Search Results (160)

Effective rhizobium–legume symbiosis depends on multiple molecular signaling pathways, integrating not only classical nodulation factors and surface polysaccharides but also diverse protein secretion systems. Among them, the Type VI Secretion System (T6SS) has emerged as a key player, due to its dual roles in interbacterial competition and interactions with eukaryotic hosts, though its contribution to symbiosis remains unclear. Key regulatory messengers, including the main autoinducer of the quorum sensing (QS) systems, the N-acyl homoserine lactones (AHLs), and the second messenger cyclic di-GMP (c-di-GMP), modulate the transition between motility and biofilm formation, especially in the context of bacteria interacting with eukaryotes, including rhizobia. While c-di-GMP’s impact on exopolysaccharide production in these organisms is well established, its influence on protein secretion systems, particularly in conjunction with QS, is largely unexplored. To contribute to the study of such interplay, we artificially increased intracellular c-di-GMP levels by overexpressing a heterologous diguanylate cyclase in three Sinorhizobium fredii strains of agronomic relevance. This engineering revealed strain-specific outcomes, since elevated c-di-GMP enhanced biofilm development in two strains, but reduced it in another. Furthermore, using β-galactosidase expression assays, we confirmed that both high c-di-GMP and/or AHL concentrations contribute to the transcriptional activation of T6SS. These results demonstrate a direct regulatory link between c-di-GMP, QS signals, and T6SS expression, shedding light on the multilayered control mechanisms that structure beneficial rhizobia–plant interactions. Full article

Molecularly imprinted polymers (MIPs) have emerged as promising materials for selectively targeting biomolecules, including quorum sensing autoinducers that regulate bacterial communication and biofilm formation. In this study, both single-template and dual-template strategies were employed to design and synthesize MIPs capable of capturing autoinducer-2 analogs using (3R,4S)-tetrahydro-3,4-furandiol (T1) or (R/S) 2,2-dimethyl-1,3-dioxolane-4-methanol (T2) as the templates. This approach offers translational potential of a complementary or non-antibiotic strategy to conventional antimicrobial therapies in mitigating biofilm-associated infections. Computational modeling guided the rational selection of functional monomers, predicting favorable interaction energies (ΔE_C up to −135 kcal·mol⁻¹) and optimal hydrogen-bonding patterns to enhance template–polymer affinity. The synthesized MIPs were characterized using spectroscopic and microscopic techniques to confirm imprinting efficiency and structural integrity. The adsorption capacity measurements demonstrated higher adsorption capacity and selectivity of MIPs compared to non-imprinted polymers, with the highest selectivity equal to 3.36 for T1 and 3.14 for T2 on MIPs fabricated from methacrylic acid. Preliminary microbiological evaluations using Chromobacterium violaceum ATCC 12472 reveal that the MIPs prepared from 2-hydroxyethyl methacrylate effectively inhibited violacein production by up to 78.2% at 5.0 mg·mL⁻¹, consistent with quorum sensing interference. These findings highlight the feasibility of employing molecular imprinting to target autoinducer-2 analogs, introducing a novel synthetic strategy for disrupting bacterial communication. This further suggests that molecular imprinting can be leveraged to develop potent quorum-sensing inhibitors, an approach that offers translational potential as an alternative to conventional antimicrobial strategies to mitigate biofilm-associated infections. Full article

This study evaluated the inhibitory activity of quercetin at sub-inhibitory concentrations on quorum-sensing (QS) molecules in vitro and the effects of dietary supplementation with quercetin (for 24 consecutive days) on enterotoxigenic Escherichia coli (ETEC)-induced inflammatory and oxidative stress responses in weaned piglets. The piglets were fed one of three diets: the basal diet (Con), ETEC challenge (K88) after the basal diet, or ETEC challenge (quercetin + K88) after the basal diet supplemented with 0.2% quercetin. In vitro experiments revealed that 5 mg/mL quercetin exhibited the strongest QS inhibitory activity and reduced pigment production by Chromobacterium violaceum ATCC12472 by 67.70%. In vivo experiments revealed that quercetin + K88 significantly increased immunoglobulin A (IgA), immunoglobulin M (IgM), and immunoglobulin G (IgG) levels in the serum, ileum mucosa, and colon mucosa; increased glutathione peroxidase (GSH-Px), catalase (CAT), and superoxide dismutase (SOD) levels in the serum, liver, and colon mucosa; and decreased cluster of differentiation 3 (CD3) and cluster of differentiation 8 (CD8)activity in the serum compared with K88 alone. Quercetin + K88 significantly alleviated pathological damage to the liver and spleen and upregulated antioxidant genes (nuclear factor erythroid 2-related factor 2 (Nrf2), heme oxygenase-1(HO-1), CAT, SOD, and glutathione s-transferase (GST)). Inducible nitric oxide synthase (iNOS) and kelch-like ech-associated protein 1 (Keap1), which cause oxidative damage to the liver and spleen, were significantly downregulated. The acetic acid content in the cecum was significantly increased, and the E. coli count and QS signal molecule autoinducer-2 (AI-2) yield were significantly reduced. In conclusion, 0.2% dietary quercetin can alleviate ETEC-induced inflammation and oxidative stress in weaned piglets. Full article

Precision nutrition-targeted gut microbiota (GM) may have therapeutic potential not only for age-related diseases but also for slowing the aging process and promoting longer healthspan. Recent studies have shown that restoring a healthy symbiosis of GM by counteracting dysbiosis (DYS) through precise nutritional intervention is becoming a major target for extending healthspan. Microbiota-accessible borate (MAB) complexes, such as boron (B)–pectins (rhamnogalacturonan–borate) and borate–phenolic esters (diester chlorogenoborate), have a significant impact on healthy host–microbiota symbiosis (HMS). The mechanism of action of MABs involves the biosynthesis of the autoinducer-2–borate (AI-2B) signaling molecule, B fortification of the mucus gel layer by the MABs diet, inhibition of pathogenic microbes, and reversal of GM DYS, strengthening the gut barrier structure, enhancing immunity, and promoting overall host health. In fact, the lack of MAB complexes in the human diet causes reduced levels of AI-2B in GM, inhibiting the Firmicutes phylum (the main butyrate-producing bacteria), with important effects on healthy HMS. It can now be argued that there is a relationship between MAB-rich intake, healthy HMS, host metabolic health, and longevity. This could influence the deployment of natural prebiotic B-based nutraceuticals targeting the colon in the future. Our review is based on the discovery that MAB diet is absolutely necessary for healthy HMS in humans, by reversing DYS and restoring eubiosis for longer healthspan. Full article

Quorum quenchers are emerging as an alternative to conventional antimicrobials, since they hinder the development of virulence or resistance mechanisms but without killing the microorganisms, thus, reducing the risk of antimicrobial resistance. Many quorum quenchers are analogs of the natural quorum-sensing signaling molecules or autoinducers. Thus, different analogs of natural N-acylhomoserine lactones (AHLs) have been reported for controlling virulence or reducing the production of biofilms in Gram-negative pathogens. Herein we report the preparation of AHL analogs with a variety of N-substituents in just two steps from readily available N-substituted hydroxyproline esters. The substrates underwent an oxidative radical scission of the pyrrolidine ring. The resulting N-substituted β-aminoaldehyde underwent reduction and in situ cyclization to give a variety of homoserine lactones, with N- and N,N-substituted amino derivatives and with high optical purity. The libraries were screened for the inhibition of violacein production in Chromobacterium violaceum, a Gram-negative pathogen. For the first time, N,N-disubstituted AHL analogs were studied. Several N-sulfonyl derivatives, one carbamoyl, and one N-alkyl-N-sulfonyl homoserine lactone displayed a promising inhibitory activity. Moreover, they did not display microbicide action against S. aureus, C. jejuni, S. enterica, P. aeruginosa, and C. albicans, confirming a pure QQ activity. The determination of structure–activity relationships and in silico ADME studies are also reported, which are valuable for the design of next generations QQ agents. Full article

Quorum sensing (QS) is a cell-to-cell communication mechanism through which microorganisms can sense their population density and adjust their physiology by producing and detecting small signaling molecules called autoinducers (AIs). QS influences various aspects of microbial physiology, including virulence and pathogenesis by bacterial pathogens, biofilm formation, sporulation, antimicrobial resistance, etc. Lactic acid bacteria (LAB) have been used for centuries in food fermentation to improve sensory and nutritional profiles and preserve against spoilage and pathogenic microflora. This study investigated the potential of foodborne LAB of various genera, including Lactococcus, Lactobacillus, Leuconostoc, Streptococcus, and Enterococcus, to interfere with the QS system of bacterial pathogens. For this, cell-free supernatants (CFSs) of 89 LAB foodborne isolates were collected by centrifugation following a 20 h culture (at 30 °C) in quarter-strength Brain Heart Infusion (BHI) broth. The pH of all CFSs was adjusted to 6.5 and sterilized by filtration. The anti-QS activity of the sterilized and neutralized CFSs was initially screened using the biosensor strains Chromobacterium violaceum 026 and Agrobacterium tumefaciens NTL4 (pZLR4) through an agar well diffusion assay that can detect the inhibition of the QS system that is based on acylated homoserine lactones (AHLs), which are used as AIs by Gram-negative bacteria. Additionally, all the CFSs were also screened for interference with the autoinducer 2 (AI-2) QS system that is mostly used for interspecies communication by both Gram-positive and Gram-negative bacteria. This was assessed using a luminescence bioassay with the Vibrio harveyi BAA-1117 biosensor strain. The results indicate that none of the LAB CFSs could inhibit AHL-based QS. However, 61.8% (55/89) of the CFSs induced luminescence in V. harveyi BAA-1117, while the remaining 38.2% (34/89) of the samples were capable of inhibiting AI-2-based QS. In the next steps, the most representative of these latter AI-2 interfering LAB isolates will be investigated for possible inhibition of biofilm formation by some important foodborne bacterial pathogens. Full article

Quorum sensing, a bacterial cell-to-cell communication mechanism, plays a key role in bacterial virulence and biofilm formation. Targeting quorum-sensing pathways represents a promising strategy for the development of novel antibacterial agents. This study evaluated the anti-quorum-sensing activities of 18 natural compounds, including cannabinoids, arylbenzofurans, flavonoids, caffeine, and chlorogenic acid, using the luminescent biosensor strain Vibrio harveyi MM30. V. harveyi MM30, a mutant strain deficient in the production of autoinducer-2 (AI-2) but responsive to exogenous AI-2, was used to assess the activity of test compounds on the AI-2 receptor pathway. Test compounds were incubated in AI-2-containing media, and luminescence was measured to evaluate quorum-sensing inhibition. Comparisons were made in the absence of AI-2 to determine AI-2-independent inhibitory activity. The most active compounds were further tested on methicillin-resistant Staphylococcus aureus (MRSA 7112) to determine their effects on AI-2 production in spent media. Among the tested compounds, the non-prenylated arylbenzofuran moracin M and the prenylated arylbenzofuran moracin C exhibited significant quorum-sensing inhibitory activity in the AI-2-mediated pathway. None of the test compounds significantly inhibited quorum sensing in the absence of AI-2. Five compounds (cannabigerol, cannabidiol, cannabigerolic acid, moracin M, and moracin C) were selected for further investigation in MRSA 7112 cultures. The spent media from MRSA 7112 cultures treated with moracin M (16, 32, 64 µg/mL) and cannabigerolic acid (16 µg/mL) showed significant inhibition of AI-2 production when transferred to V. harveyi MM30 cultures. Moracin M and cannabigerolic acid demonstrated potential as quorum-sensing inhibitors by targeting AI-2 production and signalling pathways in MRSA 7112 and V. harveyi. These findings suggest their potential for further development as antibacterial agents targeting quorum-sensing mechanisms. Full article

Inflammatory bowel disease (IBD) is increasing among mammals around the world, and domestic dogs are no exception. There is no approved cure for canine IBD with limited treatment options. Novel probiotic bacteria discovery from free-ranging animals for the treatment of IBD in domestic pets can likely yield promising probiotic candidates. Consequently, the overall aim was to isolate bacteria from free-ranging animals that could potentially be utilized as novel probiotics. Two bacteria identified as unique Paenibacillus spp. strains by small ribosomal RNA (16S) gene sequencing were isolated from the gastrointestinal tract of a North American Gray Wolf (Canis lupus). The bacteria were typed as Gram-variable, and both were catalase/oxidase positive as well as sensitive to commonly used antibiotics. The bacteria digested complex carbohydrates and lipids by standard assays. The isolated bacteria also inhibited the growth of Staphylococcus aureus and Micrococcus luteus. The whole genome sequence (WGS) length of bacterial isolate ClWae17B was 6,939,193 bp, while ClWae19 was 7,032,512 bp, both similar in size to other Paenibacillus spp. The genomes of both bacteria encoded enzymes involved with the metabolism of complex starches and lipids, such as lyases and pectinases, along with encoding antimicrobials such as lanthipeptides, lasso peptides, and cyclic-lactone-autoinducers. No pernicious virulence genes were identified in the WGS of either bacterial isolate. Phylogenetically, the most closely related bacteria based on 16S gene sequences and WGS were P. taichungensis for ClWae17B and P. amylolyticus for ClWae19. WGS analyses and phenotypic assays supported the hypothesis that the isolates described constitute two novel candidate probiotic bacteria for potential use in dogs. Full article

Biofilms are complex, highly organized structures formed by microorganisms, with functional cell arrangements that allow for intricate communication. Severe clinical challenges occur when anaerobic bacterial species establish long-lasting infections, especially those involving biofilms. These infections can occur in device-related settings (e.g., implants) as well as in non-device-related conditions (e.g., inflammatory bowel disease). Within biofilms, bacterial cells communicate by producing and detecting extracellular signals, particularly through specific small signaling molecules known as autoinducers. These quorum-sensing signals are crucial in all steps of biofilm formation: initial adhesion, maturation, and dispersion, triggering gene expression that coordinates bacterial virulence factors, stimulates immune responses in host tissues, and contributes to antibiotic resistance development. Within anaerobic biofilms, bacteria communicate via quorum-sensing molecules such as N-Acyl homoserine lactones (AHLs), autoinducer-2 (AI-2), and antimicrobial molecules (autoinducing peptides, AIPs). To effectively combat pathogenic biofilms, understanding biofilm formation mechanisms and bacterial interactions is essential. The strategy to disrupt quorum sensing, termed quorum quenching, involves methods like inactivating or enzymatically degrading signaling molecules, competing with signaling molecules for binding sites, or noncompetitively binding to receptors, and blocking signal transduction pathways. In this review, we comprehensively analyzed the fundamental molecular mechanisms of quorum sensing in biofilms formed by anaerobic bacteria. We also highlight quorum quenching as a promising strategy to manage bacterial infections associated with anaerobic bacterial biofilms. Full article

Recently, we discovered and proved the essentiality of organic boron species (OBS), such as borate–pectic polysaccharides and borate–phenolic esters, for healthy symbiosis (HS) between microbiota and human/animal (H/A) host. The essentiality of OBS will provide new options for B supplementation in H/A nutrition for a healthy and long life. New knowledge on the essentiality of naturally occurring microbiota-accessible borate species for HS between microbiota and H/A host will allow the use of natural B-based dietary supplements to target the H/A microbiome (the gut, skin, oral, scalp, and vaginal microbiome). In the literature, there is evidence that certain bacteria need B (autoinducer-2 borate) for communication and our preliminary data show that HS takes place when the colonic mucus gel layer contains B. Subsequently, OBS become novel prebiotic candidates and target the colon as novel colonic foods. Full article

Background/Objective: The anti-restriction protein ArdA-H1, found in multiresistant Staphylococcus aureus (MRSA) strains from the ST239-SCCmecIII lineage, inhibits restriction–modification systems, fostering horizontal gene transfer (HGT) and supporting genetic adaptability and resistance. This study investigates the regulatory mechanisms controlling ardA-H1 expression in S. aureus under various stress conditions, including acidic pH, iron limitation, and vancomycin exposure, and explores the roles of the Agr quorum sensing system. Methods: The expression of ardA-H1 was analyzed in S. aureus strains exposed to environmental stressors using real-time quantitative reverse transcription PCR. Comparisons were made between Agr-functional and Agr-deficient strains. In addition, Agr inhibition was achieved using a heterologous Agr autoinducing peptide. Results: The Agr system upregulated ardA-H1 expression in acidic and iron-limited conditions. However, vancomycin induced ardA-H1 activation specifically in the Agr-deficient strain GV69, indicating that an alternative regulatory pathway controls ardA-H1 expression in the absence of agr. The vancomycin response in GV69 suggests that diminished quorum sensing may offer a survival advantage by promoting persistence and HGT-related adaptability. Conclusion: Overall, our findings provide new insights into the intricate relationships between quorum-sensing, stress responses, bacterial virulence, and genetic plasticity, enhancing our understanding of S. aureus adaptability in challenging environments. Full article

Bacterial quorum sensing (QS) is a critical communication process that regulates gene expression in response to population density, influencing activities such as biofilm formation, virulence, and antibiotic resistance. This study investigates the inhibitory effects of five phytochemicals—apigenin, carnosol, chlorogenic acid, quercetin, and rosmarinic acid—on the S-ribosylhomocysteinase (LuxS) enzyme, a key player in AI-2 signaling across both Gram-positive and Gram-negative bacteria. Using molecular docking studies, we identified that these phytochemicals interact with the LuxS enzyme, with apigenin, carnosol, chlorogenic acid, and rosmarinic acid binding within the substrate-binding pocket and exhibiting binding scores below −7.0 kcal/mol. Subsequent in vitro assays demonstrated that these compounds inhibited AI-2 signaling and biofilm formation in Escherichia coli MG1655 in a concentration-dependent manner. Notably, carnosol and chlorogenic acid showed the most potent effects, with IC₅₀ values of approximately 60 μM. These findings suggest that these phytochemicals may serve as potential QS inhibitors, providing a foundation for developing new anti-pathogenic agents to combat bacterial infections without promoting antibiotic resistance. Further studies are warranted to explore the therapeutic applications of these compounds in both clinical and agricultural settings. Full article

Background: Lactiplantibacillus plantarum is a species found in a wide range of ecological niches, including vegetables and dairy products, and it may occur naturally in the human gastrointestinal tract. The precise mechanisms underlying the beneficial properties of these microbes to their host remain obscure. Although Lactic acid bacteria are generally regarded as safe, there are rare cases of the emergence of infections and antibiotic resistance by certain probiotics. Objective: An in silico whole genome sequence analysis of putative probiotic bacteria was set up to identify strains, predict desirable functional properties, and identify potentially detrimental antibiotic resistance and virulence genes. Methods: We characterized the genomes of three L. plantarum strains (54B, 54C, and 55A) isolated from Ethiopian traditional cottage cheese. Whole-genome sequencing was performed using Illumina MiSeq sequencing. The completeness and quality of the genome of L. plantarum strains were assessed through CheckM. Results: Analyses results showed that L. plantarum 54B and 54C are closely related but different strains. The genomes studied did not harbor resistance and virulence factors. They had five classes of carbohydrate-active enzymes with several important functions. Cyclic lactone autoinducer, terpenes, Type III polyketide synthases, ribosomally synthesized and post-translationally modified peptides-like gene clusters, sactipeptides, and all genes required for riboflavin biosynthesis were identified, evidencing their promising probiotic properties. Six bacteriocin-like structures encoding genes were found in the genome of L. plantarum 55A. Conclusions: The lack of resistome and virulome and their previous functional capabilities suggest the potential applicability of these strains in food industries as bio-preservatives and in the prevention and/or treatment of infectious diseases. The results also provide insights into the probiotic potential and safety of these three strains and indicate avenues for further mechanistic studies using these isolates. Full article

Quorum sensing (QS) is a mechanism by which bacteria detect and respond to cell density, regulating collective behaviors. Burkholderia plantarii, the causal agent of rice seedling blight, employs the LuxIR-type QS system, common among Gram-negative bacteria, where LuxI-type synthase produces QS signals recognized by LuxR-type regulators to control gene expression. This study aimed to elucidate the QS mechanism in B. plantarii KACC18965. Through whole-genome analysis and autoinducer assays, the plaI gene, responsible for QS signal production, was identified. Motility assays confirmed that C8-homoserine lactone (C8-HSL) serves as the QS signal. Physiological experiments revealed that the QS-defective mutant exhibited reduced virulence, impaired swarming motility, and delayed biofilm formation compared to the wild type. Additionally, the QS mutant demonstrated weakened antibacterial activity against Escherichia coli and decreased phosphate solubilization. These findings indicate that QS in B. plantarii significantly influences various pathogenicity and survival traits, including motility, biofilm formation, antibacterial activity, and nutrient acquisition, highlighting the critical role of QS in pathogen virulence and adaptability. Full article

Pseudomonas aeruginosa is an opportunistic Gram-negative bacterium that remains a prevalent clinical and environmental challenge. Quorum-sensing (QS) molecules are effective biomarkers in pinpointing the presence of P. aeruginosa. This study aimed to develop a convenient-to-use, whole-cell biosensor using P. aeruginosa reporters individually encapsulated within alginate-poly-L-lysine (alginate-PLL) microbeads to specifically detect the presence of bacterial autoinducers. The PLL-reinforced microbeads were prepared using a two-step method involving ionic cross-linking and subsequent coating with thin layers of PLL. The alginate-PLL beads showed good stability in the presence of a known cation scavenger (sodium citrate), which typically limits the widespread applications of calcium alginate. In media containing synthetic autoinducers—such as N-(3-oxo dodecanoyl) homoserine lactone (3-oxo-C₁₂-HSL) and N-butanoyl-L-homoserine lactone (C₄-HSL), or the cell-free supernatants of planktonic or the flow-cell biofilm effluent of wild P. aeruginosa (PAO1)—the encapsulated bacteria enabled a dose-dependent detection of the presence of these QS molecules. The prepared bioreporter beads remained stable during prolonged storage at 4 and −80 °C and were ready for on-the-spot sensing without the need for recovery. The proof-of-concept, optical fiber-based, and whole-cell biosensor developed here demonstrates the practicality of the encapsulated bioreporter for bacterial detection based on specific QS molecules. Full article