Does Avian Coronavirus Co-Circulate with Avian Paramyxovirus and Avian Influenza Virus in Wild Ducks in Siberia?

Avian coronaviruses (ACoV) have been shown to be highly prevalent in wild bird populations. More work on avian coronavirus detection and diversity estimation is needed for the breeding territories of migrating birds, where the high diversity and high prevalence of Orthomyxoviridae and Paramyxoviridae have already been shown in wild birds. In order to detect ACoV RNA, we conducted PCR diagnostics of cloacal swab samples from birds, which we monitored during avian influenza A virus surveillance activities. Samples from two distant Asian regions of Russia (Sakhalin region and Novosibirsk region) were tested. Amplified fragments of the RNA-dependent RNA-polymerase (RdRp) of positive samples were partially sequenced to determine the species of Coronaviridae represented. The study revealed a high presence of ACoV among wild birds in Russia. Moreover, there was a high presence of birds co-infected with avian coronavirus, avian influenza virus, and avian paramyxovirus. We found one case of triple co-infection in a Northern Pintail (Anas acuta). Phylogenetic analysis revealed the circulation of a Gammacoronavirus species. A Deltacoronavirus species was not detected, which supports the data regarding the low prevalence of deltacoronaviruses among surveyed bird species.

Synchronicity of viral shedding in molossid bat maternity colonies.

Infection dynamics in vertebrates are driven by biological and ecological processes. For bats, population structure and reproductive cycles have major effects on RNA virus transmission. On Reunion Island, previous studies have shown that parturition of pregnant females and aggregation of juvenile Reunion free-tailed bats (Mormopterus francoismoutoui) are associated with major increase in the prevalence of bats shedding RNA viruses. The synchronicity of such shedding pulses, however, is yet to be assessed between viruses but also maternity colonies. Based on 3422 fresh faeces collected every 2-5 weeks during four consecutive birthing seasons, we report the prevalence of bats shedding astroviruses (AstVs), coronaviruses (CoVs) and paramyxoviruses (PMVs) in two maternity colonies on Reunion Island. We found that the proportion of bats shedding viruses is highly influenced by sampling collection periods, and therefore by the evolution of the population age structure. We highlight that virus shedding patterns are consistent among years and colonies for CoVs and to a lesser extent for PMVs, but not for AstVs. We also report that 1% of bats harbour co-infections, with two but not three of the viruses, and most co-infections were due to CoVs and PMVs.

One Health Surveillance Highlights Circulation of Viruses with Zoonotic Potential in Bats, Pigs, and Humans in Viet Nam.

A One Health cross-sectoral surveillance approach was implemented to screen biological samples from bats, pigs, and humans at high-risk interfaces for zoonotic viral spillover for five viral families with zoonotic potential in Viet Nam. Over 1600 animal and human samples from bat guano harvesting sites, natural bat roosts, and pig farming operations were tested for coronaviruses (CoVs), paramyxoviruses, influenza viruses, filoviruses and flaviviruses using consensus PCR assays. Human samples were also tested using immunoassays to detect antibodies against eight virus groups. Significant viral diversity, including CoVs closely related to ancestors of pig pathogens, was detected in bats roosting at the human-animal interfaces, illustrating the high risk for CoV spillover from bats to pigs in Viet Nam, where pig density is very high. Season and reproductive period were significantly associated with the detection of bat CoVs, with site-specific effects. Phylogeographic analysis indicated localized viral transmission among pig farms. Our limited human sampling did not detect any known zoonotic bat viruses in human communities living close to the bat cave and harvesting bat guano, but our serological assays showed possible previous exposure to Marburg virus-like (Filoviridae), Crimean-Congo hemorrhagic fever virus-like (Bunyaviridae) viruses and flaviviruses. Targeted and coordinated One Health surveillance helped uncover this viral pathogen emergence hotspot.

The characterization of multiple novel paramyxoviruses highlights the diverse nature of the subfamily Orthoparamyxovirinae.

The subfamily Orthoparamyxovirinae is a group of single-stranded, negative-sense RNA viruses that contains many human, animal, and zoonotic pathogens. While there are currently only forty-two recognized species in this subfamily, recent research has revealed that much of its diversity remains to be characterized. Using a newly developed nested PCR-based screening assay, we report here the discovery of fifteen orthoparamyxoviruses in rodents and shrews from Belgium and Guinea, thirteen of which are believed to represent new species. Using a combination of nanopore and sanger sequencing, complete genomes could be determined for almost all these viruses, enabling a detailed evaluation of their genome characteristics. While most viruses are thought to belong to the rapidly expanding genus Jeilongvirus, we also identify novel members of the genera Narmovirus, Henipavirus, and Morbillivirus. Together with other recently discovered orthoparamyxoviruses, both henipaviruses and the morbillivirus discovered here appear to form distinct rodent-/shrew-borne clades within their respective genera, clustering separately from all currently classified viruses. In the case of the henipaviruses, a comparison of the different members of this clade revealed the presence of a secondary conserved open reading frame, encoding for a transmembrane protein, within the F gene, the biological relevance of which remains to be established. While the characteristics of the viruses described here shed further light on the complex evolutionary origin of paramyxoviruses, they also illustrate that the diversity of this group of viruses in terms of genome organization appears to be much larger than previously assumed.

Coronavirus and Paramyxovirus Shedding by Bats in a Cave and Buildings in Ethiopia.

Bats are important hosts of zoonotic viruses with pandemic potential, including filoviruses, MERS-Coronavirus (CoV), SARS-CoV -1, and likely SARS-CoV-2. Viral infection and transmission among wildlife are dependent on a combination of factors that include host ecology and immunology, life history traits, roosting habitats, biogeography, and external stressors. Between 2016 and 2018, four species of insectivorous bats from a readily accessed roadside cave and buildings in Ethiopia were sampled and tested for viruses using consensus PCR assays for five viral families/genera. Previously identified and novel coronaviruses and paramyxoviruses were identified in 99 of the 589 sampled bats. Bats sampled from the cave site were more likely to test positive for a CoV than bats sampled from buildings; viral shedding was more common in the wet season; and rectal swabs were the most common sample type to test positive. A previously undescribed alphacoronavirus was detected in two bat species from different taxonomic families, sampling interfaces, geographic locations, and years. These findings expand knowledge of the range and diversity of coronaviruses and paramyxoviruses in insectivorous bats in Ethiopia and reinforce that an improved understanding of viral diversity and species-specific shedding dynamics is important for designing informed zoonotic disease surveillance and spillover risk reduction efforts.

Behavioral–biological surveillance of emerging infectious diseases among a dynamic cohort in Thailand

Background Interactions between humans and animals are the key elements of zoonotic spillover leading to zoonotic disease emergence. Research to understand the high-risk behaviors associated with disease transmission at the human-animal interface is limited, and few consider regional and local contexts. Objective This study employed an integrated behavioral–biological surveillance approach for the early detection of novel and known zoonotic viruses in potentially high-risk populations, in an effort to identify risk factors for spillover and to determine potential foci for risk-mitigation measures. Method Participants were enrolled at two community-based sites (n = 472) in eastern and western Thailand and two hospital (clinical) sites (n = 206) in northeastern and central Thailand. A behavioral questionnaire was administered to understand participants’ demographics, living conditions, health history, and animal-contact behaviors and attitudes. Biological specimens were tested for coronaviruses, filoviruses, flaviviruses, influenza viruses, and paramyxoviruses using pan (consensus) RNA Virus assays. Results Overall 61/678 (9%) of participants tested positive for the viral families screened which included influenza viruses (75%), paramyxoviruses (15%), human coronaviruses (3%), flaviviruses (3%), and enteroviruses (3%). The most salient predictors of reporting unusual symptoms (i.e., any illness or sickness that is not known or recognized in the community or diagnosed by medical providers) in the past year were having other household members who had unusual symptoms and being scratched or bitten by animals in the same year. Many participants reported raising and handling poultry (10.3% and 24.2%), swine (2%, 14.6%), and cattle (4.9%, 7.8%) and several participants also reported eating raw or undercooked meat of these animals (2.2%, 5.5%, 10.3% respectively). Twenty four participants (3.5%) reported handling bats or having bats in the house roof. Gender, age, and livelihood activities were shown to be significantly associated with participants’ interactions with animals. Participants’ knowledge of risks influenced their health-seeking behavior. Conclusion The results suggest that there is a high level of interaction between humans, livestock, and wild animals in communities at sites we investigated in Thailand. This study highlights important differences among demographic and occupational risk factors as they relate to animal contact and zoonotic disease risk, which can be used by policymakers and local public health programs to build more effective surveillance strategies and behavior-focused interventions.

Behavioral-biological surveillance of emerging infectious diseases among a dynamic cohort in Thailand.

BACKGROUND: Interactions between humans and animals are the key elements of zoonotic spillover leading to zoonotic disease emergence. Research to understand the high-risk behaviors associated with disease transmission at the human-animal interface is limited, and few consider regional and local contexts. OBJECTIVE: This study employed an integrated behavioral-biological surveillance approach for the early detection of novel and known zoonotic viruses in potentially high-risk populations, in an effort to identify risk factors for spillover and to determine potential foci for risk-mitigation measures. METHOD: Participants were enrolled at two community-based sites (n = 472) in eastern and western Thailand and two hospital (clinical) sites (n = 206) in northeastern and central Thailand. A behavioral questionnaire was administered to understand participants' demographics, living conditions, health history, and animal-contact behaviors and attitudes. Biological specimens were tested for coronaviruses, filoviruses, flaviviruses, influenza viruses, and paramyxoviruses using pan (consensus) RNA Virus assays. RESULTS: Overall 61/678 (9%) of participants tested positive for the viral families screened which included influenza viruses (75%), paramyxoviruses (15%), human coronaviruses (3%), flaviviruses (3%), and enteroviruses (3%). The most salient predictors of reporting unusual symptoms (i.e., any illness or sickness that is not known or recognized in the community or diagnosed by medical providers) in the past year were having other household members who had unusual symptoms and being scratched or bitten by animals in the same year. Many participants reported raising and handling poultry (10.3% and 24.2%), swine (2%, 14.6%), and cattle (4.9%, 7.8%) and several participants also reported eating raw or undercooked meat of these animals (2.2%, 5.5%, 10.3% respectively). Twenty four participants (3.5%) reported handling bats or having bats in the house roof. Gender, age, and livelihood activities were shown to be significantly associated with participants' interactions with animals. Participants' knowledge of risks influenced their health-seeking behavior. CONCLUSION: The results suggest that there is a high level of interaction between humans, livestock, and wild animals in communities at sites we investigated in Thailand. This study highlights important differences among demographic and occupational risk factors as they relate to animal contact and zoonotic disease risk, which can be used by policymakers and local public health programs to build more effective surveillance strategies and behavior-focused interventions.

Surveillance for potentially zoonotic viruses in rodent and bat populations and behavioral risk in an agricultural settlement in Ghana.

BACKGROUND: In Ghana, the conversion of land to agriculture, especially across the vegetative belt has resulted in fragmented forest landscapes with increased interactions among humans, domestic animals, and wildlife. METHODS: We investigated viruses in bats and rodents, key reservoir hosts for zoonotic viral pathogens, in a small agricultural community in the vegetation belt of Ghana. We also administered questionnaires among the local community members to learn more about people's awareness and perceptions of zoonotic disease risks and the environmental factors and types of activities in which they engage that might influence pathogen transmission from wildlife. RESULTS: Our study detected the RNA from paramyxoviruses and coronaviruses in rodents and bats, including sequences from novel viruses with unknown zoonotic potential. Samples collected from Epomophorus gambianus bats were significantly more likely to be positive for coronavirus RNA during the rainy season, when higher numbers of young susceptible individuals are present in the population. Almost all community members who responded to the questionnaire reported contact with wildlife, especially bats, rodents, and non-human primates in and around their homes and in the agricultural fields. Over half of the respondents were not aware or did not perceive any zoonotic disease risks associated with close contact with animals, such as harvesting and processing animals for food. To address gaps in awareness and mitigation strategies for pathogen transmission risks, we organized community education campaigns using risk reduction and outreach tools focused around living safely with bats and rodents. CONCLUSIONS: These findings expand our knowledge of the viruses circulating in bats and rodents in Ghana and of the beliefs, perceptions, and practices that put community members at risk of zoonotic virus spillover through direct and indirect contact with bats and rodents. This study also highlights the importance of community engagement in research and interventions focused on mitigating risk and living safely with wildlife.

Poxviruses and paramyxoviruses use a conserved mechanism of STAT1 antagonism to inhibit interferon signaling.

The induction of interferon (IFN)-stimulated genes by STATs is a critical host defense mechanism against virus infection. Here, we report that a highly expressed poxvirus protein, 018, inhibits IFN-induced signaling by binding to the SH2 domain of STAT1, thereby preventing the association of STAT1 with an activated IFN receptor. Despite encoding other inhibitors of IFN-induced signaling, a poxvirus mutant lacking 018 was attenuated in mice. The 2.0 Å crystal structure of the 018:STAT1 complex reveals a phosphotyrosine-independent mode of 018 binding to the SH2 domain of STAT1. Moreover, the STAT1-binding motif of 018 shows similarity to the STAT1-binding proteins from Nipah virus, which, similar to 018, block the association of STAT1 with an IFN receptor. Overall, these results uncover a conserved mechanism of STAT1 antagonism that is employed independently by distinct virus families.

Broad-Spectrum In Vitro Antiviral Activity of ODBG-P-RVn: An Orally-Available, Lipid-Modified Monophosphate Prodrug of Remdesivir Parent Nucleoside (GS-441524).

The necessity for intravenous administration of remdesivir confines its utility for treatment of coronavirus disease 2019 (COVID-19) to hospitalized patients. We evaluated the broad-spectrum antiviral activity of ODBG-P-RVn, an orally available, lipid-modified monophosphate prodrug of the remdesivir parent nucleoside (GS-441524), against viruses that cause diseases of human public health concern, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). ODBG-P-RVn showed 20-fold greater antiviral activity than GS-441524 and had activity nearly equivalent to that of remdesivir in primary-like human small airway epithelial cells. Our results warrant in vivo efficacy evaluation of ODBG-P-RVn. IMPORTANCE While remdesivir remains one of the few drugs approved by the FDA to treat coronavirus disease 2019 (COVID-19), its intravenous route of administration limits its use to hospital settings. Optimizing the stability and absorption of remdesivir may lead to a more accessible and clinically potent therapeutic. Here, we describe an orally available lipid-modified version of remdesivir with activity nearly equivalent to that of remdesivir against emerging viruses that cause significant disease, including Ebola and Nipah viruses. Our work highlights the importance of such modifications to optimize drug delivery to relevant and appropriate human tissues that are most affected by such diseases.

Discovery and Genetic Characterization of Novel Paramyxoviruses Related to the Genus Henipavirus in Crocidura Species in the Republic of Korea.

Paramyxoviruses, negative-sense single-stranded RNA viruses, pose a critical threat to human public health. Currently, 78 species, 17 genera, and 4 subfamilies of paramyxoviruses are harbored by multiple natural reservoirs, including rodents, bats, birds, reptiles, and fish. Henipaviruses are critical zoonotic pathogens that cause severe acute respiratory distress and neurological diseases in humans. Using reverse transcription-polymerase chain reaction, 115 Crocidura species individuals were examined for the prevalence of paramyxovirus infections. Paramyxovirus RNA was observed in 26 (22.6%) shrews collected at five trapping sites, Republic of Korea. Herein, we report two genetically distinct novel paramyxoviruses (genus: Henipavirus): Gamak virus (GAKV) and Daeryong virus (DARV) isolated from C. lasiura and C. shantungensis, respectively. Two GAKVs and one DARV were nearly completely sequenced using next-generation sequencing. GAKV and DARV contain six genes (3'-N-P-M-F-G-L-5') with genome sizes of 18,460 nucleotides and 19,471 nucleotides, respectively. The phylogenetic inference demonstrated that GAKV and DARV form independent genetic lineages of Henipavirus in Crocidura species. GAKV-infected human lung epithelial cells elicited the induction of type I/III interferons, interferon-stimulated genes, and proinflammatory cytokines. In conclusion, this study contributes further understandings of the molecular prevalence, genetic characteristics and diversity, and zoonotic potential of novel paramyxoviruses in shrews.

Cospeciation of coronavirus and paramyxovirus with their bat hosts in the same geographical areas.

BACKGROUND: Bat-borne viruses are relatively host specific. We hypothesize that this host specificity is due to coevolution of the viruses with their hosts. To test this hypothesis, we investigated the coevolution of coronavirus and paramyxovirus with their bat hosts. Published nucleotide sequences of the RNA-dependent RNA polymerase (RdRp) gene of 60 coronavirus strains identified from 37 bat species, the RNA polymerase large (L) gene of 36 paramyxovirus strains from 29 bat species, and the cytochrome B (cytB) gene of 35 bat species were analyzed for coevolution signals. Each coevolution signal detected was tested and verified by global-fit cophylogenic analysis using software ParaFit, PACo, and eMPRess. RESULTS: Significant coevolution signals were detected in coronaviruses and paramyxoviruses and their bat hosts, and closely related bat hosts were found to carry closely related viruses. CONCLUSIONS: Our results suggest that paramyxovirus and coronavirus coevolve with their hosts.

Comparative Protective Efficacies of Novel Avian Paramyxovirus-Vectored Vaccines against Virulent Infectious Bronchitis Virus in Chickens.

Viral vectored vaccines are desirable alternatives for conventional infectious bronchitis virus (IBV) vaccines. We have recently shown that a recombinant Newcastle disease virus (rNDV) strain LaSota expressing the spike (S) protein of IBV strain Mass-41 (rLaSota/IBV-S) was a promising vaccine candidate for IBV. Here we evaluated a novel chimeric rNDV/avian paramyxovirus serotype 2 (rNDV/APMV-2) as a vaccine vector against IBV. The rNDV/APMV-2 vector was chosen because it is much safer than the rNDV strain LaSota vector, particularly for young chicks and chicken embryos. In order to determine the effectiveness of this vector, a recombinant rNDV/APMV-2 expressing the S protein of IBV strain Mass-41 (rNDV/APMV-2/IBV-S) was constructed. The protective efficacy of this vector vaccine was compared to that of the rNDV vector vaccine. In one study, groups of one-day-old specific-pathogenic-free (SPF) chickens were immunized with rLaSota/IBV-S and rNDV/APMV-2/IBV-S and challenged four weeks later with the homologous highly virulent IBV strain Mass-41. In another study, groups of broiler chickens were single (at day one or three weeks of age) or prime-boost (prime at day one and boost at three weeks of age) immunized with rLaSota/IBV-S and/or rNDV-APMV-2/IBV-S. At weeks six of age, chickens were challenged with a highly virulent IBV strain Mass-41. Our challenge study showed that novel rNDV/APMV-2/IBV-S provided similar protection as rLaSota/IBV-S in SPF chickens. However, compared to prime-boost immunization of chickens with chimeric rNDV/APMV-2, rLaSota/IBV-S and/or a live IBV vaccine, single immunization of chickens with rLaSota/IBV-S, or live IBV vaccine provided better protection against IBV. In conclusion, we have developed the novel rNDV/APMV-2 vector expressing S protein of IBV that can be a safer vaccine against IB in chickens. Our results also suggest a single immunization with a LaSota vectored IBV vaccine candidate provides better protection than prime-boost immunization regimens.

Nipah@20: Lessons Learned from Another Virus with Pandemic Potential.

Nipah disease is listed as one of the WHO priority diseases that pose the greatest public health risk due to their epidemic potential. More than 200 experts from around the world convened in Singapore last year to mark the 20th anniversary of the first Nipah virus outbreaks in Malaysia and Singapore. Most of these experts are now involved in responding to the coronavirus disease 2019 (COVID-19) pandemic. Here, members of the Organizing Committee of the 2019 Nipah Virus International Conference review highlights from the Nipah@20 Conference and reflect on key lessons learned from Nipah that could be applied to the understanding of the COVID-19 pandemic and to preparedness against future emerging infectious diseases (EIDs) of pandemic potential.