Resultado da pesquisa (7)
Termo utilizado na pesquisa complement fixation
#1 - Pathology, microbiology, and molecular evaluation of tissues from equids serologically positive for Burkholderia mallei in Midwestern Brazil
Abstract in English:
Glanders is a disease caused by the bacterium Burkholderia mallei that primarily affects horses, mules and donkeys. The disease can cause lesions in the skin, lungs and several other organs. However, it often manifests as an asymptomatic disease. In Brazil, serological tests of high sensitivity and specificity are used to assist in the detection of antibodies against B. mallei and to contribute to the control of the disease. However, due to the mandatory euthanasia of seroreactive animals, equids with positive serology for B. mallei and asymptomatic generated great conflicts between breeders, veterinarians and diagnostic laboratories. This study clarifies the limitations of complementary diagnostic tests for detecting B. mallei. It describes the clinical, morphological and laboratory findings in 24 equines from different municipalities in the Mato Grosso State, Brazil, which reacted to the complement fixation test and were positive in the western blotting test for glanders. Data and tissue samples were collected from 24 horses for histological, microbiological and molecular analysis. In 23 horses, no clinical signs, morphological alterations, microbiological isolation, or molecular detection would characterize B. mallei infection. On the other hand, samples from an asymptomatic horse without lesional alterations showed sequence amplification compatible with B. mallei in the PCR. Considering that the infection by B. mallei is subject to the application of animal sanitary defense measures and that, by international requirement and national legislation, the serological results are tools that should support the sanitation procedures for the error of the bacteria in the Mato Grosso State, Brazil.
Abstract in Portuguese:
Mormo é uma enfermidade causada pela bactéria Burkholderia mallei que acomete primariamente cavalos, mulas e burros. A doença pode causar lesões na pele, pulmões e em diversos outros órgãos, entretanto frequentemente manifesta-se como uma enfermidade assintomática. No Brasil são utilizados testes sorológicos de elevada sensibilidade e especificidade para auxiliar na detecção de anticorpos contra B. mallei e contribuir para controle da doença. Porém, devido à obrigatoriedade da eutanásia de animais sororeagentes, os equídeos com sorologia positiva para B. mallei e assintomáticos geraram grandes embates entre criadores, médicos-veterinários e laboratórios de diagnóstico. Este trabalho esclarece as limitações dos testes diagnósticos complementares para detecção de B. mallei e descreve os achados clínicos, morfológicos e de exames laboratoriais em 24 equídeos, procedentes de diferentes municípios do estado de Mato Grosso, Brasil, que reagiram ao teste de fixação de complemento e foram positivos no teste de “western blotting” para mormo. Foram colhidos dados e amostras de tecidos de 24 equídeos para análise histológica, microbiológica e molecular. Em 23 equídeos não existiam sinais clínicos, alterações morfológicas, isolamento microbiológico ou detecção molecular que caracterizassem infecção por B. mallei. Por outro lado, amostras de um cavalo assintomático e sem alterações lesionais apresentaram amplificação de sequência compatível com B. mallei na PCR. Considerando que a infecção por B. mallei é passível da aplicação de medidas de defesa sanitária animal e que por exigência internacional e da legislação nacional, os resultados sorológicos são ferramentas que devem amparar os procedimentos de saneamento para erradicação da bactéria no estado de Mato Grosso, Brasil.
#2 - Ocorrência e fatores de risco para Chlamydophila abortus em ovinos e caprinos no estado de Pernambuco, p.33-40
Abstract in English:
ABSTRACT.- Pereira M.F., Peixoto R.M., Piatti R.M., Medeiros E.S., Mota I.O., Azevedo S.S. & Mota R.A. 2009. [Occurrence and risk factors for Chlamydophila abortus infection in sheep and goats in Pernambuco.] Ocorrência e fatores de risco para Chlamydophila abortus em ovinos e caprinos no estado de Pernambuco. Pesquisa Veterinária Brasileira 29(1):33-40. Departamento de Medicina Veterinária, Universidade Federal Rural de Pernambuco, Rua Dom Manoel de Medeiros s/n, Recife, PE 52.171-900, Brazil. E-mail: marcia.pereira@dmv.ufrpe.br
The study aimed to identify risk factors associated with Chlamydophila abortus infection in sheep and goats from the Litoral/Zona da Mata and Agreste region of Pernambuco state, Brazil. Serum samples (n=290) were analyzed to detect Chlamydophila spp. antibodies in 12 farms. Questionnaires were applied to identify risk factors. Frequency of serum-reactive animals were 10.3% (12.0% in ewes and 8.1% in goats) and 1/12 (91.6%) infection focuses were identified. This is the first report on anti-Chlamydophhila abortus antibodies in goats and sheep in Pernambuco and Brazil, respectively. Risk factors associated with goat infection were breed (OR=9.10) and management (OR=6.41). No significant associations in any of the analyzed factors were found for sheep. In summary, Chlamydophila sp. infection is disseminated in sheep and goat herds in the region. Control measures should be established, focusing primarily risk factor identified in this study, to reduce the possibility of infection by the agent.
Abstract in Portuguese:
ABSTRACT.- Pereira M.F., Peixoto R.M., Piatti R.M., Medeiros E.S., Mota I.O., Azevedo S.S. & Mota R.A. 2009. [Occurrence and risk factors for Chlamydophila abortus infection in sheep and goats in Pernambuco.] Ocorrência e fatores de risco para Chlamydophila abortus em ovinos e caprinos no estado de Pernambuco. Pesquisa Veterinária Brasileira 29(1):33-40. Departamento de Medicina Veterinária, Universidade Federal Rural de Pernambuco, Rua Dom Manoel de Medeiros s/n, Recife, PE 52.171-900, Brazil. E-mail: marcia.pereira@dmv.ufrpe.br
The study aimed to identify risk factors associated with Chlamydophila abortus infection in sheep and goats from the Litoral/Zona da Mata and Agreste region of Pernambuco state, Brazil. Serum samples (n=290) were analyzed to detect Chlamydophila spp. antibodies in 12 farms. Questionnaires were applied to identify risk factors. Frequency of serum-reactive animals were 10.3% (12.0% in ewes and 8.1% in goats) and 1/12 (91.6%) infection focuses were identified. This is the first report on anti-Chlamydophhila abortus antibodies in goats and sheep in Pernambuco and Brazil, respectively. Risk factors associated with goat infection were breed (OR=9.10) and management (OR=6.41). No significant associations in any of the analyzed factors were found for sheep. In summary, Chlamydophila sp. infection is disseminated in sheep and goat herds in the region. Control measures should be established, focusing primarily risk factor identified in this study, to reduce the possibility of infection by the agent.
#3 - Serodiagnosis of Babesia equi in horses submitted to exercise stress, p.179-183
Abstract in English:
ABSTRACT.- Baldani C.D., Machado R.Z., Raso T.F. & Pinto A.A. 2007. Serodiagnosis of Babesia equi in horses submitted to exercise stress. Pesquisa Veterinária Brasileira27(4):179-183. Departamento de Patologia Veterinária, Universidade Estadual Paulista, Faculdade de Ciências Agrárias e Veterinárias, Via de Acesso Prof. Paulo Donato Castellane s/n, Jaboticabal, SP 14870-000, Brazil. E-mail: zacarias@fcav.unesp.br
A complement fixation test (CFT), performed in microtitre plates, based upon the use of crude antigenic preparation of Babesia equi was adapted for the detection of antibodies in serum of infected horses. The indirect fluorescent antibody test (IFAT) and enzyme-linked immunosorbent assay (ELISA) were also used for the immunodiagnosis of B. equi. Serum samples from 15 apparently healthy horses, previously conditioned to a high-speed equine treadmill, were taken before and after exercise. All the samples analyzed were positive for B. equi infection. There were no significant differences (P<0.01) between these 3 tests, or the condition of rest or stress. The combined use of CFT and IFAT or ELISA should be recommended in order to enable veterinary services to more efficiently prevent introduction of infected horses into disease-free areas.
Abstract in Portuguese:
ABSTRACT.- Baldani C.D., Machado R.Z., Raso T.F. & Pinto A.A. 2007. Serodiagnosis of Babesia equi in horses submitted to exercise stress. Pesquisa Veterinária Brasileira27(4):179-183. Departamento de Patologia Veterinária, Universidade Estadual Paulista, Faculdade de Ciências Agrárias e Veterinárias, Via de Acesso Prof. Paulo Donato Castellane s/n, Jaboticabal, SP 14870-000, Brazil. E-mail: zacarias@fcav.unesp.br
A complement fixation test (CFT), performed in microtitre plates, based upon the use of crude antigenic preparation of Babesia equi was adapted for the detection of antibodies in serum of infected horses. The indirect fluorescent antibody test (IFAT) and enzyme-linked immunosorbent assay (ELISA) were also used for the immunodiagnosis of B. equi. Serum samples from 15 apparently healthy horses, previously conditioned to a high-speed equine treadmill, were taken before and after exercise. All the samples analyzed were positive for B. equi infection. There were no significant differences (P<0.01) between these 3 tests, or the condition of rest or stress. The combined use of CFT and IFAT or ELISA should be recommended in order to enable veterinary services to more efficiently prevent introduction of infected horses into disease-free areas.
#4 - Estabilidade do antígeno de célula total de Brucella abortus para uso no diagnóstico sorológico da brucelose bovina pela reação de fixação de complemento, p.18-22
Abstract in English:
ABSTRACT.- Mathias L.A., Meirelles R.B. & Buchala F.G. 2007. [Stability of Brucella abortus whole cell antigen for use in the serological diagnosis of bovine brucellosis by the complement fixation test.] Estabilidade do antígeno de célula total de Brucella abortus para uso no diagnóstico sorológico da brucelose bovina pela reação de fixação de complemento. Pesquisa Veterinária Brasileira 27(1):18-22. Departamento de Medicina Veterinária Preventiva e Reprodução Animal, Faculdade de Ciências Agrárias e Veterinárias (FCAV), Universidade Estadual Paulista (Unesp), Jaboticabal, SP 14884-900, Brazil. E-mail: lmathias@fcav.unesp.br
The complement fixation test is used worldwide in the confirmatory diagnosis of bovine brucellosis. For this technique the antigen is the same as the one used in the tube agglutination test. However, literature is poor in information about the stability of the whole cell Brucella antigen for use in the complement fixation test to establish a time of validity of the antigen. Hence the aim of this investigation was to evaluate the stability of this antigen under refrigeration for use in the complement fixation test. Fourteen batches of antigen prepared with Brucella abortus strain 1119/3, produced from 9 months to 23 years and 11 months before, were analysed. One hundred and sixty-seven cattle sera with varying titres of antibodies to Brucella were tested through the warm complement fixation microtechnique with five 50% haemolytic units of complement. Sera with at least 25% of complement fixation in dilution 1:4 were considered positive. The results with 13 of the antigen batches were compared with the results obtained with the batch produced 9 months before by the McNemar c2 test and kappa statistic. The oldest antigen batch gave a higher proportion of sera titres which were exactly the same observed with the 9-month-batch (90.4%), and the antigen produced 4 years and 3 months before the test gave de lowest proportion of sera with the same titre of the 9-month-antigen (73.7%). The comparison of the results after being classified as positive and negative showed that the highest proportion of agreed results was observed with the antigen produced 21 years and 4 months before (98.8%, kappa 0.98). The antigen with the lowest proportion of agreed results was the one produced 3 years and 2 months before (91.6%, kappa 0.84). The results of the study show that most sera gave very similar results with all antigen batches evaluated, and that there was no relationship between the period of antigen production and the difference in test results.
Abstract in Portuguese:
ABSTRACT.- Mathias L.A., Meirelles R.B. & Buchala F.G. 2007. [Stability of Brucella abortus whole cell antigen for use in the serological diagnosis of bovine brucellosis by the complement fixation test.] Estabilidade do antígeno de célula total de Brucella abortus para uso no diagnóstico sorológico da brucelose bovina pela reação de fixação de complemento. Pesquisa Veterinária Brasileira 27(1):18-22. Departamento de Medicina Veterinária Preventiva e Reprodução Animal, Faculdade de Ciências Agrárias e Veterinárias (FCAV), Universidade Estadual Paulista (Unesp), Jaboticabal, SP 14884-900, Brazil. E-mail: lmathias@fcav.unesp.br
The complement fixation test is used worldwide in the confirmatory diagnosis of bovine brucellosis. For this technique the antigen is the same as the one used in the tube agglutination test. However, literature is poor in information about the stability of the whole cell Brucella antigen for use in the complement fixation test to establish a time of validity of the antigen. Hence the aim of this investigation was to evaluate the stability of this antigen under refrigeration for use in the complement fixation test. Fourteen batches of antigen prepared with Brucella abortus strain 1119/3, produced from 9 months to 23 years and 11 months before, were analysed. One hundred and sixty-seven cattle sera with varying titres of antibodies to Brucella were tested through the warm complement fixation microtechnique with five 50% haemolytic units of complement. Sera with at least 25% of complement fixation in dilution 1:4 were considered positive. The results with 13 of the antigen batches were compared with the results obtained with the batch produced 9 months before by the McNemar c2 test and kappa statistic. The oldest antigen batch gave a higher proportion of sera titres which were exactly the same observed with the 9-month-batch (90.4%), and the antigen produced 4 years and 3 months before the test gave de lowest proportion of sera with the same titre of the 9-month-antigen (73.7%). The comparison of the results after being classified as positive and negative showed that the highest proportion of agreed results was observed with the antigen produced 21 years and 4 months before (98.8%, kappa 0.98). The antigen with the lowest proportion of agreed results was the one produced 3 years and 2 months before (91.6%, kappa 0.84). The results of the study show that most sera gave very similar results with all antigen batches evaluated, and that there was no relationship between the period of antigen production and the difference in test results.
#5 - Evolution of serological antibody titres by the plate agglutination, rose Bengal and complement fixation tests in Nelore heifers (Bos indicus) vaccinated at 18 months of age with Brucella abortus stra
Abstract in English:
ABSTRACT.- Mathias, L.A., Chaves, L.F., Chen, A.A., Girio, R.J.S. & Valério Neto, W. 2001. [Evolution of serological antibody titres by the plate agglutination, rose Bengal and complement fixation tests in Nelore heifers (Bos indicus) vaccinated at 18 months of age with Brucella abortus strain 19] Evolução de títulos sorológicos, nas provas de soroaglutinação em placa, antígeno acidificado tamponado e fixação de complemento, em bezerras Nelore vacinadas aos 18 meses de idade com Brucella abortus amostra B 19. Pesquisa Veterinária Brasileira 21(4):139-142. Depto Medicina Veterinária Preventiva e Reprodução Animal, Faculdade de Ciências Agrárias e Veterinárias, Unesp-Campus de Jaboticabal, SP 14884-900, Brazil.
One hundred and eight Nelore heifers (Bos indicus) about 18 months of age were vaccinated with a standard dosage (60-120 x 109 viable organisms) of Brucella abortus strain 19 vaccine, airning at to study the persistence of serological antibody titres by the plate agglutination, rase Bengal and cornplement fixation tests. Serum samples were collected just before vaccination and after 45 days, 6, 12 and 18 months. Before the vaccination, all the heifers showed negative results by the three tests. After 45 days, all the heifers had titres higher than or equal to 1/100 by the plate agglutination test and all of them showed positive results by the rose Bengal plate test, while 2 animals had titre 1/2 and 106 animals had titres higher than or equal to 1/4 by the complement fixation test. One year after the vaccination, most of the heifers did not show significant serological antibody titres. Considering the risk to which animals in endemic areas and infected herds are subjected, and considering the clear reduction of the serological antibody titres shown by most of the vaccinated heifers, one rnay assume that, in case of Nelore heifers in endemic areas and that were not vaccinated between 3 and 8 months of age, it would be more reasonable to vaccinate them at 18 months of age than to expose them to a high risk of Brucella infection.
Abstract in Portuguese:
RESUMO.- Mathias, L.A., Chaves, L.F., Chen, A.A., Girio, R.J.S. & Valério Neto, W. 2001. [Evolution of serological antibody titres by the plate agglutination, rose Bengal and complement fixation tests in Nelore heifers (Bos indicus) vaccinated at 18 months of age with Brucella abortus strain 19] Evolução de títulos sorológicos, nas provas de soroaglutinação em placa, antígeno acidificado tamponado e fixação de complemento, em bezerras Nelore vacinadas aos 18 meses de idade com Brucella abortus amostra B 19. Pesquisa Veterinária Brasileira 21(4):139-142. Depto Medicina Veterinária Preventiva e Reprodução Animal, Faculdade de Ciências Agrárias e Veterinárias, Unesp-Campus de Jaboticabal, SP 14884-900, Brazil.
Com o objetivo de estudar a persistência da resposta sorológica, através das provas de soroaglutinação em placa, rosa Bengala e fixação de complemento, 108 bezerras, com idade ao redor de 18 meses, foram vacinadas com uma dose padrão da vacina preparada com Bntcella abortus amostra B 19. Foram obtidas amostras de soro sangüíneo antes da vacinação e após 45 dias, 6, 12 e 18 meses. Antes da vacinação, todas as bezerras apresentaram resultados negativos nos três testes. Após 45 dias, todas apresentaram título a partir de 1/100 na prova de soroaglutinação em placa e todas apresentaram resultado positivo no teste rosa Bengala, ao passo que dois animais apresentaram título 1/2 e os demais apresentaram título a partir de 1/4 na reação de fixação de complemento. Após um ano de vacinação, a grande maioria das bezerras já não apresentava título sorológico significativo. Considerando o risco a que estão sujeitos animais que vivem em áreas endêmicas e em propriedades onde a doença ocorre, e considerando a acentuada redução de título sorológico observada na grande maioria dos animais, pode-se concluir que, no caso bezerras de raças zebuínas em áreas endêmicas e que não tenham sido vacinadas na idade regulamentar, é mais vantajoso vaciná-las com a amostra B 19 aos 18 meses de idade do que deixá-las expostas a um elevado risco de infecção por Brucella.
#6 - Toxicological sensitivity and specificity of the micro-complement fixation test for the detection of botulinum toxins C and D in culture medium and liver of mice, 18(2):47-52
Abstract in English:
ABSTRACT.- Menegucci E.A., Dutra I.S. & Döbereiner J. 1998. [Toxicological sensitivity and specificity of the micro-complement fixation test for the detection of botulinum toxins C and D in culture medium and liver of mice.] Sensibilidade toxicológica e especificidade do teste de microfixação de complemento na detecção de toxinas botulínicas C e D em meio de cultura e fígado de camundongo. Pesquisa Veterinária Brasileira 18(2):47-52. Depto Apoio, Produção e Saúde Animal, Unesp-Campus de Araçatuba, Cx. Postal 533, Araçatuba, SP 16015-050, Brazil.
The toxicological sensitivity and specificity of the micro-complement fixation test (MCF) for the detection of botulinum toxins C and D were studied in supernatants of the bacterial cultures and in livers of mice inoculated with lethal and sublethal doses. Botulinum toxins C and D were produced in Hemoline culture medium, titered through the determination of LD50 by the mouse test and adjusted to dilutions of 10, 1, 0.1, 0.01 and 0.001 LD50 Two experimental models were used to determine the toxicological sensitivity of MCF in the supernatant of the culture medium with the dilutions described, and also in liver extracts of mice weighing 20 g and inoculated with the same dilutions. Detection of the botulinum toxins was attempted in liver extracts of mice which had received lethal doses of the respective toxins, and in others which had been inoculated with sublethal doses and were sacrificed in intervals of 5 days. The results show that the toxicological sensitivity of MCF, regarding the two types of toxins at the level of 0.001 LD50, was 100% when the supernatants of the culture medium were tested; this means that the sensitivitywas 100 times higher than with the mouse test. The toxicological sensitivity of MCF in the liver extracts of mice inoculated with 1 and 10 LD50 of botulinum toxins C and D was inferior, giving values of 100, 80, 89 and 72% respectively. By this test it was also possible to detect botulinum toxins type C and D in liver extracts of mice inoculated with sublethal doses, up to 15 days after the injection. The specificity of MCF was 88% and 92%, when liver extracts of healthy contral mice were tested and when challenged with antitoxins C and D; and 100% when challenged with the supernatant of the culture medium. These results indicate that MCF could be of importance for research and could substitute in vivo tests.
Abstract in Portuguese:
RESUMO.- Menegucci E.A., Dutra I.S. & Döbereiner J. 1998. [Toxicological sensitivity and specificity of the micro-complement fixation test for the detection of botulinum toxins C and D in culture medium and liver of mice.] Sensibilidade toxicológica e especificidade do teste de microfixação de complemento na detecção de toxinas botulínicas C e D em meio de cultura e fígado de camundongo. Pesquisa Veterinária Brasileira 18(2):47-52. Depto Apoio, Produção e Saúde Animal, Unesp-Campus de Araçatuba, Cx. Postal 533, Araçatuba, SP 16015-050, Brazil.
No presente estudo pretendeu-se verificar a sensibilidade toxicológica e especificidade do Teste de Microfixação
de Complemento (MCF) na detecção de toxinas botulínicas C e D no sobrenadante de cultivas bacterianos e em fígados de camundongos inoculados com doses letais e subletais. As toxinas foram produzidas em meio de cultura Hemoline, tituladas através da determinação da DL50 pelo Bioensaio em Camundongo e diluídas nas concentrações de 10, 1, 0, 1, 0,01 e 0,001 DL50. Desta forma, foram utilizadas em dois modelos experimentais, onde foi determinada a sensibilidade toxicológica do MCF no sobrenadante do meio de cultura com as diluições descritas acima e ainda em extratos hepáticos de camundongos com peso corporal de 20g, inoculados com as mesmas diluições. A tentativa de evidenciação das toxinas botulínicas nos extratos hepáticos de camundongos foi realizada através da sua extração após a morte pela administração das doses letais e ainda pelo sacrifício dos animais inoculados -com doses subletais, em intervalos de 5 dias. Os resultados evidenciaram uma sensibilidade toxicológica para o MCF de 100% para os dois tipos de toxinas ao nível de 0,01 DL50, quando testados os sobrenadantes de meio de cultura, portanto 100 vezes superior ao Bioensaio em Camundongo. A sensibilidade toxicológica do MCF, quando examinados extratos hepáticos de camundongos inoculados com 1 e 10 DL50 de toxinas botulínicas C e D, foi inferior, com valores de 100, 80, 89 e 72%, respectivamente. Pelo teste foi possível detectar toxinas botulínicas tipos C e D nos extratos hepáticos de camundongos inoculados com doses subletais até 15 dias após a sua inoculação. A especificidade do MCF foi de 88 e 92%, quando testados extratos hepáticos de camundongos sadios, e confrontados com as antitoxinas C e D; e 100% no sobrenadante do meio de cultura. Os resultados apontam para uma possível utilização do teste como importante instrumento de pesquisa e ainda na eventual substituição dos testes in vivo pelas suas implicações éticas e limitações práticas.
#7 - Diagnosis of botulismo of cattle in Brazil by micro-complement fixation
Abstract in English:
The heat induced micro-complement-fixation test was used for the laboratory diagnosis of botulism in cattle. The following specimens were collected in southeastem and central-westem Brazil: Thirty six samples of blood serum, liver and rumen liquid from 23 bovines with clinical signs of botulism (a condition commonly called "doença da vaca caída" - fallen cow disease) and from 1 bovine inoculated subcutaneously with botulinum type C toxin. Samples of blood serum, liver and rumen liquid collected from healthy bovines were used as negative controls. Sixteen out of 17 serum samples were positive for botulinum toxin in cattle with clinical signs of acute, subacute and chronic botulism. All 9 liver samples were positive including those from chronic cases. Two samples of rumen liquid from bovines with the acute disease gave a positive test, but 2 other chronic cases were negative. All 3 samples collected from the bovine with subacute clinical sigos gave positive results. From 36 samples examined, 31 were positive; of these 2 were of type C, 5 type D and 24 samples had to be grouped within the CD complex. All samples gave negative results with the mouse bioassay, including the material collected from the experimental case. In conclusion at least one sample of each of the 24 cases examined gave a positive result for botulinum toxin; thus this highly sensitive test confirmed the diagnosis that the "doença da vaca caída" is indeed botulism, caused by type C and D toxins.
Abstract in Portuguese:
Foi utilizada a técnica de Microfixação de Complemento Induzida pelo Aquecimento para o diagnóstico laboratorial do botulismo em bovinos. Neste sentido foram colhidas, nas regiões Sudeste e Centro-Oeste do Brasil, 36 amostras constituídas por soro sangüíneo, fígado e líquido ruminal de 23 bovinos com quadro clínico de botulismo, vulgarmente conhecido como “doença da vaca caída”, e de 1 bovino intoxicado experimentalmente pela toxina botulínica tipo C. Como controle negativo foram utilizados soro sangüíneo, fígado e líquido ruminal de bovinos sadios. De 17 soros examinados de bovinos com quadros clínicos agudo, subagudo e crônico, 16 foram positivos para a presença de toxina; todas as 9 amostras de fígado também foram, positivas, mesmo de animais com intoxicação crônica. Duas amostras de líquido ruminal de bovinos com doença aguda foram, positivas no teste, enquanto 2 amostras colhidas de animais com evolução crônica foram negativas. As 3 amostras oriundas do bovino intoxicado experimentalmente e apresentando evolução subaguda, foram positivas na Microfixação. Das 36 amostras examinadas, 31 foram positivas no teste, sendo 2 pertencentes ao tipo C, 5 ao tipo D, e 24 agrupadas dentro do complexo CD. Todas essas amostras foram negativas no Bioensaio em camundongo, inclusive o material colhido do bovino intoxicado experimentalmente. Com base nos resultados, e tendo como referência o Bioensaio em camundongo, a Microfixação de Complemento demonstrou um excelente desempenho, permitindo o diagnóstico laboratorial de botulismo em pelo menos um material de cada um dos 24 casos examinados, confirmando assim o diagnóstico de que a "doença da vaca caída" é de fato botulismo causado pelas toxinas C e D.
