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Bats are the natural reservoirs of severe acute respiratory syndrome-coronavirus (SARS-CoV). Six Alphacoronavirus and five Betacoronavirus have been detected in many bat species, including SARS-related CoV and Middle East respiratory syndrome (MERS)-related CoV. In Taiwan, SARS-related CoV, belonging to Betacoronavirus, has been detected in Rhinolophus monoceros. Scotophilus bat CoV-512, belonging to Alphacoronavirus, has been detected in Scotophilus kuhlii, Miniopterus fuliginosus, and Rhinolophus monoceros by using reverse transcription polymerase chain reaction (RT-PCR). To understand the infection history of CoV in these three insectivorous bat populations, CoV-specific antibodies were surveyed by using western blot (WB) analysis and indirect enzyme-linked immunosorbent assay (ELISA). The carboxyl terminal fragment of nucleocapsid protein (N3) of SARS-CoV and Scotophilus bat CoV-512 were used as the antigen in the assays. Of the 52 serum samples obtained from Scotophilus kuhlii, 29 samples (56%) were tested positive for Scotophilus bat CoV-512-specific antibodies through ELISA. Of the 63 serum samples obtained from Rhinolophus monoceros, 9 samples were tested positive for only SARS-CoV-specific antibodies, 7 samples were tested positive for only Scotophilus bat CoV-512-specific antibodies, and 16 samples (25.4%) were tested positive for both antibodies through WB analysis. Only 1 of 18 Miniopterus bat serum samples tested positive for Scotophilus bat CoV-512-specific antibodies through ELISA. Lactating female bats had higher positive rates of CoV-specific antibodies than non-lactating female and male bats did. Our findings were crucial for understanding CoV infection history in three insectivorous bat species and important for the control of bat-borne zoonosis diseases.

Bats can serve as natural reservoirs for severe acute respiratory syndrome (SARS) and Middle East respiratory syndrome coronavirus (MERS-CoV). Investigating the prevalence of bat CoV is critical for assessing the risks of the outbreaks of emerging CoV. Chestnut bats (Scotophilus kuhlii) were captured in this study for detecting the partial RNA-dependent RNA polymerase (RdRp) gene in their feces through reverse transcription polymerase chain reaction (RT-PCR) and antibodies to the nucleocapsid (N) protein of bat CoV through western blotting (WB) analysis. Three recombinant N protein fragments (N1, N2, N3) of the isolated Scotophilus bat CoV/CYCU-S1/TW/2013 were expressed by Escherichia coli. WB analyses were performed with bat serum samples and the sera of a patient who recovered from a SARS-CoV infection. Fragment N2 contained a highly conserved motif among CoVs whereas N1 and N3 protein fragments were specific to the S. kuhlii bat CoV. A total of 32 fecal and 19 serum samples were collected in Changhua County and Yunlin County during 2013 and 2014. About 17 fecal samples tested positive for the RdRp gene with an overall prevalence of 53%. Sequences comparison showed that the Scotophilus bat CoV isolates in Taiwan belonged to the genus Alphacoronavirus and were closest to Scotophilus bat CoV/Hainan/China/2005 and Diliman1552G1/Philippines/2008, followed by porcine epidemic diarrhea coronavirus. Only one bat serum sample reacted positively to all 3N protein fragments. Cross-reactivity was observed between N2 protein fragment and the sera of a patient recovered from a SARS-CoV infection. The results indicated that Scotophilus bat CoV was circulating endemically in chestnut bat population in Taiwan.