Detection of viruses in nasal swab samples from horses with acute, febrile, respiratory disease using virus isolation, polymerase chain reaction and serology

Objective To examine the association of viruses with acute febrile respiratory disease in horses. Design Nasal swab and serum samples were collected from 20 horses with acute febrile upper respiratory disease that was clinically assessed to have a viral origin. Methods Each of the samples was inoculated onto equine fetal kidney, RK1 3 and Vero cell cultures, and viral nucleic acid was extracted for polymerase chain reaction (PCR) or reverse transcription PCR. PCR primers were designed to amplify nucleic acid from viruses known to cause or be associated with acute febrile respiratory disease in horses in Australia. A type specific ELISA was used to measure equine herpesvirus (EHV1 and EHV4) antibody, and serum neutralisation assays were used to measure equine rhinitis A virus (ERAV) and equine rhinitis B virus 1 and 2 (ERBV1 and ERBV2) antibody titres in serum samples. Results Virus was isolated from 4 of 20 nasal swab samples. There were, three isolations of EHV4 and one of ERBV2. By PCR, virus was identified in the nasal swab samples of 12 of the 20 horses. Of those that were positive, 17 viruses were detected as follows: one triple positive (EHV4, EHV2, and EHV5), three double positives (EHV4, ERBV and EHV2, EHV5 in two horses) and eight single positives (EHV4 in two horses, EHV2 in one horse, EHV5 in six horses and ERBV in six horses). Conclusion By virus isolation and PCR, 17 viruses were identified in nasal swab samples from 12 of 20 horses that had acute febrile respiratory disease consistent with a diagnosis of virus infection. Initial PCR identification and subsequent virus isolation led to the isolation of ERBV2 for the first time in Australia and the second time anywhere of ERBV2.