Identification and characterization of virulence factors in Brucella abortus

DOHMER, PETER H.; UGALDE, JUAN E.

Tucuman, Argentina

Congreso; SAIB 45th Annual Meeting; 2009

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular

Brucella abortus is a gram-negative, intracellular bacterium, responsible of brucellosis, a worldwide zoonosis of veterinary and human concern. Virulence of this pathogen is associated with the ability to survive and replicate within professional and nonprofessional phagocytes. With the aim of finding and characterizing virulence factors, we used a bioinformatic approach to search for potential proteins that might play a role in the intracellular survival. The candidates were fused to an antigenic epitope, expressed in B. abortus and its expression and localization analyzed by western-blotting, and indirect mmunofluorescence in infection assays. We found that ORF Bab1_1492, that codes for a hypothetical protein with a molecular weight of 24 Kda, is secreted during the intracellular life cycle and that it localizes to the Brucella containing vacuole. Sequence analysis showed that it has no potential transmembrane domains or a canonical signal peptide. Analysis of the promoter allowed us to identify a potential secondary structure that could play a role in the transcriptional/translational regulation. We cloned the ORF with and without a 250 bp upstream region in a replicative plasmid and analyzed expression in E. coli and B. abortus. Our results suggest that this upstream region may play a role in the transcriptional regulation of this hypothetical protein.