polymyxin B (MP Biomedicals), 5 μg/mL kanamycin (MP Biomedicals), 7.5 μg/mL zeocin (Life Technologies). Burkholderia mallei bacteria used to inoculate mice were cultured on agar plates and suspended in PBS to the indicated concentration, as previously reported [41 (link)]. Aliquots of the bacterial suspension were immediately spread onto agar plates to determine the number of colony forming units (CFU) in the inoculum. Escherichia coli was cultured at 37°C using Low Salt Luria Bertani (LSLB) agar (Teknova) supplemented with 15 μg/mL chloramphenicol, 50 μg/mL kanamycin, 100 μg/mL ampicillin (Sigma-Aldrich), or 50 μg/mL zeocin, where applicable. The cell lines A549 (human type II alveolar epithelium; ATCC CCL85) and J774A.1 (murine macrophages; ATCC TIB-67) were cultured as described elsewhere [14 (link), 38 (link)].