Increase In Nitrogenase Activity Research Articles

Root exudates in relation to growth and nitrogenase activity of Rhizobium japonicum.

Root exudates of cowpea and soybean were collected in aerated water cultures. The ability of these exudates to support nitrogenase activity of R. japonicum was studied on defined media. When a complete nitrogenase-inducing medium was supplemented with concentrated root exudate, there was an increased nitrogenase activity. When the cowpea root exudate was substituted for glutamine or sodium succinate in the medium, nitrogenase activity was not detected. Growth was good when the root exudate was substituted for sodium succinate but was poor when the exudate replaced glutamine. There was nitrogenase activity in R. japonicum when the cowpea root exudate was substituted for arabinose in the 20E medium. Gas chromatographic analyses of cowpea root exudates revealed that ribose, arabinose, glucose, and sucrose were the main constituents. The significance of this finding in the asymbiotic nitrogen fixation by Rhizobium spp. is discussed.

Root exudates in relation to growth and nitrogenase activity of Rhizobium japonicum

AbstractRoot exudates of cowpea and soybean were collected in aerated water cultures. The ability of these exudates to support nitrogenase activity of R.japonicum was studied on defined media. When a complete nitrogenase‐inducing medium was supplemented with concentrated root exudate, there was an increased nitrogenase activity. When the cowpea root exudate was substituted for glutamine or sodium succinate in the medium, nitrogenase activity was not detected. Growth was good when the root exudate was substituted for sodium succinate but was poor when the exudate replaced glutamine. There was nitrogenase activity in R. japonicum when the cowpea root exudate was substituted for arabinose in the 20E medium. Gas chromatographic analyses of cowpea root exudates revealed that ribose, arabinose, glucose, and sucrose were the main constituents. The significance of this finding in the asymbiotic nitrogen fixation by Rhizobium spp. is discussed.

On the Mechanism of Hydrogen Evolution by Nitrogen Starved Anabaena cylindrica

Summary The low rate of hydrogen production by air grown Anabaena cylindrica can be enhanced by nitrogen starvation (Weissman and Benemann, 1977). The starved filaments produce more heterocysts, parallel with increased activity of nitrogenase catalysing hydrogen evolution. Our results show that not only the increased nitrogenase activity, but also the decreased activity of the uptake hydrogenase, may be responsible for the severalfold higher hydrogen evolution. The efficiency of conversion of light energy into the chemical energy of hydrogen by nitrogen starved Anabaena cylindrica is O,3 %.

Effect of plant spacing and soil application of aldicarb on nitrogen fixation by spring-sown field beans (Vicia faba L.)

SummaryVicia faba cv. Minden was precision planted at densities of 12–100 plants/m2 in 1977 and 30–60 plants/m2 in 1978 with and without seed-bed applications of aldicarb (10 kg/ha).Nitrogenase activity per plant decreased significantly with increased plant density but this was compensated for on an area basis by increased numbers of plants. When expressed on the basis of area, nitrogenase activity was higher with closely, than with widely spaced plants both early and late in the season. Irrespective of planting density nitrogenase activity reached a peak during vegetative growth of the plant and a second peak during rapid pod and seed development. In 1977 the second peak was interrupted by very dry soil conditions but later activity increased following rain.Aldicarb consistently increased nitrogenase activity at all harvests after Sitona damage to the nodules was evident. Over this period plots treated with aldicarb had twice the nitrogenase activity of untreated plots in 1977 and 55% more in 1978.

Azolla filiculoides Lam. as a Fallow‐Season Green Manure for Rice in a Temperate Climate

AbstractAzolla pinnata R. Brown and its N‐fixing endophyte Anabaena azollae Strasburger are cultivated as a fallow‐season green manure for rice in Vietnam. This study presents field and laboratory experiments to determine if the temperate‐climate Azolla filiculoides Lam. could be grown in fallow flooded rice fields of California's Sacramento Valley.Field experiments in an agricultural drainage ditch indicate the exponential growth rate and N content of A. filiculoides is correlated logarithmically with maximum air temperature between early February and the end of April. Laboratory studies in controlled environment further revealed the exponential growth rate of A. filiculoides increases linearly with temperature between 10/1 and 25/15 C (12 hour thermoperiod and photoperiod) and remains high up to 35/25 C. Light saturation for growth is 100 µE✕m−2✕sec−1 at 10/1 C but at 35/25 C increases to at least 1,000 µE✕m−2✕sec−1. Nitrogenase activity increases with temperature between 10/1 C and 30/20 C, but is insensitive to light over a range of 50 to 1,000 µE✕m−2✕sec−1. Optimum N fixation for A. filiculoides is between 250 and 300 mg N✕g initial dry weigh−1✕week− and occurs at 25/15 to 30/20 C and 500 µE✕m−2✕sec−1.At 40/30 C the growth and nitrogenase activity of A. filiculoides is nil. However, if ferns are grown at lower temperatures and then subjected to a stepwise increase in temperature simulating dawn to midday of a diurnal cycle, nitrogenase activity increases with temperature up to 40 C and remains high at 45 C. Similar increases in nitrogenase activity with temperature are observed for field‐grown A. filiculoides during hot (40 to 45 C) afternoon periods.Inoculation of A. filiculoides equivalent to 1.2 kg N✕ha−1 onto 22.4 m−2 plots of a fallow flooded rice paddy results in N yields between 33 and 93 kg✕ha−1. Rice paddies used for 1977 studies were a Capay silty clay (Typic Chromoxererts), while 1978 studies were on a Hillgate loam (Typic Palexeralfs). On both soil series N yield of Azolla was correlated positively with the length of time the fern cover had developed prior to sporulation. Sporulation, which was followed by termination of nitrogenase activity and frond senescence, was hastened by high temperature. Thus, a mat of A. filiculoides developing in hot summer weather fixed only one‐third as much N as a population which developed over a longer period during early spring.Incorporation of 40 kg N✕ha−1 into soil as dry A. filiculoides in spring 1977 increased rice yield about 2.0 metric tons✕ha−1 over unfertilized controls. This effect was the same as would derive from an equal amount of N fertilizer in the form of ammonium sulfate and remained true when Azolla was supplemented with 40 and 80 kg N✕ha−1 as ammonium sulfate. Incorporation of 93 kg N✕ha−1 as dry A. filiculoides grown over a 46 day period in spring 1978 increased rice yield 2.6 MT✕ha−1, or 70%, of the increase obtained with an equivalent amount of ammonium sulfate.While more data are needed for A. filiculoides at different sites and years, results from this 2‐year study suggest up to 50% of the N requirement for rice in California could be supplied by one fallow‐season crop of Azolla. Further experiments will be needed to determine the optimum levels of inorganic nutrients (P and Fe) and labor inputs needed to grow Azolla before this source of biologically fixed N can be compared to N fertilizers currently used in temperate‐zone rice culture

Nitrogen fixation and allantoin formation in soybean plants.

The activity of nitrogenase and the concentration of ammonia and allantoin (+ allantoic acid) in root nodules were measured throughout the growth period of soybean plants. Nitrogenase activity measured by acetylene reduction increased with plant growth and reached a maximum level at the flowering period. The level of ammonia and allantoin concentration in nodules was parallel with increased nitrogenase activity. At the late reproductive stage (pod-forming period), nitrogenase activity showed a marked decrease, but the ammonia and allantoin in the nodules remained at a constant level. Detached nodules from 56 day-old soybean plants were exposed to 15N2 gas, and the distribution of 15N among nitrogen compounds was investigated. Enrichment of 15N in allantoin and allantoic acid reached a fairly high level after 90 min of nitrogen fixation; ca. 22% of 15N in acid-soluble nitrogen compounds was incorporated into allantoin + allantoic acid. In contrast, enrichment of 15N in amide nitrogen was relatively low. No...

Acetylene reduction assay on white clover genotypes and on grass/clover swards

SUMMARYAcetylene reduction assay was used to measure the nitrogenase activity of white clover genotypes in pots and of grass/clover swards in situ. Much of the variation in nitrogenase activity of single genotypes and hybrid populations was associated with plant dry weight. After adjustment for plant dry weight it was concluded that there was limited scope for selection for increased nitrogenase activity. In plant breeding this technique would seem to have greatest application in the selection for continued nitrogen fixation activity in the presence of inhibitory factors such as high levels of mineral nitrogen.The in situ studies revealed differences in nitrogenase activity of grass/clover swards based on contrasting cultivars of white clover. These differences were due to variation in clover density and also to variation in activity per unit clover dry weight. It was concluded that the variation in nitrogenase activity per unit dry weight reflected differences in growth pattern of the cultivars in the autumn when these assays were made.The in situ studies offer a means whereby the nitrogenase activity of cultivars and selected families can be monitored under varying levels of mineral nitrogen and other husbandry treatments without disruption of the sward.

Effect of Early Mycorrhizal Infection on Nodulation and Nitrogen Fixation in Trifolium subterraneum L

Three experiments are described. Rapid establishment of vesicular-arbuscular mycorrhizas in roots of T. subterraneum cv. Mt Barker, using natural soil inoculum, was associated with improved nodulation, increased nitrogenase activity per plant (nmol C2H2 reduced per plant per hour) and increased nodule efficiency on the basis of nodule volume (nmol C2H2 reduced per mm� nodule per hour). In two experiments (on soil low in nutrients), this increase occurred before any positive growth response to mycorrhizal infection was apparent. In all experiments, mycorrhizal roots had a higher phosphorus concentration (%P) than did non-mycorrhizal roots. This difference, which was evident before any differences in total plant phosphorus were detected, was not accompanied by an increase in nodule phosphorus concentration, so that differences in nodule efficiency could not be attributed to differences in this parameter. In the third experiment (on soil with higher nutrient levels), establishment of mycorrhizas was also accompanied by increased growth, phosphorus and nitrogen contents within a 35-day experimental period. Phosphorus inflow into roots (moles P per cm root per second) was higher in mycorrhizal plants. Delay in formation of mycorrhizas (by reduction in the amount of inoculum in soil) was accompanied by lower inflow, and delay in both the establishment of high root phosphorus concentration and in the onset of enhanced nodulation and nitrogenase activity.

The Relationship between Nitrogenase and Glutamine Synthetase in Bacteroids of Rhizobium leguminosarum of Various Ages

The activities of nitrogenase, glutamine synthetase and other ammonia-assimilating enzymes were studied in bacteroids of Rhizobium leguminosarum isolated 9 to 21 d after infection. Total bacteroid numbers (per plant) increased proportionally with nitrogenase, but glutamine synthetase activity decreased with increasing age of the root nodule. Glutamine synthetase was adenylylated throughout the period of increasing nitrogenase activity and during the period of constant nitrogenase activity.

Activity of nitrogenase and glutamine synthetase in relation to availability of oxygen in continuous cultures of a strain of cowpea Rhizobium sp. supplied with excess ammonium

In samples from nitrogen-fixing continuous cultures of strain CB756 of the cowpea type rhizobia ( Rhizobium sp.), newly fixed NH 4 + is in equilibrium with the medium, from where it is assimilated by the glutamine synthetase/glutamate synthase pathway. In samples from steady state cultures with different degrees of oxygen-limitation, nitrogenase activity was positively correlated with the biosynthetic activity of glutamine synthetase in cell free extracts. Also, activities in biosynthetic assays were positively correlated with activities in γ-glutamyl transferase assays containing 60 mM Mg 2+. Relative adenylylation of glutamine synthetase was conveniently measured in cell free extracts as the ratio of γ-glutamyl transferase activities without and with addition of 60 mM Mg 2+. Automatic control of oxygen supply was used to facilitate the study of transitions between steady-state continuous cultures with high and low nitrogenase activities. Adenylylation of glutamine synthetase and repression of nitrogenase activity in the presence of excess NH 4 +, were masked when oxygen strongly limited culture yield. Partial relief of the limitation in cultures supplied with 10 mM NH 4 + produced early decline in nitrogenase activity and increase in relative adenylylation of glutamine synthetase. Decreased oxygen supply produced a rapid decline in relative adenylylation, followed by increased nitrogenase activity, supporting the concept that control of nitrogenase synthesis is modulated by glutamine synthetase adenylylation in these bacteria.

Associative nitrogenase activity by some Hawaiian grass roots

Nitrogenase activity by the roots of some tropical grasses was detected by the acetylene reduction technique. In some cases particular plant species, evaluated at different times from different sites, displayed marked variations in nitrogenase activity. Acetylene reduction by root samples which had undergone preincubation under low O2 displayed an increase of approximately 5-fold when compared to plant-containing soil cores. These increases in nitrogenase activity correspond to an increase in the number of nitrogen-fixing bacteria associated with the roots.

Arrangement and regulation of the nitrogen fixation genes in Klebsiella pneumoniae studied by depression kinetics.

Events underlying depression of the nitrogen fixation (nif) genes in Klebsiella pneumoniae M5A1 were analyzed in vivo by comparing the effects of selective inhibitors of transcription and translation on subsequent nitrogenase activity (rate of acetylene reduction). When batch cultures were induced for depression, an 87-min lag separated ammonium ion/oxygen removal and the appearance of activity.

Ethylene as a factor limiting the legume‐Rhizobium symbiosis in tube culture

ABSTRACTGrowth and nitrogen fixation by Trifolium subterraneum plants at 15 and 10 oC root temperatures is much less in test tubes plugged with cotton wool than in open pots of sand. Removing the ethylene produced by plants growing in cotton wool plugged flasks by adsorption onto activated charcoal, or by aerating with sterile air, increased nitrogenase activity and growth over the first 25 days, to levels approaching that of plants grown in open pots of sand.

THE EFFECTS OF FLOWERING AND FRUIT FORMATION ON THE SUPPLY OF PHOTOSYNTHETIC ASSIMILATES TO THE NODULES OF PISUM SATIVUM L. IN RELATION TO THE FIXATION OF NITROGEN

SUMMARYBoth nitrogenase activity and the accumulation of 14C‐labelled photosynthates in the nodules of pea plants in nitrogen‐free culture reached maxima shortly before flowering and fruit development. During the period from flowering to fruiting, nitrogenase activity and accumulation of 14C‐photosynthates in the nodules declined by 60%, whereas the photosynthesis of the plant doubled. Studies of the translocation of the photosynthates within the plant during this period suggested that this might be due to an increase from 17% to 50% of the photosynthates which accumulated at the shoot apex. Consistent increases in nitrogenase activity over the 24 hours following removal of the shoot apex were only achieved in fruiting plants, however, although vegetative, flowering and fruiting plants all showed increased accumulation of photosynthates in the nodules. Continuous removal of flowers as they were formed over a 2‐week period resulted in a large increase in accumulation of photosynthates in the nodules and in nitrogenase activity.Other factors which may regulate nitrogenase activity in these plants are discussed.

Nitrogen metabolism and ultrastructure in Anabaena cylindrica. II. The effect of molybdenum and vanadium.

The structural and functional symptoms of molybdenum deficiency inAnabaena cylindrica grown in a medium without combined nitrogen and thus dependent on fixation of elemental nitrogen, resemble those brought about by nitrogen starvation. However, the substantially increased rate of heterocyst differentiation in this culture is not accompanied by a corresponding increase in nitrogenase activity; on the contrary, enzyme activity is severely impaired in the absence of molybdenum. When the supply of molybdenum, or of ammonia, is restored, the alga recovers rapidly. Vanadium exerts an inhibitory effect upon nitrogen-fixing ability of the alga, and its presence in the molybdenum-deficient culture results in the amplification of the symptoms of mlybdenum deficiency.

Differentiation in Nostoc muscorum: nitrogenase is synthesized in heterocysts.

Cellular differentiation can be observed in certain filamentous blue-green algae after transfer of the cells from medium containing NH(4) (+) or NO(3) (-) to nitrogen-free medium. The appearance of differentiated cells (heterocysts) is accompanied by an increase in the activity of nitrogenase, an enzyme complex that reduces N(2) to NH(3). We have separated vegetative cells from heterocysts in differentiated filaments of Nostoc muscorum, and analyzed their proteins by polyacrylamide-gel electrophoresis. The pattern of incorporation of (35)SO(4) (-) into proteins of the two cell types during differentiation indicates that the increase in nitrogenase activity is due to the de novo synthesis of nitrogenase proteins predominantly, if not exclusively, in heterocysts.

Rhizobium Strain Influence on Disruption of Clover Nodule Development at High Root Temperature

SUMMARY: Trifolium subterraneum plants grown at 30°C root temperature nodulated 3 days after inoculation with Rhizobium trifolii and nitrogenase activity (determined by acetylene-reduction) was detected 3 days later. While all four strains of R. trifolii examined formed effective nodules at 22°C, at 30°C only the nodules formed by strain TAI showed a rapid increase in nitrogenase activity. Light and electron microscopy of the nodules formed by one temperature-sensitive strain (NA30) showed the following abnormalities: multiple branching and distortion of the infection threads; failure of many bacteria to develop into characteristic bacteroids; continued division of these bacteria to give atypical multiple occupancy of the membrane envelopes; release of polysaccharide from ruptured infection threads into the host cytoplasm; rapid degeneration of the membrane envelopes and their contents. While the development of the membrane envelopes appeared to be under host control, the effect of the higher temperature on bacteroid development was highly strain specific. Upon transfer of nodulated plants from 22 to 30°C, bacteroid tissue in nodules formed by the temperature-sensitive strain NA30 broke down rapidly but nodules formed by strain TAI were less severely affected.