PADRONIZAÇÃO E VALIDAÇÃO DA REAÇÃO DE qPCR PARA GÊNERO Parapoxvirus.

Abstract

The genus Parapoxvirus includes 4 species, including Bovine Papular Stomatitis Virus (BPSV), Orf virus (ORFV), New Zealand Parapoxvirus of red deer (PVNZ) and Pseudocowpox (PCPV). These species affect cattle and have their importance in the differential diagnosis for foot and mouth disease, besides being able to infect humans who had direct or indirect contact with the animal. This study aims to standardize and validate quantitative PCR reaction (qPCR) for implementation in the diagnostic routine of the Laboratório de Viroses de Bovideos (LVB), Instituto Biologico de São Paulo. As reference strain the Pseudocowpoxvirus (PCPV), ATCC VR-634 was used in the 4th pass in MDBK strain cells. The viral material was extracted and subjected to a series of steps for reaction optimization including determination of Primer concentrations, probe in addition to annealing temperature, sensitivity and repeatability. The efficiency of the test was 91.398%, maintaining the slope between the viral dilutions of -3.547, with a ratio coefficient (R2) of 0.948 and a high sensitivity detecting 1 TCID 50 / 50μL. Obtaining an optimized and standardized reaction for inclusion in the laboratory routine.