A first insight into genetic diversity of Mycobacterium bovis isolated from extrapulmonary tuberculosis patients in South Tunisia assessed by spoligotyping and MIRU VNTR

Abstract

In Tunisia, almost 77% of clinically and bacteriologically diagnosed cases of extrapulmonary tuberculosis (EPTB) are zoonotic TB, caused by M. bovis. Although several studies have analyzed bovine TB in cattle in Tunisia, no study has evaluated the risk of transmission to humans in such an endemic country. We aimed to study the genetic diversity of M. bovis human isolates, to ascertain the causes of human EPTB infection by M. bovis and to investigate the distribution and population structure of this species in Tunisia. A total of 110 M. bovis isolates taken from patients with confirmed EPTB were characterized by spoligotyping and MIRU-VNTR typing methods. Among the 15 spoligotypes detected in our study, 6 (SB0120, SB0121, SB2025, SB1200, SB1003 and SB0134) were the most prevalent (83.5%) of which SB0120, SB0121 and SB2025 were the most prevailing. MIRU-VNTR typing method showed a high genotypic and genetic diversity. The genetic differentiation based on MIRU-VNTR was significant between populations from South East (Tataouine, Medenine) and Central West (Gafsa, Sidi Bouzid, Kasserine) regions. Of note, 13/15 (86.7%) spoligotypes detected in our study were previously identified in cattle in Tunisia with different frequencies suggesting a peculiar ability of some genotypes to infect humans. Using combined spoligotyping and MIRU-VNTR method, a high clustering rate of 43.9% was obtained. Our results underlined that human EPTB due to M. bovis was more commonly found in female gender and in young patients. Most of our patients, 66.4% (73/110) were raw milk or derivatives consumers, whereas 30.9% (34/110) patients would have contracted EPTB through contact with livestock. The findings suggest that the transmission of Zoonotic TB caused by M. bovis to humans mainly occurred by oral route through raw milk or derivatives. Our study showed the urgent need of a better veterinary control with the implementation of effective and comprehensive strategies in order to reach a good protection of animals as well as human health. In South Tunisia, the prevalence of bovine TB is high with Mycobacterium bovis as causative agent and cattle as reservoir of the bacteria. However as previously mentioned in several studies, M. bovis is also responsible for human extrapulmonary tuberculosis (EPTB) cases in South Tunisia. Despite the veterinary and medical problems, M. bovis is still little studied. In this context, this work aimed to study the molecular epidemiology of M. bovis in EPTB patients in south Tunisia in order to determine the main risk factors of transmission. Our results underlined that SB0120, SB0121 and SB2025, previously described in cattle in Tunisia, represent the predominant genotypes. The findings highlighted that human EPTB caused by M. bovis mainly occurred through the consumption of raw milk or derivatives. These data demonstrate the urgent need to implement strategies for preventing and controlling zoonotic TB.