Estudo do envolvimento da enzima formil-transferase na biossíntese do LPS da Brucella abortus e avaliação da persistência e proteção de uma linhagem mutante para este gene de camundongos C57BL/6 e defifientes para IRF-1

Abstract

Members of genus Brucella are Gram negative, facultative intracellular bacterial pathogens that can cause chronic infections in mammals. Several attempts to get an efficient vaccine have been developed. However, in spite of conferring protection, the vaccines currently commercial available show disadvantages as diagnostic interference and resistance to antibiotics. Considering Brucellas LPS one of its main virulence factors, we disrupted Brucella abortus wbkC gene which is related to LPS biosynthesis pathway. More specifically, wbkC catalyzes the GDP-4-NH2-4,6 dideoximanose conversion in GDP-4-formamido-4,6 dideoximanose, the monomeric unit of antigen-O present in Brucella LPS. Mutants for smooth wild type Brucella abortus S2308 and vaccine strain S19 were generated by double recombination strategy. In vitro analyses were performed in bone marrow macrophages, were we observed alteration in intracellular survival and trafficking of the mutants compared to the parental strains. In vivo experiments in C56BL/6 and IRF-1 (interferon regulation factor-1) knockout mice models were also performed. wbkC mutants had a lower persistence in both mice models, showing the critical role of the selected gene for full bacterial virulence. Challenge experiments revealed that wbkC mutant strains induced lower protective immunity in C56BL/6 when compared to smooth vaccine strain S19, however ÄwbkC 2308 mutant could induce the same level of protection observed for rough vaccine strain RB51.