Comparison between multiplex PCR of the regions of difference (RD) and spoligotyping for identification of Mycobacterium bovis and Mycobacterium bovis BCG strains

Identification of Mycobacterium isolates is important for appropriate treatment of tuberculosis. BCG is difficult to distinguish from other strains of M. bovis and other members of the M. tuberculosis complex by conventional methods. The molecular methods that demonstrate the presence of RD regions, which also play a significant role in the evolutionary process, are very important in rapid diagnosis. We aimed to make this distinction by performing multiplex PCR with 7 RD regions taking into account the evolutionary tree of the Mycobacterium tuberculosis complex. The strains used for this study were randomly selected among the Mycobacterium tuberculosis complex, which includes M. tuberculosis, M. bovis and M. bovis BCG and other strains; isolated during 2007-2017 in the Regional Tuberculosis Laboratory, Çukurova University. These strains were already identified using the biochemical and growth tests. Genomic DNA was obtained by the Mickle apparatus and all strains tested by RDs-based Multiplex PCR based on the presence or absence of the seven RD regions. Spoligotyping was performed to confirm sublineages. The molecular identification by RDs-based Multiplex PCR has high sensitivity and specificity in differentiation between the MTBC subspecies. We conclude that this molecular method allows accurate and rapid identification of the MTBC members including M. bovis and BCG strains at low cost.