Establishment of serological test to detect antibody against ferret coronavirus

MINAMI, Shohei; TERADA, Yutaka; SHIMODA, Hiroshi; TAKIZAWA, Masaki; ONUMA, Mamoru; OTA, Akihiko; OTA, Yuichi; AKABANE, Yoshihito; TAMUKAI, Kenichi; WATANABE, Keiichiro; NAGANUMA, Yumiko; KANAGAWA, Eiichi; NAKAMURA, Kaneichi; OHASHI, Masanari; TAKAMI, Yoshinori; MIWA, Yasutsugu; TANOUE, Tomoaki; OHWAKI, Masao; OHTA, Jouji; UNE, Yumi; MAEDA, Ken

In this study, we attempted to clarify the seroprevalence of FRcoV in Japan and developed an ELISA using two Yamaguchi-1 strain recombinant N proteins, GST-N (1-179) and GST-N (180-374). More ferret serum samples recognized GST-N (1-179) than GST-N (180-374) (Fig. 2 ). In addition, identities of N (1-179) between Yamaguchi-1 and the other FRcoVs (96.6-98.3%) were higher than those of N (180-374) (90.7-93.8%) (data not shown). Therefore, we selected GST-N (1-179) as the ELISA antigen for our serosurvey. Surprisingly, we found that 89% (31/35) of domestic ferrets were seropositive to this antigen by ELISA (Table 1 ). There are reports of FRcoV gene detection in 56%-61% of ferrets in Japan and the Netherlands [6, 9] . These data indicate that FRcoV has already spread within the ferret population and that many ferrets may be persistently infected with FRcoV. However, there was no significant difference between seropositivity and symptoms, age or sex. Further studies are required to clarify the pathogenesis of FRcoV in ferrets. Plasma from ferret No.10 and serum from ferret No.22 showed different reactivities from those of other ferret samples in ELISA, reacting only with GST-N (1-179), but not with GST-N (180-374) (Fig. 2) . The different reactivity of ferret No. 22 serum was also confirmed by immunoblot analysis using GST-N (180-374) (Fig. 3c) . These results indicated that GST-N (1-179) is a better choice of antigen for ELISA than GST-N (180-374). ELISA using GST-N (1-179) will be useful for serological surveys for FRcoV. In future studies, this FRcoV infected with ferret No.22 should be analyzed closely.