Nitrogen fixation in paddy soils

The 4 long-term experimental plots (Umbric haplaquept) with different fertilizer treatment at Cent. Agric. Exp. Sta. in Konosu City, Saitama Prefecture, were used for the sites of investigation. The 4 plots were NF (applied with no fertilizer), IF (applied with inorganic fertilizers), GM (applied with green manure and CaCO₂), and OM (applied with manure and inorganic fertilizers). Flooded water, floating weed, upper (0-2cm) and lower (2-10cm) parts of Apg horizon and rhizosphere were collected from each plot before flooding, during flooding, and after drainage. These samples were analyzed for N₂-fixing activity by acetylene reduction method, pH, Eh, and contents of Fe²⁺, NH₄ ⁺, chlorophyll-type compounds, and water-soluble carbohydrates.

The N₂-fixing activity of all samples showed almost the same pattern of change with time: very low before flooding, rapidly increased after flooding, the maximum value at the maximum tillering stage of rice plant, declined afterwards and reached a very low value after drainage.

Rough estimation of the “N₂-fixing capacity” of each part of the paddy field revealed that the most important site of the N₂ fixation was the reduced Apg horizon, that the importance of flooded water and/or the oxidized layer in the N₂ fixation was rather low except in infertile soil, and that the role of rhizosphere in the N₂ fixation could not be neglected also in Japan.

Reduced condition and content of easily decomposable organic substances were judged to be main factors which control the N₂-fixing activity in the flooded soil on the basis of correlations between the Nt-fixing activity and several analytical data of the paddy soils.     

Influences of organic debris and rice root on the nitrogen fixation in the submerged soil

Abstract

A series of experiments has been conducted on the N2 fixation in the paddy soils by the authors (1–4). The amount of organic substrates for microorganisms and the degree of reduction of the soil are found to be two major factors affecting the N₂-fixing activity of the heterotrophic microorganisms in the submerged soil. Organic debris, rice root and their neighboring soils are identified to be the important micro-sites for the heterotrophic N₂ fixers. The organic debris and the rice root are considered to play dual roles by supplying the organic substances; (1) increase of the population of the heterotrophic N₂ fixers—the amount of nitrogenase, (2) preparation of the reduced conditions favorable for the nitrogenase activity.

However, it is not yet clearly known which of these two roles of the organic substrates is more essential to the N₂-fixing activity in the paddy soil. In addition, it is expected that there must be some differences between the organic debris and rice root in their contribution to the N₂ fixation in the paddy soil.

An experiment was carried out to clarify these problems. Moist soil sample was collected from the plough layer of the paddy field at Central Agric. Exp. Sta. in Konosu City, Saitama Pref., passed through a 5 mm sieve and placed in pots (3 kg moist soil/pot). Ammonium sulfate, calcium superphosphate, and potassium chloride at the rate of 0.4-0.4-0.4 (N-P₂O₅-K₂0) g/pot were incorporated into the soil 7 days before transplanting. Split application of ammonium sulfate at the rates of 0.2 and 0.4 g N/pot were also incorporated at 30 and 41 days after transplanting respectively. These pots were divided into three series; planted (P-series), non-planted (N-series), and non-planted and applied with organic manure (O-series). In case of O-series, 60 g of fairly rotted organic manure was applied to each pot. Each pot of P-series was planted with two 4O-day-old seedlings of rice plant at 7 days after submergence. The Nseries was regarded as a control. Each series was not replicated in this preliminary experiment.     

稻鸭共作对不同栽培环境稻季CH4和N2O排放的影响

稻田是大气温室气体甲烷(CH₄)和氧化亚氮(N₂O)的重要排放源, 稻田温室气体减排一直是生态农业研究的热点。目前, 采用水稻品种选择利用、水分控制管理、肥料运筹管理、耕作制度调整以及种养结合模式等方法来减少稻田温室气体排放有较好实践效应, 但不同稻田栽培环境(露地、网室)基础上的稻鸭共作对麦秸全量还田的稻田温室气体排放特征及相关土壤理化特性关联性的影响尚为少见。本研究采用裂区设计, 在两种栽培环境条件下, 以无鸭子放养的常规稻作和麦秸不还田为对照, 在等养分条件下分析麦秸全量还田与稻鸭共作模式对稻田土壤氧化还原电位、CH₄排放量、产CH₄潜力及CH₄氧化能力、N₂O排放量及N₂O排放高峰期土壤反硝化酶活性、全球增温潜势、水稻产量的影响, 为稻田可持续生产和温室气体减排提供参考。结果表明, 麦秆还田增加了稻田产CH₄潜力、提高了CH₄排放量, 降低了稻田土壤反硝化酶活性、土壤氧化还原电位和N₂O排放量, 整体上导致全球增温潜势上升96.89%~123.02%; 稻鸭共作模式, 由于鸭子的不间断活动提高了稻田土壤氧化还原电位, 降低了稻田产CH₄潜力, 增强了稻田CH₄氧化能力, 从而降低稻田CH₄排放量, N₂O排放量虽有提高, 整体上稻鸭共作模式的全球增温潜势较无鸭常规稻田下降8.72%~14.18%; 网室栽培模式显著提高了稻田土壤氧化还原电位, 降低稻田产CH₄潜力、CH₄氧化能力和土壤反硝化酶活性, 减少了稻田CH₄和N₂O排放量, 全球增温潜势降低6.35%~13.14%。本试验条件下, 稻田土壤的CH₄氧化能力是产CH₄潜力的2.21~3.81倍; 相同环境条件下, 稻鸭共作和麦秸还田均能增加水稻实际产量, 网室栽培的所有处理较相应的露地栽培减少了水稻实际产量1.19%~5.48%。本试验表明, 稻鸭共作和网室栽培可减缓全球增温潜势, 稻鸭共作和麦秸还田能够增加水稻实际产量。     

The effects on N2 fixation (C2H2 reduction), bacterial population and rice plant growth of two modes of straw application to a wetland rice field

Summary The effects of incorporation and surface application of straw to a wetland rice field on nitrogen fixation (C₂H₂ reduction), bacterial population and rice plant growth were studied. Rice straw (5 t ha^–1) was chopped (10- to 15-cm pieces) and applied to the field 2 weeks before transplanting IR42, a long-duration variety, and IR50, a short-duration variety. The acetylene-reducing activity (ARA) of IR42 and IR50 measured at heading stage for 3 consecutive days showed significantly higher ARA in IR42 as a result of the 2 straw application methods. Mostly up to 20 days after straw surface application and incorporation, the dark ARA in the soil, total and N₂-fixing heterotrophs, and photoorganotrophic purple nonsulphur bacteria (POPNS) in the soil and in association with degrading straw were stimulated. Higher bacterial populations were associated with straw on the surface than with straw incorporated. The POPNS counts, in particular, were increased hundreds fold in the surface-applied straw treatment. Straw applications also increased the root, shoot and total plant biomass at heading stage and the total dry matter yield at harvest in both varieties. The data show the potentials of straw as a source of substrate for the production of microbial biomass and for the non-symbiotic N₂ fixation to improve soil fertility and plant nutrition.     

Effect of grazer regulation and algal inoculation on photodependent nitrogen fixation in a wetland rice field

Summary A dry season field experiment conducted for two consecutive years highlighted problems of achieving increased populations of N₂-fixing blue-green algae (BGA) in wetland rice fields. Inoculation of non-indigenous BGA strains, either dried or as fresh viable inocula even at high levels of application, was unsuccessful. A limiting effect of grazing invertebrate populations on BGA establishment was evident, but other factors were involved. Reducing grazer pressure did not permit establishment of inoculated BGA; interspecific competition and environmental factors may explain the inoculation failure. Grazer regulation permitted the establishment of a fast-growing indigenous N₂-fixing Anabaena and the doubling of N₂-fixing activity over a control. Neither inoculation nor grazer control affected grain yields significantly.     

Effect of soil physical properties on soybean nodulation and N2 fixation at the early growth stage in heavy soil field in Hachirougata Polder,Japan

We studied the effect of the soil physical properties on soybean nodulation and N₂ fixation in the heavy soil of an upland field (UF) and an upland field converted from a paddy field (UCPF) in the Hachirougata polder, Japan. Seeds of the soybean cultivar Ryuho were sown in each field with or without inoculation of Bradyrhizobium japonicum A1017. The soybean plants were sampled at 35 (V3) and 65 (Rl) d after sowing (DAS), and then nodulation and the percentage of N derived from N₂ fixation in the xylem sap were determined. The soil physical properties were different between UF and UCPF, especially the air permeability and soil water regime. Nodule growth was restricted in UCPF irrespective of rhizobial inoculation, though rhizobial infection was not inhibited by the unfavorable soil physical conditions. Soybean plant growth was closely related to the nodule mass and N₂ fixation activity, and the inoculation of a superior rhizobium strain was effective only at 35 DAS. These results indicate that soybean nodulation and N₂ fixation was considerably affected by the physical properties of heavy soil, and that it is important to maintain the N₂ fixation activity and inoculate the soybean plants with a superior rhizobium strain at a later growth stage in order to increase soybean production in heavy soil fields.     

Changes in acetylene reduction activities and effects of inoculated rhizosphere nitrogen-fixing bacteria on rice

Acetylene reduction activities (ARAs) of soils and rice plants during rice-growing season were monitored in temperate region in northeast China. This activity was significantly higher in rhizosphere soil than that in inter-row soil after rice seedlings were transplanted. The ARA was high for most of growing season, suggesting that the native N₂-fixing bacteria responded to rice roots very quickly. Sixteen strains of free-living N₂-fixing bacteria were isolated from three different soils. The ARAs of these strains were correlated with the averaged soil ARAs, suggesting that the isolated strains were likely the active flora responsive to rice roots. The strains were inoculated by soaking seedling roots into the liquid culture for 2 h, and the seedlings were transplanted into pots. Most strains tested did not show any growth-promoting effects except Azotobacter armeniacus and Azotobacter nigricans, which showed growth-promoting effects only at late rice growth stage and only when inoculated in combination but not separately. Present data indicated the promising future applications of these two strains in combination in the region, but further research is needed to understand the underlying mechanisms.     

水稻根际和周围土壤中微生物功能基因丰度与碳、氮状况及其通量之间的关系

Rapid nitrogen（N） transformations and losses occur in the rice rhizosphere through root uptake and microbial activities. However,the relationships between rice roots and rhizosphere microbes for N utilization are still unclear. We analyzed different N forms（NH＋4,NO-3, and dissolved organic N）, microbial biomass N and C, dissolved organic C, CH4 and N2O emissions, and abundance of microbial functional genes in both rhizosphere and bulk soils after 37-d rice growth in a greenhouse pot experiment. Results showed that the dissolved organic C was significantly higher in the rhizosphere soil than in the non-rhizosphere bulk soil, but microbial biomass C showed no significant difference. The concentrations of NH＋4, dissolved organic N, and microbial biomass N in the rhizosphere soil were significantly lower than those of the bulk soil, whereas NO-3in the rhizosphere soil was comparable to that in the bulk soil. The CH4 and N2O fluxes from the rhizosphere soil were much higher than those from the bulk soil. Real-time polymerase chain reaction analysis showed that the abundance of seven selected genes, bacterial and archaeal 16 S rRNA genes, amoA genes of ammonia-oxidizing archaea and ammonia-oxidizing bacteria, nosZ gene, mcrA gene, and pmoA gene, was lower in the rhizosphere soil than in the bulk soil, which is contrary to the results of previous studies. The lower concentration of N in the rhizosphere soil indicated that the competition for N in the rhizosphere soil was very strong, thus having a negative effect on the numbers of microbes. We concluded that when N was limiting, the growth of rhizosphere microorganisms depended on their competitive abilities with rice roots for N.     

Heterotrophy-coordinated diazotrophy is associated with significant changes of rare taxa in soil microbiome

Soil heterotrophic respiration during decomposition of carbon (C)-rich organic matter plays a vital role in sustaining soil fertility. However, it remains poorly understood whether dinitrogen (N₂) fixation occurs in support of soil heterotrophic respiration. In this study, ¹⁵N₂-tracing indicated that strong N₂ fixation occurred during heterotrophic respiration of carbon-rich glucose. Soil organic ¹⁵N increased from 0.37 atom% to 2.50 atom% under aerobic conditions and to 4.23 atom% under anaerobic conditions, while the concomitant CO₂ flux increased by 12.0-fold under aerobic conditions and 5.18-fold under anaerobic conditions. Soil N₂ fixation was completely absent in soils replete with inorganic N, although soil N bioavailability did not alter soil respiration. High-throughput sequencing of the 16S rRNA gene further indicated that: i) under aerobic conditions, only 15.2% of soil microbiome responded positively to glucose addition, and these responses were significantly associated with soil respiration and N₂ fixation and ii) under anaerobic conditions, the percentage of responses was even lower at 5.70%. Intriguingly, more than 95% of these responses were originally rare with < 0.5% relative abundance in background soils, including typical N₂-fixing heterotrophs such as Azotobacter and Clostridium and well-recognized non-N₂-fixing heterotrophs such as Sporosarcina, Agromyces, and Sedimentibacter. These results suggest that only a small portion of the soil microbiome could respond quickly to the amendment of readily accessible organic C in a fluvo-aquic soil and highlighted that rare phylotypes might have played more important roles than previously appreciated in catalyzing soil C and nitrogen turnovers. Our study indicates that N₂ fixation could be closely associated with microbial turnover of soil organic C when available in excess.     

Application of N2/Ar ratio method for N2 fixation studies in submerged soil

Abstract

Recently there has been developments in the measurement of N₂ fixation due mainly to the C₂H₂ reduction method (1). This method, however, has several disadvantages, especially for submerged soil, and the estimated amount of fixed N₂ on the basis of the C₂H₂ reduction activity is not very reliable. The tracer ¹⁵N₂ technique which gives a reliable estimation of the fixed N₂ is too expensive for common use. Development of an alternative method suitable for submerged soil would therefore be desirable. The present authors expected that the measurement of the ratio N₂/Ar in the soil solution might provide advantages for the estimation of the fixed N₂ in submerged soil.     

Effect of intermittent drainage in reduction of methane emission from paddy soils in Hokkaido,northern Japan

ABSTRACT

Emission of methane (CH₄), a major greenhouse gas, from submerged paddy soils is generally reduced by introducing intermittent drainage in summer, which is a common water management in Japan. However, such a practice is not widely conducted in Hokkaido, a northern region in Japan, to prevent a possible reduction in rice grain yield caused by cold weather. Therefore, the effects of intermittent drainage on CH₄ emission and rice grain yield have not been investigated comprehensively in Hokkaido. In this study, we conducted a three-year field experiment in Hokkaido and measured CH₄ and nitrous oxide (N₂O) fluxes and rice grain yield to elucidate whether the reduction in CH₄ emission can be achieved in Hokkaido as well as other regions in Japan. Four experimental treatments, namely, two soil types [soils of light clay (LiC) and heavy clay (HC) textures] and two water management [continuous flood irrigation (CF), and intermittent drainage (ID)], were used, and CH₄ and N₂O fluxes were measured throughout the rice cultivation periods from 2016 to 2018. Cumulative CH₄ emissions in 2016 were markedly low, suggesting an initially low population of methanogens in the soils presumably due to no soil submergence or crop cultivation in the preceding years, which indicates a possible reduction in CH₄ emission by introducing paddy-upland crop rotation. Cumulative CH₄ emissions in the ID-LiC and ID-HC plots were 21–91% lower than those in the CF-LiC and CF-HC plots, respectively, whereas the cumulative N₂O emissions did not significantly differ between the different water managements. The amount of CH₄ emission reduction by the intermittent drainage was largest in 2018, with a comparatively long period of the first drainage for 12 days in summer. Rice grain yields did not significantly differ between the different water managements for the entire 3 years, although the percentage of well-formed rice grains was reduced by the intermittent drainage in 2018. These results suggest that CH₄ emission from paddy fields can be reduced with no decrease in rice grain yield by the intermittent drainage in Hokkaido. In particular, the first drainage for a long period in summer is expected to reduce CH₄ emission markedly.     

水稻根际固氮量及根系不同部位的固氮活性

采用乙炔还原法测定水稻根际原状土粒的固氮活性,以3∶1的换算比例推算为田间固氮量。每季水稻根际固氮量早稻为0.4-0.45斤氮/亩/66天;晚稻为0.6-0.79斤氮/亩/72天。其中早稻80—90%的氮素在抽穗至成熟期固定,而晚稻在这段期间固定的氮素占总数的40—52%。经多次测定表明:水稻根系不同部位固氮活性的趋势是,埋入土中带根基茎段2.5厘米活性最高,根基以下3厘米根段次之,3厘米以下根段活性很低,水稻根系结合的腐生性细菌以固氮菌占优势,同时也含有少量的丁酸固氮梭菌。     

Use of 15N isotope dilution for quantification of N2 fixation associated with roots of kallar grass (Leptochloa fusca (L.))

Summary Leptochloa fusca (L.) Kunth (kallar grass) has previously been found to exhibit high rates of nitrogen fixation. A series of experiments to determine the level of biological nitrogen fixation using ¹⁵N isotopic dilution were carried out in nutrient solution and saline soil. In the nutrient solution, E. coli inoculated plants were taken as non-nitrogen-fixing control. It was observed that nearly 60%–80% of the plant N was derived from atmospheric fixation. Estimations based on the N difference method gave much lower values (18%–35%). In experiments with saline soil which was initially sterilized with chloroform fumigation, a mixed culture of N₂-fixing rhizospheric isolates from kallar grass roots was inoculated and planted to kallar grass. Uninoculated treatments were regarded as controls. The soil was previously labelled with ¹⁵N by adding cellulose and (¹⁵NH₄)₂SO₄. The results of these studies showed fixation values of 6%–32% when estimated by ¹⁵N dilution, whereas by the N difference method 54% of the plant N was estimated to be derived from fixation. This discrepancy is due to the increase in root proliferation due to inoculation, which results in greater uptake of soil N. The distribution of ¹⁵N in different fractions of the soil-N indicted isotopic dilution due to bacterial fixation of atmospheric N₂.     

Short-term measurements of uptake of nitrogen fixed in the rhizospheres of sorghum (Sorghum bicolor) and millet (Pennisetum americanum)

Summary In a series of short-term experiments root systems of young sorghum and millet plants inoculated with N₂-fixing bacteria were exposed to ¹⁵N₂-enriched atmospheres for 72 h. The plants were grown in a normal atmosphere for up to 22 days after the end of the exposure to allow them to take up the fixed N₂. Environmental conditions and genotypes of sorghum and millet were selected to maximise N₂-fixation in the rhizosphere. Detectable amounts of fixed N (> 16 g/plant) were rapidly incorporated into sorghum plants grown in a sand/farmyard manure medium, but measurable fixation was found on only one occasion in plants grown in soil. N₂ fixation was detectable in some experiments with soil-grown millet plants but the amounts were small (2–4 g/plant) and represented less than 1 % of plant N accumulated over the same period. In many cases there was no detectable ¹⁵N₂ incorporation despite measurable increases in ethylene concentration found during an acetylene reduction assay.Published as ICRISAT Journal Article No. JA 740     

Denitrifying ability of indigenous strains of Bradyrhizobium japonicum isolated from fields under paddy-upland rotation

Summary In Japan some paddy fields are used for upland crops for several years and then returned to paddy fields (paddy-upland rotation). Soybeans (Glycine max L.) are an important summer crop. The ability to denitrify and some characteristics of denitrification by isolated strains of Bradyrhizobium japonicum were investigated to clarify the frequency of denitrifiers in indigenous populations of B. japonicum in fileds under paddyupland rotation. Eight field plots with different cropping systems at two sites were used. The fields consisted of a Gray Lowland Soil, and either soybeans or paddy rice (Oryza sativa L.) was grown as a summer crop, with barley (Hordeum distichum L.) or wheat (Triticum aestivum L.) as a winter crop. All B. japonicum strains present in the plots were able to denitrify. Isolated strains fell into two main groups (groups I and II) according to the rate of denitrification. Strains of group I evolved N₂O with C₂H₂ at a rate comparable to that of Alcaligenes denitrificans IAM 12370, whereas the denitrification activity of group II strains was 100 times lower than that of group I strains. Both group I and II strains occurred in each plot. Amounts of N₂O produced by indigenous strains with and without C₂H₂ suggested that strains of group I and II evolved N₂ or N₂O, respectively, as the end product of denitrification. One strain (S 107) that was isolated had the highest denitrifying ability with an end product of N₂O. These results indicate that indigenous bradyrhizobia may partly contribute to denitrification of field soil under a paddy-upland rotation.     

Wheat straw management affects CH4 and N2O emissions from rice fields

A 3-year field experiment was conducted in Jiangsu Province, China from 2004 to 2006 to investigate CH₄ and N₂O emissions from paddy fields as affected by various wheat straw management practices prior to rice cultivation. Five methods of returning wheat straw, no straw, evenly incorporating, burying straw, ditch mulching and strip mulching, were adopted in the experiment. Evenly incorporating is the most common management practice in the region. Results showed that compared with no straw, evenly incorporating increased CH₄ emission significantly by a factor of 3.9-10.5, while decreasing N₂O emission by 1-78%. Methane emission from burying straw was comparable with that from evenly incorporating, while N₂O emission from burying straw was 94-314% of that from evenly incorporating. Compared with evenly incorporating, CH₄ emission was decreased by 23-32% in ditch mulching and by 32% in strip mulching, while N₂O emission was increased by a factor of 1.4-3.7 in ditch mulching and by a factor of 5.1 in strip mulching. During the rice-growing season, the emitted N₂O was negligible compared to that of emitted CH₄. No significant difference in grain yield was observed between ditch mulching, burying straw, evenly incorporating and no straw. Compared with no straw, the grain yield was increased by 27% in strip mulching. Based on these results, the best management practice for returning wheat straw to the soil is strip mulching wheat straw partially or completely onto the field surface, as the method reduced CH₄ emission from rice fields with no decrease in rice yield.     

Inoculation of rice with nitrogen-fixing bacteria —problems and perspectives

Summary The present status and merits of inoculating rice with N₂-fixing bacteria are discussed in the light of recent advances. Bacterial inoculation improves plant growth and rice yield but not uniformly. The yield response to inoculation is more pronounced in the presence of moderate levels of fertilizer N. Evidence for the establishment and activity of the inoculated bacteria is limited, and the poor survival of the inoculum under field conditions further complicates the effects of inoculation. There is no clear evidence that improved growth and mineral content following inoculation are due to increased N₂ fixation. Beneficial effects of the inoculum on rice, such as plant growth promotion, N₂ fixation and antagonism effects against pathogens need to be further investigated under laboratory and field conditions. Improved management practices, such as organic amendments, suitable water and soil management, selection of efficient microbial strains, selection of effective breeding lines with high associative nitrogen fixation, and better management of agrochemicals are some of the measures suggested for deriving benefits from bacterial associations with rice.     

Nitrogen fixation and beneficial effects of some grain legumes and green-manure crops on rice

Studies were conducted on paddy soils to ascertain N₂ fixation, growth, and N supplying ability of some green-manure crops and grain legumes. In a 60-day pot trial, sunhemp (Crotalaria juncia) produced a significantly higher dry matter content and N yield than Sesbania sesban, S. rostrata, cowpeas (Vigna unguiculata), and blackgram (V. mungo), deriving 91% of its N content from the atmosphere. Dry matter production and N yield by the legumes were significantly correlated with the quantity of N₂ fixed. In a lowland field study involving sunhemp, blackgram, cowpeas, and mungbean, the former produced the highest stover yield and the stover N content, accumulating 160–250 kg N ha^-1 in 60 days, and showed great promise as a biofertilizer for rice. The grain legumes showed good adaptability to rice-based cropping systems and produced a seed yield of 1125–2080 kg ha^-1, depending on the location, species, and cultivar. Significant inter- and intraspecific differences in the stover N content were evident among the grain legumes, with blackgram having the highest N (104–155 kg N ha^-1). In a trial on sequential cropping, the groundnut (Arachis hypogaea) showed a significantly higher N₂ fixation and residual N effect on the succeeding rice crop than cowpeas, blackgram, mungbeans (V. radiata), and pigeonpeas (Cajanus cajan). The growth and N yield of the rice crop were positively correlated with the quantity of N₂ fixed by the preceding legume crop.     

N2 fixation of nodules and N absorption by soybean roots associated with ridge tillage on poorly drained upland fields converted from rice paddy fields

Abstract

Sowing on elevated ridges is effective in reducing wet injury of soybean plants cultivated in upland fields converted from rice paddy fields. Therefore, we investigated the effect of ridge tillage (RT) on soybean N accumulation properties. We compared the amounts of plant N associated with N₂ fixation of nodules and from soil and fertilizer in the RT treatment with amounts in conventional tillage (CT) in two fields in 2002–2003. Both fields were upland fields converted from rice paddy fields (Typic Hydraquents). The main difference between the fields was the presence of a field underdrain. The amounts of Rb and K accumulated in the shoots were also determined to estimate soybean root distribution. The grain yields with RT increased in both fields from 106% to 129% compared with CT. Increased pod number and seed weight were the major factors responsible for the yield increase. anova indicated that RT significantly increased the activities of both N₂ fixation of nodules and N absorption by roots until R1 (flowering stage). The ratio of Rb and K accumulated in the shoots indicated that with RT, the root distribution was more abundant in the superficial layers compared with CT. Thus, RT reduced wet injury during the rainy season that overlapped the flowering stage. Nitrogen accumulation from N₂ fixation until the R7 stage with RT was significantly higher than that with CT. We concluded that RT was effective in increasing N₂ fixation of nodules in poorly drained upland fields converted from rice paddy fields.     

Low levels of ferredoxin, ATP and leghemoglobin contribute to limited N2 fixation of peas (Pisum sativum L.) and alfalfa (Medicago sativa L.) under S deficiency conditions

Sulphur (S) has become a major limiting factor for plant production in industrial as well as in remote industrial rural areas. Limitation of S can reduce legume N₂ fixation by affecting nodule development and function. In pot experiments with pea (Pisum sativum L.) and alfalfa (Medicago sativa L.), we investigated the influence of S on growth, ferredoxin, ATP and leghemoglobin concentrations. Addition of 200 mg S pot⁻¹ increased yield of shoots, roots and nodules of both plant species significantly. However, the influence of S on nodule yield formation was most pronounced. Pea and alfalfa roots were found to have higher S concentrations than shoots and being up to 2.9 times the S concentration in the shoots of peas under S-sufficient conditions. Sulphur addition also increased N₂ fixation significantly. The ferredoxin concentration in bacteroids of root nodules of pea was increased significantly by S only 10 weeks after planting and in bacteroids of root nodules of alfalfa 10 and 17 weeks after planting, while on per pot base the amounts of ferredoxin were higher throughout the experimental period of time. The ATP concentration of bacteroids of root nodules of both plant species as well as of mitochondria of root nodules of pea were significantly higher with optimum S supply. The effects of S deficiency on N₂ fixation are likely to be caused by the shortage of ferredoxin and ATP. The amount of leghemoglobin was reduced in comparison to nodules of the S-sufficient plants.