Figure 3.

ArsR2 response to copper toxicity. (A) Growth of Brucella and its derivatives in TSB medium supplemented with copper or ferric ions. Bacteria were grown in TSB to OD₆₀₀ 0.6. Bacteria were diluted 100-fold in TSB at 37°C with continuous shaking for 72 h. OD₆₀₀ was measured at different time points. (B) Ten-fold serial dilutions of Brucella and derivatives were grown to OD₆₀₀ 0.6 and spotted on TSA containing 2 mM copper or 1 mM ferric ion. (C) Intracellular replication of Brucella and deletion derivatives. Data represent mean and standard deviation of N = 3 (independent biological replicates). *<0.05 by one-way analysis of variation followed by Tukey’s multiple-comparison test. (D) Representative confocal micrographs of rBCVs formation in RAW264.7 infected with Brucella or deletion derivatives (red) for 12 h and labelled for the LAMP1 marker protein (green). eBCVs is LAMP1-positive and rBCVs is LAMP1-negative. Results are representative of at least three independent experiments. Scale bar, 10 µm. (E) Bacterial intracellular survival was detected at 24 h post-infection supplemented with 200 μM copper. Data represent mean and standard deviation. *<0.05 by unpaired two-tailed t tests with Welch’s correction.