Search Results (109)

Background: Infections caused by multidrug-resistant bacteria and inadequate vaccine coverage against opportunistic pathogens highlight the need for interventions that broadly and durably enhance host defense beyond antigen-specific adaptive immunity. Trained immunity, driven by metabolic and epigenetic reprogramming of innate immune cells, has been predominantly characterized using Bacille Calmette–Guérin and β-glucan, whereas its induction by Gram-negative bacteria remains poorly defined. To address this gap, we aimed to determine whether heat-killed Klebsiella pneumoniae (HK Kp) induces trained immunity through metabolic and hematopoietic reprogramming to confer heterologous antibacterial protection. Methods: HK Kp-trained murine bone marrow-derived macrophages and HK Kp-immunized C57BL/6 mice were employed to interrogate functional, metabolic, and transcriptomic reprogramming in vitro, hematopoietic progenitor remodeling in vivo, and protective efficacy against systemic Salmonella Typhimurium and Staphylococcus aureus infection. Results: HK Kp-trained macrophages showed markedly enhanced IL-1β secretion across all restimulation conditions, stimulus-dependent amplification of TNF-α responses, increased phagocytosis, and improved intracellular control of S. typhimurium, together with sustained upregulation of the glycolytic enzymes-encoding genes Hk2 and Pfkfb3. Transcriptomic profiling revealed extensive reprogramming enriched in glycolysis/gluconeogenesis and hematopoietic cell lineage pathways. In vivo, HK Kp immunization shifted bone marrow stem/progenitor compartments toward a myeloid-biased state. HK Kp-trained mice challenged with lethal S. typhimurium or S. aureus exhibited less weight loss, improved survival rates, and reduced bacterial burdens. Conclusions: Inactivated K. pneumoniae orchestrates metabolic and hematopoietic reprogramming to establish enhanced innate immune responsiveness and confer heterologous protection in murine S. typhimurium and S. aureus sepsis models, supporting its potential as a potent inducer of trained immunity. These findings establish HK Kp-based trained immunity as a promising strategy for combating multidrug-resistant and vaccine-evading pathogens. Full article

Lactic acid bacteria (LAB) have dominated food fermentation globally and are ingrained in many food cultures. Obesity is a global health concern, and LAB ingestion is known to exert anti-obesity effects in animals. However, the characteristics of individual bacterial strains and their underlying mechanisms require elucidation since the anti-obesity effects can differ with variations in the strain, host, and living environment. In this study, we aimed to evaluate the safety and anti-obesity effects of Lactiplantibacillus plantarum TO-A (LPTOA), isolated from silage, using Caenorhabditis elegans as the model organism. The study findings revealed that LPTOA was non-toxic to mice, as established via subacute toxicity tests, and extended the lifespan of C. elegans. Furthermore, both LPTOA and heat-killed LPTOA reduced fat accumulation in C. elegans by 60% and 58%, respectively. However, in vitro experiments suggested that LPTOA does not decompose cholesterol and triglycerides, nor does it inhibit lipase activity. We identified that pept-1 (a dipeptide transporter) in C. elegans is involved in the anti-obesity effects of LPTOA. PEPT-1 is a protein that controls proton influx into the intestinal tract and is involved in not only peptide uptake but also free fatty acid absorption. These results demonstrate the anti-obesity effects and probiotic potential of LPTOA for application in products, including foods and supplements. Full article

Heat-killed Lactiplantibacillus plantarum SNK12 (SNK), isolated from a traditional Japanese fermented food, has been suggested to influence sleep quality, but human data on sleep improvement with heat-killed lactic acid bacteria (postbiotics) remain limited. We conducted a randomized controlled trial to test whether heat-killed SNK (≥1 × 10¹¹ cells/day for 4 weeks) improves sleep quality and alters stress-related immune and neuroendocrine biomarkers. Healthy adults received SNK or a placebo for 4 weeks. The primary outcome was the Oguri–Shirakawa–Azumi Sleep Inventory MA version (OSA-MA) factor “Sleepiness on Rising”; secondary outcomes were other OSA-MA factors and the stress-related biomarkers salivary cortisol and plasma tumor necrosis factor-α (TNF-α). Compared with placebo, SNK improved Sleepiness on Rising (p = 0.032) and Initiation and Maintenance of Sleep (p = 0.010). Salivary cortisol (p = 0.016) and plasma TNF-α (p = 0.037) were also lower with SNK, and no safety concerns emerged. These concomitant changes in subjective sleep indices and stress-related biomarkers are consistent with modulation of hypothalamic–pituitary–adrenal axis activity and inflammatory pathways along the gut–brain axis. SNK may, therefore, represent a practical postbiotic option to support sleep quality. Full article

Objective: Previous reports showed that periodontopathic bacteria induce epithelial–mesenchymal transition (EMT) in oral squamous cell carcinoma (OSCC). Fisetin, a foodborne flavonoid, is reportedly associated with anticancer potential in various carcinogenic processes. This study aimed to elucidate the effects of fisetin on Fusobacterium nucleatum- and Porphyromonas gingivalis-induced EMT in OSCC cells. Methods: OSCC cells were co-cultured with live and heat-killed forms of F. nucleatum and P. gingivalis. The concentration of fisetin was set at 10 μM. Morphological changes in the OSCC cells were observed under a light microscope. Cell viability was measured using the Cell Counting Kit-8 assay, whereas migration was examined via wound healing. The mRNA expression of EMT-related markers was quantified using quantitative real-time polymerase chain reaction (PCR), and the expression of EMT-related markers and Wnt pathway-associated proteins was examined via Western blotting. Results: At a multiplicity of infection (MOI) of 300:1 for F. nucleatum and 100:1 for P. gingivalis, OSCC cell viability remained unchanged; however, wound closure rates increased significantly relative to the control. Likewise, treatment with fisetin (10 µM) did not materially alter viability; nevertheless, it attenuated promigratory effects induced by heat-killed periodontal pathogens at 3 h and 6 h. The OSCC cells exhibited EMT-like morphological changes after 6 h of co-culture with heat-killed pathogens. Consistently, reverse-transcriptase quantitative PCR and Western blot analyses showed increased expression of TWIST, ZEB1, and N-cadherin, accompanied by decreased E-cadherin expression, which was more pronounced in F. nucleatum than in P. gingivalis. However, fisetin reversed these trends. Moreover, co-culture with heat-killed pathogens markedly elevated β-catenin protein levels. In line with modulation of canonical Wnt/β-catenin signaling, fisetin and a Wnt inhibitor reduced β-catenin expression, whereas co-treatment with a Wnt agonist restored β-catenin levels in the presence of fisetin. Conclusions: Heat-killed F. nucleatum and P. gingivalis induced EMT in OSCC cells, with F. nucleatum exerting the strongest effect. Fisetin suppressed pathogen-driven EMT, at least partly via canonical Wnt/β-catenin signaling, highlighting its potential therapeutic value and warranting further investigation. Full article

Heating at 95 °C or boiling E. coli HT115 (DE3) cells is often used to extract heterologous dsRNA or kill bacteria, although these temperatures cause dsRNA denaturation and destruction. In this study, we examined the risk of degradation of dsRNA fragments of AT-rich genes at high temperature. The expression of dsRNA fragments of AT-rich genes encoding DNA replication enzymes from the microsporidia Vairimorpha ceranae and Nosema bombycis in E. coli HT115 (DE3) was accompanied by heating the bacteria at 95 °C for 30 min. In contrast to four control fragments with normal GC content, the AT-rich dsRNAs of microsporidia were destroyed by this treatment. The in vitro synthesis and heating of the studied dsRNAs showed the degradation of both microsporidia and control fragments. The thermal degradation of in vitro-synthesized control dsRNA with a normal GC content of 47.6% was prevented by the addition of 2 × YT media, NaCl, or low concentrations of MgSO₄. This demonstrates the important role of mono- and divalent cations in stabilizing heated fragments and helps explain the preservation of their integrity and RNAi-initiating activity despite the treatment of bacteria at temperatures that denature dsRNA. Feeding Colorado potato beetle larvae with the same in vitro-synthesized dsRNA containing fragments of three Leptinotarsa decemlineata genes showed that their thermal destruction was accompanied by a decrease in pest-suppressing activity. No dsRNA degradation was observed at 80 °C or after E. coli sonication, and these treatments, as well as increasing cation content, may help to avoid the degradation of heat-sensitive dsRNA. Full article

Sleeplessness (insomnia) is a significant symptom associated with stress-induced depression/anxiety. In the present study, we selected Bifidobacterium longum P77, which increased serotonin production in corticosterone-stimulated SH-SY5Y cells, from the fecal bacteria collection of healthy volunteers and examined the effects of B. longum on depression, anxiety, and sleeplessness induced by immobilization stress or by transplantation of cultured fecal microbiota (cFM) from patients with depression. Orally administered B. longum P77 decreased depression/anxiety- and sleeplessness-like behaviors in immobilization stress-exposed mice. B. longum P77 reduced immobilization stress-induced corticosterone, tumor necrosis factor (TNF)-α, and interleukin (IL)-6 expression and the cell population of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB)⁺ in the prefrontal cortex, while the expression levels of immobilization stress-suppressed IL-10, γ-aminobutyric acid (GABA), its receptor GABA_ARα1, serotonin, and its receptor 5-HT_1AR increased. B. longum P77 also alleviated immobilization stress-induced colitis: it decreased TNF-α and IL-6 expression and increased IL-10 expression in the colon. Furthermore, B. longum P77, Lactiplantibacillus plantarum P72, and their combination decreased cFM- or immobilization stress-induced depression-, anxiety-, and sleeplessness-like behaviors. They also decreased cFM-induced, corticosterone, TNF-α, and IL-6 expression levels in the prefrontal cortex and colon, while increasing cFM- or immobilization stress-suppressed GABA, GABA_ARα1, serotonin, and 5-HT_1AR expression levels in the prefrontal cortex. In particular, the combination of B. longum P77 and L. plantarum P72 (P7277) additively or synergistically alleviated depression-, anxiety-, and sleeplessness-like behaviors, along with their associated biomarkers. Heat-killed P7277 also alleviated immobilization stress-induced depression/anxiety- and sleeplessness-like symptoms. These results imply that L. plantarum P72 and/or B. longum P77 can mitigate depression/anxiety and sleeplessness by upregulating GABAergic and serotonergic systems, along with the suppression of NF-κB activation. Full article

Background/Objectives: Antimicrobial resistance (AMR) contributes to millions of deaths globally each year, creating an urgent need for new therapeutic agents. Antimicrobial peptides (AMPs) have emerged as promising candidates due to their potential to combat AMR pathogens. This study aimed to evaluate the antimicrobial activity of an AMP from a soil-derived bacterial isolate against Gram-negative bacteria. Method: Soil bacteria were isolated and screened for antimicrobial activity. The bioactive peptide was purified and determined its structure and antimicrobial efficacy. Genomic analysis was conducted to predict the biosynthetic gene clusters (BGCs) responsible for AMP production. Results: Genomic analysis identified the isolate as Paenibacillus sp. Na14, which exhibited low genomic similarity (61.0%) to other known Paenibacillus species, suggesting it may represent a novel species. The AMP from the Na14 strain exhibited heat stability up to 90 °C for 3 h and retained its activity across a broad pH range from 3 to 11. Structural analysis revealed that the Na14 peptide consisted of 14 amino acid residues, adopting an α-helical structure. This peptide exhibited bactericidal activity at concentrations of 2–4 µg/mL within 6–12 h, and its killing rate was concentration-dependent. The peptide was found to disrupt the bacterial membranes. The Na14 peptide shared 64.29% sequence similarity with brevibacillin 2V, an AMP from Brevibacillus sp., which also belongs to the Paenibacillaceae family. Genomic annotation identified BGCs associated with secondary metabolism, with a particular focus on non-ribosomal peptide synthetase (NRPS) gene clusters. Structural modeling of the predicted NRPS enzymes showed high similarity to known NRPS modules in Brevibacillus species. These genomic findings provide evidence supporting the similarity between the Na14 peptide and brevibacillin 2V. Conclusions: This study highlights the discovery of a novel AMP with potent activity against Gram-negative pathogens and provides new insight into conserved AMP biosynthetic enzymes within the Paenibacillaceae family. Full article

The gut microbiome is vital in maintaining metabolic health, and dietary habits can significantly impact its composition. A high-fat diet (HFD) can disrupt gut microbial balance, contributing to obesity, insulin resistance, and fatty liver disease. This study explores the potential benefits of heat-killed Enterococcus faecalis EF-2001 (EF-2001) in restoring gut balance and improving metabolic health in HFD-fed mice (HFD-mice). HFD mice administered EF-2001 had 18% less body fat, 22% lower triglyceride levels, and significantly reduced liver enzyme markers, including aspartate aminotransferase (AST) by 28% and alanine aminotransferase (ALT) by 31%. Additionally, EF-2001 improved glucose metabolism, increasing glucose tolerance by 20% and insulin sensitivity by 15%, while reducing fat buildup in the liver by 24%, indicating protection against fatty liver disease. These changes correlated with better metabolic health and reduced inflammation. Our results show that EF-2001 supplementation helped counteract HFD-induced gut imbalances by increasing microbial diversity and supporting beneficial bacteria, such as Akkermansia and Ligilactobacillus spp. Our findings highlight the potential of heat-killed EF-2001 as a promising strategy to restore gut balance and mitigate diet-related metabolic issues. Furthermore, analysis of antibiotic resistance genes (ARGs) revealed that HFD mice exhibited an increased abundance of multidrug resistance genes, particularly those associated with antibiotic efflux mechanisms, such as bcrA, cdeA, and msbA. Notably, EF-2001 supplementation mitigated this increase, reducing the relative abundance of the above ARGs and suggesting a protective role in limiting the spread of antibiotic resistance linked to dysbiosis. EF-2001 offers a compelling approach to managing obesity and metabolic disorders, paving the way for microbiome-based health interventions. Full article

The global energy sector is aiming to substantially reduce CO₂ emissions to meet the UN climate goals. Among the proposed strategies, underground storage solutions such as radioactive disposal, CO₂, NH₃, and underground H₂ storage (UHS) have emerged as promising options for mitigating anthropogenic emissions. These approaches require rigorous research and development (R&D), often involving laboratory-scale experiments to establish their feasibility before being scaled up to pilot plant operations. Microorganisms, which are ubiquitous in laboratory environments, can significantly influence geochemical reactions under variable experimental conditions of porous media and a salt cavern. We have selected a consortium composed of Bacillus sp., Enterobacter sp., and Cronobacter sp. bacteria, which are typically present in the laboratory environment. These microorganisms can contaminate the rock sample and develop experimental artifacts in UHS experiments. Hence, it is pivotal to sterilize the rock prior to conduct experimental research related to effects of microorganisms in the porous media and the salt cavern for the investigation of UHS. This study investigated the efficacy of various disinfection and sterilization methods, including ultraviolet irradiation, autoclaving, oven heating, ethanol treatments, and gamma irradiation, in removing the microorganisms from silica sand. Additionally, the consideration of their effects on mineral properties are reviewed. A total of 567 vials, each filled with 9 mL of acid-producing bacteria (APB) media were used to test killing efficacy of the cleaning methods. We conducted serial dilutions up to 10⁻⁸ and repeated them three times to determine whether any deviation occurred. Our findings revealed that gamma irradiation and autoclaving were the most effective techniques for eradicating microbial contaminants, achieving sterilization without significantly altering the mineral characteristics. These findings underscore the necessity of robust cleaning protocols in hydrogeochemical research to ensure reliable, reproducible data, particularly in future studies where microbial contamination could induce artifacts in laboratory research. Full article

Background/Objectives: Alternative therapies for urinary tract infections (UTIs) have been explored, but their efficacy remains inconsistent. With rising antibiotic resistance, this study aimed to evaluate simplified postbiotic formulations derived from heat-killed probiotics for long-term protection against primary and recurrent UTIs in a murine model. Methods: We compared a multi-strain (seven-strain) versus a single-strain postbiotic in preventing Escherichia coli-induced UTIs and recurrent polymicrobial UTIs, assessed protection persistence after treatment discontinuation, and established a novel sustained UTI model via intravesical co-inoculation of three uropathogens. Mice were allocated to three experimental groups: a placebo group (PBS), Postbiotic I group (a seven-strain heat-killed probiotic formulation), and Postbiotic II group (a single-strain heat-killed probiotic). After two weeks of treatment, mice were challenged with uropathogenic E. coli (UPEC) and treated for seven days. Following a 14-day washout and bacterial clearance, they were rechallenged with multidrug-resistant UPEC, Klebsiella pneumoniae, and Staphylococcus pseudintermedius. Results: Both postbiotics significantly accelerated bacterial clearance in primary UTIs (p < 0.05). In recurrent UTIs, placebo-treated mice exhibited persistent bacteriuria, while Postbiotic I maintained a significantly higher sterile urine rate (50–80%, p < 0.01) post-treatment. Histopathological analysis confirmed reduced bladder and kidney inflammation (p < 0.05) with Postbiotic I. Conclusions: These findings demonstrate the superior efficacy of Postbiotic I in mitigating UTIs, with sustained protection post-treatment, supporting its potential as a long-term, non-antibiotic strategy. Additionally, our reproducible chronic UTI model, achieved through the co-inoculation of three uropathogens, provides a valuable tool for future research on chronic UTI pathogenesis and treatment. Full article

Aminopeptidase N (APN), an enzyme expressed in the small intestinal mucosa, is involved in dietary protein digestion. Previous studies have shown that oral administration of fermented milk containing lactic acid bacteria (LAB) enhances mucosal APN activity in young mice. This study aimed to investigate whether LAB strains stimulate mucosal APN activity in aged mice and to evaluate its relevance to age-related changes in body composition. The underlying molecular mechanisms were also explored in vitro. Experiment 1: Aged C57BL/6J mice were fed diets supplemented with heat-killed LAB strains—Enterococcus faecalis OU-23 (EF), Leuconostoc mesenteroides OU-03 (LM), or Lactiplantibacillus plantarum SNK12 (LP). Compared to the aged Control group, the ileal APN activity was significantly higher in the LP group. LP administration also elevated serum Gla-osteocalcin levels and decreased serum CTX-1 levels. Experiment 2: IEC-6 cells were co-cultured with LP that had been treated with RNase, DNase, or lysozyme. APN activity was significantly lower in cells co-cultured with DNase- or lysozyme-treated LP compared to those co-cultured with untreated LP. A specific LAB strain may enhance mucosal APN activity in the aged intestine, potentially contributing to improved bone metabolism. This effect may be mediated by bacterial DNA and peptidoglycan. Full article

Canine atopic dermatitis (AD) is a T-cell-driven inflammatory skin disease, characterized by an imbalance between the Th1 and Th2 immune responses. Probiotics (live bacteria) and postbiotics (inactivated, killed bacteria) have garnered attention for the management of AD in humans and dogs. Both probiotics and postbiotics possess immunomodulating properties that could be beneficial for allergic patients. This study aims to evaluate the immunomodulating effects of Tyndallized (heat-killed) postbiotics of Lactobacillus rhamnosus and Lactobacillus reuteri, which are active components of the Linkskin products (Nextmune, Palazzo Pignano, Cremona, Italy). Peripheral blood mononuclear cells (PBMCs) were isolated from healthy dogs and incubated separately with each postbiotic. The cytokine levels in the supernatants were measured before and after 12, 24, 48, and 72 h of incubation. Both Tyndallized lactobacilli significantly increased the levels of IL-12 and IFN-γ (Th1 cytokines) and IL-10 (associated with T regulatory cells), while the levels of the Th2 cytokine IL-4 remained stable. Overall, these two Lactobacillus postbiotics stimulated canine PBMCs to produce a cytokine profile typically associated with an anti-allergic response. Further studies are needed to evaluate the benefit of these postbiotics as an adjuvant for the reactive treatment or for the prevention of relapses of allergic flares in atopic dogs. Full article

Non-typhoidal salmonellosis (NTS) caused by multidrug-resistant (MDR) Salmonella enterica is a significant public health concern worldwide. Probiotics offer a potential alternative to antibiotics in many infectious diseases, including NTS. However, using living bacteria raises safety concerns in clinical settings, especially in the immunocompromised host. This study compared the anti-Salmonella and immunomodulatory effects between viable (probiotics) and heat-killed (paraprobiotics) lactic acid bacteria Lactiplantibacillus plantarum KUNN19-2 (KUNN19-2), isolated from Thai-style fermented pork (Nham), against several strains of MDR Salmonella. Only viable KUNN19-2 and its cell-free supernatant directly inhibited Salmonella growth by spot-on lawn and agar well diffusion assays. A significant reduction in Salmonella numbers in the co-culture assay with viable KUNN19-2 was observed at 12–14 h after the incubation. Viable and heat-killed KUNN19-2 exhibited moderate adhesion to human colonic epithelium (T84) cells. Pretreatment with either form of KUNN19-2 enhanced macrophage (RAW264.7) phagocytic activity against Salmonella and upregulated pro-inflammatory genes (Mip-2 and Nos2) and anti-inflammatory gene (IL10) expression, with viable KUNN19-2 showing a more potent effect. Collectively, viable KUNN19-2 can directly inhibit Salmonella growth. However, viable and heat-killed KUNN19-2 can modulate gut immunity against Salmonella infection, suggesting that paraprobiotic KUNN19-2 may serve as an alternative treatment against MDR Salmonella through host immune modulation. Full article

The problem of antibiotic abuse and drug resistance is becoming increasingly serious. In recent years, polydopamine (PDA) nanoparticles have been recognized as a potential antimicrobial material for photothermal therapy (PTT) due to their excellent photothermal conversion efficiency and unique antimicrobial ability. PDA is capable of rapidly converting light energy into heat energy under near-infrared (NIR) light irradiation to kill bacteria efficiently. In order to solve the problem of PDA’s tendency to aggregate and precipitate, this study improved its stability by grafting hyaluronic acid (HA) onto the surface of PDA. Using dopamine and hyaluronic acid as raw materials, hyaluronic acid (HA) was grafted onto polydopamine (PDA) nanoparticles via self-polymerization and Michael addition reactions under alkaline conditions to obtain PDA-HA-modified nanoparticles. We confirmed the successful grafting of hyaluronic acid via scanning electron microscopy (SEM), Fourier infrared spectroscopy (FTIR), nuclear magnetic hydrogen spectroscopy (¹H NMR), ultraviolet–visible spectroscopy (UV–vis), Raman spectroscopy (Raman), and dynamic light scattering (DLS) methods. Scanning electron microscopy (SEM) was used to observe the surface morphology and nanostructure of the grafted materials, providing information on the morphology and size distribution of the materials. Near-infrared performance experiments showed that the temperature of the PDA-HA solution increased rapidly under near-infrared light irradiation, demonstrating an excellent photothermal conversion performance. Antimicrobial properties were assessed via the colony counting method, and typical Gram-positive bacteria S. aureus and Gram-negative bacteria E. coli were selected as model strains. The experimental groups were tested under dark conditions and near-infrared (NIR) light irradiation. PDA/HA showed significant photothermal properties under NIR light irradiation, resulting in a rapid increase in the surrounding temperature to a level sufficient to kill bacteria. Under NIR light irradiation, PDA/HA exhibited 100% antimicrobial efficacy against both S. aureus and E. coli, while antimicrobial efficacy was limited under dark conditions. This indicates that the antibacterial activity of PDA/HA is highly dependent on NIR light activation. Full article

This study investigated whether viable cells, dead cells or cell-free supernatants (CFS) were responsible for the biocontrol effect of strains from two important bacterial genera, Pseudomonas and Lactobacillus, known for their antifungal properties against plant pathogens and food spoilage microorganisms. Specifically, the capability of these strains to produce extracellular hydrolytic enzymes on specified media was assessed, along with their effectiveness in inhibiting the mycelial growth of several phytopathogenic fungi (Fusarium oxysporum, Botrytis cinerea, Pythium ultimum and Rhizoctonia solani) using dual culture plate assays. Results from these inhibition assays revealed that P. fluorescens PF05 and L. plantarum LMG 23520 strains were the most effective in suppressing fungal growth, especially F. oxysporum. Therefore, further experiments were carried out to investigate the antifungal potential of the viable cells, heat-killed cells (HKC) and CFS from these strains against the germination of F. oxysporum spores. The viable cell trial proved successful, whereas HKC from the two bacterial isolates were ineffective against fungal spore germination. Conversely, the CFS of L. plantarum LMG 23520 was able to prevent fungal spore development for up to six days. The CFS of P. fluorescens PF05, instead, did not yield positive results. Additional studies are required to evaluate the potential inhibitory effects of the CFS from P. fluorescens PF05 and the HKC from both strains. Full article