Prevalence of canine coronavirus (CCoV) in dog in Japan: detection of CCoV RNA and retrospective serological analysis

TAKANO, Tomomi; YAMASHITA, Saya; MURATA-OHKUBO, Michiko; SATOH, Kumi; DOKI, Tomoyoshi; HOHDATSU, Tsutomu

Rectal swab samples were collected from 101 dogs between 2011 and 2014. These samples were submitted by veterinary clinics in Japan (Aomori, Tochigi, Ibaraki, Saitama, Chiba, Tokyo, Kanagawa, Osaka, Hyogo, Kochi and Okinawa). Viral RNA was extracted from rectal swabs using the High Pure Viral RNA Isolation Kit (Roche, Basel, Switzerland) following the attached instructions. cDNA was amplified by PCR using specific primers as shown in Table 1 . PCR was performed using the method of Takano et al. [13] . ELIF-ELIR used to detect CCoV-I also detects FCoV-I. Thus, the ORF3f-ORF3r primer set which specifically detects only CCoV-I was also used. The CCoV gene was detected in rectal swab samples collected from dogs. CCoV N gene was detected in 27/101 (26.7%). CCoV-I S gene, CCoV-II S gene and CCoV-I specific ORF3 gene were detected in 21/101 (20.8%), 2/101 (2.0%) and 24/101 (23.8%), respectively. On the basis of the rate of CCoV N gene-and ORF3 gene-positive CCoV-infected dogs, the type of CCoV detected in samples from CCoV-infected dogs was CCoV-I in 88.9%. The CCoV-I S gene fragments (PCR products generated using ELIF-ELIR) were sequenced as described by Takano et al. [15] . Amino acid sequences were deduced from the CCoV-I S gene detected in 4 dogs with diarrhea symptoms (Nos.9, 19, 27 and 33) and 2 dogs without diarrhea symptoms (Nos. 55 and 73) (Fig. 1A) . All of 6 CCoV strains detected in this study were classified into the clade of the other CCoV-I (Fig. 1B) .