Porcine epidemic diarrhea virus (PEDV) causes a porcine disease associated with swine epidemic diarrhea. The type I interferon (IFN-I or IFN α/β) is a key mediator of innate antiviral response during virus infection. Different antagonistic strategies have been identified and determined as to how PEDV infection inhibits the host’s IFN responses to escape the host innate immune pathway, but the pathogenic mechanisms of PEDV infection are not fully elucidated. Our preliminary results revealed that endogenous TANK-binding kinase 1 (TBK1) and interferon regulatory factor 3 (IRF3), the key components in the IFN signaling pathway were downregulated in PEDV infected IPEC-J2 cells by iTRAQ analysis. In this study, we screened nsp15 as the most important viral encoded protein involved in TBK1 and IRF3 reduction. Endoribonuclease (EndoU) activity has been well determined for coronavirus nsp15. Three residues (H226, H241, and K282) of PEDV nsp15 were identified as critical amino acids for PEDV EndoU but not D265, which was not well correlated with published results of other coronaviruses, such as severe acute respiratory syndrome virus (SARS-CoV). Moreover, PEDV nsp15 can directly degrade the RNA levels of TBK1 and IRF3 dependent on its EndoU activity to suppress IFN production and constrain the induction of IFN stimulated genes (ISGs), by which PEDV antagonizes the host innate response to facilitate its replication. Collectively, these results have confirmed that PEDV nsp15 was capable of subverting the IFN response by the RNA degradation of TBK1 and IRF3.

Coronaviruses (CoVs) are a family of the largest RNA viruses that typically cause respiratory, enteric, and hepatic diseases in animals and humans, imposing great threats to the public safety and animal health. Porcine deltacoronavirus (PDCoV), a newly emerging enteropathogenic coronavirus, causes severe diarrhea in suckling piglets all over the world and poses potential risks of cross-species transmission. Here, we use PDCoV as a model of CoVs to illustrate the reciprocal regulation between CoVs infection and host antiviral responses. In this study, downregulation of DNA polymerase delta interacting protein 3 (POLDIP3) was confirmed in PDCoV infected IPEC-J2 cells by isobaric tags for relative and absolute quantification (iTRAQ) and Western blotting analysis. Overexpression of POLDIP3 inhibits PDCoV infection, whereas POLDIP3 knockout (POLDIP3^-/-) by CRISPR-Cas9 editing significantly promotes PDCoV infection, indicating POLDIP3 as a novel antiviral regulator against PDCoV infection. Surprisingly, an antagonistic strategy was revealed that PDCoV encoded nonstructural protein 5 (nsp5) was responsible for POLDIP3 reduction via its 3C-like protease cleavage of POLDIP3 at the glutamine acid 176 (Q176), facilitating PDCoV infection due to the loss of antiviral effects of the cleaved fragments. Consistent with the obtained data in IPEC-J2 cell model in vitro, POLDIP3 reduction by cleavage was also corroborated in PDCoV infected-SPF piglets in vivo. Collectively, we unveiled a new antagonistic strategy evolved by PDCoV to counteract antiviral innate immunity by nsp5-mediated POLDIP3 cleavage, eventually ensuring productive virus replication. Importantly, we further demonstrated that nsp5s from PEDV and TGEV harbor the conserved function to cleave porcine POLDIP3 at the Q176 to despair POLDIP3-mediated antiviral effects. In addition, nsp5 from SARS-CoV-2 also cleaves human POLDIP3. Therefore, we speculate that coronaviruses employ similar POLDIP3 cleavage mechanisms mediated by nsp5 to antagonize the host antiviral responses to sustain efficient virus infection.

Porcine deltacoronavirus (PDCoV) is a novel coronavirus that can cause vomiting and watery diarrhea in pigs and death in piglets. Since PDCoV was first detected in 2009 in Hong Kong, the prevalence of PDCoV has increased in recent years, resulting in serious economic losses to the swine industry. The coronavirus spike (S) protein is an antigen that has been demonstrated to contain epitopes that induce neutralizing antibodies. The presence of serum and milk IgA antibodies against pathogens that replicate primarily on mucosal surfaces is important for mucosal immunity. Here, an indirect anti-PDCoV IgA antibody enzyme-linked immunosorbent assay (PDCoV S1 IgA ELISA) using the purified S1 portion of S protein as the coating antigen was developed to detect PDCoV IgA antibodies in serum and sow’s milk. A receiver operating characteristic (ROC) curve analysis showed high specificity and sensitivity of the PDCoV-S1-IgA-ELISA based on samples confirmed by IFA. Anti-PDCoV IgA antibodies in 152 serum samples and 65 milk samples collected from six farms that had experienced diarrhea outbreaks within previous last two years were detected by this assay, and 62.5% of the serum samples and 100% of the milk samples were positive for PDCoV. The indirect ELISA method established in this study will provide a convenient tool for measurement of serum and milk IgA levels against PDCoV in pig herds, rapid detection of PDCoV infection in pigs, and evaluation of the immunogenicity of vaccines.

Porcine epidemic diarrhea virus (PEDV) infects piglets and causes serious diarrhea as well as vomiting, dehydration, and death. The trimeric S protein plays a crucial role in the induction of neutralizing antibodies, and many neutralizing monoclonal antibodies (mAbs) against PEDV S protein have been developed. However, these mAbs exclusively target the S1 protein. In this study, we obtained a neutralizing mAb, 5F7, against the S2 protein of PEDV, and this mAb could neutralize new variant genotype 2 PEDV strains (LNCT2), as well as a genotype 1 PEDV strain (CV777), in vitro. The core sequence of the epitope was found in amino acid sequence 1261 aa~1337 aa. These findings confirm that the S2 protein possessed neutralizing epitopes and provided knowledge to aid further research on this virus.

Porcine deltacoronavirus (PDCoV) is a recently discovered enteropathogenic coronavirus and has caused significant economic impacts on the pork industry. Although studies have partly uncovered the molecular mechanism of PDCoV–host interaction, it requires further research. In this study, we explored the roles of Stromal Antigen 2 (STAG2) in PDCoV infection. We found that STAG2-deficient cells inhibited infection with vesicular stomatitis virus (VSV) and PDCoV, whereas restoration of STAG2 expression in STAG2-depleted (STAG2^−/−) IPEC-J2 cells line restored PDCoV infection, suggesting that STAG2 is involved in the PDCoV replication. Furthermore, we found that STAG2 deficiency results in robust interferon (IFN) expression. Subsequently, we found that STAG2 deficiency results in the activation of JAK-STAT signaling and the expression of IFN stimulated gene (ISG), which establish an antiviral state. Taken together, the depletion of STAG2 activates the JAK-STAT signaling and induces the expression of ISG, thereby inhibiting PDCoV replication. Our study provides new insights and potential therapeutic targets for unraveling the mechanism of PDCoV replication.