FIGURE 1.

Growth properties of Sterne and ΔNOS strains (A,D) and accumulation of nitrate/nitrite in cultures (B,C). (A) The growth was measures as OD₆₀₀ at 37°C and shaking at 300 r.p.m. after inoculation of Luria broth (LB) medium with equal amounts of bacteria from overnight cultures. (B) Nitrate and nitrite were measured using a colorimetric Griess test (Life Technologies) in the sups of cultures grown in 12-well plates upon inoculation of 3 × 10⁶ spores per ml of DMEM/F12 medium supplemented with 1 mg/ml Bovine serum albumin (BSA), with shaking in at 5% CO₂, 37°C for 18 h. The background values corresponding to the control wells with medium were subtracted from those with sups. The OD₆₀₀ of the Sterne vs. ΔNOS cultures measured in 200 μl volumes using 96-well plate were 0.48 vs. 0.36 (without BSA) and 1.07 vs. 0.77 (with BSA), respectively. No substantial acidification of the medium was detected, demonstrating the absence of acidic fermentation. (C) The appearance of nitrite as a result respiration on nitrate in the Complete Serum-Free Medium (CSFM) culture medium containing 300 μM of nitrate was measured using Griess test. Cultures were grown at 37°C and shaking at 300 r.p.m. for 17 h upon inoculation of 3 × 10⁶ spores per ml of CSFM. (D) The growth of static cultures at different glucose concentrations in the DMEM medium at 5% CO₂, 37°C for 24 h. The final pH values of the sups are shown below the corresponding bars. The error bars in all panels indicate 95% confidence intervals of triplicate measurements.