Rapid and sensitive detection of equine arteritis virus in semen and tissue samples by reverse transcription-polymerase chain reaction, dot blot hybridisation and nested polymerase chain reaction.
A reverse transcription-polymerase chain reaction (RT-PCR) assay using four different primer pairs for the detection of equine arteritis virus (EAV) RNA in semen and tissue samples was evaluated. A fragment encoding the leader sequence of the EAV genome was most successfully amplified. The specificity and sensitivity of RT-PCR was assessed by virus isolation in cell culture, restriction analysis, dot blot hybridisation and nested PCR. To this end, 23 semen samples from seropositive stallions and 11 tissue samples from 4 aborted foals were tested. Compared to the virus isolation test in cell culture, the sensitivity of the molecular methods proved to be higher. In the RT-PCR, dot blot hybridisation and nested PCR tests, semen samples from 11 stallions and tissue samples from all 4 foals were found positive, while the virus could be isolated in cell culture from only 4 semen samples and tissue samples from 1 aborted foal. The sensitivity of the dot blot hybridisation test was superior to that of the RT-PCR test. The nested PCR test proved to be the most sensitive one, because 3 semen samples were recognised as positive by this method only. Considering the sensitivity, and rapidity and reliability, RT-PCR followed by dot blot hybridisation or nested PCR represents the best method for diagnosis of EAV and should be included in the official diagnostic regimes.
Starick
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Federal Research Centre for Virus Diseases of Animals, Friedrich-Loeffler-Institute, Insel Riems, Germany.
D000818:Animals; D018174:Arterivirus Infections / Q000662:veterinary* / Q000821:virology; D002460:Cell Line; D007240:Equartevirus / Q000302:isolation & purification*; D005260:Female; D006734:Horse Diseases / Q000821:virology*; D006736:Horses; D015151:Immunoblotting / Q000379:methods; D008297:Male; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D011247:Pregnancy; D012367:RNA, Viral / Q000032:analysis; D012661:Semen / Q000821:virology*; D012680:Sensitivity and Specificity
D012367:RNA, Viral