ORIGINAL ARTICLE Detection and Molecular Characterization of Calf Diarrhoea Bovine Coronaviruses Circulating in South Korea during 20042005 S. J. Park1, G. K. Lim1, S. I. Park1, H. H. Kim1, H. B. Koh1and K. O. Cho1 1 Biotherapy Human Resources Center, College of Veterinary Medicine, Chonnam National University, Gwangju 500-757, South Korea Introduction Coronaviruses (CoVs), belonging to the family Coronavir- idae, are large, enveloped viruses 120150 nm in diameter and possess a single-stranded, plus-sense RNA genome of approximately 2630 kb in length (Lai and Holmes, 2001). Coronaviruses are now recognized as both veterin- ary and human pathogens that are associated with a wide range of economically important diseases in their respect- ive hosts. CoVs have been separated into three distinct subgroups based on the serological cross-reactivity and genomic relatedness (Lai and Holmes, 2001). The bovine coronavirus (BCoV) belongs to the second subgroup, which also contain a severe acute respiratory syndrome CoV, human respiratory CoV (HCoV-OC43), haemagglutinatingencephalomyelitisCoVofswine, turkey enteric CoV and murine hepatitis CoVs (MHV) (Lai and Holmes, 2001; Snijder et al., 2003). The BCoV causes severe calf diarrhoea (CD) and is associated with winter dysentery (WD) in adult cattle and respiratory infections in feedlot cattle (Saif and Heckert, 1990; Clark, 1993; Cho et al., 2000; Lathrop et al., 2000). Clinically different forms of BCoV infections have been reported in most cattle producing countries including those in Europe, North America and East Asia (Saif and Heckert, 1990). It is believed that CD BCoV infections have caused enormous economic losses in the cattle industry of South Korea, such as WD in adult cattle (Jeong et al., 2005b). Keywords: Bovine coronavirus; calves; genetic differences; prevalence Correspondence: K. O. Cho. Biotherapy Human Resources Center, College of Veterinary Medicine, Chonnam National University, Gwangju 500- 757, South Korea. Tel.: +82 62 530 2845; Fax: +82 62 530 0835; E-mail: chokochonnam.ac.kr Received for publication October 21, 2006 Summary Although the widespread occurrence of calf diarrhoea (CD) bovine coronavirus (BCoV) infections have been reported in most cattle producing countries, only the genetic differences in the BCoVs from American and Canadian isolates and/or strains have been identifi ed and compared. Hence, it is unclear if the BCoVs circulating in the other countries have distinct genetic characteristics. The aim of this study was to determine the prevalence and genetic diversity of CD BCoVs based on the deduced amino acid (aa) sequences of the spike (S) and haemagglutinin/esterase (HE) proteins in South Korea. RT-PCR and nested PCR using the primer pairs specifi c to the nucleocapsid gene, BCoVs detected the BCoVs in 56 (15.6%) of 359 diarrhoeic faecal samples. Phylo- genetic analysis of the entire S gene indicated that 10 Korean CD BCoV strains clustered with other Korean BCoV strains with different clinical forms but were different from the American and Canadian BCoV strains. Moreover, the phylogenetic data of the aa sequences of the HE gene revealed all the Korean CD strains to be distinct from the other Korean BCoV strains with different clinical forms. These results suggest that the Korean BCoVs cause endemic infections in diarrhoeic calves in Jeonnam province and have taken a different evolutionary pathway from the BCoVs in other countries. Moreover, the different BCoV strains are circulating in the different clinical forms in South Korea. These results also suggest that vaccines against the BCoVs can be developed with each Korean BCoV in different clinical forms. Zoonoses and Public Health 2007 The Authors Journal compilation 2007 Blackwell Verlag Zoonoses Public Health. 54 (2007) 223230223 However, little is known about the precise epidemiology of CD BCoV infections in South Korea. The BCoV contains fi ve major structural proteins: the nucleocapsid (N), transmembrane (M), spike (S), haem- agglutinin/esterase (HE) and small membrane (E) (Lai and Holmes, 2001). Although both the S and HE glyco- proteinshaemagglutinateerythrocytesbybindingto N-acetyl-9-O acetyl neuraminic acid as a receptor deter- minant, the S glycoprotein requires fewer of these receptors on the surface of the erythrocytes for agglutination than the HE protein (Schultze et al., 1991a,b). Therefore, the S glycoprotein is believed to be the main haemagglutinin of BCoV and is responsible for the primary attachment of BCoV to other cell surface receptors (Schultze et al., 1991b). The variation in the host range as well as the tis- sue tropism of CoVs is largely due to variations in the S glycoprotein (Gallagher and Buchmeier, 2001). Viruses circulating in geographically distinct areas or countries might have different antigenicity and pathogen- icity. Like other RNA viruses, CoVs are believed to mutate at a high frequency because of the high error frequencies of the RNA pol