Utilização de um painel de proteínas recombinantes para o imunodiagnóstico da infecção por corynebacterium pseudotuberculosis em pequenos ruminantes

Abstract

Caseous lymphadenitis (CLA) is a small ruminant disease that is characterized by the development of granulomatose lesions in superficial and internal lymph nodes, as well as in some organs. CLA can be found worldwide causing significant economic losses, its etiological agent is the bacterium Corynebacterium pseudotuberculosis, and there is no commercial available diagnosis tool that presents satisfactory characteristics, as high level of accuracy, low cost and rapid results. The present study had the objective to develop an indirect ELISA for the detection of C. pseudotuberculosis specific immunoglobulins in small ruminants using recombinant proteins. The electrophoretic profiles of the recombinant antigens PLD, CP40, PknG, DtxR e Grx expressed in a heterologous prokaryotic system were analyzed by SDS-PAGE, and the immune recognition of the proteins was analyzed by Dot Blot. The recombinant proteins were employed in a checkerboard system for the standardization of the ELISA conditions. 130 goat’s serum samples and 160 sheep’s serum samples were used for the obtaining of the ELISA validation parameters. Antibodies from a pool of positive serum samples recognized the recombinant proteins, and there was a weak recognition when a pool of negative serum samples was used. The ELISA that was developed for goats using a combination of PLD and CP40 and only PLD for sheep was able to discriminate between positive and negative animals, with 95% sensitivity, 97.5 specificity and 99.3% accuracy for goats and 91% sensitivity, 98.7% specificity and 96.5% accuracy for sheep, respectively. It can be concluded that these developed ELISAs can be used as a high accurate tool in the immunodiagnosis of the infection with C. pseudotuberculosis in small ruminants.