BRUCELLA-ABORTUS DEFICIENT IN COPPER-ZINC SUPEROXIDE-DISMUTASE IS VIRULENT IN BALB/C MICE

The gene encoding the Cu Zn superoxide dismutase (SOD) of Brucella abortus strain 2308 was identified in a Brucella genomic library utilizing a combination of Western blotting and native gel electrophoresis. The Cu Zn SOD gene was inactivated in vitro by ligation of a kanamycin resistance gene into the open reading frame encoding SOD. The plasmid born construct was introduced back into B. abortus by electroporation. Replacement of the wild-type Cu Zn SOD by recombination was demonstrated by showing that both the KnR gene and the Cu Zn SOD gene hybridized to the same band in a Southern analysis of genomic DNA. In addition, KnR strains were deficient in Cu Zn SOD activity as assessed by lack of Cu Zn SOD activity on a native gel and by lack of reactivity with specific serum in a Western analysis. Either strain 2308 or the Cu Zn SOD deficient mutant injected intraperitoneally into BALB/c mice, exhibited no differences in their ability to colonize the spleen at 7 and 28 days post-inoculation. Thus, the inability to produce Cu Zn SOD by B. abortus does not significantly impair its virulence in mice. © 1992.