This cross-sectional study was conducted at the slaughterhouses/slabs of Oudalan and Ouagadougou in Burkina Faso, between August and September 2013. It aimed at determining the prevalence of bovine tuberculosis (bTB) suggestive lesions in slaughtered cattle carcasses and to identify and characterize the mycobacteria isolated from these lesions. A thorough postmortem examination was conducted on carcasses of a total of 2165 randomly selected cattle. The overall prevalence of bTB suggestive lesions was 2.7% (58/2165; 95% CI 2.1–3.5%). Due to the low number of positive samples, data were descriptively presented. The lesions were either observed localized in one or a few organs or generalized (i.e., miliary bTB) in 96.6% (n = 57) and 3.4% (n = 2), respectively. The identified mycobacteria were M. bovis (44.4%, n = 20), M. fortuitum (8.9%, n = 4), M. elephantis (6.7%, n = 3), M. brumae (4.4%, n = 2), M. avium (2.2%, n = 1), M. asiaticum (2.2%, n = 1), M. terrae (2.2%, n = 1), and unknown non-tuberculous mycobacteria (NTM) (11.1%, n = 5). Moreover, eight mixed cultures with more than one Mycobacterium species growing were also observed, of which three were M. bovis and M. fortuitum and three were M. bovis and M. elephantis. In conclusion, M. bovis is the predominant causative agent of mycobacterial infections in the study area. Our study has identified a base to broaden the epidemiological knowledge on zoonotic transmission of mycobacteria in Burkina Faso by future studies investigating further samples from humans and animals, including wild animals employing molecular techniques.