Detection of alpha- and betacoronaviruses in rodents from Yunnan, China

Abstract

Background: Rodents represent the most diverse mammals on the planet and are important reservoirs of human pathogens. Coronaviruses infect various animals, but to date, relatively few coronaviruses have been identified in rodents worldwide. The evolution and ecology of coronaviruses in rodent have not been fully investigated.

Results: In this study, we collected 177 intestinal samples from thress species of rodents in Jianchuan County, Yunnan Province, China. Alphacoronavirus and betacoronavirus were detected in 23 rodent samples from three species, namely Apodemus chevrieri (21/98), Eothenomys fidelis (1/62), and Apodemus ilex (1/17). We further characterized the full-length genome of an alphacoronavirus from the A. chevrieri rat and named it as AcCoV-JC34. The AcCoV-JC34 genome was 27,649 nucleotides long and showed a structure similar to the HKU2 bat coronavirus. Comparing the normal transcription regulatory sequence (TRS), 3 variant TRS sequences upstream the spike (S), ORF3, and ORF8 genes were found in the genome of AcCoV-JC34. In the conserved replicase domains, AcCoV-JC34 was most closely related to Rattus norvegicus coronavirus LNRV but diverged from other alphacoronaviruses, indicating that AcCoV-JC34 and LNRV may represent a novel alphacoronavirus species. However, the S and nucleocapsid proteins showed low similarity to those of LRNV, with 66.5 and 77.4% identities, respectively. Phylogenetic analysis revealed that the S genes of AcCoV-JC34, LRNV, and HKU2 formed a distinct lineage with all known coronaviruses.

Conclusions: Both alphacoronaviruses and betacoronaviruses were detected in Apodemus chevrieri in the Yunnan Province of China, indicating that Apodemus chevrieri is an important host for coronavirus. Several new features were identified in the genome of an Apodemus chevrieri coronavirus. The phylogenetic distance to other coronaviruses suggests a variable origin and evolutionary route of the S genes of AcCoV-JC34, LRNV, and HKU2. These results indicate that the diversity of rodent coronaviruses is much higher than previously expected. Further surveillance and functional studies of these coronaviruses will help to better understand the importance of rodent as host for coronaviruses.

Keywords: Coronavirus; Genetic diversity; Rodent.

Fig. 1

Phylogenetic analysis of detected rodent CoVs with representative CoVs based on 440-bp partial RdRp sequences. The tree was constructed by the maximum-likelihood method with 1000 bootstrap replicates. Bootstrap values above 50% were shown. Rodent CoVs found in this study are shown in bold. CoV abbreviations: bat SL-CoV WIV1, bat SARS-like coronavirus WIV1; BCoV, bovine coronavirus; CCoV, canine coronavirus; CrCoV, canine respiratory coronavirus; ECoV, equine coronavirus; FCoV, feline coronavirus; HCoV 229E, human coronavirus 229E; HCoV HKU1, human coronavirus HKU1; HCoV NL63, human coronavirus NL63; HCoV OC43, human coronavirus OC43; IBV, infectious bronchitis virus; MERS-CoV, MERS coronavirus; MHV, murine hepatitis virus; PHEV, porcine hemagglutinating encephalomyelitis virus; SARS-CoV, SARS coronavirus; TGEV, transmissible gastroenteritis virus

Fig. 2

Comparison of the genome organizations of AcCoV-JC34, LRNV, HKU2, FCoV, 229E, MHV, HKU24, and WIV1. Predicted ORFs and 5 conserved domains are indicted by the boxes. Abbreviations: 3CL, 3C-like protease; E, envelope; HE, hemagglutinin-esterase; Hel, helicase; M, membrane; N, nucleocapsid; PL1pro and PL2P, papain-like proteases 1 and 2; RdRp, RNA-dependent RNA polymerase; S, spike

Fig. 3

Phylogenetic analyses of AcCoV-JC34 based on amino acid sequences of ORF1a, 1b, S, E, M, and N. The trees were constructed by the maximum-likelihood method with 1000 bootstrap replicates. Bootstrap values above 50% are shown. AcCoV-JC34 identified in this study is shown in bold. The abbreviations and GenBank accession numbers are the same as those used in Fig. 1