Objective To investigate the effect of inoculating the associative nitrogen-fixing bacteria strain Paraburkholderia RBCS-17 on the root-associated bacterial community of sugarcane. Methods We employed 16S rRNA gene high-throughput sequencing combined with QIIME 2-based bioinformatics analysis to investigate the effects of inoculating Paraburkholderia RBCS-17 on the alpha diversity, beta diversity, composition, and co-occurrence network of the root-associated bacterial community of sugarcane. Results Inoculating Paraburkholderia RBCS-17 did not significantly affect the diversity but significantly changed the structure of the root-associated bacterial community. Further studies showed that the inoculation significantly increased the relative abundance of Burkholderia, Dyella, and Pseudomonas, while reducing the relative abundance of certain potentially detrimental bacteria such as Ralstonia. In addition, the inoculation altered the key module composition of the bacterial co-occurrence network in sugarcane roots, which suggested that inoculation might influence potential bacterial interactions. Conclusion Inoculating the associative nitrogen-fixing bacterial strain Paraburkholderia RBCS-17 modified the root-associated bacterial community structure of sugarcane, promoting the enrichment of potentially beneficial bacteria while suppressing potentially harmful ones. These findings provide new insights into the intricate interactions among associative nitrogen-fixing bacteria, host plants, and indigenous bacteria.

Objective Screening fungi with the ability to degrade polyacrylamide (PAM) and analyzing the characteristics of their degradation products can provide a basis for clarifying the degradation mechanism. Methods Fungi capable of degrading PAM were screened from bauxite sludge and used to construct a composite fungal community, with the degradation products and morphological characteristics being determined under optimized conditions. Results The results showed that the three strains of fungi screened out were Trichoderma asperellum, Aspergillus flavus, and Aspergillus niger, which showed the degradation rates of 27.35%, 25.20%, and 23.04%, respectively, for PAM. The degradation conditions were optimized by the response surface method as initial pH 5.5, inoculum amount of 5.1%, and incubation temperature of 32 ℃, under which the fungal complex constructed with T. asperellum and A. flavus showed the PAM degradation rate of 45.44%, a viscosity reduction rate of 84.57%, and laccase and urease activities of 13.90 U/mL and 17.70 U/mL, respectively. A large number of hollows and cavities were formed on the surface of PAM after degradation. In addition, mycelial biofilm was observed on the surface. The degradation products showed -COOH and -OH functional groups. Conclusion The above results suggest that the fungal complex may degrade PAM into small molecules through the synergistic effects of mycelial physical erosion and extracellular enzymes.

Objective We compared the rhizosphere microbial interaction network structure and keystone taxon identification arising from distinct network construction algorithms, aiming to clarify the characteristics and advantages of each algorithm in inferring microbial interactions and identifying keystone taxa, thereby providing a theoretical basis for methodological selection. Methods Taking the rhizosphere microbial community of Camphora migao (a rare plant) as the model system, we constructed molecular ecological networks with three mainstream algorithms: sparse correlations for compositional data (SparCC), random matrix theory (RMT), and co-occurrence network (CoNet). We comprehensively compared network structural features and keystone taxon identification across algorithms by integrating PICRUSt2 functional prediction with keystone taxa-environmental factor correlation analysis. Results Network construction algorithms significantly influenced the topological properties of networks. SparCC generated highly modular networks (relative modularity index, RM=1.31) with distinct interaction segregation (edge connectivity=0). RMT produced a single-module structure (RM=0.78) and homogeneous connectivity (closeness centralization index=0.22). Integration of 26.0% negative correlations in CoNet reduced modularity (RM=0.95), increased network diameter (33.22 steps), and decreased robustness. Keystone taxon identification was method-dependent. Specifically, CoNet, SparCC, and RMT identified 224.00, 44.00, and 19.00 keystone taxa, respectively, with<9.2% cross-method overlap. Rhizobiales and Acidobacteriales were consistently identified as core keystone taxa by all methods, demonstrating cross-algorithm stability. The correlation analysis with environmental factors confirmed that these shared taxa significantly correlated with β-glucosidase activity, validating their role in cellulose degradation and highlighting methodological consistency in identifying key ecological processes. Conclusion The three algorithms exhibited complementary strengths: CoNet resolved complex competitive interactions; SparCC reliably assessed functional stability; RMT uncovered core functional modules. The correlation analysis with environmental factors validated the cellulose degradation function of keystone taxa, with high cross-method consistency in core ecological process identification. Our work provides a theoretical foundation for elucidating plant-microbe interactions and optimizing microbial network construction.

Objective The formation of neutrophil extracellular traps (NETs) induced by influenza A virus (IAV) subtype H1N1 was investigated both qualitatively and quantitatively. Methods Mouse bone marrow neutrophils were isolated, purified, and characterized. NETs were induced in vitro using lipopolysaccharide (LPS) and phorbol 12-myristate 13-acetate (PMA). Additionally, IAV groups with three different titers: one hundred 50% tissue culture infective doses (100 TCID₅₀), 50 TCID₅₀, and 25 TCID₅₀ as well as the normal control group were established, and the intracellular nucleoprotein (NP) mRNA expression levels of the IAV groups were detected using reverse transcription quantitative polymerase chain reaction (RT-qPCR). The effect of each factor on neutrophils was assessed by measuring the concentration of circulating cell-free DNA (cfDNA) in the supernatant of each group using the quantitative SYTOX Green staining method. The NETs structure in each group of cells was observed under a fluorescence microscope after Hoechst 33342 staining. An immunofluorescence assay was performed to detect the expression levels of NET characteristic markers citrullinated histone H3 (CitH3), peptidylarginine deiminase 4 (PAD4), myeloperoxidase (MPO), and neutrophil elastase (NE) proteins, as well as the nuclear co-localization and fluorescence intensity of PAD4 with CitH3, and MPO with NE in each group. The levels of reactive oxygen species (ROS) were determined by using a fluorescent probe assay, and the levels of intracellular CitH3 protein formation were determined by using Western blotting. Results The activity of neutrophils isolated from mouse bone marrow reached 98%, with purities of ≥87%. The expression levels of NP mRNA in the IAV groups were significantly higher than those in the control group. Compared with the control group, the cfDNA levels of the PMA, LPS, and IAV groups were significantly increased, with significant increases in the web-like structures of NETs. The immunofluorescence assay showed that the relative expression levels of MPO, NE, PAD4, and CitH3 proteins were elevated to varying degrees, with the co-localization of PAD4/CitH3 or MPO/NE increased after IAV infection. Moreover, the peak of MPO protein expression was observed before that of NE protein, whereas CitH3 expression paralleled that of PAD4 protein. Additionally, the ROS level was elevated, and the level of CitH3 protein formation was also significantly increased. Conclusion Stimulation of neutrophils by IAV (H1N1) induces NET formation, which may be related to the increased intracellular ROS and PAD4 levels.

Helicobacter pylori infection is a major causative factor for chronic gastritis and gastric cancer, while current antibiotic therapies are facing increasingly severe resistance. Probiotics have emerged as a promising approach for anti-H. pylori research due to their high safety. Notably, certain Lactobacillus strains have been demonstrated to effectively alleviate H. pylori-induced inflammatory responses, yet their underlying molecular regulatory mechanisms remain unclear. Objective To investigate the molecular mechanism by which Lactiplantibacillus plantarum ZJ316 inhibits the H. pylori-induced inflammatory response by modulating the p38 mitogen-activated protein kinase (MAPK) signaling pathway in the host cells and assess the regulatory effect of this strain on gastric microecological homeostasis, thus providing a theoretical basis for the development of probiotic therapeutics targeting H. pylori. Methods We integrated cell experiments (human gastric adenocarcinoma cell line AGS) and animal experiments (C57BL/6 mice) and employed Western blotting (to determine the phosphorylation level of p38 MAPK), transcriptome sequencing and RT-qPCR (to analyze differential gene expression), ELISA [to determine the levels of inflammatory cytokines interleukin (IL)-8 and IL-10], 16S rRNA gene sequencing (to unveil the gastric flora structure), and hematoxylin-eosin staining (to observe gastric mucosal damage) to systematically study the intervention effect of L. plantarum ZJ316 on H. pylori infection. Results At the cellular level, L. plantarum ZJ316 inhibited H. pylori-induced p38 MAPK phosphorylation, with the inhibition rates of 21.95% and 33.72% at the time points of 1 h and 2 h, respectively (P<0.01). It down-regulated the expression of pathway genes such as MAP3K8 and FOS, and lowered the mRNA levels of the pro-inflammatory cytokines interferon-γ, tumor necrosis factor-α, and IL-6 by 43.26%, 35.95%, and 51.91%, respectively (P<0.01). The combination of this strain with adezmapimod, a p38 MAPK-specific inhibitor, further enhanced the inhibitory effect. In animal experiments, L. plantarum ZJ316 significantly attenuated gastric mucosal pathological injury and inflammatory response, and 16S rRNA gene sequencing revealed that ZJ316 reduced the relative abundance of pathogenic Pseudomonadota and significantly increased the relative abundance of Bacillota [(54.8±9.9)% vs. (27.8±5.9)%, P<0.01] in the stomach. When ZJ316 was combined with adezmapimod, the relative abundance of Bacteroidota was elevated [(58.5±5.2)% vs. (47.8±6.9)%, P<0.05], and specific beneficial genera such as Alistipes were synergistically enriched (an increase of 69.52% compared with the H. pylori group). Conclusion L. plantarum ZJ316 alleviated the inflammatory response triggered by H. pylori infection by inhibiting the p38 MAPK pathway and remodeled the gastric microecological structure. The findings provide a theoretical basis for the inhibition of H. pylori-induced inflammation by lactobacilli and the development of probiotic-based functional foods.

Objective To address the environmental pollution caused by polyethylene terephthalate (PET), we screened functional bacterial strains capable of degrading PET and analyzed their growth and degradation characteristics, aiming to provide theoretical support and microbial resources for PET bioremediation. Methods Bacterial strains capable of degrading PET were isolated from the soil samples collected around a landfill site. The selected strain was identified based on morphological characteristics, physiological and biochemical properties, and 16S rRNA gene sequencing. The surface morphology and chemical group changes of PET films before and after degradation were analyzed by scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), and water contact angle (WCA) measurements. The types and concentrations of degradation products were quantified via high-performance liquid chromatography (HPLC). Results A PET-degrading strain, designated YH-1, was successfully isolated and identified as a member of the Cellulosimicrobium sp. genus based on 16S rRNA analysis. The optimal growth conditions for strain YH-1 were 35 °C, pH 7.0, and 1% salinity, and the strain exhibited robust growth within the ranges of pH 6.0-10.0 and 1%-4% salinity. After 6 days of incubation, YH-1 achieved a PET weight loss rate of 1.90%. HPLC revealed that terephthalic acid (TPA) and bis(2-hydroxyethyl) terephthalate (BHET) were the main degradation products, with concentrations of 3.87 mg/L and 4.70 mg/L, respectively. SEM images showed obvious surface roughening and cracking of the PET film after degradation, while FTIR revealed changes in functional groups. WCA measurements showed a reduction in contact angle from 79.385° to 65.052°, indicating enhanced hydrophilicity of the degraded PET film surface. Conclusion Strain YH-1 demonstrates good environmental adaptability and PET degradation potential. It can disrupt the PET film surface and generate typical degradation products. The findings lay a foundation for further development of PET-degrading microbial resources and exploration of degradation mechanisms.

Objective Saline-alkali soil is an important farmland resource in China. This study explored the effects of a bio-organic fertilizer fortified with a functional strain isolated from the crop rhizosphere of saline-alkali soil on the growth and the grain yield and quality of peanut plants in saline-alkali soil. The results are expected provide a solution for the development of specific microbial organic fertilizers for saline-alkali soil. Methods We first compared the rhizosphere bacterial communities of peanut plants growing in low-salt stress and non-salt stress soils, and identified the potential taxa improving the salt tolerance of plants that were enriched in the peanut rhizosphere under low-salt stress. A strain named HS6 capable of enhancing the salt tolerance of peanut plants was isolated from the rhizosphere soil of peanut plants. It was preliminarily identified as Bacillus paralicheniformis HS6. A microbial organic fertilizer was prepared by combining this strain with organic fertilizer. A field experiment was carried out in coastal saline-alkali land, including a control treatment (CK: decomposed organic fertilizer) and treatment 1 (T1: decomposed organic fertilizer supplemented with the cells of strain HS6). The growth and yield-related indicators of peanut plants were determined by counting and weighting, and the quality of peanuts was determined by the Kjeldahl method and the Soxhlet extraction method. Results The soil salt concentration higher than 0.3% significantly inhibited the growth of peanut plants. The principal component analysis revealed a significant difference in the peanut rhizosphere bacterial communities between low-salt stress (0.3%) and non-salt stress soils. Under low-salt stress, 22 differential taxa, mainly including Bacillaceae, were positively enriched in the peanut rhizosphere. The application of the organic fertilizer prepared with strain HS6 significantly promoted the growth, enhanced the biomass accumulation, and increased the number of nodules of peanut plants. The number of peanut nodules of T1 was 5 times that of CK. Moreover, the functional microbial fertilizer improved the yield and quality of peanuts. Compared with CK, T1 decreased the crude protein content of peanuts by 13.84%, while increasing the crude fat content of peanuts by 5.63%. Conclusion Low-salt stress can promote the enrichment of functional microbial taxa capable of enhancing salt tolerance in the peanut rhizosphere. The microbial organic fertilizer fortified with the functional strain enriched in the rhizosphere under salt stress can significantly improve the yield and quality of peanuts, demonstrating the potential to serve as a special microbial fertilizer for saline-alkali soil.

Objective To compare the stress tolerance of recombinant Mycobacterium smegmatis strains Ms-PPE61 and Ms-Vec under different external stress conditions, investigate the activation/inhibition levels of the mitogen-activated protein kinase (MAPK)/nuclear factor (NF)-κB signaling pathway following their infection of macrophages, and explore differences in inflammatory cytokine expression after infection of RAW264.7 cells. Methods Ms-Vec and Ms-PPE61 were constructed and cultured to the logarithmic growth phase before being subjected to acidic, SDS, and H₂O₂ conditions. Colony-forming units (CFUs) were measured at different time points. Proteins were extracted from cells collected 1-48 h post-infection (hpi), and the expression levels of signaling pathway marker molecules were determined by Western blotting. The interleukin (IL)-6, tumor necrosis factor (TNF)-α, and IL-1β concentrations in the supernatants of RAW264.7 cells infected with Ms-Vec and Ms-PPE61 were measured by ELISA at 24 hpi and 48 hpi. GraphPad Prism 7.0 was used for analysis of variance of the data, and P<0.05 was considered significant. Results PCR revealed the presence of a target band in Ms-PPE61 but not in Ms-Vec. Coomassie brilliant blue staining confirmed consistent protein loading. Western blotting showed that Ms-PPE61 expressed a ~42 kDa Flag fusion protein, while Ms-Vec did not. Ultra-high-speed centrifugation was performed to separate the components of M. smegmatis. Western blotting revealed that the cytoplasmic marker protein GroES was expressed in the cytoplasmic fractions of both Ms-Vec and Ms-PPE61, while the Flag-tagged target protein was exclusively present in the cell wall of Ms-PPE61. After treatment under acidic conditions (pH 3.0) for 3 h, the survival rate of Ms-PPE61 was higher than that of Ms-Vec (P<0.000 1), while the survival rate showed no significant difference after treatment for 6 h and 9 h (P>0.05). After treatment with 0.2% SDS for 3, 6, and 9 h, the survival rate of Ms-PPE61 was higher than that of Ms-Vec (P<0.000 1). Similarly, after H₂O₂ treatment for 3 h and 6 h, the survival rate of Ms-PPE61 was higher than that of Ms-Vec (P<0.000 1). Western blotting showed that the Ms-PPE61 group had significantly lower p-p38 and p-ERK levels at 48 hpi and higher IκB-α levels at all time points than the Ms-Vec group. ELISA results indicated no differences in TNF-α secretion between the Ms-PPE61 and Ms-Vec groups at 24 hpi and 48 hpi (P>0.05), while the Ms-PPE61 group had lower IL-6 levels at 24 hpi and 48 hpi (P<0.000 1) and lower IL-1β level at 48 hpi (P<0.01) than the Ms-Vec group. Conclusion PPE61 can enhance the tolerance of recombinant Mycobacterium smegmatis to acidic, SDS and H₂O₂ stress, inhibit the MAPK and NF-κB signaling pathways by down-regulating the expression of p-p38 and p-ERK and up-regulating the expression of IκB-α, and reduce the secretion of IL-6 (significantly at both 24 h and 48 h) and IL-1β (significantly at 48 h) in macrophages, but has no significant effect on the secretion of TNF-α.

Heterotrophic nitrifying-aerobic denitrifying (HN-AD) bacteria can simultaneously complete nitrification and denitrification processes under aerobic conditions, significantly simplifying wastewater treatment procedures. These strains exhibit rapid growth, tolerance to extreme environments, and are widely applied in various wastewater treatments. This review summarizes the nitrogen removal pathways of HN-AD bacteria and highlights the potential of novel immobilization carriers (composite materials, magnetic nanocarriers, and biochar) for nitrogen removal across diverse wastewater sectors. It focuses on elucidating the fundamental principles, application cases, current research status, and future prospects of emerging immobilization technologies, including biomimetic mineralization immobilization, electrospun fiber immobilization, and 3D printing carrier immobilization. The enhanced effects of novel immobilization strategies on improving nitrogen removal efficiency and system stability are discussed. Finally, challenges and future prospects for HN-AD bacterial immobilization technology during fabrication and application are outlined.

In recent years, as the antibiotic resistance of bacteria is aggravating, bacterial infections have brought severe challenges to disease prevention and control. Rapid and accurate identification of bacteria and their basic characteristics is extremely important for disease prevention and control, medical diagnosis, and scientific research. Compared with conventional detection methods such as plate culture counting, polymerase chain reaction (PCR), and adenosine triphosphate bioluminescence (ATP) bioluminescence, single-cell Raman spectroscopy has shown advantages and broad application prospects in bacterial classification and identification, bacterial pathogenicity and antibiotic resistance detection, and bacterial viability evaluation. This paper reviews the application of single-cell Raman technology in the field of bacteria, aiming to provide technical and application reference for practitioners engaged in the research on bacteria and Raman technology.