Abstract
Brucella canis is transmitted among dogs and from dogs to humans mainly through contact with vaginal secretions, fetus and placenta from infected animals. In Costa Rica, a diversity of outbreaks related to the disease have occurred in commercial kennels, causing significant economic losses as well as awareness from health and animal authorities. To date, no commercial vaccines are available against canine brucellosis. In this work, we used successive passages in different in vitro culture media as a strategy to induce spontaneous mutations in a virulent strain of B.canis. Subsequently, a genomic-based approach was performed to screen for DNA targets that could be potentially selected to assess attenuation. Distinct strains with different number of passages in in vitro culture media were selected and genetically characterized through whole genome sequencing. Genomic analysis revealed no major genetic or structural rearrangements among passages in comparison to the original strain. In addition, we found 8 different Single Nucleotide Polymorphisms (SNPs) between the original strain and strains with different number of passages as potential targets, located in different genes and in intergenic regions. Non reverting deletion mutants of selected target SNPs were derived from virulent B.canis strain and were tested for attenuation in a murine model. Significant differences in virulence profiles were observed between the deletion mutants and the virulent B.canis parental strain. These results contribute on the searching for potential DNA targets for the development of vaccine candidates for canine brucellosis.
