scholarly journals Porcine Epidemic Diarrhea Virus (PEDV) ORF3 Enhances Viral Proliferation by Inhibiting Apoptosis of Infected Cells

Viruses ◽  
2020 ◽  
Vol 12 (2) ◽  
pp. 214 ◽  
Author(s):  
Fusheng Si ◽  
Xiaoxia Hu ◽  
Chenyang Wang ◽  
Bingqing Chen ◽  
Ruiyang Wang ◽  
...  
Keyword(s):  
Porcine Epidemic Diarrhea Virus ◽  
Underlying Mechanism ◽  
Vero Cells ◽  
Orf3 Gene ◽  
Accessory Gene ◽  
Orf3 Protein ◽  
Diarrhea Virus ◽  
Viral Virulence ◽  
Porcine Epidemic Diarrhea

The genomes of coronaviruses carry accessory genes known to be associated with viral virulence. The single accessory gene of porcine epidemic diarrhea virus (PEDV), ORF3, is dispensable for virus replication in vitro, while viral mutants carrying ORF3 truncations exhibit an attenuated phenotype of which the underlying mechanism is unknown. Here, we studied the effect of ORF3 deletion on the proliferation of PEDV in Vero cells. To this end, four recombinant porcine epidemic diarrhea viruses (PEDVs) were rescued using targeted RNA recombination, three carrying the full-length ORF3 gene from different PEDV strains, and one from which the ORF3 gene had been deleted entirely. Our results showed that PEDVs with intact or naturally truncated ORF3 replicated to significantly higher titers than PEDV without an ORF3. Further characterization revealed that the extent of apoptosis induced by PEDV infection was significantly lower with the viruses carrying an intact or C-terminally truncated ORF3 than with the virus lacking ORF3, indicating that the ORF3 protein as well as its truncated form interfered with the apoptosis process. Collectively, we conclude that PEDV ORF3 protein promotes virus proliferation by inhibiting cell apoptosis caused by virus infection. Our findings provide important insight into the role of ORF3 protein in the pathogenicity of PEDV.

scholarly journals Successive Passage In Vitro Led to Lower Virulence and Higher Titer of A Variant Porcine Epidemic Diarrhea Virus

Viruses ◽  
2020 ◽  
Vol 12 (4) ◽  
pp. 391 ◽  
Author(s):  
Pengwei Zhao ◽  
Song Wang ◽  
Zhi Chen ◽  
Jiang Yu ◽  
Rongzhi Tang ◽  
...  
Keyword(s):  
Amino Acids ◽  
Porcine Epidemic Diarrhea Virus ◽  
Vero Cells ◽  
Economic Losses ◽  
Diarrhea Virus ◽  
S Gene ◽  
Successive Passage ◽  
Porcine Epidemic Diarrhea

A highly virulent porcine epidemic diarrhea virus (PEDV) appeared in China and spread rapidly to neighbor countries, which have led to great economic losses to the pig industry. In the present study, we isolated a PEDV using Vero cells and serially propagated 100 passages. PEDV SDSX16 was characterized in vitro and in vivo. The viral titers increased to 107.6 TCID50/mL (100th) by serial passages. The spike (S) gene and the whole gene of the SDSX16 virus was fully sequenced to assess the genetic stability and relatedness to previously identified PEDV. Along with successive passage in vitro, there were 18 nucleotides (nt) deletion occurred in the spike (S) gene resulting in a deletion of six amino acids when the SDSX16 strain was passaged to the 64th generation, and this deletion was stable until the P100. However, the ORF1a/b, M, N, E, and ORF3 genes had only a few point mutations in amino acids and no deletions. According to growth kinetics experiments, the SDSX16 deletion strain significantly enhanced its replication in Vero cells since it was passaged to the 64th generation. The animal studies showed that PEDV SDSX16-P10 caused more severe diarrhea and vomiting, fecal shedding, and acute atrophic enteritis than SDSX16-P75, indicating that SDSX16-P10 is enteropathogenic in the natural host, and the pathogenicity of SDSX16 decreased with successive passage in vitro. However, SDSX16-P10 was found to cause lower levels of cytokine expression than SDSX16-P75 using real-time PCR and flow cytometry, such as IL1β, IL6, IFN-β, TNF-α, indicating that SDSX16-P10 might inhibit the expression of cytokines. Our data indicated that successive passage in vitro resulted in virulent attenuation in vivo of the PEDV variant strain SDSX16.

scholarly journals In Vitro Antiviral Activities of Salinomycin on Porcine Epidemic Diarrhea Virus

Viruses ◽  
2021 ◽  
Vol 13 (4) ◽  
pp. 580
Author(s):  
Chen Yuan ◽  
Xintong Huang ◽  
Ruiyu Zhai ◽  
Yichao Ma ◽  
Anyuan Xu ◽  
...  
Keyword(s):  
Porcine Epidemic Diarrhea Virus ◽  
Mapk Pathway ◽  
Vero Cells ◽  
Dependent Manner ◽  
Diarrhea Virus ◽  
Pathway Activation ◽  
P38mapk Signaling ◽  
Porcine Epidemic Diarrhea ◽  
Post Entry

Porcine epidemic diarrhea virus (PEDV), an enteropathogenic coronavirus, has catastrophic impacts on the global pig industry. Owing to the lack of effective vaccines and specific therapeutic options for PEDV, it is pertinent to develop new and available antivirals. This study identified, for the first time, a salinomycin that actively inhibited PEDV replication in Vero cells in a dose-dependent manner. Furthermore, salinomycin significantly inhibited PEDV infection by suppressing the entry and post-entry of PEDV in Vero cells. It did not directly interact with or inactivate PEDV particles, but it significantly ameliorated the activation of Erk1/2, JNK and p38MAPK signaling pathways that are associated with PEDV infection. This implied that salinomycin inhibits PEDV replication by altering MAPK pathway activation. Notably, the PEDV induced increase in reactive oxidative species (ROS) was not decreased, indicating that salinomycin suppresses PEDV replication through a pathway that is an independent pathway of viral-induced ROS. Therefore, salinomycin is a potential drug that can be used for treating PEDV infection.

scholarly journals Porcine Epidemic Diarrhea Virus (PEDV) ORF3 Interactome Reveals Inhibition of Virus Replication by Cellular VPS36 Protein

Viruses ◽  
2019 ◽  
Vol 11 (4) ◽  
pp. 382 ◽  
Author(s):  
Challika Kaewborisuth ◽  
Yodying Yingchutrakul ◽  
Sittiruk Roytrakul ◽  
Anan Jongkaewwattana
Keyword(s):  
Virus Replication ◽  
Porcine Epidemic Diarrhea Virus ◽  
Interacting Proteins ◽  
Gene Ontology Analysis ◽  
Orf3 Gene ◽  
Orf3 Protein ◽  
Diarrhea Virus ◽  
Host Interaction ◽  
Porcine Epidemic Diarrhea ◽  
Vacuolar Protein

The accessory protein ORF3 of porcine epidemic diarrhea virus (PEDV) has been proposed to play a key role in virus replication. However, our understanding of its function regarding virus and host interaction is still limited. In this study, we employed immunoprecipitation and mass spectrometry to screen for cellular interacting partners of ORF3. Gene ontology analysis of the host interactome highlighted the involvement of ORF3 in endosomal and immune signaling pathways. Among the identified ORF3-interacting proteins, the vacuolar protein-sorting-associated protein 36 (VPS36) was assessed for its role in PEDV replication. VPS36 was found to interact with ORF3 regardless of its GLUE domain. As a result of VPS36–ORF3 interaction, PEDV replication was substantially suppressed in cells overexpressing VPS36. Interestingly, the ORF3 protein expression was diminished in VPS36-overexpressing cells, an effect that could not be restored by treatment of lysosomal inhibitors. In addition, disruption of endogenously-expressed VPS36 by siRNA could partially augment PEDV replication. Taken together, our study provides mechanistic insights into the contribution of ORF3 in PEDV replication.

scholarly journals Epigallocatechin-3-Gallate, the Main Polyphenol in Green Tea, Inhibits Porcine Epidemic Diarrhea Virus In Vitro

2021 ◽  
Vol 12 ◽  
Author(s):  
Changchao Huan ◽  
Weiyin Xu ◽  
Bo Ni ◽  
Tingting Guo ◽  
Haochun Pan ◽  
...  
Keyword(s):  
Green Tea ◽  
Western Blotting ◽  
Porcine Epidemic Diarrhea Virus ◽  
Vero Cells ◽  
Dependent Manner ◽  
Egcg Treatment ◽  
Diarrhea Virus ◽  
Porcine Epidemic Diarrhea ◽  
Dose Dependent

There are currently no licensed drugs against porcine epidemic diarrhea virus (PEDV), but vaccines are available. We identified a natural molecule, epigallocatechin-3-gallate (EGCG), the main polyphenol in green tea, which is effective against infection with PEDV. We used a variety of methods to test its effects on PEDV in Vero cells. Our experiments show that EGCG can effectively inhibit PEDV infections (with HLJBY and CV777 strains) at different time points in the infection using western blot analysis. We found that EGCG inhibited PEDV infection in a dose-dependent manner 24 h after the infection commenced using western blotting, plaque formation assays, immunofluorescence assays (IFAs), and quantitative reverse-transcriptase PCR (qRT-PCR). We discovered that EGCG treatment of Vero cells decreased PEDV attachment and entry into them by the same method analysis. Western blotting also showed that PEDV replication was inhibited by EGCG treatment. Whereas EGCG treatment was found to inhibit PEDV assembly, it had no effect on PEDV release. In summary, EGCG acts against PEDV infection by inhibiting PEDV attachment, entry, replication, and assembly.

scholarly journals Caerin1.1 Suppresses the Growth of Porcine Epidemic Diarrhea Virus In Vitro via Direct Binding to the Virus

Viruses ◽  
2018 ◽  
Vol 10 (9) ◽  
pp. 507 ◽  
Author(s):  
Nan Guo ◽  
Bingzhou Zhang ◽  
Han Hu ◽  
Shiyi Ye ◽  
Fangzhou Chen ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Porcine Epidemic Diarrhea Virus ◽  
Antiviral Drug ◽  
Vero Cells ◽  
Economic Losses ◽  
Diarrhea Virus ◽  
Direct Binding ◽  
Porcine Epidemic Diarrhea ◽  
And Control

Porcine epidemic diarrhea (PED) has re-emerged in recent years and has already caused huge economic losses to the porcine industry all over the world. Therefore, it is urgent for us to find out efficient ways to prevent and control this disease. In this study, the antiviral activity of a cationic amphibian antimicrobial peptide Caerin1.1 against porcine epidemic diarrhea virus (PEDV) was evaluated by an in vitro system using Vero cells. We found that even at a very low concentration, Caerin1.1 has the ability to destroy the integrity of the virus particles to block the release of the viruses, resulting in a considerable decrease in PEDV infections. In addition, Caerin1.1 showed powerful antiviral activity without interfering with the binding progress between PEDV and the receptor of the cells, therefore, it could be used as a potential antiviral drug or as a microbicide compound for prevention and control of PEDV.

scholarly journals Differential expression and correlation analysis of miRNA–mRNA profiles in swine testicular cells infected with porcine epidemic diarrhea virus

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Xiaoqian Zhang ◽  
Chang Li ◽  
Bingzhou Zhang ◽  
Zhonghua Li ◽  
Wei Zeng ◽  
...  
Keyword(s):  
Differential Expression ◽  
Porcine Epidemic Diarrhea Virus ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Bacterial Invasion ◽  
Sequencing Data ◽  
Diarrhea Virus ◽  
Comparison Groups ◽  
Porcine Epidemic Diarrhea

AbstractThe variant virulent porcine epidemic diarrhea virus (PEDV) strain (YN15) can cause severe porcine epidemic diarrhea (PED); however, the attenuated vaccine-like PEDV strain (YN144) can induce immunity in piglets. To investigate the differences in pathogenesis and epigenetic mechanisms between the two strains, differential expression and correlation analyses of the microRNA (miRNA) and mRNA in swine testicular (ST) cells infected with YN15, YN144, and mock were performed on three comparison groups (YN15 vs Control, YN144 vs Control, and YN15 vs YN144). The mRNA and miRNA expression profiles were obtained using next-generation sequencing (NGS), and the differentially expressed (DE) (p-value < 0.05) mRNA and miRNA were obtained using DESeq R package. mRNAs targeted by DE miRNAs were predicted using the miRanda algortithm. 8039, 8631 and 3310 DE mRNAs, and 36, 36, and 22 DE miRNAs were identified in the three comparison groups, respectively. 14,140, 15,367 and 3771 DE miRNA–mRNA (targeted by DE miRNAs) interaction pairs with negatively correlated expression patterns were identified, and interaction networks were constructed using Cytoscape. Six DE miRNAs and six DE mRNAs were randomly selected to verify the sequencing data by real-time relative quantitative reverse transcription polymerase chain reaction (qRT-PCR). Based on bioinformatics analysis, we discovered the differences were mostly involved in host immune responses and viral pathogenicity, including NF-κB signaling pathway and bacterial invasion of epithelial cells, etc. This is the first comprehensive comparison of DE miRNA–mRNA pairs in YN15 and YN144 infection in vitro, which could provide novel strategies for the prevention and control of PED.

scholarly journals Short communication: antiviral activity of porcine IFN-λ3 against porcine epidemic diarrhea virus in vitro

Virus Genes ◽  
2016 ◽  
Vol 52 (6) ◽  
pp. 877-882 ◽  
Author(s):  
Haiyan Shen ◽  
Chunhong Zhang ◽  
Pengju Guo ◽  
Zhicheng Liu ◽  
Minhua Sun ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Porcine Epidemic Diarrhea Virus ◽  
Short Communication ◽  
Diarrhea Virus ◽  
Porcine Epidemic Diarrhea

scholarly journals Emergence and evolution of highly pathogenic porcine epidemic diarrhea virus by natural recombination of a low pathogenic vaccine isolate and a highly pathogenic strain in the spike gene

2020 ◽  
Vol 6 (2) ◽  
Author(s):  
Huinan Wang ◽  
Libo Zhang ◽  
Yuanbin Shang ◽  
Rongrong Tan ◽  
Mingxiang Ji ◽  
...  
Keyword(s):  
Porcine Epidemic Diarrhea Virus ◽  
Intestinal Epithelial ◽  
Orf3 Gene ◽  
Highly Pathogenic ◽  
Spike Gene ◽  
Diarrhea Virus ◽  
Attenuated Vaccine ◽  
New Variant ◽  
Porcine Epidemic Diarrhea ◽  
Natural Recombination

Abstract Outbreaks of a new variant of porcine epidemic diarrhea virus (PEDV) at the end of 2010 have raised interest in the mutation and recombination of PEDV. A PEDV strain (CN/Liaoning25/2018) isolated from a clinical outbreak of piglet diarrhea contained a 49-bp deletion in the ORF3 gene. This deletion is considered a genetic characteristic of low pathogenic attenuated vaccine strains. However, CN/Liaoning25/2018 was highly pathogenic. Complete genome sequencing, identity analysis, phylogenetic tree construction, and recombination analysis showed that this virus was a recombinant strain containing the Spike (S) gene from the highly pathogenic CN/GDZQ/2014 strain and the remaining genomic regions from the low pathogenic vaccine isolate SQ2014. Histopathology and immunohistochemistry results confirmed that this strain was highly pathogenic and indicated that intestinal epithelial cell vacuolation was positively correlated with the intensity and density of PEDV antigens. A new natural recombination model for PEDV was identified. Our results suggest that new highly pathogenic recombinant strains in the field may be generated by recombination between low pathogenic attenuated live PEDV vaccines and pathogenic circulating PEDV strains. Our findings also highlight that the 49-bp deletion of the ORF3 gene in low pathogenic attenuated vaccine strains will no longer be a reliable standard to differentiate the classical vaccine attenuated from the field strains.

scholarly journals Characterization and pathogenicity of Vero cell-attenuated porcine epidemic diarrhea virus CT strain

2019 ◽  
Vol 16 (1) ◽  
Author(s):  
Yu Wu ◽  
Wei Li ◽  
Qingfeng Zhou ◽  
Qunhui Li ◽  
Zhichao Xu ◽  
...  
Keyword(s):  
Porcine Epidemic Diarrhea Virus ◽  
Clinical Symptoms ◽  
Vero Cells ◽  
Economic Losses ◽  
Protective Effects ◽  
Diarrhea Virus ◽  
Cytopathic Effects ◽  
Porcine Epidemic Diarrhea ◽  
Piglet Survival

Abstract Background Porcine epidemic diarrhea virus (PEDV) has caused enormous economic losses to the global pig industry. Currently available PEDV vaccine strains have limited protective effects against PEDV variant strains. Methods In this study, the highly virulent epidemic virus strain CT was serially passaged in Vero cells for up to 120 generations (P120). Characterization of the different passages revealed that compared with P10 and P64, P120 had a higher viral titer and more obvious cytopathic effects, thereby demonstrating better cell adaptability. Results Pathogenicity experiments using P120 in piglets revealed significant reductions in clinical symptoms, histopathological lesions, and intestinal PEDV antigen distribution; the piglet survival rate in the P120 group was 100%. Furthermore, whole-genome sequencing identified 13 amino acid changes in P120, which might be responsible for the attenuated virulence of P120. Conclusions Thus, an attenuated strain was obtained via cell passaging and that this strain could be used in preparing attenuated vaccines.

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