Molecular identification and characterization of novel coronaviruses infecting graylag geese (Anser anser), feral pigeons (Columbia livia) and mallards (Anas platyrhynchos)

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Abstract
In light of the finding of a previously unknown coronavirus as the aetiology of the severe acute respiratory syndrome (SARS), it is probable that other coronaviruses, than those recognized to date, are circulating in animal populations. Here, the results of a screening for coronavirus are presented, using a universal coronavirus RT-PCR, of the bird species graylag goose (Anser anser), feral pigeon (Columbia livia) and mallard (Anas platyrhynchos). Coronaviruses were found in cloacal swab samples from all the three bird species. In the graylag goose, 40 of 163 sampled birds were coronavirus positive, whereas two of 100 sampled pigeons and one of five sampled mallards tested positive. The infected graylag geese showed lower body weights compared with virus-negative birds, suggesting clinical significance of the infection. Phylogenetic analyses performed on the replicase gene and nucleocapsid protein sequences, indicated that the novel coronaviruses described in the present study all branch off from group III coronaviruses. All the novel avian coronaviruses harboured the conserved s2m RNA structure in their 3′ untranslated region, like other previously described group III coronaviruses, and like the SARS coronavirus. Sequencing of the complete nucleocapsid gene and downstream regions of goose and pigeon coronaviruses, evidenced the presence of two additional open reading frames for the goose coronavirus with no sequence similarity to known proteins, but with predicted transmembrane domains for one of the encoded proteins, and one additional open reading frame for the pigeon coronavirus, with a predicted transmembrane domain, downstream of the nucleocapsid gene.

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