Search Results (18)

Nitrogen is an important nutrient required for plant growth and development, but most of the time plants face nitrogen deficiency, all it is important to study the mechanism of low nitrogen tolerance in plants. This study addresses this gap by investigating the role of the StDWF1 gene through the generation and analysis of transgenic potato lines overexpressing StDWF1 (OE1, OE2, OE3). Exogenous BL treatment showed that the StDWF1 gene responded to oleuropein lactone. Phenotypic assessments under normal nitrogen (NN) and low nitrogen (LN) conditions demonstrated that OE2 consistently outperformed WT, showing a 43% increase in root vitality and a 23% retention of chlorophyll under LN. Additionally, OE2 transgenics accumulated significantly higher levels of nitrate nitrogen (64.1% increase) and ammonium nitrogen (53% increase) compared to WT. Enzymatic assays further confirmed elevated activities of glutamine synthetase and nitrate reductase in both OE1 and OE2 lines. Hormone analyses showed that BL content of StDWF1 overexpression lines was significantly increased under LN conditions, higher Oleandrin lactone (BL) content of OE2 improved plant stress tolerance, and WT was more affected by low nitrogen stress than OE2, resulting in higher levels of stress hormones than OE2. Temporal gene expression analysis showed significant upregulation of key nitrogen metabolism-related genes (NR, NiR, AT, NRT2.1) in OE2, with StDWF1 expression reaching 79% higher than WT at 3 h. Protein–protein interaction assays, including yeast two-hybrid and BiLC assays, verified the interaction between StDWF1 and StGRP1, suggesting the existence of a functional network to enhance low-nitrogen tolerance in potato plants. In conclusion, these findings suggest that overexpression of StDWF1 significantly enhances low-nitrogen tolerance in transgenic potato lines, providing a promising strategy for improving crop performance under nitrogen-limited conditions. Full article

Managing nitrogen and phosphorus pollution in high-salinity wastewater is a critical challenge for sustainable aquaculture and environmental protection. In this study, a novel salt-tolerant strain, Pseudomonas sediminis D4, was isolated from a brackish water environment. This strain exhibited simultaneous heterotrophic nitrification–aerobic denitrification and phosphorus removal capabilities. Biosafety assays demonstrated that the strain was antibiotic-sensitive and safe for aquatic environments. The optimal conditions for nitrogen and phosphate removal of strain D4 were carbon/nitrogen (C/N) ratio 10, phosphorus/nitrogen (P/N) ratio 0.2, pH 7, and temperature 30 °C while using sodium succinate as the carbon source. Under these conditions, strain D4 achieved removal efficiencies of 97.36% for ammonia (NH₄⁺-N), 100.00% for nitrate (NO₃⁻-N), and 98.02% for nitrite (NO₂⁻-N), along with 94.69%, 89.56%, and 97.40% removal of PO₄³⁻P, respectively. The strain exhibited strong salinity tolerance, functioning effectively within a range of 0% to 5% (w/v), and maintaining high nitrogen and phosphorus removal efficiency at a salinity of 3%. Enzyme activity assays verified the existence of key enzymes, such as ammonia nitrogen oxidase, nitrate oxidoreductase, nitrate reductase, nitrite reductase, polyphosphate kinase, and exopolyphosphatase, which are essential for the heterotrophic nitrification-aerobic denitrification and phosphorus removal capabilities of D4. These findings highlight the potential of Pseudomonas sediminis D4 for the biological treatment of high-salinity wastewater. Full article

Light is a key factor influencing the growth and quality of crops in plant factories. To explore the optimal light quality for pakchoi production, five light formulations were applied to ‘Youguan NO.3’ pakchoi: white LEDs (W; CK); white/red = 4:1 (WR); white/blue = 4:1 (WB); white/red/blue = 3:1:1 (WRB); and white/green = 4:1 (WG), all with a light intensity of 250 ± 10 µmol·m⁻²·s⁻¹. The results showed significant variations in growth indices, nutritional quality, enzyme activity, and other parameters under different light qualities. The best growth results were observed under the WRB treatment. Chloroplasts under WRB treatment appeared well-developed, with clear grana lamellae. The thylakoids in the chloroplast grana of the WRB plants were densely stacked, and a large number of starch grains were detected. The contents of total sugar, soluble sugar, soluble protein, and protein nitrogen were significantly higher under the WB, WRB, and WR treatments compared to the CK treatment, along with a significant reduction in nitrate content. Among all the treatments, WRB treatment resulted in the highest levels of total sugar, starch, free amino acids, soluble protein, total nitrogen, protein nitrogen, and ascorbic acid (AsA). Enzyme activity assays revealed that the activities of sucrose phosphate synthetase (SPS), nitrate reductase (NR), glutamine synthetase (GS), glutamate synthetase (GOGAT), and glutamate dehydrogenase (GDH) were highest under WRB treatment. Therefore, supplemental red-blue mixed light can effectively improve the growth and nutritional properties of pakchoi grown under white light. This supplementary lighting strategy provides a new way to enhance the nutritional value of leafy vegetables in plant factories. Full article

The transmembrane nitrate reductase (Nar) is the first enzyme in the dissimilatory alternate anaerobic nitrate respiratory chain in denitrifying bacteria. To date, there has been no real-time method to determine its specific activity embedded in its native membrane; here, we describe such a new method, which is useful with the inside-out membranes of Paracoccus denitrificans and other denitrifying bacteria. This new method takes advantage of the native coupling of the endogenous NADH dehydrogenase or Complex I with the reduction of nitrate by Nar through the quinone pool of the inner membranes of P. denitrificans. This is achieved under previously reached anaerobic conditions. Inner controls confirming the specific Nar activity determined by this new method were made by the total inhibition of the Nar enzyme by sodium azide and cyanide, well-known Nar inhibitors. The estimation of the Michaelis–Menten affinity of Nar for NO₃⁻ using this so-called Nar-JJ assay gave a K_m of 70.4 μM, similar to previously determined values. This new Nar-JJ assay is a suitable, low-cost, and reproducible method to determine in real-time the endogenous Nar activity not only in P. denitrificans, but in other denitrifying bacteria such as Brucella canis, and potentially in other entero-pathogenic bacteria. Full article

The release rates of different nitrogen (N) fertilizers and their matching with plant demand determine crop yields. A field experiment was conducted to investigate the effect of using no fertilizer (N0), regular urea applied at rates of 180 kg ha⁻¹ (N180) and 240 kg ha⁻¹ (N240), controlled-release urea applied at a rate of 180 kg ha⁻¹ (H180), and polyaspartic acid urea (PASP) on maize seed yield, soil microbial community diversity, and leaf N-converting enzymes. XianYu 688 was selected as the test maize variety. All cobs in the sample plots were collected per unit area to estimate maize yield. The enzyme-linked immunosorbent assay (ELISA) was used to determine leaf N-converting enzyme activities. Soil DNA was extracted using the Power Max Soil DNA Isolation Kit and subsequently sequenced using the Illumina HiSeq platform (PE 2500) to determine the microbial diversity and communities. The results showed that the highest seed yields were obtained under N240 and PASP180 treatments. The N-partial factor productivity of the PASP180 fertilizer was significantly higher than that of the other treatments. PASP treatment significantly increased maize seed yield due to the potential of storing more N in the ear leaves. Additionally, partial N productivity showed a significant positive correlation with the soil microbial Shannon, Chao1, and ACE indices, indicating that increased soil microbial diversity promoted N efficiency in maize. Further analysis revealed that PASP treatment increased seed yield by promoting leaf N-converting enzyme activity and soil microbial diversity. The results revealed that nitrate reductase (NR), glutamate synthase (GOGAT), and glutaminase (GLNS) enzyme activities in maize leaves were higher under the PASP treatment than under other fertilizer treatments. The PASP treatment significantly enhanced soil microbial diversity at different maize stages. Our study revealed the effects of using different N fertilizers on seed yield by examining their impact on soil microbial diversity and leaf N-converting enzyme activity. This study provides essential insights into maize production and soil fertility maintenance in the North China Plain. Full article

The impact of the exogenous supplementation of hydrogen sulfide (20 and 50 µM HS) on growth, enzyme activity, chlorophyll pigments, and tolerance mechanisms was studied in salinity-stressed (100 mM NaCl) wheat. Salinity significantly reduced height, fresh and dry weight, chlorophyll, and carotenoids. However, the supplementation of HS (at both concentrations) increased these attributes and also mitigated the decline to a considerable extent. The exogenous supplementation of HS reduced the accumulation of hydrogen peroxide (H₂O₂) and methylglyoxal (MG), thereby reducing lipid peroxidation and increasing the membrane stability index (MSI). Salinity stress increased H₂O₂, MG, and lipid peroxidation while reducing the MSI. The activity of nitrate reductase was reduced due to NaCl. However, the supplementation of HS alleviated the decline with obvious effects being seen due to 50 µM HS. The activity of antioxidant enzymes (superoxide dismutase, catalase, ascorbate peroxidase, and glutathione reductase) was assayed and the content of reduced glutathione (GSH) increased due to salt stress and the supplementation of HS further enhanced their activity. A decline in ascorbic acid due to salinity stress was alleviated due to HS treatment. HS treatment increased the endogenous concentration of HS and nitric oxide (NO) under normal conditions. However, under salinity stress, HS supplementation resulted in a reduction in HS and NO as compared to NaCl-treated plants. In addition, proline and glycine betaine increased due to HS supplementation. HS treatment reduced sodium levels, while the increase in potassium justified the beneficial role of applied HS in improving salt tolerance in wheat. Full article

Ustilago maydis is an important model to study intermediary and mitochondrial metabolism, among other processes. U. maydis can grow, at very different rates, on glucose, lactate, glycerol, and ethanol as carbon sources. Under nitrogen starvation and glucose as the only carbon source, this fungus synthesizes and accumulates neutral lipids in the form of lipid droplets (LD). In this work, we studied the accumulation of triacylglycerols in cells cultured in a medium containing acetate, a direct precursor of the acetyl-CoA required for the synthesis of fatty acids. The metabolic adaptation of cells to acetate was studied by measuring the activities of key enzymes involved in glycolysis, gluconeogenesis, and the pentose phosphate pathways. Since growth on acetate induces oxidative stress, the activities of some antioxidant enzymes were also assayed. The results show that cells grown in acetate plus nitrate did not increase the amount of LD, but increased the activities of glutathione reductase, glutathione peroxidase, catalase, and superoxide dismutase, suggesting a higher production of reactive oxygen species in cells growing in acetate. The phosphofructokinase-1 (PFK1) was the enzyme with the lowest specific activity in the glycolytic pathway, suggesting that PFK1 controls the flux of glycolysis. As expected, the activity of the phosphoenolpyruvate carboxykinase, a gluconeogenic enzyme, was present only in the acetate condition. In summary, in the presence of acetate as the only carbon source, U. maydis synthesized fatty acids, which were directed into the production of phospholipids and neutral lipids for biomass generation, but without any excessive accumulation of LD. Full article

Fungal AreA is a key nitrogen metabolism transcription factor in nitrogen metabolism repression (NMR). Studies have shown that there are different ways to regulate AreA activity in yeast and filamentous ascomycetes, but in Basidiomycota, how AreA is regulated is unknown. Here, a gene from Ganoderma lucidum with similarity to nmrA of filamentous ascomycetes was identified. The NmrA interacted with the C-terminal of AreA according to yeast two-hybrid assay. In order to determine the effect of NmrA on the AreA, 2 nmrA silenced strains of G. lucidum, with silencing efficiencies of 76% and 78%, were constructed using an RNA interference method. Silencing nmrA resulted in a decreased content of AreA. The content of AreA in nmrAi-3 and nmrAi-48 decreased by approximately 68% and 60%, respectively, compared with that in the WT in the ammonium condition. Under the nitrate culture condition, silencing nmrA resulted in a 40% decrease compared with the WT. Silencing nmrA also reduced the stability of the AreA protein. When the mycelia were treated with cycloheximide for 6 h, the AreA protein was almost undetectable in the nmrA silenced strains, while there was still approximately 80% of the AreA protein in the WT strains. In addition, under the nitrate culture, the content of AreA protein in the nuclei of the WT strains was significantly increased compared with that under the ammonium condition. However, when nmrA was silenced, the content of the AreA protein in the nuclei did not change compared with the WT. Compared with the WT, the expression of the glutamine synthetase gene in nmrAi-3 and nmrAi-48 strains increased by approximately 94% and 88%, respectively, under the ammonium condition, while the expression level of the nitrate reductase gene in nmrAi-3 and nmrAi-48 strains increased by approximately 100% and 93%, respectively, under the nitrate condition. Finally, silencing nmrA inhibited mycelial growth and increased ganoderic acid biosynthesis. Our findings are the first to reveal that a gene from G. lucidum with similarity to the nmrA of filamentous ascomycetes contributes to regulating AreA, which provides new insight into how AreA is regulated in Basidiomycota. Full article

Encephalartos spp. establish symbioses with nitrogen (N)-fixing bacteria that contribute to soil nutrition and improve plant growth. Despite the Encephalartos mutualistic symbioses with N-fixing bacteria, the identity of other bacteria and their contribution to soil fertility and ecosystem functioning is not well understood. Due to Encephalartos spp. being threatened in the wild, this limited information presents a challenge in developing comprehensive conservation and management strategies for these cycad species. Therefore, this study identified the nutrient-cycling bacteria in Encephalartos natalensis coralloid roots, rhizosphere, and non-rhizosphere soils. Additionally, the soil characteristics and soil enzyme activities of the rhizosphere and non-rhizosphere soils were assayed. The coralloid roots, rhizosphere, and non-rhizosphere soils of E. natalensis were collected from a population of >500 E. natalensis in a disturbed savanna woodland at Edendale in KwaZulu-Natal (South Africa) for nutrient analysis, bacterial identification, and enzyme activity assays. Nutrient-cycling bacteria such as Lysinibacillus xylanilyticus; Paraburkholderia sabiae, and Novosphingobium barchaimii were identified in the coralloid roots, rhizosphere, and non-rhizosphere soils of E. natalensis. Phosphorus (P) cycling (alkaline and acid phosphatase) and N cycling (β-(D)-Glucosaminidase and nitrate reductase) enzyme activities showed a positive correlation with soil extractable P and total N concentrations in the rhizosphere and non-rhizosphere soils of E. natalensis. The positive correlation between soil enzymes and soil nutrients demonstrates that the identified nutrient-cycling bacteria in E. natalensis coralloid roots, rhizosphere, and non-rhizosphere soils and associated enzymes assayed may contribute to soil nutrient bioavailability of E. natalensis plants growing in acidic and nutrient-poor savanna woodland ecosystems. Full article

Nitric oxide (NO) acts as a key signaling molecule in higher plants, regulating many physiological processes. Several photosynthetic algae from different lineages are also known to produce NO. However, it remains unclear whether this messenger is produced by non-photosynthetic algae. Among these organisms, the colorless alga Polytomella parva is a special case, as it has lost not only its plastid genome, but also nitrate reductase and nitrite reductase. Up to now, the question of whether NO synthesis occurs in the absence of functional nitrate reductase (NR) and the assimilation of nitrates/nitrites in P. parva has not been elucidated. Using spectrofluorometric assays and confocal microscopy with NO-sensitive fluorescence dye, we demonstrate L-arginine-dependent NO synthesis by P. parva cells. Based on a pharmacological approach, we propose the existence of arginine-dependent NO synthase-like activity in this non-photosynthetic alga. GC-MS analysis provides primary evidence that P. parva synthesizes putrescine, which is not an NO source in this alga. Moreover, the generated NO causes the S-nitrosation of protein cysteine thiol groups. Together, our data argue for NR-independent NO synthesis and its active role in S-nitrosation as an essential post-translational modification in P. parva. Full article

Salt stress has the most severe impact on plant growth and development, including seed germination. However, little is known about the mechanism of NR (nitrate reductase)-associated nitric oxide (NO) regulates salt tolerance during seed germination in rice. Herein, we shown that inhibition of seed germination by salt stress was significantly impaired by sodium nitroferricyanide (SNP), a NO donor. Then a triple mutant, nr1/nr2/nr3, was generated. Results shown that germination of triple mutants were delayed and were much more sensitive to salt stress than WT plant, which can be rescued by application of SNP. qPCR analysis revealed that expressions of abscisic acid (ABA) catabolism gene, OsABA8ox1, was suppressed in triple mutants under salt stress, resulting in an elevated ABA content. Similar to SNP, application of nitrate also rescued seed germination under salt stress, which, however, was blocked in the triple mutants. Further study revealed that a nitrate responsive transcript factor, OsNLP2, was induced by salt stress, which thus up-regulates the expression of OsNRs and NR activity, resulting in promoted salt tolerance during seed germination. In addition, nitrate-mediated salt tolerance was impaired in mutant of aba8ox1, a target gene for NLP2. Transient trans-activation assays further revealed NLP2 can significantly activate the expression of OsABA8ox1 and OsNR1, suggesting that NLP2 activates expression of ABA catabolism gene directly or indirectly via NR-associated NO. Taken together, our results demonstrate that NLP2-NR associated NO was involved in salt response by increasing ABA catabolism during seed germination and highlight the importance of NO for stress tolerance of plants. Full article

α-Casein (α-CN) is considered the main allergen in bovine milk. Lactic acid bacteria (LAB) fermentation can hydrolyze milk protein and therefore reduce the antigenicity. In this paper, a LAB reducing the antigenicity of casein, identified as LactiplantibacillusPlantarum 7-2 (L. plantarum 7-2), was primarily identified by screening for protein hydrolysis ability using a method involving the determination of released free amino acid with further selection for the ideal antigenicity-reducing capability by enzyme-linked immunosorbent assay (ELISA). In order to verify the capability of L. plantarum 7-2 in inhibiting antigenicity, the standard milk proteins α-LA, β-LG, α-CN, β-CN and κ-CN were cultured with L. plantarum 7-2 for 18 h; The results of SDS-PAGE show that all the bands corresponding to the full length tested proteins became unclear or completely disappeared indicating that these proteins were hydrolyzed by L. plantarum 7-2. Correspondingly, the antigenicities of α-CN and β-LG were significantly reduced. L. plantarum 7-2 demonstrated negative hemolysis and nitrate reductase capabilities and was sensitive to the commonly used antibiotics ampicillin clindamycin tetracycline chloramphenicol, and erythromycin, demonstrating that L. plantarum 7-2 could be used in dairy product fermentation to reduce the antigenicity of milk protein. Full article

Bacterial stress responses are closely associated with the survival and colonization of anaerobes in the human gut. Mesosutterella multiformis JCM 32464^T is a novel member of the family Sutterellaceae, an asaccharolytic bacterium. We previously demonstrated energy generation via heme biosynthesis, which is coupled with nitrate reductase. Here, physiological and morphological changes in M. multiformis induced by exposure to nitrate were investigated. The ability of M. multiformis to reduce nitrate was determined using a colorimetric assay. A unique morphology was observed during nitrate reduction under anaerobic conditions. The association between nitrate concentration and cell size or cellular fatty acid composition was evaluated. Nitrate-induced responses of M. multiformis were compared to those of related species. An increase in cellular filamentation and the ratio of saturated: unsaturated fatty acids was mediated specifically by nitrate. This indicates a decrease in cell fluidity and low leakage. Furthermore, a similar response was not observed in other related species cultured in the presence of nitrate. Hence, the nitrate-induced stress response in new anaerobes such as M. multiformis was demonstrated. The response could also be involved in the conservation of menaquinones and the maximization of nitrate reduction. Full article

Fe-dependent nitrate reduction by anammox consortia could serve as a valuable autotrophic denitrification process for wastewater treatment. However, influences of temperature, pH, and Fe/NO₃-N ratio on this biochemical process have not been studied. The present study investigated individual and interactive effects of aforementioned parameters on nitrate removal performance and bioactivity of anammox consortia via a series of batch assays. Enzymes activity of nitrate reductase (NAR) and hydrazine dehydrogenase (HDH) of anammox consortia had high consistency with nitrogen removal performance and significantly depended on temperature and Fe/NO₃-N ratio, while the narG and hdh genes expression were drastically depressed by extreme temperature. Models developed by response surface methodology (RSM) showed the significance of individual parameter followed by Fe/NO₃-N ratio > temperature > pH, while combined effects of temperature versus Fe/NO₃-N ratio exerted the most significant impacts. The pH in range of 4.0–8.0 had less influence. The optimum condition for nitrate removal efficiency (NRE) > 90% and total nitrogen removal efficiency (TNRE) > 75% was 4.0–7.4 for pH, 25.5–30.0 °C for temperature, and 31–48 for Fe/NO₃-N molar ratio. The maximum NRE and TNRE could be 98.68% and 79.42%, respectively, under the condition of pH = 4.00, temperature = 28.5 °C and Fe/NO₃-N ratio = 37.4. The models showed good dependability for simulation nitrogen removal performance by anammox in the real semiconductor wastewater. Full article

Uncharacterized protein STY1099, encoded by the yccT gene, was previously identified as the most altered (i.e., upregulated) protein among the ZnO nanoparticle (NP) stimulon of Salmonella enterica serovar Enteritidis. Here we combined various stress response-related assays with functional genetics, global transcriptomic and proteomic analyses to characterize the yccT gene and its STY1099 product. Exposure of S. enterica Enteritidis to H₂O₂ (i.e., hydrogen peroxide) resulted in a significant (p < 0.0001) upregulation of the yccT gene, whereas exposure to paraquat (i.e., superoxide) did not alter the expression of the yccT gene. The ∆yccT mutant of S. enterica Enteritidis exposed to 0.75 mM H₂O₂, showed significantly reduced (p < 0.05) viability compared to the wild type strain. Further, comparative transcriptome analyses supported by Co-immunoprecipitation (Co-IP) assay revealed that STY1099 protein plays a role in redox homeostasis during the peroxide stress assault via involvement in the processes of respiratory nitrate reductase, oxidoreductase activities, cellular uptake and stress response. In addition, we found that the STY1099 protein has the monopolar subcellular location and that it interacts with key cell division proteins, MinD, and FtsH, as well as with a rod shape-determining protein MerB. Full article