e Recently, we reported the discovery of three novel coronaviruses, bulbul coronavirus HKU11, thrush coronavirus HKU12, and munia coronavirus HKU13, which were identified as representatives of a novel genus, Deltacoronavirus, in the subfamily Coronavirinae. In this territory-wide molecular epidemiology study involving 3,137 mammals and 3,298 birds, we discovered seven additional novel deltacoronaviruses in pigs and birds, which we named porcine coronavirus HKU15, white-eye coronavirus HKU16, sparrow coronavirus HKU17, magpie robin coronavirus HKU18, night heron coronavirus HKU19, wigeon coronavirus HKU20, and common moorhen coronavirus HKU21. Complete genome sequencing and comparative genome analysis showed that the avian and mammalian deltacoronaviruses have similar genome characteristics and structures. They all have relatively small genomes (25.421 to 26.674 kb), the smallest among all coronaviruses. They all have a single papain-like protease domain in the nsp3 gene; an accessory gene, NS6 open reading frame (ORF), located between the M and N genes; and a variable number of accessory genes (up to four) downstream of the N gene. Moreover, they all have the same putative transcription regulatory sequence of ACACCA. Molecular clock analysis showed that the most recent common ancestor of all coronaviruses was estimated at approximately 8100 BC, and those of Alphacoronavirus, Betacoronavirus, Gammacoronavirus, and Deltacoronavirus were at approximately 2400 BC, 3300 BC, 2800 BC, and 3000 BC, respectively. From our studies, it appears that bats and birds, the warm blooded flying vertebrates, are ideal hosts for the coronavirus gene source, bats for Alphacoronavirus and Betacoronavirus and birds for Gammacoronavirus and Deltacoronavirus, to fuel coronavirus evolution and dissemination.

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Despite extensive laboratory investigations in patients with respiratory tract infections, no microbiological cause can be identified in a significant proportion of patients. In the past 3 years, several novel respiratory viruses, including human metapneumovirus, severe acute respiratory syndrome (SARS) coronavirus (SARSCoV), and human coronavirus NL63, were discovered. Here we report the discovery of another novel coronavirus, coronavirus HKU1 (CoV-HKU1), from a 71-year-old man with pneumonia who had just returned from Shenzhen, China. Quantitative reverse transcription-PCR showed that the amount of CoV-HKU1 RNA was 8.5 to 9.6 ؋ 10 6 copies per ml in his nasopharyngeal aspirates (NPAs) during the first week of the illness and dropped progressively to undetectable levels in subsequent weeks. He developed increasing serum levels of specific antibodies against the recombinant nucleocapsid protein of CoV-HKU1, with immunoglobulin M (IgM) titers of 1:20, 1:40, and 1:80 and IgG titers of <1:1,000, 1:2,000, and 1:8,000 in the first, second and fourth weeks of the illness, respectively. Isolation of the virus by using various cell lines, mixed neuron-glia culture, and intracerebral inoculation of suckling mice was unsuccessful. The complete genome sequence of CoV-HKU1 is a 29,926-nucleotide, polyadenylated RNA, with G؉C content of 32%, the lowest among all known coronaviruses with available genome sequence. Phylogenetic analysis reveals that CoV-HKU1 is a new group 2 coronavirus. Screening of 400 NPAs, negative for SARS-CoV, from patients with respiratory illness during the SARS period identified the presence of CoV-HKU1 RNA in an additional specimen, with a viral load of 1.13 ؋ 10 6 copies per ml, from a 35-year-old woman with pneumonia. Our data support the existence of a novel group 2 coronavirus associated with pneumonia in humans.Since no microbiological cause can be identified for a significant proportion of patients with respiratory tract infections (18, 29), research has been conducted to identify novel agents. Of the three novel agents identified in recent 3 years, including human metapneumovirus (36), severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV) (25), and human coronavirus NL63 (HCoV-NL63) (6, 37), two were coronaviruses. Coronaviruses possess the largest genomes of all RNA viruses, consisting of about 30 kb. As a result of their unique mechanism of viral replication, coronaviruses have a high frequency of recombination.Based on genotypic and serological characterization, coronaviruses were divided into three distinct groups, with human coronavirus 229E (HCoV-229E) being a group 1 coronavirus and human coronavirus OC43 (HCoV-OC43) being a group 2 coronavirus (16). They account for 5 to 30% of human respiratory tract infections. In late 2002 and 2003, the epidemic caused by SARS-CoV affected more than 8,000 people with 750 deaths (23-25, 44, 45, 51). We have also reported the isolation of SARS-CoV-like viruses from Himalayan palm civets, which suggested that animals could be the reservoir fo...