Caracterização genotípica de isolados de Mycobacterium bovis provenientes de bovídeos leiteiros diagnosticados com tuberculose clínica

Abstract

Interest in molecular methods for diagnosis of bovine tuberculosis plays a relevant role in the epidemiological knowledge of this disease as they allow the comparison of mycobacterial genome sequences, identifying genetic similarities among strains of Mycobacterium bovis (M. bovis) circulating in a herd and/or geographic regions. Bovine tuberculosis caused by M. bovis represents a potential risk to human health due to the zoonotic character related to the consumption of raw milk and unpasteurized dairy products. This study aimed to carry out the genotypic characterization of M. bovis isolated from cattle clinically diagnosed with tuberculosis, which belonged to dairy herds in the state of Pernambuco. They were sent for material microbiological cultivation (granulomas) of 30 bovines. The animals whose M. bovis was isolated presented as clinical findings apathy, inappetence, low body score, seromucous nasal discharge, dry cough, dyspnea, tachypnea, polypnea, crackles and areas of silence in the lung fields. Post mortem findings revealed that the animals had generalized lesions suggestive of tuberculosis distributed mainly in the lungs, mediastinal and tracheobronchial lymph nodes, liver, mesenteric lymph nodes, mammary gland and uterus. M. bovis isolates were submitted to genotypic characterization using Spoligotyping and MIRU-VNTR techniques. Microbiological culture revealed the growth of colonies compatible with Mycobacterium spp. in 17/30 (56.66%) which were confirmed by molecular techniques as M. bovis. Spoligotyping analysis ranked the 17 isolates into five distinct profiles of M. bovis (SB0121), (SB0295), (SB0852), (SB0120) and a spoligotype not yet classified in the database, grouping them into 3 clusters and 2 orphan patterns. The MIRU-VNTR 24-loci analysis grouped the same isolates into two clusters and 13 unique profiles, being more discriminatory compared to spoligotyping. One locus (ETR-A) among the 24 analyzed by MIRU-VNTR showed higher discriminatory power with (h> 0.6) and five loci (ETR-B, ETR-C, MIRU 16, MIRU 27 and QUB 26) displayed moderate allelic diversity with (0.3 ≤ h ≤ 0.6). The discriminatory power of MIRU-VNTR loci-24 was higher than spoligotyping. The combination of the two techniques ratified the best strategy for M. bovis molecular typing, allowing the identification of different genotypes, revealing the high genotypic diversity of circulating M. bovis strains in the state of Pernambuco, corroborating the importance of this microorganism as an agent of tuberculosis and its zoonotic potential, being this epidemiological tool crucial in the rigor of health practices for controlling the disease in dairy herds.