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Mycobacterium bovis is the agent of bovine tuberculosis, a disease endemic to all Brazilian states. Molecular typing techniques help to stratify and refine data, providing information that facilitates epidemiological research. In this study, MIRU-VNTR, targeting 24 loci, was employed to identify and characterize the genetic groups of M. bovis isolates obtained from an outbreak of bovine tuberculosis. Eighteen acid-fast bacilli isolates, obtained from bovine tissue samples, and reactive to the comparative cervical tuberculin test, were identified as species of the M. tuberculosis complex, and were genotyped by MIRU-VNTR with 24 primer pairs. Genotyping revealed three genetic profiles comprising one with 15 isolates (83.3%), one with two isolates (11.1%), and one profile with one unique isolate (5.6%). This distinction was achieved with the MIRU 31 primer, which resulted in clustering of two isolates into the same profile, and ETR A, B, and C, which discriminated the isolate with a unique profile. The occurrence of clustered isolates is indicative of recent transmission, whereas isolates with a unique profile suggest reactivation of latent infection. The presence of different M. bovis genotypes in the same herd suggests movement of infected animals or different sources of intra-herd infection. Use of the MIRU-VNTR molecular epidemiology technique in M. bovis isolates obtained from an outbreak of bovine tuberculosis in Rio Grande do Sul state demonstrated the genetic diversity of circulating strains, despite the presence of a predominant group. Made available in DSpace on 2018-02-22T00:40:19Z (GMT). No. of bitstreams: 1 28GeneticprofilesofMycobacteriumbovisfromacattleherdinsouthernmostBrazil2016.pdf: 346859 bytes, checksum: 715d486ddd5b39e86904192a643f9403 (MD5) Previous issue date: 2018-02-21 Título em português: Perfis genéticos de Mycobacterium bovis em um rebanho bovino no extremo sul do Brasil.

Realizou-se um estudo para caracterizar a situação epidemiológica da brucelose bovina. O Estado foi dividido em sete regiões. Em cada região foram amostradas aleatoriamente cerca de 300 propriedades, e dentro dessas foi escolhido de forma aleatória um número pré-estabelecido de animais, dos quais foi obtida uma amostra de sangue. No total foram amostrados 16.072 animais, provenientes de 1.957 propriedades. Em cada propriedade amostrada foi aplicado um questionário epidemiológico para verificar o tipo de exploração e as práticas zootécnicas e sanitárias que poderiam estar associadas ao risco de infecção pela doença. O protocolo de testes utilizado foi o da triagem com o teste do antígeno acidificado tamponado e o reteste dos positivos com o teste do 2-mercaptoetanol. O rebanho foi considerado positivo se pelo menos um animal foi reagente às duas provas sorológicas. Para o Estado, as prevalências de focos e de animais infectados foram, respectivamente, 2,1% [1,5-2,6%] e 1,0% [0,60-1,4%]. Para os circuitos, a prevalência de focos e a de animais foram, respectivamente: circuito 1, 3,1% [1,4-5,7%] e 0,95% [0,0-2,0%]; circuito 2, 7,7% [4,9-11,3%] e 1,0% [0,40-1,7%]; circuito 3, 5,7% [3,4-8,8%] e 2,1% [0,41-3,8%]; circuito 4, 0,66% [0,08-2,4%] e 0,66% [0,0-1,8%]; circuito 5, 0,66% [0,08-2,4%] e 0,05% [0,0-0,13%]; circuito 6, 0,0% [0,0-1,3%] e 0,0% [0,0-0,25%]; circuito 7, 5,4% [2,5-10,1%] e 2,9% [0,49-5,3%]. Os fatores de risco (odds ratio, OR) associados à condição de foco foram: exploração de corte (OR= 4,27 [1,82-10,01]) e histórico de aborto (OR= 3,27, [1,71-6,25]). A study to characterize the epidemiological status of bovine brucellosis was carried out in the State of Rio Grande do Sul. The State was divided in seven regions. Three hundred herds were randomly sampled in each region and a pre-established number of animals were sampled in each of these herds. A total of 16,072 serum samples from 1,957 herds, were collected. In each herd, it was applied an epidemiological questionnaire focused on herd traits as well as husbandry and sanitary practices that could be associated with the risk of infection. The serum samples were screened for antibodies against Brucella spp. by the Rose-Bengal Test and all positive sera were re-tested by the 2-mercaptoethanol test. The herd was considered positive if at least one animal was positive on both tests. The prevalences of infected herds and animals in the State were, respectively 2.1% [1.5-2.6%] and 1.0% [0.60-1.4%]. In the regions, the prevalences of infected herds and animals were, respectively: region 1, 3.1% [1.4-5.7%] and 0.95% [0.0-2.0%]; region 2, 7.7% [4.9-11.3%] and 1.0% [0.40-1.7%]; region 3, 5.7% [3.4-8.8%] and 2.1% [0.41-3.8%]; region 4, 0.66% [0.08-2.4%] and 0.66% [0.0-1.8%]; region 5, 0.66% [0.08-2.4%] and 0.05% [0.0-0.13%]; region 6, 0.0% [0.0-1.3%] and 0.0% [0.0-0.25%]; and region 7, 5.4% [2.5-10.1%] and 2.9% [0.49-5.3%]. The risk factors (odds ratio, OR) associated with the presence of infection were: beef herd (OR= 4.27 [1.82-10.01]) and recent history of abortion (OR= 3.27-1.71-6.25]).

One-hundred eight Mycobacterium avium isolates from pigs, humans, birds, and bovines were typed by the IS1245-based restriction fragment length polymorphism (RFLP) method and PCR-restriction enzyme analysis (PRA) of hsp65. Nine clusters of isolates showing more than 80% similarity in their RFLP profiles were detected. The largest cluster (cluster B) included 32 of 79 pig isolates (40.5%), 3 of 25 human isolates (12%), and 1 of 2 bovine isolates, comprising 33% of all isolates. The second largest cluster (cluster A) included 18 pig isolates (22.8%) and 6 human isolates (24%). Six smaller clusters included six pig isolates (clusters C and D), four and two human isolates (clusters E and F, respectively), two pig isolates (cluster I), and two pig isolates plus one bovine isolate and the avian purified protein derivative strain (cluster H). Cluster G represented the "bird-type" profile and included the bird isolate in this series, one pig isolate, plus reference strain R13. PRA revealed four allelic variants. Seventy-seven isolates were identified as M. avium PRA variant I, 24 were identified as M. avium PRA variant II, 6 were identified as M. avium PRA variant III, and 1 was identified as M. avium PRA variant IV. Except for three isolates from cluster B, each of the RFLP clusters was associated with a single PRA pattern. Isolates with unique (nonclustered) RFLP profiles were distributed between PRA variants I and II, and there was one unique isolate of PRA variant IV. These observations are consistent with divergent evolution within M. avium, resulting in the emergence of distinct lineages with particular competence to infect animals and humans.

The objective of the present study was to improve the detection of B. abortus by PCR in organs of aborted fetuses from infected cows, an important mechanism to find infected herds on the eradication phase of the program. So, different DNA extraction protocols were compared, focusing the PCR detection of B. abortus in clinical samples collected from aborted fetuses or calves born from cows challenged with the 2308 B. abortus strain. Therefore, two gold standard groups were built based on classical bacteriology, formed from: 32 lungs (17 positives), 26 spleens (11 positives), 23 livers (8 positives) and 22 bronchial lymph nodes (7 positives). All samples were submitted to three DNA extraction protocols, followed by the same amplification process with the primers B4 and B5. From the accumulated results for organ, the proportion of positives for the lungs was higher than the livers (p=0.04) or bronchial lymph nodes (p=0.004) and equal to the spleens (p=0.18). From the accumulated results for DNA extraction protocol, the proportion of positives for the Boom protocol was bigger than the PK (pO objetivo do presente estudo foi aperfeiçoar a detecção de Brucella abortus pela PCR em homogeneizados de órgãos de fetos abortados por vacas infectadas, importante mecanismo para descobrir focos da doença na fase de erradicação. Assim, foram comparados diferentes protocolos de extração de DNA, visando à detecção de B. abortus pela PCR em amostras clínicas colhidas de abortos ou de bezerros oriundos de vacas desafiadas com a estirpe 2308 de B. abortus. Para tanto, foram construídos dois grupos padrão ouro com base na bacteriologia clássica, constituídos por: 32 pulmões (17 positivos), 26 baços (11 positivos), 23 fígados (8 positivos) e 22 linfonodos bronquiais (7 positivos). Todas essas amostras foram submetidas a três protocolos de extração de DNA, seguidos do mesmo processo de amplificação com os primers B4 e B5. Nos resultados acumulados por órgão, a proporção de positivos nos pulmões foi maior que a encontrada nos fígados (p=0,04) e nos linfonodos bronquiais (p=0,004), e igual a verificada nos baços (p=0,18). Nos resultados acumulados por método de extração de DNA, a proporção de positivos para o protocolo de Boom foi maior que a verificada para o PK (p