Interruption of the cydB focus in Brucella abortus attenuates intracellular survival and virulence in the mouse model of infection

Brucellosis is characterized by abortion in ruminants and a protracted undu lant fever in humans, which often results in severe pathological manifestat ions. Scant information exists about the molecular mechanisms employed by B rucella abortus to combat host defenses or to persist and replicate within host cells. Transposon (Tn5) mutagenesis of B. abortus and the subsequent s creening of mutants for sensitivity to killing in murine macrophages and in the mouse model fed to the identification of mutants which were severely a ttenuated for intracellular survival. One group of mutants was interrupted in cydB, a gene that is part of the cydAB operon encoding cytochrome bd oxi dase, which catalyzes an alternate terminal electron transport step in bact erial respiration. The elevated affinity for molecular oxygen of this enzym e in Escherichia coli has suggested that it is involved in the protection o f sensitive enzymatic activities such as those of hydrogenases and nitrogen ases from damage. B. abortus cydB::Tn5 strains exhibited heightened sensiti vity to the respiratory inhibitors zinc and azide, highly reactive oxygen s pecies such as hydrogen peroxide, low pH, and attenuated virulence in the m ouse model of infection. Virulence was restored by an intact copy of cydAB or by B. abortus genes encoding the oxidative radical-scavenging enzyme Cu/ Zn superoxide dismutase or catalase. These results suggest a bifunctional r ole for the products of the cydAB operon, both in preventing the buildup of oxidative free radicals and in detoxifying the intracellular compartment, thus indicating the importance of these products in preventing intracellula r destruction. Intracellular conditions that favor expression of the cydAB operon are under investigation and may be linked to the acid sensitivity al so observed in this strain.