Nested and multiplex real-time PCR using dual-labeled probes: detecting and discriminating Mycobacterium tuberculosis complex members in cultures and animal tissues.

Members of the Mycobacterium tuberculosis complex (MTC) are causative agents of tuberculosis (TB) in both humans and animals. In the last two decades, the accumulating knowledge of the nucleotide sequences of several genes, and of the whole genomes, of MTC members has allowed the development of novel molecular assays able to detect and discriminate between these species. However, despite the significant advances in the development of molecular assays for detecting MTC members in human samples, only a few assays have been described for detecting these agents in animal tissues. In this chapter we describe the use of two TaqMan (®)-based real-time PCR approaches, highly sensitive and specific and easy to perform, to detect and identify veterinary-relevant MTC species in both animal tissue samples and cultures.