Molecular Typing of Human and Animal Brucella Isolates from Georgia

Brucellosis, an ancient worldwide zoonosis and significant economic and public health problem in Georgia, is caused by Brucella, a genus with Category B agents because of their high infectivity in mammals. Biotyping Brucella by microbiological methods alone has limitations, so molecular typing was implemented in this study to confirm species. Isolates from human blood and ruminant milk or blood were identified by a bacteriological algorithm and confirmed by real-time PCR (Brucella T1, Idaho Technology). Isolates were then typed using AMOS PCR, which differentiates four human pathogenic species but cannot recognize certain Brucella biovars. This gap was addressed by using more universal species-specific Single Nucleotide Polymorphism (SNP) assays. 86 strains (48 human, 38 animal isolates) obtained 2009-2011 were confirmed as Brucella by real-time PCR. AMOS PCR supported biochemical test results for 53 Brucella melitensis and four Brucella abortus strains, but not for 29 suspected B. abortus human and animal isolates. SNP typing of all 86 isolates supported the AMOS PCR results but also confirmed the species of the 29 strains not amplified by AMOS PCR. Combined AMOS PCR and SNP typing in this study provided the first genetic confirmation that both B. abortus and B. melitensis are actively circulating in humans and animals in Georgia. Since these SNP assays successfully determined species of Georgian Brucella isolates, this approach should be incorporated for this purpose in future surveillance. Methods