Bovine mastitis is a key factor restricting the high-quality development of China’s dairy industry, while the prevalence of antimicrobial-resistant bacteria and recurrent infections have become severe challenges for the current prevention and control system. Although long-term antibiotic selection pressure may temporarily alleviate clinical symptoms, it significantly accelerates the evolution and dissemination of multidrug-resistant (MDR) pathogens, leading to rising treatment failure rates and a transition toward chronic disease. Epidemiological data indicate that the pathogen spectrum of bovine mastitis in China is dominated by Staphylococcus aureus, Streptococcus spp., and Enterobacteriaceae, with resistance genes propagating across species and regions through horizontal gene transfer (HGT) mediated by plasmids, transposons, and integrons. Furthermore, mechanisms such as biofilm formation, efflux pump activation, target modification, and intracellular escape interact with the mammary microenvironment to establish a robust defense barrier against host immune clearance and antimicrobial agents. Incorporating the principles of endogenous inflammation and metabolic dysregulation mediated by the “gut-mammary axis”, this paper systematically reviews the regional prevalence and molecular dissemination mechanisms of resistant pathogens in China. It provides an in-depth analysis of the pathological basis of recurrence and prospectively proposes comprehensive management strategies—ranging from precision diagnosis to alternative therapies—aiming to provide a theoretical foundation for the scientific control of bovine mastitis.

Alzheimer’s disease (AD) is a complex neurodegenerative disorder characterized by progressive cognitive impairment. In recent years, the “gut-brain axis” has been recognized to play an important role in the pathogenesis of AD, with the gut microbiota-derived metabolites serving as key mediators of gut-brain communication. This review systematically summarizes the alterations and underlying molecular mechanisms of five classes of key metabolites—short-chain fatty acids, bile acids, indole derivatives, trimethylamine N-oxide, and lipopolysaccharides—during the progression of AD. This review not only provides new perspectives for understanding the pathological processes of AD but also lays a theoretical foundation for the development of diagnostic and therapeutic strategies based on microbiota modulation.

Neurodegenerative diseases, a group of highly prevalent central nervous system disorders, are closely linked to gut microbiota dysbiosis and impaired gut-brain axis function. The emerging gut-brain axis theory offers a novel perspective to explain how microbes exert remote, cross-organ regulation over the central nervous system. Recent studies have indicated that butyrate-producing bacteria, a core functional component of the gut microbiota, exhibit dynamic changes temporally and spatially correlated with the onset and progression of these diseases. These bacteria are thought to modulate the central nervous system microenvironment and disease pathology via the gut-brain axis. This article systematically reviews the bidirectional signaling mechanisms between the gut microbiota and the brain, analyzing the complex, multidimensional relationship between butyrate-producing bacteria and neurodegenerative diseases. This analysis encompasses population heterogeneity in patients with diverse clinical features and the dynamic evolution of these bacterial communities across different disease stages. We summarize the key mechanisms by which butyrate-producing bacteria regulate disease progression, including barrier protection, immunomodulation, metabolic regulation, and epigenetic modification. Furthermore, we explore their clinical potential as predictive biomarkers and therapeutic targets. We propose that future research should prioritize the development of targeted intervention strategies for gut-derived butyrate-producing bacteria. This review aims to provide a theoretical foundation and novel insights to advance both the fundamental investigation and clinical translation of these bacteria in the context of neurodegenerative diseases.

Coronavirus infections pose a serious threat to human health and have resulted in substantial economic losses to the livestock industry. Coronaviruses invade host cells primarily through two pathways: cell surface membrane fusion and endosomal membrane fusion. During viral entry, the spike protein subunit 1 (S1) recognizes and binds to cellular receptors, while the spike protein subunit 2 (S2) facilitates membrane fusion between the viral envelope and host cell membrane. Due to its high sequence conservation across different coronaviruses, S2 represents an attractive target for the development of broad-spectrum antiviral agents. Blocking S2-mediated membrane fusion can effectively inhibit viral infection. This review summarizes recent advances in understanding the mechanisms of coronavirus entry into host cells, the structure and function of the spike protein, and the development of membrane fusion inhibitors. In addition, this paper discusses the challenges and future prospects in targeting S2 for antiviral drug development, aiming to provide insights for coronavirus prevention and the discovery of novel antiviral therapeutics.

Salicylic acid (SA) is an important phenolic compound that plays a key role in plant defenses and is widely used in pharmaceuticals, cosmetics, and personal care products due to its significant anti-inflammatory and antimicrobial activities. Currently, the production of SA mainly relies on plant extraction and chemical synthesis, which suffers from complex processes, severe environmental pollution, and high dependence on petrochemical resources. With the rapid development of synthetic biology, metabolic engineering, and artificial intelligence (AI) technologies, the green synthesis of SA through intelligently designed microbial cell factories, empowered by machine learning algorithms and automated platforms, has become an important research direction to replace conventional production methods. This review systematically summarizes the microbial biosynthetic pathways of SA. With a focus on the intelligent design theme, this paper highlights the application of AI and synthetic biology tools in the discovery and utilization of natural SA-producing microbial resources and the rational reconstruction and optimization of the SA biosynthetic pathway in model microorganisms via intelligent metabolic engineering strategies. Furthermore, it introduces the key intelligent technologies for enhancing yields and the challenges faced. Finally, it discusses the future trends in this field.

Streptococcus suis is a major zoonotic pathogen that can infect both pigs and humans, causing severe diseases such as meningitis in humans. Its pathogenicity depends on the ability to rapidly adapt to environmental stress and host immune responses. Serine/threonine kinases and their corresponding phosphatases constitute a eukaryotic-like signal transduction system in bacteria and play a key regulatory role in S. suis serotype 2, closely related to its biological characteristics and pathogenic mechanisms. Through precise regulation of the phosphorylation and dephosphorylation of downstream substrates, serine/threonine kinases/phosphatases form complex signaling networks, thereby influencing various physiological and pathogenic processes of the bacterium. This article systematically reviews the currently known substrates of serine/threonine kinases in S. suis serotype 2, with a focus on elucidating how these kinases precisely regulate bacterial growth and division, capsule synthesis, stress tolerance, adhesion, invasion, and pathogenicity by modulating the phosphorylation status of functional substrates. This review aims to provide new perspectives for deciphering the pathogenic mechanism of S. suis serotype 2 and the development of novel antibacterial strategies.

Rice is one of the most widely cultivated and highest-yielding crops worldwide, and its yield stability is closely linked to global food security. As a typical mycorrhizal crop, rice exhibits enhanced growth and stress tolerance after colonization of arbuscular mycorrhizal fungi (AMF). Rhizosphere bacteria, a major component of the rhizosphere microbiome, interact synergistically with AMF, collectively contributing to enhanced nutrient acquisition and improved rice performance. This review summarizes recent advances in the interactions between AMF and rhizosphere bacteria. We highlight that their synergistic interactions regulate rice growth, nutrient acquisition, and stress tolerance, which contribute to the ecosystem stability and biodiversity of the rhizosphere. However, most available studies on the interactions between AMF and rhizosphere bacteria have been conducted under controlled conditions, which limits their applicability to the complex natural field environments. Therefore, this review proposes several suggestions for future research. First, utilize multi-omics technologies such as metagenomics and isotope tracing techniques to elucidate the molecular mechanisms underlying the AMF-bacterial interaction. Second, in practical applications, long-term field positioning trials should be conducted to screen out superior microbial agents that meet the requirements. Finally, optimize regulatory conditions to achieve large-scale propagation of AMF and overcome the bottleneck in AMF field propagation provides theoretical support for promoting the practical application of this technology in agricultural production.

Dimorphic prosthecate bacteria (DPB) are a group of prokaryotes that reproduce through asymmetric division, producing two morphologically and functionally highly differentiated daughter cells. This unique lifestyle strategy endows them with a competitive advantage in nutrient-poor environments and makes them ideal models for studying bacterial developmental regulation, morphological evolution, and ecological adaptation. This review systematically summarizes the research progress in DPB in terms of their taxonomy, molecular mechanisms of lifestyle regulation, extensive ecological distribution, and environmental adaptation strategies. Additionally, it discusses the application potential of DPB in environmental remediation and biotechnology. Finally, this review makes an outlook on future research directions, aiming to provide a reference for deeply understanding the biological characteristics of these bacteria and expanding their application value.

Ciliates of the family Balantidiidae widely parasitize the digestive systems of various invertebrates and vertebrates. Among them, Balantioides coli with cosmopolitan distribution is a major zoonotic pathogen. For a long time, due to the similar morphology of trophozoites of Balantidiidae ciliates from various hosts and the influences of various growing stages and host intestinal microenvironments on morphological characteristics, there have been some inconsistencies and confusions in their classification and nomenclature in literature. Recently, significant progress has been achieved in the study of the phylogenetic relationship and genetic diversity of Balantidiidae ciliates, providing new evidences for their taxonomic reconstruction. On the basis of a systematic review of literature and our findings, this article summarizes the taxonomic status and genetic diversity of Balantidiidae ciliates from different hosts in terms of the morphological characteristics, encystation ability, host specificity, and phylogenetic relationships. The existing evidence supports the division of the family Balantidiidae into two genera Balantidium and Balantioides and indicates that the ciliate infecting humans and various mammals should be classified as Balantioides coli. The related findings and understanding help clarify the taxonomy and nomenclature system of Balantidiidae ciliates and provide an important reference for studying the biological characteristics of Balantidiidae ciliates as well as the diagnosis and control of balantiosis in humans and animals.

Neopestalotiopsis cubana Maharachch. is the primary pathogen responsible for leaf spot and fruit ring rot in guava (Psidium guajava L.). The ADP-ribosylation factor-like protein 8 (Arl8) belonging to the ADP-ribosylation factor (Arf) superfamily is associated with lysosomal processes including localization, trafficking, and fusion. The function of Arl8 in phytopathogenic fungi remains poorly understood. Objective To elucidate the function of Arl8 in N. cubana. Methods An Arl8 gene was identified from the genome of N. cubana and designated as NecArl8. A NecArl8 knockout mutant (ΔNecArl8) and a complemented strain (ΔNecArl8-C) were generated via clustered regularly interspaced short palindromic repeats (CRISPR)-associated 9, (CRISPR/Cas9)-mediated genome editing. These strains were subsequently analyzed through phenotypic experiments and RNA-seq. Results ΔNecArl8 showed a reduction of 34.93% in sporulation capacity and significant increases of 5.74% and 7.39% in inhibition rates by salt and osmotic stresses, respectively. In addition, this mutant exhibited weakened pathogenicity, as indicated by a reduction of 5.74 mm in the average spot diameter. In contrast, the mutant displayed significantly enhanced resistance to oxidative stress, which showed an inhibition rate of -19.22%. After long-term culture, a large number of vacuolated cells were observed in the mutant. RNA-seq analysis was performed on ΔNecArl8 and the wild-type strain to identify differentially expressed genes. Transcriptomic analysis showed that 2 330 genes were significantly down-regulated and 1 355 genes were significantly up-regulated. Gene set enrichment analysis further revealed that transmembrane transport proteins were broadly suppressed, and the expression levels of 16 fungal-type vacuolar-related proteins were significantly down-regulated, suggesting that the structure and functions of fungal-type vacuoles were seriously damaged. The reverse transcription quantitative PCR (RT-qPCR) results showed the expression patterns of 9 randomly selected transmembrane transport genes were consistent with those obtained by transcriptome sequencing, indicating that the transcriptome data were highly reliable. Conclusion NecArl8 may affect the pathogenicity of N. cubana on guava by regulating function of transmembrane transport. This study lays a theoretical foundation for exploring the pathogenic mechanism of N. cubana and developing new control strategies.

Objective To overcome the limitations such as high costs and restricted substrate utilization of mono-culture fermentation, bacterium-alga co-culture based on resource complementarity offers a promising new avenue for ectoine production. This study investigated the co-culture conditions of Dunaliella pseudosalina ZBY-1 and Halomonas campaniensis XH26 and the variations in ectoine yield, aiming to elucidate the metabolic regulation mechanism of ectoine biosynthesis in the co-culture system. Methods Strains XH26 and ZBY-1 were co-cultured at different inoculation ratios (1/0, 1/5, 1/10, 1/15, and 1/20) to screen the ratio yielding the highest ectoine production. Targeted metabolomics analysis was performed on the co-culture group [H group (H)], the bacterial control group [XH26 group (X)], and the algal control group [ZBY-1 group (D)] to identify significant differential metabolites. Results The highest ectoine yield was achieved at a bacterium-to-alga ratio of 1:15, while the pigment content of the algal strain was lower than that of the control group. Metabolomics analysis identified 15 (H vs. D), 16 (H vs. X), and 16 (X vs. D) significant differential metabolites, including L-alanine, L-asparagine, L-aspartic acid, L-phenylalanine, malic acid, and pyruvic acid. Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis revealed that alanine, aspartate and glutamate metabolism, glyoxylate and dicarboxylate metabolism, and arginine biosynthesis were the significantly altered metabolic pathways. Conclusion The co-culture system exhibited an asymmetric pattern characterized by bacterial proliferation and algal inhibition. The co-culture system significantly activated the central carbon metabolic network of the bacteria. Notably, aspartic acid and glutamic acid were significantly accumulated in cells, serving as the direct carbon skeleton and amino donor, respectively, to directly promote the efficient synthesis of ectoine.

Objective To clone and express a gene encoding the β-xylosidase from Enterobacter cloacae GX-3, a putative member of the glycoside hydrolase family 3, systematically characterize the recombinant enzyme, and improve the xylose tolerance by molecular engineering of key amino acid residues involved in xylose binding. Methods On the basis of the whole-genome sequencing data of E. cloacae GX-3, primers were designed to amplify the β-xylosidase gene annotated as GH3. The target gene was amplified by PCR and cloned into the pQE30 expression vector, and the resulting recombinant plasmid was transformed into Escherichia coli M15 for induced expression. The recombinant enzyme was purified by nickel-affinity chromatography, and its enzymatic properties were studied. Site-directed mutagenesis was conducted on amino acid residues associated with xylose tolerance. Results The β-xylosidase gene belonging to the GH3 family was successfully cloned from E. cloacae GX-3 and heterologously expressed in E. coli M15. Substrate specificity analysis revealed that the recombinant enzyme EXYL was a multifunctional enzyme exhibiting β-xylosidase, β-glucosidase, and α-L-arabinofuranosidase activities. EXYL showed the optimal performance with the substrate of pNPX and at pH 5.5 and 45 ℃. The K_m and V_max values of this enzyme were (0.73±0.06) mmol/L and (130.00±6.85) μmol/(mg·min), respectively. The inhibition constant (K_i) for xylose was (51.95±2.36) mmol/L. When EXYL acted on xylooligosaccharides (X3-X5), the main products were xylose and xylobiose, each accounting for approximately 50% of the yield. Site-directed mutagenesis of xylose tolerance-related residues yielded positive mutants W138C and W138A, which showed 2.38-fold and 1.83-fold improvements in xylose tolerance, respectively. Conclusion This study provides insights into the multifunctional activities of β-xylosidases and offers new strategies for enhancing the xylose tolerance of β-xylosidases in the GH3 family.

Objective Bacillomycin L, a cyclic lipopeptide antibiotic produced by Bacillus velezensis Bs916, has been demonstrated to possess strong antifungal activity. However, its low yield has become a critical bottleneck limiting its large-scale application. This study aims to identify the negative transcriptional regulators involved in bacillomycin L biosynthesis and, based on this, adopt a dual strategy combining genetic engineering and fermentation process optimization to promote its large-scale production. Methods Homologous recombination was employed to construct single (ΔresD, ΔabrB) and double (ΔresDΔabrB) knockout strains. The regulatory characteristics were elucidated by HPLC, inhibition zone measurement, and RT-qPCR. Furthermore, EMSA and DNase I footprinting assays were conducted to investigate the binding activity and identify the specific binding sites of these transcription factors with the flanking sequences of the transcription initiation site of the Bac gene cluster. Finally, fermentation process optimization was performed in a bioreactor to further enhance the production of bacillomycin L. Results HPLC results demonstrated that the bacillomycin L yields of all the mutant strains cultured in the LB media significantly increased by 5.8, 11.3, and 12.0 folds compared with that of the wild-type strain. The antagonistic activities of the mutants against plant pathogenic fungi exhibited corresponding increases. RT-qPCR results further confirmed that both ResD and AbrB acted as negative regulators of bacillomycin L biosynthesis. EMSA experiments revealed that both ResD and AbrB possessed strong binding activities with their target sequences. DNase I footprinting assays further elucidated that AbrB exhibited a propensity for binding to A+T-rich gene fragments and displayed extensive DNA-binding capabilities, directly interacting with the promoter region, 5′ UTR, and coding regions of the Bac gene cluster. Unfortunately, the specific binding site of ResD remained to be identified. Under fermentation conditions, the genetically engineered strain ΔresDΔabrB achieved gram-per-liter level production of bacillomycin L in an optimized glucose-mineral salts medium. This result greatly promoted the large-scale production of bacillomycin L. Conclusion Adopting a dual strategy that combines genetic engineering with fermentation process optimization effectively breaks the bottleneck of low yields for Bacillus-derived antimicrobial peptides. This study provides a reference for the mass production of other microbial secondary metabolites.

Objective To achieve high-level expression and purification of the recombinant 9 kDa and 15 kDa chicken NK-lysin proteins (designated as cNKL9 and cNKL15) and evaluate their in vitro antibacterial and immunomodulatory activities. Methods The target genes—cNKL9 and cNKL15—were individually cloned into the pPIC9K vector to construct recombinant expression plasmids for heterologous expression in Pichia pastoris. After confirming the correct expression of the recombinant proteins via SDS-PAGE and Western blotting, we used nickel affinity chromatography to obtain highly purified recombinant cNKL9 and cNKL15 proteins. The in vitro antibacterial activitiesof recombinant proteins against Salmonella typhimurium and Escherichia coli O157:H7 were assessed by colony forming unit (CFU) assays. We measured the mRNA levels by RT-qPCR and analyzed the transcriptional changes of relevant cytokines after applying the recombinant proteins to the chicken macrophage cell line HD11. High-level expression of the recombinant proteins was achieved in a 15 L fermenter for scale-up culture via optimization of the high-density fermentation process. Results The P. pastoris expression plasmids pPIC9K-cNKL9 andpPIC9K-cNKL15 were successfully constructed. SDS-PAGE and Western blotting results revealed specific bands of recombinant cNKL9 and cNKL15 proteins in the culture supernatant, confirming the successful secretory expression of the target proteins. Antibacterial activity assay demonstrated that the recombinant cNKL9 protein exhibited significant inhibitory activities against both S. typhimurium and E. coli O157:H7, whereas the recombinant cNKL15 protein showed no obvious inhibitory effects on either pathogenic strain. RT-qPCR analysis showed that the recombinant cNKL15 protein significantly upregulated the transcription levels of cytokines such as CCL4 and CCL20 in HD11 cells, while the recombinant cNKL9 protein did not exhibit immunomodulatory activity. Furthermore, in the 15 L fermenter, the heterologous expression yields of cNKL9 and cNKL15 reached 0.64 g/L and 0.53 g/L, respectively. Conclusion For the first time, we achieved high-level expression of recombinant cNKL9 and cNKL15 proteins in the P. pastoris system. Specifically, cNKL9 exhibited inhibitory activities against S. typhimurium and E. coli O157:H7, while cNKL15 exerted immunomodulatory effects by upregulating the mRNA levels of cytokines in the chicken macrophage cell line HD11. These findings provide experimental evidence for the development of cNKL9 and cNKL15 as antibacterial or immunomodulatory agents, indicating their considerable application potential.

Objective To investigate the effects of ferric uptake regulator (Fur) on iron homeostasis, oxidative stress response, and virulence in Vibrio parahaemolyticus. Methods BLASTp and multiple sequence alignment analyses were employed to analyze the conservation of Fur proteins in several Gram-negative bacteria. A fur gene deletion mutant (Δfur) and a complementation strain (CΔfur) were constructed. The growth of the wild-type (WT), Δfur, and CΔfur was compared under normal (iron-replete), iron-excess, iron-restricted, and oxidative stress conditions. Inductively coupled plasma mass spectrometry was employed to measure intracellular metal content in each strain. A zebrafish survival assay and a competition infection assay were performed to assess the impact of Fur on the virulence of V. parahaemolyticus. RNA sequencing was conducted to identify the genes regulated by Fur. Results The Fur proteins were highly conserved among several Gram-negative bacteria. The deletion of fur attenuated the growth of V. parahaemolyticus under normal (iron-replete) and iron-excess conditions, and reduced its sensitivity to iron restriction. The intracellular iron content in Δfur was significantly lower than that in the WT and CΔfur strains. Fur regulated the oxidative stress response in V. parahaemolyticus. Fur played no significant role in the virulence of V. parahaemolyticus in the zebrafish model. The expression of multiple genes related to iron uptake, iron storage, and type Ⅲ secretion system 1 (T3SS1) was significantly upregulated, whereas genes encoding iron-containing proteins and type Ⅵ secretion system 2 (T6SS2) components were significantly downregulated in Δfur. Conclusion Fur regulates iron homeostasis and oxidative stress response in V. parahaemolyticus, but does not affect its virulence in zebrafish.

Objective To clarify the role and molecular mechanism of the host protein solute carrier family 25 member 6 (SLC25A6) during fowl adenovirus serotype-4 (FAdV-4) infection, thus providing a theoretical basis for elucidating the pathogenic mechanism of FAdV-4 and developing novel prevention and control strategies. Methods First, we confirmed that infection of LMH cells with FAdV-4 at different time points and multiplicities of infection (MOI) resulted in obvious cytopathic effects (CPE). Second, on the basis of the host protein SLC25A6 identified in previous screening, Western blotting was employed to examine the effect of FAdV-4 on the expression of endogenous SLC25A6 in cells. Subsequently, transfection experiments were performed to regulate the expression of SLC25A6 (overexpression or interference with endogenous expression). RT-qPCR and Western blotting were employed to analyze the effect of SLC25A6 on FAdV-4 replication from the aspects of mRNA level, protein level, and viral titer. Finally, co-immunoprecipitation (Co-IP) was employed to verify the interaction between SLC25A6 and the core capsid protein Hexon of FAdV-4. Results FAdV-4 significantly inhibited the expression of endogenous SLC25A6 in cells. The overexpression of SLC25A6 markedly inhibited FAdV-4 replication, while interference with endogenous SLC25A6 promoted viral replication. SLC25A6 could directly interact with the Hexon protein of FAdV-4. Conclusion The host protein SLC25A6 inhibits FAdV-4 replication through its interaction with the viral Hexon protein. The results provide a theoretical basis for further elucidating the pathogenic mechanism of FAdV-4 and developing novel prevention and control strategies.

Objective To address the limitations of current hair-loss treatments, we employed an in vitro human dermal papilla cell (HDPC) model and a telogen C57BL/6 mouse model to study the effects of Lactiplantibacillus plantarum CCFM1352 fermentation metabolites and bacterial preparations on the hair follicle cycle and decipher the associated molecular mechanisms. Methods HDPCs were treated with different concentrations of CCFM1352 fermentation metabolites or bacterial lysates in vitro, and cell viability as well as the expression of molecules related to Wnt/β-catenin signaling, anagen/catagen regulation, and apoptosis was assessed. Telogen C57BL/6 mice were administrated with the fermentation metabolites or heat-killed bacterial cells by gavage for 21 days, and changes in hair coverage, follicular structure, and associated signaling molecules were analyzed. Results CCFM1352 fermentation metabolites at a volume fraction of 10% increased the viability of HDPCs to 135.30%, up-regulated Wnt10b and FGF-7, down-regulated DKK1 and TGF-β1, and increased Bcl-2 and decreased Bax, inducing a favorable change pattern associated with the anagen phase. Continuous gavage of the fermentation metabolites promoted the transition of hair follicles from telogen to anagen, increased the hair coverage to 61.62%, and enhanced hair follicle density, dermal thickness, and β-catenin accumulation and nuclear translocation. However, the regulatory effects of bacterial lysate and heat-killed bacteria were weaker in both models and showed only limited improvements in some indices. Conclusion CCFM1352 fermentation metabolites could modulate Wnt/β-catenin-related molecules and the expression of key factors such as FGF-7, TGF-β1, Bcl-2, and Bax, which favors the transition of hair follicles from telogen to anagen and suggests an important role in promoting hair growth. This study provides experimental evidence for the application of bioactive metabolites from probiotics in the field of hair health.

Objective To investigate the effects of Chlamydia trachomatis plasmid protein pORF5 on cellular mitophagy and mitochondrial fission and to elucidate whether its mechanism is related to Drp1 activation. Methods HeLa cells stably expressing pORF5 and control cells were constructed by lentiviral transfection. After serum starvation treatment, the expression levels of autophagy-related proteins—microtubule-associated protein 1 light chain 3 (LC3), Beclin-1, and p62—were determined by Western blotting. Co-localization of LC3 and translocase of outer mitochondrial membrane 20 (TOMM20) was assessed by indirect immunofluorescence. Mitochondria were stained with MitoTracker Red CMXRos, and mitochondrial morphology was observed and analyzed through confocal laser scanning microscopy. Dynamin-related protein 1 (Drp1) phosphorylation and its translocation to mitochondria were examined by Western blotting and indirect immunofluorescence. To explore the role of Drp1 in autophagy and mitochondrial fission, we pretreated cells with the Drp1-specific mitochondrial division inhibitor 1 (Mdivi-1). Changes in mitochondrial morphology and Drp1 translocation were evaluated by confocal microscopy and indirect immunofluorescence. Then, Western blotting was employed to determine the expression levels of autophagy-related proteins, and indirect immunofluorescence assay to analyze LC3 fluorescence intensity and its co-localization with TOMM20. Results Compared with the control group, pORF5 significantly upregulated the expression of LC3-Ⅱ and Beclin-1, downregulated the expression of p62, and enhanced the co-localization of LC3 and TOMM20. pORF5 expression led to the fragmentation of the mitochondrial network structure. It promoted Drp1 phosphorylation at Ser616 and enhanced Drp1 translocation to mitochondria. Inhibition of Drp1 with Mdivi-1 attenuated Drp1 phosphorylation and translocation, resulting in elongated mitochondrial morphology. In addition, the Mdivi-1 inhibitor group showed downregulated expression of LC3-Ⅱ and Beclin-1, upregulated the expression of p62, and attenuated co-localization of LC3 and TOMM20. Conclusion The C. trachomatis plasmid protein pORF5 may induce mitophagy and mitochondrial fission by promoting Drp1 phosphorylation and its mitochondrial translocation.

Objective To investigate whether cinnamaldehyde affects the acquisition of exogenous plasmids via conjugative transfer in Vibrio harveyi and to elucidate the potential mechanism. Methods V. harveyi 345 was used as the recipient strain, and an Escherichia coli strain harboring the shuttle plasmid pMMB207 served as the donor. The efficiency of exogenous plasmid acquisition by V. harveyivia conjugative transfer was analyzed before and after cinnamaldehyde treatments at three different stages: Treatment 1 (cinnamaldehyde added during the early logarithmic phase of V. harveyi), Treatment 2 (cinnamaldehyde added during the plate-mating process), and Treatment 3 (cinnamaldehyde added during both stages). Changes in biofilm-forming ability of V. harveyi before and after cinnamaldehyde treatments were assessed. Furthermore, qPCR was used to monitor the changes in the expression of quorum sensing (QS)-related genes under Treatment 1 and Treatment 2. Results Cinnamaldehyde at 0.5, 1, 2, 4, and 8 µg/mL reduced the conjugative transfer efficiency by 59%, 98%, 87%, 85%, and 83% under Treatment 1 and by 51%, 85%, 36%, 93%, and 49% under Treatment 2, respectively. Under Treatment 3, cinnamaldehyde at 1, 2, and 8 µg/mL decreased the conjugative transfer efficiency by 71%, 14%, and 75%, respectively. Cinnamaldehyde treatments did not significantly alter biofilm formation. Under Treatment 1, cinnamaldehyde led to no significant changes in the expression of QS-related genes (P>0.05). Under Treatment 2, after 4 h of conjugation, the treatment with 8 µg/mL cinnamaldehyde downregulated the expression of luxM, luxN, luxS, and luxP by 1.47 to 2.94 folds (P<0.05). Meanwhile, treatments with 0.5-8 µg/mL cinnamaldehyde downregulated luxR expression by 1.16 to 3.19 folds (P<0.05). Conclusion Cinnamaldehyde suppresses conjugative plasmid transfer, most likely by attenuating quorum sensing (QS). This study elucidates the role of cinnamaldehyde as a natural compound in regulating bacterial gene transfer and the potential spread of antibiotic resistance, providing a reference for the development of novel antibacterial adjuvants.

Heyndrickxia coagulans, formerly known as Bacillus coagulans, exhibits pronounced intraspecific heterogeneity and represents an important probiotic candidate for animal feed. Objective To isolate high-quality H. coagulans strains and evaluate their potential for feed applications. Methods H. coagulans strains were isolated from spoiled fruits, soil, and feces through the plate streaking method. The isolates were identified by 16S rRNA gene sequencing. Functional assays were conducted to comprehensively evaluate their feed potential, including acid production, gastrointestinal tolerance, biosurfactant production, antimicrobial activity, protease activity, and utilization of different carbohydrates and feed raw materials. Results A total of 133 strains were isolated, among which 23 strains were identified as H. coagulans. Nine strains with strong acid-producing ability were further selected. Among them, strain N9 isolated from rotten Artocarpus heterophyllus exhibited the highest acid production, decreasing the pH to 3.89±0.05 after 24 h of incubation at 37 °C and achieving a lactic acid yield of (3 370.00±87.36) μg/mL. Following treatment with simulated gastric and intestinal fluids and bile salts, the survival rates of N9 were (73.50±1.54)% and (83.20±1.66)%, respectively. Furthermore, strain N9 showed the strongest biosurfactant production, with an oil displacement diameter of (43.00±0.46) mm. Antimicrobial assays revealed that N9 effectively inhibited three intestinal pathogens: Salmonella enterica subsp. enterica, Escherichia coli, and Staphylococcus aureus. In addition, strain N9 exhibited notable protease activity, with enzymatic characteristics well adapted to the animal intestinal environment. It was capable of utilizing carbohydrates such as xylo-oligosaccharides, as well as feed ingredients including palm kernel meal, cottonseed meal, sunflower meal, soybean meal, and wheat bran for acid production. Conclusion H. coagulans N9, isolated from rotten A. heterophyllus, demonstrates good potential for feed additive, serving as a new candidate strain for the research on probiotic application.

Objective Cyanocobalamin (CN-CbI) requires to be converted into adenosylcobalamin (Ado-CbI) to exert neuroprotective effects, yet its conversion efficiency is impaired under conditions such as chronic sleep deprivation (CSD). This study aimed to obtain a bacterial strain with high efficiency in converting CN-CbI to Ado-CbI that could enhance neuroprotective effects. Methods Lactobacillus paragasseri CCFM1526, a strain capable of converting CN-CbI to Ado-CbI, was isolated via UPLC. A mouse model of CSD was established, and the cognitive functions of mice were evaluated by the novel object recognition and Morris water maze tests. The neuronal structure in the hippocampal dentate gyrus (DG) region was observed via histological staining. Tissue vitamin B₁₂ levels were measured, and the ERK/mTOR signaling pathway along with related neural protein expression was analyzed to assess the neuroprotective mechanism of the fermentation broth. Results Compared with CN-CbI supplementation alone, the fermentation broth of L. paragasseri CCFM1526 significantly improved the cognitive function and alleviated the structural damage in the hippocampal DG region of CSD mice. Intervention with the fermentation broth increased the total vitamin B₁₂ content in the liver, serum, and brain by 10.2%, 16.3%, and 29.0%, respectively (P<0.05). Meanwhile, it activated the ERK/mTOR signaling pathway, leading to increases of 21.5%, 52.4%, 17.3%, and 19.7% in the content of myelin basic protein, postsynaptic density protein 95, brain-derived neurotrophic factor, and nerve growth factor, respectively (P<0.05). Conclusion L. paragasseri CCFM1526 converts CN-CbI into Ado-CbI through fermentation, subsequently activating the ERK/mTOR signaling pathway and upregulating the expression of neurotrophic and myelin repair-related proteins, thereby alleviating CSD-induced nerve injury.

Objective Curcumin exhibits diverse pharmacological activities, yet its clinical and industrial applications are largely limited by its poor water solubility and low oral bioavailability. The present study was designed to evaluate the anti-fatigue efficacy of curcumin fermented by Lactobacillus delbrueckii subsp. bulgaricus CCFM1520 and to preliminarily elucidate the underlying mechanism responsible for its anti-fatigue effects. Methods The bioconversion efficiency of curcumin and its metabolites by CCFM1520 were quantified viain vitro bioconversion assays. A mouse model of fatigue was established by combining forced swimming with chronic restraint stress. Mice were randomized into six groups: blank control, model, curcumin, CCFM1520, CCFM1520-curcumin synbiotic, and CCFM1520-fermented curcumin. The anti-fatigue efficacy of fermented curcumin was comprehensively assessed from behavioral parameters, serum level of oxidative stress markers [superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), malondialdehyde (MDA)], and cerebral levels of neurotransmitters [dopamine (DA) and norepinephrine (NE)]. Real-time quantitative PCR was performed to examine the mRNA expression profiles of genes associated with the nuclear factor erythroid 2-related factor 2/heme oxygenase-1 (Nrf2/HO-1) signaling pathway in the skeletal muscle and the brain-derived neurotrophic factor/tyrosine kinase receptor B (BDNF/TrkB) signaling pathway in the brain tissue. Results CCFM1520 converted curcumin at an efficiency of 62.39%, efficiently converting it into tetrahydrocurcumin. Compared with unfermented curcumin, the fermented product significantly improved behavioral outcomes, including prolonged exhaustive swimming time, shortened escape latency in the water maze, and increased swimming distance. In addition, the fermented product significantly enhanced the activities of SOD and GSH-Px and reduced the content of MDA in the serum, and simultaneously elevated the levels of DA and NE in the mouse brain. Mechanism studies revealed that fermented curcumin up-regulated the mRNA levels of Nrf2 and HO-1 in the muscle tissue and BDNF and TrkB in the brain tissue. Conclusion L. delbrueckii subsp. bulgaricus CCFM1520efficiently converts curcumin into more bioactive metabolites. These metabolites exert a pronounced anti-fatigue effect by enhancing antioxidant capacity and modulating neurotransmitter balance. This study provides a novel strategy and a theoretical basis for developing high-efficacy anti-fatigue functional foods.

Phytoplankton are key primary producers in marine ecosystems, and their community composition is influenced not only by environmental factors but also by biological factors such as parasitic organisms. However, the relationships between parasitic eukaryotes and phytoplankton in coastal bays remain poorly understood. Objective To investigate the community composition, seasonal succession, and interactions of dominant phytoplankton, including diatoms, dinoflagellates, and their parasitic eukaryotes, Syndiniales, in Sanmen Bay, Zhejiang Province during summer and autumn. Methods Surface water samples were collected from Sanmen Bay in September (summer) and November (autumn) 2019. High-throughput sequencing was performed for the V4 region of the 18S rRNA gene of eukaryotic microbes. Principal coordinate analysis, co-occurrence network analysis, and bipartite network analysis were employed to explore the succession and interactions of diatom, dinoflagellate, and Syndiniales communities. Results Diatom communities exhibited pronounced seasonal variations, with the relative abundance of 36.9% in summer and 5.5% in autumn, being dominated by Cyclotella, Skeletonema, and Thalassiosira. Dinoflagellates showed similar relative abundance in summer (28.0%) and autumn (28.5%), being dominated by Gyrodinium, Heterocapsa, and Gymnodinium. Four Syndiniales groups (group I-IV) were detected in Sanmen Bay, with higher relative abundance in autumn (10.5%) than in summer (7.6%). Among Syndiniales, group I dominated in summer (49.1%), while group Ⅱ dominated in autumn (75.2%). Salinity, nitrate, nitrite, N:P ratio, and Si:N ratio were identified as the main drivers of the dominant phytoplankton and Syndiniales communities. Network analysis indicated that Syndiniales mainly interacted with diatoms and dinoflagellates, with higher interaction frequency in autumn than in summer. Syndiniales were identified as keystone taxa in the interaction network. Conclusion Seasonal succession of diatoms in Sanmen Bay was more pronounced than that of dinoflagellates. Syndiniales groups I and Ⅱ exhibited significant seasonal succession and maintained close interactions with diatoms and dinoflagellates, serving as key taxa in the interaction network. High-throughput sequencing overcame the limitations of microscopy in detecting Syndiniales, providing systematic insights into the community features and potential interactions of dominant phytoplankton and Syndiniales in Sanmen Bay. This study offers important insights into phytoplankton adaptation and interspecies interactions in eutrophic coastal bays.

Objective Planktonic bacteria are essential for marine ecosystem health, yet how mussel aquaculture influences planktonic microbial communities remains unclear. This study aims to clarify the effects of mussel aquaculture on the community structure, diversity, and assembly mechanisms of marine planktonic bacteria. Methods Sixty-eight water samples were collected from a mussel aquaculture area and surrounding areas in Shengsi, Zhejiang, during summer 2024. Integrated analyses of 16S rRNA gene amplicon sequencing data and environmental factors were performed. Results The alpha diversity of particle-attached bacteria (PAB) significantly reduced in the aquaculture area, whereas free-living bacteria (FLB) showed no significant change in alpha diversity but exhibited clear shifts in beta diversity and phylogenetic structure. Random forest analysis identified Pseudomonadales and Bdellovibrionaceae as indicator taxa within the aquaculture area, and their changes might be associated with organic matter inputs and altered nutrient conditions. Functional prediction indicated enhanced nitrogen cycling (especially nitrification and aerobic ammonia oxidation) and a shift toward reductive acetogenesis in carbon cycling, alongside suppressed methanogenesis in the aquaculture area. Microbial community assembly was governed mainly by deterministic processes (e.g., heterogeneous selection) in the aquaculture area but by stochastic processes in surrounding waters. Conclusion This study demonstrates that mussel aquaculture can reconfigure the structures, functions, and assembly mechanisms of planktonic bacterial communities, providing insights for ecological impact assessment of mariculture.

Excessive nitrogen input caused by eutrophication in nearshore waters is a major environmental stressor driving the global degradation of seagrass beds. Objective To screen and identify efficient aerobic denitrifying bacteria from seagrass bed ecosystems and elucidate their nitrogen removal performance and mechanisms, thus providing microbial resources for alleviating nitrogen loading and restoring eutrophic seagrass beds. Methods Aerobic denitrifying bacteria were isolated and screened from seagrass rhizosphere sediments in Zhifu Bay, Yantai by enrichment-domestication culture and bromothymol blue assay. The taxonomic status of the strains was determined by 16S rRNA gene sequencing. On the basis of nitrogen removal performance, an efficient aerobic denitrifying strain was selected. Single-factor and orthogonal experiments were conducted to optimize its denitrification conditions, and nitrogen balance experiments and whole-genome sequencing were employed to elucidate its nitrogen removal pathways and key functional genes. Results A total of 34 denitrifying strains were isolated from seagrass rhizosphere sediments in Zhifu Bay, Yantai. The dominant genera were Pseudomonas and Acinetobacter. A strain designated as Pseudomonas sp. S22 with high denitrification performance was selected. The denitrification conditions of this strain were optimized as follows: sodium succinate as the carbon source, C/N=15, pH 9.0, salinity (S)=30‰, and T=28 ℃. Under these conditions, the strain achieved a removal rate of 99.99% for 140 mg/L nitrate nitrogen within 36 h, demonstrating excellent nitrogen removal efficiency. Nitrogen balance analysis revealed that approximately 59.64% of the initial nitrate nitrogen was converted to gaseous nitrogen, confirming that denitrification was the dominant nitrogen removal pathway. Genomic sequencing revealed that strain S22 carried key functional genes for aerobic denitrification, including napA and nirS, providing a genetic basis for its denitrification phenotype at the molecular level. Conclusion This study systematically isolated and identified aerobic denitrifying bacteria from seagrass beds in northern China. Strain S22 exhibits outstanding nitrogen removal performance and environmental adaptability. Nitrogen balance and genomic analyses confirm that denitrification is its primary nitrogen removal pathway and the strain carries key functional genes for aerobic denitrification. Strain S22 can serve as a potential microbial resource for reducing nitrogen loading in seagrass beds. This study provides both a valuable strain and a theoretical basis for the future development of microbe-seagrass synergistic remediation technologies.

Objective To elucidate the tolerance mechanisms of Penicillium oxalicum Z2 to flavonoids present in licorice residues and to analyze its transcriptional responses under gradient stress concentrations, thereby providing a theoretical basis for breeding resistant strains and improving fermentation efficiency. Methods P. oxalicum Z2 was treated with different concentrations of licorice flavonoids (0, 0.25, 0.50, 1.00, and 2.00 mg/mL). Physiological indicators were systematically measured. Transcriptome sequencing was conducted to identify differentially expressed genes and enriched KEGG/GO pathways, revealing the molecular adaptation mechanisms of P. oxalicum Z2 under flavonoid stress. Results As the flavonoid concentration increased, P. oxalicum Z2 maintained high biomass and cellulase activity. Under the highest flavonoid concentration (2.00 mg/mL), its biomass reached 4.840 g/L, and the activities of endoglucanase, β-glucosidase, filter paper enzyme, and xylanase reached 74.78, 3.24, 6.99, and 562.5 U/mL, respectively, which were more than ten folds of those in the conventional P. oxalicum strain. Transcriptome analysis showed that the number of differentially expressed genes increased in a concentration-dependent manner, from 818 in the Low group to 3 945 in the Super group. Under low stress (≤1.00 mg/mL), the strain reprogrammed carbohydrate metabolism and ABC transporter pathways to adjust carbon utilization and sustain essential cellular activities. Under high stress (2.00 mg/mL), it further activated the ribosome and oxidative phosphorylation pathways, with 65 genes upregulated in the ribosome pathway. Conclusion This study identified key genes (e.g., EF-Tu, SecY,and FtsY) and core pathways (e.g., oxidative phosphorylation and ABC transporter), demonstrating that P. oxalicum Z2 employed tiered responses to licorice flavonoid stress. Specifically, the strain remodels sugar metabolism and transmembrane transport under low flavonoid stress to maintain survival and activates ribosome and energy metabolism compensation mechanisms under high flavonoid stress.

Sugarcane smut is a severe fungal disease caused by Sporisorium scitamineum, resulting in yield reduction and economic losses. Reversible protein phosphorylation plays a crucial role in the sexual mating and pathogenicity of S. scitamineum. Protein phosphatases, as key regulators of reversible protein phosphorylation, remain poorly characterized in S. scitamineum. Objective To elucidate the biological functions of the protein phosphatase SsPpe1 in S. scitamineum, providing a potential target for effective control of sugarcane smut. Methods We constructed overexpression mutants OE-Ssppe1 by Agrobacterium-mediated genetic transformation technology and analyzed the sporidium morphology, sexual mating ability, stress tolerance, and pathogenicity. Results The OE-Ssppe1 sporidia exhibited pseudohyphal morphology with multiple nuclei and abnormal chitin accumulation. The OE-Ssppe1 mutants showed reduced tolerance to NaCl and SDS, sexual mating, and pathogenicity. RT-qPCR and RNA-seq analyses revealed that Ssppe1 overexpression affected the expression of genes related to pheromone response, MAPK, and cAMP-PKA signaling pathways. In addition, Ssppe1 overexpression affected protein synthesis and folding process. Conclusion The protein phosphatase SsPpe1 is involved in regulating the sporidium morphology, stress responses, sexual mating, and pathogenicity of S. scitamineum. These findings provide a theoretical basis for thoroughly elucidating the pathogenic mechanisms of S. scitamineum and developing targeted disease control strategies.

Objective To prepare the chlamydospore wettable powder with significant control efficacy against tomato bacterial wilt from Cladophialophora guangxiense HX2, a dark septate endophyte (DSE). Methods Single-factor experiments were carried out to screen the types and dosages of carriers, wetting agents, dispersants, and ultraviolet protectants for the wettable powder. Pot experiments were conducted to evaluate the effects of soaking tomato seeds with four concentrations (T1: 1×10⁸ CFU/mL; T2: 1×10⁷ CFU/mL; T3: 1×10⁶ CFU/mL; T4: 1×10⁵ CFU/mL) for 30 min on tomato plant growth, tomato bacterial wilt, and activities of five defense enzymes—peroxidase (POD), polyphenol oxidase (PPO), phenylalanine ammonialyase (PAL), catalase (CAT), and superoxide dismutase (SOD). Results The formulation was optimized as follows: 25% chlamydospore suspension, 8% dispersant polyethylene glycol (PEG8000) and wetting agent Tween-60 at a mass ratio of 1:3, 0.5%-1.5% UV protectant ascorbic acid (VC), and white carbon black as the carrier to make up the remaining proportion to 100%. The wettable powder prepared according to this formulation had the chlamydospore content of 2.35×10⁸ CFU/g, the wetting time of 24.25 s, a suspension rate of 73.8%, pH 5.71, the moisture content of 16.67%, and the fineness of 98.81%. All indicators met the requirements of the national standard GB 20287—2006 Microbial Inoculants in Agriculture. The results of pot experiments indicated that the T2 treatment exhibited a significant plant growth-promoting effect, increasing the root length, plant height, stem diameter, fresh weight, and dry weight by 47.39%, 31.82%, 24.64%, 89.45%, and 90.97%, respectively, compared with the control group. On day 30 after pathogen inoculation, the control efficacy of this treatment against tomato bacterial wilt reached 50.9%, which was significantly higher than that of the Trichoderma harzianum treatment. Moreover, the T2 treatment significantly enhanced the activities of the five defense enzymes. Conclusion The HX2 chlamydospore wettable powder prepared in this study has good control efficacy against tomato bacterial wilt. This study provides a technical basis for the large-scale popularization and application of this agent.

Objective To address the problems of rhizospheric microenvironment deterioration and medicinal quality decline caused by continuous cropping obstacles of Fritillaria unibracteata Hsiao et K. C. Hsia, this study explored the regulatory effects of different concentrations of salicylic acid (SA) under continuous and non-continuous cropping patterns and clarified the optimal SA concentration and underlying mechanism for alleviating continuous cropping obstacles, aiming to provide a theoretical basis for optimizing cultivation techniques. Methods A pot experiment was conducted with two cultivation patterns (continuous cropping and non-continuous cropping) and six SA concentration gradients (0, 20, 50, 100, 200, and 500 μmol/L). The changes in root exudates, soil physicochemical properties, soil enzyme activities, alkaloid content, and microbial community structure were determined. Correlation analysis and redundancy analysis (RDA) were performed to elucidate the regulation mechanism. Results SA exerted significant concentration-specific regulatory effects on the rhizospheric microenvironment and alkaloid biosynthesis of F. unibracteata, showcasing a significant interaction effect with cultivation patterns. Total phenolic acids in root exudates increased under 20 μmol/L and 500 μmol/L SA treatments (P<0.05), and organic acids reached the peak under 200 μmol/L SA treatment. The soil organic carbon and soil organic matter in the non-continuous cropping group were significantly higher than those in the continuous cropping group. SA at 50 μmol/L optimized soil pH, increased the supply of available phosphorus and ammonium nitrogen, and enhanced the activities of urease and acid phosphatase. Pseudomonadota and Ascomycota were the dominant phyla in bacterial and fungal communities, respectively. SA at 500 μmol/L enriched beneficial microorganisms such as Streptomyces, inhibited pathogens, and specifically increased the content of peimisine and sipeimine. RDA results showed that SA remodeled the microbial community by regulating the composition of root exudates, thereby mediating alkaloid biosynthesis. Conclusion SA at 50 μmol/L SA is suitable for optimizing rhizosphere nutrient supply and enzyme activities, and that at 500 μmol/L is suitable for effectively alleviating continuous cropping obstacles and promoting the accumulation of medicinal alkaloids. SA achieves rhizospheric ecological restoration and medicinal quality improvement through a synergistic pathway of regulating root exudate composition, remodeling microbial community structure, and repairing rhizospheric interaction network. This study provides a new approach for the management of continuous cropping obstacles for F. unibracteata.

Objective To investigate the antagonistic activity of Bacillus amyloliquefaciens X60 against tobacco phyllosphere microorganisms and its effects on the phyllosphere microbial community of tobacco. Methods Bioactivity assays were conducted to evaluate the antagonistic effects of B. amyloliquefaciens X60 against 20 species of pathogenic fungi, 15 species of non-pathogenic fungi, 2 specialized forms of pathogenic bacteria, and 15 species of non-pathogenic bacteria. Amplicon sequencing was employed to assess the influence of this strain on the phyllosphere microbial community structure. Results B. amyloliquefaciens X60 exhibited strong antagonistic activity (inhibition rates of 60.00%-80.00%) against 13 species of pathogenic fungi (e.g., Rhizopus oryzae) and 12 species of non-pathogenic fungi (e.g., Trichoderma harzianum). Moderate antagonism (inhibition rates of 10.00%-59.00%) was observed against 7 species of pathogenic fungi (e.g., Alternaria tenuissima) and 3 species of non-pathogenic fungi (e.g., Thielavia microspora). Significant antibacterial activity (inhibition zone diameter >20 mm) was detected against 2 specialized forms of pathogenic bacteria (Pseudomonas syringae pv. tabaci and pv. angulata) and 7 non-pathogenic bacteria (e.g., Exiguobacterium). After application, X60 showed the control efficacy of 52.35% against tobacco leaf spot. Following treatment, the relative abundance of Pantoea—a genus of opportunistic bacteria dominating the infected tissue—increased, whereas bacterial diversity and richness initially declined and then recovered. Fungal richness decreased throughout the observation period, while fungal diversity exhibited a transient decrease followed by a rebound. The relative abundance of phytopathogenic fungi declined from 44.87% to 6.71%. Conclusion B. amyloliquefaciens X60 possesses a broad antimicrobial spectrum and exerts strong antagonistic activity against 25 fungal and 9 bacterial species colonizing the tobacco phyllosphere. Under field conditions, the strain provided 52.35% control of tobacco leaf spot and significantly reduced the abundance of foliar phytopathogens, demonstrating the potential as a biocontrol agent for the management of this disease.

Objective To achieve the targeted isolation of the cyclooctapeptides, surugamides, from the deep-sea-derived Streptomyces sp. NA13 and explore their biological activities. Methods An approach integrating genome mining and LC-MS/MS molecular networking was employed to discover cyclopeptides from Streptomyces sp. NA13. Through systematic natural product isolation and characterization, these compounds were identified as surugamides. Their growth-promoting effects on Oryza sativa and Zea mays were assessed. Results Four cyclooctapeptides (surugamides A, B, D, and E) were isolated and identified. They had significant effects of promoting root growth in Z. mays and O. sativa seedlings. Notably, surugamide A at a concentration of 0.1 µmol/L demonstrated particularly outstanding growth-promoting effects on Z. mays roots. Conclusion This study uncovers the novel plant growth-promoting activity of surugamides, offering lead compounds for the development of innovative marine microbial-derived plant growth regulators.

Objective Southern blight, caused by Sclerotium rolfsii Sacc., is a major soil-borne disease that limits peanut production. At present, its management relies mainly on chemical fungicides. To improve the field control efficacy, reduce fungicide application rates, and lower pesticide residues in peanuts, this study screened antagonistic microorganisms against S. rolfsii and evaluated their synergistic effects with chemical fungicides in the control of peanut southern blight. The results are expected to provide technical support for the green and sustainable management of this disease. Methods This study first used a plate-based screening assay to obtain biocontrol strains showing strong antagonistic activity against S. rolfsii and high compatibility with commonly used fungicides. Next, a fungicide screening assay was conducted to identify control agents that effectively inhibited S. rolfsii without adversely affecting the growth of the biocontrol strains. Subsequently, an in vitro combined toxicity assay was performed. The pathogen was inoculated onto PDA plates containing different concentrations of thifluzamide (0.15 and 0.30 μg/mL), sterile 10% fermentation broth of H02, or their mixtures. Fungal growth was observed, and the inhibition rate and inhibition ratio (IR) were calculated to evaluate the interaction type of the combined treatment. Finally, a pot experiment was carried out to compare the control efficacy against peanut southern blight among the fermentation broth, the full-dose fungicide (recommended field rate), and the combination of fermentation broth with a half-dose fungicide. Results A Burkholderia gladioli strain H02 with high antagonistic activity and good fungicide compatibility was screened out, exhibiting an inhibition rate of 76.03%. Concurrently, thifluzamide was identified as a fungicide with strong inhibitory activity against S. rolfsii and no negative impact on the growth of strain H02, exhibiting an EC₅₀ of 0.151 3 μg/mL. Results from the in vitro combined toxicity assay indicated that the combination of the H02 fermentation broth and thifluzamide showed an IR greater than 1, exhibiting a synergistic effect. Moreover, the combination of half the concentration of thifluzamide with the fermentation broth significantly demonstrated higher inhibition rates against pathogen mycelia and sclerotia (74.91% and 95.58%, respectively) than the full concentration of thifluzamide alone (68.04% and 83.67%, respectively). Pot experiment results showed that the combination of H02 fermentation broth and half dosage of thifluzamide had the control efficacy comparable to that of the full dosage of thifluzamide (68.94% vs. 66.63%), whereas the single application of H02 fermentation broth provided the control efficacy of 61.17%. Conclusion The combined application of B. gladioli H02 and thifluzamide can achieve synergistic control of peanut southern blight and reduce chemical fungicide usage while maintaining effective disease control, thus showing promising potential for practical application.

Meloidogyne incognita is one of the most destructive plant-parasitic nematodes worldwide, causing severe economic losses in agricultural production. Biocontrol bacteria can effectively control M. incognita, with significant differences in control efficacy among different strains. However, the mechanisms underlying differences in control efficacy remain unclear. Objective To explore the mechanisms responsible for the different efficacy of various biocontrol bacteria against nematodes. Methods The differences in nematicidal activity between two biocontrol bacterial strains, B133 and B104, were analyzed. Comparative genomics and metabolomics techniques were employed to investigate the genetic composition and metabolic mechanisms influencing the nematicidal activity of the two strains. Result From 24 h to 120 h of fermentation, the nematicidal activity of strain B133 was significantly higher than that of strain B104, reaching peaks of 77% and 54%, respectively, at the time point of 60 h. Whole-genome comparative analysis revealed that strain B133 possessed a larger genome size and a greater number of coding genes than strain B104. The phylogenetic trees conducted based on 16S rRNA gene or the housekeeping gene gyrB indicated that strains B133 and B104 were two different subspecies of Priestia megaterium. Predictions based on the virulence factors database (VFDB) and Kyoto encyclopedia of genes and genomes (KEGG) database showed that strain B133 harbored 22 unique virulence genes and 75 unique metabolism genes compared with strain B104. Meanwhile, the metabolites in the fermentation filtrate (60 h) were determined. Principal component analysis demonstrated significant differences in metabolite profiles between the two strains. Compared with that of strain B104, the fermentation filtrate of strain B133 had 40 increased metabolites (P<0.05), such as galactinol, 4-aminobenzoic acid, lumichrome, anthranilic acid, trehalose, and 3-methylthiopropionic acid. Moreover, through integrated genomics-metabolomics analyses, cysteine and methionine metabolism was identified as a key pathway influencing nematicidal activity. This pathway involves an L-lactic dehydrogenase (LDH) gene unique to strain B133 and 3-methylthiopropanoic acid with an elevated level and a positive correlation with the nematicidal effect of the strain. Conclusion By coupling genomics and metabolomics, this study reveals the different functional gene clusters and potential related metabolites of different subspecies of P. megaterium, laying a theoretical foundation and a practical basis for the targeted screening, modification, and industrial development of efficient biocontrol agents for nematodes.

Objective To examine the anti-lung cancer activity of Cordyceps militaris and predict its potential quality markers. Methods We systematically reviewed the current studies on the anti-lung cancer effects of C. militaris and summarized its chemical components. Homo sapiens lung cancer microarray data were integrated with network pharmacology to build a “component-target-pathway” network, followed by molecular docking analysis. On this basis, the potential quality markers of C. militaris for lung cancer treatment were predicted. Results We predicted 11 potential quality markers, which were grouped into six categories: (1) cordycepin and its analog O^5′-acetylcordycepin; (2) adenosine and its analogs, including N⁶-[β-(acetylcarbamoyloxy)ethyl]-adenosine, N⁶-(2-hydroxyethyl)-adenosine, N⁶-(4-methylbutyrate)-adenosine, and 5′-(3″-deoxy-β-D-ribofuranosyl)-3′-deoxyadenosine; (3) ergosta-7,22-dien-3β,5α-dihydroxy-6-one; (4) cordycepisosalt A; (5) pentostatin; and (6) cordyrrole B. Conclusion This study integrates literature review and bioinformatics analysis to predict potential anti-lung cancer quality markers of C. militaris. The suggested mechanisms and candidate components are theoretical and need further experimental validation to confirm their effectiveness. This work offers a reference for developing a quality standard system of C. militaris for anti-lung cancer applications.

A reverse genetics platform for foot-and-mouth disease virus (FMDV) is an indispensable tool for studying the pathogenic mechanism, protein function, and vaccine development. However, the conventional method of constructing infectious clones of FMDV is usually laborious, time-consuming, and costly. Objective To establish a new reverse genetics platform for rapid rescue of FMDV based on infectious subgenomic amplicons (ISA), which can avoid in vitro ligation and bacterial cloning. Methods The whole gene of FMDV O/GDLeiZh/2020 strain was divided into five overlapping fragments and then individually amplified by high-fidelity PCR. The T7 promoter sequence was added to the 5′-end gene and the poly(A) tail was introduced at the 3′-end. At the same time, the poly(C) sequence and molecular markers were introduced by fusion PCR. Two large fragments covering the whole gene of FMDV were obtained by multiple rounds of fusion PCR amplification and co-transfected into BSR/T7 cells expressing T7 RNA polymerase. The cell supernatant was collected 72 h post-transfection. The rescued virus was identified and characterized by RT-PCR, indirect immunofluorescence, electron microscopy, plaque assay, and one-step growth curve assay. Results The typical cytopathic effect of FMDV was observed 60 h post-transfection. Sequencing, immunofluorescence, and electron microscopy collectively confirmed that infectious FMDV was successfully rescued. Furthermore, one-step growth curve and plaque assays demonstrated that the rescued virus retained replication kinetics and biological characteristics comparable to those of the wild-type virus. Conclusion This study successfully establishes a new method for rapid and efficient rescue of FMDV based on ISA, which will lay a solid foundation for further improving FMDV rescue technology and rapidly expanding its application in the future.

To provide valuable information on the current status of research and the hot topics in the field of lactic acid bacteria (LAB) in Baijiu brewing, this study conducted a literature review and screening of relevant publications from 2006 to 2025, based on the China National Knowledge Infrastructure (CNKI) and the Web of Science (WOS) core collection with “Baijiu” and “lactic acid bacteria” as search terms. We employed bibliometrics to analyze the annual number of publications, authors, research institutions, and keyword co-occurrence and bursts of 284 Chinese publications and 324 English publications in this field, and visualized the results in a knowledge map. The results showed that the annual number of publications on LAB in Baijiu brewing exhibited an upward trend overall, with the number of English publications significantly surpassing that of Chinese publications after 2020. Chinese scholars constituted the research mainstream, and XU Yan, ZHANG Suyi, SUN Baoguo and other scholars were major contributors to this research field. Universities such as Jiangnan University, Sichuan University, and Sichuan University of Science and Engineering formed the core research forces, exhibiting a distinct pattern of production-education-research collaboration. Research hotspots gradually shifted from the early isolation and identification of LAB to the in-depth analysis of community structure, dynamic succession, metabolism, and flavor substance formation mechanisms, driven by multi-omics technologies. Future research can focus on the multi-omics mechanisms of LAB, breeding of functional strains, intelligent regulation of synthetic microbial communities, and technological transformation for green brewing to boost the high-quality development of the Baijiu industry. As the first systematic bibliometric and visual analysis of this specific field, this study provides a reference for scholars to grasp the development trends and optimize academic layout, while also offering informational support to promote the scientific and intelligent upgrading of the Baijiu brewing industry.