Detection of bovine herpesvirus-1 infection in breeding bull semen by virus isolation and polymerase chain reaction

Abstract

Serum samples from 51 apparently healthy breeding bulls were screened for bovine herpesvirus-1 (BHV-1) antibodies using an avidin-biotin enzyme-linked immunosorbent assay, revealing a sero-positive prevalence rate of 45.09%. Semen samples were then collected from 12 of the sero-positive and 12 of the sero-negative bulls and tested for BHV-1 antigen using both a virus isolation assay and a polymerase chain reaction (PCR) assay; PCR was applied to detect BHV-1 deoxyribonucleic acid by using primers selected from the relatively conserved sequence of the gl glycoprotein gene to amplify a 468 base pair fragment. The PCR-amplified products were confirmed as BHV-1 by restriction enzyme, Dde 1, which produced fragments of predictable sizes, namely 340 and 128 base pairs. Positive virus isolation test results, confirmed by virus neutralisation, found BHV-1 antigen in the semen of five sero-positive and six sero-negative bulls. In comparison, positive PCR results found BHV-1 genome in the semen of six sero-positive and eight sero-negative bulls. From the 24 semen samples tested, 14 were shown to be positive by PCR and 11 by virus isolation. The sensitivity and specificity of virus isolation were 57.14% and 70% respectively, and were significantly lower than PCR. In the semen samples taken from sero-negative bulls, BHV-1 was detected more often by PCR methods than by virus-isolation, suggesting that PCR is a more sensitive method for BHV-1 screening in bulls.