Development of reverse transcriptase PCR and nested PCR to detect porcine hemagglutinating encephalomyelitis virus.
Porcine hemagglutinating encephalomyelitis virus (HEV) causes encephalomyelitis, or vomiting and wasting disease, in suckling piglets. The mortality rate for piglets under 3 weeks old is 100%, but they are usually protected by maternal antibodies. Recently, the risk of an HEV outbreak has increased in the pig industry, because of widely using specific pathogen-free pigs that have no antibodies to HEV. We developed reverse transcription (RT) PCR and nested PCR to detect HEV. Primer sets of polymerase, non-structural protein, and spike protein were designed for RT-PCR and nested PCR based on the nucleotide sequences of the HEV 67N strain. The PCR designated primer sets of spike protein detected only HEV viral RNA among other related nidoviruses. Detection of HEV viral RNA by nested PCR was more sensitive than virus isolation in cell cultures. Nested PCR detected HEV viral RNA from experimentally infected samples of mice and field samples of piglets. The RT-PCR and nested PCR methods to detect HEV is considered a good way to show the HEV etiology on pig farms.
Sekiguchi
Yoshiko
Y
Department of Veterinary Microbiology, Tokyo University of Agriculture and Technology, Fuchu, Japan.
Shirai
Junsuke
J
Taniguchi
Takahide
T
Honda
Eiichi
E
D000383:Agriculture; D000818:Animals; D002460:Cell Line; D017934:Coronavirus / Q000235:genetics*; D017931:DNA Primers; D007564:Japan; D051379:Mice; D016133:Polymerase Chain Reaction / Q000379:methods*; D012313:RNA / Q000302:isolation & purification*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D034421:Sus scrofa / Q000821:virology*
D017931:DNA Primers; D012313:RNA