A natural gene drive system confers reproductive isolation in rice.

Hybrid sterility restricts the utilization of superior heterosis of indica-japonica inter-subspecific hybrids. In this study, we report the identification of RHS12, a major locus controlling male gamete sterility in indica-japonica hybrid rice. We show that RHS12 consists of two genes (iORF3/DUYAO and iORF4/JIEYAO) that confer preferential transmission of the RHS12-i type male gamete into the progeny, thereby forming a natural gene drive. DUYAO encodes a mitochondrion-targeted protein that interacts with OsCOX11 to trigger cytotoxicity and cell death, whereas JIEYAO encodes a protein that reroutes DUYAO to the autophagosome for degradation via direct physical interaction, thereby detoxifying DUYAO. Evolutionary trajectory analysis reveals that this system likely formed de novo in the AA genome Oryza clade and contributed to reproductive isolation (RI) between different lineages of rice. Our combined results provide mechanistic insights into the genetic basis of RI as well as insights for strategic designs of hybrid rice breeding.

Caloric Restriction Reprograms the Single-Cell Transcriptional Landscape of Rattus Norvegicus Aging.

Aging causes a functional decline in tissues throughout the body that may be delayed by caloric restriction (CR). However, the cellular profiles and signatures of aging, as well as those ameliorated by CR, remain unclear. Here, we built comprehensive single-cell and single-nucleus transcriptomic atlases across various rat tissues undergoing aging and CR. CR attenuated aging-related changes in cell type composition, gene expression, and core transcriptional regulatory networks. Immune cells were increased during aging, and CR favorably reversed the aging-disturbed immune ecosystem. Computational prediction revealed that the abnormal cell-cell communication patterns observed during aging, including the excessive proinflammatory ligand-receptor interplay, were reversed by CR. Our work provides multi-tissue single-cell transcriptional landscapes associated with aging and CR in a mammal, enhances our understanding of the robustness of CR as a geroprotective intervention, and uncovers how metabolic intervention can act upon the immune system to modify the process of aging.

Single-Cell Transcriptomic Atlas of Primate Ovarian Aging.

Molecular mechanisms of ovarian aging and female age-related fertility decline remain unclear. We surveyed the single-cell transcriptomic landscape of ovaries from young and aged non-human primates (NHPs) and identified seven ovarian cell types with distinct gene-expression signatures, including oocyte and six types of ovarian somatic cells. In-depth dissection of gene-expression dynamics of oocytes revealed four subtypes at sequential and stepwise developmental stages. Further analysis of cell-type-specific aging-associated transcriptional changes uncovered the disturbance of antioxidant signaling specific to early-stage oocytes and granulosa cells, indicative of oxidative damage as a crucial factor in ovarian functional decline with age. Additionally, inactivated antioxidative pathways, increased reactive oxygen species, and apoptosis were observed in granulosa cells from aged women. This study provides a comprehensive understanding of the cell-type-specific mechanisms underlying primate ovarian aging at single-cell resolution, revealing new diagnostic biomarkers and potential therapeutic targets for age-related human ovarian disorders.

Deep learning-assisted LI-RADS grading and distinguishing hepatocellular carcinoma (HCC) from non-HCC based on multiphase CT: a two-center study.

OBJECTIVES: To develop a deep learning (DL) method that can determine the Liver Imaging Reporting and Data System (LI-RADS) grading of high-risk liver lesions and distinguish hepatocellular carcinoma (HCC) from non-HCC based on multiphase CT. METHODS: This retrospective study included 1049 patients with 1082 lesions from two independent hospitals that were pathologically confirmed as HCC or non-HCC. All patients underwent a four-phase CT imaging protocol. All lesions were graded (LR 4/5/M) by radiologists and divided into an internal (n = 886) and external cohort (n = 196) based on the examination date. In the internal cohort, Swin-Transformer based on different CT protocols were trained and tested for their ability to LI-RADS grading and distinguish HCC from non-HCC, and then validated in the external cohort. We further developed a combined model with the optimal protocol and clinical information for distinguishing HCC from non-HCC. RESULTS: In the test and external validation cohorts, the three-phase protocol without pre-contrast showed κ values of 0.6094 and 0.4845 for LI-RADS grading, and its accuracy was 0.8371 and 0.8061, while the accuracy of the radiologist was 0.8596 and 0.8622, respectively. The AUCs in distinguishing HCC from non-HCC were 0.865 and 0.715 in the test and external validation cohorts, while those of the combined model were 0.887 and 0.808. CONCLUSION: The Swin-Transformer based on three-phase CT protocol without pre-contrast could feasibly simplify LI-RADS grading and distinguish HCC from non-HCC. Furthermore, the DL model have the potential in accurately distinguishing HCC from non-HCC using imaging and highly characteristic clinical data as inputs. CLINICAL RELEVANCE STATEMENT: The application of deep learning model for multiphase CT has proven to improve the clinical applicability of the Liver Imaging Reporting and Data System and provide support to optimize the management of patients with liver diseases. KEY POINTS: • Deep learning (DL) simplifies LI-RADS grading and helps distinguish hepatocellular carcinoma (HCC) from non-HCC. • The Swin-Transformer based on the three-phase CT protocol without pre-contrast outperformed other CT protocols. • The Swin-Transformer provide help in distinguishing HCC from non-HCC by using CT and characteristic clinical information as inputs.

Pterygium combined with corneal perforation: a case report.

BACKGROUND: Pterygium is a common ocular surface disease. Pterygium combined with corneal perforation is rare. CASE PRESENTATION: A 28-year-old female patient visited our outpatient clinic due to sudden onset of blurred vision and increased tearing in her left eye. The visual acuity was 1.0 OD and intraocular pressure (IOP) of 19.5 mmHg for the right eye with no significant abnormalities found in the anterior and posterior segments. The visual acuity of her left eye was 0.06, and IOP was 6.2 mmHg. A triangular vascular membranous tissue was seen in her left eye below the nose growing into the cornea and the pupil area was not touched. Slit-lamp examination revealed a tiny round corneal perforation in 8 o'clock position of the lesion area. Hospital diagnosis was given as pterygium combined with corneal perforation. The patient was treated with levofloxacin eye drops and autologous serum-based eye drops. CONCLUSIONS: We report a rare case of pterygium combined with corneal perforation. Perforation is a very rare complication of pterygium. This patient received proper treatment and good result was seen. This article aimed to improve clinicians' understanding of pterygium.

Clinicopathologic and mutational profiles of primary breast diffuse large B cell lymphoma in a male patient: case report and literature review.

INTRODUCTION: Primary breast lymphoma (PBL) is rare, and most cases occur in female patients, with few reported cases in male patients. The clinical presentation is similar to that of breast cancer, but the condition needs to be well understood, as treatment options and clinical course vary. Hence, we provide a relatively rare case of primary breast diffuse large B cell lymphoma (PB-DLBCL) in a male, including its complete clinicopathological features, radiological findings, genomic mutational profiles, and clinical course. CASE PRESENTATION: A 45-year-old male presented with a lump in his right breast for 1 week and was pathologically diagnosed with breast malignancy after a breast puncture biopsy at the local hospital. He came to our hospital for further treatment and underwent breast ultrasound and systemic positron emission tomography/computed tomography (PET/CT) imaging, followed by right mastectomy and sentinel lymph node biopsy. Histomorphology showed diffuse hyperplasia of tumor cells with clear boundaries and surrounding normal breast ducts. The adhesion of tumor cells was poor with obvious atypia. Immunohistochemical results showed that the tumor cells were positive for CD20, Bcl6, and MUM-1 but negative for CK (AE1/AE3), ER, PR, CD3, and CD10. Forty percent of the tumor cells were positive for c-Myc, and 80% of tumor cells were positive for Bcl2. The Ki-67 proliferation index was up to 80%. The tumor cells were negative for MYC and BCL2 rearrangements but positive for BCL6 rearrangement by fluorescent in situ hybridization. No abnormality was found in the pathological examination of bone marrow aspiration. Therefore, the male was diagnosed with PB-DLBCL, nongerminal center (non-GCB) phenotype, dual-expression type. The sample were sequenced by a target panel of 121 genes related to lymphoma. Next-generation sequencing revealed six tumor-specific mutated genes (IGH/BCL6, TNFAIP3, PRDM1, CREBBP, DTX1, and FOXO1). The patient was given six cycles of orelabrutinib plus R-CHOP chemotherapy and two cycles of intrathecal injection of cytarabine. The last follow-up was on April 13, 2023 (17 months). No recurrence or metastasis was found in laboratory and imaging examinations. CONCLUSION: We reported a relatively rare PB-DLBCL in a male, non-GBC phenotype, dual-expression type. It is worth mentioning that this case had IgH/BCL6 fusion, nonsense mutations in TNFAIP3, frameshift mutations in PRDM1, and missense mutations in CREBBP, DTX1, and FOXO1. To the best of our knowledge, this case is the first report of genomic mutational profiles of PB-DLBCL in males.

Sanggenon C Suppresses Tumorigenesis of Gastric Cancer by Blocking ERK-Drp1-Mediated Mitochondrial Fission.

Sanggenon C is a flavonoid extracted from the root bark of white mulberry, which is a traditional Chinese medicine with anti-inflammatory, antioxidative, and antitumor pharmacological effects. In this study, sanggenon C was found to inhibit human gastric cancer (GC) cell proliferation and colony formation, induce GC cell cycle arrest in the G0-G1 phase, and promote GC cell apoptosis. Moreover, sanggenon C was found to decrease the level of mitochondrial membrane potential in GC cells and inhibit mitochondrial fission. Mechanistically, RNA sequencing, bioinformatics analysis, and a series of functional analyses confirmed that sanggenon C inhibited mitochondrial fission to induce apoptosis by blocking the extracellular regulated protein kinases (ERK) signaling pathway, and constitutive activation of ERK significantly abrogated these effects. Finally, sanggenon C was found to suppress the growth of tumor xenografts in nude mice without obvious side effects to the vital organs of animals. This study reveals that sanggenon C could be a novel therapeutic strategy for GC treatment.

Roles of Resolvins in Chronic Inflammatory Response.

An inflammatory response is beneficial to the organism, while an excessive uncontrolled inflammatory response can lead to the nonspecific killing of tissue cells. Therefore, promoting the resolution of inflammation is an important mechanism for protecting an organism suffering from chronic inflammatory diseases. Resolvins are a series of endogenous lipid mediums and have the functions of inhibiting a leukocyte infiltration, increasing macrophagocyte phagocytosis, regulating cytokines, and alleviating inflammatory pain. By promoting the inflammation resolution, resolvins play an irreplaceable role throughout the pathological process of some joint inflammation, neuroinflammation, vascular inflammation, and tissue inflammation. Although a large number of experiments have been conducted to study different subtypes of resolvins in different directions, the differences in the action targets between the different subtypes are rarely compared. Hence, this paper reviews the generation of resolvins, the characteristics of resolvins, and the actions of resolvins under a chronic inflammatory response and clinical translation of resolvins for the treatment of chronic inflammatory diseases.

Synthesis of nitrogenase by Paenibacillus sabinae T27 in presence of high levels of ammonia during anaerobic fermentation.

Biological nitrogen fixation is usually inhibited by fixed nitrogen. Paenibacillus sabinae T27, a Gram-positive, spore-forming diazotroph, possesses high nitrogenase activity and has 3 copies of nifH (nifH, nifH2, nifH3), a copy of nifDK, and multiple nifHDK-like genes. In this study, we found that P. sabinae T27 showed nitrogenase activities not only in low (0-3 mM) concentrations of NH4+ but also in high (30-300 mM) concentrations of NH4+, no matter whether this bacterium was grown in a flask or in a fermenter on scale cultivation. qRT-PCR and western blotting analyses supported that Fe protein and MoFe protein were synthesized under both low (0-3 mM) and high (30-300 mM) concentrations of NH4+. Liquid chromatography-mass spectrometry (LC-MS) analysis revealed that MoFe protein was encoded by nifDK and Fe protein was encoded by both nifH and nifH2. The cross-reaction suggested the purified Fe and MoFe components from P. sabinae T27 grown in both nitrogen-limited and nitrogen-excess conditions were active. This is the first time to report that diazotrophs show nitrogenase activity in presence of high (30-300 mM) concentrations of NH4+. Our study will provide a clue for studying the mechanisms of nitrogen fixation in presence of the high concentration of NH4+. KEY POINTS: • P. sabinae T27 can synthesize active nitrogenase in presence of high levels of ammonia. •Fe and MoFe proteins of nitrogenase purified in absence of ammonia are the same as those purified from the high concentration of ammonia. • Fe protein is encoded by nifH and nifH2, and MoFe protein is encoded by nifDK.

Effect of Andrographolide and Its Analogs on Bacterial Infection: A Review.

Bacterial infections remain the leading cause of death in children, the elderly, and immunocompromised patients. Andrographolide (AG), the main active component of the herb Andrographis paniculata, has been used for many years for anti-inflammatory and antibacterial infections. AG has an antibacterial effect on a wide variety of bacteria, which is reflected in the inhibition of bacterial pathogenic factors and the regulation of immunity to downregulate infectious inflammation caused by bacteria. In the current climate of frequently occurring antibiotic resistance, AG might be considered a promising lead for new antibacterial drug development. This review outlines the therapeutic potential of AG and its analogs in combating various bacterial infections, focusing on the mechanisms of action.

Study on the Release Potential of BPA and Steroid Estrogens in the Sediments of Erhai Lake, a Typical Plateau Lake of China.

On-site sampling analysis and laboratory-scale experiments were conducted to study the pollution status and release potential of EDCs in Erhai Lake. We found that nitrogen and phosphorus pollution in Erhai Lake sediment were both at a high level, as well as EDCs pollution. The concentrations of BPA, E2α, E1, E2ß, EE2, and E3 were 36.84 ng/g(DW), 13.04 ng/g(DW), 128.97 ng/g(DW), 52.57 ng/g(DW), 18.48 ng/g(DW) and 5.36 ng/g(DW), respectively. The concentrations of E2α, E1, E2ß and EE2 in the bottom water were higher than the surface water due to the impact of sediment release. The results of the 20 days release test indicated that BPA release from the sediment had a greater correlation with the original concentration and the particle size of sediment, while the steroid EDCs had no obvious correlation with these two factors, probably due to the difference in hydrophobicity between them. Under hydraulic disturbance and aerobic conditions, the release process of EDCs was accompanied by a large amount of microbial degradation, and degradation amount > released amount. BPA was released quickly, 9.56% was released in 20 days, but only 3.37% of steroid EDCs released. In comparison, the release process of steroids was longer and posed a greater threat to aquatic ecology.

LncRNA HOTAIR-mediated Wnt/ß-catenin network modeling to predict and validate therapeutic targets for cartilage damage.

BACKGROUND: Cartilage damage is a crucial feature involved in several pathological conditions characterized by joint disorders, such as osteoarthritis and rheumatoid arthritis. Accumulated evidences showed that Wnt/ß-catenin pathway plays a role in the pathogenesis of cartilage damage. In addition, it is experimentally documented that lncRNA (long non-coding RNA) HOTAIR plays a key role in the regulation of Wnt/ß-catenin pathway based on directly decreased WIF-1 expression. Further, it is reported that Wnt/ß-catenin pathway is a potent pathway to regulate the expression of MMP-13, which is responsible for degradation of collagen type II in articular cartilage. It is increasingly recognized that systems modeling approach provides an opportunity to understand the complex relationships and direct quantitative analysis of dynamic network in various diseases. RESULTS: A dynamic network of lncRNA HOTAIR-mediated Wnt/ß-catenin pathway regulating MMP-13 is developed to investigate the dynamic mechanism of the network involved in the pathogenesis of cartilage damage. Based on the network modeling, the potential therapeutic intervention point Axin is predicted and confirmed by the experimental validation. CONCLUSIONS: Our study provides a promising strategy for revealing potential dynamic mechanism and assessing potential targets which contribute to the prevention of the pathological conditions related to cartilage damage.

Identification of Constituents Affecting the Secretion of Pro-Inflammatory Cytokines in LPS-Induced U937 Cells by UHPLC-HRMS-Based Metabolic Profiling of the Traditional Chinese Medicine Formulation Huangqi Jianzhong Tang.

Within non-communicable diseases, chronic inflammatory conditions represent one of the biggest challenges for modern medicine. Traditional Chinese Medicine (TCM) has been practiced over centuries and has accumulated tremendous empirical knowledge on the treatment of such diseases. Huangqi Jianzhong Tang (HQJZT) is a famous TCM herbal formula composed of Radix Astragali, Ramulus Cinnamomi, Radix et Rhizoma Glycyrrhizae Praeparata cum Melle, Radix Paeoniae Alba, Rhizoma Zingiberis Recens, Fructus Jujubae and Saccharum Granorum (maltose), which has been used for the treatment of various chronic inflammatory gastrointestinal diseases. However, there is insufficient knowledge about its active constituents and the mechanisms responsible for its effects. The present study aimed at identifying constituents contributing to the bioactivity of HQJZT by combining in vitro cytokine production assays and LC-MS metabolomics techniques. From the HQJZT decoction as well as from its single herbal components, extracts of different polarities were prepared. Phytochemical composition of the extracts was analyzed by means of UPLC-QTOF-MS/MS. The inhibitory effects of the extracts on TNF-α, IL-1ß and IFN-γ production were studied in U937 cells. Phytochemical and pharmacological bioactivity data were correlated by orthogonal projection to latent structures discriminant analysis (OPLS-DA) in order to identify those HQJZT constituents which may be relevant for the observed pharmacological activities. The investigations resulted in the identification of 16 HQJZT constituents, which are likely to contribute to the activities observed in U937 cells. Seven of them, namely calycosin, formononetin, astragaloside I, liquiritigenin, 18ß-glycyrrhetinic acid, paeoniflorin and albiflorin were unambiguously identified. The predicted results were verified by testing these compounds in the same pharmacological assays as for the extracts. In conclusion, the anti-inflammatory activity of HQJZT could be substantiated by in vitro pharmacological screening, and the predicted activities of the OPLS-DA hits could be partially verified. Moreover, the benefits and limitations of MVDA for prediction pharmacologically active compounds contributing to the activity of a TCM mixture could be detected.

La2O3-modified MCM-41 for efficient phosphate removal synthesized using natural diatomite as precursor.

In this study, an ordered mesoporous silica modified with lanthanum oxide was synthesized using diatomite as silica source and applied for adsorption of phosphate from aqueous solution. By taking cost-effectiveness for practical application into consideration, the adsorbent with a theoretical La/SiO2 molar ratio of 0.2 (La0.2M41) possessed a promising performance. In the batch adsorption tests, the adsorbents with La2O3 loading possessed markedly enhanced adsorption capacities. Phosphate uptake by La0.2M41 was pH-dependent with the highest sorption capacities observed over a pH range of 3.0-6.0. Coexistent anions displayed an adverse effect on phosphate adsorption following the order of CO3 2-  > F-  > NO3 - > Cl- > SO4 2-. In the kinetic study, phosphate adsorption onto La0.2M41 followed the pseudo-second-order equation better than the pseudo-first-order, suggesting chemisorption. The Langmuir isothermal model well described the adsorption isotherm data, showing a maximum adsorption capacity for phosphate of up to 263.16 mg/g at 298 K. In a real treated wastewater effluent with phosphate concentration of 2.5 mg P/L, La0.2M41 efficiently reduced the phosphate concentration to 28 µg P/L.

[Bioinformatics analysis on immune response pathways of triptolide in treating AIDS].

The aim of this paper was to study the influence of triptolide in the immune response pathways of acquired immune deficiency syndrome( AIDS). Target proteins of triptolide and related genes of AIDS were searched in PubChem and Gene databases on line. Molecular networks and canonical pathways comparison analyses were performed by bioinformatics software( IPA). There were 15 targets proteins of triptolide and 258 related genes of AIDS. Close biological relationships of molecules of triptolide and AIDS were established by networks analysis. There were 21 common immune response pathways of triptolide and AIDS,including neuroinflammation signaling pathway,Th1 and Th2 activation pathway and role of pattern recognition receptors in recognition of bacteria and viruses. Triptolide stimulated immune response pathways by the main molecules of IFNγ,JAK2,NOD1,PTGS2,RORC. IFNγ is the focus nodes of triptolide and AIDS,and regulates genes of AIDS directly or indirectly. Triptolide may against AIDS by regulating molecules IFNγ in immune response pathways.

The Effect of Triptolide in Rheumatoid Arthritis: From Basic Research towards Clinical Translation.

Triptolide (TP), a major extract of the herb Tripterygium wilfordii Hook F (TWHF), has been shown to exert potent pharmacological effects, especially an immunosuppressive effect in the treatment of rheumatoid arthritis (RA). However, its multiorgan toxicity prevents it from being widely used in clinical practice. Recently, several attempts are being performed to reduce TP toxicity. In this review, recent progress in the use of TP for RA, including its pharmacological effects and toxicity, is summarized. Meanwhile, strategies relying on chemical structural modifications, innovative delivery systems, and drug combinations to alleviate the disadvantages of TP are also reviewed. Furthermore, we also discuss the challenges and perspectives in their clinical translation.

Cyclin-Dependent Kinase 5 Regulates Dendritic Spine Formation and Maintenance of Cortical Neuron in the Mouse Brain.

Cyclin-dependent kinase 5 (Cdk5) activity is dependent on its association with 1 of 2 neuron-specific activators, p35 or p39. Cdk5 and its activators play an important role in brain development as well as higher functions like synaptic plasticity, learning, and memory. Reduction in p35 was reported in postmortem schizophrenia brain, in which reduced dendritic spine density was observed. Previous in vitro experiments have shown that Cdk5 is involved in dendritic spine formation, although in vivo evidence is limited. We examined dendritic spine formation in inducible-p35 conditional knockout (p35 cKO); p39 KO mice. When we deleted the p35 gene either during early postnatal days or at adult stage, we observed reduced spine densities of layer V neurons in the cerebral cortex and CA1 pyramidal neurons in the hippocampus. We further generated CA1-specific p35 conditional knockout (CA1-p35 cKO) mice and also CA1-p35 cKO; p39 KO mice in which have specific deletion of p35 in the CA1 region of hippocampus. We found a greater reduction in spine densities in CA1 pyramidal neurons in CA1-p35 cKO; p39 KO mice than in CA1-p35 cKO mice. These results indicate that dendritic spine formation and neuronal maintenance are dependent on Cdk5 activity.

Combination therapeutics in complex diseases.

The biological redundancies in molecular networks of complex diseases limit the efficacy of many single drug therapies. Combination therapeutics, as a common therapeutic method, involve pharmacological intervention using several drugs that interact with multiple targets in the molecular networks of diseases and may achieve better efficacy and/or less toxicity than monotherapy in practice. The development of combination therapeutics is complicated by several critical issues, including identifying multiple targets, targeting strategies and the drug combination. This review summarizes the current achievements in combination therapeutics, with a particular emphasis on the efforts to develop combination therapeutics for complex diseases.

Genome-wide transcriptome profiling of nitrogen fixation in Paenibacillus sp. WLY78.

BACKGROUND: Diazotrophic (nitrogen-fixing) Gram-positive and endospore-formed Paenibacillus spp. have potential uses as a bacterial fertilizer in agriculture. The transcriptional analysis of nitrogen fixation in Paenibacillus is lacking, although regulation mechanisms of nitrogen fixation have been well studied in Gram-negative diazotrophs. RESULTS: Here we report a global transcriptional profiling analysis of nitrogen fixation in Paenibacillus sp. WLY78 cultured under N2-fixing condition (without O2 and NH4(+)) and non-N2-fixing condition (air and 100 mM NH4(+)). The nif (nitrogen fixation) gene operon composed of 9 genes (nifBHDKENXhesAnifV) in this bacterium was significantly up-regulated in N2-fixing condition compared to non-N2-fixing condition, indicating that nif gene transcription is strictly controlled by NH4(+) and O2. qRT-PCR confirmed that these nif genes were differently expressed. Non-nif genes specifically required in nitrogen fixation, such as mod, feoAB and cys encoding transporters of Mo, Fe and S atoms, were coordinately transcribed with nif genes in N2-fixing condition. The transcript abundance of suf operon specific for synthesis of Fe-S cluster was up-regulated in N2-fixing condition, suggesting that Sul system, which takes place of nifS and nifU, plays important role in the synthesis of nitrogenase. We discover potential specific electron transporters which might provide electron from Fe protein to MoFe protein of nitrogenase. The glnR whose predicted protein might mediate nif transcription regulation by NH4(+) is significantly up-regulated in N2-fixing condition. The transcription levels of nitrogen metabolism and anaerobic respiration were also analyzed. CONCLUSIONS: The nif gene operon (nifBHDKENXhesAnifV) in Paenibacillus sp. WLY78 is significantly up-regulated in N2-fixing condition compared to non-N2-fixing condition. Non-nif genes specifically required in nitrogen fixation were also significantly up-regulated in N2-fixing condition. Fur and Fnr which are involved in anaerobic regulation and GlnR which might mediate nif gene transcription regulation by NH4(+) were significantly up-regulated in N2-fixing condition. This study provides valuable insights into nitrogen fixation process and regulation in Gram-positive firmicutes.