, and "Mycobacterium tuberculosis subsp. canettii" ("M. canettii") (7, 29) (the name "M. canettii" is in quotation marks since it does not appear on the official List of Bacterial Names with Standing in Nomenclature [http://www.bacterio.cict.fr]). Members of the MtbC are highly related mycobacteria exhibiting remarkable nucleotide sequence level homogeneity despite varying in pathogenicity, geographic range, certain physiological features (such as colony morphology as well as profiles of resistance and susceptibility to inhibitors), epidemiology, and host preference (10,11,39). Notably, M. bovis has a wide host range but is primarily a bovid pathogen, goats are the natural host of M.caprae, and M. microti is most often isolated from small rodents, while M. tuberculosis is the predominant cause of human tuberculosis (2,3,39,44,45). However, each of the MtbC subspecies is known to infect humans (16,17,20,22,27,33,(43)(44)(45), and since most laboratories do not fully identify MtbC isolates, the true cause of tuberculosis in these patients and its source often remain undiscovered. An important health concern is the zoonotic transmission of some MtbC subspecies from animals to humans and vice versa. Of particular significance is the transmission of M. bovis to humans from cattle and unpasteurized milk as well as M. bovis BCG infection of immunocompromised individuals (1,21,27,33). M. bovis is naturally resistant to pyrazidamide, a first-line antituberculosis drug (31,36). Therefore, complete identification of MtbC isolates at the subspecies level is required in order to collect information on their epidemiology and also to enable appropriate patient treatment and public health measures.Various biological and molecular mycobacterial characteristics have been utilized to identify MtbC isolates but have limited applicability as MtbC taxonomical tools. Although certain Mycobacterium species-specific gene sequence differences work well to differentiate mycobacteria other than MtbC (MOTT) from each other and from the MtbC, to date none can discriminate the individual MtbC subspecies due to genetic invariance in the target loci (8,11,26,28,34,37,39,41). In contrast, a