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Virus Research Feb 2024Porcine deltacoronavirus (PDCoV) is an emerging swine enteropathogenic coronavirus (CoV) that mainly causes acute diarrhea/vomiting, dehydration, and mortality in...
Porcine deltacoronavirus (PDCoV) is an emerging swine enteropathogenic coronavirus (CoV) that mainly causes acute diarrhea/vomiting, dehydration, and mortality in piglets, possessing economic losses and public health concerns. However, there are currently no proven effective antiviral agents against PDCoV. Cepharanthine (CEP) is a naturally occurring alkaloid used as a traditional remedy for radiation-induced symptoms, but its underlying mechanism of CEP against PDCoV has remained elusive. The aim of this study was to investigate the anti-PDCoV effects and mechanisms of CEP in LLC-PK1 cells. The results showed that the antiviral activity of CEP was based on direct action on cells, preventing the virus from attaching to host cells and virus replication. Importantly, Surface Plasmon Resonance (SPR) results showed that CEP has a moderate affinity to PDCoV receptor, porcine aminopeptidase N (pAPN) protein. AutoDock predicted that CEP can form hydrogen bonds with amino acid residues (R740, N783, and R790) in the binding regions of PDCoV and pAPN. In addition, RT-PCR results showed that CEP treatment could significantly reduce the transcription of ZBP1, cytokine (IL-1β and IFN-α) and chemokine genes (CCL-2, CCL-4, CCL-5, CXCL-2, CXCL-8, and CXCL-10) induced by PDCoV. Western blot analysis revealed that CEP could inhibit viral replication by inducing autophagy. In conclusion, our results suggest that the anti-PDCoV activity of CEP is not only relies on competing the virus binding with pAPN, but also affects the proliferation of the virus in vitro by downregulating the excessive immune response caused by the virus and inducing autophagy. CEP emerges as a promising candidate for potential anti-PDCoV therapeutic development.
Topics: Animals; Swine; Swine Diseases; Coronavirus; Coronavirus Infections; CD13 Antigens; Deltacoronavirus; Benzylisoquinolines; Benzodioxoles
PubMed: 38145807
DOI: 10.1016/j.virusres.2023.199303 -
Journal of Virology Jan 2024Coronaviruses (CoVs) pose a major threat to human and animal health worldwide, which complete viral replication by hijacking host factors. Identifying host factors...
Coronaviruses (CoVs) pose a major threat to human and animal health worldwide, which complete viral replication by hijacking host factors. Identifying host factors essential for the viral life cycle can deepen our understanding of the mechanisms of virus-host interactions. Based on our previous genome-wide CRISPR screen of α-CoV transmissible gastroenteritis virus (TGEV), we identified the host factor dual-specificity tyrosine phosphorylation-regulated kinase 1A (DYRK1A), but not DYRK1B, as a critical factor in TGEV replication. Rescue assays and kinase inhibitor experiments revealed that the effect of DYRK1A on viral replication is independent of its kinase activity. Nuclear localization signal modification experiments showed that nuclear DYRK1A facilitated virus replication. Furthermore, DYRK1A knockout significantly downregulated the expression of the TGEV receptor aminopeptidase N () and inhibited viral entry. Notably, we also demonstrated that DYRK1A is essential for the early stage of TGEV replication. Transmission electron microscopy results indicated that DYRK1A contributes to the formation of double-membrane vesicles in a kinase-independent manner. Finally, we validated that DYRK1A is also a proviral factor for mouse hepatitis virus, porcine deltacoronavirus, and porcine sapelovirus. In conclusion, our work demonstrated that DYRK1A is an essential host factor for the replication of multiple viruses, providing new insights into the mechanism of virus-host interactions and facilitating the development of new broad-spectrum antiviral drugs.IMPORTANCECoronaviruses, like other positive-sense RNA viruses, can remodel the host membrane to form double-membrane vesicles (DMVs) as their replication organelles. Currently, host factors involved in DMV formation are not well defined. In this study, we used transmissible gastroenteritis virus (TGEV) as a virus model to investigate the regulatory mechanism of dual-specificity tyrosine phosphorylation-regulated kinase 1A (DYRK1A) on coronavirus. Results showed that DYRK1A significantly inhibited TGEV replication in a kinase-independent manner. DYRK1A knockout (KO) can regulate the expression of receptor aminopeptidase N () and endocytic-related genes to inhibit virus entry. More importantly, our results revealed that DYRK1A KO notably inhibited the formation of DMV to regulate the virus replication. Further data proved that DYRK1A is also essential in the replication of mouse hepatitis virus, porcine deltacoronavirus, and porcine sapelovirus. Taken together, our findings demonstrated that DYRK1A is a conserved factor for positive-sense RNA viruses and provided new insights into its transcriptional regulation activity, revealing its potential as a candidate target for therapeutic design.
Topics: Animals; Humans; Mice; CD13 Antigens; Coronavirus; Coronavirus Infections; Deltacoronavirus; Murine hepatitis virus; Swine; Transmissible gastroenteritis virus; Tyrosine; Virus Replication; Dyrk Kinases
PubMed: 38099687
DOI: 10.1128/jvi.01239-23 -
The Journal of Biological Chemistry Jan 2024Porcine deltacoronavirus (PDCoV) is an emerging enteropathogenic coronavirus. It causes mortality in neonatal piglets and is of growing concern because of its broad host...
Porcine deltacoronavirus (PDCoV) is an emerging enteropathogenic coronavirus. It causes mortality in neonatal piglets and is of growing concern because of its broad host range, including humans. To date, the mechanism of PDCoV infection remains poorly understood. Here, based on a genome-wide CRISPR screen of PDCoV-infected cells, we found that HSP90AB1 (heat shock protein 90 alpha family class B1) promotes PDCoV infection. Knockdown or KO of HSP90AB1 in LLC-PK cells resulted in a significantly suppressed PDCoV infection. Infected cells treated with HSP90 inhibitors 17-AAG and VER-82576 also showed a significantly suppressed PDCoV infection, although KW-2478, which does not affect the ATPase activity of HSP90AB1, had no effect on PDCoV infection. We found that HSP90AB1 interacts with the N, NS7, and NSP10 proteins of PDCoV. We further evaluated the interaction between N and HSP90AB1 and found that the C-tail domain of the N protein is the HSP90AB1-interacting domain. Further studies showed that HSP90AB1 protects N protein from degradation via the proteasome pathway. In summary, our results reveal a key role for HSP90AB1 in the mechanism of PDCoV infection and contribute to provide new host targets for PDCoV antiviral research.
Topics: Animals; Humans; Deltacoronavirus; Host Specificity; HSP90 Heat-Shock Proteins; Swine; HEK293 Cells; Virus Replication
PubMed: 38092149
DOI: 10.1016/j.jbc.2023.105536 -
Frontiers in Microbiology 2023Coinfection of porcine epidemic diarrhea virus (PEDV) and porcine deltacoronavirus (PDCoV) is common in pig farms, but there is currently no effective vaccine to prevent...
Developing a multi-epitope vaccine candidate to combat porcine epidemic diarrhea virus and porcine deltacoronavirus co-infection by employing an immunoinformatics approach.
Coinfection of porcine epidemic diarrhea virus (PEDV) and porcine deltacoronavirus (PDCoV) is common in pig farms, but there is currently no effective vaccine to prevent this co-infection. In this study, we used immunoinformatics tools to design a multi-epitope vaccine against PEDV and PDCoV co-infection. The epitopes were screened through a filtering pipeline comprised of antigenic, immunogenic, toxic, and allergenic properties. A new multi-epitope vaccine named , comprising cytotoxic T lymphocyte-, helper T lymphocyte-, and B cell epitopes, was constructed. To enhance immunogenicity, the TLR2 agonist Pam2Cys and the TLR4 agonist RS09 were added to . Molecular docking and dynamics simulation were performed to reveal the stable interactions between and TLR2 as well as TLR4. Additionally, the immune stimulation prediction indicated that could stimulate T and B lymphocytes to induce a robust immune response. Finally, to ensure the expression of the vaccine protein, the sequence of was optimized and further performed cloning. These studies suggest that has the potential to be a vaccine candidate against PEDV and PDCoV co-infection as well as a new strategy for interrupting the spread of both viruses.
PubMed: 38075906
DOI: 10.3389/fmicb.2023.1295678 -
Veterinarni Medicina Mar 2023Porcine deltacoronavirus (PDCoV) and porcine sapelovirus (PSV) are two viruses that can cause diarrhoea in pigs and bring great economic loss to the pig industry....
Development and application of a low-priced duplex quantitative PCR assay based on SYBR Green I for the simultaneous detection of porcine deltacoronavirus and porcine sapelovirus.
Porcine deltacoronavirus (PDCoV) and porcine sapelovirus (PSV) are two viruses that can cause diarrhoea in pigs and bring great economic loss to the pig industry. In this research, a duplex real-time quantitative polymerase chain reaction (qPCR) assay based on SYBR Green I was developed to simultaneously detect PDCoV and PSV. No specific melting peaks were found in other porcine diarrhoea-associated viruses, indicating that the method developed in this study had good specificity. The detection limits of PDCoV and PSV were 1.0 × 10 copies μl and 1.0 × 10 copies μl, respectively. The duplex real-time qPCR assay tested two hundred and three (203) intestinal and faecal samples collected from diarrhoeal and asymptomatic pigs. The positive rates of PDCoV and PSV were 20.2% and 23.2%, respectively. The co-infection rate of PDCoV and PSV was 13.8%. To evaluate the accuracy of the developed method, conventional PCR and singular TaqMan real-time qPCR assays for PDCoV/PSV were also used to detect the samples. The results showed that the duplex real-time qPCR assay was consistent with the singular assays, but its sensitivity was higher than conventional PCR methods. This duplex real-time qPCR assay provides a rapid, sensitive and reliable method in a clinic to simultaneously detect PDCoV and PSV.
PubMed: 37981902
DOI: 10.17221/79/2022-VETMED -
Microbiology Spectrum Dec 2023Porcine deltacoronavirus (PDCoV) is a newly emerged enteric virus threatening pig industries worldwide. Our previous work showed that PDCoV enters porcine kidney (PK-15)...
Porcine deltacoronavirus (PDCoV) is a newly emerged enteric virus threatening pig industries worldwide. Our previous work showed that PDCoV enters porcine kidney (PK-15) cells through a caveolae-dependent pathway, but the entry mechanism for PDCoV into swine testicle (ST) cells remains unclear. Mechanisms of virus entry can be different with different virus isolates and cell types. Here, we determined that PDCoV enters ST cells via clathrin-mediated endocytosis. Additionally, we found that PDCoV entry does not require Rab5, Rab7, or Rab11. These findings provide additional understanding of the entry mechanisms of PDCoV and possible antiviral targets.
Topics: Animals; Swine; Endocytosis; Deltacoronavirus; Virus Internalization; Clathrin; Swine Diseases; Coronavirus Infections
PubMed: 37962380
DOI: 10.1128/spectrum.02553-23 -
Viruses Sep 2023Emerging and re-emerging swine coronaviruses (CoVs), including porcine epidemic diarrhea virus (PEDV), porcine deltacoronavirus (PDCoV), and swine acute diarrhea... (Review)
Review
Emerging and re-emerging swine coronaviruses (CoVs), including porcine epidemic diarrhea virus (PEDV), porcine deltacoronavirus (PDCoV), and swine acute diarrhea syndrome-CoV (SADS-CoV), cause severe diarrhea in neonatal piglets, and CoV infection is associated with significant economic losses for the swine industry worldwide. Reverse genetics systems realize the manipulation of RNA virus genome and facilitate the development of new vaccines. Thus far, five reverse genetics approaches have been successfully applied to engineer the swine CoV genome: targeted RNA recombination, in vitro ligation, bacterial artificial chromosome-based ligation, vaccinia virus -based recombination, and yeast-based method. This review summarizes the advantages and limitations of these approaches; it also discusses the latest research progress in terms of their use for virus-related pathogenesis elucidation, vaccine candidate development, antiviral drug screening, and virus replication mechanism determination.
Topics: Animals; Swine; Coronavirus; Reverse Genetics; Coronavirus Infections; Porcine epidemic diarrhea virus; RNA; Diarrhea; Swine Diseases
PubMed: 37896780
DOI: 10.3390/v15102003 -
Journal of Virology Nov 2023As a member of the δ-coronavirus family, porcine deltacoronavirus (PDCoV) is a vital reason for diarrhea in piglets, which can contribute to high morbidity and...
As a member of the δ-coronavirus family, porcine deltacoronavirus (PDCoV) is a vital reason for diarrhea in piglets, which can contribute to high morbidity and mortality rates. Initially identified in Hong Kong in 2012, the virus has rapidly spread worldwide. During PDCoV infection, the virus employs evasion mechanisms to evade host surveillance, while the host mounts corresponding responses to impede viral replication. Our research has revealed that PDCoV infection down-regulates the expression of PGAM5 to promote virus replication. In contrast, PGAM5 degrades PDCoV N through autophagy by interacting with the cargo receptor P62 and the E3 ubiquitination ligase STUB1. Additionally, PGAM5 interacts with MyD88 and TRAF3 to activate the IFN signal pathway, resulting in the inhibition of viral replication.
Topics: Animals; Coronavirus Infections; Interferon Type I; Signal Transduction; Swine; Swine Diseases; Ubiquitin-Protein Ligases; Virus Replication; Coronavirus Nucleocapsid Proteins; Deltacoronavirus; Phosphoprotein Phosphatases; Mitochondrial Proteins; Down-Regulation; Immune Evasion; RNA-Binding Proteins; Proteolysis
PubMed: 37882521
DOI: 10.1128/jvi.01470-23 -
Journal of Virology Nov 2023As an emerging porcine enteropathogenic coronavirus that has the potential to infect humans, porcine deltacoronavirus (PDCoV) is receiving increasing attention. However,...
As an emerging porcine enteropathogenic coronavirus that has the potential to infect humans, porcine deltacoronavirus (PDCoV) is receiving increasing attention. However, no effective commercially available vaccines against this virus are available. In this work, we designed a spike (S) protein and receptor-binding domain (RBD) trimer as a candidate PDCoV subunit vaccine. We demonstrated that S protein induced more robust humoral and cellular immune responses than the RBD trimer in mice. Furthermore, the protective efficacy of the S protein was compared with that of inactivated PDCoV vaccines in piglets and sows. Of note, the immunized piglets and suckling pig showed a high level of NAbs and were associated with reduced virus shedding and mild diarrhea, and the high level of NAbs was maintained for at least 4 months. Importantly, we demonstrated that S protein-based subunit vaccines conferred significant protection against PDCoV infection.
Topics: Animals; Female; Humans; Mice; Coronavirus; Coronavirus Infections; Deltacoronavirus; Swine; Swine Diseases; Vaccines, Subunit
PubMed: 37846983
DOI: 10.1128/jvi.00958-23 -
Journal of Virology Oct 2023Retrograde transport has been reported to be closely associated with normal cellular biological processes and viral replication. As an emerging enteropathogenic...
Retrograde transport has been reported to be closely associated with normal cellular biological processes and viral replication. As an emerging enteropathogenic coronavirus with zoonotic potential, porcine deltacoronavirus (PDCoV) has attracted considerable attention. However, whether retrograde transport is associated with PDCoV infection remains unclear. Our present study demonstrates that retromer protein VPS35 acts as a critical host factor that is required for PDCoV infection. Mechanically, VPS35 interacts with PDCoV NS6, mediating the retrograde transport of NS6 from endosomes to the Golgi and preventing it from lysosomal degradation. Recombinant PDCoVs with an NS6 deletion display resistance to VPS35 deficiency. Our work reveals a novel evasion mechanism of PDCoV that involves the manipulation of the retrograde transport pathway by VPS35, providing new insight into the mechanism of PDCoV infection.
Topics: Animals; Coronavirus; Coronavirus Infections; Deltacoronavirus; Swine; Swine Diseases; Virus Replication; Viral Regulatory and Accessory Proteins; Vesicular Transport Proteins
PubMed: 37815351
DOI: 10.1128/jvi.00957-23