RESUMEN
Turkey rotaviruses are one of the major pathogens responsible for the poult enteritis syndrome (PES). In this study a one step real-time reverse transcription polymerase chain reaction (RT-qPCR) assay targeting the rotaviral non-structural protein 4 (NSP4) was developed. The NSP4 is a highly conserved gene inside the turkey rotavirus genome and contains an internal control system to monitor any potential RT-qPCR inhibitors. The detection limit of the optimized NSP4-RT-qPCR assay ranged from 8.15 to 8.15 × 10(5) copy numbers. In total 149 faecal samples were collected from eight different flocks of commercial turkey farms. Faecal samples from hens and toms were collected separately at 2-week intervals from the 2nd week of age through the 16th and 20th week of age (age of slaughter for female and male, respectively) and tested. One farm reared only hens. The samples were tested previously using conventional RT-PCR targeting the same gene. When the conventional RT-PCR was compared with the developed NSP4-RT-qPCR, the results revealed that 11% of the samples of the conventional RT-PCR were false negative. The results indicate that this NSP4-RT-qPCR is highly sensitive for the detection of turkey rotaviruses in faeces. In addition, it could be suitable for the development of high-throughput screening.
Asunto(s)
Enfermedades de las Aves de Corral/diagnóstico , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Infecciones por Rotavirus/diagnóstico , Rotavirus/aislamiento & purificación , Pavos/virología , Animales , Secuencia de Bases , Heces/virología , Femenino , Masculino , Datos de Secuencia Molecular , Enfermedades de las Aves de Corral/virología , Rotavirus/genética , Infecciones por Rotavirus/virología , Sensibilidad y Especificidad , Alineación de Secuencia , Proteínas no Estructurales Virales/genéticaAsunto(s)
Viruela Vacuna/complicaciones , Dedos/patología , Linfadenitis/etiología , Linfadenitis/patología , Adolescente , Animales , Viruela Vacuna/diagnóstico , Viruela Vacuna/transmisión , Virus de la Viruela Vacuna/clasificación , Virus de la Viruela Vacuna/genética , Femenino , Hemaglutininas Virales/genética , Humanos , Ganglios Linfáticos/patología , Linfadenitis/diagnóstico , Filogenia , RatasRESUMEN
Diagnosis of porcine reproductive and respiratory syndrome virus (PRRSV) is often performed by serological testing, but ELISA does not differentiate between infections with wild-type or vaccine virus. Two attenuated live vaccines [European (EU) or North American (NA) genotype] are used. In addition to wild-type isolates, vaccine or vaccine-derived viruses occur frequently. This is often not considered when the ELISA results are used to differentiate between epizootic and enzootic infections. In this study, an infection with the NA genotype vaccine-derived virus was detected in two herds previously PRRSV negative and ELISA results [sample to positive (s/p) ratios] were analysed. The virus was identified by RT-PCR and nucleotide sequences of ORF5 had 97% (herd A) and 99% (herd B) identity with the genome of a ML PRRSV vaccine belonging to the NA genotype. Pigs of different age became positive with an average s/p ratio of 2.24 (A) and 1.18 (B). The data clearly demonstrate that spontaneous infection with a vaccine-derived virus of the NA genotype induces ELISA s/p ratios similar to those induced by vaccination or by infection with wild-type virus. We conclude that for a correct interpretation of serological results the circulation of vaccine or vaccine-derived virus isolates has to be excluded by RT-PCR, even if vaccination is not ongoing.