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BioRxiv : the Preprint Server For... Sep 2023Small to mid-sized carnivores, or meso-carnivores, comprise a group of diverse mammals, many of which can adapt to anthropogenically disturbed environments. Wild...
Small to mid-sized carnivores, or meso-carnivores, comprise a group of diverse mammals, many of which can adapt to anthropogenically disturbed environments. Wild meso-carnivores living in urban areas may get exposed to or spread pathogens to other species, including stray/feral domestic animals. Several coronaviruses (CoVs) have been detected in domesticated and farmed meso-carnivores, but knowledge of CoVs circulating in free-ranging wild meso-carnivores remains limited. In this study, we analyzed 321 samples collected between 2016 and 2022 from 9 species of free-ranging wild meso-carnivores and stray/feral domestic cats in the northeastern United States. Using a pan-CoV PCR, we screened tissues, feces, and saliva, nasal, and rectal swabs. We detected CoV RNA in fecal and saliva samples of animals in four species: fisher (), bobcat (), red fox (), and domestic cat . Next-generation sequencing revealed that all these viruses belonged to the subgenus ( genus), previously reported only in rodents and lagomorphs (i.e., rabbits). Genetic comparison of the 3'-end of the genome (~12,000bp) revealed that although the viruses detected group with, and have a genetic organization similar to other luchacoviruses, they are genetically distinct from those from rodents and lagomorphs. Genetic characterization of the spike protein revealed that the meso-carnivore luchacoviruses do not have an S1/S2 cleavage motif but do have highly variable structural loops containing cleavage motifs similar to those identified in certain pathogenic CoVs. This study highlights the importance of characterizing the spike protein of CoVs in wild species for further targeted epidemiologic monitoring.
PubMed: 37745528
DOI: 10.1101/2023.05.31.541188 -
Viruses Sep 2023Although the involvement of the ubiquitin-proteasome system (UPS) in several coronavirus-productive infections has been reported, whether the UPS is required for...
Although the involvement of the ubiquitin-proteasome system (UPS) in several coronavirus-productive infections has been reported, whether the UPS is required for infectious bronchitis virus (IBV) and porcine epidemic diarrhea virus (PEDV) infections is unclear. In this study, the role of UPS in the IBV and PEDV life cycles was investigated. When the UPS was suppressed by pharmacological inhibition at the early infection stage, IBV and PEDV infectivity were severely impaired. Further study showed that inhibition of UPS did not change the internalization of virus particles; however, by using R18 and DiOC-labeled virus particles, we found that inhibition of UPS prevented the IBV and PEDV membrane fusion with late endosomes or lysosomes. In addition, proteasome inhibitors blocked the degradation of the incoming viral protein N, suggesting the uncoating process and genomic RNA release were suppressed. Subsequently, the initial translation of genomic RNA was blocked. Thus, UPS may target the virus-cellular membrane fusion to facilitate the release of incoming viruses from late endosomes or lysosomes, subsequently blocking the following virus uncoating, initial translation, and replication events. Similar to the observation of proteasome inhibitors, ubiquitin-activating enzyme E1 inhibitor PYR-41 also impaired the entry of IBV, enhanced the accumulation of ubiquitinated proteins, and depleted mono-ubiquitin. In all, this study reveals an important role of UPS in coronavirus entry by preventing membrane fusion and identifies UPS as a potential target for developing antiviral therapies for coronavirus.
Topics: Animals; Swine; Proteasome Endopeptidase Complex; Cell Line; Ubiquitin; Coronavirus; Proteasome Inhibitors; Membrane Fusion; Coronavirus Infections; Endosomes; Porcine epidemic diarrhea virus; RNA; Virus Replication
PubMed: 37896778
DOI: 10.3390/v15102001 -
International Journal of Biological... Jan 2024Porcine epidemic diarrhea virus (PEDV) infection causes immunosuppression and clinical symptoms such as vomiting, watery diarrhea, dehydration, and even death in...
Porcine epidemic diarrhea virus (PEDV) infection causes immunosuppression and clinical symptoms such as vomiting, watery diarrhea, dehydration, and even death in piglets. TRIM28, an E3 ubiquitin ligase, is involved in the regulation of autophagy. However, the role of TRIM28 in PEDV infection is unknown. This study aimed to determine whether TRIM28 acts as a host factor for PEDV immune escape. We found that depletion of TRIM28 inhibited PEDV replication, whereas overexpression of TRIM28 promoted the viral replication in host cells. Furthermore, knockdown of TRIM28 reversed PEDV-induced downregulation of the JAK/STAT1 pathway. Treatment with the mitophagic activator carbonyl cyanide 3-chlorophenylhydrazone (CCCP) attenuated the activating effect of TRIM28 depletion on the expression of the STAT1 pathway-related proteins. Treatment with CCCP also reduced the nuclear translocation of pSTAT1. Moreover, TRIM28, via its RING domain, interacted with PEDV N. Overexpression of TRIM28 induced mitophagy, which could be enhanced by co-expression with PEDV N. The results indicate that PEDV infection upregulates the expression of TRIM28, which induces mitophagy, leading to inhibition of the JAK-STAT1 pathway. This research unveils a new mechanism by which PEDV can hijack host cellular TRIM28 to promote its own replication.
Topics: Animals; Swine; Chlorocebus aethiops; Porcine epidemic diarrhea virus; Mitophagy; Carbonyl Cyanide m-Chlorophenyl Hydrazone; Coronavirus Infections; Virus Replication; Vero Cells
PubMed: 37907173
DOI: 10.1016/j.ijbiomac.2023.127722 -
Virus Research Sep 2023Coronaviruses (CoVs) are responsible for sporadic, epidemic and pandemic respiratory diseases worldwide. Bats have been identified as the reservoir for CoVs. To increase...
Coronaviruses (CoVs) are responsible for sporadic, epidemic and pandemic respiratory diseases worldwide. Bats have been identified as the reservoir for CoVs. To increase the number of complete coronavirus genomes in Africa and to comprehend the molecular epidemiology of bat Alphacoronaviruses (AlphaCoVs), we used deep metagenomics shotgun sequencing to obtain three (3) near-complete genomes of AlphaCoVs from Mops condylurus (Angolan free-tailed) bat in Nigeria. Phylogenetic and pairwise identity analysis of open reading frame 1ab (ORF1ab), spike (S), envelope (E), membrane (M) and nucleocapsid (N) genes of AlphaCoV in this study to previously described AlphaCoVs subgenera showed that the Nigerian AlphaCoVs may be members of potentially unique AlphaCoV subgenera circulating exclusively in bats in the Molossidae bat family. Recombination events were detected, suggesting the evolution of AlphaCoVs within the Molossidae family. The pairwise identity of the S gene in this study and previously published S gene sequences of other AlphaCoVs indicate that the Nigerian strains may have a genetically unique spike protein that is distantly related to other AlphaCoVs. Variations involving non-polar to polar amino acid substitution in both the Heptad Repeat (HR) regions 1 and 2 were observed. Further monitoring of bats to understand the host receptor use requirements of CoVs and interspecies CoV transmission in Africa is necessary to identify and prevent the potential danger that bat CoVs pose to public health.
Topics: Animals; Alphacoronavirus; Chiroptera; Phylogeny; Nigeria; Genome, Viral; Coronavirus; Coronavirus Infections; Genomics
PubMed: 37467933
DOI: 10.1016/j.virusres.2023.199174 -
Nature Communications Feb 2024Cepharanthine is a secondary metabolite isolated from Stephania. It has been reported that it has anti-conronaviruses activities including severe acute respiratory...
Cepharanthine is a secondary metabolite isolated from Stephania. It has been reported that it has anti-conronaviruses activities including severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Here, we assemble three Stephania genomes (S. japonica, S. yunnanensis, and S. cepharantha), propose the cepharanthine biosynthetic pathway, and assess the antiviral potential of compounds involved in the pathway. Among the three genomes, S. japonica has a near telomere-to-telomere assembly with one remaining gap, and S. cepharantha and S. yunnanensis have chromosome-level assemblies. Following by biosynthetic gene mining and metabolomics analysis, we identify seven cepharanthine analogs that have broad-spectrum anti-coronavirus activities, including SARS-CoV-2, Guangxi pangolin-CoV (GX_P2V), swine acute diarrhoea syndrome coronavirus (SADS-CoV), and porcine epidemic diarrhea virus (PEDV). We also show that two other genera, Nelumbo and Thalictrum, can produce cepharanthine analogs, and thus have the potential for antiviral compound discovery. Results generated from this study could accelerate broad-spectrum anti-coronavirus drug discovery.
Topics: Animals; Swine; Stephania; China; SARS-CoV-2; Antiviral Agents; Alphacoronavirus; Benzylisoquinolines; Benzodioxoles
PubMed: 38378731
DOI: 10.1038/s41467-024-45690-5 -
Microbiology Spectrum Dec 2023Understanding the role of the endoribonuclease non-structural protein 15 (nsp15) (EndoU) in coronavirus (CoV) infection and pathogenesis is essential for vaccine target...
Understanding the role of the endoribonuclease non-structural protein 15 (nsp15) (EndoU) in coronavirus (CoV) infection and pathogenesis is essential for vaccine target discovery. Whether the EndoU activity of CoV nsp15, as a virulence-related protein, has a diverse effect on viral virulence needs to be further explored. Here, we found that the transmissible gastroenteritis virus (TGEV) and feline infectious peritonitis virus (FIPV) nsp15 proteins antagonize SeV-induced interferon-β (IFN-β) production in human embryonic kidney 293 cells. Interestingly, compared with wild-type infection, infection with EnUmt-TGEV or EnUmt-FIPV did not change the IFN-β response or reduce viral propagation in immunocompetent cells. The results of animal experiments showed that EnUmt viruses did not reduce the clinical presentation and mortality caused by TGEV and FIPV. Our findings enrich the understanding of nsp15-mediated regulation of alpha-CoV propagation and virulence and reveal that the conserved functions of nonstructural proteins have diverse effects on the pathogenicity of CoVs.
Topics: Animals; Humans; Coronavirus; Virulence; Endoribonucleases; Uridylate-Specific Endoribonucleases; Coronavirus Infections
PubMed: 37938022
DOI: 10.1128/spectrum.02209-23 -
Journal of Virology Dec 2023Coronaviruses are important pathogens of humans and animals, and vaccine developments against them are imperative. Due to the ability to induce broad and prolonged...
Coronaviruses are important pathogens of humans and animals, and vaccine developments against them are imperative. Due to the ability to induce broad and prolonged protective immunity and the convenient administration routes, live attenuated vaccines (LAVs) are promising arms for controlling the deadly coronavirus infections. However, potential recombination events between vaccine and field strains raise a safety concern for LAVs. The porcine epidemic diarrhea virus (PEDV) remodeled TRS (RMT) mutant generated in this study replicated efficiently in both cell culture and in pigs and retained protective immunogenicity against PEDV challenge in pigs. Furthermore, the RMT PEDV was resistant to recombination and genetically stable. Therefore, RMT PEDV can be further optimized as a backbone for the development of safe LAVs.
Topics: Animals; Coronavirus Infections; Porcine epidemic diarrhea virus; Recombination, Genetic; Swine; Swine Diseases; Vaccines, Attenuated; Viral Vaccines; Virus Replication; Cells, Cultured; Mutation
PubMed: 37971221
DOI: 10.1128/jvi.01193-23 -
PLoS Pathogens Mar 2024Alphacoronaviruses are the primary coronaviruses responsible for causing severe economic losses in the pig industry with the potential to cause human outbreaks....
Alphacoronaviruses are the primary coronaviruses responsible for causing severe economic losses in the pig industry with the potential to cause human outbreaks. Currently, extensive studies have reported the essential role of endosomal sorting and transport complexes (ESCRT) in the life cycle of enveloped viruses. However, very little information is available about which ESCRT components are crucial for alphacoronaviruses infection. By using RNA interference in combination with Co-immunoprecipitation, as well as fluorescence and electron microscopy approaches, we have dissected the role of ALIX and TSG101 for two porcine alphacoronavirus cellular entry and replication. Results show that infection by two porcine alphacoronaviruses, including porcine epidemic diarrhea virus (PEDV) and porcine enteric alphacoronavirus (PEAV), is dramatically decreased in ALIX- or TSG101-depleted cells. Furthermore, PEDV entry significantly increases the interaction of ALIX with caveolin-1 (CAV1) and RAB7, which are crucial for viral endocytosis and lysosomal transport, however, does not require TSG101. Interestingly, PEAV not only relies on ALIX to regulate viral endocytosis and lysosomal transport, but also requires TSG101 to regulate macropinocytosis. Besides, ALIX and TSG101 are recruited to the replication sites of PEDV and PEAV where they become localized within the endoplasmic reticulum and virus-induced double-membrane vesicles. PEDV and PEAV replication were significantly inhibited by depletion of ALIX and TSG101 in Vero cells or primary jejunal epithelial cells, indicating that ALIX and TSG101 are crucial for PEDV and PEAV replication. Collectively, these data highlight the dual role of ALIX and TSG101 in the entry and replication of two porcine alphacoronaviruses. Thus, ESCRT proteins could serve as therapeutic targets against two porcine alphacoronaviruses infection.
Topics: Animals; Alphacoronavirus; Cell Line; Chlorocebus aethiops; Endosomal Sorting Complexes Required for Transport; Epithelial Cells; Porcine epidemic diarrhea virus; Swine; Vero Cells; Virus Replication; Calcium-Binding Proteins
PubMed: 38489378
DOI: 10.1371/journal.ppat.1012103 -
Veterinary Medicine and Science Nov 2023To investigate intestinal injury, repair and vasculitis biomarkers that may illuminate the progression and/or pathogenesis of feline infectious peritonitis (FIP) or...
OBJECTIVE
To investigate intestinal injury, repair and vasculitis biomarkers that may illuminate the progression and/or pathogenesis of feline infectious peritonitis (FIP) or feline enteric coronavirus (FECV) infection.
MATERIALS AND METHODS
A total of 40 cats with effusive FIP (30 with abdominal effusion, AE group; 10 with thoracic effusion, TE group) and 10 asymptomatic but FECV positive cats (FECV group), all were confirmed by reverse transcription polymerase chain reaction either in faeces or effusion samples. Physical examinations and effusion tests were performed. Trefoil factor-3 (TFF-3), intestinal alkaline phosphatase (IAP), intestinal fatty acid binding protein (I-FABP), myeloperoxidase-anti-neutrophilic cytoplasmic antibody (MPO-ANCA) and proteinase 3-ANCA (PR3-ANCA) concentrations were measured both in serum and effusion samples.
RESULTS
Rectal temperature and respiratory rate were highest in the TE group (p < 0.000). Effusion white blood cell count was higher in the AE group than TE group (p < 0.042). Serum TFF-3, IAP and I-FABP concentrations were higher in cats with effusive FIP than the cats with FECV (p < 0.05). Compared with the AE group, TE group had lower effusion MPO-ANCA (p < 0.036), higher IAP (p < 0.050) and higher TFF-3 (p < 0.016) concentrations.
CLINICAL SIGNIFICANCE
Markers of intestinal and epithelial surface injury were higher in cats with effusive FIP than those with FECV. Compared to cats with abdominal effusions, markers of apoptosis inhibition and immunostimulation to the injured epithelium were more potent in cats with thoracic effusion, suggesting the possibility of a poorer prognosis or more advanced disease in these patients.
Topics: Cats; Animals; Coronavirus, Feline; Feline Infectious Peritonitis; Antibodies, Antineutrophil Cytoplasmic; Coronavirus Infections; Biomarkers; Cat Diseases
PubMed: 37872840
DOI: 10.1002/vms3.1299 -
Vaccine Oct 2023Porcine epidemic diarrhea virus (PEDV) is a main cause of severe enteric disease in piglets, leading to millions of dollars lost annually in the global pig industry....
Integrative transcriptomic and metabolomic analysis in mice reveals the mechanism by which ginseng stem-leaf saponins enhance mucosal immunity induced by a porcine epidemic diarrhea virus vaccination.
Porcine epidemic diarrhea virus (PEDV) is a main cause of severe enteric disease in piglets, leading to millions of dollars lost annually in the global pig industry. Parenteral vaccination is limited in generating sufficient mucosal immunity, which is crucial for early defense against PEDV. Here, we orally administered ginseng stem-leaf saponins (GSLS) to mice before parenteral vaccination and found that GSLS significantly enhanced the phagocytosis of dendritic cells, promoted the activities of CD4 T cells and increased PEDV-specific IgA antibodies in the intestinal mucosa. Transcriptomic results showed that the altered genes following GSLS treatment were mostly related to the immune response and metabolism. In addition, integrated analysis of the transcriptome and metabolome revealed that the mechanism by which GSLS enhances mucosal immunity may be associated with progesterone-related pathways. Further studies are needed to explore the detailed molecular mechanisms.
Topics: Animals; Swine; Mice; Immunity, Mucosal; Transcriptome; Porcine epidemic diarrhea virus; Panax; Saponins; Vaccination; Plant Leaves; Coronavirus Infections; Swine Diseases
PubMed: 37704497
DOI: 10.1016/j.vaccine.2023.09.017