Effects of Antrodia cinnamomea solid culture mycelium by-products on growth performance and immune response in weaning black piglets.

This study evaluated the effects of supplementation with Antrodia cinnamomea mycelium by-product (ACBP) on growth performance and immune response in weaning piglets. Total available content and antioxidant capacity of ACBP were determined. Ninety-six black pigs were randomly distributed to 24 pens. Study compared four groups which were supplemented with ACBP at 0%, 2.5%, 5%, or 10% for 6 weeks after weaning at 4 weeks. Results showed that ACBP on total phenolic, total flavonoid, and total triterpenoids contents were 13.68 mg GAE/g DW, 1.67 µg QE/g DW, and 15.6 mg/g, respectively. Weaning piglets fed 2.5% ACBP showed a significant decreased body weight gain compared with those supplemented with 5% ACBP, 10% ACBP, and control groups. Results showed that all ACBP groups increased the villi height of jejunum significantly. Incidence of diarrhea in 11 weeks with supplementation with 5% and 10% ACBP diets were lower than in control group. The 10% ACBP group showed significantly lower expression of immune response genes (IL-1ß, IL-6, IL-8, TNF-α, and IFN-γ) than the 2.5% and 5% ACBP groups. Based on results, dietary supplementation with 10% ACBP did not significantly affect body weight but could decrease piglet diarrhea condition and expression of IL-1ß and IL-6 genes.

The role of hormones in the regulation of lactogenic immunity in porcine and bovine species.

Colostrum and milk offer a complete diet and vital immune protection for newborn mammals with developing immune systems. High immunoglobulin levels in colostrum serve as the primary antibody source for newborn piglets and calves. Subsequent milk feeding support continued local antibody protection against enteric pathogens, as well as maturation of the developing immune system and provide nutrients for newborn growth. Mammals have evolved hormonal strategies that modulate the levels of immunoglobulins in colostrum and milk to facilitate effective lactational immunity. In addition, hormones regulate the gut-mammary gland-secretory immunoglobulin A (sIgA) axis in pregnant mammals, controlling the levels of sIgA in milk, which serves as the primary source of IgA for piglets and helps them resist pathogens such as PEDV and TGEV. In the present study, we review the existing studies on the interactions between hormones and the gut-mammary-sIgA axis/lactogenic immunity in mammals and explore the potential mechanisms of hormonal regulation that have not been studied in detail, to draw attention to the role of hormones in influencing the immune response of pregnant and lactating mammals and their offspring, and highlight the effect of hormones in regulating sIgA-mediated anti-infection processes in colostrum and milk. Discussion of the relationship between hormones and lactogenic immunity may lead to a better way of improving lactogenic immunity by determining a better injection time and developing new vaccines.

Differential antigenicity of individual Mycoplasma hyorhinis variable lipoproteins.

The Mycoplasma hyorhinis (Mhr) variable lipoprotein (Vlp) family, comprising Vlps A, B, C, D, E, F, and G, are highly variable in expression, size, and cytoadhesion capabilities across Mhr strains. The 'Vlp system' plays a crucial role in cytoadhesion, immune evasion, and in eliciting a host immunologic response. This pilot study described the development of Vlp peptide-based ELISAs to evaluate the antigenic reactivity of individual Vlps against Mhr antisera collected throughout a longitudinal study focused on Mhr strain 38983, reproducing Mhr-associated disease under experimental conditions. Specifically, serum samples were collected at day post-inoculation 0, 7, 10, 14, 17, 21, 24, 28, 35, 42, 49, and 56 from Mhr- and mock (Friis medium)-inoculated cesarean-derived, colostrum-deprived pigs. Significant Mhr-specific IgG responses were detected at specific time points throughout the infection, with some variations for each Vlp. Overall, individual Vlp ELISAs showed consistently high accuracy rates, except for VlpD, which would likely be associated with its expression levels or the anti-Vlp humoral immune response specific to the Mhr strain used in this study. This study provides the basis and tools for a more refined understanding of these Vlp- and Mhr strain-specific variations, which is foundational in understanding the host immune response to Mhr.

Identifying major histocompatibility complex class II-DR molecules in bovine and swine peripheral blood monocyte-derived macrophages using mAb-L243.

Major histocompatibility complex class II (MHC-II) molecules are involved in immune responses against pathogens and vaccine candidates' immunogenicity. Immunopeptidomics for identifying cancer and infection-related antigens and epitopes have benefited from advances in immunopurification methods and mass spectrometry analysis. The mouse anti-MHC-II-DR monoclonal antibody L243 (mAb-L243) has been effective in recognising MHC-II-DR in both human and non-human primates. It has also been shown to cross-react with other animal species, although it has not been tested in livestock. This study used mAb-L243 to identify Staphylococcus aureus and Salmonella enterica serovar Typhimurium peptides binding to cattle and swine macrophage MHC-II-DR molecules using flow cytometry, mass spectrometry and two immunopurification techniques. Antibody cross-reactivity led to identifying expressed MHC-II-DR molecules, together with 10 Staphylococcus aureus peptides in cattle and 13 S. enterica serovar Typhimurium peptides in swine. Such data demonstrates that MHC-II-DR expression and immunocapture approaches using L243 mAb represents a viable strategy for flow cytometry and immunopeptidomics analysis of bovine and swine antigen-presenting cells.

Semen Protein CRISP3 Promotes Reproductive Performance of Boars through Immunomodulation.

Semen proteins play an important role in male reproductive performance and sperm fertilization ability and can be used as potential biomarkers to evaluate male fertility. The role of cysteine-rich secretory protein 3 (CRISP3) in male reproduction remains unknown. This study aimed to investigate the role of CRISP3 in the reproductive performance of boars. Our results showed that the CRISP3 protein content was significantly and positively correlated with boar fertility, sow delivery rate, and litter size. CRISP3 is highly expressed in the bulbourethral gland of adult boars and is enriched in the seminal plasma. It is localized in the post-acrosomal region of the sperm head and migrates to the anterior end of the tail after capacitation. The CRISP3 recombinant protein did not affect sperm motility and cleavage rate, but it significantly downregulated the mRNA expression of inflammatory factors IL-α, IL-1ß, and IL-6 and the protein expression of IL-α and IL-6 in lipopolysaccharide (LPS)-induced RAW264.7 cells, indicating that CRISP3 has an immunomodulatory function. In conclusion, our study suggests that semen CRISP3 protein levels positively correlate with reproductive performance, which may be achieved by regulating immune responses in the female reproductive tract.

Porcine deltacoronavirus nsp5 antagonizes type I interferon signaling by cleaving IFIT3.

Porcine deltacoronavirus (PDCoV) has caused enormous economic losses to the global pig industry. However, the immune escape mechanism of PDCoV remains to be fully clarified. Transcriptomic analysis revealed a high abundance of interferon (IFN)-induced protein with tetratricopeptide repeats 3 (IFIT3) transcripts after PDCoV infection, which initially implied a correlation between IFIT3 and PDCoV. Further studies showed that PDCoV nsp5 could antagonize the host type I interferon signaling pathway by cleaving IFIT3. We demonstrated that PDCoV nsp5 cleaved porcine IFIT3 (pIFIT3) at Gln-406. Similar cleavage of endogenous IFIT3 has also been observed in PDCoV-infected cells. The pIFIT3-Q406A mutant was resistant to nsp5-mediated cleavage and exhibited a greater ability to inhibit PDCoV infection than wild-type pIFIT3. Furthermore, we found that cleavage of IFIT3 is a common characteristic of nsp5 proteins of human coronaviruses, albeit not alphacoronavirus. This finding suggests that the cleavage of IFIT3 is an important mechanism by which PDCoV nsp5 antagonizes IFN signaling. Our study provides new insights into the mechanisms by which PDCoV antagonizes the host innate immune response.IMPORTANCEPorcine deltacoronavirus (PDCoV) is a potential emerging zoonotic pathogen, and studies on the prevalence and pathogenesis of PDCoV are ongoing. The main protease (nsp5) of PDCoV provides an excellent target for antivirals due to its essential and conserved function in the viral replication cycle. Previous studies have revealed that nsp5 of PDCoV antagonizes type I interferon (IFN) production by targeting the interferon-stimulated genes. Here, we provide the first demonstration that nsp5 of PDCoV antagonizes IFN signaling by cleaving IFIT3, which affects the IFN response after PDCoV infection. Our findings reveal that PDCoV nsp5 is an important interferon antagonist and enhance the understanding of immune evasion by deltacoronaviruses.

A recombination-resistant genome for live attenuated and stable PEDV vaccines by engineering the transcriptional regulatory sequences.

IMPORTANCE: Coronaviruses are important pathogens of humans and animals, and vaccine developments against them are imperative. Due to the ability to induce broad and prolonged protective immunity and the convenient administration routes, live attenuated vaccines (LAVs) are promising arms for controlling the deadly coronavirus infections. However, potential recombination events between vaccine and field strains raise a safety concern for LAVs. The porcine epidemic diarrhea virus (PEDV) remodeled TRS (RMT) mutant generated in this study replicated efficiently in both cell culture and in pigs and retained protective immunogenicity against PEDV challenge in pigs. Furthermore, the RMT PEDV was resistant to recombination and genetically stable. Therefore, RMT PEDV can be further optimized as a backbone for the development of safe LAVs.

African swine fever virus B175L inhibits the type I interferon pathway by targeting STING and 2'3'-cGAMP.

IMPORTANCE: African swine fever virus (ASFV), the only known DNA arbovirus, is the causative agent of African swine fever (ASF), an acutely contagious disease in pigs. ASF has recently become a crisis in the pig industry in recent years, but there are no commercially available vaccines. Studying the immune evasion mechanisms of ASFV proteins is important for the understanding the pathogenesis of ASFV and essential information for the development of an effective live-attenuated ASFV vaccines. Here, we identified ASFV B175L, previously uncharacterized proteins that inhibit type I interferon signaling by targeting STING and 2'3'-cGAMP. The conserved B175L-zf-FCS motif specifically interacted with both cGAMP and the R238 and Y240 amino acids of STING. Consequently, this interaction interferes with the interaction of cGAMP and STING, thereby inhibiting downstream signaling of IFN-mediated antiviral responses. This novel mechanism of B175L opens a new avenue as one of the ASFV virulent genes that can contribute to the advancement of ASFV live-attenuated vaccines.

Heterogeneity of Phenotypic and Functional Changes to Porcine Monocyte-Derived Macrophages Triggered by Diverse Polarizing Factors In Vitro.

Swine are attracting increasing attention as a biomedical model, due to many immunological similarities with humans. However, porcine macrophage polarization has not been extensively analyzed. Therefore, we investigated porcine monocyte-derived macrophages (moMΦ) triggered by either IFN-γ + LPS (classical activation) or by diverse "M2-related" polarizing factors: IL-4, IL-10, TGF-ß, and dexamethasone. IFN-γ and LPS polarized moMΦ toward a proinflammatory phenotype, although a significant IL-1Ra response was observed. Exposure to IL-4, IL-10, TGF-ß, and dexamethasone gave rise to four distinct phenotypes, all antithetic to IFN-γ and LPS. Some peculiarities were observed: IL-4 and IL-10 both enhanced expression of IL-18, and none of the "M2-related" stimuli induced IL-10 expression. Exposures to TGF-ß and dexamethasone were characterized by enhanced levels of TGF-ß2, whereas stimulation with dexamethasone, but not TGF-ß2, triggered CD163 upregulation and induction of CCL23. Macrophages stimulated with IL-10, TGF-ß, or dexamethasone presented decreased abilities to release proinflammatory cytokines in response to TLR2 or TLR3 ligands: IL-10 showed a powerful inhibitory activity for CXCL8 and TNF release, whereas TGF-ß provided a strong inhibitory signal for IL-6 production. While our results emphasized porcine macrophage plasticity broadly comparable to human and murine macrophages, they also highlighted some peculiarities in this species.

Development and Characterization of New Monoclonal Antibodies Against Porcine Interleukin-17A and Interferon-Gamma.

Current research efforts require a broad range of immune reagents, but those available for pigs are limited. The goal of this study was to generate priority immune reagents for pigs and pipeline them for marketing. Our efforts were aimed at the expression of soluble swine cytokines and the production of panels of monoclonal antibodies (mAbs) to these proteins. Swine interleukin-17A (IL-17A) and Interferon-gamma (IFNγ) recombinant proteins were produced using yeast expression and used for monoclonal antibody (mAb) production resulting in panels of mAbs. We screened each mAb for cross-species reactivity with orthologs of IL-17A or IFNγ and checked each mAb for inhibition by other related mAbs, to assign mAb antigenic determinants. For porcine IL-17A, the characterization of a panel of 10 mAbs identified eight different antigenic determinants; interestingly, most of the mAbs cross-reacted with the dolphin recombinant ortholog. Likewise, the characterization of a panel of nine anti-PoIFNγ mAbs identified four different determinants; most of the mAbs cross-reacted with dolphin, bovine, and caprine recombinant orthologs. There was a unique reaction of one anti-PoIFNγ mAb that cross-reacted with the zebrafish recombinant ortholog. The αIL-17A mAbs were used to develop a quantitative sandwich ELISA detecting the yeast expressed protein as well as native IL-17A in stimulated peripheral blood mononuclear cell (PBMC) supernatants. Our analyses showed that phorbol myristate acetate/ionomycin stimulation of PBMC induced significant expression of IL-17A by CD3+ T cells as detected by several of our mAbs. These new mAbs expand opportunities for immunology research in swine.

Development of an indirect competitive enzyme linked immunosorbent assay for the quantitative detection of Mycoplasma hyopneumoniae during the vaccine production process.

Inactivated Mycoplasma hyopneumoniae vaccine is used extensively to control M. hyopneumoniae infection worldwide. Quantification techniques are essential in the process of standardizing and validating vaccines. In this study, we developed and optimized an indirect competitive enzyme linked immunosorbent assay (ic-ELISA) for the rapid quantification of M. hyopneumoniae antigen during vaccine production. Briefly, whole M. hyopneumoniae antigen was coated onto microtiter plates, and a polyclonal antibody against M. hyopneumoniae recombinant elongation factor thermo unstable (EF-Tu) protein was prepared and added with the samples to be tested. The methods were optimized and showed significant reproducibility, with coefficients of variation of 4.01% and 6.14% for the intra-and inter-assays, respectively. Quantification of M. hyopneumoniae cultures at different growth stages using the ic-ELISA test showed a similar curve to that of the traditional color changing units (CCU) assay, with a delay in the time when the amount reached the peak and started to fall. In the inactivated vaccine production process, the cultures could be harvested later than that for the live vaccine, at about 12 h after the end of the logarithmic growth phase. Different batches of cultures were measured for their relative potency value compared with the in-house reference vaccine, which was used to determine whether the cultures met the antigen amount requirements for vaccine preparation. The curves of the CCU titer and ic-ELISA titer in the logarithmic phase correlated strongly and a linear regression equation was established to calculate the CCU values rapidly using the ic-ELISA results. In conclusion, an ic-ELISA method was established to rapidly assess the amount of antigen in an M. hyopneumoniae culture during the vaccine production process.

Dietary Chito-oligosaccharides Improve Intestinal Immunity via Regulating Microbiota and Th17/Treg Balance-Related Immune Signaling in Piglets Challenged by Enterotoxigenic E. coli.

This study was conducted to investigate how chito-oligosaccharides (COSs) affect the growth performance and immune stress response and to further explain their mechanisms. A total of 32 boars that were 28 days old and three-way weaned were randomly allotted to four equal groups [CON (basal diet) group, enterotoxigenic Escherichia coli (ETEC) group, COS group, and COS*ETEC group]. The results showed that COS partially reversed the negative changes in the average daily gain and average daily feed intake caused by the ETEC challenge and thereby alleviated the increase in the feed conversion ratio. Dietary COS increased the villus length as compared with the CON group and improved the ileal morphological structure. Additionally, it increased the bacterial diversity and Bacteroidetes abundance and lowered the Firmicutes abundance and Firmicutes-to-Bacteroidetes ratio at the phylum level. COS treatment lowered the abundance of Lactobacillus, Streptococcus, and Anarovovrio in the intestines of piglets, while it increased Muribaculaceae_unclassified and Prevotella at the genus level. COS had a significant inhibitory effect on the increase in the relative expression abundance of STAT3 mRNA caused by ETEC. The IL-10 and FOXP3 mRNAs were found to be significantly lower in the COS, ETEC, and COS*ETEC groups as compared to the CON group. These results demonstrate that COS could be beneficial for improving the growth performance and attenuating ETEC-challenged intestinal inflammation via regulating microbiota and Th17/Treg balance-related immune signaling pathways.

Fruit and Vegetable Supplemented Diet Modulates the Pig Transcriptome and Microbiome after a Two-Week Feeding Intervention.

A study was conducted to determine the effects of a diet supplemented with fruits and vegetables (FV) on the host whole blood cell (WBC) transcriptome and the composition and function of the intestinal microbiome. Nine six-week-old pigs were fed a pig grower diet alone or supplemented with lyophilized FV equivalent to half the daily recommended amount prescribed for humans by the Dietary Guideline for Americans (DGA) for two weeks. Host transcriptome changes in the WBC were evaluated by RNA sequencing. Isolated DNA from the fecal microbiome was used for 16S rDNA taxonomic analysis and prediction of metabolomic function. Feeding an FV-supplemented diet to pigs induced differential expression of several genes associated with an increase in B-cell development and differentiation and the regulation of cellular movement, inflammatory response, and cell-to-cell signaling. Linear discriminant analysis effect size (LEfSe) in fecal microbiome samples showed differential increases in genera from Lachnospiraceae and Ruminococcaceae families within the order Clostridiales and Erysipelotrichaceae family with a predicted reduction in rgpE-glucosyltransferase protein associated with lipopolysaccharide biosynthesis in pigs fed the FV-supplemented diet. These results suggest that feeding an FV-supplemented diet for two weeks modulated markers of cellular inflammatory and immune function in the WBC transcriptome and the composition of the intestinal microbiome by increasing the abundance of bacterial taxa that have been associated with improved intestinal health.

Fc-Mediated E2-Dimer Subunit Vaccines of Atypical Porcine Pestivirus Induce Efficient Humoral and Cellular Immune Responses in Piglets.

Congenital tremor (CT) type A-II in piglets is caused by an emerging atypical porcine pestivirus (APPV), which is prevalent in swine herds and a serious threat to the pig production industry. This study aimed to construct APPV E2 subunit vaccines fused with Fc fragments and evaluate their immunogenicity in piglets. Here, APPV E2Fc and E2ΔFc fusion proteins expressed in Drosophila Schneider 2 (S2) cells were demonstrated to form stable dimers in SDS-PAGE and western blotting assays. Functional analysis revealed that aE2Fc and aE2ΔFc fusion proteins could bind to FcγRI on antigen-presenting cells (APCs), with the affinity of aE2Fc to FcγRI being higher than that of aE2ΔFc. Moreover, subunit vaccines based on aE2, aE2Fc, and aE2ΔFc fusion proteins were prepared, and their immunogenicity was evaluated in piglets. The results showed that the Fc fusion proteins emulsified with the ISA 201VG adjuvant elicited stronger humoral and cellular immune responses than the IMS 1313VG adjuvant. These findings suggest that APPV E2 subunit vaccines fused with Fc fragments may be a promising vaccine candidate against APPV.

Antibody-Mediated Targeting of Antigens to Intestinal Aminopeptidase N Elicits Gut IgA Responses in Pigs.

Many pathogens enter the host via the gut, causing disease in animals and humans. A robust intestinal immune response is necessary to protect the host from these gut pathogens. Despite being best suited for eliciting intestinal immunity, oral vaccination remains a challenge due to the gastrointestinal environment, a poor uptake of vaccine antigens by the intestinal epithelium and the tolerogenic environment pervading the gut. To improve uptake, efforts have focused on targeting antigens towards the gut mucosa. An interesting target is aminopeptidase N (APN), a conserved membrane protein present on small intestinal epithelial cells shown to mediate epithelial transcytosis. Here, we aimed to further optimize this oral vaccination strategy in a large animal model. Porcine APN-specific monoclonal antibodies were generated and the most promising candidate in terms of epithelial transcytosis was selected to generate antibody fusion constructs, comprising a murine IgG1 or porcine IgA backbone and a low immunogenic antigen: the F18-fimbriated E. coli tip adhesin FedF. Upon oral delivery of these recombinant antibodies in piglets, both mucosal and systemic immune responses were elicited. The presence of the FedF antigen however appeared to reduce these immune responses. Further analysis showed that F18 fimbriae were able to disrupt the antigen presenting capacity of intestinal antigen presenting cells, implying potential tolerogenic effects of FedF. Altogether, these findings show that targeted delivery of molecules to epithelial aminopeptidase N results in their transcytosis and delivery to the gut immune systems. The results provide a solid foundation for the development of oral subunit vaccines to protect against gut pathogens.

Human-like Response of Pig T Cells to Superagonistic Anti-CD28 Monoclonal Antibodies.

Because of its size, anatomical similarities, and now also accessibility to genetic manipulations, pigs are used as animal models for human diseases and immune system development. However, expression and function of CD28, the most important costimulatory receptor expressed by T cells, so far is poorly understood in this species. Using a newly generated mAb (mAb 3D11) with specificity for pig CD28, we detected CD28 on CD8+ and CD4+ αß T cells. Among γδ T cells, CD28 expression was restricted to a small CD2+ subpopulation of phenotypically naive cells. Functionally, CD28 ligation with mAb 3D11-costimulated porcine T cells, enhanced proliferation and cytokine secretion in vitro. We used a second, likewise newly generated but superagonistic, anti-CD28 mAb (CD28-SA; mAb 4D12) to test the function of CD28 on porcine T cells in a pilot study in vivo. Injection of the CD28-SA into pigs in vivo showed a very similar dose-response relationship as in humans (i.e., 100 µg/kg body weight [BW]) of CD28-SA induced a cytokine release syndrome that was avoided at a dose of 10 µg/kg BW and below. The data further suggest that low-dose (10 µg/kg BW) CD28-SA infusion was sufficient to increase the proportion of Foxp3+ regulatory T cells among CD4+ T cells in vivo. The pig is thus a suitable animal model for testing novel immunotherapeutics. Moreover, data from our pilot study in pigs further suggest that low-dose CD28-SA infusion might allow for selective expansion of CD4+ regulatory T cells in humans.

Development of a Dendrimeric Peptide-Based Approach for the Differentiation of Animals Vaccinated with FlagT4G against Classical Swine Fever from Infected Pigs.

Classical swine fever virus (CSFV) causes a viral disease of high epidemiological and economical significance that affects domestic and wild swine. Control of the disease in endemic countries is based on live-attenuated vaccines (LAVs) that induce an early protective immune response against highly virulent CSFV strains. The main disadvantage of these currently available LAVs is the lack of serological techniques to differentiate between vaccinated and infected animals (DIVA concept). Here, we describe the development of the FlagDIVA test, a serological diagnostic tool allowing for the differentiation between animals vaccinated with the FlagT4G candidate and those infected with CSFV field strains. The FlagDIVA test is a direct ELISA based on a dendrimeric peptide construct displaying a conserved epitope of CSFV structural protein E2. Although FlagDIVA detected anti-CSFV anti-bodies in infected animals, it did not recognize the antibody response of FlagT4G-vaccinated animals. Therefore, the FlagDIVA test constitutes a valuable accessory DIVA tool in implementing vaccination with the FlagT4G candidate.

Composition and physiological functions of the porcine colostrum.

The first secretion, 24-h post parturition of the mammary glands of sows, known as colostrum, is high in protein and low in lactose and fat. As a consequence of an insufficient ingestion of colostrum, more than 50% of piglets fail to reach weaning and die. The composition and some functions of colostrum have been previously reported. For example, colostrum carbohydrates consist of mainly lactose. Lipids in the colostrum are mostly triacylglycerols, but <1% is fatty acids, which may act as homeostasis regulators. Similarly, proteins are found mostly as casein and whey, the latter being ≥80% immunoglobulins. Colostrum-derived immunoglobulins and bioactive proteins such as azurocidin help the immune system of the piglet fend off infections. In addition, leukocytes and exosomes are other minor but nonetheless equally crucial bioactive components in the porcine colostrum. Modern pig farming has achieved increases in pig productivity and litter size, but this has been accomplished in detriment of the health and the survival rate of piglets. Therefore, porcine colostrum is now even more important in pig farming. In the present review, we discuss the current knowledge on the composition and physiological functions of the porcine colostrum and briefly propose future research directions.

Exosome-Based Vaccines: Pros and Cons in the World of Animal Health.

Due to the emergence of antibiotic resistance and new and more complex diseases that affect livestock animal health and food security, the control of epidemics has become a top priority worldwide. Vaccination represents the most important and cost-effective measure to control infectious diseases in animal health, but it represents only 23% of the total global animal health market, highlighting the need to develop new vaccines. A recent strategy in animal health vaccination is the use of extracellular vesicles (EVs), lipid bilayer nanovesicles produced by almost all living cells, including both prokaryotes and eukaryotes. EVs have been evaluated as a prominent source of viral antigens to elicit specific immune responses and to develop new vaccination platforms as viruses and EVs share biogenesis pathways. Preliminary trials with lymphocytic choriomeningitis virus infection (LCMV), porcine reproductive and respiratory syndrome virus (PRRSV), and Marek's disease virus (MDV) have demonstrated that EVs have a role in the activation of cellular and antibody immune responses. Moreover, in parasitic diseases such as Eimeria (chickens) and Plasmodium yoelii (mice) protection has been achieved. Research into EVs is therefore opening an opportunity for new strategies to overcome old problems affecting food security, animal health, and emerging diseases. Here, we review different conventional approaches for vaccine design and compare them with examples of EV-based vaccines that have already been tested in relation to animal health.

Isolation and characterization of porcine monoclonal antibodies revealed two distinct serotype-independent epitopes on VP2 of foot-and-mouth disease virus.

Pigs are susceptible to foot-and-mouth disease virus (FMDV), and the humoral immune response plays an essential role in protection against FMDV infection. However, little information is available about FMDV-specific mAbs derived from single B cells of pigs. This study aimed to determine the antigenic features of FMDV that are recognized by antibodies from pigs. Therefore, a panel of pig-derived mAbs against FMDV were developed using fluorescence-based single B cell antibody technology. Western blotting revealed that three of the antibodies (1C6, P2-7E and P2-8G) recognized conserved antigen epitopes on capsid protein VP2, and exhibited broad reactivity against both FMDV serotypes A and O. An alanine-substitution scanning assay and sequence conservation analysis elucidated that these porcine mAbs recognized two conserved epitopes on VP2: a linear epitope (2KKTEETTLL10) in the N terminus and a conformational epitope involving residues K63, H65, L66, F67, D68 and L81 on two ß-sheets (B-sheet and C-sheet) that depended on the integrity of VP2. Random parings of heavy and light chains of the IgGs confirmed that the heavy chain is predominantly involved in binding to antigen. The light chain of porcine IgG contributes to the binding affinity toward an antigen and may function as a support platform for antibody stability. In summary, this study is the first to reveal the conserved antigenic profile of FMDV recognized by porcine B cells and provides a novel method for analysing the antibody response against FMDV in its natural hosts (i.e. pigs) at the clonal level.