RESUMO
Pestivirus can contaminate cell cultures and sera and cause serious problems that evolve the integrity of studies, confidence in diagnostic results, and safety of human and animal vaccines. Contaminations by Pestivirus and other viruses may occur at any time and regular assays of monitoring in cell cultures and your supplies are necessary. This study aimed to analyze the phylogeny of Pestivirus detected from cell cultures, calf serum, and standard strains of three laboratories in Brazil that carry out frequent tests for the monitoring of cellular contaminations. These samples were submitted to phylogenetic analysis to understand the genetic relationship between contaminants occurring in these facilities. As result, the Pestivirus found in samples were Bovine viral diarrhea virus (BVDV-1 and BVDV-2), Hobi-like viruses (often named BVDV-3), and Classical swine fever virus (CSFV), and the phylogenetic analysis help us to infer at three possible routes of contamination in this work.
Assuntos
Vírus da Diarreia Viral Bovina Tipo 1 , Vírus da Diarreia Viral Bovina , Pestivirus , Animais , Suínos , Humanos , Pestivirus/genética , Filogenia , Vírus da Diarreia Viral Bovina/genética , Vírus da Diarreia Viral Bovina Tipo 1/genética , Linhagem CelularRESUMO
Foot-and-mouth-disease (FMD) is a highly contagious disease of domestic animals which can result in substantial economic losses, caused by the FMD virus (FMDV). The aim of this study was to develop and standardize a novel reverse transcriptase droplet digital PCR (RT-ddPCR) assay for the quantification of FMDV RNA. This assay was based upon an OIE-recognized real-time RT-PCR that detects the 3D-encoding region of FMDV. The limit of detection at 101.4 TCID50/mL and 26.5 copies was determined using FMDV-A24-Cruzeiro-virus and a plasmid containing the 3D-FMDV sequences, respectively. FMDV O, A and C serotypes and 11 species of non-FMDV were used to confirm the sensitivity and specificity of the assay. The RT-ddPCR was standardized using 60 bovine samples (representing negative and positive samples of epithelium and/or oesophageal-pharyngeal [OP] fluid) from animals suspected of vesicular diseases and previously tested by RT-qPCR. The RT-ddPCR showed robustness, sensitivity, specificity and accuracy, with similar results to the RT-qPCR. Moreover, the new RT-ddPCR diagnostic tool allowed the absolute quantification of FMDV RNA from epithelium and OP-fluid samples, as well as having the advantages of direct quantification by endpoint, eliminating the need for a calibration standard curve required in quantitative real-time RT-PCR.
Assuntos
Vírus da Febre Aftosa/isolamento & purificação , Febre Aftosa/virologia , RNA Viral/análise , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Carga Viral/métodos , Animais , Bovinos , Doenças dos Bovinos/virologia , Vírus da Febre Aftosa/genética , RNA Viral/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa/normas , Sensibilidade e Especificidade , Sorogrupo , Carga Viral/normasRESUMO
Senecavirus A (SV-A) may cause vesicular disease and neonatal mortality in pigs, and was first detected in Brazil in 2015. Samples including tissues and serum from pigs with suspected vesicular diseases were collected from January to August in 2015 from farms in the states of Minas Gerais, Santa Catarina, Goiás and Rio Grande do Sul, Brazil, and tested for the presence of SV-A by reverse transcriptase PCR. All samples were negative for foot and mouth disease virus, as well as 13 other infectious agents associated with vesicular diseases in pigs. SV-A was detected by PCR in 65/265 (24.5%) specimens. A 530 base pair fragment sequenced from the VP1 protein coding region indicated a high genetic distance from SV-A in other countries, but a common origin among the Brazilian isolates.
