Molecular Characterization of virulence factors of Brucella abortus

DOHMER P.; CZIBENER C; UGALDE JE.

Buenos Aires

Congreso; Brucellosis 2011-International Research Conference; 2011

Asociación Argentina de Microbiología

Virulence of Brucella depends on the capacity to survive within the host cells. To achieve this the bacterium has developed strategies to escape recognition by the innate immune system and, specifically, to avoid killing within phagocytic cells. Although much has been done on the mechanisms that allow the bacterium to survive within the host cell, few bonafide virulence factors have been identified to date. The aim of this work was to study and characterize new virulence factors involved in the intracellular survival or replication within host cells. Applying a bioinformatic approach we searched the Brucella genome for genomic regions with transposons or recombinases close to transfer RNA genes indicative of possible horizontal transfer with the premise that horizontally transmitted regions might contain putative novel virulence factors. With this approach we identified several gene candidates that were flag-tagged to determine protein localization in infected cells. One of them, Bab1_1492, with unknown function and no conserved domains, is secreted from the bacteria during infection and, most probably, localizes to the Brucella containing vacuole. To gain insight into the probable function of this gene we generated a deletion mutant and determined its intracellular replication capacity in phagocytic cells. The results showed that the mutant has a significant defect in the intracellular replication rate specially during the first hours of infection. To further characterize this gene we have transfected HeLa cells with a Bab1_1492-GFP fusion to determine protein localization and generated an amino-terminal truncated version of the protein and assayed secretion of it in Brucella. Our results indicate that the fusion protein localizes to the endoplasmic reticulum.