Comparative assessment of nitrogen fixation rate by ^(15)N_(2) tracer assays in the South China Sea

Nitrogen fixation is one of the most important sources of new nitrogen in the ocean and thus profoundly affects the nitrogen and carbon biogeochemical processes.The distribution,controlling factors,and flux of N2 fixation in the global ocean remain uncertain,partly because of the lack of methodological uniformity.The^(15)N_(2)tracer assay(the original bubble method→the^(15)N_(2)-enriched seawater method→the modified bubble method)is the mainstream method for field measurements of N2 fixation rates(NFRs),among which the original bubble method is the most frequently used.However,accumulating evidence has suggested an underestimation of NFRs when using this method.To improve the availability of previous data,we compared NFRs measured by three^(15)N_(2)tracer assays in the South China Sea.Our results indicate that the relationship between NFRs measured by the original bubble method and the^(15)N_(2)-enriched seawater method varies obviously with area and season,which may be influenced by incubation time,diazotrophic composition,and environmental factors.In comparison,the relationship between NFRs measured by the original bubble method and the modified bubble method is more stable,indicating that the N2 fixation rates based on the original bubble methods may be underestimated by approximately 50%.Based on this result,we revised the flux of N2 fixation in the South China Sea to 40 mmol/(m2·a).Our results improve the availability and comparability of literature NFR data in the South China Sea.The comparison of the^(15)N_(2)tracer assay for NFRs measurements on a larger scale is urgently necessary over the global ocean for a more robust understanding of the role of N2 fixation in the marine nitrogen cycle.

Effect of nitrogen fertilization on yield, N content, and nitrogen fixation of alfalfa and smooth bromegrass grown alone or in mixture in greenhouse pots

Planting grass and legume mixtures on improved grasslands has the potential advantage of realizing both higher yields and lower environmental pollution by optimizing the balance between applied N fertilizer and the natural process of legume biological nitrogen fixation. However, the optimal level of N fertilization for grass-legume mixtures, to obtain the highest yield, quality, and contribution of N2 fixation, varies with species. A greenhouse pot experiment was conducted to study the temporal dynamics of N2 fixation of alfalfa （Medicago sativa L.） grown alone and in mixture with smooth bromegrass （Bromus inermis Leyss.） in response to the addition of fertilizer N. Three levels of N （0, 75, and 150 kg ha-1） were examined using 15N-labeled urea to evaluate N2 fixation via the 15N isotope dilution method. Treatments were designated NO （0.001 g per pot）, N75 （1.07 g per pot） and N150 （2.14 g per pot）. Alfalfa grown alone did not benefit from the addition of fertilizer N; dry matter was not significantly increased. In contrast, dry weight and N content of smooth bromegrass grown alone was increased significantly by N application. When grown as a mixture, smooth bromegrass biomass was increased significantly by N application, resulted in a decrease in alfalfa biomass. In addition, individual alfalfa plant dry weight （shoots＋roots） was significantly lower in the mixture than when grown alone at all N levels. Smooth bromegrass shoot and root dry weight were significantly higher when grown with alfalfa than when grown alone, regardless of N application level. When grown alone, alfalfa＇s N2 fixation was reduced with N fertilization （R2=0.9376,P=0.0057）. When grown in a mixture with smooth bromegrass, with 75 kg ha-1 of N fertilizer, the percentage of atmospheric N2 fixation contribution to total N in alfalfa （%Ndfa） had a maximum of 84.07 and 83.05% in the 2nd and 3rd harvests, respectively. Total 3-harvest %Ndfa was higher when alfalfa was grown in a mixture than when grown alone （shoots： ｜t｜=3.39, P=0.0096; root： ｜t｜=3.57, P=0.0073）. We believe this was due to smooth bromegrass being better able to absorb available soil N （due to its fibrous root system）, resulting inlower soil N availability and allowing alfalfa to develop an effective N2 fixing symbiosis prior to the 1st harvest. Once soil N levels were depleted, alfalfa was able to fix N2, resulting in the majority of its tissue N being derived from biological nitrogen fixation （BNF） in the 2nd and 3rd harvests. When grown in a mixture, with added N, alfalfa established an effective symbiosis earlier than when grown alone; in monoculture BNF did not contribute a significant portion of plant N in the N75 and N150 treatments, whereas in the mixture, BNF contributed 17.90 and 16.28% for these treatments respectively. Alfalfa has a higher BNF efficiency when grown in a mixture, initiating BNF earlier, and having higher N2 fixation due to less inhibition by soil-available N. For the greatest N-use-efficiency and sustainable production, grass-legume mixtures are recommended for imDrovino orasslands, usino a moderate amount of N fertilizer （75 kq N ha-l） to provide optimum benefits.

Fertilization practices affect biological nitrogen fixation by modulating diazotrophic communities in an acidic soil in southern China

Biological nitrogen(N)fixation(BNF)driven by diazotrophs is an important pathway for N input in agricultural ecosystems.However,free-living BNF and its associated diazotrophic communities under different fertilization practices in acidic soils are poorly studied.Here,we conducted a long-term(29 years)fertilization experiment to explore how fertilization affected free-living BNF via changing biotic and abiotic variables.The fertilization treatments included an unfertilized control(CK),chemical N,phosphorus(P),and potassium(K)fertilizers(NPK),NPK plus lime(NPKL),NPK plus straw(NPKS),NPK plus straw and lime(NPKSL),and NPK plus manure(NPKM).Compared with CK(1.51 nmol C_(2)H_(2)g^(-1)d^(-1)),BNF rate was significantly(P<0.05)higher in NPKM(1.99 nmol C_(2)H_(2)g^(-1)d^(-1))but lower in NPK(0.55 nmol C_(2)H_(2)g^(-1)d^(-1)),NPKL(0.61 nmol C_(2)H_(2)g^(-1)d^(-1)),and NPKS(0.69nmol C_(2)H_(2)g^(-1)d^(-1)).Similarly,chemical fertilization treatments without manure reduced the gene abundance(0.71×10^(8)-1.18×10^(8)copies g^(-1))andα-diversity(Shannon index,1.11-2.43)of diazotrophic communities,whereas the treatment with manure had a positive effect on diazotrophic abundance(3.23×10^(8)copies g^(-1))and Shannon index(3.36).Non-parametric multivariate analysis of variance(PERMANOVA)suggested that manure application(R^(2)=0.212,P=0.001)had a stronger influence on diazotrophic community composition than the addition of lime(R^(2)=0.115,P=0.019)or straw(R^(2)=0.064,P=0.161).Random forest modeling revealed that BNF rates can be significantly(P<0.05)explained by soil pH(9.9%),diazotrophic community attributes(composition,8.5%;Chao 1 index,8.1%;abundance,6.0%;Shannon index,5.7%),and soil total carbon(5.1%).Partial least squares path modeling(PLS-PM)suggested that the diazotrophic community attributes and soil properties mainly provided direct and indirect contributions to the variations in BNF rates,respectively.The dominant genera,Pelomonas,Azospirillum,and Dechloromonas,were positively associated with BNF rates,with their members being observed as keystone species in the community network.Application of chemical fertilizers combined with manure is an effective practice for improving BNF in acidic soils by affecting soil diazotrophic communities.

Invasive Spartina alterniflora accelerates the increase in microbial nitrogen fixation over nitrogen removal in coastal wetlands of China

Salt marsh plants play a vital role in mediating nitrogen(N)biogeochemical cycle in estuarine and coastal ecosystems.However,the effects of invasive Spartina alterniflora on N fixation and removal,as well as how these two processes balance to determine the N budget,remain unclear.Here,simultaneous quantifications of N fixation and removal via^(15)N tracing experiment with native Phragmites australis,invasive S.alterniflora,and bare flats as well as corresponding functional gene abundance by qPCR were carried out to explore the response of N dynamics to S.alterniflora invasion.Our results showed that N fixation and removal rates ranged from 0.77±0.08 to 16.12±1.13 nmol/(g·h)and from 1.42±0.14 to 16.35±1.10 nmol/(g·h),respectively,and invasive S.alterniflora generally facilitated the two processes rates.Based on the difference between N removal and fixation rates,net N_(2)fluxes were estimated in the range of-0.39±0.14 to 8.24±2.23 nmol/(g·h).Estimated net N_(2)fluxes in S.alterniflora stands were lower than those in bare flats and P.australis stands,indicating that the increase in N removal caused by S.alterniflora invasion may be more than offset by N fixation process.Random forest analysis revealed that functional microorganisms were the most important factor associated with the corresponding N transformation process.Overall,our results highlight the importance of N fixation in evaluating N budget of estuarine and coastal wetlands,providing valuable insights into the ecological effect of S.alterniflora invasion.

High piezo/photocatalytic efficiency of Ag/Bi_(5)O_(7)I nanocomposite using mechanical and solar energy for N2 fixation and methyl orange degradation

In this work,Ag/Bi_(5)O_(7)I nanocomposite was prepared and firstly applied in piezo/photocatalytic reduction of N2 to NH3 and methyl orange(MO)degradation.Bi_(5)O_(7)I was synthesized via a hydrothermal-calcination method and shows nanorods morphology.Ag nanoparticles(NPs)were photo deposited on the Bi_(5)O_(7)I nanorods as electron trappers to improve the spatial separation of charge carriers,which was confirmed via XPS,TEM,and electronic chemical analyses.The catalytic test indicates that Bi_(5)O_(7)I presents the piezoelectric-like behavior,while the loading of Ag NPs can strengthen the character.Under ultrasonic vibration,the optimal Ag/Bi_(5)O_(7)I presents high efficiency in MO degradation.The degradation rate is determined to be 0.033 min
1,which is 4.7 folds faster than that of Bi_(5)O_(7)I.The Ag/Bi_(5)O_(7)I also presents a high performance in piezocatalytic N2 fixation.The piezocatalytic NH3 generation rate reaches 65.4 μmol L^(-1)g^(-1)h^(-1)with water as a hole scavenger.The addition of methanol can hasten the piezoelectric catalytic reaction.Interestingly,when ultrasonic vibration and light irradiation simultaneously act on the Ag/Bi_(5)O_(7)I catalyst,higher performance in NH3 generation and MO degradation is observed.However,due to the weak adhesion of Ag NPs,some Ag NPs would fall off from the Bi_(5)O_(7)I surface under long-term ultrasonic vibration,which would greatly reduce the piezoelectric catalytic performance.This result indicates that a strong binding force is required when preparing the piezoelectric composite catalyst.The current work provides new insights for the development of highly efficient catalysts that can use multiple energies.

Symbionts in Mucuna pruriens stimulate plant performance through nitrogen fixation and improved phosphorus acquisition

Aims South Africa is mainly dominated by savanna and grasslands ecosystems which have been previously reported to be acidic and nutrient deficient,specifically with regard to phosphorus(P)and nitrogen(N).Mucuna pruriens(L.)DC,commonly known as velvet bean,is an indigenous legume in most African countries and has been reported to withstand these adverse soil conditions.The legume is used in many countries of the world for their medicinal value as well as for soil fertilization purposes.Although there are reports on M.pruriens growth and establishment in nutrient stressed ecosystems,no investigation has been conducted on M.pruriens symbiotic interactions,N source preference and associated growth carbon costs when subjected to P deficiency.In this study,we determined the impact of microbial symbionts on N nutrition and growth carbon costs of M.pruriens under P deficiency.Methods Microbe inoculation soils were collected from four geographical distinct KwaZulu-Natal locations.Thereafter,seeds were germinated in these natural soils and in early stages of nodule development,then seedlings were transferred in sterile quartz sand and supplied with Long Ashton nutrient media with varying P concentrations.Important Findings The 16S RNA sequence results revealed that M.pruriens was nodulated by Burkholderia sp.,Paenibacillus sp.and Bacillus irrespective of P concentrations.Even though P deficiency resulted in decreased overall biomass/growth,the root biomass,nodule number and carbon costs increased.In addition,low P supplied saplings showed the highest arbuscular mycorrhiza fungi percentage root colonization.In M.pruriens,nitrogen derived from atmosphere had a positive correlation with P level and the saplings had a dual reliance on atmospheric derived N and soil derived N with increased reliance on soil N in low P supplied plants.Therefore,M.pruriens exhibited different morphological and microbial symbiosis when subjected to P deficiency.

N2 fixation rate and diazotroph community structure in the western tropical North Pacific Ocean

In the present study, we report N2 fixation rate(15N isotope tracer assay) and the diazotroph community structure(using the molecular method) in the western tropical North Pacific Ocean(WTNP)(13°–20°N, 120°–160°E). Our independent evidence on the basis of both in situ N2 fixation activity and diazotroph community structure showed the dominance of unicellular N2 fixation over majority of the WTNP surface waters during the sampling periods.Moreover, a shift in the diazotrophic composition from unicellular cyanobacteria group B-dominated to Trichodesmium spp.-dominated toward the western boundary current(Kuroshio) was also observed in 2013. We hypothesize that nutrient availability may have played a major role in regulating the biogeography of N2 fixation.In surface waters, volumetric N2 fixation rate(calculated by nitrogen) ranged between 0.6 and 2.6 nmol/(L·d) and averaged(1.2±0.5) nmol/(L·d), with <10 μm size fraction contributed predominantly(88%±6%) to the total rate between 135°E and 160°E. Depth-integrated N2 fixation rate over the upper 200 m ranged between 150 μmol/(m^2·d)and 480 μmol/(m^2·d)average(225±105) μmol/(m^2·d). N2 fixation can account for 6.2%±3.7% of the depthintegrated primary production, suggesting that N2 fixation is a significant N source sustaining new and export production in the WTNP. The role of N2 fixation in biogeochemical cycling in this climate change-vulnerable region calls for further investigations.

Determining N supplied sources and N use efficiency for peanut under applications of four forms of N fertilizers labeled by isotope^15N

Rational application of different forms of nitrogen（N） fertilizer for peanut（Arachis hypogaea L.） requires tracking the N supplied sources which are commonly not available in the differences among the three sources：root nodule,soil and fertilizer.In this study,two kinds of peanut plants（nodulated variety（Huayu 22） and non-nodulated variety（NN-1）） were choosed and four kinds of N fertilizers：urea-N（CONH_2-N）,ammonium-N（NH_4~＋-N）,nitrate-N（NO_3^--N） and NH_4~＋ ＋NO_3^--N labeled by^（15）N isotope were applied in the field barrel experiment in Chengyang Experimental Station,Shandong Province,China,to determine the N supplied sources and N use efficiency over peanut growing stages.The results showed that intensities and amounts of N supply from the three sources were all higher at middle growing stages（pegging phase and podding phase）.The accumulated amounts of N supply from root nodule,soil and fertilizer over the growing stages were 8.3,5.3 and 3.8g m^（-2） in CONH_2-N treatment,which are all significantly higher than in the other three treatments.At seedling phase,soil supplied the most N for peanut growth,then root nodule controlled the N supply at pegging phase and podding phase,but soil mainly provided N again at the last stage（pod filling phase）.For the whole growing stages,root nodule supplied the most N（47.8 and 43.0%） in CONH_2-N and NH_4~＋-N treatments,whereas soil supplied the most N（41.7 and 40.9%） in NH_4~＋ ＋NO_3^--N and NO_3^--N treatments.The N use efficiency was higher at pegging phase and podding phase,while accumulated N use efficiency over the growing stages was higher in CONH_2-N treatment（42.2%） than in other three treatments（30.4%in NH_4~＋-N treatment,29.4%in NO_3^--N treatment,29.4%in NH_4~＋ ＋NO_3^--N treatment）.In peanut growing field,application of CONH_2-N is a better way to increase the supply of N from root nodule and improve the N use efficiency.

Optimization of medium and cultivation conditions for enhanced exopolysaccharide yield by marine Cyanothece sp. 113

Cyanothece sp. 113 is a unicellular, aerobic, diazotrophic and photosynthetic marine cyanobacterium. The optimal medium for exopolysaccharide yield by the strain was 70.0 g/L of NaCl, and 0.9 g/L of MgSO4 based on the modified F/2 medium for cultivation of marine algae. The optimal cultivation condition for exopolysaccharide yield by this cyanobacterial strain was 29℃, aeration, and continuous illumination at 86.0 μE/M^2/S. Under the optimal conditions, over 18.4 g/L of exopolysaccharide was produced within 12 days. This was so far the highest exopolysaccharide yield produced with strains of Cyanothece sp. obtained.

Decoration of CdMoO_(4) micron polyhedron with Pt nanoparticle and their enhanced photocatalytic performance in N_(2) fixation and water purification

This study aimed to prepare and apply a novel Pt/CdMoO_(4) composite photocatalyst for photocatalytic N2 fixation and tetracycline degradation. The Pt/CdMoO_(4) composite was subjected to comprehensive investigation on the morphology, structure, optical properties, and photoelectric chemical properties. The results demonstrate the dispersion of Pt nanoparticles on the CdMoO_(4) surface. Close contact between CdMoO_(4) and Pt was observed, resulting in the formation of a heterojunction structure at their contact region. Density functional theory calculation and Mott-Schottky analysis revealed that Pt possesses a higher work function value than CdMoO_(4), resulting in electron drift from CdMoO_(4) to Pt and the formation of a Schottky barrier. The presence of this barrier increases the separation efficiency of electron-hole pairs, thereby improving the performance of the Pt/CdMoO_(4) composite in photocatalysis. When exposed to simulated sunlight, the optimal Pt/CdMoO_(4) catalyst displayed a photocatalytic nitrogen fixation rate of 443.7 μmol·L‒^(1)·g‒^(1)·h‒^(1), which is 3.2 times higher than that of pure CdMoO_(4). In addition, the composite also exhibited excellent performance in tetracycline degradation, with hole and superoxide species identified as the primary reactive species. These findings offer practical insights into designing and synthesizing efficient photocatalysts for photocatalytic nitrogen fixation and antibiotics removal.

Dimensional heterojunction design:The rising star of 2D bismuthbased nanostructured photocatalysts for solar-to-chemical conversion

With the fast-pace digitalization evolution in the current generation’s lifestyle and the industry revolution,the energy demand has been skyrocketed.Recently,the two-dimensional(2D)bismuth-based nanomaterials emerged as a promising photocatalyst candidate in solar fuel conversion,not only for its exceptional light absorption capability and tunable optical properties,but it also can be synthesized into diverse variety of nanomaterials with different ranges of potential gap and band position to fulfill the potential requirement of wide range of photocatalytic reaction.Yet,the weak light harvesting ability and ultrafast charge recombination has restricted its potential in commercial application.Thus,recent researches have been focusing on tackling these issues by incorporating some modification strategies such as heteroatom doping,vacancy engineering,facet engineering,bismuth rich strategy and heterojunction engineering.Herein,this review article presents the state-of-the-art modifications on 2D bismuth-based parent material,specifically on the relationship between each of the modification strategy on the electronic properties and surface chemistry in achieving boosted photocatalytic performance.In the view of the unique charge interaction between two semiconductors with different dimensions,we systematically discuss the rational heterostructure design from the dimensionality perspective,namely,point-to-face,line-to-face,face-to-face,and bulk-to-face in solar fuel conversion to provide inspiring insights for future interface engineering.Finally,the challenges and the future research outlook in the solar-to-fuel conversion are highlighted to push forward the design of high-performance bismuth-based photocatalyst in realizing commercialscale solar-to-fuel conversion.

Cyanobacterial photo-driven mixotrophic metabolism and its advantages for biosynthesis

Cyanobacterium offers a promising chassis for phototrophic production of renewable chemicals. Although engineered cyanobacteria can achieve similar product carbon yields as heterotrophic microbial hosts, their production rate and titer under photoautotrophic conditions are 10 to 100 folds lower than those in fast growing E. colt. Cyanobacterial factories face three indomitable bottlenecks. First, photosynthesis has limited ATP and NADPH generation rates. Second, CO2 fixation by ribulose-l,5-bisphosphate carboxylase/oxygenase （RuBisCO） has poor efficiency. Third, CO2 mass transfer and light supply are deficient within large photobioreac- tors. On the other hand, cyanobacteria may employ organic substrates to promote phototrophic cell growth, Nz fixation, and metabolite synthesis. The photo-fermenta- tions show enhanced photosynthesis, while CO2 loss from organic substrate degradation can be reused by the Calvin cycle. In addition, the plasticity of cyanobacterial path- ways （e.g., oxidative pentose phosphate pathway and the TCA cycle） has been recently revealed to facilitate the catabolism. The use of cyanobacteria as ＂green E. colt＂ could be a promising route to develop robust photobiorefineries.

Phosphorus-Mobilizing Rhizobacterial Strain Bacillus cereus GS6 Improves Symbiotic Efficiency of Soybean on an Aridisol Amended with Phosphorus-Enriched Compost

Legume plants are an essential component of sustainable farming systems.Phosphorus(P) deficiency is a significant constraint for legume production, especially in nutrient-poor soils of arid and semi-arid regions.In the present study, we conducted a pot experiment to evaluate the effects of a phosphorus-mobilizing plant-growth promoting rhizobacterial strain Bacillus cereus GS6, either alone or combined with phosphate-enriched compost(PEC) on the symbiotic(nodulation-N_2 fixation) performance of soybean(Glycine max(L.) Merr.) on an Aridisol.The PEC was produced by composting food waste with addition of single super phosphate.The bacterial strain B.cereus GS6 showed considerable potential for P solubilization and mobilization by releasing carboxylates in insoluble P(rock phosphate)-enriched medium.Inoculation of B.cereus GS6 in combination with PEC application significantly improved nodulation and nodule N_2 fixation efficiency.Compared to the control(without B.cereus GS6 and PEC), the combined application of B.cereus GS6 with PEC resulted in significantly higher accumulation of nitrogen(N), P, and potassium(K) in grain, shoot, and nodule.The N:P and P:K ratios in nodules were significantly altered by the application of PEC and B.cereus GS6, which reflected the important roles of P and K in symbiotic performance of soybean.The combined application of PEC and B.cereus GS6 also significantly increased the soil dehydrogenase and phosphomonoesterase activities, as well as the soil available N, P, and K contents.Significant positive relationships were found between soil organic carbon(C) content, dehydrogenase and phosphomonoesterase activities, and available N, P, and K contents.This study suggests that inoculation of P-mobilizing rhizobacteria, such as B.cereus GS6, in combination with PEC application might enhance legume productivity by improving nodulation and nodule N_2 fixation efficiency.

Relationship between acid phosphatase activity and P concentration in organs of Cyclopia and Aspalathus species,and a non-legume of the Cape Floristic Region

Aims The role of tissue acid phosphatase(APase)activity of legumes and non-legumes in their P nutrition and adaptation to low-P soils is not well understood.To better understand this,a relationship between APase activ-ity and P concentration in leaves,stems,roots and nodules of legumes,Cyclopia and Aspalathus and a non-legume,Leucadendron strictum,all native to the P-poor soils of the Cape fynbos biome,was assessed.Methods Plants were collected and each separated into leaves,stems and roots.Phosphatase enzyme activity was assayed in soil using the p-nitrophenol method,while soil P and shoot P were measured using ICP-MS.To measure tissue APase activity,an acetate buffer was added into ground plant material and contents filtered.An ace-tate buffer and a p-nitrophenyl solution were added to the superna-tant and contents incubated.After incubation,NaOH(0.5 M)was added and absorbance read at 405 nm.Important Findings At Koksrivier,Cyclopia genistoides exhibited the highest leaf enzyme activity whilst Aspalathus aspalathoides showed the highest enzyme activity in the stems.At both Kleinberg and Kanetberg,Cyclopia subternata and Cyclopia longifolia showed the highest APase activity in leaves,followed by stems and low-est in roots.P concentration closely mirrored enzyme activity in organs of all test species from each site.APase activity posi-tively correlated with P concentration in organs of all the test Cyclopia and Leucadendron species,indicating that intracellular APase activity is directly linked to P mobilization and transloca-tion in these species.Percentage of N derived from fixation was positively correlated with tissue APase activity in C.genistoides(r=0.911^(*)),A.aspalathoides(r=0.868^(*))and Aspalathus cal-edonensis(r=0.957^(*)),suggesting that APase activity could be directly or indirectly linked to symbiotic functioning in these fynbos legumes,possibly via increased P supply to sites of N2 fixation.