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Tumour necrosis factor (TNF) superfamily member 11 (TNFSF11), also known as RANKL, plays a crucial role in regulating several physiological and pathological activities. Additionally, it is a vital factor in bone physiology, and the sex hormone progesterone regulates the expansion of stem cells and the proliferation of mammary epithelial cells. It is essential for animal growth and reproductive physiological processes. This study aimed to evaluate the tissue-specific expression characteristics and promoter activity of the TNFSF11 gene in pigs. As a result, the study examined the presence of TNFSF11 expression in the tissues of Xiangsu pigs at 0.6 and 12 months of age. Moreover, the core promoter region of TNFSF11 was also identified by utilizing a combination of bioinformatic prediction and dual-luciferase activity tests. Finally, the effect of transcription factors on the transcriptional activity of the core promoter region was determined using site-directed mutagenesis. TNFSF11 was uniformly expressed in all tissues; however, its expression in muscles was comparatively low. The core promoter region of TNFSF11 was located in the −555 to −1 region. The prediction of the transcription start site of TNFSF11 gene-2000 ∼ + 500bp showed that there was a CpG site in 17 ∼ + 487bp. Analysis of mutations in the transcription factor binding sites revealed that mutations in the Stat5b , Myog , Trl , and EN1 binding sites had significant effects on the transcriptional activity of the TNFSF11 gene, particularly following the EN1 binding site mutation (P < 0.001). This study provides insights into both the tissue-specific expression patterns of TNFSF11 in the tissues of Xiangsu pigs and the potential regulatory effects of transcription factors on its promoter activity. These results may be helpful for future research aimed at clarifying the expression and role of the porcine TNFSF11 gene. [ABSTRACT FROM AUTHOR]

In vitro embryo production (IVP) is of great importance to the porcine industry, as well as for basic research and biomedical applications. Despite the large efforts made in laboratories worldwide to address suboptimal culture conditions, porcine IVP remains inefficient. Nobiletin (Nob, 5,6,7,8,3′,4′ hexamethoxyflavone) supplementation to in vitro culture (IVC) medium, enhances in vitro embryo development in various species. However, its impact on the quality and developmental capacity of in vitro-produced pig embryos is yet to be established. This study evaluated the effects of different concentrations (2.5 and 5 μM) of Nob during the early culture of in vitro -produced pig embryos on embryo developmental competence, mitochondrial activity, lipid content, intracellular Reactive Oxygen Species (ROS) and Glutathione (GSH) content, Total Cell Number (TCN) per blastocyst, and expression of genes related to embryo development, quality and oxidative stress. Embryos cultured in medium without Nob supplementation and in medium supplemented with 0.01 % dimethyl sulfoxide (DMSO-vehicle for Nob) constituted the Control and DMSO groups, respectively. Embryo development rates were evaluated on Days 2, 6 and 7 of IVC. Additionally, a representative group of embryos was selected to assess mitochondrial activity, lipid, ROS and GSH content (on Days 2 and 6 of IVC), TCN assessment and gene expression analyses (on Day 6 of IVC). No significant differences were observed in any of the parameters evaluated on Day 2 of IVC. In contrast, embryos cultured under the presence of Nob 2.5 showed higher developmental rates on Days 6 and 7 of IVC. In addition, Day 6 embryos showed increased mitochondrial activity, with decreased levels of ROS and GSH in the Nob 2.5 group compared to the other groups. Both Nob 2.5 and Nob 5 embryos showed higher TCN compared to the Control and DMSO groups. Furthermore, Nob 2.5 and Nob 5 upregulated the expression of Superoxide dismutase type 1 (SOD1) and Glucose-6-phosphate dehydrogenase (G6PDH) genes, which could help to counteract oxidative stress during IVC. In conclusion, the addition of Nob during the first 48 h of IVC increased porcine embryo development rates and enhanced their quality, including the upregulation of relevant genes that potentially improved the overall efficiency of the IVP system. • In vitro embryo production is of great importance to the porcine industry and biomedical applications. • The efficiency of porcine in vitro embryo production falls below optimal levels. • Nobiletin supplementation in vitro increased porcine embryo development and quality. • Nobiletin is a potential solution for addressing ROS-related issues in porcine in vitro embryo production. [ABSTRACT FROM AUTHOR]

The present Porcine circovirus type 2 virus (PCV 2) vaccine adjuvants suffer from numerous limitations, such as adverse effects, deficient cell-mediated immune responses, and inadequate antibody production. In this study, we explored the potential of a novel nanoparticle (CS-Au NPs) based on gold nanoparticles (Au NPs) and chitosan (CS) that modified Viola philippica polysaccharide (VPP) as efficient adjuvants for PCV 2 vaccine. The characterization demonstrated that CS-Au-VPP NPs had a mean particle size of 507.42 nm and a zeta potential value of −21.93 mV. CS-Au-VPP NPs also exhibited good dispersion and a stable structure, which did not alter the polysaccharide properties. Additionally, the CS-Au-VPP NPs showed easy absorption and utilization by the organism. To investigate their immune-enhancing potential, mice were immunized with a mixture of CS-Au-VPP NPs and PCV 2 vaccine. The evaluation of relevant immunological indicators, including specific IgG antibodies and their subclasses, cytokines, and T cell subpopulations, confirmed their immune-boosting effects. The in vivo experiments revealed that the medium-dose CS-Au-VPP NPs significantly elevated the levels of specific IgG antibodies and their subclasses, cytokines, and T cell subpopulations in PCV 2 -immunized mice. These findings suggest that CS-Au-VPP NPs can serve as a promising vaccine adjuvant due to their stable structure and immunoenhancement capabilities. [ABSTRACT FROM AUTHOR]

Porcine reproductive and respiratory syndrome virus (PRRSV), a highly infectious virus, causes severe losses in the swine industry by regulating the inflammatory response, inducing tissue damage, suppressing the innate immune response, and promoting persistent infection in hosts. Interleukin-13 (IL-13) is a cytokine that plays a critical role in regulating immune responses and inflammation, particularly in immune-related disorders, certain types of cancer, and numerous bacterial and viral infections; however, the underlying mechanisms of IL-13 regulation during PRRSV infection are not well understood. In this study, we demonstrated that PRRSV infection elevates IL-13 levels in porcine alveolar macrophages. PRRSV enhances m6A-methylated RNA levels while reducing the expression of fat mass and obesity associated protein (FTO, an m6A demethylase), thereby augmenting IL-13 production. PRRSV nonstructural protein 9 (nsp9) was a key factor for this modulation. Furthermore, we found that the residues Asp567, Tyr586, Leu593, and Asp595 were essential for nsp9 to induce IL-13 production via attenuation of FTO expression. These insights delineate PRRSV nsp9's role in FTO-mediated IL-13 release, advancing our understanding of PRRSV's impact on host immune and inflammatory responses. [ABSTRACT FROM AUTHOR]

Edwardsiellosis is one of the most important bacterial diseases in fish, sometimes causing extensive economic losses in the aquaculture industry. Our previous studies demonstrated that the Cu,Zn-SOD (sod1) activity has significantly increased in Japanese flounder, Paralichthys olivaceus , hepatopancreas infected by causative bacteria of edwardsiellosis Edwardsiella tarda NUF251. In this study, NUF251 stimulated intracellular superoxide radical production in mouse macrophage RAW264.7 cells, which was reduced by N -acetylcysteine. This result suggests that NUF251 infection causes oxidative stress. To evaluate the regulatory mechanism of Jf sod1 at transcriptional levels under oxidative stress induced by NUF251 infection, we cloned and determined the nucleotide sequence (1124 bp) of the 5′-flanking region of the Jf sod1 gene. The sequence analysis demonstrated that the binding sites for the transcription factors C/EBPα and NF-IL6 involved in the transcriptional regulation of the mammalian sod1 gene existed. We constructed a luciferase reporter system with the 5′-flanking region (−1124/−1) of the Jf sod1 gene, and a highly increased transcriptional activity of the region was observed in NUF251 - infected RAW264.7 cells. Further studies using several mutants indicated that deletion of the recognition region of NF-IL6 (−272/−132) resulted in a significant decrease in the transcriptional activity of the Jf sod1 gene in NUF251-infected RAW264.7 cells. In particular, the binding site (−202/−194) for NF-IL6 might play a major role in upregulating the transcriptional activity of the 5′-flanking region of the Jf sod1 gene in response to oxidative stress induced by NUF251 infection. These results could be provided a new insight to understand the pathogenic mechanism of causative bacteria of edwardsiellosis. [Display omitted] • NUF251 infection caused oxidative stress in RAW264.7 cells. • 5′-flanking region of Jf sod1 gene was firstly sequenced and found some cis -elements. • Causative bacteria of edwardsiellosis infection upregulated transcription of Jf sod1. • NF-IL6 might play a role in activation of Jf sod1 transcription by NUF251 infection. [ABSTRACT FROM AUTHOR]

Granulocyte colony-stimulating factor (G-CSF), a pleiotropic cytokine, is secreted by the reproductive tract. Furthermore, our previous study indicated that human recombinant G-CSF (hrG-CSF) supplementation during porcine oocyte in vitro maturation (IVM) or during embryo in vitro culture (IVC) improved their quality and development potential when using cumulus-oocyte complexes (COCs) with more than three cumulus cell layers (CCL >3). Thus, in this study, we investigate the optimal conditions of hrG-CSF supplementation throughout the in vitro production (IVP: IVM + IVC) system to improve the embryo production efficiency of "poor-quality (CCL ≤3)" oocytes. COCs were classified into two groups according to the number of CCL (>3 and ≤3) and embryonic viability was analyzed after treatment with hrG-CSF during IVC. The mRNA transcription levels of G-CSF in COCs were compared based on their type and the period of IVM. Finally, developmental capacity and quality were evaluated after treatment with hrG-CSF for different periods of IVP. No marked effects on the developmental potential of embryos when using CCL ≤3 type COCs were observed after supplementing hrG-CSF only during IVC. Moreover, the mRNA transcription level of G-CSF increased gradually with IVM culture time and was higher in CCL ≤3 COCs than in >3. Supplementing hrG-CSF only during the IVM period resulted in the best embryo developmental potential, while supplementing hrG-CSF during the IVP period resulted in the best quality embryos, reflected in the increased total cell number and decreased apoptotic nuclei index of blastocysts. These findings indicate that "poor-quality" COCs may have a greater demand for G-CSF than "good-quality", meanwhile hrG-CSF supplementation throughout IVP improves resource utilization efficiency in poor-quality COCs. • The hrG-CSF treatment during IVC showed different embryonic development efficiency depending on the quality of oocytes. • The expression level of G-CSF increased gradually with the culture time of IVM and was relatively high in CCL ≤ 3 COCs. • CCL ≤ 3-derived embryos showed the best development efficiency after IVM period treatment while the best quality after IVP. [ABSTRACT FROM AUTHOR]

• Trimeric S, S1, COEs, and two adjuvants were selected to evaluate immune effects. • S/M103 elicited neutralizing antibodies, T cells, B cells, and NK cells in mice. • S/M103 induced high levels of neutralizing antibodies, IFN-γ, and IL-4 in piglets. • S/M103 provided protection against viral challenge in piglets. Porcine epidemic diarrhea virus (PEDV) has caused serious economic losses to the pig husbandry worldwide, and the effects of existing commercialized vaccines are suboptimal. Therefore, research to develop an efficacious vaccine for prevention and control of PEDV is essential. In this study, we designed and produced trimerized proteins of full-length PEDV spike (S) protein, S1 subunit, and a tandem of multiple epitopes of S protein using an efficient mammalian expression vector system in HEK 293F cells. The immunogenicity of two commercial adjuvants, M401 and M103, was also evaluated in mice. Enzyme-linked immunosorbent assays demonstrated that all immunized mice generated highly systemic PEDV S-specific IgG and IgA antibodies. Mice in S/M103-immunized group generated the highest neutralizing antibody titer with 1:96. Compared with control group, the subunit vaccines elicited multifunctional CD3+CD4+ and CD3+CD8+ T cells, B220+CD19+ B cells, and CD3-CD49b+ natural killer cells in the spleen. PEDV S/M103 vaccine, which had the best immune effect, was selected for further evaluation in piglets. Immunization with S/M103 vaccine induced high levels of S-specific IgG, IgA, and neutralizing antibodies, and increased the proliferation of peripheral blood mononuclear cells and the expression levels of interferon-γ and interleukin-4 in peripheral blood of piglets. Virus challenge test results showed significantly lower diarrheal index scores and fecal viral loads, and less pathological damage to the intestines in S/M103-immunized piglets than in controls, indicating that S/M103 provides good protection against the virulent virus challenge. Our findings suggest that trimeric PEDV S/M103 has potential as a clinical vaccine candidate. [ABSTRACT FROM AUTHOR]

A renewable source of porcine macrophages derived from pluripotent stem cells (PSCs) would be a valuable alternative to primary porcine alveolar macrophages (PAMs) in the research of host-pathogen interaction mechanisms. We developed an efficient and rapid protocol, within 11 days, to derive macrophages from porcine PSCs (pPSCs). The pPSC-derived macrophages (pPSCdMs) exhibited molecular and functional characteristics of primary macrophages. The pPSCdMs showed macrophage-specific surface protein expression and macrophage-specific transcription factors, similar to PAMs. The pPSCdMs also exhibited the functional characteristics of macrophages, such as endocytosis, phagocytosis, porcine respiratory and reproductive syndrome virus infection and the response to lipopolysaccharide stimulation. Furthermore, we performed transcriptome sequencing of the whole differentiation process to track the fate transitions of porcine PSCs involved in the signaling pathway. The activation of transforming growth factor beta signaling was required for the formation of mesoderm and the inhibition of the transforming growth factor beta signaling pathway at the hematopoietic endothelium stage could enhance the fate transformation of hematopoiesis. In summary, we developed an efficient and rapid protocol to generate pPSCdMs that showed aspects of functional maturity comparable with PAMs. pPSCdMs could provide a broad prospect for the platforms of host-pathogen interaction mechanisms. [ABSTRACT FROM AUTHOR]

Early cleavage (EC) influences the development of the pre-implantation and post-implantation embryo. Symmetric cleavage (Sym) and asymmetric cleavage (Asy) have been observed in EC, but its molecular mechanism remains unclear. This study was designed to pick out the key candidate genes and signaling pathway between Sym and Asy embryos by applying Smart-seq2 technique. In in-vitro fertilization (IVF) 2-cell embryos, Sym embryos and Asy embryos accounted for 62.55% and 37.45%, respectively. The 2-cell rate, blastocyst rate and total blastocyst cells of Sym group were significantly higher than those of Asy group (31.38% vs 18.79%, 47.55% vs 29.5%, 71.33 vs 33.67, P < 0.05). The 2-cell rate, blastocyst rate and total blastocyst cell number in parthenogenetic activation (PA) embryos in Sym group were significantly higher than those in Asy group (40.61% vs 23.64%, 63.15% vs 30.11%, 50.75 vs 40.5, P < 0.05). A total of 216 differentially expressed genes (DEGs) incorporating 147 genes up-regulated and 69 genes down-regulated genes were screened under the p-value <0.05 and |log2 (fold change)| ≥ 1 when compared with Sym group. Further Gene Ontology (GO) analysis showed that these DEGs were related to the regulation of metabolic process, cell cycle, chromosome segregation, centromeric region and microtubule cytoskeleton. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that the DEGs were mainly enriched to oocyte meiosis, cell cycle, p53 and Hippo signaling pathways. We concluded that asymmetric cleavage is a consequence of altered gene expression. Atg4c, Sesn2, Stk11ip, Slc25a6, Cep19 and Cep55 associated with mitochondrial function and cytoskeletal structure were probably the key candidate genesto determine the zygote cleavage pattern. • Symmetric cleavage (Sym) and asymmetric cleavage (Asy) embryos had different development capacity, and Asy is detrimental to the development of porcine embryos. • 216 differentially expressed genes (DEGs) were found between Sym and Asy embryos, which were mainly associated with the regulation of metabolic process, cell cycle, chromosome segregation, centromeric region and microtubule cytoskeleton. • Genes related to mitochondrial function and cytoskeletal structure probably were the candidate genes to determine the zygote cleavage patterns. [ABSTRACT FROM AUTHOR]

• The protective efficacy of the inactivated PRRS vaccine candidate was assessed against a homologous virus challenge in pigs at short and long durations post-booster. • An earlier onset of PRRSV-specific virus neutralizing antibodies and cell-mediated immune responses was observed in immunized pigs compared to unimmunized controls. • Piglets that received booster immunizations six months prior showed stronger PRRSV-specific immune responses when challenged, in comparison to piglets from the short-term interval group Porcine Reproductive and Respiratory Syndrome (PRRS) is an important viral disease of swine that causes significant mortality in piglets and production losses in adult pigs. In this study, we investigated the protective efficacy of an inactivated PRRS virus vaccine candidate and evaluated the differences in PRRSV specific anamnestic response in piglets when challenged with live PRRSV at two different intervals post-immunization. Six-week-old piglets were immunized intramuscularly with an inactivated, Montanide ISA-206 adjuvanted Indian PRRSV isolate, followed by a booster dose at 21 days post-immunization. Homologous live PRRS virus challenge was done on 60 and 180 days post-booster (dpb). We assessed humoral and cell-mediated immune responses at various intervals post-immunization and after challenge. Viraemia, virus shedding in nasal secretions and lung lesion scores were studied to assess the efficacy of the vaccine candidate. All the immunized pigs developed PRRSV-specific antibodies upon booster dose administration. Neutralizing antibody (NA) titres before challenge, in most animals, ranged between 0 and 4. Potentially protective NA titre of 8 was observed in serum of seven out of the 12 immunized piglets after challenge, across the immunized groups. A significant increase in the mean T-helper, T-cytotoxic, memory or activated T-helper and NK cell populations was observed in immunized piglets challenged at 180 dpb, from 4 to 11 dpc, 5 to 11 dpc, 5 to 7 dpc and 6 to 11 dpc, respectively as compared to the challenge controls. Protective efficacy of the inactivated PRRSV antigen against the homologous virus challenge was evidenced by earlier onset of PRRSV specific virus neutralizing antibodies and cell mediated immune responses, reduced viremia, nasal virus shedding and severity of lung lesions in immunized piglets as compared to unimmunized controls post-challenge. Our results indicated that the inactivated PRRSV antigen elicited better virus specific anamnestic immune responses in piglets when challenged at six months after the single booster dose, due to age related increase in antigen-specific memory T helper cell responses, as compared to those challenged at 2 months post booster. [ABSTRACT FROM AUTHOR]

Porcine epidemic diarrhea virus (PEDV) is a main cause of severe enteric disease in piglets, leading to millions of dollars lost annually in the global pig industry. Parenteral vaccination is limited in generating sufficient mucosal immunity, which is crucial for early defense against PEDV. Here, we orally administered ginseng stem-leaf saponins (GSLS) to mice before parenteral vaccination and found that GSLS significantly enhanced the phagocytosis of dendritic cells, promoted the activities of CD4+ T cells and increased PEDV-specific IgA antibodies in the intestinal mucosa. Transcriptomic results showed that the altered genes following GSLS treatment were mostly related to the immune response and metabolism. In addition, integrated analysis of the transcriptome and metabolome revealed that the mechanism by which GSLS enhances mucosal immunity may be associated with progesterone-related pathways. Further studies are needed to explore the detailed molecular mechanisms. [ABSTRACT FROM AUTHOR]

Extracellular vesicles (EVs) are lipid bilayer nanovesicles generated by almost all living cells which possess various size ranges depending on producer cells and biogenesis mechanisms. Several EV markers were determined including tetraspanins (e.g., CD9, CD63 and CD81), heat shock proteins (HSP70 and HSP90), some 14–3–3 proteins (a family of conserved regulatory molecules), major histocompatibility complex molecules (MHC-I/-II), and enzymes (Glyceraldehyde 3-phosphate dehydrogenase and enolase-1). EVs are known as an abundant source of antigens and immune molecules that can be used for vaccine development in human and animals. EV-based immunization could significantly activate immune responses in different infections such as Porcine reproductive and respiratory syndrome virus (PRRSV), Lymphocytic choriomeningitis virus (LCMV), Marek's disease virus (MDV), and SARS-CoV-2 infections. The engineered and modified EVs showed a promising potential in development of anti-tumor vaccines and therapeutics, protection against parasitic diseases (e.g., Eimeria, and Plasmodium yoelii) and viral diseases (e.g., COVID-19), and improvement of biomarkers. Also, EVs possess a crucial role in antigen presentation in vivo. In this review, we describe the roles of EVs in vaccine development and therapeutic approaches for viral diseases. [Display omitted] • Extracellular vesicles (EVs) provide abundant source of antigens and immune molecules for vaccine development. • Encapsulating cargo into EVs is performed through cell- and non-cell-based loading methods. • EVs harboring biomolecules contribute to cellular communication and tissue repair. • Antiviral drug-loaded exosomes combat respiratory viruses through various strategies. • Safety of different cargos-loaded EVs should be determined for any clinical intervention. [ABSTRACT FROM AUTHOR]

• Immunoinformatics tools enable rapid and accurate analysis of microbial better epitopes, and accelerate vaccine design. • rPMT is a multi-epitope recombinant toxin (PMT) antigen of P. multocida. • Vaccination of rPMT induced humoral and Th1-type cellular immune response in a mouse model. • rPMT confers 56% protection against P. multocida virulent strain. Pasteurella multocida (P. multocida) infection frequently results in porcine atrophic rhinitis and swine plague, leading to large economic losses for the swine industry worldwide. P. multocida toxin (PMT, 146 kDa) is a highly virulent key virulence factor that plays a vital role in causing lung and turbinate lesions. This study developed a multi-epitope recombinant antigen of PMT (rPMT) that showed excellent immunogenicity and protection in a mouse model. Using bioinformatics to analyse the dominant epitopes of PMT, we constructed and synthesized rPMT containing 10 B-cell epitopes, 8 peptides with multiple B-cell epitopes and 13 T-cell epitopes of PMT and a rpmt gene (1,974 bp) with multiple epitopes. The rPMT protein (97 kDa) was soluble and contained a GST tag protein. Immunization of mice with rPMT stimulated significantly elevated serum IgG titres and splenocyte proliferation, and serum IFN-γ and IL-12 were upregulated by 5-fold and 1.6-fold, respectively, but IL-4 was not. Furthermore, the rPMT immunization group exhibited alleviated lung tissue lesions and a significantly decreased degree of neutrophil infiltration compared with the control groups post-challenge. In the rPMT vaccination group, 57.1% (8/14) of the mice survived the challenge, similar to the bacterin HN06 group, while all the mice in the control groups succumbed to the challenge. Thus, rPMT could be a suitable candidate antigen for developing a subunit vaccine against toxigenic P. multocida infection. [ABSTRACT FROM AUTHOR]

Mixed infections of multiple viruses significantly contribute to the prevalence of swine diseases, adversely affecting global livestock production and the economy. However, effectively monitoring multiple viruses and detecting mixed infection samples remains challenging. This study describes a method that combines single-base extension PCR with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) to detect important porcine viruses. Our approach accurately and simultaneously identified 14 porcine viruses, including porcine circovirus types 1–3, porcine bocaviruses groups 1–3, African swine fever virus, pseudorabies virus, porcine parvovirus, torque teno sus virus, swine influenza virus, porcine reproductive and respiratory syndrome virus, classical swine fever virus, and foot-and-mouth disease virus. The low limit of detection for multiplex identification ranges from 13.54 to 1.59 copies/μL. Inter- and intra-assay stability was found to be ≥98.3 %. In a comprehensive analysis of 114 samples, the assay exhibited overall agreement with qPCR results of 97.9 %. The developed MALDI-TOF NAMS assay exhibits high sensitivity, specificity, and reliability in detecting and distinguishing a wide spectrum of porcine viruses in complex matrix samples. This underscores its potential as an efficient diagnostic tool for porcine-derived virus surveillance and swine disease control. • Developed MALDI-TOF NAMS detects 14 porcine viruses simultaneously. • The method distinguishes PCV1 and PCV2 via SNP site. • Applicable for monitoring pathogens in clinical samples, feed, and pork. [ABSTRACT FROM AUTHOR]

• PCV2 ORF5 promotes inflammatory responses by upregulating miR-21. • Nuclear miR-30d inhibits miR-21 by targeting pri-miR-21 to suppress the inflammation. • PCV2 ORF5 inhibits miR-30d levels by direct binding rather than the circRNA pathway. Porcine circovirus type 2 (PCV2) infection leads to multi-system inflammation in pigs, and this effect can be achieved by upregulating host miR-21. The underlying mechanism of miR-21 regulates PCV2-induced inflammation is already known, however, how PCV2 regulates miR-21 levels and function using both autonomic and host factors remains to be further revealed. Here we present the first evidence that PCV2 ORF5 induces an inflammatory response by up-regulating miR-21 level through targeting nuclear miR-30d. In this study, we found that overexpression of ORF5 significantly increased miR-21 level and promoted the expression of inflammatory cytokines and activation of the NF-κB pathway, while ORF5 mutation had the opposite effect. Moreover, the differential expression of miR-21 could significantly change the pro-inflammatory effect of ORF5, indicating that ORF5 promotes inflammatory response by up-regulating miR-21. Bioinformatics analysis and clinical detection found that nuclear miR-30d was significantly down-regulated after ORF5 overexpression and PCV2 infection, and targeted pri-miR-21 and PCV2 ORF5. Functionally, we found that miR-30d inhibited the levels of miR-21 and inflammatory cytokines in cells. Mechanistically, we demonstrated that ORF5 inhibits miR-30d expression levels through direct binding but not via the circRNA pathway, and miR-30d inhibits miR-21 levels by targeting pri-miR-21. In summary, the present study revealed the molecular mechanism of ORF5 upregulation of miR-21, further refined the molecular chain of PCV2-induced inflammatory response and elucidated the role of miRNAs in it. [ABSTRACT FROM AUTHOR]

This study was designed to investigate the different sequential order of infection for porcine circovirus type 2 (PCV2) and porcine reproductive and respiratory syndrome virus (PRRSV). Thirty-six pigs were randomly assigned to six different treatment groups. The first (hereafter referred to as PRRSV-PCV2) group was inoculated with PRRSV first followed by PCV2d. The second (hereafter referred to as PCV2+PRRSV) group was co-infected with both viruses at the same timepoint (42 days of age). The third (hereafter referred to as PCV2-PRRSV) group was inoculated with PCV2d first followed by PRRSV. A fourth group was only inoculated with PCV2d at 42 days of age, while a fifth group was only inoculated with PRRSV at the same timepoint. The sixth group served as a negative control group. The most important observation discovered that PRRSV only had a potentiation effect on PCV2 in both PRRS-PCV2 and PCV2+PRRSV groups. Both PRRSV-PCV2 and PCV2+PRRSV groups experienced a significant reduction in growth performance compared with control pigs. In addition, PRRSV-PCV2 and PCV2+PRRSV groups exhibited a greater severity in their clinical signs, and/or had higher PCV2 blood and lymphoid viral loads that resulted in a stronger severity of lymphoid lesions compared with PCV2-PRRSV group. Serum TNF-α levels were significantly higher in both PRRS-PCV2 and PCV2+PRRSV groups compared with those in PCV2-PRRS, PCV2, and PRRSV groups. The results of this study demonstrated that divergent clinical outcomes are dependent on the sequential infection order of PCV2 and PRRSV. • Infection order of PRRSV before PCV2d enhances clinical outcomes. • Infection order of concurrent PRRSV and PCV2d enhances clinical outcomes. • Infection order of PCV2d before PRRSV do not enhance clinical outcomes. [ABSTRACT FROM AUTHOR]

Porcine reproductive and respiratory syndrome (PRRS) is one of the costliest diseases to pork producers worldwide. We tested samples from the pregnant gilt model (PGM) to better understand the fetal response to in-utero PRRS virus (PRRSV) infection. Our goal was to identify critical tissues and genes associated with fetal resilience or susceptibility. Pregnant gilts (N=22) were infected with PRRSV on day 86 of gestation. At 21 days post maternal infection, the gilts and fetuses were euthanized, and fetal tissues collected. Fetuses were characterized for PRRS viral load in fetal serum and thymus, and preservation status (viable or meconium stained: VIA or MEC). Fetuses (N=10 per group) were compared: uninfected (UNIF; <1 log/µL PRRSV RNA), resilient (HV_VIA, >5 log virus/µL but viable), and susceptible (HV_MEC, >5 log virus/µL with MEC). Gene expression in fetal heart, kidney, and liver was investigated using NanoString transcriptomics. Gene categories investigated were hypothesized to be involved in fetal response to PRRSV infection: renin- angiotensin-aldosterone, inflammatory, transporter and metabolic systems. Following PRRSV infection, CCL5 increased expression in heart and kidney, and ACE2 decreased expression in kidney, each associated with fetal PRRS susceptibility. Liver revealed the most significant differential gene expression: CXCL10 decreased and IL10 increased indicative of immune suppression. Increased liver gene expression indicated potential associations with fetal PRRS susceptibility on several systems including blood pressure regulation (AGTR1), energy metabolism (SLC16A1 and SLC16A7), tissue specific responses (KL) and growth modulation (TGFB1). Overall, analyses of non-lymphoid tissues provided clues to mechanisms of fetal compromise following maternal PRRSV infection. [Display omitted] • Fetal response to late gestation PRRSV infection is dependent on viral load and preservation status. • Infection alters gene expression associated with blood pressure, glucose transport, sodium homeostasis and immune response. • Gene expression changes associated with fetal susceptibility were prominent in fetal liver, followed by kidney and heart. [ABSTRACT FROM AUTHOR]

Porcine reproductive and respiratory syndrome (PRRS) is one of the most detrimental contagious swine ailments worldwide. Currently, no effective drugs are available for its treatment. Targeting the structural and non-structural proteins (NSP) of the type 2 PRRS virus (PRRSV-2) with small interfering RNA (siRNA) is an effective approach to inhibit PRRSV replication. NSP4, which is highly conserved and possesses 3 C-like serine protease activity (3CLSP), can cleave PRRSV self-proteins, thereby contributing to viral replication. To investigate the mechanism by which NSP4 regulates PRRSV-2 replication and screen for effective siRNA inhibitors of PRRSV-2 replication, the recombinant plasmid pEGFP-C1-NSP4 was constructed, and a control siRNA pair and two siRNA pairs targeting the PRRSV-2 NSP4 gene (shRNA-ctr, shRNA-150, and shRNA-536) were synthesized and cloned into the pSilencer4.1-CMV vector. After 24 h of incubation, Marc-145 cells were transfected with recombinant plasmids, and subsequently infected with different PRRSV-2 (XH-GD, ZQ-GD, GDr180, and JXA1-R). Subsequently, the effects of NSP4 overexpression, shRNA on PRRSV-2 replication were evaluated by assessing cytopathic effects (CPE), TCID 50 , quantitative real-time PCR (qPCR), immunofluorescence assays (IFA), and Western blotting. The data from these CPE, TCID 50 , qPCR, and IFA experiments revealed that NSP4 overexpression significantly enhanced PRRSV-2 replication and shRNA targeting NSP4 can inhibit PRRSV-2 replication in Marc-145 cells, indicating that shRNA could serve as candidate molecules for fundamental research on PRRSV-2. • PRRS is an extremely severe, contagious disease that poses a significant threat to the global swine industry, and it is caused by a virus known as PRRSV. • NSP4, which is highly conserved and possesses 3 C-like serine protease activity (3CLSP), can cleave PRRSV self-proteins, thereby contributing to viral replication. • By assessing cytopathic effects (CPE), TCID50, immunofluorescence assays (IFA), and western blotting, we found that NSP4 overexpression promotes PRRSV-2 replication and shRNA inhibits PRRSV-2 replication. • These results provide a theoretical foundation for prevention and control of the spread of PRRSV. [ABSTRACT FROM AUTHOR]

Pseudorabies virus (PRV), an alphaherpesvirus, is a neglected zoonotic pathogen. Dectin-1 sensing of β-glucan (BG) induces trained immunity, which can possibly form a new strategy for the prevention of viral infection. However, alphaherpesvirus including PRV have received little to no investigation in the context of trained immunity. Here, we found that BG pretreatment improved the survival rate, weight loss outcomes, alleviated histological injury and decreased PRV copy number of tissues in PRV-infected mice. Type I interferons (IFNs) including IFN-α/β levels in serum were significantly increased by BG. However, these effects were abrogated in the presence of Dectin-1 antagonist. Dectin-1-mediated effect of BG was also confirmed in porcine and murine macrophages. These results suggested that BG have effects on type I IFNs with antiviral property involved in Dectin-1. In piglets, oral or injected immunization with BG and PRV vaccine could significantly elevated the level of PRV-specific IgG and type I IFNs. And it also increased the antibody levels of porcine reproductive and respiratory syndrome virus vaccine and classical swine fever vaccine that were later immunized, indicating a broad-spectrum effect on improving vaccine immunity. On the premise that the cost was greatly reducing, the immunological effect of oral was better than injection administration. Our findings highlighted that BG induced type I IFNs related antiviral effect against PRV involved in Dectin-1 and potential application value as a feed additive to help control the spread of PRV and future emerging viruses. [Display omitted] • Dectin-1 sensing of β-glucan (BG) induces trained immunity, which protected against pseudorabies virus (PRV) infection in mice. • Type I IFNs (IFN-α/β) played a role in BG-induced protection via Dectin-1 in vivo and in vitro. • BG enhanced PRV vaccine potency in piglets and BG as feed additives has higher monetary returns. • BG also enhanced non-specific immune responses to vaccines of other swine viruses. [ABSTRACT FROM AUTHOR]

Considerable loss of life, economic slowdown, and public health risk associated with the transmission of airborne respiratory pathogens was underscored by the recent COVID-19 pandemic. Airborne transmission of zoonotic diseases such as the highly pathogenic avian influenza (HPAI) and porcine reproductive and respiratory syndrome virus (PRRSV) has caused major disruptions to domestic and global food security. Current ambient air pathogen monitoring systems involves the collection of air samples from indoor settings suspected of viral contamination, followed by subsequent processing of capture samples to determine the presence and species of airborne viral matter. Nucleic acid amplification techniques are considered the gold standard for pathogen diagnostics. Currently, the necessary extraction and purification of viral RNA from air collector systems prior to sample analysis is both time consuming and performed manually. A monitoring system with separate air sampling and biochemical detection procedures is prone to delay the response to emergent viral threats. In this paper, we present a pathogen monitoring system that overcomes these limitations related to extraction and purification of viral samples and lays the groundwork for a real-time monitor for airborne viral pathogens. We demonstrate a high flow electrostatic precipitator system, that uses small collection wells as counter electrodes for pathogen collection. Integrated reverse-transcriptase loop-mediated isothermal amplification (RT-LAMP) is used for detection of captured viral matter within wells. On-chip heating of collection wells is enabled by integrated planar heaters and small volumes of reagent (30 μ L) directly to the collection wells. We present the design of such a system and show experimental results that demonstrate the use of this device for detection of aerosolized SARS-CoV-2 virus like particles (VLPs), a model pathogen for SARV-CoV-2. • A system for collection and real-time detection of airborne viral pathogens. • Integrates airborne viral capture with isothermal nucleic acid amplification. • Electrostatic capture of airborne viruses into a custom well-plate. • No need for viral extraction from collection surface (one-pot detection). • Experimental verification of real-time detection using aerosolized engineered viral model. [ABSTRACT FROM AUTHOR]

[Display omitted] • Prolidase represents a valuable biological target. • Few ligands for prolidase have been identified so far. • Triggering prolidase has a great therapeutic potential. • Modified aminoacids are among the most promising ligands for prolidase. Prolidase (EC.3.4.13.9) is a Mn+2-dependent dipeptidase that is well known to play a crucial role in several physiological and pathological processes affecting humans. More in particular, this enzyme is involved in the cleavage of proline- and hydroxyproline-containing dipeptides (imidodipeptides), providing a fine regulation of the homeostasis of these two amino acids. Hyperactivity or deficiency of prolidase have been clearly associated to the development and progress of several acute and chronic syndromes (e.g. chronic liver fibrosis, viral and acute hepatitis, cancer, neurological disorders, inflammation, skin diseases, intellectual disability, respiratory infection). Thus, targeting prolidase and modulating its activity is an intriguing field of research with a great therapeutic potential for the next future and for the design of specific and selective drugs. Prolidase can be exploited in two essential ways: as an activator of proline containing prodrugs and by direct interaction. In this latter case, few specific ligands for the title enzyme have been described, but with no reports about their structure–activity relationship. The aim of this comprehensive review is to gather all available information on prolidase targeting so far reported in the literature, to rationalize the observed data and effect into a preliminary structure-relationship picture, to comment about the effectiveness of each reported ligands, and to address future research activities providing new potential and putative natural, semisynthetic, and purely synthetic molecules able to trigger prolidase as the main biological target. [ABSTRACT FROM AUTHOR]

Algae-derived β-glucan has been widely used as a feed additive in the swine industry. The supplementation of β-glucan aims to improve growth performance and modulate the immunity of pigs. However, the potential effects of supplementing β-glucan from algae on immune responses in pigs—specifically antigen-specific immunity—must be determined. In this study, the effects of algae-derived β-glucan supplementation on growth performance, virus neutralising antibody and virus-specific T lymphocytes responses were investigated in pigs. Piglets (n=112 per treatment) were assigned to three treatments including non-supplemented group (control), β-glucan 100 g/ton supplemented group (BG100), and β-glucan 200 g/ton supplemented group (BG200). In this study, production performance of pigs was not found to be different between the experimental groups. Pigs supplemented with β-glucan exhibited high levels of classical swine fever virus (CSFV)-specific producing T lymphocytes and neutralising antibody titer, compared to the control group. Interestingly, supplementation of β-glucan significantly enhanced porcine reproductive and respiratory syndrome virus (PRRSV)-specific interferon-gamma (IFN-γ) producing T lymphocytes, including CD4+, CD8+, and CD4+CD8+ T lymphocyte subpopulations. Moreover, PRRS modified live vaccine (MLV) viremia was reduced in earlier for β-glucan-supplemented pigs compared to the control group. The findings indicate that the algae-derived β-glucan possesses biological potential as an immunomodulatory substance to enhance antiviral immunity, which may contribute to disease resistance in pigs. • The study showed the immunomodulatory effects of algae-derived β-glucan supplementation on enhancing porcine antiviral immunity. • β-glucan supplementation enhanced levels of classical swine fever virus (CSFV)-specific IFN-γ producing T lymphocytes and neutralising antibody. • β-glucan supplementation increased number of porcine reproductive and respiratory syndrome virus (PRRSV)-specific IFN-γ producing T lymphocytes. • β-glucan supplementation shortened the viremia of PRRSV modified live vaccine (MLV). [ABSTRACT FROM AUTHOR]

• CLEVir-X inhibits PRRSV via disruption of IMPDH in vitro. • CLEVir-X induced in vitro mutagenesis of PRRSV, reducing infectivity. • Oral administration of CLEVir-X alleviated clinical signs, lung lesions and PRRSV load in experimentally infected pigs. The aim of this study was to investigate the in vitro and in vivo antiviral effects of CLEVir-X, against porcine reproductive and respiratory syndrome virus (PRRSV). CLEVir-X is a nucleoside analogue and a dialdehyde form of xanthosine. CLEVir-X demonstrated antiviral action during the in vitro portion of this experiment with its inosine monophosphate dehydrogenase (IMPDH) inhibition against PRRSV. The anti-PRRSV effect of CLEVir-X was recovered through supplementation with guanosine. This suggests that PRRSV replication may be regulated through IMPDH and its guanosine biosynthetic pathway. CLEVir-X treatment in cultures resulted in mutation frequency increase of up to 7.8-fold within the viral genomes (e.g. ORF6) compared to their parallel, untreated cultures. The incorporation of CLEVir-X into the viral genome causes lethal mutagenesis and subsequent decrease in specific infectivity. During the in vivo antiviral experiment, 21-day-old pigs began oral administration of 5 mL of phosphate buffered saline containing CLEVir-X (with purity of 68 % and dosage of 40 mg/kg body weight). This treatment was provided twice daily at 9:00AM and 5:00PM for 14 days. Pigs were simultaneously intranasally inoculated with PRRSV at the beginning of CLEVir-X treatment (21 days of age). Several beneficial effects from the oral administration of CLEVir-X were observed including reduction of body temperature, alleviation of respiratory clinical signs, decreased PRRSV load in both blood and lung tissues, and mitigation of lung interstitial pneumonia lesions. The results of the present study demonstrated that CLEVir-X has mutagenic and nonmutagenic modes of antiviral action against PRRSV based on both in vitro and in vivo antiviral experiments. [ABSTRACT FROM AUTHOR]

Porcine reproductive and respiratory syndrome virus (PRRSV) is a major pathogen affecting pigs and belongs to the enveloped plus-stranded RNA virus family Arteriviridae. A unique feature of Arteriviruses is that the genes encoding the structural proteins overlap at their 3' and 5' ends. This impedes mutagenesis opportunities and precludes the binding of short peptides for antibody detection, as this would alter the amino acids encoded by the overlapping gene. In this study, we aimed to generate infectious PRRSV variants with separated genes encoding the minor glycoproteins Gp2, Gp3, and Gp4, accompanied by appended tags for detection. All recombinant genomes facilitate the release of infectious virus particles into the supernatant of transfected 293 T cells, as evidenced by immunofluorescence of infected MARC-145 cells using anti-nucleocapsid antibodies. Furthermore, expression of Gp2-Myc and Gp3-HA was confirmed through immunofluorescence and western blot analysis with tag-specific antibodies. However, after two passages of Gp2-Myc and Gp3-HA viruses, the appended tags were completely removed as indicated by sequencing the viral genome. Recombinant viruses with separated Gp2 and Gp3 genes remained stable for at least nine passages, while those with Gp3 and Gp4 genes separated reverted to wild type after only four passages. Notably, this virus exhibited significantly reduced titers in growth assays. Furthermore, we introduced a tag to the C-terminus of Gp4. The Gp4-HA virus was consistently stable for at least 10 passages, and the HA-tag was detectable by western blotting and immunofluorescence. • Most genes encoding the structural proteins are overlapping in the PRRSV genome. • We separated the genes encoding Gp2, Gp3 and Gp4 of PRRSV-2 and added small tags. • Although initially viable, recombinant Gp2-Myc and Gp3-HA viruses lose the tags. • Even viruses with separated genes rapidly reverted to the wild-type sequence. • ORF4 does not overlap with the subsequent genes and Gp4-HA virus is stable. [ABSTRACT FROM AUTHOR]

Pig production is increasing annually in Africa as it is recognized as a significant source of income, livelihood and food security, particularly in rural communities. Understanding the circulating swine pathogens is crucial for the success of this emerging industry. Although there is extensive data available on the African swine fever virus due to its devastating impact on pig production, knowledge about the presence of other viral swine pathogens on the continent is still extremely limited. This review discusses what is currently known about six swine pathogens in Africa: classical swine fever virus, porcine reproductive and respiratory syndrome virus, porcine circovirus-2, porcine circovirus-3, porcine parvovirus-1, and pseudorabies virus. Gaps in our knowledge are identified and topics of future focus discussed. • Pig production is increasing annually in Africa. • Limited knowledge on the presence of viral swine pathogens in Africa. • Gaps in our knowledge on the presence of six viral swine pathogens are identified. • Topics of future focus discussed. [ABSTRACT FROM AUTHOR]

Glaesserella parasuis (G. parasuis) is the causative agent of porcine Glässer's disease, resulting in high mortality rates in pigs due to excessive inflammation-induced tissue damage. Previous studies investigating the protective effects of G. parasuis vaccination indicated a possible role of ApoA1 in reflecting disease progression following G. parasuis infection. However, the mechanisms of ApoA1 expression and its role in these infections are not well understood. In this investigation, newborn porcine tracheal (NPTr) epithelial cells infected with G. parasuis were used to elucidate the molecular mechanism and role of ApoA1. The study revealed that the AMPK pathway activation inhibited ApoA1 expression in NPTr cells infected with G. parasuis for the first time. Furthermore, Egr1 was identified as a core transcription factor regulating ApoA1 expression using a CRISPR/Cas9-based system. Importantly, it was discovered that APOA1 protein significantly reduced apoptosis, pyroptosis, necroptosis, and inflammatory factors induced by G. parasuis in vivo. These findings not only enhance our understanding of ApoA1 in response to bacterial infections but also highlight its potential in mitigating tissue damage caused by G. parasuis infection. • High-virulent G. parasuis SH0165 stain reduces the expression of ApoA1 in vivo and in vitro. • The AMPK signaling pathway regulates ApoA1 expression via transcription factor Egr1 during G. parasuis infection. • ApoA1 inhibited programmed cell death and pro-inflammatory factors production during G. parasuis infection in vivo [ABSTRACT FROM AUTHOR]

Porcine reproductive and respiratory syndrome (PRRS) caused by the PRRS virus (PRRSV) is one of the most severe swine diseases causing great economic losses for the international swine industry. Non-structural protein 4 (NSP4) is critical to the life cycle of PRRSV and contains dominant B cell epitopes. This study prepared a monoclonal antibody against Nsp4, and 2D11, which contained the sequence 138KQGGGIVTRPSGQFCN153, was confirmed as the epitope. A 2D11-based double antibody sandwich enzyme-linked immunosorbent assay (dasELISA) was next developed with a cut value of 0.1987. A total of 1354 pig serum samples were detected by dasELISA and compared to a commercial ELISA kit (N-coated iELISA), resulting in a positive coincidence rate of 98.8% and negative coincidence rate of 96.9%. A total of 119 sera were positive by dasELISA while negative by iELISA. Higher positive rates by dasELISA were found in pig farms where PRRSV antibody levels varied widely. These results indicated that the dasELISA was a useful tool to detect PRRSV antibody in clinical samples. • A monoclonal antibody (2D11) to PRRSV NSP4 was prepared. • 138KQGGGIVTRPSGQFCN15 was identified as a dominant epitope. • A dasELISA based on 2D11 was developed. • The developed dasELISA was an effective tool for clinical samples. [ABSTRACT FROM AUTHOR]

Porcine respiratory disease complex, which is caused by a combination of pathogens, including swine influenza A virus (SIV) and porcine reproductive respiratory syndrome virus (PRRSV), results in significant economic losses in pig production systems. Nasopharynx-associated lymphoid tissue (NALT) plays an important role in the uptake of pathogens and defence of the nasal mucosa in rodents and humans. We characterized NALT M cells in pigs and detected SIV antigen and PRRSV nucleic acid in NALT using histopathological, immunohistochemical and in-situ hybridization analyses. All SIV- and PRRSV-positive cases examined had suppurative nasopharyngitis and pneumonia. M cells were detected by immunohistochemistry and the distribution of M cells showed an increase in the middle section of NALT. SIV antigen was detected in M cells and PRRSV nucleic acid was demonstrated in the cytoplasm of macrophages in NALT. We believe that SIV and PRRSV infection in the upper respiratory tract induces local immunosuppression and these results confirm that swine NALT is a location for virus replication and may be strongly associated with the development of pneumonia in pigs. [Display omitted] [ABSTRACT FROM AUTHOR]

A new porcine circovirus type 2e (PCV2e) genotype was recently isolated from diseased pigs. To investigate the pathogenicity of PCV2e, groups of conventional pigs were inoculated in one of three ways: with PCV2e only, with Mycoplasma hyopneumoniae and PCV2e, or with PCV2e and porcine reproductive and respiratory syndrome virus (PRRSV). Pigs were examined post mortem at 21 days post inoculation. Pigs in the M. hyopeumoniae /PCV2e group were inoculated intratracheally with M. hyopneumoniae at 4 weeks of age followed by an intranasal inoculation with PCV2e at 6 weeks of age. Pigs in the PCV2e/PRRSV group were inoculated intranasally with PCV2e and PRRSV at 6 weeks of age. Significant differences in PCV2e loads in blood or lymph nodes, or in the severity of lymphoid lesions, were not detected between the M. hyopneumoniae /PCV2e and PCV2e/PRRSV groups. All pigs co-infected with either M. hyopneumoniae /PCV2e or PCV2e/PRRSV developed mild porcine circovirus-associated disease (PCVAD), whereas none of the pigs infected with PCV2e alone developed PCVAD. Production of PCVAD in pigs therefore appears to require PCV2e infection simultaneously with an additional infectious agent such as M. hyopneumoniae or PRRSV for full disease expression in pigs. These results demonstrate that PCV2e is not associated with significant clinical disease as assessed by levels of PCV2e viraemia and severity of lymphoid lesions. [ABSTRACT FROM AUTHOR]

Reproductive failure caused by porcine reproductive and respiratory syndrome virus (PRRSV) is characterized by embryonic death and weak-born piglets and is associated with placental cell apoptosis and impairment of endometrial integrity. Here, we aimed to determine whether endometrial epithelial barrier function and viability were altered following PRRSV type 1 or type 2 infection. PRRSV inoculation was examined at the apical or basolateral side of porcine glandular endometrial epithelial cell cultures isolated from 4- to 6-month-old PRRSV-free herd gilts (n = 7 pigs). On the apical side, four days postinfection (4 dpi) with type 2 PRRSV, transepithelial electrical resistance decreased by 31% ± 5%, and paracellular permeability to fluorescein isothiocyanate-dextran (4 kDa) increased by 10-fold as compared with the mock and type 1 infection. Real-time polymerase chain reaction results revealed that both PRRSV types upregulated the mRNA expression of the barrier builder tight junction protein (TJ) Cldn5 , but downregulated pore-forming TJ Cldn7. Additionally, the expression of other TJ genes, i.e., Cldn3 and Cldn8 , was differentially increased by PRRSV type 1 and that of zonula occludens-1 was increased by PRRSV type 2. MTT assays indicated an increase in porcine glandular endometrial epithelial cell culture at 2–6 dpi following type 2 infection. Analysis of apoptosis using Annexin/propidium iodide staining combined with flow cytometry showed that the percentage of viable cells decreased, accompanied by a significantly higher dead cell population following PRRSV type 2 infection at 2–4 dpi. PRRSV type 1 infection also induced dead cells (>4%) at 2 dpi; however, the cell population recovered at 4 dpi. In conclusion, PRRSV type 2 infection caused more severe TJ barrier dysfunction and reduced cell viability compared with PRRSV type 1 infection in the porcine endometrium. Impairment in the membrane integrity of the maternal glandular endometrium may be the underlying mechanism of PRRSV-induced reproductive failure in pregnant sows. [Display omitted] • PRRSV inducing dysfunction and apoptosis of endometrial cells were determined. • PRRSV type 2 induced TJ disruption with cell proliferation and apoptosis. • PRRSV type 1 and 2 altered TJ family protein expression, differently. • PRRSV disrupting PEG function may associate with reproductive failure. [ABSTRACT FROM AUTHOR]

African swine fever (ASF) caused by African swine fever virus (ASFV) is becoming a serious threat to the swine industry worldwide. CD2v is a key pathogenic factor of ASFV and the protective antigen with low immunogenicity, whereas viral protein-based nanoparticles have advantages of precise assembly and high immunogenicity. In this study, the CD2v protein fused with Norovirus (NoV) P particle assembled into nanoparticle for improved immunogenicity. Then, CD2v protein nanoparticle and monomer CD2v protein were expressed in HEK293F cells. The former induced higher levels of antibodies, and thus highly potent monoclonal antibodies (mAbs) were generated and characterized. The highest antibody titration of mAb 10A3 reached 1:2048000, and mAb 2E9 had the highest inhibition percent of 84% when competed with ASFV positive serum. Meanwhile, all mAbs reacted specifically with the denatured CD2v protein, and the linear epitope with the location of amino acids 28th to 51st of CD2v extracellular domain sequence was identified. In summary, this study produced a highly immunogenic CD2v protein and generated high-titer mAbs, the precise location of linear epitope on the CD2v was further determined. These findings may provide a powerful help for etiology and serological detection of ASFV. • Highly immunogenic CD2v protein nanoparticle was obtained. • High-titer mAbs induced by CD2v protein nanoparticle were generated, and mAb 10A3 had the highest antibody titration of 1:2048000. • The linear epitope from the 28th to the 51st position of amino acids locating in the CD2v extracellular domain sequence was first reported. [ABSTRACT FROM AUTHOR]

PRRSV causes major economic losses to swine industry world-wide, which requires innovative antiviral agents. Porcine scavenger receptor CD163 has been identified as an essential fusion receptor for Porcine reproductive and respiratory Syndrome Virus (PRRSV) infection. In this study, novel antiviral peptides from pCD163 against PRRSV were developed based on broad neutralizing monoclonal antibodies. SRCR-5-9 of pCD163 from baculovirus efficiently binds to PRRSVs of lineage 8 and lineage 1, blocking infection in PAMs. A batch of monoclonal antibodies targeting SRCR-5-9 were generated and characterized. 8H2 and 4H7 block PRRSV infection by the disruption in viral attachment to PAMs. Virus titer reduced 100–1000 folds in average and the virus copy number decreased about 104 folds with these antibodies. Linear epitopes of 8H2 and 4H7 were individually localized in SRCR6 (1-30 aa) and PSTI(1-15aa) of pCD163. Mutations of SRCR6 NI1718KT and PST SS1314AA abolished the recognition of 8H2 and 4H7 to the corresponding region individually. Peptides derived from the linear epitopes displayed a broad inhibitory effect on PRRSVs of different lineages in a dose-dependent manner and further modulated PRRSV-related NF-κB pathway. In conclusion, these findings deepen the understanding in the interaction between PRRSV and pCD163 receptor and provide alternative universal antiviral strategies against PRRSV. [ABSTRACT FROM AUTHOR]

Understanding why intrauterine growth restricted (IUGR) fetuses are more resilient to transplacental porcine reproductive and respiratory syndrome virus-2 (PRRSV-2) infection compared to normal fetuses may lead to alternative approaches to control PRRS. Our objective was to compare gene expression of a subset of tight junction proteins in the endometrium (END) and placenta (PLC) of i) IUGR vs N-IUGR fetuses, and ii) across disease progression phenotypes following PRRSV-2 infection. In experiment 1, snap frozen END and PLC from fetuses of non-infected control dams (CTRL) and from high viral load viable (HVL-VIA) fetuses, with both groups further classified as either IUGR or non(N)-IUGR based on brain: liver weight ratio were strategically selected from a large challenge trial. In experiment 2, similar tissues were randomly selected from CTRL and from uninfected thymus (UNIF), (HVL-VIA) and HVL meconium-stained in the body (HVL-MEC-B) of PRRSV-infected dams. The expression of claudin (CLDN) 1, 3, 4, 5, 6, 7, 10, tight junction protein 1 (TJP1) and occludin (OCLN) genes were evaluated by PCR. There were no significant group differences between IUGR and N-IUGR groups, regardless of infection status, that explained the resilience of IUGR fetuses. Regarding disease progression, elevated CLDN3 was observed in END of UNIF, CLDN6 expression was lower in PLC when the fetus became infected (HVL-VIA), and CLDN10 elevated in PLC in fetuses showing evidence of compromise (HVL-MEC-B). Lastly, OCLN gene expression was higher in the END and PLC following maternal infection. In conclusion, differences in TJ integrity were mainly observed following PRRSV-2 infection with stepwise changes corresponding with disease progression. • PRRS resilience in IUGR fetuses was not related to tight junction expression. • Tight junction expression was modified in diseased progression groups. • OCLN expression increased in endometrium and placenta after maternal infection. • CLDN6 expression decreased in placenta when the fetus became infected. • CLDN10 expression increased in placenta in fetuses showing evidence of compromise. [ABSTRACT FROM AUTHOR]

• Recombinant epitope protein rEP-PM contains some B cell epitopes and T cell epitopes of PRRSV-GP5, PRRSV-M, Mhp-P46 and Mhp-P65 protein. • rEP-PM could induce the production of high levels of antibodies. • rEP-PM could induce the Th1-dominant cellular immune response in mice. • rEP-PM has good immunoreactivity with anti-rEP-PM, anti-PRRSV and anti-Mhp antibodies. • Anti-rEP-PM antibody could specifically recognize proteins from PRRSV and Mhp. Porcine reproductive and respiratory syndrome virus (PRRSV) and Mycoplasma hyopneumoniae (M. hyopneumoniae , Mhp) are two of the most common pathogens involved in the porcine respiratory disease complex (PRDC) resulting in significant economic losses worldwide. Vaccination is the most effective approach to disease prevention. Since PRRSV and Mhp co-infections are very common, an efficient dual vaccine against these pathogens is required for the global swine industry. Compared with traditional vaccines, multi-epitope vaccines have several advantages, they are comparatively easy to produce and construct, are chemically stable, and do not have an infectious potential. In this study, to develop a safe and effective vaccine, B cell and T cell epitopes of PRRSV-GP5, PRRSV-M, Mhp-P46, and Mhp-P65 protein had been screened to construct a recombinant epitope protein rEP-PM that has good hydrophilicity, strong antigenicity, and high surface accessibility, and each epitope is independent and complete. After immunization in mice, rEP-PM could induce the production of high levels of antibodies, and it had good immunoreactivity with anti-rEP-PM, anti-PRRSV, and anti-Mhp antibodies. The anti-rEP-PM antibody specifically recognizes proteins from PRRSV and Mhp. Moreover, rEP-PM induced a Th1-dominant cellular immune response in mice. Our results showed that the rEP-PM protein could be a potential candidate for the development of a safe and effective multi-epitope peptide combined vaccine to control PRRSV and Mhp infections. [ABSTRACT FROM AUTHOR]

Piglet pre-weaning mortality (PWM) is a significant issue in the U.S. swine industry, causing economic losses and raising sustainability and animal welfare concerns. This study conducted a multivariable analysis to identify factors associated with PWM in a Midwestern U.S. swine production system. Weekly data from 47 sow farms (7207 weaning weeks) were captured from January 2020 to December 2022. Initially, 29 variables regarding farm infrastructure, productivity parameters, health status, and interventions were selected for univariate analysis to assess their association with PWM. The initial multivariable analysis included the variables with P < 0.20 in the univariate analyses. A backward stepwise model selection was conducted by excluding variables with P > 0.05, and the final multivariable model consisted of 19 significant risk factors and 6 interaction terms. The overall average PWM for the study population was 14.02 %. Yearly variations in PWM were observed, with the highest recorded in 2020 (16.61 %) and the lowest in 2021 (15.78 %). Cohorts with a pond water source, lower farrowing rate (71.9 %), higher farrowing parity (5.1), shorter gestation length (116.2 days), and using oxytocin during farrowing had increased PWM. The higher productivity parameters such as mummies rate, stillborn rate, and average total born, the higher the PWM was. Additionally, health status and intervention-related factors were associated with PWM, where higher PWM rates were observed in herds facing porcine reproductive and respiratory syndrome virus (PRRSV) outbreaks, porcine epidemic diarrhea virus (PEDV) positive, the weeks before and during feed medication, and weeks without using Rotavirus vaccine or Rotavirus feedback. Altogether, these results corroborate that PWM is a multifactorial problem, and a better understanding of the risk factors is essential in developing strategies to improve survival rates. Therefore, this study identified the major risk factors associated with PWM for groups of pigs raised under field conditions, and the results underscore the significance of data analysis in comprehending the unique challenges and opportunities inherent to each system. [ABSTRACT FROM AUTHOR]

• A coinfection study was conducted using Atypical porcine pestivirus (APPV). • APPV in pigs has been characterised recently. • Rectal temperatures were significantly higher in the coinfected group. • Coinfected pigs had enhanced lung pathology at post-mortem. • APPV infection did not significantly impact virological or immunological outcomes. Atypical porcine pestivirus (APPV) is a novel member of the Pestivirus genus detected in association with congenital tremor (CT) type A-II outbreaks and from apparently healthy pigs, both as singular infection and as part of multi-pathogen infections. 'Classical' pestiviruses are known to cause immunosuppression of their host, which can increase susceptibility to secondary infections, severely impacting health, welfare, and production. To investigate APPV's effect on the host's immune system and characterise disease outcomes, 12 piglets from a natural APPV CT type A-II outbreak were experimentally infected with porcine reproductive and respiratory syndrome virus (PRRSV), a significant porcine pathogen. Rectal temperatures indicating febrile responses, viremia and viral-specific humoral and cellular responses were assessed throughout the study. Pathological assessment of the lungs and APPV-PRRSV co-localisation within the lungs was performed at necropsy. Viral co-localisation and pathological assessment of the lungs (Immunohistochemistry, BaseScope in situ hybridisation) were performed post-mortem. APPV status did not impact virological or immunological differences in PRRSV-infected groups. However, significantly higher rectal temperatures were observed in the APPV+ve/PRRSV+ve group over four days, indicating APPV increased the febrile response. Significant differences in the lung consolidation of the apical and intermediate lobes were also present, suggesting that APPV co-infection may augment lung pathology. [ABSTRACT FROM AUTHOR]

The Porcine reproductive and respiratory syndrome (PRRS) causes severe financial losses to the global swine industry. Due to continuous virus evolution, the protection against the PRRS provided by current vaccines is limited. In order to find new antiviral strategies, this study investigated the antiviral potential of antimicrobial peptides (AMPs) against PRRSV. Given the diversity of PRRSV strains and the limited effectiveness of existing vaccines in controlling PRRSV, this study evaluated the inhibitory effects of KLAK, Cecropin B, Piscidin1, and Caerin1.1 on 3 strains of PRRSV (lineage 5 classical strain, lineage 8 highly pathogenic strain, and lineage 1 NADC30-like strain). Caerin1.1 exhibited significant dose-dependent antiviral activity, with an effective concentration (EC 50) of 7.5 μM. Caerin1.1 effectively inhibited PRRSV replication when added before or in early infection but showed reduced effectiveness when added in late infection, indicating its potential involvement in targeting early transcription mechanisms of viral RNA polymerase and significantly upregulating cytokine gene expression. In the NADC30 strain-based animal infection model, Caerin1.1 treatment significantly reduced lung viral loads and inflammation in the lungs of PRRSV-infected pigs, with a mortality rate of 0 % (0/5) in the treated group compared to 66.67 % (4/6) in the untreated group, indicating a reduction in the mortality rate. Additionally, compared with the untreated group, the Caerin1.1-treated group showed significant improvements, such as lighter fever, more daily weight gain, less clinical symptoms, less viral load in blood, and less virus oral shedding (P < 0.05). These findings reveal the potential of antimicrobial peptides as PRRSV therapeutic agents and suggest that Caerin1.1 is a promising candidate for a novel anti-PRRSV drug. • Caerin1.1 exhibits a dose-dependent inhibitory effect on PRRSV replication. • Caerin1.1 shows broad-spectrum antiviral activity against various PRRSV subtypes. • Caerin1.1 alleviates symptoms, reduces tissue damage and lowers mortality in infected pigs. [ABSTRACT FROM AUTHOR]

Circular RNA (circRNA) is abundantly expressed in preimplantation embryos and embryonic stem cells in mice and humans. However, its function and mechanism in early development of mammalian embryos remain unclear. Here, we showed that circHIRA mediated miR-196b-5p to regulate porcine early embryonic development. We verified the circular feature of circHIRA by sanger sequencing, and proved the authenticity of circHIRA by enzyme digestion test. HIRA and circHIRA were expressed in porcine early embryos, and its expression levels significantly increased from 8-cell stage onwards and reached the maximum at the blastocyst stage. Functional studies revealed that circHIRA knockdown not only significantly reduced the developmental efficiency of embryos from 8-cell stage to blastocyst stage, but also impaired the blastocyst quality. Mechanistically, integrated analysis of miRNA prediction and gene expression showed that circHIRA knockdown significantly increased the expression of miR-196b-5p in porcine early embryos. Furthermore, miR-196b-5p inhibitor injection could rescue the early development of circHIRA knockdown embryos. Taken together, the findings reveal that circHIRA regulates porcine early embryonic development via inhibiting the expression of miR-196b-5p. [ABSTRACT FROM AUTHOR]

Porcine reproductive and respiratory syndrome (PRRS) is one of the most widespread illnesses in the world's swine business. To detect the antibodies against PRRSV-2, a blocking enzyme-linked immunosorbent assay (B-ELISA) was developed, utilizing a PRRSV-2 N protein monoclonal antibody as the detection antibody. A checkerboard titration test was used to determine the optimal detection antibody dilution, tested pig serum dilution and purified PRRSV coated antigen concentration. After analyzing 174 negative pig sera and 451 positive pig sera, a cutoff value of 40 % was selected to distinguish between positive and negative sera using receiver operating characteristic curve analysis. The specificity and sensitivity of the assay were evaluated to equal 99.8 % and 96 %, respectively. The method had no cross-reaction with PCV2, PRV, PPV, CSFV, PEDV, TGEV, and PRRSV-1 serum antibodies, and the coefficients of variation of intra-batch and inter-batch repeatability experiments were both <10 %. A total of 215 clinical serum samples were tested, and the relative coincidence rate with commercial ELISA kit was 99.06 %, and the kappa value was 0.989, indicating that these two detection results exhibited high consistency. Overall, the B-ELISA should serve as an ideal method for large-scale serological investigation of PRRSV-2 antibodies in domestic pigs. • A monoclonal antibody that can specifically recognize PRRSV-2 N protein was successfully screened using recombinant N protein as immunogen. • A blocking enzyme-linked immunosorbent assay (B-ELISA) for specific detection of PRRSV-2 antibody was established. • The B-ELISA assay was evaluated in the laboratory and showed comparable performance to commercial ELISA kit, and even had higher analytical sensitivity. [ABSTRACT FROM AUTHOR]

porcine reproductive and respiratory syndrome (PRRS), caused by porcine reproductive and respiratory syndrome virus (PRRSV) infection, is an important swine infectious disease that causes substantial losses worldwide each year. PRRSV is a positive-sense single-stranded RNA virus that is highly susceptible to mutation and recombination, making vaccine and drug research for the disease extremely difficult. In this study, the binding of PRRSV nsp2 to HSP71 protein was detected by using the IP/MS technique. And the inhibitory effect of HSP71 on nsp2 antagonistic activity was validated by measuring NF-kB luciferase reporter. According to stress from inhibitory effects, the amino acid variation profile of PRRSV nsp2 under HSP71 stress was further analyzed using second-generation sequencing. Surprisingly, the results indicated that HSP71 pressure limits the random mutations of PRRSV nsp2 and maintains the dominant PRRSV strain within the population. Mutant strain showed weaker antagonistic activity and replication capability in cell. These results imply the binding of HSP71 with PRRSV nsp2 may lead to maintain the stability of highly virulent strains of PRRSV. • Hsp71 binds and inhibits PRRSV nsp2. • hsp71 binding contributes to the stabilization of PRRSV nsp2. • PRRSV nsp2 is prone to random mutations in normal cells. [ABSTRACT FROM AUTHOR]

Porcine epidemic diarrhea virus (PEDV) is a devastating pathogen of acute- gastrointestinal infectious diseases, which can cause vomiting, diarrhea, dehydration and high morbidity and mortality among neonatal piglets. Humoral immunity plays a vital role in the host anti-PEDV infection process, but the mechanism of PEDV-induced B-cell immune response remains unknown. In this study, the effects of PEDV infection on CD21+ B cell activation were systematically analyzed through animal experiments. Enzyme-linked immunosorbent assays (ELISA) revealed that low levels of serum-specific IgA, IgM, or IgG were detected in piglets after PEDV infection, respectively. Serum interleukin (IL)-6 levels increased significantly at 4 d after infection, and the levels of IL-4, B-cell activating factor (BAFF), interferon (IFN)-γ, transforming growth factor (TGF)-β and IL-10 decreased at 7 d after infection. Fluorescence-activated cell sorting (FACS) showed that expression levels of CD21, MHC Ⅱ, CD40, and CD38 on B cell surfaces were significantly higher. In contrast, the proportions of CD21+IgM+ B cells were decreased in peripheral blood mononuclear cells (PBMCs) from the infected piglets. No differences were found in the percentage of CD21+CD80+ and CD21+CD27+ B cells in PBMCs from the infected piglets. In addition, the number of CD21+B cells in PBMCs stimulated with PEDV in vitro was significantly lower. No significant change in the mRNA expression of BCR molecules was found while the expression levels of paired immunoglobulin-like receptor B (PIR-B), B cell adaptor molecule of 32 kDa (Bam32) and BAFF were decreased. In conclusion, our research demonstrates that virulent strains of PEDV profoundly impact B cell activation, leading to alterations in phenotypic expression and BCR signaling molecules. Furthermore, this dysregulation results in compromised specific antibody secretion and perturbed cytokine production, highlighting the intricate immunological dysfunctions induced by PEDV infection. • PEDV virulent strains significantly affected B cell activation in infected piglets. • PEDV caused the insufficient expression of BCR signaling molecules Bam and BAFF. [ABSTRACT FROM AUTHOR]

Gene editing (GnEd) involves using a site-directed nuclease to introduce a double-strand break (DSB) at a targeted location in the genome. A literature search was performed on the use of GnEd in animals for agricultural applications. Data was extracted from 212 peer-reviewed articles that described the production of at least one living animal employing GnEd technologies for agricultural purposes. The most common GnEd system reported was CRISPR/Cas9, and the most frequent type of edit was the unguided insertion or deletion resulting from the repair of the targeted DSB leading to a knock-out (KO) mutation. Animal groups included in the reviewed papers were ruminants (cattle, sheep, goats, n =63); monogastrics (pigs and rabbits, n =60); avian (chicken, duck, quail, n =17); aquatic (many species, n =65), and insects (honeybee, silkworm, n =7). Yield (32%), followed by reproduction (21%) and disease resistance (17%) were the most commonly targeted traits. Over half of the reviewed papers had Chinese first-authorship. Several countries, including Argentina, Australia, Brazil, Colombia and Japan, have adopted a regulatory policy that considers KO mutations introduced following GnEd DSB repair as akin to natural genetic variation, and therefore treat these GnEd animals analogously to those produced using conventional breeding. This approach has resulted in a non-GMO determination for a small number of GnEd food animal applications, including three species of GnEd KO fast-growing fish, (red sea bream, olive flounder and tiger pufferfish in Japan), KO fish and cattle in Argentina and Brazil, and porcine reproductive and respiratory syndrome (PRRS) virus disease-resistant KO pigs in Colombia. • Gene editing (GnEd) can introduce useful genetic alterations in food animals • A total of 212 papers using GnEd in animals for agriculture were identified • Yield, reproduction, and disease resistance were the most commonly targeted traits • CRISPR/Cas9 was by far the most prevalent GnEd system used to edit animal genomes • Three species of GnEd fast-growing finfish have already been commercialized in Japan [ABSTRACT FROM AUTHOR]

Respiratory diseases due to viral or bacterial agents, either alone or in combination, cause substantial economic burdens to the swine industry worldwide. Rapid and reliable detection of causal pathogens is crucial for effective epidemiological surveillance and disease management. This research aimed to employ the multiplex ligation-dependent probe amplification (MLPA) assay for simultaneous detection of seven distinct pathogens causing respiratory problems in swine, porcine reproductive and respiratory syndrome virus (PRRSV), swine influenza virus (SIV), porcine respiratory coronavirus (PRCV), porcine circovirus type 2 (PCV2), Pasteurella multocida , Actinobacillus pleuropneumoniae , and Glässerella parasuis. The results indicated no probe cross-reactivity among the seven target agents with other swine pathogens. The detection limits ranged from 5 to 34 copies per assay for the target organisms. The MLPA assay was evaluated with 88 samples and compared to real-time or multiplex PCR for the target pathogens. The MLPA assay demonstrated high relative test sensitivities (100 %) and reasonable to good relative specificities at 62.5 %, 95.1 %, 86.8 %, and 97.6 % for PRRSV, P. multocida , G. parasuis , and PCV2, respectively, relative to comparator PCR assays. In 71 samples where MLPA and comparator PCR assays matched exactly, infections were detected in 64 samples (90.1 %), with PRRSV being the most commonly found virus and 50.7 % of the samples showing co-infection with two to five of the pathogens. This approach serves as a valuable tool for conducting differential diagnoses and epidemiological investigations of pathogen prevalence within swine populations. • An MLPA method was developed to detect seven swine pathogens simultaneously. • PRRSV, PCV2, P. multocida and G. parasuis were detected in nasal swabs. • SIV, porcine respiratory coronavirus and A. pleuropneumoniae were also targeted. • MLPA assay is rapid for specific detection of multiple pathogens in pigs. • The assay has value in epidemiological surveillance of swine diseases. [ABSTRACT FROM AUTHOR]

The cytotoxic mycotoxin deoxynivalenol (DON) reportedly has adverse effects on oocyte maturation and embryonic development in pigs. Recently, the interplay between cell apoptosis and endoplasmic reticulum (ER) stress has garnered increasing attention in embryogenesis. However, the involvement of the inositol-requiring enzyme 1 (IRE1)/c-jun N-terminal kinase (JNK)/C/EBP homologous protein (CHOP) pathways of unfolded protein response (UPR) signaling in DON-induced apoptosis in porcine embryos remains unknown. In this study, we revealed that exposure to DON (0.25 μM) substantially decreased cell viability until the blastocyst stage in porcine embryos, concomitant with initiation of cell apoptosis through the IRE1/JNK/CHOP pathways in response to ER stress. Quantitative PCR confirmed that UPR signaling-related transcription factors were upregulated in DON-treated porcine blastocysts. Western blot analysis showed that IRE1/JNK/CHOP signaling was activated in DON-exposed porcine embryos, indicating that ER stress-associated apoptosis was instigated. The ER stress inhibitor tauroursodeoxycholic acid protected against DON-induced ER stress in porcine embryos, indicating that the toxic effects of DON on early developmental competence of porcine embryos can be prevented. In conclusion, DON exposure impairs the developmental ability of porcine embryos by inducing ER stress-mediated apoptosis via IRE1/JNK/CHOP signaling. • DON exposure reduces the developmental capacity of porcine blastocysts. • DON exposure decreases ER-phagy. • 0.1 μM DON induces the IRE1 pathway due to Ca2+ imbalance. • 0.25 μM DON triggers the JNK/CHOP pathways by increasing mitochondrial Ca2+. • TUDCA protects ER stress on development of DON-exposed porcine embryos. [ABSTRACT FROM AUTHOR]

Porcine epidemic diarrhea virus (PEDV) causes immensely large economic losses worldwide in the swine industry. PEDV attacks the intestine, disrupts intestinal epithelium morphology and barrier integrity, and results in profound diarrhea and high mortality. A commercially available isotonic protein solution (IPS) (Tonisity Px) has anecdotally been reported to be effective in supportive treatment of piglets with active PEDV infections. This study evaluated the effects of supplementing (or not) the drinking water of 14 day old PEDV-infected piglets with the IPS on the content of E-cadherin, fibronectin, interferon-alpha (IFN-α), and matrix metalloproteinase 9 (MMP-9) in duodenal tissue. The content of PEDV DNA in feces was also measured. Though both groups had similar PEDV shedding at day 1, IPS piglets had significantly lower PEDV shedding at day 5, 14 and 21. The IPS group also had a shorter duration of PEDV virus shedding. Levels of E-cadherin and fibronectin, both of which are structural proteins in the intestine, remained unchanged from baseline in the IPS group, whereas the same molecules decreased significantly in the control group. IFN-α, an antiviral cytokine, and MMP-9, an enzyme that aids in tissue remodeling, were increased at days 5 and 14 post infection, and then decreased at day 21 post-infection in the IPS group compared to control. Overall, the IPS used in this study enhanced epithelial intercellular adhesion (E-cadherin) and extracellular matrix structure (fibronectin), resulted in significantand favorable changes in MMP-9 activity, and favorably modulated IFN-α production. This is the first report of this panel of biomarkers, especially MMP-9 and IFN-α, in the face of in vivo PEDV infection. This is also the first report to investigate a commercially available swine product that does not need to be administered in solid feed, and that is already registered for use throughout Asia, Europe, South America, and North America. Overall, the results of this study serve to clarify the behavior of 4 key biomarkers in the presence of in vivo PEDV infection. The results also indicate that IPS (Tonisity Px) supplementation is a viable intervention to modulate the porcine intestinal immune response with favorable effects on the intestine. [Display omitted] • The Porcine Epidemic Diarrhea Virus (PEDV) results in significant losses in the pig industry globally. • An intervention was tested in the form of a commercially available Isotonic Protein Solution (Tonisity Px; IPS). • Two groups of PEDV-infected pigs (n=16/group) were either administered IPS or not. The IPS significantly enhanced. • Cell adhesion and gutintegrity markers as well as immune response and gut remodeling markers. • The IPS significantly reduced PEDV shedding in vivo. [ABSTRACT FROM AUTHOR]

In this computational study, we advanced the understanding of the antigenic properties of the NADC-34-like isolate of the Porcine Reproductive and Respiratory Syndrome Virus (PRRSV), named YC-2020, relevant in veterinary pathology. We utilized sequence comparison analyses of the M and N proteins, comparing them with those of NADC34, identifying substantial amino acid homology that allowed us to highlight conserved epitopes and crucial variants. Through the application of Clustal Omega for multiple sequence alignment and platforms like Vaxijen and AllerTOP for predicting antigenic and allergenic potential, our analyses revealed important insights into the conservation and variation of epitopes essential for the development of effective diagnostic tools and vaccines. Our findings, aligned with initial experimental studies, underscore the importance of these epitopes in the development of targeted immunodiagnostic platforms and significantly contribute to the management and control of PRRSV. However, further studies are required to validate the computational predictions of antigenicity for this new viral isolate. This approach underscores the potential of computational models to enable ongoing monitoring and control of PRRSV evolution in swine. While this study provides valuable insights into the antigenic properties of the novel PRRSV isolate YC-2020 through computational analysis, it is important to acknowledge the limitations inherent to in silico predictions, specifically, the absence of laboratory validation. [ABSTRACT FROM AUTHOR]

Porcine reproductive and respiratory syndrome (PRRS), which has posed substantial threats to the swine industry worldwide, is primarily characterized by interstitial pneumonia. A disintegrin and metalloproteinase 17 (ADAM17) is a multifunctional sheddase involved in various inflammatory diseases. Herein, our study showed that PRRS virus (PRRSV) infection elevated ADAM17 activity, as demonstrated in primary porcine alveolar macrophages (PAMs), an immortalized PAM cell line (IPAM cells), and the lung tissues of PRRSV-infected piglets. We found that PRRSV infection promoted ADAM17 translocation from the endoplasmic reticulum to the Golgi by enhancing its interaction with inactive rhomboid protein 2 (iRhom2), a newly identified ADAM17 regulator, which in turn elevated ADAM17 activity. By screening for PRRSV-encoded structural proteins, viral envelope (E) and nucleocapsid (N) proteins were identified as the predominant ADAM17 activators. E and N proteins bind with both ADAM17 and iRhom2 to form ternary protein complexes, ultimately strengthening their interactions. Additionally, we demonstrated, using an ADAM17-knockout cell line, that ADAM17 augmented the shedding of soluble TNF-α, a pivotal inflammatory mediator. We also discovered that ADAM17-mediated cleavage of porcine TNF-α occurred between Arg-78 and Ser-79. By constructing a precision mutant cell line with Arg-78-Glu/Ser-79-Glu substitution mutations in TNF-α, we further revealed that the ADAM17-mediated production of soluble TNF-α contributed to the induction of inflammatory responses by PRRSV and its E and N proteins. Taken together, our results elucidate the mechanism by which PRRSV infection activates the iRhom2/ADAM17/TNF-α axis to enhance inflammatory responses, providing valuable insights into the elucidation of PRRSV pathogenesis. • PRRSV infection upregulates ADAM17 activity both in vivo and in vitro. • iRhom2 plays an important role in PRRSV infection-induced ADAM17 activation. • PRRSV E and N proteins are predominant ADAM17 activators. • ADAM17 cleaves porcine TNF-α at Arg-78/Ser-79. • iRhom2/ADAM17/TNF-α axis is involved in inflammation induced by E and N proteins. [ABSTRACT FROM AUTHOR]

Porcine reproductive and respiratory syndrome virus (PRRSV) has caused significant economic losses in the swine industry. In this study, the high-throughput sequencing, microRNAs (miRNAs) mimic, and lentivirus were used to screen for potential miRNAs that can promote PRRSV infection in porcine alveolar macrophages or Marc-145 cells. It was observed that novel-216, a previously unidentified miRNA, was upregulated through the p38 signaling pathway during PRRSV infection, and its overexpression significantly increased PRRSV replication. Further analysis revealed that novel-216 regulated PRRSV replication by directly targeting mitochondrial antiviral signaling protein (MAVS), an upstream molecule of type Ⅰ IFN that mediates the production and response of type Ⅰ IFN. The proviral function of novel-216 on PRRSV replication was abolished by MAVS overexpression, and this effect was reversed by the 3′UTR of MAVS, which served as the target site of novel-216. In conclusion, this study demonstrated that PRRSV-induced upregulation of novel-216 served to inhibit the production and response of typeⅠ IFN and facilitate viral replication, providing new insights into viral immune evasion and persistent infection. • Porcine reproductive and respiratory syndrome virus (PRRSV) is characterized by immunosuppression and persistent infection. • PRRSV upregulates novel-216 expression to facilitate self-replication. • Novel-216 targets MAVS to negatively regulates the antiviral activity of type Ⅰ IFN. [ABSTRACT FROM AUTHOR]

Porcine reproductive and respiratory syndrome virus (PRRSV) is a pathogen for swine, resulting in substantial economic losses to the swine industry. However, there has been little success in developing effective vaccines or drugs for PRRSV control. In the present study, we discovered that Diltiazem HCl, an inhibitor of L-type Ca2+ channel, effectively suppresses PRRSV replication in MARC-145, PK-15CD163 and PAM cells in dose-dependent manner. Furthermore, it demonstrates a broad-spectrum activity against both PRRSV-1 and PRRSV-2 strains. Additionally, we explored the underlying mechanisms and found that Diltiazem HCl -induced inhibition of PRRSV associated with regulation of calcium ion homeostasis in susceptible cells. Moreover, we evaluated the antiviral effects of Diltiazem HCl in PRRSV-challenged piglets, assessing rectal temperature, viremia, and gross and microscopic lung lesions. Our results indicate that Diltiazem HCl treatment alleviates PRRSV-induced rectal temperature spikes, pulmonary pathological changes, and serum viral load. In conclusion, our data suggest that Diltiazem HCl could serve as a novel therapeutic drug against PRRSV infection. [ABSTRACT FROM AUTHOR]

Group A Rotavirus (RVA) is a major cause of diarrhea in infants and piglets. β2-microglobulin (β2 M), encoded by the B2M gene, serves as a crucial subunit of the major histocompatibility complex class I (MHC-I) molecules. β2 M is indispensable for the transport of MHC-I to the cell membrane. MHC-I, also known as swine leukocyte antigen class I (SLA-I) in pigs, presents viral antigens to the cell surface. In this study, RVA infection down-regulated β2 M expression in both porcine intestinal epithelial cells-J2 (IPEC-J2) and MA-104 cells. RVA infection did not down-regulate the mRNA level of the B2M gene, indicating that the down-regulation of β2 M occurred on the protein level. Mechanismly, RVA infection triggered β2 M aggregation in the endoplasmic reticulum (ER) and enhanced the Lys48 (K48)-linked ubiquitination of β2 M, leading to the degradation of β2 M through ERAD-proteasome pathway. Furthermore, we found that RVA infection significantly impeded the level of SLA-I on the surface, and the overexpression of β2 M could recover its expression. In this study, our study demonstrated that RVA infection degrades β2 M via ERAD-proteasome pathway, consequently hampering SLA-I expression on the cell surface. This study would enhance the understanding of the mechanism of how RVA infection induces immune escape. • RVA infection degrades β2 M via ERAD-proteasome pathway. • RVA infection inhibits SLA-I expression on the cell surface. • RVA infection promotes the Lys48 (K48)-linked ubiquitination of β2 M. • RVA infection may inhibit host adapted immune response. [ABSTRACT FROM AUTHOR]