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.

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 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 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.

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.

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.

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 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 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.