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Excessive ammonium disrupts the biological and physical characteristics of aquatic freshwater ecosystems, causing nutrient imbalances and toxicity. Different macrophytes exhibit varying tolerance levels to ammonium stress, influenced by species-specific adaptations. However, eutrophic water bodies not only have high nutrient loads but also exhibit low light transparency, necessitating an understanding of how submerged macrophytes cope with both high ammonium concentrations and low light conditions. In this study, we explored the tolerance of submerged macrophytes under these challenging conditions by testing various ammonium concentrations and light intensities. Our findings reveal that Myriophyllum spicatum demonstrates high ammonium tolerance under both optimal and low light intensities. Specifically, under optimal light, the primary ammonium assimilation pathway is catalyzed by NADH-GDH (Nicotinamide Adenine Dinucleotide-dependent Glutamate Dehydrogenase), with its activity increasing 4-fold at 50 mg L-1 [NH4+-N] compared to the control. Conversely, under low light intensity, the GS (Glutamine Synthetase)-catalyzed pathway becomes predominant, with GS activity rising 3-fold at 50 mg L-1 [NH4+-N] compared to the control. These results provide new insights into the adaptive mechanisms of M. spicatum, highlighting its flexible strategies for ammonium assimilation and its potential application in water restoration efforts. This study offers valuable information on the enzymatic pathways involved in ammonium detoxification, which is essential for developing effective strategies to manage and restore eutrophic aquatic systems.

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This study investigated the effect of nitrogen application on the rhizosphere soil microenvironment of sunflower and clarified the relationship between ammonium assimilation and the microenvironment. In a field experiment high (HN, 190 kg/hm2), medium (MN, 120 kg/hm2) and low nitrogen (CK, 50 kg/hm2) treatments were made to replicate plots of sunflowers using drip irrigation. Metagenomic sequencing was used to analyze the community structure and functional genes involved in the ammonium assimilation pathway in rhizosphere soil. The findings indicated that glnA and gltB played a crucial role in the ammonium assimilation pathway in sunflower rhizosphere soil, with Actinobacteria and Proteobacteria being the primary contributors. Compared with CK treatment, the relative abundance of Actinobacteria increased by 15.57% under MN treatment, while the relative abundance decreased at flowering and maturation stages. Conversely, the relative abundance of Proteobacteria was 28.57 and 61.26% higher in the MN treatment during anthesis and maturation period, respectively, compared with the CK. Furthermore, during the bud stage and anthesis, the abundance of Actinobacteria, Proteobacteria, and their dominant species were influenced mainly by rhizosphere soil EC, ammonium nitrogen (NH4+-N), and nitrate nitrogen (NO3--N), whereas, at maturity, soil pH and NO3--N played a more significant role in shaping the community of ammonium-assimilating microorganisms. The MN treatment increased the root length density, surface area density, and root volume density of sunflower at the bud, flowering, and maturity stages compared to the CK. Moreover, root exudates such as oxalate and malate were positively correlated with the dominant species of Actinobacteria and Proteobacteria during anthesis and the maturation period. Under drip irrigation, applying 120 kg/hm2 of nitrogen to sunflowers effectively promoted the community structure of ammonium-assimilating microorganisms in rhizosphere soil and had a positive influence on the rhizosphere soil microenvironment and sunflower root growth.

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A pilot-scale continuous-flow modified anaerobic-anoxic-oxic (MAAO) process examined the impact of external carbon sources (acetate, glucose, acetate/propionate) on ammonium assimilation, denitrifying phosphorus removal (DPR), and microbial community. Acetate exhibited superior efficacy in promoting the combined process of ammonia assimilation and DPR, enhancing both to 50.0 % and 60.0 %, respectively. Proteobacteria and Bacteroidota facilitated ammonium assimilation, while denitrifying phosphorus-accumulating organisms (DPAOs) played a key role in nitrogen (N) and phosphorus (P) removal. Denitrifying glycogen-accumulating organisms (DGAOs) aided N removal in the anoxic zone, ensuring stable N and P removal and recovery. Acetate/propionate significantly enhanced DPR (77.7 %) and endogenous denitrification (37.9 %). Glucose favored heterotrophic denitrification (29.6 %) but had minimal impact on ammonium assimilation. These findings provide valuable insights for wastewater treatment plants (WWTPs) seeking efficient N and P removal and recovery from low-strength wastewater.

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The use of slow-release fertilizers and seed-fertilizers cause localized high-ammonium (NH4 +) environments in agricultural fields, adversely affecting wheat growth and development and delaying its yield. Thus, it is important to investigate the physiological responses of wheat and its tolerance to NH4 + stress to improve the adaptation of wheat to high NH4 + environments. In this study, the physiological mechanisms of ammonium tolerance in wheat (Triticum aestivum) were investigated in depth by comparative analysis of two cultivars: NH4 +-tolerant Xumai25 and NH4 +-sensitive Yangmai20. Cultivation under hydroponic conditions with high NH4 + (5 mM NH4 +, AN) and nitrate (5 mM NO3 -, NN), as control, provided insights into the nuanced responses of both cultivars. Compared to Yangmai20, Xumai25 displayed a comparatively lesser sensitivity to NH4 + stress, as evident by a less pronounced reduction in dry plant biomass and a milder adverse impact on root morphology. Despite similarities in NH4 + efflux and the expression levels of TaAMT1.1 and TaAMT1.2 between the two cultivars, Xumai25 exhibited higher NH4 + influx, while maintaining a lower free NH4 + concentration in the roots. Furthermore, Xumai25 showed a more pronounced increase in the levels of free amino acids, including asparagine, glutamine, and aspartate, suggesting a superior NH4 + assimilation capacity under NH4 + stress compared to Yangmai20. Additionally, the enhanced transcriptional regulation of vacuolar glucose transporter and glucose metabolism under NH4 + stress in Xumai25 contributed to an enhanced carbon skeleton supply, particularly of 2-oxoglutarate and pyruvate. Taken together, our results demonstrate that the NH4 + tolerance of Xumai25 is intricately linked to enhanced glucose metabolism and optimized glucose transport, which contributes to the robust NH4 + assimilation capacity.

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Nutrient removal from sewage is transitioning to nutrient recovery. However, biological treatment technologies to remove and recover nutrients from domestic sewage are still under investigation. This study delved into the integration of ammonium assimilation with denitrifying phosphorus removal (DPR) as a method for efficient nutrient management in sewage treatment. Results indicated this approach eliminated over 80 % of the nitrogen in the influent, simultaneously recovering over 60 % of the nitrogen as the activated sludge through ammonia assimilation, and glycerol facilitated this process. The nitrification/denitrifying phosphorus removal ensured the stability of both nitrogen and phosphorus removal. The phosphorus removal rate exceeded 96 %, and the DPR rate reached over 90 %. Network analysis highlighted a stable community structure with Proteobacteria and Bacteroidota driving ammonium assimilation. The synergistic effect of fermentation bacteria, denitrifying glycogen-accumulating organisms, and denitrifying phosphorus-accumulating organisms contributed to the stability of nitrogen and phosphorus removal. This approach offers a promising method for sustainable nutrient management in sewage treatment.

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In natural and agricultural situations, ammonium ( NH 4 + ) is a preferred nitrogen (N) source for plants, but excessive amounts can be hazardous to them, known as NH 4 + toxicity. Nitrate ( NO 3 - ) has long been recognized to reduce NH 4 + toxicity. However, little is known about Brassica napus, a major oil crop that is sensitive to high NH 4 + . Here, we found that NO 3 - can mitigate NH 4 + toxicity by balancing rhizosphere and intracellular pH and accelerating ammonium assimilation in B. napus. NO 3 - increased the uptake of NO 3 - and NH 4 + under high NH 4 + circumstances by triggering the expression of NO 3 - and NH 4 + transporters, while NO 3 - and H+ efflux from the cytoplasm to the apoplast was enhanced by promoting the expression of NO 3 - efflux transporters and genes encoding plasma membrane H+ -ATPase. In addition, NO 3 - increased pH in the cytosol, vacuole, and rhizosphere, and down-regulated genes induced by acid stress. Root glutamine synthetase (GS) activity was elevated by NO 3 - under high NH 4 + conditions to enhance the assimilation of NH 4 + into amino acids, thereby reducing NH 4 + accumulation and translocation to shoot in rapeseed. In addition, root GS activity was highly dependent on the environmental pH. NO 3 - might induce metabolites involved in amino acid biosynthesis and malate metabolism in the tricarboxylic acid cycle, and inhibit phenylpropanoid metabolism to mitigate NH 4 + toxicity. Collectively, our results indicate that NO 3 - balances both rhizosphere and intracellular pH via effective NO 3 - transmembrane cycling, accelerates NH 4 + assimilation, and up-regulates malate metabolism to mitigate NH 4 + toxicity in oilseed rape.

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To investigate the effects of nitrogen source supply on microbial protein (MP) production by hydrogen-oxidizing bacteria (HOB) under continuous feed gas provision, a sequencing batch culture comparison (N2 fixation versus ammonium assimilation) was performed. The results confirmed that even under basic cultivation conditions, N2-fixing HOB (NF-HOB) communities showed higher levels of CO2 and N2 fixation (190.45 mg/L Δ CODt and 11.75 mg/L Δ TNbiomass) than previously known, with the highest biomass yield being 0.153 g CDW/g COD-H2. Rich ammonium stimulated MP synthesis and the biomass accumulation of communities (increased by 7.4 ～ 14.3 times), presumably through the enhancement of H2 and CO2 absorption. The micro mechanism may involve encouraging the enrichment of species like Xanthobacter and Acinetobacter then raising the abundance of nitrogenase and glutamate synthase to facilitate the nitrogen assimilation. This would provide NF-HOB with ideas for optimizing their MP synthesis activity.

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BACKGROUND: Phosphorus is one of the essential nutrients for plant growth. Phosphate-solubilizing microorganisms (PSMs) can alleviate available P deficiency and enhance plant growth in an eco-friendly way. Although ammonium toxicity is widespread, there is little understanding about the effect of ammonium stress on phosphorus solubilization (PS) of PSMs. RESULTS: In this study, seven PSMs were isolated from mangrove sediments. The soluble phosphate concentration in culture supernatant of Bacillus aryabhattai NM1-A2 reached a maximum of 196.96 mg/L at 250 mM (NH4)2SO4. Whole-genome analysis showed that B. aryabhattai NM1-A2 contained various genes related to ammonium transporter (amt), ammonium assimilation (i.e., gdhA, gltB, and gltD), organic acid synthesis (i.e., ackA, fdhD, and idh), and phosphate transport (i.e., pstB and pstS). Transcriptome data showed that the expression levels of amt, gltB, gltD, ackA and idh were downregulated, while gdhA and fdhD were upregulated. The inhibition of ammonium transporter and glutamine synthetase/glutamate synthase (GS/GOGAT) pathway contributed to reducing energy loss. For ammonium assimilation under ammonium stress, accompanied by protons efflux, the glutamate dehydrogenase pathway was the main approach. More 2-oxoglutarate (2-OG) was induced to provide abundant carbon skeletons. The downregulation of formate dehydrogenase and high glycolytic rate resulted in the accumulation of formic acid and acetic acid, which played key roles in PS under ammonium stress. CONCLUSIONS: The accumulation of 2-OG and the inhibition of GS/GOGAT pathway played a key role in ammonium detoxification. The secretion of protons, formic acid and acetic acid was related to PS. Our work provides new insights into the PS mechanism, which will provide theoretical guidance for the application of PSMs.

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To remove ammonium and tetracycline (TC) from wastewater, a new strain, DX-21, was isolated and exhibited simultaneous removal ability. The performance of DX-21 in TC removal, its removal mechanism, and the potential toxicities of the degradation products were investigated with genomics, mass spectrometry, density functional theory calculations, quantitative structure-activity relationship analyses, and Escherichia coli exposure experiments. DX-21 exhibited removal of ammonium (9.64 mg·L-1·h-1) via assimilation, and TC removal (0.85 mg·L-1·h-1) primarily occurred through cell surface bio-adsorption and biodegradation. Among the 12 identified degradation products, the majority exhibited lower toxicities than TC. Moreover, potential degradation pathways were proposed, including hydroxylation and deamination. Furthermore, DX-21 possessed TC resistance genes, various oxygenases and peroxidases that could potentially contribute to TC degradation. DX-21 colonized activated sludge and significantly enhanced the biodegradation of TC. Therefore, DX-21 showed potential for treating wastewater containing both ammonium and TC.

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Although ammonium (NH4+-N) is an important nutrient for plants, increases in soil nitrogen (N) input and atmospheric deposition have made ammonium toxicity a serious ecological problem. In this study, we explored the effects of NH4+-N stress on the ultrastructure, photosynthesis, and NH4+-N assimilation of Ottelia cordata (Wallich) Dandy, an endangered heteroblastic plant native to China. Results showed that 15 and 50 mg L-1 NH4+-N damaged leaf ultrastructure and decreased the values of maximal quantum yield (Fv/Fm), maximal fluorescence (Fm), and relative electron transport rate (rETR) in the submerged leaves of O. cordata. Furthermore, when NH4+-N was ≥ 2 mg L-1, phosphoenolpyruvate carboxylase activity (PEPC) and soluble sugar and starch contents decreased significantly. The content of dissolved oxygen in the culture water also decreased significantly. The activity of the NH4+-N assimilation enzyme glutamine synthetase (GS) significantly increased when NH4+-N was ≥ 10 mg L-1 and NADH-glutamate synthase (NADH-GOGAT) and Fd-glutamate synthase (Fd-GOGAT) increased when NH4+-N was at 50 mg L-1. However, the activity of nicotinamide adenine dinucleotide-dependent glutamate dehydrogenase (NADH-GDH) and nicotinamide adenine dinucleotide phosphate-dependent glutamate dehydrogenase (NADPH-GDH) did not change, indicating that GS/GOGAT cycle may play an important role in NH4+-N assimilation in the submerged leaves of O. cordata. These results show that short-term exposure to a high concentration of NH4+-N is toxic to O. cordata.

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Enteric bacteria use up to 15% of their cellular energy for ammonium assimilation via glutamine synthetase (GS)/glutamate synthase (GOGAT) and glutamate dehydrogenase (GDH) in response to varying ammonium availability. However, the sensory mechanisms for effective and appropriate coordination between carbon metabolism and ammonium assimilation have not been fully elucidated. Here, we report that in Salmonella enterica, carbon metabolism coordinates the activities of GS/GDH via functionally reversible protein lysine acetylation. Glucose promotes Pat acetyltransferase-mediated acetylation and activation of adenylylated GS. Simultaneously, glucose induces GDH acetylation to inactivate the enzyme by impeding its catalytic centre, which is reversed upon GDH deacetylation by deacetylase CobB. Molecular dynamics (MD) simulations indicate that adenylylation is required for acetylation-dependent activation of GS. We show that acetylation and deacetylation occur within minutes of "glucose shock" to promptly adapt to ammonium/carbon variation and finely balance glutamine/glutamate synthesis. Finally, in a mouse infection model, reduced S. enterica growth caused by the expression of adenylylation-mimetic GS is rescued by acetylation-mimicking mutations. Thus, glucose-driven acetylation integrates signals from ammonium assimilation and carbon metabolism to fine-tune bacterial growth control.

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Utilization of arbuscular mycorrhizal (AM) fungi (AMF) as a sustainable strategy in redeeming arsenic (As) toxicity in plants is a promising approach. Low As accumulation, restoration of physiological processes, and As tolerance by AMF have been documented in crop plants. However, to comprehend AM-mediated As tolerance in plants, understanding the biochemical responses of host to the symbiont is crucial. The study evaluated the effect of an AM fungus, Rhizophagus intraradices on tricarboxylic acid cycle (TCA) and nitrogen metabolism of Triticum aestivum under three As concentrations (0, 25, and 50 mg As kg-1 soil). Results showed that TCA cycle and nitrogen metabolism were severely impaired by As that resulted into a higher C/N ratio. However, colonization by R. intraradices attenuated As mediated alterations in TCA cycle by augmenting the activity of pyruvate dehydrogenase that provided sufficient substrate for the TCA cycle. Furthermore, mycorrhizal (M) plants reinstated the activities of isocitrate dehydrogenase, succinate dehydrogenase, fumarase, and malate dehydrogenase even under high As level. Although citrate synthase and oxoglutarate dehydrogenase activities declined upon As exposure in M-plants, these were nevertheless higher than their non-mycorrhizal (NM) counterparts, ensuring higher levels of citric acid and succinic acid in M-plants. AM colonization also moderated the As-mediated disturbances in nitrogen assimilation by augmenting the activity of nitrate reductase, nitrite reductase, glutamine synthase, and glutamine-2-oxoglutarate amino transferase. Overall findings of the study point out that colonization by R. intraradices favourably regulated the TCA cycle and nitrogen metabolism and confronted As-mediated alterations in C/N ratio.

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Nitrogen (N) removal from high-salinity wastewater is a major challenge. The aerobic-heterotrophic nitrogen removal (AHNR) process has been demonstrated to be feasible for treating hypersaline wastewater. In this study, Halomonas venusta SND-01, a halophilic strain capable of performing AHNR, was isolated from saltern sediment. The strain achieved ammonium, nitrite, and nitrate removal efficiencies of 98%, 81%, and 100%, respectively. The N balance experiment suggests that this isolate removes N mainly via assimilation. Various functional genes related to N metabolism were found in the genome of the strain, establishing a complex AHNR pathway that includes ammonium assimilation, heterotrophic nitrification-aerobic denitrification, and assimilatory nitrate reduction. Four key enzymes in the N removal process were successfully expressed. The strain exhibited high-adaptability under C/N ratios of 5-15, salinities of 2%-10% (m/v), and pH of 6.5-9.5. Therefore, the strain shows high potential for treating saline wastewater with different inorganic N compositions.

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Clostridium thermocellum is a cellulolytic thermophile that is considered for the consolidated bioprocessing of lignocellulose to ethanol. Improvements in ethanol yield are required for industrial implementation, but the incompletely understood causes of amino acid secretion impede progress. In this study, amino acid secretion was investigated via gene deletions in ammonium-regulated, nicotinamide adenine dinucleotide phosphate (NADPH)-supplying and NADPH-consuming pathways as well as via physiological characterization in cellobiose-limited or ammonium-limited chemostats. First, the contribution of the NADPH-supplying malate shunt was studied with strains using either the NADPH-yielding malate shunt (Δppdk) or a redox-independent conversion of PEP to pyruvate (Δppdk ΔmalE::Peno-pyk). In the latter, branched-chain amino acids, especially valine, were significantly reduced, whereas the ethanol yield increased from 46 to 60%, suggesting that the secretion of these amino acids balances the NADPH surplus from the malate shunt. The unchanged amino acid secretion in Δppdk falsified a previous hypothesis on an ammonium-regulated PEP-to-pyruvate flux redistribution. The possible involvement of another NADPH-supplier, namely, NADH-dependent reduced ferredoxin:NADP+ oxidoreductase (nfnAB), was also excluded. Finally, the deletion of glutamate synthase (gogat) in ammonium assimilation resulted in the upregulation of NADPH-linked glutamate dehydrogenase activity and decreased amino acid yields. Since gogat in C. thermocellum is putatively annotated as ferredoxin-linked, a claim which is supported by the product redistribution observed in this study, this deletion likely replaced ferredoxin with NADPH in ammonium assimilation. Overall, these findings indicate that a need to reoxidize NADPH is driving the observed amino acid secretion, likely at the expense of the NADH needed for ethanol formation. This suggests that metabolic engineering strategies that simplify the redox metabolism and ammonium assimilation can contribute to increased ethanol yields. IMPORTANCE Improving the ethanol yield of C. thermocellum is important for the industrial implementation of this microorganism in consolidated bioprocessing. A central role of NADPH in driving amino acid byproduct formation was demonstrated by eliminating the NADPH-supplying malate shunt and separately by changing the cofactor specificity in ammonium assimilation. With amino acid secretion diverting carbon and electrons away from ethanol, these insights are important for further metabolic engineering to reach industrial requirements on ethanol yield. This study also provides chemostat data that are relevant for training genome-scale metabolic models and for improving the validity of their predictions, especially considering the reduced degree-of-freedom in the redox metabolism of the strains generated here. In addition, this study advances the fundamental understanding on the mechanisms underlying amino acid secretion in cellulolytic Clostridia as well as on the regulation and cofactor specificity in ammonium assimilation. Together, these efforts aid in the development of C. thermocellum for the sustainable consolidated bioprocessing of lignocellulose to ethanol with minimal pretreatment.

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It is unknown whether nirBDs, which conventionally encode an NADH nitrite reductase, play other novel roles in nitrogen cycling. In this study, we explored the role of nirBDs in the nitrogen cycling of Pseudomonas putida Y-9. nirBDs had no effect on organic nitrogen transformation by strain Y-9. The △nirBD strain exhibited higher ammonium removal efficiency (90.7%) than the wild-type strain (76.1%; P < 0.05) and lower end gaseous nitrogen (N2O) production. Moreover, the expression of glnA (control of the ammonium assimilation) in the △nirBD strain was higher than that in the wild-type strain (P < 0.05) after being cultured in ammonium-containing medium. Furthermore, nitrite noticeably inhibited the ammonium elimination of the wild-type strain, with a corresponding removal rate decreasing to 44.8%. However, no similar impact on ammonium transformation was observed for the △nirBD strain, with removal efficiency reaching 97.5%. In conclusion, nirBDs in strain Y-9 decreased the ammonium assimilation and increased the ammonium oxidation to nitrous oxide.

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Ammonium (NH4+) is essential to generate the nitrogenous building blocks of life. It gets assimilated via the canonical biosynthetic routes to glutamate and is further distributed throughout metabolism via a network of transaminases. To study the flexibility of this network, we constructed an Escherichia coli glutamate auxotrophic strain. This strain allowed us to systematically study which amino acids serve as amine sources. We found that several amino acids complemented the auxotrophy either by producing glutamate via transamination reactions or by their conversion to glutamate. In this network, we identified aspartate transaminase AspC as a major connector between many amino acids and glutamate. Additionally, we extended the transaminase network by the amino acids ß-alanine, alanine, glycine, and serine as new amine sources and identified d-amino acid dehydrogenase (DadA) as an intracellular amino acid sink removing substrates from transaminase reactions. Finally, ammonium assimilation routes producing aspartate or leucine were introduced. Our study reveals the high flexibility of the cellular amination network, both in terms of transaminase promiscuity and adaptability to new connections and ammonium entry points.

Nitrogen is an essential part of many of the cell's building blocks, including amino acids and nucleotides, which form proteins and DNA respectively. Therefore, nitrogen has to be available to cells so that they can survive and grow. In nature, some microorganisms convert the gaseous form of nitrogen into ammonium, which then acts as the nitrogen source of most organisms, including bacteria, plants and animals. Once cells take up ammonium, it is 'fixed' by becoming part of an amino acid called glutamate, which has a so-called 'amine group' that contains a nitrogen. Glutamate then becomes the central source for passing these amines on to other molecules, distributing nitrogen throughout the cell. This coupling between ammonium fixation and glutamate production evolved over millions of years and occurs in all organisms. However, the complete metabolic network that underlies the distribution of amines remains poorly understood despite decades of research. Furthermore, it is not clear whether ammonium can be fixed in a way that is independent of glutamate. To answer these questions, Schulz-Mirbach et al. used genetic engineering to create a strain of the bacterium E. coli that was unable to make glutamate. These mutant cells could only grow in the presence of certain amino acids, which acted as alternative amine sources. Schulz-Mirbach et al. found that enzymes called transaminases, and one called AspC in particular, were required for the cells to be able to produce glutamate using the amine groups from other amino acids. Notably, Schulz-Mirbach et al. showed that AspC, which had previously been shown to use an amino acid called aspartate as a source of amine groups, is indispensable if the cell is to use the amine groups from other amino acids ­ including histidine, tyrosine, phenylalanine, tryptophan, methionine, isoleucine and leucine. Schulz-Mirbach et al. also discovered that if they engineered the E. coli cells to produce transaminases from other species, the repertoire of molecules that the cells could use as the source of amines to generate glutamate increased. In a final set of experiments, Schulz-Mirbach et al. were able to engineer the cells to fix ammonium by producing aspartate and leucine, thus entirely bypassing the deleted routes of glutamate synthesis. These data suggest that fixing ammonium and distributing nitrogen in E. coli can be very flexible. The results from these experiments may shed light on how cells adapt when there is not a lot of ammonium available. Moreover, this study could advance efforts at metabolic engineering, for example, to create molecules through new pathways or to boost the production of amino acids needed for industrial purposes.

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Microbial immobilization of fertilizer nitrogen (N) can effectively reduce N losses in soil. However, the effects of crop residue on microbial assimilation of fertilizer-N and the underlying microbial mechanisms in upland soils are unclear. We evaluated the influence of maize residue (13C) addition on the microbial assimilation of ammonium-N (15N) in DNA from fertilizer, and quantified the bacterial 13C or 15N assimilation by quantitative stable isotope probing (DNA-qSIP). We found that the straw addition did increase total microbial assimilation of ammonium from fertilizer during the 2-week incubation. However, bacterial taxa varied in their responses to straw addition: Bacteriodetes and Proteobacteria accounted for large fractions of ammonium assimilation and their N assimilations were increased, while N assimilations of Acidobacteria were decreased. We revealed that highly 13C-labeled taxa were the main contributors of N assimilation under straw addition. The straw primarily enhanced the contributions of bacterial taxa to ammonium assimilation through increasing the extent of N assimilation, or enhancing the abundance of the N-assimilating bacterial taxa. Overall, our study elucidated an interaction between microbial assimilation of fertilizer-N and straw-C, showing a close element coupling of the keystone functional microbial taxa in N immobilization driven by organic carbon.

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As a paradoxical nutrient in water ecosystems, ammonium can promote plants growth under moderate concentration, but excess of it causes phytotoxic effects. Previous research has revealed that glutamate dehydrogenase in the above-ground part of submerged macrophytes plays an important role in ammonium detoxification. However, the strategies of ammonium utilization at the whole plant level of submerged macrophytes are still unclear and the role of the above-ground part in nutrient utilization has not been clearly elucidated in previous studies, hence, directly influencing the application of previous theory to practice. In the present research, we combined the methods of isotopic labeling and enzyme estimation to investigate strategies of ammonium utilization by the submerged macrophytes. The results showed that when [NH4 +-N] was 50 mg L-1, 15N taken up through the above-ground parts was 13.24 and 17.52 mg g-1 DW, while that of the below-ground parts was 4.24 and 8.54 mg g-1 DW in Potamogeton lucens and Myriophyllum spicatum, respectively. The ratios of 15N acropetal translocation to uptake were 25.75 and 35.69%, while those of basipetal translocation to uptake were 1.93 and 4.09% in P. lucens and M. spicatum, respectively. Our results indicated that the above-ground part was not only the main part for ammonium uptake, but also the major pool of exogenous ammonium. Besides, the dose-response curve of GDH (increased by 20.9 and 50.2% under 15 and 50 mg L-1 [NH4 +-N], respectively) exhibited by the above-ground parts of M. spicatum indicates that it is the main site for ammonium assimilation of the tolerant species. This study identifies the ammonium utilization strategy of submerged macrophytes and reveals the important role of the above-ground part in nutrient utilization providing new insight into the researches of nutrient utilization by plants and theoretical supports for water restoration by phytoremediation.

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The contradiction between theoretical metabolism of ammonium assimilation and experiential understanding of conventional biosystems makes the rational optimization of the ammonium-assimilating microbiome through carbon to nitrogen (C/N) ratios perplexing. The effect of different C/N ratios on ammonium-assimilating biosystems was investigated in saline wastewater treatment. C/N ratios significantly hindered the nutrient removal efficiency, but ammonium-assimilating biosystems maintained functional stability in nitrogen conversions and microbial communities. With sufficient biomass, higher than 86% ammonium and 73% phosphorus were removed when C/N ratios were higher than 25. Ammonium assimilation dominated the nitrogen metabolism in all biosystems even under relatively low C/N ratios, evidenced by the extremely low abundances of nitrification functional genes. Different C/N ratios did not significantly change the bacterial community structure of ammonium-assimilating biosystems. It is anticipated that the ammonium-assimilating biosystem with advantages of clear metabolic pathway and easy optimization can be applied to nutrient removal and recovery in saline environments.

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Ammonium translocation through biological membranes, by the ubiquitous Amt-Mep-Rh family of transporters, plays a key role in all domains of life. Two highly conserved histidine residues protrude into the lumen of the pore of these transporters, forming the family's characteristic Twin-His motif. It has been hypothesized that the motif is essential to confer the selectivity of the transport mechanism. Here, using a combination of in vitro electrophysiology on Escherichia coli AmtB, in silico molecular dynamics simulations, and in vivo yeast functional complementation assays, we demonstrate that variations in the Twin-His motif trigger a mechanistic switch between a specific transporter, depending on ammonium deprotonation, to an unspecific ion channel activity. We therefore propose that there is no selective filter that governs specificity in Amt-Mep-Rh transporters, but the inherent mechanism of translocation, dependent on the fragmentation of the substrate, ensures the high specificity of the translocation. We show that coexistence of both mechanisms in single Twin-His variants of yeast Mep2 transceptors disrupts the signaling function and so impairs fungal filamentation. These data support a signaling process driven by the transport mechanism of the fungal Mep2 transceptors. IMPORTANCE Fungal infections represent a significant threat to human health and cause huge damage to crop yields worldwide. The dimorphic switch between yeast and filamentous growth is associated with the virulence of pathogenic fungi. Of note, fungal Mep2 proteins of the conserved Amt-Mep-Rh family play a transceptor role in the induction of filamentation; however, the signaling mechanism remains largely unknown. Amt-Mep-Rh proteins ensure the specific scavenging of NH4+ through a mechanism relying on substrate deprotonation, thereby preventing competition and translocation of similar-sized K+. Our multidisciplinary approaches using E. coli AmtB, Saccharomyces cerevisiae, and Candida albicans Mep2 show that double variation of the family-defining Twin-His motif triggers a mechanistic switch from a specific transporter to an unspecific ion channel with both mechanisms still coexisting in single variants. Moreover, we show that this mechanistic alteration is associated with loss of signaling ability of Mep2, supporting a transport mechanism-driven process in filamentation induction.

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