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.

Porcine epidemic diarrhea virus (PEDV) belongs to the genus Alphacoronavirus of the family Coronaviridae that causes severe diarrhea and high mortality in neonatal suckling piglets. Currently, there is no effective medication against this pathogen. Cepharanthine (CEP), tetrandrine (TET), and fangchinoline (FAN) are natural bis-benzylisoquinoline alkaloids with anti-inflammatory, antitumor, and antiviral properties. Here, we first found that CEP, TET, and FAN had anti-PEDV activity with IC₅₀ values of 2.53, 3.50, and 6.69 μM, respectively. The compounds could block all the processes of viral cycles, but early application of the compounds before or during virus infection was advantageous over application at a late stage of virus replication. FAN performed inhibitory function more efficiently through interfering with the virus entry and attachment processes or through attenuating the virus directly. CEP had a more notable effect on virus entry. With the highest SI index of 11.8 among the three compounds, CEP was chosen to carry out animal experiments. CEP in a safe dosage of 11.1 mg/kg of body weight could reduce viral load and pathological change of piglet intestinal tracts caused by PEDV field strain challenge, indicating that CEP efficiently inhibited PEDV infection in vivo. All of these results demonstrated that the compounds of bis-benzylisoquinoline alkaloids could inhibit PEDV proliferation efficiently and had the potential of being developed for PED prevention and treatment.

Porcine epidemic diarrhea virus (PEDV) is a lethal pathogen for newborn piglets, and an efficient vaccine is needed urgently. However, propagating wild-type PEDV in cultured cells for vaccine development is still challenging due to the lack of knowledge about the mechanism of the cell adaptation of PEDV.

The spike protein (S) plays a crucial role in porcine epidemic diarrhea virus (PEDV) infection and induces neutralizing antibodies. Mutations of the S protein are supposed to provide the main antigenic shift leading to the antigenic escape of PEDVs. It is therefore a significant question how much accumulation of antigenic shift could lead to the antigenic escape of the variant PEDV. To provide an answer in the study, B cell epitopes (BCEs) on the S protein of the PEDV vaccine strain CV777 (S^CV777) and variant strain SD2014 (S^SD2014) were mapped using biosynthetic peptides and rabbit anti-PEDV S serum. Seventy-nine and 68 linear BCEs were identified from S^CV777 and S^SD2014, respectively. While 66.2% of the BCEs of S^SD2014 could be recognized by anti-S^CV777 serum and 67.1% of S^CV777 BCEs could be recognized by anti-S^SD2014 serum, more than 40% of the BCEs identified using anti-S^CV777 serum on S^CV777 could not be recognized by anti-S^SD2014 serum and vice versa. The completely shared BCEs took low percentages of 29.4% and 25.3% for S^SD2014 and S^CV777, respectively. These results indicate a low conservation of antigenicity of the S protein compared to a relatively high amino acid sequence similarity of 92.2% between the two strains. The study provided a BCE shift reference of PEDV antigenic escape and surveillance control.

Porcine epidemic diarrhea virus (PEDV) is the etiology of porcine epidemic diarrhea (PED), a highly contagious digestive disease in pigs and especially in neonatal piglets, in which a mortality rate of up to 100% will be induced. Immunizing pregnant sows remains the most promising and effective strategy for protecting their neonatal offspring from PEDV. Although half a century has passed since its first report in Europe and several prophylactic vaccines (inactivated or live attenuated) have been developed, PED still poses a significant economic concern to the swine industry worldwide. Hence, there is an urgent need for novel vaccines in clinical practice, especially live attenuated vaccines (LAVs) that can induce a strong protective lactogenic immune response in pregnant sows. Reverse genetic techniques provide a robust tool for virological research from the function of viral proteins to the generation of rationally designed vaccines. In this review, after systematically summarizing the research progress on virulence-related viral proteins, we reviewed reverse genetics techniques for PEDV and their application in the development of PED LAVs. Then, we probed into the potential methods for generating safe, effective, and genetically stable PED LAV candidates, aiming to provide new ideas for the rational design of PED LAVs.

Coronaviruses (CoVs) are a large class of positively stranded RNA viruses that pose a significant threat to public health, livestock farming, and wild animals. These viruses have the ability to cross species barriers and cause devastating epidemics. Animals are considered to be intermediate hosts for many coronaviruses, and many animal coronaviruses also have the potential for cross-species transmission to humans. Therefore, controlling the epidemic transmission of animal coronaviruses is of great importance to human health. Vaccination programs have proven to be effective in controlling coronaviruses infections, offering a cost-effective approach to reducing morbidity and mortality, so the re-emergence of lethal coronaviruses emphasizes the urgent need for the development of effective vaccines. In this regard, we explore the progress in animal coronavirus vaccine development, covering the latest taxonomy of the main animal coronaviruses, spillover events, diverse vaccine development platforms, potential main targets for animal coronavirus vaccine development, and primary challenges facing animal coronavirus vaccines. We emphasize the urgent need to create a “dual-effect” vaccine capable of eliciting both cellular and humoral immune responses. The goal is to highlight the contributions of veterinary scientists in this field and emphasize the importance of interdisciplinary collaboration between the veterinary and medical communities. By promoting communication and cooperation, we can enhance the development of novel and super vaccines to combat human and animal coronavirus infections in the future.