Porcine epidemic diarrhea virus (PEDV) causes substantial economic losses in the swine industry globally. Host factors regulating the intracellular replication of PEDV, particularly early RNA synthesis and structural protein production, are not well understood, limiting antiviral strategies. We report that glycoprotein non‑metastatic melanoma protein B (GPNMB) is a key host factor promoting PEDV infection. Initially identified as a PEDV S1-binding partner, GPNMB was confirmed to enhance infection via loss-and gain-of-function experiments in Vero and IPEC-J2 cells. Genetic knockout of GPNMB inhibited PEDV replication without affecting viral attachment or internalization. We show that GPNMB is necessary for the accumulation of double-membrane vesicles (DMVs) and promotes early viral RNA synthesis. Notably, GPNMB directly interacts with the PEDV spike (S) and nucleocapsid (N) proteins, increases their abundance, and facilitates their transport from the endoplasmic reticulum (ER) to the Golgi apparatus, implicating it in structural protein maturation. Our work reveals a pivotal role for GPNMB in PEDV replication and nominates it as a target for host-directed antiviral intervention.

Background Porcine epidemic diarrhea virus (PEDV) primarily infects intestinal epithelial cells, causing severe diarrhea and even death in piglets, which results in substantial economic losses for the global swine industry. Recent studies have suggested that PEDV may exhibit new tissue tropism, including the spleen. Results This study aimed to investigate the infection of the Pandemic PEDV in the spleen based on animal experiments and perform a transcriptomic analysis of response of the piglets spleen infected PEDV using RNA sequencing (RNAseq). The result demonstrated that PEDV primarily colonized the jejunum, but spleen had been also detected high viral RNA titers. Furthermore, the PEDV-N antigen was detectable in the spleens of infected pigs, no significant pathological changes were observed in the regions where the antigen was present. Gene expression analysis identified a total of 21,695 genes in spleen tissue, with 392 differentially expressed genes (DEGs), including 162 upregulated and 230 downregulated. Gene Ontology (GO) analysis revealed significant enrichment in terms related to immune system processes; and, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis indicated that the DEGs were enriched in Complement and coagulation cascades, as well as Viral protein interactions with. To validate the RNAseq results, 20 randomly selected genes and nine representative DEGs associated with PEDV-infection were confirmed using real-time quantitative PCR (RT-qPCR), and the results were consistent with the trends observed in RNAseq. Conclusions In conclusion, the piglets spleen infected PEDV activated the innate immune response, and the spleen expression of genes associated with PEDV-infected had significant changes. These findings provide new insights into the extra-intestinal pathogenic mechanisms of PEDV infection.

Porcine reproductive and respiratory syndrome (PRRS) is a highly contagious swine disease and poses a severe economic burden to the global swine industry. Owing to the inadequacy of current prevention and control measures, developing effective antiviral strategies for preventing PRRS epidemics is imperative. Honokiol (HNK) is a bioactive component of the dietary supplement magnolia extract. Here, we demonstrated that HNK exhibited potent antiviral effects against different PRRSV strains in vitro. Mechanistically, HNK was shown to interfere with PRRSV replication rather than attachment, internalization, or release. HNK was further determined to target PRRSV RNA-dependent RNA polymerase to impair viral genomic RNA synthesis. In addition, HNK was found to decrease PRRSV-triggered inflammatory responses. More importantly, HNK was revealed to reduce viral loads and alleviate lung damage in vivo. This study highlights the potential of HNK as an inhibitor of PRRSV and provides novel insights into its antiviral properties.

BackgroundPorcine reproductive and respiratory syndrome virus (PRRSV) is a significant pathogen in the swine industry that causes reproductive failure in sows and respiratory distress in pigs of all ages, leading to substantial economic losses globally. PRRSV manipulates host cellular processes, particularly those associated with endoplasmic reticulum (ER) function. ER-phagy plays a crucial role in maintaining ER homeostasis and enabling cellular adaptations to stress. Whether and how PRRSV modulates ER-phagy remains incompletely understood.MethodsER-phagy was monitored by western blotting for free mCherry from the mCherry-Sec61B reporter. FAM134B mRNA and protein levels were examined by RT-qPCR and western blotting, respectively. All 12 PRRSV Nsps were screened for FAM134B-suppressing activity by co-transfection, followed by western blotting. Co-immunoprecipitation (Co-IP) was performed to assess interactions between candidate Nsps and FAM134B, as well as their impact on FAM134B- microtubule-associated proteins light chain 3 (LC3) binding. Viral replication was evaluated by RT-qPCR targeting ORF7 and TCID50 assays.ResultsWe investigated the interplay between PRRSV and ER-phagy and discovered that PRRSV suppresses ER-phagy during the late stages of infection. Further analysis revealed that PRRSV employs its Nsps to inhibit the expression of FAM134B. Specifically, PRRSV Nsp2 and Nsp5 interact with FAM134B, promote its degradation and disrupt its binding to microtubule-LC3, thereby impairing ER-phagy.ConclusionsCollectively, our findings uncover a novel viral strategy to subvert host ER-phagy and provide new insights into PRRSV pathogenesis.

Toxoplasmosis is a globally prevalent zoonosis caused by Toxoplasma gondii (T. gondii), a parasite that infects nearly all warm-blooded animals, including humans, cats, and pigs. Beyond incurring substantial economic losses to the swine industry, T. gondii infection poses a severe threat to public health, as pigs, which are key intermediate hosts for T. gondii transmission, serve as a major source of human infection via the food chain. Thus, developing specific, sensitive, rapid, and easy-to-perform detection methods for porcine T. gondii is of paramount importance for the prevention, control, and eventual eradication of this pathogen in swine populations. To address this need, a novel colloidal gold immunochromatographic strip (CGIS) was developed for the rapid serological detection of T. gondii infection in swine. The strip was constructed using recombinant surface antigen 1 (SAG1) expressed in a baculovirus-insect cell system, which ensures that the protein retains native antigenic epitopes. The purified SAG1 protein was conjugated to colloidal gold nanoparticles and loaded onto a conjugate pad, while staphylococcal protein A and a polyclonal anti-SAG1 antibody were immobilized on a nitrocellulose membrane to form the test line (T line) and control line (C line), respectively. Clinical validation demonstrated that the developed CGIS exhibits high specificity, with no cross-reactivity to sera positive for 13 common porcine pathogens. The strip achieved a visual limit of detection of 1:25,600 dilution of positive serum. In parallel, a commercial enzyme-linked immunosorbent assay kit yielded positive results up to a 1:800 dilution according to the manufacturer’s diagnostic cutoff. Importantly, evaluation of 300 clinical porcine serum samples demonstrated a 98.3

Given the pivotal role of noncovalent interactions in protein-protein interactions (PPIs), exploring the hidden patterns underlying the interaction data has become essential for deciphering and evaluating PPIs. In the current study, different types of noncovalent interaction data were generated from 44848 pdb files collected from the RCSB-PDB database, based on which twenty-five machine learning algorithms were benchmarked using default parameters, with top performers selected for subsequent hyperparameter optimization. Then, optimized models underwent feature selection and were subsequently ensembled via stacking and voting classifiers before comprehensive performance evaluation on test data. Finally, 12 models were built to evaluate the relationship between PPIs and noncovalent interactions after optimization. Among them, ETsO achieved the best performance across all eight metrics (>0.9, only Specificity and MCC < 0.9), followed closely by the three stacking models (SM_et487, SM_se375 and SM_dt415) and ETsO_FS. The SHAP analysis was used for elucidating the contribution of noncovalent interactions in PPIs, which indicated that PPIs depend inherently on synergistic effects among multiple noncovalent interactions. Further feature analysis indicated a notable divergence in features using behaviors among the three models after FS, with varying frequencies of different interactions observed among the top 20 polynomial features. The current study provided new practical tools for PPI prediction and supplied valuable insights into the molecular determinants of protein recognition.

Since its discovery, porcine epidemic diarrhea virus (PEDV) has significantly affected the agricultural economy worldwide. The available commercialized coronavirus vaccines cannot adequately control emerging strains. Therefore, investigating the correlation between viruses and antiviral host factors is necessary. In this study, we showed that zinc finger protein 219 (ZNF219) was upregulated by viral nonstructural protein 12 (nsp12) upon PEDV challenge. Moreover, ZNF219 inhibited the replication of PEDV through selective autophagic degradation of the PEDV S2 protein. ZNF219 recruited TRAF6, the ubiquitin E3 ligase, to ubiquitinate the PEDV S2 protein. After recognition, the ubiquitinated PEDV S2 protein was delivered to autolysosomes via the cargo receptor p62 for degradation by autophagy, thus inhibiting the proliferation of PEDV. To summarize, after sensing PEDV infection by recognizing the viral nsp12 protein, host cells upregulated the intracellular expression of ZNF219, which degraded the viral S2 protein by activating autophagy, thus suppressing viral replication. Our study revealed a novel antiviral mechanism involving ZNF219 and provided a novel target for preventing and treating PEDV.

Porcine epidemic diarrhea virus (PEDV) induces acute enteric disease in swine, finally leading to neonatal piglets dehydration to death, and which has a great side effect on China's pig-breeding industry. Therefore, it is urgent to find an economical and effective treatment to prevent and control PEDV. In this study, we identified a high-affinity peptide P6 that specifically targets the C-terminal domain (CTD) of PEDV S1 protein. Subsequently, peptide P6 was conjugated to poly (lactic-co-glycolic acid) (PLGA) via dehydration synthesis to generate PLGA-P6 nanoparticles. The antiviral effect of PLGA-P6 nanoparticles was evaluated through cell counting kit-8 (CCK-8), real-time fluorescence quantitative PCR (qRT-PCR), Western blot and indirect immunofluorescence (IFA). Results showed that peptide P6 exhibited high binding affinity to PEDV S1 protein, among which hydrogen and electrostatic interactions are the key between peptide P6 and PEDV S1 active pocket. Toxicity test suggested that cell viability was >95 % when treated with PLGA-P6 nanoparticles at concentrations not exceeding 1000 μg/mL. Furthermore, absolute quantitative PCR demonstrated that 400 μg/mL PLGA-P6 nanoparticles significantly reduce viral load of PEDV compared to the virus group (P < 0.0001). Similarly, Western blot and indirect immunofluorescence also suggested that the antiviral effect of PLGA-P6 nanoparticles at 400 μg/mL is still significant. Based on the above research, the strategy that PLGA nanoparticles combined with antiviral peptides can reduce PEDV infection to a certain extent, which not only lays a foundation for the development of new PEDV drugs, but also provides an auxiliary comprehensive prevention and control method for the prevalence of PEDV.

Porcine epidemic diarrhea virus (PEDV) leads to a high mortality in neonatal piglets and causes serious harm to the global swine industry. PEDV has been shown to exploit diverse strategies for antagonism of host innate immunity and promotion of self-replication. However, the underlying mechanisms involved in PEDV immunosuppression remain to be fully elucidated. The current study reveals that PEDV triggers mitophagy to suppress host innate immune responses and facilitate viral proliferation. Mechanistically, PEDV non-structural protein (Nsp) 14 was identified to mediate the interaction between the mitophagy receptor NDP52 and mitochondrial outer membrane protein TOM20 to induce mitophagy. Subsequently, Nsp14-induced mitophagy resulted in the degradation of mitochondrial antiviral signaling protein (MAVS) to suppress interferon-β (IFN-β) production and promote viral propagation. These findings deepen the understanding of PEDV pathogenesis and provide novel targets for the development of antiviral avenues. IMPORTANCE:The global pig farming industry has suffered huge economic losses from PEDV, underscoring an urgent need for in-depth research on its pathogenesis. Host innate immunity functions as the first line of defense against PEDV propagation, and PEDV has developed multiple countermeasures to dampen host antiviral responses. Here, we found that PEDV Nsp14 induced mitophagy via mediating the interaction between NDP52 and TOM20, which led to MAVS degradation and hampered IFN-β production. Therefore, our work unveils a novel mechanism by which PEDV antagonizes host innate immunity to facilitate its proliferation and is beneficial for the prevention and control of the virus.

Porcine epidemic diarrhea (PED) is a highly pathogenic and infectious intestinal disease caused by the PED virus (PEDV) and has inflicted substantial economic losses on the global swine industry. Therefore, it is imperative to explore appropriate targets to restrain PEDV infection. PEDV spike (S) protein is crucial for viral infection and is regarded as an ideal target for the development of vaccines and antiviral therapeutics. Palmitoylation is a significant post-translational modification implicated in multiple viral replication cycles. Despite the fact that palmitoylation of certain coronavirus S proteins has been reported, the specific biological significance and underlying molecular mechanisms of PEDV S protein palmitoylation have not been fully defined. In the present study, we uncover that palmitoylation enhances the stability of PEDV S protein to promote viral proliferation. Mechanistically, we identify that a cysteine-rich region within the cytoplasmic tail of PEDV S protein is palmitoylated by the zinc finger Asp-His-His-Cys domain palmitoyltransferase 5 (ZDHHC5). We further illustrate that palmitoylation prevents the recognition of Lys-Phe-Glu-Arg-Gln (KFERQ)-like motif in PEDV S protein by heat shock cognate protein of 70 kDa (HSC70), thereby antagonizing its degradation via chaperone-mediated autophagy (CMA). Collectively, our findings underscore the importance of palmitoylation for PEDV pathogenesis and provide prospective targets for the development of antiviral interventions.IMPORTANCEPEDV poses a serious threat to pig farming worldwide. As a consequence, a comprehensive investigation of PEDV pathogenesis is of great significance for the prevention and control of the virus. Here, we verify that ZDHHC5-mediated palmitoylation of PEDV S protein enhances its stability through impeding recognition by HSC70 and antagonizing degradation via CMA to facilitate viral propagation. Our findings highlight the important role of palmitoylation in PEDV proliferation and support palmitoylation as a promising target for the development of antiviral strategies.

BackgroundFowl adenovirus serotype 4 (FAdV-4) is the main pathogen of hepatitis-hydropericardium syndrome (HHS), which brings huge economic losses to the poultry industry worldwide. Fiber-1 protein plays an important role in viral infection and pathogenesis by binding directly to cellular receptors of FAdV-4. In particular, the knob domain of fiber-1 protein has been reported to induce the production of neutralizing antibodies and arouse protection against the lethal challenge of chickens with FAdV-4.MethodsThe fiber-1 knob (F1K) protein was expressed in a prokaryotic expression system and purified using Ni-NTA affinity chromatography. Monoclonal antibodies (mAbs) against FAdV-4 were generated by immunizing BALB/c mice with the purified F1K protein and screened using a series of immunoassays. Potential B cell epitopes on the knob domain of fiber-1 protein were mapped using enzyme-linked immunosorbent assay (ELISA) and dot-blot. Precious location and crucial amino acids of the identified epitopes were determined using peptide array scanning, truncations and alanine-scanning mutagenesis. The epitopes were analyzed and visualized on the knob trimer of FAdV-4 fiber-1 protein using the PyMOL software.ResultsWater-soluble recombinant fiber-1 knob (F1K) protein was obtained with the assistance of chaperone. Four monoclonal antibodies (5C10, 6F8, 8D8, and 8E8) against FAdV-4 were generated and characterized using indirect ELISA, Western blot, dot-blot, and immunological fluorescence assay (IFA). The mAbs were demonstrated to be from different hybridoma cell lines based on the sequences of the variable regions. Meanwhile, three distinct novel linear B-cell epitopes (319SDVGYLGLPPH329, 328PHTRDNWYV336, and 407VTTGPIPFSYQ417) on the knob domain of fiber-1 protein were identified and the key amino acid residues in the epitopes were determined. Structural analysis showed that the two adjacent epitopes 319SDVGYLGLPPH329 and 328PHTRDNWYV336 were exposed on the surface of the fiber-1 knob trimer, whereas the epitope 407VTTGPIPFSYQ417 was located inside of the spatial structure.ConclusionThis was the first identification of B-cell epitopes on the knob domain of fiber-1 protein and these findings provided a sound basis for the development of subunit vaccines, therapeutics, and diagnostic methods to control FAdV infections.

Classical swine fever (CSF), caused by CSF virus (CSFV), is a highly contagious disease affecting pigs and causing massive pig production losses with severe global economic recession. The immunization of live-attenuated vaccines is still one of the key measures to CSFV management in endemic countries. However, there are also strong controversies about the usage of live-attenuated vaccines, particularly in pregnant sows and young pigs, such as in Europe, where domestic pigs are routinely not vaccinated until severe outbreaks occur. Here, we report a CSF outbreak in a pig farm in China, which affected more than 90% of the delivery sows and led to ∼45% birth loss. Surprisingly, phylogenetic analysis showed that the CSFV isolate (named CSFV/HeNLY2022, GenBank No. OR195698) was clustered into subgenotype 1.1a, closely together with the live-attenuated vaccine strains. Further genomic analysis also revealed that the isolate CSFV/HeNLY2022 shared the highest nucleotide identity of 99.7% with the C/HVRI vaccine strain (C-strain, GenBank No. AY805221). Moreover, compared to the C/HVRI strain, a total of eight amino acid mutations, distributed in Erns (H436thY and S476thR), E1 (T502thI and P581thT), E2 (M979thK and A1061thS), NS5A (A2980thT), and NS5B (I3818thM), were characterized in the CSFV/HeNLY2022 isolate. Our results suggested that the CSF outbreak was most likely caused by the live-attenuated CSFV vaccine or its derivative. It raises concern that the unscientific application of CSFV vaccines could potentially lead to CSFV spread in pigs. It is needed to perform a more rigorous evaluation of the safety of the C-strain-derived vaccines in combination with other different live-attenuated vaccines.

Background Porcine epidemic diarrhea virus (PEDV) can cause diarrhea, dehydration and death in suckling piglets, which seriously affects the economic benefits of the production line. Therefore, it is urgent to find an economical and effective treatment to prevent and control PEDV. Methods peptide (P6), which could specifically target the S1 C-terminal domain (CTD) protein of porcine epidemic diarrhea virus (PEDV), was subsequently conjugated to poly (lactic-co-glycolic acid) (PLGA) by dehydration synthesis generating P6-PLGA nanoparticles and used cell counting kit-8 (CCK-8), real-time fluorescence quantitative PCR (qRT-PCR), Western blot and indirect immunofluorescence to further study the inhibitory effect of different concentrations of P6-PLGA nanoparticles on PEDV. Results The results showed that cell viability was > 95% when treated with P6-PLGA nanoparticles at concentrations not exceeding 1000 µg/ml. Results of the absolute quantitative PCR revealed that the concentration of P6-PLGA nanoparticles at 400 µg/ml could significantly reduce the viral load of PEDV compared with the virus group (p < 0.05 or p < 0.001). Similarly, results of Western blot and indirect immunofluorescence also suggested that the antiviral effect of P6-PLGA nanoparticles at 400 µg/ml is still significant. Based on the above research, high affinity peptide (P6) was covalently coupled with PLGA particles to obtain P6-PLGA nanoparticles. Conclusions PLGA as a drug delivery carrier combined with peptide (P6) can overcome the problems of poor stability, easy degradation or low bioavailability of peptide after entering the body, and provide a new strategy for the development of PEDV antiviral drugs.

Because of their high affinity, specificity, and environmental stability, nanobodies (Nbs) have continuously received attention from the field of biological research. However, it is tough work to obtain high-affinity Nbs using experimental methods. In the current study, 12 machine learning algorithms were compared in parallel to explore the potential patterns between Nb-ligand affinity and eight noncovalent interactions. After model comparison and optimization, four optimized models (SVMrB, RotFB, RFB, and C50B) and two stacked models (StackKNN and StackRF) based on nine uncorrelated (correlation coefficient <0.65) optimized models were selected. All the models showed an accuracy of around 0.70 and high specificity. Compared to the other models, RotFB and RFB were not capable of predicting nonaffinitive Nbs with lower precision (<0.44) but showed higher sensitivity at 0.6761 and 0.3521 and good model robustness (F1 score and MCC values). On the contrary, SVMrB, C50B, and StackKNN were able to effectively predict the future nonaffinitive Nbs (specificity >0.92) and reduce the number of true affinitive Nbs (precision >0.5). On the other hand, StackRF showed intermediate model performance. Furthermore, an in-depth feature analysis indicated that hydrogen bonding and aromatic-associated interactions were the key noncovalent interactions in determining Nb-ligand binding affinity. In summary, the current study provides, for the first time, a tool that can effectively predict whether there is an affinity between nanobodies and their intended ligands and explores the key factors that influence their affinity, which could improve the screening and design process of Nbs and accelerate the development of Nb drugs and applications.

ABSTRACT Viruses employ various evasion strategies to establish prolonged infection, with evasion of innate immunity being particularly crucial. Porcine reproductive and respiratory syndrome virus (PRRSV) is a significant pathogen in swine industry, characterized by reproductive failures in sows and respiratory distress in pigs of all ages, leading to substantial economic losses globally. In this study, we found that the non-structural protein 5 (Nsp5) of PRRSV antagonizes innate immune responses via inhibiting the expression of type I interferon (IFN-I) and IFN-stimulated genes (ISGs), which is achieved by degrading multiple proteins of RIG-I-like receptor (RLR) signaling pathway (RIG-I, MDA5, MAVS, TBK1, IRF3, and IRF7). Furthermore, we showed that PRRSV Nsp5 is located in endoplasmic reticulum (ER), where it promotes accumulation of RLR signaling pathway proteins. Further data demonstrated that Nsp5 activates reticulophagy (ER-phagy), which is responsible for the degradation of RLR signaling pathway proteins and IFN-I production. Mechanistically, Nsp5 interacts with one of the ER-phagy receptor family with sequence similarity 134 member B (FAM134B), promoting the oligomerization of FAM134B. These findings elucidate a novel mechanism by which PRRSV utilizes FAM134B-mediated ER-phagy to elude host antiviral immunity. IMPORTANCE Innate immunity is the first line of host defense against viral infections. Therefore, viruses developed numerous mechanisms to evade the host innate immune responses for their own benefit. PRRSV, one of the most important endemic swine viruses, poses a significant threat to the swine industry worldwide. Here, we demonstrate for the first time that PRRSV utilizes its non-structural protein Nsp5 to degrade multiple proteins of RLR signaling pathways, which play important roles in IFN-I production. Moreover, FAM134B-mediated ER-phagy was further proved to be responsible for the protein’s degradation. Our study highlights the critical role of ER-phagy in immune evasion of PRRSV to favor replication and provides new insights into the prevention and control of PRRSV.

Macroautophagy/autophagy is a cellular degradation and recycling process that maintains the homeostasis of organisms. A growing number of studies have reported that autophagy participates in infection by a variety of viruses. Porcine reproductive and respiratory syndrome virus (PRRSV) causes severe financial losses to the global swine industry. Although much research has shown that PRRSV triggers autophagy for its own benefits, the exact molecular mechanisms involved in PRRSV-triggered autophagy remain to be fully elucidated. In the current study, we demonstrated that PRRSV infection significantly induced Golgi apparatus (GA) fragmentation, which promoted autophagy to facilitate viral self-replication. Mechanistically, PRRSV nonstructural protein 2 was identified to interact with and degrade the Golgi reassembly and stacking protein 65 dependent on its papain-like cysteine protease 2 activity, resulting in GA fragmentation. Upon GA fragmentation, GA-resident Ras-like protein in brain 2 was disassociated from Golgi matrix protein 130 and subsequently bound to unc-51 like autophagy activating kinase 1 (ULK1), which enhanced phosphorylation of ULK1 and promoted autophagy. Taken together, all these results expand the knowledge of PRRSV-triggered autophagy as well as PRRSV pathogenesis to support novel potential avenues for prevention and control of the virus. More importantly, these results provide the detailed mechanism of GA fragmentation-mediated autophagy, deepening the understanding of autophagic processes.IMPORTANCEPorcine reproductive and respiratory syndrome virus (PRRSV) infection results in a serious swine disease affecting pig farming worldwide. Despite that numerous studies have shown that PRRSV triggers autophagy for its self-replication, how PRRSV induces autophagy is incompletely understood. Here, we identify that PRRSV Nsp2 degrades GRASP65 to induce GA fragmentation, which dissociates RAB2 from GM130 and activates RAB2-ULK1-mediated autophagy to enhance viral replication. This work expands our understanding of PRRSV-induced autophagy and PRRSV replication, which is beneficial for anti-viral drug development.

RESEARCH HIGHLIGHTS:A sandwich ELISA was developed to detect EDSV using the mAbs 5G4 and HRP-6G6.The sandwich ELISA maintained high specificity and sensitivity.The sandwich ELISA had equivalent consistency with real-time PCR assay.

Background Porcine deltacoronavirus (PDCoV) is one of the emerging swine enteric coronaviruses (SECoVs), which has been widely prevalent in the North America and Asia. In addition to causing severe diarrhea in piglets, PDCoV also shows the potential to infect diverse host species, including calves, chickens, turkey poults, and humans. However, the clinical pathogenicity and genetic evolution of PDCoV is still not fully understood. Results Here, we recorded an outbreak of a novel recombinant PDCoV strain (CHN-HeN06-2022) in a large nursery fattening pig farm. Genomic analysis showed that the CHN-HeN06-2022 strain shared 98.3-98.7% sequence identities with the Chinese and American reference strains. To clarify the evolutionary relationships, phylogenetic analysis was performed using the PDCoV genome sequences available in the GenBank database. Based on genetic distance and geographical distribution, the phylogenetic tree clearly showed that all the PDCoV sequences could be divided into lineage 1 and lineage 2, which were further classified into sublineage 1.1 (Chinese strains), 1.2 (the North American strains), 2.1 (the Southeast Asian strains), and 2.2 (Chinese strains). Corresponding to the evolutionary tree, we found that, compared to lineage 1, lineage 2 strains usually contain a continuous 6-nt deletion in Nsp2 and a 9-nt deletion in Nsp 3, respectively. Furthermore, recombination analysis suggested that the CHN-HeN06-2022 occurred segments exchange crossed Nsp2 and Nsp3 region between sublineage 1.1 and sublineage 2.1. Combined with previously reported recombinant strains, the highest recombination frequency occurred in Nsp2 , Nsp3 , and S gene. Additionally, we identified a total of 14 amino acid sites under positive selection in spike protein, most of which are located in the regions related with the viral attachment, receptor binding, and membrane fusion. Conclusions Taken together, our studies provide novel insights into the genetic diversity and adaptive evolution of PDCoV. It would be helpful to the development of vaccine and potential antiviral agent.

This review aims to elucidate the mechanisms employed by antiviral peptides in inhibiting coronaviruses as well as related limitations their therapeutic applications, and how to address the limitations faced as potential drugs in targeted therapy infectious diseases. The epidemic of MERS-CoV, SARS-CoV or COVID-19 have attracted more attention due to its the high mortality rate, among which, SARS-CoV-2 infections led that more than 625 million confirmed cases and more than 6 million deaths, the mortality rate of MERS-CoV reached 35.5