The novel hemagglutinin-esterase genes of human torovirus and Breda virus

Human torovirus (HTV) and Breda virus (BRV), members of the genus torovirus in the family Coronaviridae, are established infectious agents of humans a nd cattle, respectively. The hemagglutinin-esterase (HE) gene of Breda viru s serotype 2 (BRV-2) has been identified and the nucleotide sequence for BR V serotype 1 (BRV-1) genome which contains the open reading frames for the viral structural proteins has been reported revealing the presence of a 1.2 5 kb gene whose nucleotide sequence is identical to that of the BRV-2 HE ge ne. In this study, we amplified the 1.2kb HE gene from the HTV genome using long RT-PCR and sequenced the amplicon directly. At the nucleotide level, the HTV HE gene manifests 85% sequence identity to the HE genes of BRV-1 an d BRV-2 and 89% identity with the X pseudogene sequence of BEV. The 1.25 kb amplicons which contained the HE genes of BRV-1 and HTV were cloned and ex pressed in a baculovirus system and the proteins purified by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. Hyperimmune sera prepared in guinea pigs against these proteins were reactive with both bovine torovirus (BTV) and human torovirus (HTV) antigens. By immunoblot, they reacted spec ifically with a 65 kDa protein corresponding in size to the torovirus HE pr otein. Furthermore, the hyperimmune sera but not the preimmune sera reacted with a series of BTV-positive and HTV-positive fecal specimens by immunobl ot and dot blot analysis. By immunoelectron microscopy (IEM) torovirus part icles from BTV-positive specimens from calves with diarrhea and HTV-positiv e specimens from patients were aggregated by the hyperimmune sera. Human co nvalescent sera and gnotobiotic calf post-infection sera reacted by immunob lot with the expressed 65 kDa protein. The expressed HE protein of HTV has important diagnostic potential. (C) 1999 Elsevier Science B.V. All rights r eserved.