Insights and progress on epidemic characteristics, genotyping, and preventive measures of PEDV in China: A review.

Porcine Epidemic Diarrhoea (PED) is an acute, extremely infectious intestinal disease of pigs caused by the Porcine Epidemic Diarrhoea Virus (PEDV). The virus can affect pigs of all breeds and age groups and shows varying degrees of symptoms, with piglets, in particular, being infected with mortality rates of up to 100%. PEDV was first identified in China in the 1980s and in October 2010 a large-scale PED outbreak caused by a variant of PEDV occurred in China, resulting in huge economic losses. Initially, vaccination can effectively prevent the classical strain, but since December 2010, the PEDV variant has caused "persistent diarrhoea" with severe vomiting, watery diarrhoea, and high morbidity and mortality in newborn piglets as the dominant clinical features, with a significant increase in morbidity and mortality. This indicates that PEDV strains have mutated during evolution and that traditional vaccines no longer provide effective cross-immune protection, so it is necessary to optimize immunization programs and find effective treatments through epidemiological surveys of PEDV to reduce the economic losses caused by infections with mutated strains. This article reviews the progress of research on the aetiology, epidemiological characteristics, genotyping, pathogenesis, transmission routes, and comprehensive control of PEDV infection in China.

Construction and immunogenicity of a trypsin-independent porcine epidemic diarrhea virus variant.

Porcine epidemic diarrhea virus (PEDV) is a re-emerging enteropathogenic coronavirus that causes high mortality in neonatal piglets. The addition of trypsin plays a crucial role in the propagation of PEDV, but also increases the complexity of vaccine production and increases its cost. Previous studies have suggested that the S2' site and Y976/977 of the PEDV spike (S) protein might be the determinants of PEDV trypsin independence. In this study, to achieve a recombinant trypsin-independent PEDV strain, we used trypsin-dependent genotype 2 (G2) PEDV variant AJ1102 to generate three recombinant PEDVs with mutations in S (S2' site R894G and/or Y976H). The three recombinant PEDVs were still trypsin dependent, suggesting that the S2' site R894 and Y976 of AJ1102 S are not key sites for PEDV trypsin dependence. Therefore, we used AJ1102 and the classical trypsin-independent genotype 1 (G1) PEDV strain JS2008 to generate a recombinant PEDV carrying a chimeric S protein, and successfully obtained trypsin-independent PEDV strain rAJ1102-S2'JS2008, in which the S2 (amino acids 894-1386) domain was replaced with the corresponding JS2008 sequence. Importantly, immunization with rAJ1102-S2'JS2008 induced neutralizing antibodies against both AJ1102 and JS2008. Collectively, these results suggest that rAJ1102-S2'JS2008 is a novel vaccine candidate with significant advantages, including no trypsin requirement for viral propagation to high titers and the potential provision of protection for pigs against G1 and G2 PEDV infections.

miRNAs derived from milk small extracellular vesicles inhibit porcine epidemic diarrhea virus infection.

Porcine epidemic diarrhea virus (PEDV), a member of the genus Alphacoronavirus in the family Coronaviridae, causes acute diarrhea and/or vomiting, dehydration, and high mortality in neonatal piglets. It has caused huge economic losses to animal husbandry worldwide. Current commercial PEDV vaccines do not provide enough protection against variant and evolved virus strains. No specific drugs are available to treat PEDV infection. The development of more effective therapeutic anti-PEDV agents is urgently needed. Our previous study suggested that porcine milk small extracellular vesicles (sEV) facilitate intestinal tract development and prevent lipopolysaccharide-induced intestinal injury. However, the effects of milk sEV during viral infection remain unclear. Our study found that porcine milk sEV, which was isolated and purified by differential ultracentrifugation, could inhibit PEDV replication in IPEC-J2 and Vero cells. Simultaneously, we constructed a PEDV infection model for piglet intestinal organoids and found that milk sEV also inhibited PEDV infection. Subsequently, in vivo experiments showed that milk sEV pre-feeding exerted robust protection of piglets from PEDV-induced diarrhea and mortality. Strikingly, we found that the miRNAs extracted from milk sEV inhibited PEDV infection. miRNA-seq, bioinformatics analysis, and experimental verification demonstrated that miR-let-7e and miR-27b, which were identified in milk sEV targeted PEDV N and host HMGB1, suppressed viral replication. Taken together, we revealed the biological function of milk sEV in resisting PEDV infection and proved its cargo miRNAs, miR-let-7e and miR-27b, possess antiviral functions. This study is the first description of the novel function of porcine milk sEV in regulating PEDV infection. It provides a better understanding of milk sEV resistance to coronavirus infection, warranting further studies to develop sEV as an attractive antiviral.

Multiepitope Subunit Peptide-Based Nanovaccine against Porcine Circovirus Type 2 (PCV2) Elicited High Antibody Titers in Vaccinated Mice.

Porcine circovirus 2 (PCV2) infection is one of the most serious threats to the swine industry. While the disease can be prevented, to some extent, by commercial PCV2a vaccines, the evolving nature of PCV2 necessitates the development of a novel vaccine that can compete with the mutations of the virus. Thus, we have developed novel multiepitope vaccines based on the PCV2b variant. Three PCV2b capsid protein epitopes, together with a universal T helper epitope, were synthesized and formulated with five delivery systems/adjuvants: complete Freund's adjuvant, poly(methyl acrylate) (PMA), poly(hydrophobic amino acid), liposomes and rod-shaped polymeric nanoparticles built from polystyrene-poly(N-isopropylacrylamide)-poly(N-dimethylacrylamide). Mice were subcutaneously immunized with the vaccine candidates three times at three-week intervals. All vaccinated mice produced high antibody titters after three immunizations as analyzed by the enzyme-linked immunosorbent assay (ELISA), while mice vaccinated with PMA-adjuvanted vaccine elicited high antibody titers even after a single immunization. Thus, the multiepitope PCV2 vaccine candidates designed and examined here show strong potential for further development.

Modeling between-farm transmission dynamics of porcine epidemic diarrhea virus: Characterizing the dominant transmission routes.

The role of transportation vehicles, pig movement between farms, proximity to infected premises, and feed deliveries has not been fully considered in the dissemination dynamics of porcine epidemic diarrhea virus (PEDV). This has limited efforts for disease prevention, control and elimination restricting the development of risk-based resource allocation to the most relevant modes of PEDV dissemination. Here, we modeled nine pathways of between-farm transmission represented by a contact network of pig movements between sites, farm-to-farm proximity (local transmission), four distinct contact networks of transportation vehicles (trucks that transport pigs from farm-to-farm and farm-to-markets, as well as trucks transporting feed and staff), the volume of animal by-products in feed diets (e.g., fat and meat-and-bone-meal) to reproduce PEDV transmission dynamics. The model was calibrated in space and time with weekly PEDV outbreaks. We investigated the model performance to identify outbreak locations and the contribution of each route in the dissemination of PEDV. The model estimated that 42.7% of the infections in sow farms were related to vehicles transporting feed, 34.5% of infected nurseries were associated with vehicles transporting pigs between farms, and for both farm types, local transmission or pig movements were the next most relevant transmission routes. On the other hand, finishers were most often (31.4%) infected via local transmission, followed by the vehicles transporting feed and pigs between farms. Feed ingredients did not significantly improve model calibration metrics, sensitivity, and specificity; therefore, it was considered to have a negligible contribution in the dissemination of PEDV. The proposed modeling framework provides an evaluation of PEDV transmission dynamics, ranking the most important routes of PEDV dissemination and granting the swine industry valuable information to focus efforts and resources on the most important transmission routes.

Three Amino Acid Substitutions in the Spike Protein Enable the Coronavirus Porcine Epidemic Diarrhea Virus To Infect Vero Cells.

Porcine epidemic diarrhea virus (PEDV), a continuously evolving pathogen, causes severe diarrhea in piglets, with high mortality rates. To prevent or mitigate the disease, it is common practice to develop live or inactivated PEDV vaccines based on cell-adapted viral variants. Propagating wild-type PEDV in cultured cells is, however, often challenging due to the lack of knowledge about the requirements for the cell adaptation of PEDV. In the present study, by using the RNA-targeted reverse genetic system for PEDV to apply S protein swapping followed by the rescue of the recombinant viruses, three key amino acid mutations in the S protein, A605E, E633Q, and R891G, were identified, which enable attenuated PEDV strain DR13 (DR13att) to efficiently and productively infect Vero cells, in contrast to the parental DR13 strain (DR13par). The former two key mutations reside inside and in the vicinity of the receptor binding domain (RBD), respectively, while the latter occurs at the N-terminal end of the fusion peptide (FP). Besides the three key mutations, other mutations in the S protein further enhanced the infection efficiency of the recombinant viruses. We hypothesize that the three mutations changed PEDV tropism by altering the S2' cleavage site and the RBD structure. This study provides basic molecular insight into cell adaptation by PEDV, which is also relevant for vaccine design. IMPORTANCE 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. In this study, we found that three amino acid mutations, A605E, E633Q, and R891G, in the spike protein of the Vero cell-adapted PEDV strain DR13att were critical for its cell adaptation. After analyzing the mutation sites in the spike protein, we hypothesize that the cell adaptation of DR13att was achieved by altering the S2' cleavage site and the RBD structure. This study provides new molecular insight into the mechanism of PEDV culture adaptation and new strategies for PEDV vaccine design.

Innate Immune Evasion of Porcine Epidemic Diarrhea Virus through Degradation of the FBXW7 Protein via the Ubiquitin-Proteasome Pathway.

Porcine epidemic diarrhea virus (PEDV) causes a porcine disease associated with swine epidemic diarrhea. Different antagonistic strategies have been identified, and the mechanism by which PEDV infection impairs the production of interferon (IFN) and delays the activation of the IFN response to escape host innate immunity has been determined, but the pathogenic mechanisms of PEDV infection remain enigmatic. Our preliminary results revealed that endogenous F-box and WD repeat domain-containing 7 (FBXW7) protein, the substrate recognition component of the SCF-type E3 ubiquitin ligase, is downregulated in PEDV-infected Vero E6 cells, according to the results from an isobaric tags for relative and absolute quantification (iTRAQ) analysis. Overexpression of FBXW7 in target cells makes them more resistant to PEDV infection, whereas ablation of FBXW7 expression by small interfering RNA (siRNA) significantly promotes PEDV infection. In addition, FBXW7 was verified as an innate antiviral factor capable of enhancing the expression of RIG-I and TBK1, and it was found to induce interferon-stimulated genes (ISGs), which led to an elevated antiviral state of the host cells. Moreover, we revealed that PEDV nonstructural protein 2 (nsp2) interacts with FBXW7 and targets FBXW7 for degradation through the K48-linked ubiquitin-proteasome pathway. Consistent with the results proven in vitro, FBXW7 reduction was also confirmed in different intestinal tissues from PEDV-infected specific-pathogen-free (SPF) pigs. Taken together, the data indicated that PEDV has evolved with a distinct antagonistic strategy to circumvent the host antiviral response by targeting the ubiquitin-proteasome-mediated degradation of FBXW7. Our findings provide novel insights into PEDV infection and pathogenesis. IMPORTANCE To counteract the host antiviral defenses, most viruses, including coronaviruses, have evolved with diverse strategies to dampen host IFN-mediated antiviral response, by interfering with or evading specific host regulators at multiple steps of this response. In this study, a novel antagonistic strategy was revealed showing that PEDV infection could circumvent the host innate response by targeted degradation of endogenous FBXW7 in target cells, a process that was verified to be a positive modulator for the host innate immune system. Degradation of FBXW7 hampers host innate antiviral activation and facilitates PEDV replication. Our findings reveal a new mechanism exploited by PEDV to suppress the host antiviral response.

The between-farm transmission dynamics of porcine epidemic diarrhoea virus: A short-term forecast modelling comparison and the effectiveness of control strategies.

A limited understanding of the transmission dynamics of swine disease is a significant obstacle to prevent and control disease spread. Therefore, understanding between-farm transmission dynamics is crucial to developing disease forecasting systems to predict outbreaks that would allow the swine industry to tailor control strategies. Our objective was to forecast weekly porcine epidemic diarrhoea virus (PEDV) outbreaks by generating maps to identify current and future PEDV high-risk areas, and simulating the impact of control measures. Three epidemiological transmission models were developed and compared: a novel epidemiological modelling framework was developed specifically to model disease spread in swine populations, PigSpread, and two models built on previously developed ecosystems, SimInf (a stochastic disease spread simulations) and PoPS (Pest or Pathogen Spread). The models were calibrated on true weekly PEDV outbreaks from three spatially related swine production companies. Prediction accuracy across models was compared using the receiver operating characteristic area under the curve (AUC). Model outputs had a general agreement with observed outbreaks throughout the study period. PoPS had an AUC of 0.80, followed by PigSpread with 0.71, and SimInf had the lowest at 0.59. Our analysis estimates that the combined strategies of herd closure, controlled exposure of gilts to live viruses (feedback) and on-farm biosecurity reinforcement reduced the number of outbreaks. On average, 76% to 89% reduction was seen in sow farms, while in gilt development units (GDU) was between 33% to 61% when deployed to sow and GDU farms located in probabilistic high-risk areas. Our multi-model forecasting approach can be used to prioritize surveillance and intervention strategies for PEDV and other diseases potentially leading to more resilient and healthier pig production systems.

First expert elicitation of knowledge on drivers of emergence of influenza D in Europe.

The influenza D virus (IDV) was first identified and characterized in 2011. Considering the virus' zoonotic potential, its genome nature (segmented RNA virus), its worldwide circulation in livestock and its role in bovine respiratory disease, an increased interest is given to IDV. However, few data are available on drivers of emergence of IDV. We first listed fifty possible drivers of emergence of IDV in ruminants and swine. As recently carried out for COVID-19 in pets (Transboundary and Emerging Diseases, 2020), a scoring system was developed per driver and scientific experts (N = 28) were elicited to (a) allocate a score to each driver, (b) weight the drivers' scores within each domain and (c) weight the different domains among themselves. An overall weighted score was calculated per driver, and drivers were ranked in decreasing order. Drivers with comparable likelihoods to play a role in the emergence of IDV in ruminants and swine in Europe were grouped using a regression tree analysis. Finally, the robustness of the expert elicitation was verified. Eight drivers were ranked with the highest probability to play a key role in the emergence of IDV: current species specificity of the causing agent of the disease; influence of (il)legal movements of live animals (ruminants, swine) from neighbouring/European Union member states and from third countries for the disease to (re-)emerge in a given country; detection of emergence; current knowledge of the pathogen; vaccine availability; animal density; and transport vehicles of live animals. As there is still limited scientific knowledge on the topic, expert elicitation of knowledge and multi-criteria decision analysis, in addition to clustering and sensitivity analyses, are very important to prioritize future studies, starting from the top eight drivers. The present methodology could be applied to other emerging animal diseases.

Epidemic situation and control measures of African Swine Fever Outbreaks in China 2018-2020.

As of 21 April 2020, 176 ASF outbreaks have occurred in China. For each outbreak, an investigation was conducted, including historical data retrieval and traceability of potential contacts. The purpose of this study is to conduct a preliminary analysis of the data obtained from the outbreak investigations, including an investigation of the possible contributing factors of the spread of ASF in China. Based on the epidemic situation and the policies issued, the entire epidemic can be divided into three phases. 71 outbreaks were reported between 3 August 2018 and 17 November 2018; 44 outbreaks between 19 November 2018 and 30 March 2019; and 61 outbreaks between 4 April 2019 and 12 April 2020. Based on the reported outbreaks, the proportional rate of outbreaks in small farms (livestock ≤ 500, 127/168) is significantly higher than that of medium (501 ≤ livestock < 2,000, 14/168; 2001 ≤ livestock ≤ 5,000, 9/168) and large farms (livestock ≥ 5,001, 18/168). The odds of infection related to swill feeding (OR = 2.5, 95% CI, 1.5-4.3) and the mechanical dissemination of vehicles and personnel (OR = 2.7, 95% CI, 1.6-4.5) are significantly higher than those of pigs and pig production transportation. Swill feeding is the major contributing factor for small farms while mechanical dissemination of vehicles and personnel is the major contributing factor for large farms. The average duration from the beginning of the infection to the official outbreak report is gradually decreasing, which means that response speed of industry entities and the animal husbandry and veterinary departments from the beginning of the infection to the outbreak report is gradually increasing. Based on the analysis for ASF outbreaks, some policies and suggestions were put forward, such as improving the biosecurity level of the farms, as well as strengthening the supervision of breeding, transportation and slaughter.

A case study of ventilation shutdown with the addition of high temperature and humidity for depopulation of pigs.

CASE DESCRIPTION: In February 2020, a swine farm operating in multiple states throughout the Midwest began to evaluate emergency plans to respond to potential impacts of the COVID-19 pandemic. CLINICAL FINDINGS: From February through April, extensive mitigation strategies were implemented in anticipation of market disruption. The farm consulted the AVMA Guidelines for the Depopulation of Animals to identify preferred methods for depopulation of swine; however, none of these methods were feasible. When the first US packing plant closed in April 2020 because of human COVID-19 infection, the farm began to evaluate whether ventilation shutdown plus (VSD+) could be used for depopulation. TREATMENT AND OUTCOME: Through proof-of-concept trials, a method for ventilation shutdown with the addition of supplemental temperature and humidity was developed. A single location with 4 barns that could be retrofitted for the process was selected, and between April and June 2020, 243,016 pigs were depopulated (59,478 nursery and 183,538 finishing pigs). Mean ± SD time to silent (the time when no sounds could be heard and no motion seen) was 55.4 ± 14.5 minutes for the nursery pigs and 65.0 ± 18.1 minutes for the finishing pigs. Only 728 (0.300%) pigs required manual euthanasia at the end of the depopulation process. CLINICAL RELEVANCE: Efficacy exceeded AVMA recommendations for the use of VSD+ (> 95% mortality rate in < 1 hour). Findings could potentially guide the use of this method for mass depopulation in the event of a foreign animal disease outbreak or severe market disruption in the future.

Vaccines That Reduce Viral Shedding Do Not Prevent Transmission of H1N1 Pandemic 2009 Swine Influenza A Virus Infection to Unvaccinated Pigs.

Swine influenza A virus (swIAV) infection causes substantial economic loss and disease burden in humans and animals. The 2009 pandemic H1N1 (pH1N1) influenza A virus is now endemic in both populations. In this study, we evaluated the efficacy of different vaccines in reducing nasal shedding in pigs following pH1N1 virus challenge. We also assessed transmission from immunized and challenged pigs to naive, directly in-contact pigs. Pigs were immunized with either adjuvanted, whole inactivated virus (WIV) vaccines or virus-vectored (ChAdOx1 and MVA) vaccines expressing either the homologous or heterologous influenza A virus hemagglutinin (HA) glycoprotein, as well as an influenza virus pseudotype (S-FLU) vaccine expressing heterologous HA. Only two vaccines containing homologous HA, which also induced high hemagglutination inhibitory antibody titers, significantly reduced virus shedding in challenged animals. Nevertheless, virus transmission from challenged to naive, in-contact animals occurred in all groups, although it was delayed in groups of vaccinated animals with reduced virus shedding.IMPORTANCE This study was designed to determine whether vaccination of pigs with conventional WIV or virus-vectored vaccines reduces pH1N1 swine influenza A virus shedding following challenge and can prevent transmission to naive in-contact animals. Even when viral shedding was significantly reduced following challenge, infection was transmissible to susceptible cohoused recipients. This knowledge is important to inform disease surveillance and control strategies and to determine the vaccine coverage required in a population, thereby defining disease moderation or herd protection. WIV or virus-vectored vaccines homologous to the challenge strain significantly reduced virus shedding from directly infected pigs, but vaccination did not completely prevent transmission to cohoused naive pigs.

Intranasal Delivery of Inactivated Influenza Virus and Poly(I:C) Adsorbed Corn-Based Nanoparticle Vaccine Elicited Robust Antigen-Specific Cell-Mediated Immune Responses in Maternal Antibody Positive Nursery Pigs.

We designed the killed swine influenza A virus (SwIAV) H1N2 antigen (KAg) with polyriboinosinic:polyribocytidylic acid [(Poly(I:C)] adsorbed corn-derived Nano-11 particle based nanovaccine called Nano-11-KAg+Poly(I:C), and evaluated its immune correlates in maternally derived antibody (MDA)-positive pigs against a heterologous H1N1 SwIAV infection. Immunologically, in tracheobronchial lymph nodes (TBLN) detected enhanced H1N2-specific cytotoxic T-lymphocytes (CTLs) in Nano-11-KAg+Poly(I:C) vaccinates, and in commercial vaccinates detected CTLs with mainly IL-17A+ and early effector phenotypes specific to both H1N2 and H1N1 SwAIV. In commercial vaccinates, activated H1N2- and H1N1-specific IFNγ+&TNFα+, IL-17A+ and central memory T-helper/Memory cells, and in Nano-11-KAg+Poly(I:C) vaccinates H1N2-specific central memory, IFNγ+ and IFNγ+&TNFα+, and H1N1-specific IL-17A+ T-helper/Memory cells were observed. Systemically, Nano-11-KAg+Poly(I:C) vaccine augmented H1N2-specific IFNγ+ CTLs and H1N1-specific IFNγ+ T-helper/Memory cells, and commercial vaccine boosted H1N2- specific early effector CTLs and H1N1-specific IFNγ+&TNFα+ CTLs, as well as H1N2- and H1N1-specific T-helper/Memory cells with central memory, IFNγ+&TNFα+, and IL-17A+ phenotypes. Remarkably, commercial vaccine induced an increase in H1N1-specific T-helper cells in TBLN and naive T-helper cells in both TBLN and peripheral blood mononuclear cells (PBMCs), while H1N1- and H1N2-specific only T-helper cells were augmented in Nano-11-KAg+Poly(I:C) vaccinates in both TBLN and PBMCs. Furthermore, the Nano-11-KAg+Poly(I:C) vaccine stimulated robust cross-reactive IgG and secretory IgA (SIgA) responses in lungs, while the commercial vaccine elicited high levels of serum and lung IgG and serum hemagglutination inhibition (HI) titers. In conclusion, despite vast genetic difference (77% in HA gene identity) between the vaccine H1N2 and H1N1 challenge viruses in Nano-11-KAg+Poly(I:C) vaccinates, compared to over 95% identity between H1N1 of commercial vaccine and challenge viruses, the virus load and macroscopic lesions in the lungs of both types of vaccinates were comparable, but the Nano-11-KAg+Poly(I:C) vaccine cleared the virus from the nasal passage better. These data suggested the important role played by Nano-11 and Poly(I:C) in the induction of polyfunctional, cross-protective cell-mediated immunity against SwIAV in MDA-positive pigs.

Virus-like particle vaccine with B-cell epitope from porcine epidemic diarrhea virus (PEDV) incorporated into hepatitis B virus core capsid provides clinical alleviation against PEDV in neonatal piglets through lactogenic immunity.

Porcine epidemic diarrhea virus (PEDV) has had a negative economic impact on the global swine industry for decades since its first emergence in the 1970s in Europe. In 2013, PEDV emerged for the first time in the United States, causing immense economic losses to the swine industry. Efforts to protect U.S. swine herds from PEDV infection and limit PEDV transmission through vaccination had only limited success so far. Following the previous success in our virus-like particle (VLP) based vaccine in mouse model, in this study we determined the immunogenicity and protective efficacy of a VLP-based vaccine containing B-cell epitope 748YSNIGVCK755 from the spike protein of PEDV incorporated into the hepatitis B virus core capsid (HBcAg), in a comprehensive pregnant gilt vaccination and piglet challenge model. The results showed that the vaccine was able to induce significantly higher virus neutralization response in gilt milk, and provide alleviation of clinical signs for piglets experimentally infected with PEDV. Piglets from pregnant gilt that was vaccinated with the VLP vaccine had faster recovery from the clinical disease, less small intestinal lesions, and higher survival rate at 10 days post-challenge (DPC).

Effect of dietary medium-chain fatty acids on nursery pig growth performance, fecal microbial composition, and mitigation properties against porcine epidemic diarrhea virus following storage.

An experiment was conducted to evaluate the effect of dietary medium-chain fatty acid (MCFA) addition on nursery pig growth performance, fecal microbial composition, and mitigation of porcine epidemic diarrhea virus (PEDV) following storage. A total of 360 pigs (DNA 400 × 200, Columbus, NE; initially 6.7 ± 0.07 kg) were randomized to pens (5 pigs per pen) on the day of weaning (approximately 20 d of age), allowed a 6-d acclimation, blocked by BW, and randomized to dietary treatment (9 pens per treatment). All MCFA (Sigma-Aldrich, St. Louis, MO) were guaranteed ≥98% purity, including hexanoic (C6:0), octanoic (C8:0), and decanoic (C10:0) acids. Treatment diets were formulated in 2 phases (7 to 11 and 11 to 23 kg BW) and formulated to meet or exceed NRC requirement estimates. Treatments (n = 8) were a dose response including 0%, 0.25%, 0.5%, 1.0%, and 1.5% added MCFA blend (1:1:1 ratio C6:0, C8:0, and C10:0), as well as treatments with individual additions of 0.5% C6:0, C8:0, or C10:0. Fecal samples were collected from pigs fed control and 1.5% MCFA blend diets on days 0 and 14 and analyzed using 16s rDNA sequencing. Following feed manufacture, feed was stored in bags at barn temperature and humidity for 40 d before laboratory inoculation with PEDV. Subsamples of retained feed were inoculated with PEDV to achieve a titer of 104 TCID50/g and separate sample bottles were analyzed on 0 and 3 d post-inoculation (dpi). Overall, ADG and ADFI were increased (linear, P ≤ 0.010) and feed efficiency (G:F) improved (linear, P = 0.004) with increasing MCFA blend. Pigs fed 0.5% C8:0 had greater (P = 0.038) ADG compared with pigs fed the control diet, and G:F was improved (P ≤ 0.024) when pigs were fed 0.5% C6:0, 0.5% C8:0, or 0.5% C10:0 compared with control. An inclusion level × day interaction was observed (quadratic, P = 0.023), where PEDV Ct values increased (quadratic, P = 0.001) on 0 dpi with increasing levels of MCFA blend inclusion and also increased on 3 dpi (linear, P < 0.001). Fecal microbial diversity and composition were similar between control and 1.5% MCFA blend. In summary, the use of MCFA in nursery pig diets improves growth performance, provides residual mitigation activity against PEDV, and does not significantly alter fecal microbial composition.

Recombinant adenovirus carrying a core neutralizing epitope of porcine epidemic diarrhea virus and heat-labile enterotoxin B of Escherichia coli as a mucosal vaccine.

Porcine epidemic diarrhea virus (PEDV) targets the intestinal mucosa in pigs. To protect against PEDV invasion, a mucosal vaccine is utilized effectively. In this study, we generated a recombinant adenovirus vaccine encoding the heat-labile enterotoxin B (LTB) and the core neutralizing epitope (COE) of PEDV (rAd-LTB-COE). The fusion protein LTB-COE was successfully expressed by the recombinant adenovirus in HEK293 cells, and the immunogenicity of the vaccine candidate was assessed in BALB/c mice and piglets. Three intramuscular or oral vaccinations with rAd-LTB-COE at two-week intervals induced robust humoral and mucosal immune responses. Moreover, a cell-mediated immune response was promoted in immunized mice, and the neutralizing antibody inhibited both the vaccine strain and the emerging PEDV isolate. Immunization experiments in piglets revealed that rAd-LTB-COE was immunogenic and induced good immune responses in piglets. Further studies are required to evaluate the efficacy of rAd-LTB-COE against a highly virulent PEDV challenge.

Updated phylogenetic analysis of the spike gene and identification of a novel recombinant porcine epidemic diarrhoea virus strain in Taiwan.

New variants of porcine epidemic diarrhoea virus (PEDV) causing a highly contagious intestinal disease, porcine epidemic diarrhoea virus (PED), have resulted in high mortality in suckling pigs across several countries since 2013. After 2015, the prevalence of the genogroup 2b (G2b) PEDVs decreased in a cyclical pattern with endemic seasonal outbreaks occasionally seen. To better understand the genetic diversity of PEDVs recently circulating in Taiwan, full-length spike (S) genes of 31 PEDV strains from 28 pig farms collected during 2016-2018 were sequenced. While the majority of S gene sequences (from 27/28 farms) were closely related to the previous G2b PEDV strains, increased genetic diversities leading to several nonsynonymous mutations scattering in the neutralizing epitopes of the S gene were detected in PEDVs recently circulating in Taiwan. Furthermore, novel recombinant variants, the PEDV TW/Yunlin550/2018 strains exhibiting recombinant events between a previously isolated Taiwan PEDV G2b strain and a wild-type PEDV G1a strain, were identified and further classified into a new genogroup, G1c. These results provide updated information about the genetic diversity of currently circulating PEDVs in the field and could help to develop more suitable strategies for controlling this disease.

Porcine epidemic diarrhea virus: Molecular mechanisms of attenuation and vaccines.

Porcine epidemic diarrhea virus (PEDV) causes an emerging and re-emerging coronavirus disease characterized by vomiting, acute diarrhea, dehydration, and up to 100% mortality in neonatal suckling piglets, leading to huge economic losses in the global swine industry. Vaccination remains the most promising and effective way to prevent and control PEDV. However, effective vaccines for PEDV are still under development. Understanding the genomic structure and function of PEDV and the influence of the viral components on innate immunity is essential for developing effective vaccines. In the current review, we systematically describe the recent developments in vaccine against PEDV and the roles of structural proteins, non-structural proteins and accessory proteins of PEDV in affecting viral virulence and regulating innate immunity, which will provide insight into the rational design of effective and safe vaccines for PEDV or other coronaviruses.

Effects of medium chain fatty acids as a mitigation or prevention strategy against porcine epidemic diarrhea virus in swine feed.

Feed has been shown to be a vector for viral transmission. Four experiments were conducted to: 1) determine if medium chain fatty acids (MCFA) are effective mitigants when applied to feed both pre- and post-porcine epidemic diarrhea virus (PEDV) inoculation measured by quantitative reverse transcription polymerase chain reaction (qRT-PCR), 2) evaluate varying levels and combinations of MCFA measured by qRT-PCR, and 3) evaluate selected treatments in bioassay to determine infectivity. In exp. 1, treatments were arranged in a 2 × 2 + 1 factorial with main effects of treatment (0.3% commercial formaldehyde [CF] product, Sal CURB [Kemin Industries, Inc.; Des Moines, IA], or 1% MCFA blend (Blend) of 1:1:1 C6:C8:C10 [PMI, Arden Hills, MN]) and timing of application (pre- or post-inoculation with PEDV) plus a positive control (PC; feed inoculated with PEDV and no treatment). All combinations of treatment and timing decreased detectable PEDV compared with the PC (P < 0.05). Pre-inoculation treatment elicited decreased magnitude of PEDV detection (cycle threshold value) compared with post-inoculation (P = 0.009). Magnitude of PEDV detection was decreased for CF compared with Blend (P < 0.0001). In exp. 2, pre-inoculation treatments consisted of: 1) PC, 2) 0.3% CF, 3 to 5) 0.125% to 0.33% C6:0, 6 to 8) 0.125% to 0.33% C8:0, 9 to 11) 0.125% to 0.33% C10:0, and 12 to 15) 0.125% to 0.66% C5:0. Treating feed with 0.33% C8:0 resulted in decreased (P < 0.05) PEDV detection compared with all other treatments. Increasing concentration of each individual MCFA decreased PEDV detectability (P < 0.042). In exp. 3, pre-inoculation treatments consisted of: 1) PC, 2) 0.3% CF, 3 to 7) 0.25% to 1% Blend, 8 to 10) 0.125% to 0.33% C6:0 + C8:0, 11 to 13) 0.125% to 0.33% C6:0 + C10:0, and 14 to 16) 0.125% to 0.33% C8:0 + C10:0. Treating feed with CF, 0.5% Blend, 0.75% Blend, 1% Blend, all levels of C6:0+C8:0, 0.25% C6:0 + 0.25% C10:0, 0.33% C6:0 + 0.33% C10:0, 0.25% C8:0 + 0.25% C10:0, or 0.33% C8:0 + 0.33% C10:0 elicited decreased detection of PEDV compared with PC (P < 0.05). Increasing concentration of each MCFA combination decreased PEDV detectability (linear, P < 0.012). In exp. 4, feed was treated pre-inoculation with: 1) no treatment (PC), 2) 0.3% CF, 3) 0.5% Blend, or 4) 0.3% C8:0 and analyzed via qRT-PCR and bioassay. Adding 0.5% Blend or 0.3% C8:0 resulted in decreased PEDV compared with PC and only PC resulted in a positive bioassay. Therefore, MCFA can decrease detection of PEDV in feed. Further, inclusion of lower levels of MCFA than previously evaluated are effective against PEDV.

Surface-Displayed Porcine IFN-λ3 in Lactobacillus plantarum Inhibits Porcine Enteric Coronavirus Infection of Porcine Intestinal Epithelial Cells.

Interferon (IFN)-λ plays an essential role in mucosal cells which exhibit strong antiviral activity. Lactobacillus plantarum (L. plantarum) has substantial application potential in the food and medical industries because of its probiotic properties. Alphacoronaviruses, especially porcine epidemic diarrhea virus (PEDV) and transmissible gastroenteritis virus (TGEV), cause high morbidity and mortality in piglets resulting in economic loss. Co-infection by these two viruses is becoming increasingly frequent. Therefore, it is particularly important to develop a new drug to prevent diarrhea infected with mixed viruses in piglets. In this study, we first constructed an anchored expression vector with CWA (C-terminal cell wall anchor) on L. plantarum. Second, we constructed two recombinant L. plantarum strains that anchored IFN-λ3 via pgsA (N-terminal transmembrane anchor) and CWA. Third, we demonstrated that both recombinant strains possess strong antiviral effects against coronavirus infection in the intestinal porcine epithelial cell line J2 (IPEC-J2). However, recombinant L. plantarum with the CWA anchor exhibited a more powerful antiviral effect than recombinant L. plantarum with pgsA. Consistent with this finding, Lb.plantarum-pSIP-409-IFN-λ3-CWA enhanced the expression levels of IFN-stimulated genes (ISGs) (ISG15, OASL, and Mx1) in IPEC-J2 cells more than did recombinant Lb.plantarum-pSIP-409-pgsA'-IFN-λ3. Our study verifies that recombinant L. plantarum inhibits PEDV and TGEV infection in IPEC-J2 cells, which may offer great potential for use as a novel oral antiviral agent in therapeutic applications for combating porcine epidemic diarrhea and transmissible gastroenteritis. This study is the first to show that recombinant L. plantarum suppresses PEDV and TGEV infection of IPEC-J2 cells.