Objective To identify the key amino acid residues of the TetR family transcription factor BcPDR1 in Botrytis cinerea, thereby laying a foundation for elucidating the mechanism by which BcPDR1 regulates the growth, development, and pathogenicity of this pathogen. Methods The key amino acid sites of BcPDR1 were analyzed by bioinformatics methods, and four conserved regions (32-34 aa, 76-95 aa, 140-150 aa, and 189 aa) were selected for site-directed mutagenesis. On the basis of the knockout mutant ΔBcpdr1, the mutants BcPDR1-M1 (Δ32-34), BcPDR1-M2 (Δ76-95), BcPDR1-M3 (Δ140-150), and BcPDR1-M4 (mutation of Ile to Lys at 189 aa) were constructed. A comparative analysis of the phenotypic characteristics and pathogenicity was conducted on the four aforementioned mutants and the wild-type strain of B. cinerea, ΔBcpdr1, the complemented strain CE. Results The colony morphology, mycelial morphology, and growth rates of BcPDR1-M1, BcPDR1-M2, BcPDR1-M3, and BcPDR1-M4 were similar to those of ΔBcpdr1, but significantly different from those of BC22 and CE. These mutants could form lesions on tomato fruits and tobacco leaves, while their lesion areas were significantly smaller than those of BC22 and CE. Conclusion The regions 32-34, 76-95, 140-150, and the 189th amino acid are the regulatory sites for BcPDR1 to exert its functions.

Probiotic additives for feed play a crucial role in maintaining the health and improving the production performance of livestock and poultry. However, the application of most probiotics is limited by their sensitivity to environmental stresses (e.g., acid, bile salt, and temperature) in the animal intestinal tract, and microencapsulation serves as a key approach to enhance their stability. Objective This study constructed a metal-polyphenol-prebiotic (Fe-TA-GN) composite coating system for the probiotic strain Enterococcus faecium PL84 isolated by us and verified its protective effect on PL84, aiming to provide technical support for the industrial application of the strain. Methods Coating parameters (Fe³⁺-TA molar ratio and GN concentration) were optimized. Scanning electron microscopy (SEM) and the CCK-8 assay were employed to evaluate the effects of coating materials on the viability of mouse intestinal epithelial cells (IEC-6). The protective effect of the coating system was assessed through in vitro tolerance tests under acidic, bile salt, thermal conditions, as well as in simulated gastrointestinal fluids. Results E. faecium PL84 exhibited the highest cell viability during the logarithmic growth phase, being suitable for microencapsulation. When the molar ratio of Fe³⁺ to TA was 1:3, the PL84-Fe-TA composite particles showed the smallest nanoscale particle size and formed a dense metal-polyphenol network. At a GN concentration of 0.4 mg/mL, the Fe-TA-GN coating layer achieved the highest zeta potential and optimal structural stability. SEM revealed a uniform and continuous surface coating layer of PL84-Fe-TA-GN. In vitro tolerance assays demonstrated that the survival rate of PL84-Fe-TA-GN was higher than that of uncoated PL84 under conditions of pH 3.0 and 0.6% bile salt (P<0.01). After treatment at 60 ℃, the survival rate of the coated strain increased by 16.29% compared with that of uncoated PL84. Additionally, the survival rates of PL84-Fe-TA-GN in simulated gastric fluid and simulated intestinal fluid improved by 20.8% and 13.53%, respectively. The coating materials (Fe-TA, GN, and Fe-TA-GN) had no significant effect on the viability of PL84 (P>0.05). Conclusion When the molar ratio of Fe³⁺ to TA is 1:3 and the GN concentration is 0.4 mg/mL, the metal-polyphenol-prebiotic composite coating system is stable and can significantly enhance the environmental tolerance of E. faecium PL84. Moreover, the coating materials possess good biocompatibility, laying a solid technical foundation for the industrial application of E. faecium PL84.

[Objective] By examining intracellular and extracellular metabolite changes in ectomycorrhizal fungi (ECMF) under acidic aluminum stress, we identified key resistance-related metabolites and pathways, aiming to elucidate the aluminum tolerance mechanisms from the perspective of metabolic physiology and offer a theoretical basis for using ECMF in restoring aluminum-contaminated forests. [Methods] Pisolithus tinctorius was cultured in vitro in the acidic medium (pH 3.8) containing 0.0 mmol/L or 1.0 mmol/L Al³⁺. Untargeted metabolomics was employed to analyze changes in intracellular and extracellular metabolite levels. [Results] Compared with that under the 0.0 mmol/L Al³⁺ treatment, the colony diameter of P. tinctorius under 1.0 mmol/L Al³⁺ stress decreased significantly by 23.67%. In addition, the intracellular levels of nucleotides including uridylic acid, cytidine monophosphate, uridine, uridine diphosphate, cytidine, and guanosine were upregulated under 1.0 mmol/L Al³⁺ stress. Extracellular levels of organic acids such as shikimic acid, fumaric acid, heptanoic acid, and tartaric acid, along with carbohydrates including l-arabinose, trehalose, sucrose, and glucose, were also upregulated. Pyrimidine metabolism and citric acid cycle pathways were enriched intracellularly, while ABC transporters and phosphotransferase system pathways were enriched extracellularly. The potential biomarkers identified in the intracellular environment was citric acid, and those identified in the extracellular environment were trehalose and tartaric acid. [Conclusion] Acidic aluminum stress inhibits the growth of P. tinctorius. Intracellularly, P. tinctorius maintains cellular homeostasis and energy supply through enhanced nucleotide accumulation and activation of the citric acid cycle. Extracellularly, P. tinctorius promotes organic acid secretion and carbohydrate efflux to resist aluminum toxicity and associated oxidative damage.

[Objective] To investigate the regulatory effect of the cAMP receptor protein (CRP), a transcription factor in Acinetobacter baumannii, on the cas3 gene. [Methods] CRP was expressed and purified via a prokaryotic expression system. EMSA was employed to examine CRP binding to the cas3 promoter. qPCR was conducted to evaluate the regulatory effect of CRP on cas3 expression. To further confirm the regulatory function of CRP, we constructed a mutant strain Δcrp. The impact of crp deletion on A. baumannii virulence was then analyzed via the biofilm formation assay, adhesion and invasion assays with A549 cells, a Galleria mellonella model, and a murine model of bacterial infection. [Results] EMSA demonstrated that CRP specifically bound to the cas3 promoter. The qPCR results showed that cas3 transcription was downregulated (P<0.001) in Δcrp. Compared with the wild-type strain, Δcrp exhibited no significant difference in growth capacity but enhanced biofilm formation (P<0.001) as well as strengthened adhesion (P=0.003<0.050) and invasion (P<0.001) in A549 cells. Furthermore, Δcrp demonstrated a markedly increased lethality rate in G. mellonella within 72 h. Furthermore, the murine infection experiment revealed that Δcrp possessed higher colonization capacity in the lungs than the wild-type strain (P<0.001). [Conclusion] CRP acts as a transcriptional activator that directly binds to the cas3 promoter to activate its transcription, thereby attenuating the virulence and pathogenicity of A. baumannii.

[Objective] The type VI secretion system (T6SS) is a novel virulence factor of Edwardsiella tarda. E. tarda virulence protein P (EvpP) is an effector of the T6SS. To date, the functional mechanisms of EvpP are still poorly understood. This study aimed to comprehensively investigate the biological functions of EvpP and elucidate the roles of T6SS in the pathogenicity of E. tarda. [Methods] We constructed the evpP-deleted mutant (ΔevpP) and complemented strain (ΔevpP-C) to study the effects of evpP deletion on the biological characteristics of E. tarda and infection in macrophages. [Results] No significant differences were observed in the growth curves or physiological and biochemical properties among the wild-type (WT), ΔevpP, and ΔevpP-C. However, compared with WT, ΔevpP exhibited significantly reduced motility, biofilm formation, adhesion rate to RAW264.7 macrophages, intracellular proliferation capacity, and ability to induce host cell autophagy, while triggering increased secretion of tumor necrosis factor-α (TNF-α) by macrophages. The complementation of evpP-C did not fully restore the intracellular proliferation capability, but completely rescued the other phenotypic defects. [Conclusion] EvpP does not affect the growth or physiological and biochemical properties of E. tarda. However, it could enhance the bacterial motility, biofilm formation, adhesion to macrophages, intracellular proliferation, and autophagy, while suppressing TNF-α secretion in E. tarda-infected macrophages. These findings confirm that EvpP plays a critical role in the pathogenicity of E. tarda.

Soil colloidal phosphorus (CP) is an active component that cannot be overlooked in the soil phosphorus cycle. Its occurrence forms and migration behavior significantly influence phosphorus bioavailability and environmental risks. This paper systematically reviews the multiscale regulatory mechanisms of microbial actions in CP transformation and migration. It focuses on chemical effects (e.g., proton secretion, iron reduction, and organic acid coordination), physical effects (e.g., extracellular polymer trapping and biofilm pore remodeling), and microbial community dynamics and molecular ecology three dimensions to elucidate how microbes drive CP activation, immobilization, and migration through interfacial reactions, functional gene expression, and community interactions. The paper further explores the synergistic effects of multiple factors on microbial regulation processes, including soil physicochemical properties, agricultural management practices, and emerging pollutants. It identifies current research gaps in cross-scale coupling, in situ characterization, and mechanism modeling, while providing theoretical foundations and research directions for enhancing soil phosphorus utilization and pollution control.

[Objective] To explore the influence of Acinetobacter pittii LSQ 3 on the biofilm formation of Bacillus velezensis LSQ 19 and analyze the genomic characteristics of strain LSQ 3. [Methods] The effect of the cell-free supernatant (CFS) of LSQ 3 on the biofilm formation of LSQ 19 was analyzed by crystal violet staining, cell surface property analysis, the phenol-sulfuric acid method, XTT reduction assay, and scanning electron microscopy (SEM). Whole-genome sequencing was employed to determine the taxonomic status of strain LSQ 3, and the biosynthetic gene clusters for secondary metabolites were predicted based on the whole-genome data. [Results] The CFS of strain LSQ 3 significantly inhibited the biofilm formation of strain LSQ 19, and the volume ratio of 10 µL bacterial suspension to 190 µL CFS was determined as the minimum inhibitory concentration (MIC). Compared with the control, the CFS of strain LSQ 3 at MIC significantly reduced the surface hydrophobicity, adhesion, extracellular polymeric substances (EPS) production rate, and biofilm metabolic activity, while significantly improving the self-aggregation ability of LSQ 19 cells. In the presence of the CFS at MIC, LSQ 19 failed to form a biofilm on the glass surface. Strain LSQ 3 was identified as A. pittii based on whole-genome sequencing data. Its genome size was 3 939 365 bp, with the G+C content of 38.82% and 3 601 DNA coding sequences. The genome contained multiple genes involved in biofilm formation and virulence factors. The antiSMASH analysis showed that the genome of strain LSQ 3 contained seven biosynthetic gene clusters for secondary metabolites. [Conclusion] The CFS of A. pittii LSQ 3 can inhibit the biofilm formation of B. velezensis LSQ 19. This study provides a theoretical basis and reference for the construction of synthetic bacterial communities from the perspective of biofilms.

As a subterranean, soil-dwelling insect, Odontotermes formosanus is susceptible to infection by diverse soil-borne pathogenic fungi. Its symbiotic actinobacteria produce bioactive compounds to combat these pathogens, thereby maintaining a stable symbiotic system of O. formosanus with Termitomyces spp. [Objective] To screen and characterize antifungal metabolites from symbiotic actinobacteria of O. formosanus and to elucidate their defensive role in the termite-fungus symbiotic system. [Methods] Actinobacteria were isolated from O. formosanus-associated samples via diverse culture media. Antifungal strains were screened via dual-culture confrontation assays. The active strain OFGS46 was selected for whole-genome sequencing. Biosynthetic gene clusters (BGCs) were predicted by antiSMASH, and secondary metabolites were analyzed by HPLC-MS. [Results] Twenty-three actinobacteria strains were isolated. Strain OFGS46 exhibited inhibition rates of (62.05±0.98)% against Xylaria sp. and (79.99±0.58)% against Talaromyces sp. Whole-genome sequencing on the Illumina NovaSeq platform yielded approximately 1 Gb of high-quality data. De novo assembly generated a draft genome of 8 470 479 bp. CheckM evaluation demonstrated high assembly quality with 99.91% completeness and 4.63% contamination. HPLC-MS results revealed that strain OFGS46 was capable of producing multiple bioactive secondary metabolites, including enduracidin A, WS9326A, kitacinnamycin A, WAP-8294A2, skyllamycin A, cahuitamycin A, pentamycin, scabichelin, coprisamide C, and frankobactin A1. [Conclusion] This study systematically reveals that the symbiotic actinobacterium OFGS46 from O. formosanus suppresses nest pathogens through the production of diverse antimicrobial compounds. These findings not only elucidate, from a perspective of chemical ecology, the role of symbiotic bacteria in host immune defenses through suppressing multiple pathogenic fungi, but also provide a scientific basis for developing novel antimicrobial resources derived from the termite symbiotic system.

Antibiotic resistance (AR) in microorganisms has become a crucial challenge to global public health security. The acquirement of AR in microorganisms is often accompanied by a fitness cost. Typically, in an environment without antibiotics, the bacterial community with AR shows weaker competitiveness than susceptible bacteria, which makes antibiotic resistance genes (ARGs) a burden for bacteria. This article elaborates on the mechanisms underlying the generation of fitness costs and the corresponding measurement methods, provides examples to illustrate the fitness costs mediated by ARGs under different acquisition methods, and introduces the differences in fitness costs between chromosome-mediated and plasmid-mediated ARGs as well as their molecular mechanisms. Finally, it proposes prevention and control strategies for antibiotic resistant bacteria (ARB) based on fitness costs. This article reveals the biological law of the trade-off between antibiotic resistance and bacterial fitness and provides ideas for preventing and controlling the global public health security issues caused by ARB and ARGs.

The gut structure and functional performance rely on the stable type and quantity of gut mucosal epithelial cells, whose stability depends on continuous proliferation, differentiation, and migration of intestinal stem cells (ISCs) located in the crypts. Interactions between gut microbiota and ISCs support the homeostasis of the gut ecosystem. Intestinal epithelial cells (IECs) differentiated from ISCs influence the composition and function of gut microbiota through secreting immunoglobulins, mucins, etc. Meanwhile, the microorganism-associated molecular patterns (such as lipoteichoic acid and lipopolysaccharides) and metabolites (such as short-chain fatty acids and bile acids) from gut microbiota form the unique gut microenvironment to regulate the activity of ISCs and the homeostasis of IECs. Regulating the activity of ISCs and gut health by modifying gut microbiota has become a focus of current research. Thus, this review elaborates on the impact of ISCs on the gut microbiota as well as the regulatory roles of gut microbiota and related metabolites on the proliferation and differentiation fate of ISCs, aiming to broaden the understanding of the interaction between gut microbiota and ISCs. It is expected to provide strategies and targets for the regulation on gut health.