Objective To investigate the effect of yeast dietary fiber (YDF) on arsenic-induced apoptosis in Saccharomyces cerevisiae and decipher the possible mechanism. Methods The relative survival rate, apoptosis, and antioxidant indicators were determined by the spread plate method, spectrophotometry, fluorescence microscopy, and RT-qPCR. Results The exposure to arsenic significantly decreased the relative survival rate, elevated the intracellular reactive oxygen species (ROS) and malondialdehyde (MDA) levels, and induced apoptosis. However, in the presence of YDF (0.5 mg/mL or 1.0 mg/mL) and arsenic, the arsenic-induced toxic effects were effectively attenuated, which was evidenced by increases in the relative survival rate, content of glutathione, activities of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPX), and relative expression of antioxidant enzyme genes (SOD1, CTA1, CTT1, and GPX2). Moreover, the treatment with both YDF and arsenic lowered the ROS and the MDA levels, significantly down-regulated the relative expression levels of pro-apoptotic genes (AIF1, NMAⅢ, and NUC1), and significantly reduced apoptotic cells compared with the treatment with arsenic alone. Conclusion YDF regulates the antioxidant system to attenuate the arsenic-induced cytotoxicity, thereby alleviating the arsenic-induced apoptosis.

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 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.

Objective Developing plant-microbe combined techniques is significant for addressing the problem of declining cropland quality in China and enriching the approaches for biological remediation of degraded soils. Methods The plant growth-promoting traits of Burkholderia sp. YQ9 were determined by assessing its cellulase, protease, and ammonia production. Subsequently, a pot experiment was conducted to evaluate the effects of different dilutions of Burkholderia sp. YQ9 inoculant and culture medium on the growth of white clover (Trifolium repens) and the physicochemical properties of the rhizosphere soil. Furthermore, high-throughput sequencing was employed to analyze the impacts of different treatments on the structure of the rhizosphere microbial community. Results Burkholderia sp. YQ9 exhibited plant growth-promoting traits, being capable of producing cellulase, protease, and ammonia. White clover improved the pH environment of the rhizosphere soil. Application of the original inoculant of Burkholderia sp. YQ9 promoted white clover growth, significantly enhancing the content of soluble protein and soluble sugar in the shoots and leaves, as well as the levels of available phosphorus and available potassium in the rhizosphere soil, thereby facilitating organic matter decomposition. Analysis of microbial alpha diversity in the rhizosphere soil revealed that both the original inoculant of Burkholderia sp. YQ9 and the culture medium significantly reduced the richness, diversity, and evenness of both fungal and bacterial communities in the rhizosphere soil and altered the composition of the soil microbial community. The correlation analysis further indicated that the microbial community in the rhizosphere soil was correlated with white clover growth and soil physicochemical properties. Conclusion Burkholderia sp. YQ9 not only promoted the growth of white clover but also modified the composition of the rhizosphere microbial community and improved the soil fertility. These findings provide microbial augmentation-based technical support for enhancing cropland quality.

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.

Agrobacteriumtumefaciens, a classic model organism for plant-microbe interaction research, is a valuable transgenic tool for plants. Phenolic acids secreted by plants after injury can affect the infection of the host by A. tumefaciens. Objective This study investigated the transcription factor PcaR of A. tumefaciens regarding its effects on the metabolism of simple phenolic acids, regulation of the target gene, and effect on the bacterial tumorigenicity in host plants. Methods The A. tumefaciens strain with atu4546 knockout (Δatu4546) and the complement strain C-Δatu4546 were constructed via the suicide plasmid pEX18Km and the plasmid pUCA19 with a strong promoter, respectively. Both Δatu4546 and C-Δatu4546 were tested for growth with p-hydroxybenzoic acid or protocatechuic acid as the sole carbon source and tumorigenicity on carrot stems and Kalanchoe pinnata leaves. In the wild-type strain C58 and Δatu4546, the reporter gene was in situ inserted into the downstream region of the metabolic target gene atu4549. The regulatory link between atu4546 and the target gene was examined based on the β-galactosidase activity. To investigate the self-regulation of PcaR, we constructed the atu4546 self-promoter reporter plasmid. To identify the binding sites of PcaR, we constructed the upstream promoter region reporter plasmid of the target gene to remove or replace the predicted binding sites and then determined the β-galactosidase activity. Results The knockout of atu4546 did not affect the growth of A. tumefaciens on sucrose, but led to the inability to use p-hydroxybenzoic acid or protocatechuic acid as the sole carbon source. The growth was restored after atu4546 was complemented. The tumor weights of carrot stems and K. pinnata leaves infected by Δatu4546 decreased by 34.90% and 52.58%, respectively, and the number of colonies per 0.1 g tumor decreased by 72.19% and 80.54%, respectively. The knockout of atu4546 led to a 102.04% increase in its own promoter activity, which suggested that atu4546 negatively regulated its own expression. Atu4546 boosted the expression of the atu4547-atu4549 gene cluster, as evidenced by a 74.86% decrease in β-galactosidase activity downstream of the target gene in Δatu4546 compared with that in the wild type. The promoter region sequence alteration experiment identified GTGCGATATATACGAAC as the binding site of PcaR. Conclusion This study shows that the transcription factor PcaR is involved in phenolic acid catabolism, negatively regulates itself and stimulates the transcription of the downstream gene pcaIJF. The binding site of PcaR to the target gene is GTGCGATATACGAAC. The knockout of PcaR attenuates the pathogenicity of A. tumefaciens. This study reveals the dual regulation mechanism in the phenolic acid metabolism-pathogenic signaling pathway and expands the theoretical cognition of plant-microbe interactions.

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 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 Nitrous oxide (N₂O)-reducing microbes are the only known microbial group capable of eliminating N₂O. The abundance, diversity, community structure, and influencing factors of their functional gene (nosZ) are critical for N₂O removal. Cunninghamia lanceolata is a widely planted timber species in southern China, and its rhizosphere represents a hotspot for both N₂O production and reduction. However, the spatial distribution pattern of nosZ Ⅰ genes and their driving factors in the rhizosphere soils of C. lanceolata plantations remain unclear. Methods We investigated the rhizosphere soils of C. lanceolata plantations from five state-owned forest farms—Qiujiashan, Wuyi, Guanzhuang, Xiayang, and Xiapu—in Fujian Province. Quantitative PCR and amplicon sequencing were employed to analyze the abundance, diversity, and community structure of nosZ Ⅰ genes and to identify their key environmental drivers. Results Dissolved organic carbon concentrations in rhizosphere soils ranged from 6.91 mg/kg to 23.52 mg/kg, being significantly lower in Guanzhuang and Xiayang than in Wuyi, Qiujiashan, and Xiapu. The nosZ I gene abundance ranged from 4.76×10⁶ copies/g to 36.50×10⁶ copies/g, reaching 36.50×10⁶ copies/g and 29.08×10⁶ copies/g in Guanzhuang and Xiayang, respectively, which significantly exceeded those in Qiujiashan, Wuyi, and Xiapu. Dissolved organic carbon emerged as the primary driver of nosZ I gene abundance, which implied that low dissolved organic carbon may promote the proliferation of N₂O-reducing bacteria. The Shannon index of nosZ I genes ranged from 4.41 to 5.67, being significantly higher in Xiayang than in Wuyi and Xiapu and the lowest in Xiapu. Total carbon was the key factor affecting the Shannon index. The nosZ I community structures in Qiujiashan, Guanzhuang, and Xiapu were similar, whereas that of Xiayang was significantly different from the others. Soil pH was identified as the main driver of community structure, and Xiayang had a significantly higher pH than the other sites. The dominant bacterial class in the rhizosphere soils of all five forest farms was Gammaproteobacteria. Xiayang had significantly lower relative abundance of Gammaproteobacteria but significantly higher relative abundance of Alphaproteobacteria than other farms. Conclusion Soil carbon content and pH are key environmental factors regulating the abundance, diversity, and community structure of N₂O-reducing bacteria in the rhizosphere soils of C. lanceolata plantations, potentially influencing N₂O removal and mitigation potential. Therefore, the management strategies for C. lanceolata plantations should consider regulating soil carbon content and pH to optimize N₂O mitigation effects and alleviate global climate change.

Objective To investigate the community structure, network complexity, and stability of soil bacteria harboring the alkaline phosphatase gene (phoD) under the application of organic amendments, elucidating their regulatory mechanisms in microbially mediated soil phosphorus (P) transformation and availability. Methods We conducted the experiment within a 13-year long-term maize field trial located in Ya’an, Sichuan. The experiment comprised three mineral P fertilizer treatments: 0, 75, and 150 kg/hm² (designated as P0, P1, and P2, respectively). In 2018, a split-plot design was implemented with organic amendment treatments, where mineral P application was reduced by 30% and supplemented with pig manure (P0+M, 70% P1+M, and 70% P2+M treatments). The phoD-harboring bacterial community structure was characterized by high-throughput sequencing and bioinformatic analyses, which revealed the effects of organic amendments with varying P supply levels on phoD-harboring bacterial communities and their regulation of soil available P. Results As the P supply level increased, both mineral and organic amendments significantly increased the content of soil organic matter (SOM), Olsen-P, and organic P (Po), while significantly decreasing soil pH. P levels and organic amendments markedly altered the community composition and network characteristics of phoD-harboring bacteria. Under low-P conditions (P0, P0+M), Bradyrhizobium icense emerged as both the dominant and indicator species, with its relative abundance decreasing significantly as P application increased. Under P-amended treatments (P1, P2, 70% P1+M, and 70% P2+M), Bradyrhizobium diazoefficiens and Roseateles depolymerans became the predominant species, exhibiting significant increases in relative abundance with higher P inputs. Notably, the relative abundance of all the three dominant species under the application of organic amendments was higher than that in corresponding inorganic P treatments. Furthermore, organic amendments increased the network nodes and connectivity links compared with corresponding mineral P treatments. Random forest analysis further identified B. icense as the strongest predictor of soil available P. The stability of phoD-harboring bacterial networks showed no significant difference across treatments. However, after the removal of dominant species, the network stability declined significantly in all treatments. Conclusion Organic amendments increase the relative abundance of dominant species within the phoD-harboring bacterial community across different P supply levels. They enhance the network complexity of phoD-harboring bacteria, thereby improving the network stability of these bacterial communities and ultimately influencing the availability of soil P.