Fish gut-derived probiotic Pediococcus pentosaceus alleviates gossypol-induced intestinal inflammation by inhibiting NLRC3/NF-κB/IL-1ß signal pathway in Nile tilapia.

Cottonseed meal (CSM) and cottonseed protein concentrate (CPC) serve as protein alternatives to fish meal and soybean meal in the feed industry. However, the presence of gossypol residue in CSM and CPC can potentially trigger severe intestinal inflammation, thereby restricting the widespread utilization of these two protein sources. Probiotics are widely used to prevent or alleviate intestinal inflammation, but their efficacy in protecting fish against gossypol-induced enteritis remains uncertain. Here, the protective effect of Pediococcus pentosaceus, a strain isolated from the gut of Nile tilapia (Oreochromis niloticus), was evaluated. Three diets, control diet (CON), gossypol diet (GOS) and GOS supplemented with P. pentosaceus YC diet (GP), were used to feed Nile tilapia for 10 weeks. After the feeding trial, P. pentosaceus YC reduced the activity of myeloperoxidase (MPO) in the proximal intestine (PI) and distal intestine (DI). Following a 7-day exposure to Aeromonas hydrophila, the addition of P. pentosaceus YC was found to increase the survival rate of the fish. P. pentosaceus YC significantly inhibited the oxidative stress caused by gossypol, which was evidenced by lower reactive oxygen species (ROS) and malondialdehyde (MDA), as well as higher activities of glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) in PI and DI. Addition of P. pentosaceus YC significantly inhibited enteritis, with the lower expression of pro-inflammatory cytokines (il-1ß, il-6, il-8) and higher expression of anti-inflammatory cytokines tgf-ß. RNA-seq analysis indicated that P. pentosaceus YC supplementation significantly inhibited nlrc3 and promoted nf-κb expression in PI and DI, and the siRNA interference experiment in vivo demonstrated that intestinal inflammation was mediated by NLRC3/NF-κB/IL-1ß signaling pathway. Fecal bacteria transplantation experiment demonstrated that gut microbiota mediated the protective effect of P. pentosaceus YC. These findings offer valuable insights into the application of P. pentosaceus YC for alleviating gossypol-induced intestinal inflammation in fish.

Balancing immune responses: regulatory cells in eosinophilic gastrointestinal disorders.

Eosinophilic gastrointestinal disorders (EGIDs) are a group of conditions characterized by an abnormal accumulation of eosinophils in the gastrointestinal tract, leading to inflammation and tissue damage. Regulatory cells are a subset of immune cells that are crucial in maintaining the balance of the immune system and preventing the occurrence of autoimmune diseases. In EGIDs, regulatory cells are believed to play a key role in controlling the immune response and overseeing the growth and activation of eosinophils in the gastrointestinal tract. There is evidence indicating that regulatory T cells (Tregs) and regulatory eosinophils may play a role in suppressing the inflammatory response in EGIDs. Regulatory eosinophils are a subgroup of eosinophils that possess an anti-inflammatory role. Recent studies have shown that enhancing the number or effectiveness of regulatory eosinophils can reduce the severity of EGIDs. Regulatory eosinophils dampen inflammation through their regulatory mediators, such as galectin-10 and growth factor beta (TGF-ß), which promote Treg expansion and inhibit effector T cell function. Further research on regulatory cells in EGIDs may have significant implications for the advancement of novel therapies for these uncommon and intricate disorders. The aim of this review is to provide complete view of the immune responses connected to EGIDs, examine the regulatory cells that control these responses, and evaluate their potential as therapeutic targets for EGID treatment.

Genomic insights into pediatric intestinal inflammatory and eosinophilic disorders using single-cell RNA-sequencing.

Introduction: Chronic inflammation of the gastrointestinal tissues underlies gastrointestinal inflammatory disorders, leading to tissue damage and a constellation of painful and debilitating symptoms. These disorders include inflammatory bowel diseases (Crohn's disease and ulcerative colitis), and eosinophilic disorders (eosinophilic esophagitis and eosinophilic duodenitis). Gastrointestinal inflammatory disorders can often present with overlapping symptoms necessitating the use of invasive procedures to give an accurate diagnosis. Methods: This study used peripheral blood mononuclear cells from individuals with Crohn's disease, ulcerative colitis, eosinophilic esophagitis, and eosinophilic duodenitis to better understand the alterations to the transcriptome of individuals with these diseases and identify potential markers of active inflammation within the peripheral blood of patients that may be useful in diagnosis. Single-cell RNA-sequencing was performed on peripheral blood mononuclear cells isolated from the blood samples of pediatric patients diagnosed with gastrointestinal disorders, including Crohn's disease, ulcerative colitis, eosinophilic esophagitis, eosinophilic duodenitis, and controls with histologically healthy gastrointestinal tracts. Results: We identified 730 (FDR < 0.05) differentially expressed genes between individuals with gastrointestinal disorders and controls across eight immune cell types. Discussion: There were common patterns among GI disorders, such as the widespread upregulation of MTRNR2L8 across cell types, and many differentially expressed genes showed distinct patterns of dysregulation among the different gastrointestinal diseases compared to controls, including upregulation of XIST across cell types among individuals with ulcerative colitis and upregulation of Th2-associated genes in eosinophilic disorders. These findings indicate both overlapping and distinct alterations to the transcriptome of individuals with gastrointestinal disorders compared to controls, which provide insight as to which genes may be useful as markers for disease in the peripheral blood of patients.

Clostridium perfringens antigens and challenges for development of vaccines against necrotic enteritis in poultry.

INTRODUCTION: Chickens with Necrotic Enteritis (NE), caused by Clostridium perfringens, exhibit acute and chronic symptoms that are difficult to diagnose, leading to significant economic losses. Vaccination is the best method for controlling and preventing NE. However, only two vaccines based on the CPA and NetB toxins have been commercialized, offering partial protection, highlighting the urgent need for more effective vaccines. OBJECTIVE: This review aimed to identify promising antigens for NE vaccine formulation and discuss factors affecting their effectiveness. METHODS: A systematic review using five scientific databases identified 30 eligible studies through the Rayyan tool, which were included for quality review. RESULTS: We identified 25 promising antigens, including CPA, NetB, FBA, ZMP, CnaA, FimA, and FimB, categorized by their role in disease pathogenesis. This review discusses the biochemical, physiological, and genetic traits of recombinant antigens used in vaccine prototypes, their expression systems, and immunization potential in chickens challenged with virulent C. perfringens strains. Market supply challenges, immunogenic potential, vaccine platforms, adjuvants, and factors related to vaccination schedules-such as administration routes, dosing intervals, and age at immunization-are also addressed. Additionally, the study notes that vaccine formulations tested under mild challenges may not offer adequate field-level protection due to issues replicating aggressive conditions, strain virulence loss, and varied methodologies. CONCLUSIONS: An ideal NE vaccine should incorporate multiple antigens, molecular adjuvants, and delivery systems via in ovo and oral routes. The review underscores the challenges in developing and validating NE vaccines and the urgent need for a standardized protocol to replicate aggressive challenges accurately.

Effect of arginine supplementation on the growth performance, intestinal health, and immune responses of broilers during necrotic enteritis challenge.

The objective of this study was to evaluate the effect of 25% arginine supplementation as a functional amino acid in partially alleviating the detrimental effects of necrotic enteritis (NE) on the growth performance, serum biochemistry, gut integrity, and the relative gene expression of tight junction proteins and inflammatory cytokines in broilers during NE. Three hundred and sixty 1-day-old chicks were randomly allocated to 4 treatments in a 2 × 2 factorial arrangement -basal diet and 125% arginine diet, with or without NE challenge. NE was induced by inoculating 1 × 104Eimeria maxima sporulated oocysts on d 14 and 1 × 108 CFU/bird C. perfringens on d 19, 20, and 21. The NE challenge had a significant effect on the BWG (p < 0.05), FCR (p < 0.05), serum AST (p < 0.05), GLU (p < 0.05), and K+ (p < 0.05) levels, and intestinal permeability (p < 0.05) and jejunal lesion score (p < 0.05). A significant challenge × diet interaction effect was observed in the cecal tonsil CD8+: CD4+ T-cell ratio on d 21 (p < 0.05) and 28 (p < 0.05) and spleen CD8+: CD4+ T-cell ratio on d 21 (p < 0.05) and 35 (p < 0.05). Arginine supplementation significantly increased the CD8+: CD4+ T-cell ratio in uninfected birds but decreased the CD8+: CD4+ T-cell ratio in infected birds. On d 21, a significant interaction effect was observed on the relative expression of the iNOS gene (p < 0.05). Arginine supplementation significantly downregulated the expression of the iNOS gene in infected birds. A significant effect of the challenge (p < 0.05) was observed on the relative gene expression of the ZO-1 gene in the jejunum. NE challenge significantly downregulated the expression of the ZO-1 gene on d 21. In conclusion, arginine supplementation did not alleviate the depression in growth performance and disease severity during the NE challenge. However, arginine downregulated the expression of inflammatory cytokines and enzymes, preventing inflammatory injury to the tissues during NE. Hence, arginine might be supplemented with other alternatives to downregulate inflammatory response during NE in poultry.

Effect of 125% and 135% arginine on the growth performance, intestinal health, and immune responses of broilers during necrotic enteritis challenge.

The objective of this study was to evaluate the effects of 25% and 35% arginine supplementation in partially alleviating the effects of necrotic enteritis (NE) challenge on the production performance, intestinal integrity, and relative gene expression of tight junction proteins and inflammatory cytokines in broilers. Four hundred and eighty 1-day-old chicks were randomly allocated to the 4 treatments- Uninfected + Basal, NE + Basal, NE + Arg 125%, and NE + Arg 135%. NE was induced by inoculating 1 × 104Eimeria maxima sporulated oocysts on d 14 and 1 × 108 CFU/bird C. perfringens on d 19, 20, and 21 of age by oral gavage. The NE challenge significantly decreased body weight gain (BWG) (p < 0.05) and increased the feed conversion ratio (FCR) (p < 0.05). On d 21, the NE challenge also increased the jejunal lesion score (p < 0.05) and relative gene expression of IL-10 and decreased the expression of the tight junction proteins occludin (p < 0.05) and claudin-4 (p < 0.05). The 125% arginine diet significantly increased intestinal permeability (p < 0.05) and the relative gene expression of iNOS (p < 0.05) and IFN-γ (p < 0.05) on d 21 and the bile anti-C. perfringens IgA concentration by 39.74% (p < 0.05) on d 28. The 135% arginine diet significantly increased the feed intake during d 0 - 28 (p < 0.05) and 0 to 35 (p < 0.05) and increased the FCR on d 0 to 35 (p < 0.05). The 135% and 125% arginine diet increased the spleen CD8+: CD4+ T-cell ratio on d 28 (p < 0.05) and 35 (p < 0.05), respectively. The 135% arginine diet increased the CT CD8+:CD4+ T-cell ratio on d 35 (p < 0.05). In conclusion, the 125% and 135% arginine diets did not reverse the effect of the NE challenge on the growth performance. However, the 125% arginine diet significantly increased the cellular and humoral immune response to the challenge. Hence, the 125% arginine diet could be used with other feed additives to improve the immune response of the broilers during the NE challenge.

Supplementation with organic yeast-derived selenium provides immune protection against experimental necrotic enteritis in broiler chickens.

Necrotic enteritis (NE) is a potentially fatal poultry disease that causes enormous economic losses in the poultry industry worldwide. The study aimed to evaluate the effects of dietary organic yeast-derived selenium (Se) on immune protection against experimental necrotic enteritis (NE) in commercial broilers. Chickens were fed basal diets supplemented with different Se levels (0.25, 0.50, and 1.00 Se mg/kg). To induce NE, Clostridium perfringens (C. perfringens) was orally administered at 14 days of age post hatch. The results showed that birds fed 0.25 Se mg/kg exhibited significantly increased body weight gain compared with the non-supplemented/infected birds. There were no significant differences in gut lesions between the Se-supplemented groups and the non-supplemented group. The antibody levels against α-toxin and NetB toxin increased with the increase between 0.25 Se mg/kg and 0.50 Se mg/kg. In the jejunal scrapings and spleen, the Se-supplementation groups up-regulated the transcripts for pro-inflammatory cytokines IL-1ß, IL-6, IL-8, iNOS, and LITAF and avian ß-defensin 6, 8, and 13 (AvBD6, 8 and 13). In conclusion, supplementation with organic yeast-derived Se alleviates the negative consequences and provides beneficial protection against experimental NE.

Exploring the underlying mechanisms of enteritis impact on golden pompano (Trachinotus ovatus) through multi-omics analysis.

Enteritis posed a significant health challenge to golden pompano (Trachinotus ovatus) populations. In this research, a comprehensive multi-omics strategy was implemented to elucidate the pathogenesis of enteritis by comparing both healthy and affected golden pompano. Histologically, enteritis was characterized by villi adhesion and increased clustering after inflammation. Analysis of the intestinal microbiota revealed a significant increase (P < 0.05) in the abundance of specific bacterial strains, including Photobacterium and Salinivibrio, in diseased fish compared to the healthy group. Metabolomic analysis identified 5479 altered metabolites, with significant impacts on terpenoid and polyketide metabolism, as well as lipid metabolism (P < 0.05). Additionally, the concentrations of several compounds such as calcitetrol, vitamin D2, arachidonic acid, and linoleic acid were significantly reduced in the intestines of diseased fish post-enteritis (P < 0.05), with the detection of harmful substances such as Efonidipine. In transcriptomic profiling, enteritis induced 68 upregulated and 73 downregulated genes, predominantly affecting steroid hormone receptor activity (P < 0.05). KEGG pathway enrichment analysis highlighted upregulation of SQLE and CYP51 in steroidogenesis, while the HSV-1 associated MHC1 gene exhibited significant downregulation. Integration of multi-omics results suggested a potential pathogenic mechanism: enteritis may have resulted from concurrent infection of harmful bacteria, specifically Photobacterium and Salinivibrio, along with HSV-1. Efonidipine production within the intestinal tract may have blocked certain calcium ion channels, leading to downregulation of MHC1 gene expression and reduced extracellular immune recognition. Upregulation of SQLE and CYP51 genes stimulated steroid hormone synthesis within cells, which, upon binding to G protein-coupled receptors, influenced calcium ion transport, inhibited immune activation reactions, and further reduced intracellular synthesis of anti-inflammatory substances like arachidonic acid. Ultimately, this cascade led to inflammation progression, weakened intestinal peristalsis, and villi adhesion. This study utilized multi-level omics detection to investigate the pathological symptoms of enteritis and proposed a plausible pathogenic mechanism, providing innovative insights into enteritis verification and treatment in offshore cage culture of golden pompano.

Intestinal microbiota and gene expression alterations in leopard coral grouper (Plectropomus leopardus) under enteritis.

Enteritis poses a significant threat to fish farming, characterized by symptoms of intestinal and hepatic inflammation, physiological dysfunction, and dysbiosis. Focused on the leopard coral grouper (Plectropomus leopardus) with an enteritis outbreak on a South China Sea farm, our prior scrutiny did not find any abnormalities in feeding or conventional water quality factors, nor were any specific pathogen infections related to enteritis identified. This study further elucidates their intestinal flora alterations, host responses, and their interactions to uncover the underlying pathogenetic mechanisms and facilitate effective prevention and management strategies. Enteritis-affected fish exhibited substantial differences in intestinal flora compared to control fish (P = 0.001). Notably, norank_f_Alcaligenaceae, which has a negative impact on fish health, predominated in enteritis-affected fish (91.76 %), while the probiotic genus Lactococcus dominated in controls (93.90 %). Additionally, certain genera with pathogenesis potentials like Achromobacter, Sphingomonas, and Streptococcus were more abundant in diseased fish, whereas Enterococcus and Clostridium_sensu_stricto with probiotic potentials were enriched in control fish. At the transcriptomic level, strong inflammatory responses, accompanied by impaired metabolic functions, tissue damage, and iron death signaling activation were observed in the intestines and liver during enteritis. Furthermore, correlation analysis highlighted that potential pathogen groups were positively associated with inflammation and tissue damage genes while presenting negatively correlated with metabolic function-related genes. In conclusion, dysbiosis in the intestinal microbiome, particularly an aberrantly high abundance of Alcaligenaceae with pathogenic potential may be the main trigger for this enteritis outbreak. Alcaligenaceae alongside Achromobacter, Sphingomonas, and Streptococcus emerged as biomarkers for enteritis, whereas some species of Lactococcus, Clostridium_sensu_stricto, and Enterococcus showed promise as probiotics to alleviate enteritis symptoms. These findings enhance our understanding of enteritis pathogenesis, highlight intestinal microbiota shifts in leopard coral grouper, and propose biomarkers for monitoring, probiotic selection, and enteritis management.

Extracellular adenosine triphosphate skews the T helper cell balance and enhances neutrophil activation in mice with food allergies.

Exposure to food allergens elicits fast changes in the intestinal microenvironment, which guides the development of allergic reactions. Investigating the key information about these changes may help in better understanding food allergies. In this research, we explored the relationship between a food allergy and extracellular adenosine triphosphate (ATP), a danger molecule that has been proved to regulate the onset of allergic asthma and dermatitis but has not been studied in food allergies, by developing a unique animal model through allergen-containing diet feeding. After consuming an allergen-containing diet for 7 days, the allergic mice exhibited severe enteritis with elevated luminal ATP levels. The dysregulated luminal ATP worsened food-induced enteritis by enhancing Th17 cell responses and increasing mucosal neutrophil accumulation. In vitro experiments demonstrated that ATP intervention facilitated Th17 cell differentiation and neutrophil activation. In addition, the diet-induced allergy showed noticeable gut dysbiosis, characterized by decreased microbial diversity and increased diet-specific microbiota signatures. As the first, we show that food-induced enteritis is associated with an elevated concentration of luminal ATP. The dysregulated extracellular ATP exacerbated the enteritis of mice to a food challenge by manipulating intestinal Th17 cells and neutrophils.

LTA and PGN from Bacillus siamensis can alleviate soybean meal-induced enteritis and microbiota dysbiosis in Lateolabrax maculatus.

An eight-week feeding trial was designed to assess which component of commensal Bacillus siamensis LF4 can mitigate SBM-induced enteritis and microbiota dysbiosis in spotted seabass (Lateolabrax maculatus) based on TLRs-MAPKs/NF-кB signaling pathways. Fish continuously fed low SBM (containing 16 % SBM) and high SBM (containing 40 % SBM) diets were used as positive (FM group) and negative (SBM group) control, respectively. After feeding high SBM diet for 28 days, fish were supplemented with B. siamensis LF4-derived whole cell wall (CW), cell wall protein (CWP), lipoteichoic acid (LTA) or peptidoglycan (PGN) until 56 days. The results showed that a high inclusion of SBM in the diet caused enteritis, characterized with significantly (P < 0.05) decreased muscular thickness, villus height, villus width, atrophied and loosely arranged microvillus. Moreover, high SBM inclusion induced an up-regulation of pro-inflammatory cytokines and a down-regulation of occludin, E-cadherin, anti-inflammatory cytokines, apoptosis related genes and antimicrobial peptides. However, dietary supplementation with CW, LTA, and PGN of B. siamensis LF4 could effectively alleviate enteritis caused by a high level of dietary SBM. Additionally, CWP and PGN administration increased beneficial Cetobacterium and decreased pathogenic Plesiomonas and Brevinema, while dietary LTA decreased Plesiomonas and Brevinema, suggesting that CWP, LTA and PGN positively modulated intestinal microbiota in spotted seabass. Furthermore, CW, LTA, and PGN application significantly stimulated TLR2, TLR5 and MyD88 expressions, and inhibited the downstream p38 and NF-κB signaling. Taken together, these results suggest that LTA and PGN from B. siamensis LF4 could alleviate soybean meal-induced enteritis and microbiota dysbiosis in L. maculatus, and p38 MAPK/NF-κB pathways might be involved in those processes.

Minimally Invasive Approaches to Diagnose and Monitor Eosinophilic GI Diseases.

PURPOSE OF REVIEW: This review seeks to understand novel avenues for eosinophilic GI disease management. Biomarkers offer a unique and non-invasive approach to tracking EoE disease progression. While no biomarkers have definitively met the diagnostic criteria for eosinophilic GI diseases, some biomarkers have been shown to be associated with disease activity. Here, we examine the potential of recently studied biomarkers. RECENT FINDINGS: Current research shows advancements in blood, luminal fluid, and breath testing. Particular areas of interest include mRNA analyses, protein fingerprinting, amplicon sequence variants (ASVs), T cells and IgE receptors, eosinophilic cationic proteins, cytokines, and nitric oxide exhalation. Preliminary results showed that mucosal biomarkers, directly captured from the esophagus, may reflect the best representation of biopsy-based results, in contrast to biomarkers obtained from indirect or peripheral (blood, breath) methods. However, this is based on limited clinical studies without sufficient numbers to evaluate true diagnostic accuracy. Large-scale randomized trials are needed to fully ascertain both the optimal sampling technique and the specific biomarkers that reflect diagnostic status of the disease.

Common and disparate clinical presentations and mechanisms in different eosinophilic gastrointestinal diseases.

Eosinophilic gastrointestinal diseases (EGIDs) are a group of diseases characterized by selective eosinophil infiltration of the gastrointestinal (GI) tract in the absence of other causes of eosinophilia. These diseases are generally driven by type 2 inflammation, often in response to food allergen exposure. Among all EGIDs, the clinical presentation often includes a history of atopic disease with a variety of GI symptoms. EGIDs are traditionally separated into eosinophilic esophagitis (EoE) and non-EoE EGIDs. EoE is relatively better understood and now associated with clinical guidelines and 2 US Food and Drug Administration-approved treatments, whereas non-EoE EGIDs are rarer and less well-understood diseases without US Food and Drug Administration-approved treatments. Non-EoE EGIDs are further subclassified by the area of the GI tract that is involved; they comprise eosinophilic gastritis, eosinophilic enteritis (including eosinophilic duodenitis), and eosinophilic colitis. As with other GI disorders, the disease presentations and mechanisms differ depending on the involved segment of the GI tract; however, the differences between EoE and non-EoE EGIDs extend beyond which GI tract segment is involved. The aim of this article is to summarize the commonalities and differences between the clinical presentations and disease mechanisms for EoE and non-EoE EGIDs.

Treatment of chickens with lactobacilli prior to challenge with Clostridium perfringens modifies innate responses and gut morphology.

Necrotic enteritis caused by Clostridium perfringens (CP), is a common enteric disease of poultry that has been previously controlled by in-feed antibiotics. However, due to the rapid emergence of antimicrobial resistance, alternatives to antibiotics such as probiotics have received considerable attention because of their immunomodulatory and intestinal health benefits. The present study investigated the effects of probiotic lactobacilli on gut histomorphology and intestinal innate responses in chickens. Day-old male broiler chickens were treated with 1 × 107 or 1 × 108 colony-forming units (CFU) of a lactobacilli cocktail on days 1, 7, 14, and 20 post-hatch, while control groups were not treated with lactobacilli. On day 21, birds in all groups (except the negative control) were challenged with 3 × 108 CFU of CP for 3 days. Intestinal tissue samples were collected before and after the CP challenge to assess gene expression and for histomorphological analysis. Lactobacilli treatment at a dose of 1 × 108 CFU conferred partial protection against NE by lowering lesion scores, increasing villus height in the ileum and reducing crypt depth in the jejunum. In addition, 1 × 108 CFU of lactobacilli enhanced the expression of Toll-like receptor (TLR) 2, interferon-gamma (IFN-γ), interleukin (IL)-10, IL-12, and IL-13 in both the jejunum and ileum at different timepoints and subsequently decreased the expression of transforming growth factor beta (TGF-ß) and IL-1ß post-CP challenge. In conclusion, the results indicate that treatment with lactobacilli mitigated NE in a dose-dependent manner via improvement of intestinal morphology and modulation of innate immune response in chickens.

Severe Immune-Related Enteritis after In Utero Exposure to Pembrolizumab.

Immune checkpoint blockade has become standard treatment for many types of cancer. Such therapy is indicated most often in patients with advanced or metastatic disease but has been increasingly used as adjuvant therapy in those with early-stage disease. Adverse events include immune-related organ inflammation resembling autoimmune diseases. We describe a case of severe immune-related gastroenterocolitis in a 4-month-old infant who presented with intractable diarrhea and failure to thrive after in utero exposure to pembrolizumab. Known causes of the symptoms were ruled out, and the diagnosis of pembrolizumab-induced immune-related gastroenterocolitis was supported by the results of histopathological assays, immunophenotyping, and analysis of the level of antibodies against programmed cell death protein 1 (PD-1). The infant's condition was successfully treated with prednisolone and infliximab.

A Recombinant Duck Plague Virus Containing the ICP27 Deletion Marker Provides Robust Protection in Ducks.

Duck plague virus (DPV) is a member of Alphaherpesvirus genus and poses a major threat to waterfowl breeding. Genetic engineered vaccines that are capable of distinguishing naturally infected from vaccine-immunized animals are useful for eradicating duck plague. In this study, reverse genetics was used to develop an ICP27-deficient strain (CHv-ΔICP27), and its potential as a marker vaccination candidate was evaluated. The results showed that the CHv-ΔICP27 generated in this study exhibited good genetic stability in vitro and was highly attenuated both in vivo and in vitro. The level of neutralizing antibody generated by CHv-ΔICP27 was comparable to that induced by a commercial DPV vaccine, suggesting that it could protect ducks from virulent DPV attack. By using molecular identification techniques such as PCR, restriction fragment length polymorphism, immunofluorescence, Western blotting, and others, it is possible to differentiate the CHv-ΔICP27 from wild-type strains. Moreover, ICP27 can also be a potential target for the genetic engineering vaccine development of alphavirus or perhaps the entire herpesvirus family members due to the highly conservative of ICP27 protein in all herpesvirus family members. IMPORTANCE The development of distinguishable marker vaccines from natural infection is a key step toward eradicating duck plague. Here, we generated a recombinant DPV that carries an ICP27 deletion marker that could be easily distinguished from wild-type strain by molecular biological methods. It was highly attenuated in vitro and in vivo and could provide comparable protection to ducks after a single dose of immunizations, as commercial vaccines did. Our findings support the use of the ICP27-deficient virus as a marker vaccine for DPV control and future eradication.

Duck Enteritis Virus Inhibits the cGAS-STING DNA-Sensing Pathway To Evade the Innate Immune Response.

Cyclic GMP-AMP synthase (cGAS), a key DNA sensor, detects cytosolic viral DNA and activates the adaptor protein stimulator of interferon genes (STING) to initiate interferon (IFN) production and host innate antiviral responses. Duck enteritis virus (DEV) is a duck alphaherpesvirus that causes an acute and contagious disease with high mortality in waterfowl. In the present study, we found that DEV inhibits host innate immune responses during the late phase of viral infection. Furthermore, we screened DEV proteins for their ability to inhibit the cGAS-STING DNA-sensing pathway and identified multiple viral proteins, including UL41, US3, UL28, UL53, and UL24, which block IFN-ß activation through this pathway. The DEV tegument protein UL41, which exhibited the strongest inhibitory effect, selectively downregulated the expression of interferon regulatory factor 7 (IRF7) by reducing its mRNA accumulation, thereby inhibiting the DNA-sensing pathway. Ectopic expression of UL41 markedly reduced viral DNA-triggered IFN-ß production and promoted viral replication, whereas deficiency of UL41 in the context of DEV infection increased the IFN-ß response to DEV and suppressed viral replication. In addition, ectopic expression of IRF7 inhibited the replication of the UL41-deficient virus, whereas IRF7 knockdown facilitated its replication. This study is the first report identifying multiple viral proteins encoded by a duck DNA virus, which inhibit the cGAS-STING DNA-sensing pathway. These findings expand our knowledge of DNA sensing in ducks and reveal a mechanism through which DEV antagonizes the host innate immune response. IMPORTANCE Duck enteritis virus (DEV) is a duck alphaherpesvirus that causes an acute and contagious disease with high mortality, resulting in substantial economic losses in the commercial waterfowl industry. The evasion of DNA-sensing pathway-mediated antiviral innate immunity is essential for the persistent infection and replication of many DNA viruses. However, the mechanisms used by DEV to modulate the DNA-sensing pathway remain poorly understood. In the present study, we found that DEV encodes multiple viral proteins to inhibit the cGAS-STING DNA-sensing pathway. The DEV tegument protein UL41 selectively diminished the accumulation of interferon regulatory factor 7 (IRF7) mRNA, thereby inhibiting the DNA-sensing pathway. Loss of UL41 potently enhanced the IFN-ß response to DEV and impaired viral replication in ducks. These findings provide insights into the host-virus interaction during DEV infection and help develop new live attenuated vaccines against DEV.

Respiratory Influenza Virus Infection Causes Dynamic Tuft Cell and Innate Lymphoid Cell Changes in the Small Intestine.

Influenza A viruses (IAV) can cause severe disease and death in humans. IAV infection and the accompanying immune response can result in systemic inflammation, leading to intestinal damage and disruption of the intestinal microbiome. Here, we demonstrate that a specific subset of epithelial cells, tuft cells, increase across the small intestine during active respiratory IAV infection. Upon viral clearance, tuft cell numbers return to baseline levels. Intestinal tuft cell increases were not protective against disease, as animals with either increased tuft cells or a lack of tuft cells did not have any change in disease morbidity after infection. Respiratory IAV infection also caused transient increases in type 1 and 2 innate lymphoid cells (ILC1 and ILC2, respectively) in the small intestine. ILC2 increases were significantly blunted in the absence of tuft cells, whereas ILC1s were unaffected. Unlike the intestines, ILCs in the lungs were not altered in the absence of tuft cells. This work establishes that respiratory IAV infection causes dynamic changes to tuft cells and ILCs in the small intestines and that tuft cells are necessary for the infection-induced increase in small intestine ILC2s. These intestinal changes in tuft cell and ILC populations may represent unexplored mechanisms preventing systemic infection and/or contributing to severe disease in humans with preexisting conditions. IMPORTANCE Influenza A virus (IAV) is a respiratory infection in humans that can lead to a wide range of symptoms and disease severity. Respiratory infection can cause systemic inflammation and damage in the intestines. Few studies have explored how inflammation alters the intestinal environment. We found that active infection caused an increase in the epithelial population called tuft cells as well as type 1 and 2 innate lymphoid cells (ILCs) in the small intestine. In the absence of tuft cells, this increase in type 2 ILCs was seriously blunted, whereas type 1 ILCs still increased. These findings indicate that tuft cells are necessary for infection-induced changes in small intestine type 2 ILCs and implicate tuft cells as regulators of the intestinal environment in response to systemic inflammation.