key: cord-1022199-h29u51hp authors: Zhu, Wentao; Song, Wentao; Fan, Guoyin; Yang, Jing; Lu, Shan; Jin, Dong; Luo, Xue-lian; Pu, Ji; Chen, Haiying; Xu, Jianguo title: Genomic Characterization of a New Coronavirus from Migratory Birds in Jiangxi Province of China date: 2021-07-08 journal: Virol Sin DOI: 10.1007/s12250-021-00402-x sha: a536dabf168fa38c14fd26fd8951179d6de6e0aa doc_id: 1022199 cord_uid: h29u51hp nan . These analyses identified AFCoV as a new member of the species of Goose coronavirus CB17. The genome of AFCoV has a 38.5% G ? C content and typical CoV genomic organization consisting of 5 0 UTR (untranslated region), replicase ORF1ab, spike (S), envelope (E), membrane (M), nucleocapsid (N), several accessory proteins, and 3 0 UTR (Fig. 1A) . The 5 0 UTR of AFCoV is 519 nt in length and shares the highest nucleotide identity (94.9%) with that of CGCoV. Like other coronaviruses, the ORF1ab gene occupies a large portion of the AFCoV genome and has a frameshift site at position 11,940 based on the conserved heptamer 'UUUAAAC'. The polyprotein ORF1ab of AFCoV is cleaved into 16 nonstructural proteins (NSPs) by two viral proteases, with almost the same putative protease cleavage sites as those of CGCoV (Supplementary Table S1 ). The AFCoV genome contains nine predicted accessory proteins, one less than those in CGCoV (Papineau et al. 2019) (Supplementary Table S2 ). The 3 0 UTR (excluding the poly(A) tail) of AFCoV (255 nt in length) shows 98.8% nt identity with that of CGCoV. The protein ORF1ab of AFCoV (6500 aa) shares 90.0% aa identity with that of CGCoV. The deduced S protein (1186 aa) of AFCoV shares a low identity (68.5%) with the S protein of CGCoV (1184 aa), and even lower identities with that of other coronaviruses (\ 58.1%). The deduced E, M, and N proteins of AFCoV have 100%, 97.8%, and 94.9% aa identities with those of CGCoV respectively, and relatively low identities with the proteins of other Gammacoronaviruses ranging from 29.7 to 62.4% for E protein, 30.5-65.8% for M protein, and 36.4%-60.8% for N protein. To assess the phylogenetic relationship of AFCoV with other coronaviruses, the full-length genome sequences of alphacoronaviruses, betacoronaviruses, gammacoronaviruses, and deltacoronaviruses were aligned with MAFFT and used to reconstruct the phylogenetic tree by the maximum-likelihood method with JC best-fit model. The phylogenetic tree (Fig. 1B) reveals that AFCoV clusters in the genus Gammacoronaviruses and groups closely with CGCoV. Phylogenetic trees based on proteins ORF1ab (LG model), and S (Dayhoff model) were also constructed, and show similar topologies as that of the genomes, indicating that AFCoV is closely related to CGCoV ( Supplementary Fig. S1-S2) . To determine the prevalence of the new Gammacoronavirus in bird samples, we designed primers (F: 5 0 -CTTTTTGGTCTCTACCCTGTTC-3 0 , R: 5 0 -CCA-TAAAAATCCAGGACTTGTT-3 0 ) to specifically amplify an 836 bp segment located in the replicase ORF1ab of AFCoV genome. All fecal samples were tested using the One-Step RT-PCR Kit Ver.2 (Takara, Japan). Five out of 48 (10.4%) samples were found to be positive for coronavirus and were also verified by Sanger sequencing. Result of alignment showed that four of the five sequences were absolutely the same, and the remaining one sequence has seven nucleotide differences with the consensus sequence. The five hosts of those coronavirus positive samples belong to four species, all from family Anatidae, including Cygnus columbianus (one host) and Anser albifrons (one host) from Nanchang City, Anas zonorhyncha (two hosts), and Anser fabalis (one host) from Jiujiang City. Identifying viruses that have ability to spillover from wildlife into other animals and/or humans promotes our understanding of viral families that are very likely to be new pathogens in the future (Anthony et al. 2015) . Here, we detected a new Gammacoronavirus (AFCoV) in four different bird species from two sites with a high (10.4%) detection rate. The ORF1ab, E, M, and N proteins of AFCoV had [ 90% aa identities to those of CGCoV which caused a die-off of Canada and snow geese in a site near same habitat. Bootscan analysis using genomes of avian coronavirus, IBV, CGCoV, and AFCoV (as the query sequence) indicated the absence of any obvious recombination event ( Supplementary Fig. S3 ). To summarize, this study describes a new member of Goose coronavirus CB17 species in migratory birds from China and promotes our understanding of the diversity of gammacoronaviruses. We did not have the permission to collect tissue samples, and the isolation of virus was unsuccessful. Therefore, the specific pathogenicity of AFCoV still needs further study. Discovery of a novel hepatovirus (phopivirus of seals) related to human hepatitis a virus Fulllength transcriptome assembly from RNA-Seq data without a reference genome Infectious bronchitis Characterization of turkey coronavirus from turkey poults with acute enteritis Global patterns of influenza a virus in wild birds Genome organization of Canada goose coronavirus, a novel species identified in a mass die-off of canada geese A comprehensive phylogeny of birds (Aves) using targeted next-generation DNA sequencing Birds, migration and emerging zoonoses: west nile virus, lyme disease, influenza A and enteropathogens Coronavirus: epidemiology, genome replication and the interactions with their hosts Discovery of seven novel Mammalian and avian coronaviruses in the genus deltacoronavirus supports bat coronaviruses as the gene source of alphacoronavirus and betacoronavirus and avian coronaviruses as the gene source of gammacoronavirus and deltacoronavirus Beta-and novel delta-coronaviruses are identified from wild animals in the Qinghai-Tibetan Plateau Conflict of interest The authors declare that they have no conflict of interest. The fecal samples of migratory birds were collected according the Chinese law and regulations and authorized by Forestry Bureau of Jiangxi Province of China. The study practices were approved by Nanchang Center for Diseases Prevention and Control.