Yi-Ning Chena*, Hong Chang Chenb, Cheng-Han Choub, Hsi-Chi Chengb. aDepartment of Bioscience Technology, Chung-Yuan Christian University, Chung-Li ...
Detection of bat coronavirus in the bat population in Taiwan Yi-Ning Chena*, Hong Chang Chenb, Cheng-Han Choub, Hsi-Chi Chengb aDepartment bEndemic
of Bioscience Technology, Chung-Yuan Christian University, Chung-Li, Taiwan 32023
Species Research Institute, Council of Agriculture, Chichi Township, Nantou, Taiwan 55244 Abstract
Background. The outbreaks of severe acute respiratory syndrome (SARS) during 2002 to 2003 and the Middle East respiratory syndrome (MERS) 10 years later have drawn much needed attention to the investigations for the natural reservoirs of coronaviruses, especially bats. Because bats can migrate for a very long distance and roost within human community, it is very important to assess the risk of host transition and the outbreaks of emerging coronavirus by investigating the prevalence of bat coronavirus (BtCoV). Bat coronaviruses have been detected around the world, including the countries surrounding Taiwan. The current study is to fill the gap of knowledge on the ecology of BtCoV in Taiwan. Methods & Materials. Individual feces were collected from 49 net-captured bats during 2013. The conserved region (440bp) of RNA-dependent RNA polymerase (RdRp) gene was targeted by reverse transcription-polymerase chain reaction (RT-PCR) in the extracted viral RNA in fecal samples. Results. Coronaviral RNA was detected in 10 out of 17 (59%) fecal samples from endemic bat species Rhinolophus monoceros in Tainan City, 8 out of 14 fecal samples (57%) from migrating bat species Scotophilus kuhlii in Changhua County, 2 out of 12 (17%) fecal samples from endemic bat species Miniopterus schreibersii fuliginosus in New Taipei City, and 1 out of 6 (17%) fecal samples from endemic bat species Kerivoula sp. in I-Lan County. Phylogenetic analysis by BLASTn and MEGA5 software generated the Maximum likelihood tree by the neighbor-joining model indicating that the detected BtCoV formed two clades in the genus Alphacoronavirus. The RdRp sequences of the first glade showed over 95% identity to species Scotophilus BtCoV 512 and Rhinolophus BtCoV HKU2, and the second clade had over 95% identity to species Miniopterus BtCoV 1. Conclusion. The findings of this study suggest that Bat Coronaviruses exit endemically in at least four bat species in Taiwan and further genomics and serological surveillance of coronaviruses in bats throughout Taiwan are necessary to understand the role of bat coronaviruses to the public public health.
Background
Habitant of Bats
Outbreaks of SARS and MERS → Bats as reservoirs? → Bat coronavirus in Taiwan ? 2/12 (17%)
8/14 (57%)
TAIWAN
1/6 (17%)
37 Deaths/346 Cases
Bat Catch/Capture & Sampling
10/17 (59%)
Results Location TAINAN
Bat Species Rhinolophus monoceros
CHANGHUA Scotophilus kuhlii TAIPEI ILAN
TAIWAN?
Miniopterus schreibersii fuliginosus Kerivoula sp.
YES!
Pos. #/Total (Prevalence) 10/17 (59%) 8/14 (57%)
Sample ID R2, R5, R6, R7, R9, R10, R11, R13, R14, R16 S1, S3, S5, S6, S8, S9, S11, S13
2/12 (17%)
M4, M8
1/6 (17%)
K9
440nt
440nt
440nt
Fig 4. PCR reaction for RdRp (440nt) from fecal samples. NC: negative control; M:100bp marker; -positive. Lane # is loaded with PCR product from the corresponding sample ID in the table. Some samples showing negative PCR are not shown here.
Lane # Sample ID 1-3 M1-M3 2-11 S1-S8 12-16, 31 S10-13, S9, S14 17-22 M4-M9 23-30 R2, R5-R11, R13 32-34 R14-R16 35 K9
Methods & Materials
Fig 5. Phylogenetic relationships of four representing bat coronaviruses isolated from four bat species in Taiwan. The Maximum likelihood tree was generated based on 440 nucleotides of the RNA-dependent RNA polymerase by the Neighbor-joining model in the MEGA5 program. Numbers above branches are bootstrap percentage calculated from 1000 bootstrap replicates. Scale bar, 0.2, represents nucleotide substitution per site.
Fig 6. Clustal W alignment (RdRp gene) of four bat coronavirus isolated from four bat species in Taiwan. Red letters are consensus sequences. Green letters are dominate sequences. Black letters are different sequences.
