Multiplex RT-PCR for detection of the three RNA viruses CSFV,JEV and PRRSV

Classical swine fever virus(CSFV),Japanese encephalitis virus(JEV)and porcine reproductive and respiratory syndrome virus(PRRSV)are the major causes of some serious reproductive diseases and co-infection is common.Therefore rapid and reliable diagnosis is crucial for effective prevention and treatment.In this study,a multiplex PCR was developed using primers designed according to published CSFV,JEV and PRRSV sequences.PCR products of 508 bp,380 bp and 263 bp were obtained for respective viruses.Restriction digestion of PCR products with EcoR V and Sau3A I produced expected target DNA fragments.Compared to CSFV antigen test kit,the multiplex RT-PCR showed 96.7% coincidence with the commercial kit based on clinical test results.As low as 12.5 TCID_(50) JEV,10 TCID_(50) PRRSV and 10~(-3) ng total RNA including CSFV RNA could be detected by multiplex PCR.The result showed that the Multiplex RT-PCR is a rapid,sensitive and specific method for detecting virus from clinical samples.