JVI Accepts, published online ahead of print on 7 March 2007

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J. Virol. doi:10.1128/JVI.02361-06
Copyright (c) 2007, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Biologic, antigenic and full length genomic characterization of a bovine-like coronavirus isolated from a giraffe

Mustafa Hasoksuz, Konstantin Alekseev, Anastasia Vlasova, Xinsheng Zhang, David Spiro, Rebecca Halpin, Shiliang Wang, Elodie Ghedin, and Linda J. Saif^*

Food Animal Health Research Program, Department of Veterinary Preventive Medicine, Ohio Agricultural Research and Development Center, The Ohio State University, 1680 Madison Avenue, Wooster, Ohio, 44691-4096; The Institute for Genomic Research (TIGR), Rockville, MD

^* To whom correspondence should be addressed. Email: saif.2{at}osu.edu.

Coronaviruses (CoVs) possess large RNA genomes and exist as quasispecies, which increases the possibility of adaptive mutations and interspecies transmission. Recently, CoVs were recognized as important pathogens in captive wild ruminants. This is the first report of the isolation and detailed genetic, biologic and antigenic characterization of a bovine-like CoV from a giraffe (Giraffa camelopardalis) in a wild animal park in the USA. Coronavirus particles were detected by immune electron microscopy in fecal samples from 3 giraffes with mild to severe diarrhea. From one of the three giraffe samples, a CoV (GiCoV-OH3) was isolated and successfully adapted to serial passage in human rectal tumor-18 cell cultures. Hemagglutination assays, receptor-destroying enzyme activity, hemagglutination inhibition and fluorescence-focus neutralization tests revealed close biological and antigenic relationships between the GiCoV-OH3 isolate and selected respiratory and enteric bovine coronavirus (BCoV) strains. When orally inoculated into a BCoV-seronegative gnotobiotic calf, GiCoV-OH3 caused severe diarrhea and virus shedding within 2-3 days. Sequence comparisons and phylogenetic analyses were performed to assess its genetic relatedness to other CoVs. Molecular characterization confirmed that the new isolate belongs to group 2a of the mammalian CoVs, and revealed closer genetic relatedness between GiCoV-OH3 and the enteric BCoVs, BCoV-ENT and BCoV-DB2, whereas BCoV-Mebus was more distantly related. Detailed sequence analysis of the GiCoV-OH3 spike gene demonstrated the presence of a deletion in the variable region of the S1 subunit (from 543 to 547 aa), which is a region associated with pathogenicity and tissue tropism for other CoVs. The identified point mutations in the structural proteins (comparing GiCoV-OH3, BCoV-ENT, BCoV-DB2 and BCoV-Mebus) were most conserved between GiCoV-OH3, BCoV-ENT and BCoV-DB2, whereas most point mutations in the nonstructural proteins were unique to GiCoV-OH3. Our results confirm the existence of a bovine-like CoV transmissible to cattle from a wild ruminant, namely giraffes, but with certain distinctive genetic properties from BCoVs.

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