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Marcos Alexandre De ,oliveira - One of the best experts on this subject based on the ideXlab platform.
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Diagnosis of brucellosis in Milk using collective Milk samples
Universidade Federal de Uberlândia, 2014Co-Authors: Marcos Alexandre De ,oliveiraAbstract:A dissertação foi dividida em três estudos. O primeiro objetivou verificar a existência de correlação entre altas contagens de células somáticas e reações positivas no Teste do anel em leite (TAL). Foram coletadas 181 amostras de leite oriundas de tanques de expansão provenientes de diferentes propriedades rurais e submetidas ao Teste de Anel em Leite (TAL). Também foi realizada a contagem de células somáticas (CCS) através do método de citometria de fluxo. O TAL resultou em 11 amostras positivas (6%), da quais, 5 (45,4%) encontravam-se na faixa de 6x105 cel/mL. Os resultados das análises demonstraram não haver associação estatística entre altas contagens de células somáticas e reações positivas no Teste de Anel em Leite. O segundo estudo teve como objetivo avaliar a eficiência de um kit de teste imunoenzimático indireto (ELISA) para diagnóstico de brucelose em amostras coletivas de leite. Amostras de leite in natura, totalizando 181, foram submetidas ao teste ELISA indireto. O teste ELISA identificou 42 (23,2%) amostras positivas, 11 (6%) suspeitas e 128 (70,7%) negativas. Para comparação foi utilizado o TAL. O ELISA apresentou sensibilidade de 72,7%, especificidade de 78,6% e um índice de concordância Kappa de 0,22. Posteriormente 9 rebanhos foram selecionados para realização de testes individuais de Antígeno Acidificado Tamponado (AAT) e 2-Mercaptoetanol 2(ME), por meio de amostragem aleatória dos animais em lactação. Foram encontrados animais reagentes apenas em rebanhos considerados positivos no TAL e no ELISA simultaneamente. Constatou-se que o teste ELISA apresentou valores de sensibilidade e especificidade inferiores aos relatados em outros países onde o teste ELISA já é utilizado, assim como foi confirmada a existência de focos de brucelose em rebanhos leiteiros. No terceiro estudo, a Reação em Cadeia de Polimerase (PCR) foi empregada para detecção de Brucella abortus em amostras de leite in natura. Foram selecionadas 90 amostras para realização da técnica de PCR, sendo 8 positivas no TAL, 37 positivas no ELISA, 11 suspeitas no ELISA e 34 negativas em ambos os testes. A extração foi realizada por meio de lise enzimática, seguida de amplificação utilizando primers BAB e IS 711, que amplificam fragmentos de 498 pares de bases. A visualização foi realizada utilizando gel de agarose 1% em fotodocumentador Alpha Digi Doc. Não foi amplificado DNA de Brucella abortus, demonstrando a PCR não se a melhor técnica para o diagnóstico de brucelose em amostras coletivas de leite.The dissertation was divided in three studies. Our first study determined the correlation between high somatic cell counts and positive Milk Ring Tests (MRT). The Milk Ring test was performed on 181 Milk samples from expansion tanks located at various rural sites in the regions of Brazil. Somatic cell counts (SCC) via flow cytometry were also performed. MRT identified 11 positive samples (6%) where the majority were reactive and 5 (2.8%) were found in the 6x105 cel/mL range. The results showed that there was no statistical relationship between high somatic cell counts and positive Milk Ring Tests. Our second study evaluated the efficiency of an enzyme linked immunosorbent assay kit (ELISA) in diagnosing brucellosis in collective Milk samples. The indirect ELISA test was used on 181 samples of fresh Milk and identified 42 positive, 11 suspect and 128 negative samples. These results were compared to the MR test, the only test approved for collective Milk samples by the Brazilian Ministry of Agriculture, Livestock and Supply (Ministério da Agricultura Pecuária e Abastecimento). The ELISA test had sensitivity of 72.7%, specificity of 78.6% and a Kappa concordance index of 0.22. Subsequently, random samples from lactating animals in nine herds were tested individually by the rose Bengal test (RBT) and the 2-mercaptoethanol 2(ME) Tests. Positive animals were found only in herds that tested positive in both ELISA and MRT. The ELISA test was neither as sensitive nor as specific as reported in other countries where the ELISA test is used and where brucellosis outbreaks have been confirmed in dairy herds. Our third study used the Polymerase Chain Reaction (PCR) technique to detect Brucella abortus in samples of fresh Milk. Ninety samples were preselected for the PCR test including 45 that tested positive in ELISA and/or MRT, 11 suspect in ELISA and 34 negative in both Tests. Extraction was carried out by enzymatic lysis followed by amplification using BAB and IS 711 primers, which amplify fragments of 498 base pairs. Visualization was accomplished with 1% agarose gel and an Alpha Digidoc Photo Documentation system. Brucella abortus DNA was not amplified in any of the samples, demonstrating that PCR is not the best screening technique for brucellosis in Milk samples at the herd level, given that these microorganisms do not always escape into the Milk
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Diagnosis of brucellosis in Milk using collective Milk samples
UFU, 2014Co-Authors: Marcos Alexandre De ,oliveiraAbstract:The dissertation was divided in three studies. Our first study determined the correlation between high somatic cell counts and positive Milk Ring Tests (MRT). The Milk Ring test was performed on 181 Milk samples from expansion tanks located at various rural sites in the regions of Brazil. Somatic cell counts (SCC) via flow cytometry were also performed. MRT identified 11 positive samples (6%) where the majority were reactive and 5 (2.8%) were found in the 6x105 cel/mL range. The results showed that there was no statistical relationship between high somatic cell counts and positive Milk Ring Tests. Our second study evaluated the efficiency of an enzyme linked immunosorbent assay kit (ELISA) in diagnosing brucellosis in collective Milk samples. The indirect ELISA test was used on 181 samples of fresh Milk and identified 42 positive, 11 suspect and 128 negative samples. These results were compared to the MR test, the only test approved for collective Milk samples by the Brazilian Ministry of Agriculture, Livestock and Supply (Ministério da Agricultura Pecuária e Abastecimento). The ELISA test had sensitivity of 72.7%, specificity of 78.6% and a Kappa concordance index of 0.22. Subsequently, random samples from lactating animals in nine herds were tested individually by the rose Bengal test (RBT) and the 2-mercaptoethanol 2(ME) Tests. Positive animals were found only in herds that tested positive in both ELISA and MRT. The ELISA test was neither as sensitive nor as specific as reported in other countries where the ELISA test is used and where brucellosis outbreaks have been confirmed in dairy herds. Our third study used the Polymerase Chain Reaction (PCR) technique to detect Brucella abortus in samples of fresh Milk. Ninety samples were preselected for the PCR test including 45 that tested positive in ELISA and/or MRT, 11 suspect in ELISA and 34 negative in both Tests. Extraction was carried out by enzymatic lysis followed by amplification using BAB and IS 711 primers, which amplify fragments of 498 base pairs. Visualization was accomplished with 1% agarose gel and an Alpha Digidoc Photo Documentation system. Brucella abortus DNA was not amplified in any of the samples, demonstrating that PCR is not the best screening technique for brucellosis in Milk samples at the herd level, given that these microorganisms do not always escape into the Milk.Conselho Nacional de Desenvolvimento Científico e TecnológicoMestre em Ciências VeterináriasA dissertação foi dividida em três estudos. O primeiro objetivou verificar a existência de correlação entre altas contagens de células somáticas e reações positivas no Teste do anel em leite (TAL). Foram coletadas 181 amostras de leite oriundas de tanques de expansão provenientes de diferentes propriedades rurais e submetidas ao Teste de Anel em Leite (TAL). Também foi realizada a contagem de células somáticas (CCS) através do método de citometria de fluxo. O TAL resultou em 11 amostras positivas (6%), da quais, 5 (45,4%) encontravam-se na faixa de 6x105 cel/mL. Os resultados das análises demonstraram não haver associação estatística entre altas contagens de células somáticas e reações positivas no Teste de Anel em Leite. O segundo estudo teve como objetivo avaliar a eficiência de um kit de teste imunoenzimático indireto (ELISA) para diagnóstico de brucelose em amostras coletivas de leite. Amostras de leite in natura, totalizando 181, foram submetidas ao teste ELISA indireto. O teste ELISA identificou 42 (23,2%) amostras positivas, 11 (6%) suspeitas e 128 (70,7%) negativas. Para comparação foi utilizado o TAL. O ELISA apresentou sensibilidade de 72,7%, especificidade de 78,6% e um índice de concordância Kappa de 0,22. Posteriormente 9 rebanhos foram selecionados para realização de testes individuais de Antígeno Acidificado Tamponado (AAT) e 2-Mercaptoetanol 2(ME), por meio de amostragem aleatória dos animais em lactação. Foram encontrados animais reagentes apenas em rebanhos considerados positivos no TAL e no ELISA simultaneamente. Constatou-se que o teste ELISA apresentou valores de sensibilidade e especificidade inferiores aos relatados em outros países onde o teste ELISA já é utilizado, assim como foi confirmada a existência de focos de brucelose em rebanhos leiteiros. No terceiro estudo, a Reação em Cadeia de Polimerase (PCR) foi empregada para detecção de Brucella abortus em amostras de leite in natura. Foram selecionadas 90 amostras para realização da técnica de PCR, sendo 8 positivas no TAL, 37 positivas no ELISA, 11 suspeitas no ELISA e 34 negativas em ambos os testes. A extração foi realizada por meio de lise enzimática, seguida de amplificação utilizando primers BAB e IS 711, que amplificam fragmentos de 498 pares de bases. A visualização foi realizada utilizando gel de agarose 1% em fotodocumentador Alpha Digi Doc. Não foi amplificado DNA de Brucella abortus, demonstrando a PCR não se a melhor técnica para o diagnóstico de brucelose em amostras coletivas de leite
Edmondson, Anita J. - One of the best experts on this subject based on the ideXlab platform.
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Evaluation of North American Antibody Detection Tests for Diagnosis of Brucellosis in Goats
American Society for Microbiology, 1998Co-Authors: Mikolon, Andrea B., Gardner, Ian A., Hietala, Sharon K., Hernandez De Anda, Jorge, Chamizo Pestaña Elpidio, Hennager, Steven G., Edmondson, Anita J.Abstract:The sensitivities and specificities of 17 antibody detection Tests for brucellosis in goats were estimated. Tests evaluated included the U.S. Department of Agriculture (USDA) card test with 8% cell concentration (8%Card), USDA rapid automated presumptive test (RAP), Mexican rose bengal plate Tests with 8 and 3% cell concentrations (8%RB and 3%RB), French rose bengal plate test with 4.5% cell concentration (4.5%RB), USDA standard plate test (SPT), USDA buffered acidified plate agglutination test (BAPA), USDA and Mexican rivanol Tests (URIV and MRIV), USDA standard tube Tests with Brucella abortus and Brucella melitensis antigens (SATA and SATM), serum enzyme-linked immunosorbent assay (ELISA), USDA cold-fixation complement fixation Tests with B. abortus and B. melitensis antigens (CFA and CFM), USDA and Mexican Milk Ring Tests (UBRT and MBRT), and a Milk ELISA. Test sensitivity was evaluated by using two groups of 10 goats experimentally infected with B. melitensis or B. abortus and monitored for 24 weeks. Specificity was evaluated by using 200 brucellosis-free nonvaccinated goats from 10 California herds. The 3%RB was considered a good screening test because of high sensitivity at week 24 postinfection (90%), ease of performance, and low cost. The cold-fixation CFA and CFM had 100% specificity in the field study and were considered appropriate confirmatory Tests. The Milk ELISA was significantly more sensitive (P < 0.05) than the UBRT and significantly more specific (P < 0.05) than the MBRT. The Milk ELISA also had the advantage of objectivity and ease of interpretation
Claude Saegerman - One of the best experts on this subject based on the ideXlab platform.
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The unexpected discovery of Brucella abortus Buck 19 vaccine in goats from Ecuador underlines the importance of biosecurity measures
Tropical Animal Health and Production, 2017Co-Authors: Jorge Ron-román, Dirk Berkvens, Daniela Barzallo-rivadeneira, Alexandra Angulo-cruz, Pablo González-andrade, Elizabeth Minda-aluisa, Washington Benítez-ortíz, Richar Rodríguez-hidalgo, Jef Brandt, Claude SaegermanAbstract:Very few, mostly old, and only preliminary serological studies of brucellosis in goats exist in Ecuador. In order to assess the current epidemiological situation, we performed a cross-sectional serological study in the goat populations of Carchi ( n = 160 animals), Pichincha ( n = 224 animals), and Loja provinces ( n = 2024 animals). Only two positive serological results (RB negative and SAT-EDTA ≥400 IU/ml) were obtained in lactating goats from the same farm in Quito (Pichincha province). Additionally, Milk was sampled from 220 animals in Pichincha province. The present study indicates a low apparent prevalence in Pichincha province and absence in Carchi and Loja provinces. A total of 25 positive Milk Ring Tests (MRT) were obtained in Pichincha province yielding a prevalence of MRT of 11.16%. Subsequent culture was performed on the positive MRT samples. All results were negative, apart from a single sample, obtained from a serologically positive goat in Quito, that was positive for Brucella abortus strain 19 (B19). Several hypotheses are forwarded concerning this unexpected result. The most likely hypothesis is the possible accidental use of a needle, previously used for vaccination of cattle with the said vaccine, for the administration of drug treatment to the goat. This hypothesis underlines the necessity of biosecurity measures to prevent this type of accidents.
Mikolon, Andrea B. - One of the best experts on this subject based on the ideXlab platform.
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Evaluation of North American Antibody Detection Tests for Diagnosis of Brucellosis in Goats
American Society for Microbiology, 1998Co-Authors: Mikolon, Andrea B., Gardner, Ian A., Hietala, Sharon K., Hernandez De Anda, Jorge, Chamizo Pestaña Elpidio, Hennager, Steven G., Edmondson, Anita J.Abstract:The sensitivities and specificities of 17 antibody detection Tests for brucellosis in goats were estimated. Tests evaluated included the U.S. Department of Agriculture (USDA) card test with 8% cell concentration (8%Card), USDA rapid automated presumptive test (RAP), Mexican rose bengal plate Tests with 8 and 3% cell concentrations (8%RB and 3%RB), French rose bengal plate test with 4.5% cell concentration (4.5%RB), USDA standard plate test (SPT), USDA buffered acidified plate agglutination test (BAPA), USDA and Mexican rivanol Tests (URIV and MRIV), USDA standard tube Tests with Brucella abortus and Brucella melitensis antigens (SATA and SATM), serum enzyme-linked immunosorbent assay (ELISA), USDA cold-fixation complement fixation Tests with B. abortus and B. melitensis antigens (CFA and CFM), USDA and Mexican Milk Ring Tests (UBRT and MBRT), and a Milk ELISA. Test sensitivity was evaluated by using two groups of 10 goats experimentally infected with B. melitensis or B. abortus and monitored for 24 weeks. Specificity was evaluated by using 200 brucellosis-free nonvaccinated goats from 10 California herds. The 3%RB was considered a good screening test because of high sensitivity at week 24 postinfection (90%), ease of performance, and low cost. The cold-fixation CFA and CFM had 100% specificity in the field study and were considered appropriate confirmatory Tests. The Milk ELISA was significantly more sensitive (P < 0.05) than the UBRT and significantly more specific (P < 0.05) than the MBRT. The Milk ELISA also had the advantage of objectivity and ease of interpretation
Jorge Ron-román - One of the best experts on this subject based on the ideXlab platform.
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The unexpected discovery of Brucella abortus Buck 19 vaccine in goats from Ecuador underlines the importance of biosecurity measures
Tropical Animal Health and Production, 2017Co-Authors: Jorge Ron-román, Dirk Berkvens, Daniela Barzallo-rivadeneira, Alexandra Angulo-cruz, Pablo González-andrade, Elizabeth Minda-aluisa, Washington Benítez-ortíz, Richar Rodríguez-hidalgo, Jef Brandt, Claude SaegermanAbstract:Very few, mostly old, and only preliminary serological studies of brucellosis in goats exist in Ecuador. In order to assess the current epidemiological situation, we performed a cross-sectional serological study in the goat populations of Carchi ( n = 160 animals), Pichincha ( n = 224 animals), and Loja provinces ( n = 2024 animals). Only two positive serological results (RB negative and SAT-EDTA ≥400 IU/ml) were obtained in lactating goats from the same farm in Quito (Pichincha province). Additionally, Milk was sampled from 220 animals in Pichincha province. The present study indicates a low apparent prevalence in Pichincha province and absence in Carchi and Loja provinces. A total of 25 positive Milk Ring Tests (MRT) were obtained in Pichincha province yielding a prevalence of MRT of 11.16%. Subsequent culture was performed on the positive MRT samples. All results were negative, apart from a single sample, obtained from a serologically positive goat in Quito, that was positive for Brucella abortus strain 19 (B19). Several hypotheses are forwarded concerning this unexpected result. The most likely hypothesis is the possible accidental use of a needle, previously used for vaccination of cattle with the said vaccine, for the administration of drug treatment to the goat. This hypothesis underlines the necessity of biosecurity measures to prevent this type of accidents.