New insights are provided into the physiological role of azurin in environmental bacteria and its involvement in bacterium–eukaryote interactions is suggested, thereby opening new perspectives for biotechnological and biomedical applications.
Abstract
Azurin is a promising antitumor agent that selectively enters cancer cells and inhibits tumor progression. It is also known to participate in cellular processes involving single-electron transfer, including protection against oxidative stress, anaerobic respiration, and denitrification. However, the physiological role of azurin remains poorly understood. In this work, a multifaceted phenotypic characterization of an azurin-deficient mutant (Δazu) of the plant endophytic bacterium Pseudomonas sp. OHS18 was performed, using complementary approaches and technologies. Deletion of the azu gene did not affect resistance to antibiotics, copper, or hydrogen peroxide, while nuclear magnetic resonance-based metabolomic analysis revealed that the Δazu strain was moderately impaired in maintaining metabolic homeostasis from the exponential to the stationary growth phase. Phenotype microarray analyses showed that the two strains exhibited largely similar metabolic and resistance profiles, except for bromosuccinic acid utilization, under which the Δazu strain displayed reduced growth. This phenotype was further associated with a reduced ability of the mutant to colonize Arabidopsis thaliana, suggesting a role for azurin in maintaining the plant–bacterium association. Overall, these findings provide new insights into the physiological role of azurin in environmental bacteria and suggest its involvement in bacterium–eukaryote interactions, thereby opening new perspectives for biotechnological and biomedical applications.
Plant-associated Pseudomonas species are widely recognised for their ecological versatility and potential as sustainable biocontrol and plant growth-promoting agents. Among their diverse metabolites, hydrogen cyanide (HCN) is traditionally considered a potent antimicrobial compound, yet its broader role in plant-microbe interactions remains underexplored. This study investigated Pseudomonas urmiensis A32, an endophytic bacterium previously shown to enhance drought tolerance and disease resistance in pepper (Capsicum annuum). Whole-genome analysis revealed genes involved in nutrient acquisition, stress protection, and competitive exclusion, including secondary metabolite clusters for carotenoid and HCN biosynthesis. Wild-type A32 showed increased tolerance to oxidative stress, highlighting a redox-active role for HCN. Seed inoculation with A32 reduced disease symptoms and pathogen load in pepper plants challenged with Xanthomonas euvesicatoria, indicating that HCN contributes not only to direct pathogen antagonism but also to modulation of host immune responses. Biochemical analyses showed decreased activity of antioxidant enzymes and increased phenylalanine ammonia-lyase activity. Gene expression analyses revealed upregulation of PR1 and the antimicrobial peptide gene DEF1, indicating enhanced activation of defence genes. These findings demonstrate that HCN acts as a multifunctional metabolite that fine-tunes the balance between reactive oxygen species detoxification and signalling, and integrates microbial biocontrol with host immune priming. In summary, this work highlights the ecological adaptability of P. urmiensis A32 and its potential as an effective bioinoculant for sustainable crop protection.
Aleksandra Mesaroš, Marija Nedeljković, Iva Atanasković et al.· Microbiology Research· 0 citations
Endophytic fungi are a reservoir for cryptic metabolites to discover new bioactive compounds through their research on inactive biosynthetic gene clusters. In this study, Aspergillus sp. EGP214 was used to identify Aspergillus sp. EGP214, which they isolated from Hyoscyamus muticus during their study. The expression of cryptic genes needed stimulation through the use of two epigenetic modifiers, which included trichostatin A as a histone deacetylase inhibitor and 5-aza-2'-deoxycytidine as a DNA methyltransferase inhibitor, during fermentation. GC-MS analysis revealed that both treatments significantly altered the metabolic profiles, leading to the production of several unique compounds absent in the control culture. The treated extracts showed improved antimicrobial, antibiofilm, antioxidant, and DNA gyrase inhibitory effects with trichostatin A, treated cultures demonstrating the strongest activity at IC50 = 2.2 μM against DNA Gyrase-B. The induced metabolites demonstrated permanent strong bonds to bacterial DNA gyrase and HDAC enzymes' active sites according to molecular docking and molecular dynamics simulations. The evaluation of ADMET characteristics and toxicity levels showed moderate lipophilicity, together with minimal systemic toxicity but restricted oral bioavailability. The research shows that epigenetic modulation serves as an effective method to activate dormant fungal biosynthetic pathways, which produce valuable secondary compounds with antimicrobial and antioxidant properties.
A. Abd-elaziz, Mai A.M.A. Mwaheb, Gehad H. El Sayed et al.· Microbial Pathogenesis· 0 citations
INTRODUCTION
Lysobacter enzymogenes OH11 is widely recognized for its potent antagonistic activity against diverse plant pathogens, including bacteria, fungi, and oomycetes. However, the specific compound responsible for its activity against Gram-negative bacteria had remained unidentified.
OBJECTIVES
This study aimed to identify the specific anti-Gram-negative bacterial compound produced by OH11 and to evaluate its potential as a biocontrol agent.
METHODS
Methods included MS/NMR for structure elucidation, antagonism/greenhouse/UV-vis assays for function evaluation, medium/precursor optimization for yield enhancement, and gene knockout/enzyme assays for biosynthetic pathway identification.
RESULTS
The isolated compound was identified as a spermidine-containing siderophore, designated Sperbactin. Through broad-spectrum screening, Sperbactin was discovered to exert highly targeted antagonistic effects specifically against rice pathogens (Xanthomonas oryzae). In greenhouse trials, Sperbactin demonstrated remarkable efficacy against rice bacterial blight, achieving a disease control efficacy of 85.31%, which is comparable to conventional chemical agents. The Sperbactin-Fe(III) complex was found to have a 1:1 M ratio and displayed a high affinity for ferric iron, with an association constant (Ka) of 2.16 × 1029 M-1. Furthermore, the fermentation yield of Sperbactin was increased by 3.29-fold, reaching 290.46 ± 6.22 mg/L. spbB and spbD were identified as essential genes for Sperbactin biosynthesis.
CONCLUSION
This study identifies Sperbactin as a highly promising, narrow-spectrum biocontrol candidate for the targeted management of rice bacterial diseases. Its development offers a sustainable and eco-friendly alternative to chemical pesticides, supporting the advancement of sustainable agriculture.
B. Tang, P. Laborda, Xian Chen et al.· Journal of Advanced Research· 0 citations
The development of green alternatives to chemical nematicides is an urgent need in sustainable agriculture. This study constructed an engineered strain, LG209, by overexpressing the transcriptional regulator OmpR, which significantly enhanced nematicidal activity against Meloidogyne incognita J2s and increased secondary metabolite production. From its fermentation broth, 20 compounds were isolated, including a new naphthoquinone derivative. Notably, phenylacetic acid (3), 2-(4-methoxyphenyl)acetic acid (5), and 9H-carbazole (18) exhibited strong nematicidal activity, with LC50 values of 52.34, 88.52, and 15.18 μg/mL at 96 h, respectively. These compounds induced ROS burst and lipid peroxidation, leading to necrotic cell death in nematodes. Additionally, certain indole derivatives modulated nematode chemotaxis. In soil trials, LG209 fermentation broth achieved 68.9% control efficacy against root-knot nematode in tomato, surpassing the wild-type strain. This work provides a promising engineered strain and lead compounds for developing novel microbial nematicides.
Yue-Yu Ma, Jin-Fang Li, Jian Wu et al.· Journal of Agricultural and...· 0 citations
ABSTRACT Flavonoids are plant-derived polyphenols with diverse biological activities; however, the mechanisms by which individual compounds inhibit intestinal pathogens remain unclear. In this study, we found that taxifolin (dihydroquercetin) showed pronounced inhibitory activity against Clostridium perfringens, a major gram-positive enteropathogen, whereas most other bacterial species tested were less affected. Notably, at concentrations below those causing complete growth inhibition, taxifolin significantly impaired mucin adhesion and biofilm formation, indicating suppression of colonization-associated phenotypes rather than bactericidal activity. In an antibiotic-pretreated mouse infection model, oral administration of taxifolin significantly reduced C. perfringens intestinal colonization during the early phase of infection. Mechanistically, proteomic analysis of C. perfringens revealed marked alterations in surface-associated and extracellular proteins, including stress response factors (MreB, LytR, and ClpB), and enzymes linked to iron- and redox-dependent metabolism, consistent with an iron-limiting stress response. Scanning electron microscopy further revealed pronounced cell elongation in taxifolin-treated cells, consistent with impaired cell division under stress conditions. Importantly, Fe2+ supplementation partially restored bacterial growth, normal morphology, mucin adhesion, and biofilm formation, whereas supplementation with other divalent ions failed to restore growth. Similar phenotypes were observed following treatment with the iron-specific chelator 2,2′-bipyridyl. Collectively, our findings support a working model in which iron limitation-associated physiological stress contributes to the inhibitory effects of taxifolin on C. perfringens growth and colonization-related phenotypes. This study provides insights into the interaction between dietary flavonoids and clostridial pathogens, highlighting the potential of taxifolin as a plant-derived compound for limiting C. perfringens intestinal colonization. IMPORTANCE Clostridium perfringens, a clinically important pathogen in both humans and animals, causes various histotoxic and enteric diseases, including gas gangrene and foodborne or non-foodborne diarrhea. Its control remains challenging due to increasing antimicrobial resistance and the need to preserve beneficial gut microbiota. Here, we identified taxifolin, a naturally occurring flavonoid, as a potential inhibitor of C. perfringens that suppressed its growth and colonization-associated phenotypes while exerting comparatively limited effects on most other tested bacterial species, including commensal bacteria. Proteomic and physiological analyses revealed stress responses associated with iron metabolism, redox balance, and cell division, which were accompanied by reduced mucin adhesion, biofilm formation, and bacterial proliferation. Our findings suggest that taxifolin can reduce the colonization capacity of C. perfringens and support its potential use as a plant-derived approach for limiting C. perfringens intestinal colonization in humans and animals. Clostridium perfringens, a clinically important pathogen in both humans and animals, causes various histotoxic and enteric diseases, including gas gangrene and foodborne or non-foodborne diarrhea. Its control remains challenging due to increasing antimicrobial resistance and the need to preserve beneficial gut microbiota. Here, we identified taxifolin, a naturally occurring flavonoid, as a potential inhibitor of C. perfringens that suppressed its growth and colonization-associated phenotypes while exerting comparatively limited effects on most other tested bacterial species, including commensal bacteria. Proteomic and physiological analyses revealed stress responses associated with iron metabolism, redox balance, and cell division, which were accompanied by reduced mucin adhesion, biofilm formation, and bacterial proliferation. Our findings suggest that taxifolin can reduce the colonization capacity of C. perfringens and support its potential use as a plant-derived approach for limiting C. perfringens intestinal colonization in humans and animals.
H. Fadhilatunnur, Weichen Gong, Haruna Sato et al.· Microbiology spectrum· 0 citations
Endophytic fungi can produce metabolites with potential pharmaceutical applications through biosynthetic pathways strongly influenced by culture conditions. In this study, growth parameters (media composition and cultivation time) were systematically optimized to maximize 3-O-methylfunicone (3OMF) production by the endophytic fungus Talaromyces pinophilus J6. Chemical profiling by high-resolution mass spectrometry (HRMS) demonstrated that supplementation of potato dextrose agar (PDA) with 45% (w/w) ammonium sulfate and cultivation for 7 days significantly increased the accumulation of 3OMF compared to standard growth conditions. The optimized culture condition enabled the isolation of the target compound in sufficient yield for subsequent biological assays. The compound exhibited anti-Helicobacter pylori activity against one reference strain and three clinical isolates, with minimum inhibitory concentrations ranging from 0.165 to 0.660 mM and bactericidal concentrations up to 1.32 mM. In combination assays, 3OMF showed an additive effect with clarithromycin, indicating potential as an adjuvant compound in eradication therapies. Cytotoxicity assays of 3OMF demonstrated selective activity against gastric adenocarcinoma cells, with an IC50 of 0.24 mM and selectivity index of 2.37 relative to fibroblasts. Molecular docking suggested favorable interactions between 3OMF and H. pylori β-clamp protein, suggesting a possible association with interference in bacterial DNA replication. These findings demonstrate that culture optimization is an efficient biotechnological strategy to improve 3OMF production and support the further investigation of this compound as a scaffold for anti-H. pylori applications.
M. Marques, Dalila N. Loose, Crislaine S. Lima et al.· Archives of Microbiology· 1 citation