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. 2024 Jul 21;12(7):1490.
doi: 10.3390/microorganisms12071490.

Identification and Characterization of an Alphacoronavirus in Rhinolophus sinicus and a Betacoronavirus in Apodemus ilex in Yunnan, China

Qian Liu  1 , Dan-Shu Wang  1 , Zhong-Hao Lian  1 , Jie Fang  1 , Pei-Yu Han  2 , Ye Qiu  1 , Jun-Ying Zhao  2 , Li-Dong Zong  2 , Yun-Zhi Zhang  2 , Xing-Yi Ge  1
Affiliations

Identification and Characterization of an Alphacoronavirus in Rhinolophus sinicus and a Betacoronavirus in Apodemus ilex in Yunnan, China

Qian Liu et al. Microorganisms. .

Abstract

Coronaviruses (CoVs), the largest positive-sense RNA viruses, have caused infections in both humans and animals. The cross-species transmission of CoVs poses a serious threat to public health. Rodents and bats, the two largest orders of mammals, serve as significant natural reservoirs for CoVs. It is important to monitor the CoVs carried by bats and rodents. In this study, we collected 410 fecal samples from bats and 74 intestinal samples from rats in Yunnan Province, China. Using RT-PCR, we identified one positive sample for alphacoronavirus (TC-14) from Rhinolophus sinicus (Chinese rufous horseshoe bat) and two positive samples for betacoronavirus (GS-53, GS-56) from Apodemus ilex (Rodentia: Muridae). We successfully characterized the complete genomes of TC-14 and GS-56. Phylogenetic analysis revealed that TC-14 clustered with bat CoV HKU2 and SADS-CoV, while GS-56 was closely related to rat CoV HKU24. The identification of positive selection sites and estimation of divergence dates further helped characterize the genetic evolution of TC-14 and GS-56. In summary, this research reveals the genetic evolution characteristics of TC-14 and GS-56, providing valuable references for the study of CoVs carried by bats and rodents in Yunnan Province.

Keywords: CoV; HKU2; HKU24; bats and rodents; phylogenetic analysis.

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Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
The maximum likelihood tree of RNA-dependent RNA polymerase (RdRp) sequences of TC-14, GS-56, and other CoVs. The tree was constructed using IQ-tree with LG + F + I + G4 substitution model and 10,000 ultrafast bootstraps. The scale bar indicates amino acid substitution per site, and four genera of coronavirus are marked on the side. The TC-14 and GS-56 are marked in red and with a triangle.
Figure 2
Figure 2
Phylogenetic trees of amino acid sequences of S and N proteins of TC-14 and GS-56 CoVs. These trees were constructed using IQ-tree with 10,000 ultrafast bootstraps, and their substitution models are as follows: WAG + F + I + G4, pfam + F + I + G4, WAG + F + G4, pfam + F + G4, respectively. The accession number, taxonomy, and host of each sequence are displayed. TC-14 and GS-56 are marked in red and with a triangle.
Figure 3
Figure 3
Multiple alignments of S1 region of TC-14, SADS-CoV, and SADSr-CoV. The accession number and host of the selective sequences are shown. The annotation of the NTD domain in S1 is with reference to SADS-CoV; the NTD domain is marked with a blue line, and the CTD domain is marked with a green line. The short horizontal line indicated the identical sites. * indicates odd multiples of ten of the amino acid.
Figure 4
Figure 4
Multiple alignment of S1 region of GS-56, ChRCoV-HKU24, and RtAp-CoV. The accession number and host of the genomes are shown. The annotation of NTD and CTD domains in S1 is a reference to ChRCoV HKU24. The NTD domain is marked with a blue line, and the CTD domain is marked with a green line. The short horizontal line indicates the identical sites. * indicates odd multiples of ten of the amino acid.
Figure 5
Figure 5
Maximum clade credibility (MCC) tree with divergence time based on RdRp nucleotide sequence. The substitution model was GTR + F + F + I + G4, the tree was obtained by Model Finder under the BIC standard [37]. The value near the node indicates the age of the node, and the label on the branch represents the Bayesian posterior probability. The arrows are used to accurately locate points of divergence. The species and sampling time of the selected sequences are labeled. The TC-14 and GS-56 are marked in red.
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