To the problem of brucellosis diagnostics: development of a primer system for multiplex PCR

Abstract

According to WHO, more than 0.5 million cases of brucellosis registered annually among the population of 73 countries. According to the trend of brucellosis, Kazakhstan is a hyperendemic country. Although bacteriological seeding remains the “gold standard” of laboratory diagnostics, the diagnosis of brucellosis confirmed by the results of seeding only in 15-24% of cases. The effectiveness of immunological diagnostic methods reduced due to insufficient specificity. The problem solved by the introduction of a genetic method – PCR. Aim. The main goal of the work was to improve the system of laboratory diagnostics of brucellosis, detection and identification of microorganisms of the genus Brucella. Material and methods. The used reference and vaccine strains of Brucella from the Museum's collection of live cultures of NSCEDI. The main method used was the method of molecular diagnostics – PCR. Results. Three pairs of primers were obtained flanking fragments in 428 (Br1), 329 (Br2) and 179 (Br3) base pairs. Conclusion. The developed system of primers Br1 + Br2 + Br3 based on the nucleotide sequences of the genes BCAN_B0369, BSPT1_II0384 and BruAb1_0072 makes it possible to differentiate strains to the species B. abortus, B. melitensis and B. suis, and the experimental series of the drug showed high specificity. Keywords: brucellosis, brucella, molecular diagnostics, PCR, primer.