Brevibacillus laterosporus (B. laterosporus) is a significant probiotic bacterium known for producing multiple secondary metabolites with notable antibacterial activity. In this study, bioinformatic tools such as FastANI and Roary were employed to analyze genomic similarity and construct the B. laterosporus pan-genome. The software platforms antiSMASH and BAGEL4 were used to predict biosynthetic gene clusters (BGCs) and identify bacteriocins. Core candidate bacteriocins were subsequently subjected to heterologous expression in E. coli, followed by nickel column purification and concentration. Their antibacterial activity was systematically evaluated using agar diffusion assays, minimum inhibitory concentration (MIC) determinations, time-kill curves, scanning electron microscopy (SEM), and live/dead bacterial staining in Listeria monocytogenes (L. monocytogenes). Pan-genome analysis revealed an open genome for B. laterosporus, comprising 2,840 core genes. KEGG enrichment analyses indicated that these core genes were significantly associated with pathways responsible for the biosynthesis of antimicrobial compounds, including secondary metabolite, pan-quinone, and terpenoid quinone biosynthesis. AntiSMASH predicted 1,654 BGCs, predominantly non-ribosomal peptide synthase (NRPS), polyketide synthase (PKS), and heterocyclic amine-associated antimicrobial protein synthesis clusters. Moreover, BAGEL4 predicted three putative core candidate bacteriocin variants (BreL1, BreL2, and BreL3), all of which exhibited pronounced antibacterial activity against L. monocytogenes. Among these, BreL1 demonstrated the most robust activity, causing morphological changes consistent with membrane-associated damage. Collectively, this study elucidates the open pan-genome architecture and the metabolically rich secondary metabolome of B. laterosporus, identifies three putative bacteriocin variants with active antibacterial efficacy against L. monocytogenes, which provides a theoretical foundation for developing natural antimicrobial agents.

Salmonella Typhimurium (S. Typhimurium) is a major foodborne Gram-negative pathogen, presenting significant health risks to both humans and animals. LuxR family regulatory factors are critically involved in bacterial adaption to environmental changes and virulence regulation. Nevertheless, the biological function of YjjQ, a member of this family, remains uncharacterized in S. Typhimurium. To clarify the regulatory function of YjjQ in S. Typhimurium, λ-Red homologous recombination was employed to construct the yjjQ gene deletion strain (SL1344-ΔyjjQ) and its complementation strain (SL1344-ΔyjjQ/comp). Then, the growth characteristics, biofilm formation, motility, cell adhesion and invasion, intracellular survival and proliferation, as well as virulence, were systematically analyzed, respectively. Furthermore, the expression profiles of those genes associated with biofilm, flagella, and virulence were evaluated, and the interaction between YjjQ and predicted target promoter was experimentally validated. Compared to SL1344 and SL1344-ΔyjjQ/comp, SL1344-ΔyjjQ exhibited notably enhanced biofilm formation, motility, cell invasion, and intracellular proliferation under hyperosmotic conditions, in addition to heightened virulence in mice. RT-qPCR analysis revealed these genes related to flagellar (flhD, fliA), biofilm (csgD, fliC), and SPI-1 (invF) were significantly upregulated. The EMSA confirmed that YjjQ protein directly binds the flhDC promoter region, suppressing transcription of the flhDC operon, thereby negatively regulating flagellar, biofilm, and virulence. These findings provide new insights into the molecular mechanisms underlying the coordinately modulated strategy of biofilm formation, flagellar biosynthesis and virulence in S. Typhimurium.

Mycoplasma bovis is a major etiological agent of bovine respiratory disease, causing substantial economic losses to the cattle industry. To address this challenge, we performed a pan-genome analysis of 80 globally sourced M. bovis genomes and identified 1058 core genes. Reverse vaccinology and immunoinformatics approaches were subsequently integrated to screen three potential vaccine candidates: namely lipoate protein ligase 2 (LplA), elongation factor 4 (LepA) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Codon-optimized gene sequences were cloned into the pET-28a vector and successfully expressed and purified using E. coli expression system. Thirty female BALB/c mice were randomized into five groups (n = 6 per group). For primary immunization, each mouse received 100 μg of recombinant protein emulsified in Freund's complete adjuvant via subcutaneous injection, followed by booster immunizations with 50 μg of protein formulated with Freund's incomplete adjuvant at weeks 2 and 4. Fourteen days after primary immunization, all vaccinated groups exhibited significantly elevated antigen-specific antibody levels, which remained high through week 6. Notably, rLplA and rGAPDH induced higher antibody titers than rLepA. Analysis of the IgG2a/IgG1 subtype ratio indicated that rLplA, rGAPDH, and rLepA predominantly elicited Th2-type immune responses. In addition, all three recombinant proteins significantly increased cytokine production. Moreover, no adverse physiological reactions or histopathological abnormalities were observed throughout the immunization period, confirming the favorable safety profile of the candidates. Collectively, these findings identify promising targets for the development of M. bovis subunit vaccines to address pathogen variability, antibiotic resistance, and the limited efficacy of current vaccines.

This study aimed to identify and characterize a high-efficiency chitinase gene from Brevibacillus laterosporus (B. laterosporus), characterize its enzymatic traits, and assess its degrading activity against Caenorhabditis elegans (C. elegans) and Parascaris equorum (P. equorum) eggs. A nematicidal B. laterosporus isolate was subjected to whole-genome sequencing for chitinase gene screening, cloning, and molecular identification. The target gene was heterologously expressed in Escherichia coli BL21 (DE3), and biochemical properties including temperature, pH, metal ions, and substrate conditions were investigated, its kinetic parameters were determined, and its biological effects on C. elegans and P. equorum eggs were analyzed. The protein BLChi79 (79.3 kDa) belongs to GH18 chitinases with a typical carbohydrate-binding module. Its optimum activity occurred at 60 °C and pH 6.0, with colloidal chitin as the optimal substrate. Mg2+, Fe2+, and Mn2+ boosted its activity, while K+, Cu2+, Zn2+, and Ca2+ suppressed catalysis. Its Km was 6.14 mg·mL-1, Vmax 7.78 μmol·min-1·mg-1, and kcat 10.16 min-1. The purified recombinant enzyme could degradethe C. elegans chitin layer and the P. equorum egg vitelline envelope. In summary, the BLChi79 derived from B. laterosporus targets chitin-enriched eggshells, acting as a green biocatalyst for controlling livestock gastrointestinal nematodes.

Listeria monocytogenes (L. monocytogenes) is a zoonotic foodborne pathogen that triggers life-threatening invasive illnesses. MarR-type transcriptional regulators widely control bacterial stress adaptation, antibiotic tolerance and virulence, playing crucial roles in the pathogenesis of diverse bacterial pathogens. Lmo0840 is a conserved member of this family; however, its physiological functions and regulatory networks in L. monocytogenes remain completely unexplored. To this end, we constructed an lmo0840 deletion mutant and its complemented strain in the L. monocytogenes EGD-e background and systematically characterized the biological functions of Lmo0840 using phenotypic assays, transcriptomic profiling, and electrophoretic mobility shift assays (EMSA). Phenotypic characterization revealed that loss of lmo0840 impaired growth rate under hyperosmotic, oxidative, and iron-limitation stresses, while unexpectedly promoting growth rate at 30 °C and under tetracycline stress. Consistently, Disk diffusion assays further showed that the Δlmo0840 mutant exhibited significantly reduced inhibition zone diameters against tetracycline antibiotics compared to the EGD-e strain. Moreover, the Δlmo0840 mutant exhibited reduced early biofilm formation and attenuated macrophage adhesion, yet showed enhanced cellular invasion and decreased virulence in a murine model. EMSA further demonstrated that Lmo0840 directly binds to the promoter region of lmo0839 and represses its transcription, thereby contributing to bacterial adaptation to tetracycline stress. Collectively, these findings establish Lmo0840 as a pleiotropic regulator involved in environmental stress responses, tetracycline adaptation, and virulence in L. monocytogenes. Notably, this study identifies for the first time an Lmo0840–Lmo0839 regulatory cascade underlying tetracycline adaptation. These insights not only deepen our understanding of the molecular mechanisms governing Lm environmental adaptation and pathogenicity but also provide a potential target for the prevention and control of listeriosis.

The antibacterial activities of numerous medium and long-chain unsaturated fatty acids are well documented. However, the natural short-chain unsaturated fatty acid, 4-pentenoic acid (C5:1 Δ4) has not been systematically evaluated for its antibacterial activity, mechanism of action, or application potential. This study investigated these to provide a basis for novel preservatives. 4-Pentenoic acid showed broad-spectrum activity against 21 strains of pathogenic bacteria, with MICs 1.5-3 mg/mL, MBCs 3-6 mg/mL, and low resistance risk. It disrupted bacterial membrane integrity, inhibited biofilm formation, and regulated the pdu, nar and RND efflux pump genes, thereby affecting bacterial metabolism and stress responses. Checkerboard broth microdilution susceptibility assays confirmed that the combination of 4-pentenoic acid with antibiotics (such as meropenem) exerted additive antibacterial effects. At effective antibacterial concentrations, oral administration of 4-pentenoic acid caused no observable toxicity. It can markedly inhibit the proliferation of pathogenic bacteria in milk, beef, and peanut butter, while dosages close to the minimum inhibitory concentration exerted minimal adverse effects on the sensory properties and pH of these foods. In summary, this study provides theoretical and experimental basis for the development of new food preservatives and antibacterial agents.

Helminths can manipulate their host’s gut microbiota, with the expansion of the lactobacilli population being a common feature. This process profoundly influences host immunoregulation, yet the underlying mechanisms remain almost unknown. Using a tissue-dwelling helminth model (larval Echinococcus multilocularis) while validating key findings from other helminth infections, we show that helminths harness the antibacterial program of host innate immunity to transform the host gut microbiome and control gut microbiota-mediated immunity. Using multifaceted techniques, we elucidate that cathelicidin-related antimicrobial peptide (CRAMP), derived from the expanded CD11b+CD206+ macrophages rather than the intestinal epithelial cells, is the key component that enters into the gut ecological system and enhances the fitness of Lactobacillus by selectively killing gram-negative microbes like enterobacteria. Furthermore, through in vitro cell culturing and in vivo dietary intervention experiments, we demonstrate that this regulation from innate immunity is boosted via toll-like receptor signaling by helminth’s secretory products, which could be sufficiently tuned down by dietary vitamin D through its receptor and cyp27b1. Importantly, using microbiota-targeted treatment methods, we prove that this signaling bolsters gut microbiota-mediated host intestinal Foxp3+ Treg cell expansion and parasite survival and that therapies targeting this signaling are effective in treating infection. We outline a dietary micronutrient-dependent mechanism by which helminths leverage host innate immunity to edit the host gut microbiome and thereby control immunosuppression precisely.

Salmonella Typhimurium (S. typhimurium) is a significant zoonotic pathogen responsible for gastroenteritis and severe systemic infections in various hosts. The AraC family transcription factors are key gene expression regulators in prokaryotes, essential for bacterial adaptation to the environment and virulence. Despite their importance, the role of yeaM, a member of this family in S. typhimurium, remains unexplored. To elucidate yeaM regulatory function in virulence and biofilm formation, we engineered mutant and complementary strains of the yeaM gene using homologous recombination. We assessed their capabilities in biofilm formation under different conditions, macrophage adherence and invasion, and virulence in mice. Additionally, we identified potential target genes regulated by yeaM through transcriptome sequencing and confirmed these findings using an electrophoretic mobility shift assay (EMSA) and a dual-luciferase reporter assay. Our results demonstrate that, compared to the parental strain SL1344 and the complemented strain C Delta yeaM, the Delta yeaM strain exhibited significantly enhanced biofilm formation, increased invasion of mouse intestinal epithelial cells, enhanced intracellular proliferation within macrophages, and elevated induction of macrophage apoptosis. Furthermore, the Delta yeaM deletion strain displayed significantly increased virulence in mice and enhanced proliferation in milk. Transcriptome analysis revealed that S. typhimurium pathogenicity island 4 (SPI4) genes (siiA, siiB, siiC, siiD, siiF, and siiE) were significantly upregulated following the deletion of the yeaM gene. EMSA and dual-luciferase reporter assays further showed that the yeaM protein can bind to the promoter of the siiA gene and suppress its expression, thereby modulating the biofilm formation and virulence of S. typhimurium.

Introduction:Nematode-trapping fungi (NTFs) can produce various chitinases to degrade nematode body wall and eggshell chitin during predation. However, the regulatory mechanisms of their expression of chitinases still remain unclear. The primary objective of this study was to elucidate the differential protein profile of A. oligospora, an NTF, in response to chitin. Material and Methods:Colloidal chitin was added to induce the culture of A. oligospora, and the phenotypic differences before and after induction were observed under inverted microscope. The differential proteins before and after mycelium induction were screened by liquid chromatography-tandem mass spectrometry. The differentially expressed chitinase was expressed in Pichia yeast, and the recombinant enzyme was incubated with Caenorhabditis elegans and its egg suspension to explore its biological activity. Results:It was found that there was a significant acceleration in the mycelial growth post chitin interaction in A. oligospora. A total of 1,124 differentially expressed proteins (DEPs) were identified between the control group (AO-c) and the experimental group (AO-e), with 183 upregulated and 941 downregulated. Gene Ontology analysis revealed that the DEPs acted in various metabolic processes with catalysis and binding functions. Kyoto Encyclopedia of Genes and Genomes analysis associated these proteins primarily with signalling pathways related to glucose metabolism. Three chitinases were significantly modulated among DEPs. Moreover, enzymatic activity assays demonstrated that one of them effectively degraded C. elegans and its eggs. Conclusion:These findings suggest that A. oligospora can significantly alter its protein expression profile in response to chitin, thereby facilitating its sugar metabolism and mycelial development. Our study provided new insights into the regulatory mechanisms of nematode predation in A. oligospora.

Cystic echinococcosis (CE), caused by Echinococcus granulosus sensu stricto (s.s.), poses a substantial risk to both humans and domestic animals globally. Here, we compared the whole genomes of 111 E. granulosus s.s. samples from China. Genomic variation data revealed frequent cross-fertilization in the hermaphroditic E. granulosus. The G1 and G3 genotypes represent distinct mitochondrial lineages, while showing no differentiation in the nuclear genome, suggesting mito-nuclear discordance caused by historical geographic separation and subsequent fusion. Population structure, demographic history, and gene flow among populations reflected the transmission route of E. granulosus s.s. from the Middle East to Qinghai-Xizang Plateau through the migration of nomadic people, followed by introgression during secondary contact. Genomic variations highlighted selection signatures within the genome prone to balancing selection, particularly impacting genes encoding membrane-related proteins, representing a potential evolutionary strategy for adaptation to parasitic life. Balancing selection pressure on the gene-coding sodium/bile acid cotransporter led to its high level of genetic stability, which may play a crucial role in the survival and development of E. granulosus during the parasitic stage, making it a potential drug target for the treatment of CE. Meanwhile, other genomic regions under strong balancing selection may provide potential targets for protective immunity. These findings offer valuable insights into the complex dynamics and adaptive evolution of E. granulosus s.s. in China.IMPORTANCEEchinococcus granulosus sensu stricto (s.s.) is the primary cause of cystic echinococcosis (CE), a parasitic disease affecting humans and livestock with significant health and economic impacts. Previous studies on this parasite relied on mitochondrial DNA to classify its genotypes and understand its genetic diversity. However, these studies cannot capture the full complexity of its evolutionary dynamics and adaptation strategies. Our research employs comprehensive genome-wide sequencing, offering a more nuanced view of its genetic landscape. We discovered that cross-fertilization appears to be a prevalent reproductive strategy in the hermaphroditic E. granulosus, underpinning the observed deep mitochondrial divergence between genotypes G1 and G3, as well as gene flow among populations. The transmission history of E. granulosus s.s. in China and its widespread genetic mixing were likely facilitated by the migrations of nomadic peoples. Furthermore, we identified genes under balancing selection, including the gene involved in the uptake of host bile acids, which play a crucial role in the parasite's survival and development, potentially offering new targets for intervention. Our research advances the understanding of the genetic diversity and evolutionary strategies of E. granulosus, laying the foundation for improved control measures of CE.

The nematode-trapping fungus (NTF) Arthrobotrys oligospora (A. oligospora) is a promising biocontrol agent, but the transcriptional regulators governing its predation remain poorly understood. Here, we demonstrated that the APSES transcription factor AoMbp1 is a master regulator of its development and stress adaptation. Deletion of AoMbp1 severely impaired mycelial growth, conidiation, trap formation, and tolerance to oxidative and osmotic stresses. Transcriptome analysis revealed that these defects were associated with the widespread downregulation of genes, including those within the MAPK signaling pathway. Crucially, we showed that AoMbp1 directly binds to the promoter of AoSho1, a key upstream sensor of the MAPK cascade, and activates its expression. This finding establishes a direct AoMbp1-AoSho1 regulatory axis controlling trap morphogenesis and environmental adaptation. Our study provides novel mechanistic insights into the regulation of nematode trapping and identifies a potential target for enhancing the efficacy of A. oligospora as a biocontrol agent.

Exosomes are nanoscale vesicles secreted by cells that play vital regulatory roles in intercellular communication and immune responses. Listeria monocytogenes (L. Monocytogenes, LM) is a notable Gram-positive intracellular parasitic bacterium that infects humans and diverse animal species. However, the specific biological function of exosomes secreted by macrophages during L. Monocytogenes infection (hereafter EXO-LM) remains elusive. Here, we discovered that EXO-LM stimulated the secretion of inflammation-associated cytokines by macrophages, facilitating the intracellular survival of L. monocytogenes within macrophages. Transcriptomic analysis shows that EXO-LM significantly upregulates immune recognition and inflammation-related signaling pathways in macrophages. Furthermore, a ceRNA regulatory network comprising exosomal ncRNAs and macrophage RNAs was constructed through EXO-LM transcriptome sequencing. Utilizing bioinformatics and dual-luciferase reporter assays, we identified two potential binding sites between lncRNA Rpl13a-213 and miR-132-3p. Cell transfection experiments demonstrated that Rpl13a-213 overexpression augmented pro-inflammatory cytokine expression in macrophages, in contrast to the suppression by miR-132-3p overexpression. The decrease in Rpl13a-213 upon EXO-LM stimulation enhances miR-132-3p expression, dampening the inflammatory response in macrophages and aiding L. monocytogenes intracellular survival. This study unveils the immunomodulatory function of exosomal ncRNAs originating from macrophages, which provides fresh perspectives into the mechanisms underlying macrophage inflammatory response regulation by L. monocytogenes-infected cell-derived exosomes.

Liver diseases often coincide with dysregulated gut homeostasis and Streptococcus overgrowth, yet the underlying mechanisms remain unclear. Here, we study patients with liver echinococcosis and other liver conditions. We observe that these patients frequently exhibit a co-occurrence of an ectopic expansion of orally derived Streptococcus species implicated in intestinal inflammation, alongside a bile acid deficiency in the gut. This association is typically characterized by the reduction of 12-ketolithocholic acid (12-KetoLCA), which exerts potent membrane-disrupting activity. We show that liver disorders compromise gut resistance to oral microbes due to a loss of ecological control from bile acids, particularly 12-KetoLCA. This bile-acid-conferred gut barrier is regulated by cytochrome P450 enzymes and can be reconstituted through adeno-associated virus (AAV) gene therapy targeting these genes. Additionally, we reveal that supplementation with 12-KetoLCA prevents oral Streptococcus-driven gut inflammation through antibacterial activity. Our findings underscore the essential role of bile acids in maintaining the oral-gut barrier.

ABSTRACT Alveolar echinococcosis (AE) is caused by the invasive growth of the metacestodes of Echinococcus multilocularis ( E. multilocularis ). The early diagnosis, management, and treatment of AE remains challenging. Herein, we integrated bulk RNA-seq, scRNA-seq, and ST technologies to reveal the immune characteristics both spatially and chronologically in E. multilocularis infected mouse liver. An unprecedented high-resolution spatial atlas of the E. multilocularis infection foci was obtained, revealing the pivotal role of neutrophils, Spp1 + monocyte-derived macrophages (MoMFs), and fibroblasts in AE progression. We observed continuous recruitment of neutrophils and macrophages into the infected tissues, indicating prolonged immune stimulation by the growing metacestodes. However, their spatial distribution patterns and functions changed during the infection cause. At the early infection stage, Il1b hi neutrophils aggregated in the lesion displayed the highest pattern-recognition receptor activity and may kill the metacestodes through NETosis, but their death in the infection foci might also trigger immune suppression. Meanwhile, Spp1 + MoMFs attached to the protoscoleces to clear the infection. At the late infection stage, the neutrophils failed to infiltrate into the lesion although their continuous recruitment into the infected liver. Even though the Spp1 + MoMFs enclosing the microcysts of E. multilocularis may retain their pathogen-killing ability, they may also promote fibrosis and angiogenesis through interaction with fibroblasts. The pro- and anti-inflammatory phenotype balance of these immune cells may be associated with the transition of parasite control strategy from “active killing” to “negative segregation” by the host. Our findings identify key molecular traits involved in AE development, which may benefit the treatment of echinococcosis.

Introduction. Salmonella Typhimurium (STM) is a food-borne Gram-negative bacterium, which can infect humans and a wide range of livestock and poultry, causing a variety of diseases such as septicaemia, enteritis and abortion.Hypothesis/Gap Statement. We will decipher the impacts of sRNA STnc1280 on STM virulence and provide a theoretical basis to reveal the regulatory role and molecular mechanism of STnc1280.Aim. The main objective of this study was to clarify whether sRNA STnc1280 exerts regulatory roles on STM pathogenicity.Methodology. The STnc1280 gene was amplified and its molecular characteristics were analysed in this study. Then, STnc1280 gene deletion strain (STM-ΔSTnc1280) and the complementary strain (ΔSTnc1280/STnc1280) were constructed by λ-Red homologous recombination method, respectively, to analyse of adhesion and invasive ability and pathogenicity of different strains. Subsequently, the potential target gene regulated by STnc1280 was predicted using target RNA2 software, followed by the verification of the interaction between STnc1280 and target mRNA using the dual plasmid reporter system (DPRS). Furthermore, the mRNA and protein level of target gene was determined using qRT-PCR and Western blot, respectively.Results. The results revealed that the cell adhesion and invasive ability and pathogenicity of STM-ΔSTnc1280 were significantly reduced compared to STM-SL1344 strain, indicating that the deficiency of STnc1280 gene significantly influenced STM pathogenicity. The DPRS results showed that STnc1280 can interact with the mRNA of target gene gldA, thus suppressing the expression of lacZ gene. Furthermore, the level of gldA mRNA was not influenced in STM-ΔSTnc1280, but the expression of GldA protein decreased significantly.Conclusion. Combining the bioinformatic analysis, these findings suggested that STnc1280 may bind to the SD sequence of gldA mRNA, hindering the binding of ribosomes to gldA mRNA, thereby inhibiting the expression of GldA protein to modulate the virulence of STM.

Salmonella Typhimurium (STM) is an important zoonotic Gram-negative pathogen that can cause infection in a variety of livestock and poultry. Meanwhile, as an important foodborne pathogen, the bacterium can survive in various stressful environments and transmits through the fecal-oral route, posing a serious threat to global food safety. To investigate the roles of STM1863, a member of the DUFs protein family, involved in STM environmental adaptation, biofilm formation, and virulence. We analyzed the molecular characteristics of the protein encoded by STM1863 gene and examined intra- and extracellular expression levels of STM1863 gene in mouse macrophages. Furthermore, we constructed STM1863 gene deletion and complementation strains and determined its environmental adaptation under stressful conditions such as acid, alkali, high salt, bile salt, and oxidation. And the capacity of biofilm formation and pathogenicity of those strains were analyzed and compared. In addition, the interaction between the promoter of STM1863 gene and RcsB protein was analyzed using DNA gel electrophoresis migration assay (electrophoretic mobility shift assay [EMSA]). The experiments revealed that acid adaptability and biofilm formation ability of STM1863 gene deletion strain were significantly weakened compared with the parental and complementary strains. Moreover, the adhesion and invasion ability of STM1863 deletion strain to mouse macrophages was significantly decreased, while the median lethal dose (LD50) increased by 2.148-fold compared with the parental strain. In addition, EMSA confirmed that RcsB protein could bind to the promoter sequence of STM1863 gene, suggesting that the expression of STM1863 gene might be modulated by RcsB. The present study demonstrated for the first time that STM1863, a member of the DUFs protein family, is involved in the modulation of environmental adaptation, biofilm formation, and virulence.

Extracellular proteases, such as chitinases secreted by Arthrobotrys oligospora (A. oligospora), play a crucial role in the process of nematode infection. However, post-transcriptional regulation of gene expression involving microRNAs (miRNAs) in A. oligospora remains scarcely described. Hereto, transcriptome sequencing was carried out to analyze the expression profiles of chitin-responsive miRNAs in A. oligospora. Based on the RNA-seq data, the differential expression of miRNAs (DEmiRNAs) in response to chitin was screened, identified and characterized in A. oligospora. Meanwhile, the potential target genes were predicted by the online tools miRanda and Targetscan, respectively. Furthermore, the interaction of DEmiRNA with it’s target gene was validated by a dual-luciferase reporter assay system. Among 85 novel miRNAs identified, 25 miRNAs displayed significant differences in expression in A. oligospora in response to chitin. Gene Ontology (GO) analysis showed that the potential genes targeted by DEmiRNAs were enriched in the biological processes such as bio-degradation, extracellular components and cell cycle. KEGG analysis revealed that the target genes were mainly involved in Hippo, carbon and riboflavin metabolic pathway. Outstandingly, chitinase AOL_s00004g379, which is involved in the hydrolysis metabolic pathway of chitin, was confirmed to be a target gene of differential miR_70. These findings suggest that chitin-responsive miRNAs are involved in the regulation of cell proliferation, predator hyphae growth and chitinase expression through the mechanisms of post-transcriptional regulation, which provides a new perspective to the molecular mechanisms underlying miRNAs-mediated control of gene expression in A. oligospora.

As important post-transcriptional regulators of gene expression, sRNAs play important modulatory roles in the environmental adaptation and virulence of bacteria. To investigate the regulatory role of sRNA STnc3020 in the virulence of Salmonella typhimurium (S. typhimurium). This study analyzed the impacts of STnc3020 deletion on adherence, invasion, intracellular survival, macrophage apoptosis, and pathogenicity of S. typhimurium in mice. Furthermore, potential regulatory target genes of STnc3020 were identified and its regulatory mechanism was validated. The results showed that at the cellular level, the deletion of STnc3020 significantly reduced the adhesion ability of S. typhimurium to intestinal epithelial cells (P < 0.01), as well as its proliferation and apoptosis-inducing abilities within macrophages (P < 0.01). Meanwhile, animal experiment results indicated that the deletion of STnc3020 significantly reduced the colonization rate of S. typhimurium in the liver and cecum of mice (P < 0.01), and increased the median lethal dose (4.28 × 105) in mice. Regulatory mechanism research results showed that STnc3020 can interact with the target gene prgJ of the Type III secretion system (T3SS), and the protein level of PrgJ significantly decreased after the deletion of STnc3020 (P < 0.01). These findings offer new insights into sRNA-mediated virulence control and may aid in developing new vaccines and drugs for S. typhimurium.

The 2023 International African Swine Fever Workshop (IASFW) took place in Beijing, China, on 18–20 September 2023. It was jointly organized by the U.S.-China Center for Animal Health (USCCAH) at Kansas State University (KSU) and the Chinese Veterinary Drug Association (CVDA) and sponsored by the United States Department of Agriculture Foreign Agricultural Service (USDA-FAS), Harbin Veterinary Research Institute, and Zoetis Inc. The objective of this workshop was to provide a platform for ASF researchers around the world to unite and share their knowledge and expertise on ASF control and prevention. A total of 24 outstanding ASF research scientists and experts from 10 countries attended this meeting. The workshop included presentations on current ASF research, opportunities for scientific collaboration, and discussions of lessons and experiences learned from China/Asia, Africa, and Europe. This article summarizes the meeting highlights and presents some critical issues that need to be addressed for ASF control and prevention in the future.