While bats are increasingly recognized as a source of coronavirus epidemics, the diversity and emergence potential of bat coronaviruses remains to be fully understood. Among 1779 bat samples ...collected in China, diverse coronaviruses were detected in 32 samples from five different bat species by RT-PCR. Two novel alphacoronaviruses,
bat coronavirus HKU32 (Rs-BatCoV HKU32) and
bat coronavirus HKU33 (Tr-BatCoV HKU33), were discovered from Chinese horseshoe bats in Hong Kong and greater bamboo bats in Guizhou Province, respectively. Genome analyses showed that Rs-BatCoV HKU32 is closely related to BatCoV HKU10 and related viruses from diverse bat families, whereas Tr-BatCoV HKU33 is closely related to BtNv-AlphaCoV and similar viruses exclusively from bats of
family. The close relatedness of Rs-BatCoV HKU32 to BatCoV HKU10 which was also detected in Pomona roundleaf bats from the same country park suggests that these viruses may have the tendency of infecting genetically distant bat populations of close geographical proximity with subsequent genetic divergence. Moreover, the presence of SARSr-CoV ORF7a-like protein in Rs-BatCoV HKU32 suggests a common evolutionary origin of this accessory protein with SARS-CoV, also from Chinese horseshoe bats, an apparent reservoir for coronavirus epidemics. The emergence potential of Rs-BatCoV HKU32 should be explored.
Previous findings of Middle East Respiratory Syndrome coronavirus (MERS-CoV)-related viruses in bats, and the ability of Tylonycteris-BatCoV HKU4 spike protein to utilize MERS-CoV receptor, human ...dipeptidyl peptidase 4 hDPP4, suggest a bat ancestral origin of MERS-CoV. We developed 12 primary bat cell lines from seven bat species, including Tylonycteris pachypus, Pipistrellus abramus and Rhinolophus sinicus (hosts of Tylonycteris-BatCoV HKU4, Pipistrellus-BatCoV HKU5, and SARS-related-CoV respectively), and tested their susceptibilities to MERS-CoVs, SARS-CoV, and human coronavirus 229E (HCoV-229E). Five cell lines, including P. abramus and R. sinicus but not T. pachypus cells, were susceptible to human MERS-CoV EMC/2012. However, three tested camel MERS-CoV strains showed different infectivities, with only two strains capable of infecting three and one cell lines respectively. SARS-CoV can only replicate in R. sinicus cells, while HCoV-229E cannot replicate in any bat cells. Bat dipeptidyl peptidase 4 (DPP4) sequences were closely related to those of human and non-human primates but distinct from dromedary DPP4 sequence. Critical residues for binding to MERS-CoV spike protein were mostly conserved in bat DPP4. DPP4 was expressed in the five bat cells susceptible to MERS-CoV, with significantly higher mRNA expression levels than those in non-susceptible cells (P = 0.0174), supporting that DPP4 expression is critical for MERS-CoV infection in bats. However, overexpression of T. pachypus DPP4 failed to confer MERS-CoV susceptibility in T. pachypus cells, suggesting other cellular factors in determining viral replication. The broad cellular tropism of MERS-CoV should prompt further exploration of host diversity of related viruses to identify its ancestral origin.
The COVID-19 pandemic response has shown how vaccine platform technologies can be used to rapidly and effectively counteract a novel emerging infectious disease. The speed of development for mRNA and ...vector-based vaccines outpaced those of subunit vaccines, however, subunit vaccines can offer advantages in terms of safety and stability. Here we describe a subunit vaccine platform technology, the molecular clamp, in application to four viruses from divergent taxonomic families: Middle Eastern respiratory syndrome coronavirus (MERS-CoV), Ebola virus (EBOV), Lassa virus (LASV) and Nipah virus (NiV). The clamp streamlines subunit antigen production by both stabilising the immunologically important prefusion epitopes of trimeric viral fusion proteins while enabling purification without target-specific reagents by acting as an affinity tag. Conformations for each viral antigen were confirmed by monoclonal antibody binding, size exclusion chromatography and electron microscopy. Notably, all four antigens tested remained stable over four weeks of incubation at 40°C. Of the four vaccines tested, a neutralising immune response was stimulated by clamp stabilised MERS-CoV spike, EBOV glycoprotein and NiV fusion protein. Only the clamp stabilised LASV glycoprotein precursor failed to elicit virus neutralising antibodies. MERS-CoV and EBOV vaccine candidates were both tested in animal models and found to provide protection against viral challenge.
So far, dromedary camels are the only known animal reservoir for Middle East respiratory syndrome (MERS) coronavirus (MERS-CoV). Previous published serological studies showed that sera of Bactrian ...camels were all negative for MERS-CoV antibodies. However, a recent study revealed that direct inoculation of Bactrian camels intranasally with MERS-CoV can lead to infection with abundant virus shedding and seroconversion. In this study, we examined the presence of MERS-CoV antibodies in Bactrian and hybrid camels in Dubai, the United Arab Emirates (where dromedaries are also present), and Bactrian camels in Xinjiang, China (where dromedaries are absent). For the 29 serum samples from Bactrian camels in Dubai tested by the MERS-CoV spike (S) protein-based enzyme-linked immunosorbent assay (S-ELISA) and neutralization antibody test, 14 (48%) and 12 (41%), respectively, were positive for MERS-CoV antibodies. All the 12 serum samples that were positive with the neutralization antibody test were also positive for the S-ELISA. For the 11 sera from hybrid camels in Dubai tested with the S-ELISA and neutralization antibody test, 6 (55%) and 9 (82%), respectively, were positive for MERS-CoV antibodies. All the 6 serum samples that were positive for the S-ELISA were also positive with the neutralization antibody test. There was a strong correlation between the antibody levels detected by S-ELISA and neutralizing antibody titers, with a Spearman coefficient of 0.6262 (
< 0.0001; 95% confidence interval, 0.5062 to 0.7225). All 92 Bactrian camel serum samples from Xinjiang were negative for MERS-CoV antibodies tested using both S-ELISA and the neutralization antibody test. Bactrian and hybrid camels are potential sources of MERS-CoV infection.
Since its first appearance in 2012, Middle East respiratory syndrome (MERS) has affected >25 countries, with >2,400 cases and an extremely high fatality rate of >30%. The total number of mortalities due to MERS is already greater than that due to severe acute respiratory syndrome. MERS coronavirus (MERS-CoV) has been confirmed to be the etiological agent. So far, dromedaries are the only known animal reservoir for MERS-CoV. Previously published serological studies showed that sera of Bactrian camels were all negative for MERS-CoV antibodies. In this study, we observed that 41% of the Bactrian camel sera and 55% of the hybrid camel sera from Dubai (where dromedaries are also present), but none of the sera from Bactrian camels in Xinjiang (where dromedaries are absent), were positive for MERS-CoV antibodies. Based on these results, we conclude that in addition to dromedaries, Bactrian and hybrid camels are also potential sources of MERS-CoV infection.
Highlights • Respiratory virus coinfections cause severe disease in immunocompromised patients. • Such infections may be missed without the use of molecular diagnostics. • Intravenous immunoglobulin ...may be an effective therapeutic option in such patients.
Compared to other human coronaviruses, the genetic diversity and evolution of human coronavirus 229E (HCoV-229E) are relatively understudied. We report a fatal case of COVID-19 pneumonia coinfected ...with HCoV-229E in Hong Kong. Genome sequencing of SARS-CoV-2 and HCoV-229E from a nasopharyngeal sample of the patient showed that the SARS-CoV-2 strain HK13 was most closely related to SARS-CoV-2 type strain Wuhan-Hu-1 (99.99% nucleotide identity), compatible with his recent history of travel to Wuhan. The HCoV-229E strain HK20-42 was most closely related to HCoV-229E strain SC0865 from the United States (99.86% nucleotide identity). To investigate if it may represent a newly emerged HCoV-229E genotype in Hong Kong, we retrieved 41 archived respiratory samples that tested positive for HCoV-229E from 2004 to 2019. Pneumonia and exacerbations of chronic airway diseases were common among infected patients. Complete RdRp, S, and N gene sequencing of the 41 HCoV-229E strains revealed that our contemporary HCoV-229E strains have undergone significant genetic drift with clustering of strains in chronological order. Two novel genogroups were identified, in addition to previously described genogroups 1 to 4, with recent circulating strains including strain HK20-42 belonging to novel genogroup 6. Positive selection was detected in the spike protein and receptor-binding domain, which may be important for viral evolution at the receptor-binding interphase. Molecular dating analysis showed that HCoV-229E shared the most recent common ancestor with bat and camel/alpaca 229E-related viruses at ∼1884, while camel/alpaca viruses had a relatively recent common ancestor at ∼1999. Further studies are required to ascertain the evolutionary origin and path of HCoV-229E.
Since its first appearance in the 1960s, the genetic diversity and evolution of human coronavirus 229E (HCoV-229E) have been relatively understudied. In this study, we report a fatal case of COVID-19 coinfected with HCoV-229E in Hong Kong. Genome sequencing revealed that our SARS-CoV-2 strain is highly identical to the SARS-CoV-2 strain from Wuhan, compatible with the patient's recent travel history, whereas our HCoV-229E strain in this study is highly identical to a recent strain in the United States. We also retrieved 41 archived HCoV-229E strains from 2004 to 2019 in Hong Kong for sequence analysis. Pneumonia and exacerbations of chronic airway diseases were common diagnoses among the 41 patients. The results showed that HCoV-229E was evolving in chronological order. Two novel genogroups were identified in addition to the four preexisting HCoV-229E genogroups, with recent circulating strains belonging to novel genogroup 6. Molecular clock analysis dated bat-to-human and bat-to-camelid transmission to as early as 1884.
In addition to human cases, cases of COVID-19 in captive animals and pets are increasingly reported. This raises the concern for two-way COVID-19 transmission between humans and animals. Here, we ...developed a SARS-CoV-2 nucleocapsid protein-based competitive enzyme-linked immunosorbent assay (cELISA) for serodiagnosis of COVID-19 which can theoretically be used in virtually all kinds of animals. We used 187 serum samples from patients with/without COVID-19, laboratory animals immunized with inactive SARS-CoV-2 virions, COVID-19-negative animals, and animals seropositive to other betacoronaviruses. A cut-off percent inhibition value of 22.345% was determined and the analytical sensitivity and specificity were found to be 1:64–1:256 and 93.9%, respectively. Evaluation on its diagnostic performance using 155 serum samples from COVID-19-negative animals and COVID-19 human patients showed a diagnostic sensitivity and specificity of 80.8% and 100%, respectively. The cELISA can be incorporated into routine blood testing of farmed/captive animals for COVID-19 surveillance.
We conducted a six-year epidemiological study on human coronaviruses（HCoVs） circulating in Hong Kong, using 8275 nasopharyngeal samples from patients with acute respiratory tract infections. HCoVs ...were detected in 77（0.93%） of the samples by a pan-HCoV RT-PCR assay. The most frequently detected HCoV species was HCoV-OC43（0.58%）, followed by HCoV-229E（0.15%）,HCoV-HKU1（0.13%） and HCoV-NL63（0.07%）. HCoVs were detected throughout the study period（September 2008–August 2014）, with the highest detection rate from September 2010 to August2011（22/1500, 1.47%）. Different seasonal patterns of each HCoV species in Hong Kong were noted.HCoV-OC43 was predominant in the fall and winter, whereas HCoV-HKU1 showed peak activity in winter, with a few cases occurred in spring and summer. HCoV-229 E mainly occurred in winter and spring, while HCoV-NL63 was predominant in summer and autumn. HCoVs most commonly infect the elderly and young children, with median age of 79.5 years（range, 22 days to 95 years）.Intriguingly, the detection rate of HCoV-OC43 in the age group of ＆gt; 80 years（26/2380, 1.09%） was significantly higher than that in the age group of 0–10 years（12/2529, 0.47%）（P 〈 0.05）. These data provides new insight into the epidemiology of coronaviruses.
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