Caracterización Bioquimica Y Perfil De Sensibilidad Antimicrobiana in Vitro De Muestras De Burkholderia Mallei Aisladas De Équidos De La Región Nordeste De Brasil

Glanders is an infectious-contagious disease of acute or chronic character which principally affects horses, causing enormous losses in the productive chain of this animal. To control the disease, the Ministry of Agriculture, Husbandry and Supply instituted mandatory sanitation measures in the entire national territory which include an ofϐicial diagnosis through the complement ϐixation (CF) test, maleinization and sacriϐice of the animals that are positive. Nowadays the kits used for the diagnosis of the disease are imported, making their routine application difϐicult and more expensive. The objective of this study was to standardize an indirect ELISA test, using the proteic extract of Burkholderia mallei isolated from a carrier horse in the state of Pernambuco. The samples were cultivated in 10% blood agar and incubated for 48h at 37°C; later, one of the isolated colonies was characterized phenotypically and genotypically and immediately cultivated in brain heart infusion (BHI) for enrichment; then it was peaked (repicada) for the Dor-set Henley medium which was incubated at 37ºC under 60rpm for eight weeks. To standardize the test the Protein G Peroxidase Sigma Conjugate was used in the dilution of 1:90.000, with serums diluted in 1:100 and the antigen in 1:400. Sixty serums were used as negative controls, tested before the CF to determine the cutting point which was 0.042nm. After establishing the standardization, 300 samples were tested, of which 99% (297) were in agreement with the results obtained in the CF. At the end, of assay presented 100% sensibility and 98.2% speciϐicity, with predictive (preditivo) positive and negative values of 97.7% and 100% respectively. The Kappa concordance test was 0.98 and the intra and interplac repeatability were 8.8% and 10.3% respectively. From the results obtained, it is possible to afϐirm that the indirect ELISA test can be used as an efϐicient diagnosis tool. However, more essays must be carried out to consolidate the reliability of this test.

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Produção e purificação parcial de PPD-maleína para diagnóstico do mormo em equídeos

Silva

Campos-Takaki

Teles³

et al. 2014

Pesq. Vet. Bras.

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Pesq. Vet. Bras. 34(1):57-61, janeiro 2014 57 RESUMO.-Objetivou-se com este estudo produzir e purificar parcialmente a PPD-maleína a partir de amostras de Burkholderia mallei isoladas de equídeos no Brasil com potencial para uso no diagnóstico do mormo. As linhagens de B. mallei fenotipicamente caracterizadas e de virulência comprovada foram inoculadas em caldo Dorset-Henley para crescer e metabolizar. Em seguida, as proteínas foram separadas por precipitação com ácido tricloroacético e precipitadas com sulfato de amônia. As PPDs-maleínas foram concentradas em 1,0mg/ mL e na avaliação realizada em cobaios foi eficaz no desenvolvimento da hipersensibilidade do tipo tardia e consequentemente na identificação de animais verdadeiro positivos e exclusão dos verdadeiro negativos, sendo uma possibilidade em potencial para utilização no diagnóstico do mormo. INTRODUÇÃOO mormo é considerado a principal doença bacteriana dos equídeos, sendo transmissível ao homem e a outras espé-cies animais. Registrada sua reemergência no Brasil (Mota et al. 2000), atualmente a doença constitui um sério problema sanitário para os equídeos das regiões Norte e Nordeste do País (Mota et al. 2005, Brasil 2008 The objective of this study was to produce and partially purify Malleo-protein from Burkholderia mallei samples isolates from Equidae in Brazil with potential for use in the diagnosis of glanders. The strain B. mallei phenotypically characterized and proven virulent was inoculated into broth Dorset-Henley to grow and metabolize. The proteins were separated by trichloroacetic acid precipitation and amonium sulfate precipitation. The PPD mallein was concentrated 1.0mg/mL and biologically tested in guinea pigs. It was effective in the development of delayed-type hypersensitivity and consequently to identify true--positive animals and to exclude of true negatives. There is the possibility for potential use in the glanders diagnosis in Equidae.

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Caracterización Bioquimica Y Perfil De Sensibilidad Antimicrobiana in Vitro De Muestras De Burkholderia Mallei Aisladas De Équidos De La Región Nordeste De Brasil

Cited by 4 publications

References 16 publications

Current status of glanders in Brazil: recent advances and challenges

Current status of glanders in Brazil: recent advances and challenges

Desenvolvimento e avaliação de um teste ELISA indireto para o diagnóstico sorológico do mormo em equídeos

Produção e purificação parcial de PPD-maleína para diagnóstico do mormo em equídeos

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