Objective The rapid increase in wastewater discharge from animal husbandry has caused severe environmental pollution. Identifying efficient heterotrophic nitrifying-aerobic denitrifying bacteria and investigating their denitrification mechanisms are of great theoretical and practical importance for mitigating nitrogen pollution in the wastewater. Methods A strain exceling in heterotrophic nitrification-aerobic denitrification (HN-AD) was isolated and from activated sludge in pig farms. Culture conditions were optimized by response surface methodology. We evaluated the inorganic nitrogen-transforming capacity of the strain by assessing its utilization efficiency of single and mixed nitrogen sources and through nitrogen balance analysis. The completeness of the denitrification process was confirmed via gas chromatographic measurements of N₂ and N₂O. Finally, the nitrogen removal pathways and underlying mechanisms were elucidated through whole-genome analysis. Results The successfully isolated strain Klebsiella sp. WH-E exhibited excellent HN-AD capabilities. The growth conditions of the strain were optimized as follows: sodium citrate as the carbon source, 34.18 °C, initial pH 7.1, a C/N ratio of 14.53, and a shaking speed of 159.59 r/min. When the strain was cultured with ammonium, nitrate, or nitrite as the sole nitrogen source, the nitrogen removal rates were 99.80%, 81.54%, and 80.00%, respectively. Furthermore, when ammonium was the sole nitrogen source, 35.84% and 35.91% of nitrogen were converted into cellular nitrogen and gaseous nitrogen, respectively. When ammonia nitrogen was combined with nitrate nitrogen as mixed nitrogen sources, the nitrogen removal rate was 100.00%; When ammonia nitrogen was combined with nitrite nitrogen as mixed nitrogen sources, the ammonia nitrogen removal rate was 100.00%, and the nitrite nitrogen removal rate was 91.97%, respectively. Whole-genome sequencing identified several nitrogen metabolism-related functional genes, including glnB, norVWR, narGHI, nasBC, and nirBD. Conclusion Klebsiella sp. WH-E possesses three nitrogen metabolism pathways: ammonium assimilation, nitrification-denitrification, and nitrate assimilation and dissimilation. This study confirms the applicability of Klebsiella sp. WH-E for nitrogen removal from full-scale piggery wastewater and establishes a solid theoretical foundation for its engineering applications.

Nitrogen deposition is a major driver shaping the structures and functions of forest ecosystems worldwide. When nitrogen inputs exceed ecosystem critical loads (CLs), significant changes in the diversity and abundance of understory herbaceous plants can occur. This study aims to systematically compile and integrate critical load data for understory herbaceous plants in response to nitrogen deposition across three mycorrhizal types: arbuscular mycorrhiza (AM), ectomycorrhiza (ECM), and mixed arbuscular-ectomycorrhizal forests (AM+ECM), in forests. By establishing a dedicated and standardized database, this work facilitates comparisons of herbaceous plant responses to nitrogen inputs among different mycorrhizal types in forests and provides a scientific basis for assessing the impacts of nitrogen deposition on forest microbe-plant systems. On the basis of the published literature and the global nitrogen deposition critical load database developed by Wilkins et al., relevant data were systematically collected, screened, and standardized to construct the Database of Critical Loads of Nitrogen Deposition for Understory Herbaceous Plants across Different Mycorrhizal Types in Forests. All critical load values were consistently derived via the threshold indicator taxa analysis (TITAN) method. A rigorous quality control workflow was applied, including cross-validation of mycorrhizal types, outlier detection and treatment, and data standardization. The database contains 3 592 standardized records. The core data table includes the following fields: Latin name of herbaceous plant species, forest alliance, mycorrhizal types (AM, ECM, or AM+ECM), species-level critical load values (zenv.cp) estimated by TITAN with corresponding bootstrap uncertainty intervals (5th, 10th, 50th, 90th, and 95th percentiles), response direction (increase or decrease in abundance), purity and reliability metrics, community-level change points (CCP), and associated environmental metadata. The database covers the three major mycorrhizal types as well as graminoid and non-graminoid herbaceous functional groups in forests. This database represents the first large-scale, standardized database explicitly focusing on the relationships among mycorrhizal types, understory herbaceous plants, and nitrogen deposition critical loads in forests. Its standardized structure, transparent metadata, and stringent quality control procedures ensure its reliability for future research and applications, including nitrogen deposition risk assessment, comparative analyses of mycorrhizal functions, ecological model parameterization, and the formulation of biodiversity conservation strategies.

As a crucial group of probiotics, lactic acid bacteria (LAB) play a vital role in the gut microbial ecosystem of insects. This article comprehensively reviewed the species composition, ecological functions, and practical values of LAB in the guts of major insect orders, including Hymenoptera, Diptera, Coleoptera, Hemiptera, Lepidoptera, Blattodea, and Orthoptera. To date, multiple LAB genera including Lactobacillus, Lactococcus, Leuconostoc, Pediococcus, Enterococcus, Bifidobacterium, and Weissella were successfully identified from insect guts. The community composition of these bacteria was shaped by factors such as host phylogeny, dietary traits, developmental stages, gut microenvironment, and external ecological conditions. The LAB in insect guts not only assist the hosts in degrading recalcitrant complexes by secreting extracellular enzymes but also inhibit pathogens through the synthesis of antimicrobial substances such as bacteriocins. Additionally, they modulate host immune responses, promote growth and development, regulate host behavior, and participate in the metabolic detoxification of xenobiotics, thereby enhancing host survival and adaptability. Furthermore, insect-derived LAB held great potential in the production of resource insects, pest management, agricultural waste utilization, and green manufacturing. In summary, insect guts represent an important reservoir for the discovery and isolation of novel LAB.

The gut microbiota plays a crucial role in promoting food digestion in animals. However, the impact of cross-species microbiota transplantation from donors with different dietary habits on the host food digestion capacity remains unclear. Objective To investigate the role of cross-species microbiota transplantation in regulating the digestive system adaptability, metabolic functions, reproduction, stress responses, and gut microbiota structure of the host. Methods We utilized New Zealand white rabbits (Oryctolagus cuniculus), a herbivorous species, and C57BL/6J mice, an omnivorous species, as donors and recipients of gut microbiota, respectively. The mice were allocated into three groups: a control group on a normal diet (Con), a group on a high-fiber diet (TS), and a group on a high-fiber diet supplemented with rabbit fecal microbiota transplantation (OC). This study was designed to evaluate various physiological and biochemical parameters, including body weight, food intake, absolute and relative organ weights (both wet weight and organ-to-body weight ratio), morphometric indices (length and diameter) of the small intestine, sperm concentration, and serum corticosterone level, in mice. Additionally, we performed 16S rRNA gene sequencing targeting the V3-V4 hypervariable region to characterize the composition of fecal microbiota. Results A high-fiber diet significantly increased the food intake, small intestine length, and serum corticosterone level, while significantly reducing the body weight, liver and spleen wet weights, liver/body weight ratio, spleen/body weight ratio, and sperm concentration in mice. Moreover, it increased the alpha diversity of the gut microbiota, decreased the Bacillota-to-Bacteroidota ratio, and reduced the relative abundance of probiotics (such as Ligilactobacillus). Transplantation of the gut microbiota from rabbits increased the wet weight of the epididymis and the epididymis/body weight ratio, while significantly reducing the liver/body weight ratio and the serum corticosterone level in recipient mice. Furthermore, a high-fiber diet significantly increased the relative abundance of the fiber-degrading bacterial family (Oscillospiraceae) and the gut health-associated bacterial genus (Colidextribacter). After the transplantation of rabbit gut microbiota into mice, the relative abundance of Oscillospiraceae and Colidextribacter in mice increased significantly. Conclusion The high-fiber diet has adverse effects on omnivores. Although the microbiota transplantation from herbivores does not significantly improve the host ability to digest fiber, it changes the gut microbiota structure of omnivores, playing a positive role in improving their digestion, reproduction, metabolism, and stress responses. Future research needs to further determine the optimal levels of dietary fiber for omnivores and the dosage of microbiota transplantation from herbivores, as well as their synergistic effects and underlying mechanisms in improving animal health. This study provides a reference for exploring the role of gut microbiota in animal adaptation to dietary changes in natural environments and lays a foundation for future research on improving the utilization of high-fiber foods by omnivorous domestic animals.

Objective To prepare reference material (RM) for positive serum specific for genotype Ⅱ African swine fever virus (ASFV) for serological detection, quality control, and proficiency testing (PT). Methods The anti-serum collected from specific pathogen-free (SPF) swine immunized with inactivated genotype Ⅱ ASFV was used as raw material for the preparation of RM. Indirect enzyme-linked immunosorbent assay (iELISA) was employed to evaluate the purity, specificity, homogeneity, and stability of RM. In addition, RM was characterized by nine laboratories and applied in clinical trials by three laboratories. Results A total of five hundred bottles of RM for positive serum specific for genotype Ⅱ ASFV strain HLJ/18 were successfully prepared. The results indicated that the RM we prepared was pure, homogenous, and free of exogenous virus contamination, showing good specificity, homogeneity, and stability. The RM was stable for at least 18 months when it was stored at -20 ℃ and for at least 7 days at 4 ℃, 25 ℃, and 37 ℃. The characterization by the nine laboratories showed that the RM was positive for antibodies against genotype Ⅱ ASFV. Conclusion The positive serum specific for genotype Ⅱ ASFV strain HLJ/18 has successfully been prepared, providing critical material for ASF detection and diagnosis.

Objective In view of the production and environmental issues caused by excessively high nicotine content in upper tobacco leaves, this study aims to decipher the molecular mechanism of nicotine degradation by an efficient nicotine-degrading strain Pu17 screened out in the previous study via genomic approaches. Methods The taxonomic status of the strain was determined by average nucleotide identity (ANI) analysis. Whole genome sequencing and annotation were employed to clarify the nicotine metabolic pathway. Key intermediates during degradation were detected by MS/MS. Live plant trials were conducted to explore the optimal application method for nicotine reduction. Results Phylogenetic analysis revealed an ANI value of 96.51% between Pu17 and Peanarthrobacter ureafaciens, identifying Pu17 as a strain of P. ureafaciens. The genome of Pu17 was 4.47 Mb in length, with the G+C content of 63.34%, encoding 4 155 proteins. Functional annotation and comparative genomics identified unique gene clusters related to heavy metal resistance, cell surface synthesis, and metabolic potential in Pu17, which constituted its environmental adaptation strategy. Metabolite analysis detected key intermediates such as 6-hydroxypseudooxynicotine. This result, combined with that of genomic analysis, confirmed that Pu17 degraded nicotine via the pyridine pathway, with key genes (e.g., nboR, mao, and 6-hlno) primarily located on plasmids. Efficacy evaluation demonstrated that the Pu17 fermentation broth effectively reduced nicotine content in tobacco plants through both foliar spraying and root irrigation, achieving a maximum degradation rate of 14.00% in live leaves. Conclusion This study systematically elucidates the molecular mechanism and application potential of P. ureafaciens Pu17 for nicotine degradation from genomic, metabolomic, and application perspectives. It provides a theoretical basis and microbial resources for the development of bioremediation technologies for tobacco waste and harm reduction.

Vulvovaginal candidiasis (VVC) is a prevalent fungal infection affecting the female reproductive tract. Although conventional therapeutic approaches for VVC are relatively well-established, they still exhibit certain limitations. Pulsatilla decoction, a classic traditional Chinese medicine formula, has demonstrated significant clinical efficacy in the treatment of VVC. However, its precise mechanism of action remains incompletely elucidated. Objective To clarify the therapeutic mechanism of the n-butanol extract of Pulsatilla decoction (BEPD) on VVC through network pharmacology and animal experiments. Methods A mouse model of VVC was established and the therapeutic effect of BEPD on VVC was evaluated. Network pharmacology was employed to screen the potential signaling pathways of BEPD on VVC. Western blotting, immunofluorescence, immunohistochemistry, and real-time fluorescence quantitative PCR were employed to measure the changes in autophagy, apoptosis, and related pathway proteins in the vaginal mucosa of mice. Results Network pharmacology analysis identified PIK3R1 and AKT1 as key targets of Pulsatilla decoction in exerting antifungal activity against VVC. KEGG pathway enrichment analysis indicated that Pulsatilla decoction exerted its therapeutic effects on VVC by regulating the PI3K-Akt signaling pathway. Animal experiments confirmed that compared with the VVC model group, the BEPD treatment down-regulated the expression of PI3K, p-Akt, and p-mTOR, significantly up-regulated the expression of autophagy-related proteins LC3B and ATG5, significantly inhibited the expression of apoptosis-related proteins Bax and Cleaved-Caspase-3, and significantly promoted the expression of anti-apoptosis-related protein Bcl-2. Conclusion BEPD may promote autophagy and inhibit apoptosis of vaginal epithelial cells by inhibiting the PI3K-Akt-mTOR signaling pathway, thereby restoring the homeostasis of the vaginal mucosal epithelial barrier and alleviating VVC.

Objective To analyze the expression profile of the β-glucan-binding protein (Acβ-GBP) gene of Apis cerana cerana in response to Ascosphaera apis infection and to investigate the impacts of Acβ-GBP knockdown on the larval mortality and the incidence of chalkbrood disease following A. apis infection. These findings will provide a foundation for further functional research. Methods The sequence and structural characteristics of Acβ-GBP were analyzed via bioinformatics approaches. RT-qPCR was employed to investigate the expression profiles of Acβ-GBP in the larval midgut following A. apis infection. Furthermore, RNA interference (RNAi) was utilized to explore the impacts of Acβ-GBP on the larval mortality and the incidence of chalkbrood disease. Results The CDS length of Acβ-GBP was 1 440 bp, encoding a protein with a molecular weight of 54.68 kDa and a grand average of hydropathicity value of -0.22, which contained a typical transmembrane domain and signal peptide. Phylogenetic analysis revealed that β-GBP of A. c. cerana, Apis florea and Apis dorsata clustered into a single major clade. After A. apis infection, the expression level of Acβ-GBP in the midgut of A. c. cerana worker larvae was downregulated at 1-3 days post-infection (dpi) (P<0.05). Following RNA interference (RNAi)-mediated silencing of Acβ-GBP, its expression level was lower than that in the ds-egfp group at 2 and 3 dpi (P<0.01). The cumulative larval mortality and the incidence of chalkbrood disease both increased over the infection time, and the overall mortality was higher than that of the control group (P<0.000 1). Conclusion Acβ-GBP was capable of responding to A. apis infection, and knockdown of Acβ-GBP expression significantly impaired the resistance of honeybee larvae to A. apis. Collectively, β-GBP acts as an important immune recognition protein in A. c. cerana, and plays an important role in defending against fungal invasion.

Objective To address problems such as the poor structure and fertility degradation in strongly acidic soils, we isolated acid-tolerant exopolysaccharide (EPS)-producing microorganisms, constructed a composite microbial inoculant, and evaluated its improvement effects on the structure and comprehensive fertility of acidic soils. Methods Target strains were obtained through primary and secondary screening from strongly acidic soils (pH<4.5) in Nanchuan, Chongqing and Qujing, Yunnan. After their antagonistic activity and plant growth-promoting traits were assessed, a composite microbial inoculant was constructed. A laboratory soil culture experiment was conducted to investigate changes in nutrient contents and aggregate composition in strongly acidic soils treated with different inoculants, comprehensively evaluate the fertility-improving effects of the inoculants, and clarify the correlation between aggregate formation and EPS content. Results Three strains—Paraburkholderia fungorum C3, Burkholderia cepacia A13, and Cystobasidium minutum B14—with acid tolerance and high EPS-producing capabilities were successfully isolated and screened out. All the strains exhibited the capabilities of secreting indole-3-acetic acid (IAA), synthesizing siderophores, and solubilizing phosphorus. When these strains were applied individually or as a composite inoculant to soils, the composite inoculant showed the best effect of improving soil nutrients, increasing the content of soil organic matter, alkaline-hydrolyzable nitrogen, available phosphorus, and available potassium by 4.51%, 13.92%, 4.92%, and 3.71%, respectively. Application of all the inoculants effectively promoted the formation of soil macro-aggregates, among which the single-strain inoculant C3 had the most significant effect in promoting aggregate formation, increasing the soil mean weight diameter (MWD) by 19.39%. The integrated fertility index of the soil treated with the composite inoculant reached 0.61, indicating the optimal comprehensive improvement effect. The single-strain inoculant C3 and the composite inoculant significantly increased the soil EPS content by 53.17% and 35.79%, respectively. Correlation analysis results showed that soil EPS had significantly positive correlations with macro-aggregate content and MWD, significantly promoting the formation and enhancing the stability of soil macro-aggregates. Conclusion The composite inoculant composed of the three acid-tolerant EPS-producing strains screened in this study effectively improved the soil structure and enhanced the integrated soil fertility. These findings lay a theoretical foundation for the development of biological agents for acidic soil remediation.