Establishment and Application of PCR to Detect Wild-Type Pseudorabies Virus
Abstract: In order to effectively differentiate between gene-deleted vaccine and wild-type isolates of Pseudorabies virus(PrV), a PCR method, based on sequences of gE and gI of the PrV genome, was established after selection of the optimal reaction conditions. By applying this technique, the 848bp gene fragment was amplified from PrV RA strain, SH strain and LA strain, respectively. The negative results were achieved from Bartha-K61, RK-13, VSV, HCV, PRRSV, JEV, PPV and PCV2. Sequencing of the amplified products showed that the PCR method was specific. The sensitivity of PCR reached to 5pg DNA of PrV RA strain . We applied the PCR method to detect 172 tissue samples from 37 pig farms in Jiangsu, An’hui, Zhejiang, Fujian and Shanghai during 2003-2004, wild-type isolates of PrV were found in 35 tissue samples (20.34%) and 15 pig farms(40.54%). PrV distributed widely in naturally infected pig’s tissues including brain, liver, spleen, kidney, lung and lymphnodes. The PrV positive detection rate in lymphnodes was the highest in all tissues examined and reached 83.33%. The established PCR method provided a more sensitive, specific and reliable method to rapidly detect wild-type PrV and epizootic study of PrV.