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.

摘要

A reverse transcription-polymerase chain reaction (RT-PCR) assay using 4 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 hybridization and nested PCR. 23 semen samples from seropositive stallions and 11 tissue samplesfrom 4 aborted foals were tested. Compared with the virus isolation test in cell culture, the sensitivity of the molecular methods was higher. In the RT-PCR, dot blot hybridization and nested PCR tests, semen samples from 11 stallions and tissue samplesfrom all 4 foals were positive, while the virus was isolated in cell culture from only 4 semen samples and tissue samples from 1 aborted foal. The sensitivity of the dot blot hybridization test was superior to that of the RT-PCR test. The nested PCR testwas most sensitive; 3 semen samples were positive by this method only. It is concluded that RT-PCR is a rapid, sensitive and reliable method for the diagnosis of EAV. RT-PCR results can be further improved by subsequent dot blot hybridization or nestedPCR tests.