Assessment of the effects of the photosynthesis-inhibiting herbicides Diuron,DCMU, metamitron and metribuzin on growth and nitrogenase activity of Nostoc muscorum and a new cyanobacterial isolate,strain G4

Summary The influence of the photosynthesis-inhibiting herbicides Diuron, DCMU, metamitron, and metribuzin on growth and nitrogenase activity of Nostoc muscorum and a new cyanobacterial isolate, strain G4, was studied. The experiments were performed under N₂-fixing photoautotrophic conditions. Both cyanobacteria showed a high degree of tolerance towards the herbicides tested. In the presence of metamitron (35, 70, and 140 ppm) and metribuzin (7, 14, 35, and 70 ppm) strain G4 proliferated as well as the control culture. Metamitron and metribuzin had no influence on the nitrogenase synthesis of strain G4 and N. muscorum. When treated with 1 M DCMU, strain G4 and N. muscorum showed partial inhibition for the first few days, but entirely recovered during succeeding incubation. In the presence of 10 ppm Diuron, N. muscorum and strain G4, compared to the control, showed 30% and 80% nitrogenase activity, respectively.     

Free-living nitrogen-fixing bacteria in temperate cropping systems: Influence of nitrogen source

Sustainable cropping systems rely on a minimum of external inputs. In these systems N is largely acquired in animal manures and leguminous green manures. Little is known of how these organic forms of N fertilizer influence the presence and activity of free-living N₂-fixing bacteria. High concentrations of inorganic N in soil inhibit N₂-fixation in cyanobacteria and Azotobacter spp. It is likely that manure and fertilizer applications would result in concentrations of inorganic N capable of inhibiting N₂ fixation and, ultimately, the presence of these organisms. We investigated the effect of synthetic and organic N fertilizer sources on the populations and N₂-fixation potential of free-living N₂-fixing bacteria in the Farming Systems Trial at the Rodale Research Institute. Field plots received the following N treatments prior to corn (Zea mays L.) production: (1) Legume rotations and green manures supplying about 165 kg N ha^-1; (2) beef cattle manure applied at a rate of 220 kg N ha^-1 (plus 60 kg N ha^-1 from 1994 hay plow-down); or (3) fertilizer N (urea and NH₄NO₃) applied at a rate of 145 kg N ha^-1. Soil samples were collected at two depths from corn plots four times during the growing season, and analyzed for soil moisture, soil pH, numbers of N₂-fixing cyanobacteria and Azotobacter spp., extractable NH _inf4 ^sup+ and NO _inf3 ^sup- , and potentially mineralizable N. Soil samples collected in mid-July were analyzed for nitrogenase activity (by C₂H₂ reduction) and total C and N. Populations of Azotobacter spp. and cyanobacteria were influenced only slightly by treatment; however, cyanobacteria species composition was notably influenced by treatment. Nitrogenase activity in surface soils was greatest in legume-N plots and in subsurface plots levels were greatest in fertilizer-N plots. Populations and activity of free-living N-fixing bacteria appeared to be somewhat reduced in all plots as a result of low soil pH levels and high concentrations of inorganic N across all treatments. Annual applications of N to all plots resulted in high levels of potentially mineralizable N that in turn may have reduced non-symbiotic N₂-fixation in all plots.     

The effect of earthworms (Lumbricidae) on nitrogenase activity in soil

Summary Nitrogenase activity associated with earthworms, their faeces and activity in soil was measured by the acetylene reduction technique. A clear increase in nitrogenase activity was found in field-deposited casts of Aporrectodea caliginosa in comparison with surrounding soil, although potential nitrogenase activity was significantly higher in soil than in casts. Nitrogenase activity associated directly with earthworms (Lumbricus rubellus ) was detected, indicating the presence of active N₂-fixing bacteria on the body surface and/or in the gut. Laboratory experiments showed that nitrogenase activity in the casts of L. rubellus was higher than in unmodified soil, and that nitrogenase activity in soil was significantly increased by the burrowing and feeding activity of these worms. This paper discusses the possible causes of these earthworm effects on soil nitrogenase activity and some methodological problems of determining the nitrogenase activity.     

Choice of nitrogen fertilizer affects grain yield and agronomic nitrogen use efficiency of wheat cultivars

Abstract

Nitrogen use efficiency (NUE) is low in cereals especially in wheat. Different wheat cultivars may vary in NUE due to inherited biological nitrification inhibition (BNI) potential. In this study, three wheat cultivars (Punjab-2011, ARRI-2011 and Millat-2011) were fertilized at the rate of 140?kg ha^?1 with three N sources [nitrophos (NP), urea and calcium ammonium nitrate (CAN)]. The soil nitrate (NO₃^?)-N contents were significantly enhanced coupled with simultaneous decrease in ammonium (NH₄⁺)-N contents in the rhizosphere of cultivar Punjab-2011, fertilized with NP; however, cultivar Millat-2011 receiving urea behaved in contrast. Wheat cultivar Punjab-2011 fertilized with NP had the highest grain yield and agronomic NUE than other treatments due to significant increase in chlorophyl contents, allometric and yield parameters. The highest net benefit was recorded from the cultivar Punjab-2011 fertilized with CAN. In conclusion, use of NP in Punjab-2011 enhanced the grain yield and agronomic NUE.     

A bradyrhizobial-<Emphasis Type="Italic">Penicillium</Emphasis> spp. biofilm with nitrogenase activity improves N<Subscript>2</Subscript> fixing symbiosis of soybean

A bradyrhizobial-fungal biofilm (i.e. Bradyrhizobium elkanii SEMIA 5019-Penicillium spp.) developed in vitro was assayed for its nitrogenase activity and was evaluated for N₂-fixing symbiosis with soybean under greenhouse conditions. The biofilm showed nitrogenase activity, but the bradyrhizobial strain alone did not. Shoot and root growth, nodulation and N accumulation of soybean increased significantly with an inoculum developed from the biofilm. This study concludes that such biofilmed inoculants can improve N₂-fixing symbiosis in legumes, and can also directly contribute to soil N fertility in the long term. Further studies should be conducted to investigate the performance of these inoculants under field conditions.     

Occurrence and characterization of nitrogen-fixing Azospirilla in some soils of Egypt

The occurrence and characterization of N₂-fixing azospirilla in some Egyptian soils has been investigated. Seven soils, representing a wide range in texture and properties were selected from different localities in Egypt. The highest nitrogenase activity reported for soil samples under investigation were related to numbers of N₂-fixing microorganisms (Azotobacter spp., Azospirillum spp., and Clostridium spp.). Seven strains of azospirilla were isolated and purified. Based on morphological characteristics, three types of cell morphology were distinguished. Cultural and physiological characteristics as well as nitrogenase activity of representative isolates in presence of different concentrations of NaCl were determined. According to the physiological properties studied, all isolates were classified as members of Azospirillum brasilense.     

Nitrogen fixation by Azospirillium brasilense in soil and the rhizosphere under controlled environmental conditions

Summary Acetylene reduction activity by Azospirillum brasilense, either free-living in soils or associated with wheat roots, was determined in a sterilised root environment at controlled levels of O₂ tension and with different concentrations of mineral N. In an unplanted, inoculated soil nitrogenase activity remained low, at approximately 40 nmol C₂H₄ h^-1 per 2kg fresh soil, increasing to 300 nmol C₂H₄ h^-1 when malic acid was added as a C source via a dialyse tubing system. The N₂ fixation by A. brasilense in the rhizosphere of an actively growing plant was much less sensitive to the repressing influence of free O₂ than the free-living bacteria were. An optimum nitrogenase activity was observed at 10 kPa O₂, with a relatively high level of activity remaining even at an O₂ concentration of 20 kPa. Both NO _inf3 ^sup- and NH _inf4 ^sup+ repressed nitrogenase activity, which was less pronounced in the presence than in the absence of plants. The highest survival rates of inoculated A. brasilense and the highest rates of acetylene reduction were found in plants treated with azospirilli immediately after seedling emergence. Plants inoculated at a later stage of growth showed a lower bacterial density in the rhizosphere and, as a consequence, a lower N₂-fixing potential. Subsequent inoculations with A. brasilense during plant development did not increase root colonisation and did not stimulate the associated acetylene reduction. By using the ¹⁵N dilution method, the affect of inoculation with A. brasilense in terms of plant N was calculated as 0.067 mg N₂ fixed per plant, i.e., 3.3% of the N in the root and 1.6% in the plant shoot were of atmospheric origin. This ¹⁵N dilution was comparable to that seen in plants inoculated with non-N₂-fixing Psudomonas fluorescens.     

Nitrogenase activity (Acetylene reduction) during the growth cycle of spring barley (Hordeum vulgare L.)

The nitrogenase activity (C₂H₂-reduction) was measured during the growth cycle of field grown spring barley in soil cores both with and without barley plants, and at two levels of nitrogen application, 30 and 120 kg N ha^?1 year^?1 respectively. The main purpose of the investigation was to study the effects of the growing barley plants on nitrogenase activity in the soil, and temperature and moisture contents were kept constant in all experiments. Therefore, the results cannot be used to calculate actual amounts of fixed nitrogen in the field, but should be considered rather as potential values. The nitrogenase activity was found to vary during the growth cycle, and seemed to be correlated to the photosynthetic activity of the plants. Relatively low nitrogenase activity was found in the early growth stages, and the activity increased up to a maximum in the late reproductive stage, followed by a rapid decrease during the grain filling stage. The mean values of nitrogenase activity in samples without barley plants and with barley plants were 40 and 78 nmoles C₂H₄ g soil dwt^?1 24 h^?1 respectively. The positive effect of barley plants on nitrogenase activity was stronger at 120 kg N than at 30 kg N fertilization. As a mean of the whole growth cycle the ratio between samples with and without barley plants was 1.7 with 30 kgN and 2.3 with 120 kg N fertilization. The inhibitory effect of nitrogen application on nitrogenase activity was measurable until 6–7 weeks after application, and it was strongest in cores without plants.     

Influence of the photosynthesis-inhibiting herbicides goltix and sencor on growth and nitrogenase activity of Anabaena cylindrica and Nostoc muscorum

Summary The influence of the photosynthesis-inhibiting herbicides Goltix and Sencor on growth and nitrogenase activity of Anabaena cylindrica and Nostoc muscorum was studied. The cyanobacteria were grown under N₂-fixing photoautotrophic conditions. A. cylindrica was entirely inhibited even in the presence of low field concentrations (Sencor 10 ppm, Goltix 50 ppm). In contrast, Goltix (50 ppm and 100 ppm) and Sencor (10 ppm, 20 ppm, 50 ppm and 100 ppm) did not exert an inhibitory influence on growth and nitrogenase activity of N. muscorum. In the presence of normal field rates of the studied herbicides increased light intensities during incubation had no detrimental effect on N. muscorum. Only at 20 W/m² light intensity did the higher concentrations of Sencor (50 ppm and 100 ppm) cause partial inhibition for the first 10–12 days, which was compensated during subsequent incubation. The delayed inhibitory effects caused by higher concentrations of Goltix (200 ppm and 500 ppm) are supposed to be due to degradation product(s). In the natural environment other species of cyanobacteria may occur which are able to tolerate photosynthesis-inhibiting herbicides.     

Plant,Microbial and Soil Factors,Determining Nitrogen Fixation in the Rhizosphere

A genotype effect on associative (rhizosphere) N₂-fixation was observed with two cultivars of Sorghum bicolor (nutans) with a maximum rate of 8 μmol C₂H₄ · h^?1 · plant^?1 in one genotype compared to 0.9 μmol in the other. Characteristics of the high fixing genotype were a reduced transpiration rate, a lower number of stomata and increased root exudate production per gram root dry weight with higher concentration of dicarboxylic acids. The bacterial rhizosphere composition revealed a three times higher number of N₂-fixing bacteria, a tenfold reduction of actinomycetes and a threefold reduction of Arthrobacter associated with the high fixing cultivar compared to the low fixing genotype. From these and other plant rhizospheres two new nitrogen fixing bacteria, Pseudomonas stutzeri and Erwinia herbicola, were characterized. With the N₂-fixing bacteria Azospirillum brasilense and Klebsiella pneumoniae an enhancement of specific nitrogenase activity by aromatic compounds, for example phenolics, the herbicide alachlor and the insecticide carbofuran was demonstrated. An oscillating nitrogenase activity in Azospirillum brasilense under microaerobic conditions was found, resulting from an encystation and deencystation under those conditions. Experiments with wheat roots demonstrated that reduced oxygen tensions, essential for a maximum rhizosphere N₂-fixation, reduced root growth significantly and altered the N-metabolism of the roots.     

Soil mineral nitrogen concentration within cycles of flood irrigation: Effect of rice stubble and fertilization management

Two field experiments using macro (3.5 × 12m) plots and ¹⁵N-labelled fertilizer on micro (155 mm internal diameter) plots were undertaken to measure the effects of rice (Oryza sativa L.) stubble management and nitrogen fertilizer strategies on N transformations within a series of intermittent flood irrigations. Nitrate concentration fell by 90% during each flooding, and analysis of ¹⁵N micro plots showed the loss was due to denitrification rather than leaching. Over 52% of the ¹⁵N was lost. Apparent loss over four irrigations from macro plots receiving 60 kg urea-N ha⁻¹ was 37 kg N ha⁻¹ while unfertilized plots lost 19 kg N ha⁻. Stubble incorporation reduced nitrate accumulation rate and increased immobilization, thereby reducing denitrification losses by 23%. Nitrate concentration in the 0–100 mm soil layer increased after the soil water content fell below field capacity during the drying portion of each cycle, but the net nitrification rate fell with increasing number of cycles.Ammonium content in the top 100mm of soil fell from 35kgNha⁻¹ to 3kgNha⁻¹ over four irrigations. This fall was ascribed to the combined effects of nitrification and immobilization. Immobilization was greatest on plots where large quantities of rice stubble had been incorporated, and over 50% of the applied ¹⁵N was retained in the soil on these plots compared with 40% (SED = 3.5%) on plots where stubble had been burnt. We conclude that the poor response of rice to fertilization at sowing is due to a combination of denitrification and immobilization of applied nitrogen during cycles of wetting and drying prior to permanent flood.     

Diurnal and seasonal patterns of nitrogenase activity of red alder in comparison with white clover in silvopastoral agroforestry systems

 Simultaneous measurements were made to assess the diurnal and seasonal patterns of nitrogenase activity of red alder (Alnus rubra Bong.) and white clover (Trifolium repens L.) growing together in a silvopastoral agroforestry system using the acetylene reduction assay. Diurnal measurements were made in the summer and autumn at 3-h intervals whereas seasonal nitrogenase activity was assessed based on observations made at midday in July, September and January to represent the summer, autumn and winter seasons, respectively. No obvious diurnal patterns of nitrogenase activity were found in either red alder or white clover in summer and no significant variations in nitrogenase activity were observed between day and night. However, in autumn, pronounced diurnal patterns were observed in both species. Significantly higher rates of nitrogenase activity per unit dry weigh (dwt) of nodules were detected at 1500 hours in red alder, whereas, in white clover, significantly higher rates were obtained at 2100 hours. There was no significant correlation between diurnal nitrogenase activity and air temperature, photosynthetically active radiation and soil temperature at 10 cm depth in either red alder or white clover. Seasonal rates of nitrogenase activity showed significantly higher activity in summer, which subsequently decreased in autumn, to reach very low levels in the winter. The rates of nitrogenase activity of white clover were consistently higher than those of red alder both diurnally and seasonally. In the three seasons sampled, the average nitrogenase activity for white clover was 66.42 μmol C₂H₄ g dwt^–1 h^–1, which was 3.5 times higher than the 18.67 μmol C₂H₄ g dwt^–1 h^–1 obtained for red alder. Received: 11 November 1997     

Influence of tillage system on denitrification in maize-cropped soils

Denitrification losses show an irregular pattern through the year, often being caused by climatic conditions and management practices. The objectives of the present work were to quantify denitrification losses and to determine the influence of tillage system on the factors that control denitrification in fertilized soils. The modal profile of the soil was an Vertic Argiudoll, clay loam texture, located in Buenos Aires province, Argentina. The treatments were: (a) fertilized, (b) incorporated fertilization and (c) without fertilization for both no tillage and conventional tillage systems. Chambers were placed in the field to measure denitrification. In this clayish soil the estimated mean values of accumulated denitrification during the crop cycle (90 days) were 0.190kgNha^–1 for conventional tillage and 0.350kgNha^–1 for no tillage. In treatments with no tillage, losses by denitrification were approximately twice those of conventional tillage. These differences were also evidenced by the number of microorganisms, which were significantly higher (P<>;5%) for no tillage on all dates, except for at flowering. The increase at flowering coincided with the period of highest rainfall and consequently the highest water contents in the soil. The highest denitrification losses, except for sowing, were measured when soil moisture content was more than 30% (v/v). Denitrification increased in conjunction with an increase in the availability of carbon that is consumed by the heterotrophic microorganisms (including the denitrifiers). Received: 30 July 1996     

Estimating N2 fixation by Sesbania rostrata and S. cannabina (syn. S. aculeata) in lowland rice soil by the 15N dilution method

Summary A field experiment in concrete-based plots was conducted to estimate the contribution of N derived from air (Ndfa) or biological N₂ fixation in Sesbania rostrata and S. cannabina (syn. S. aculeata), using various references, by the ¹⁵N dilution method. The two Sesbania species as N₂-fixing reference plants and four aquatic weed species as non-N₂-fixing references were grown for 65 days after sowing in two consecutive crops, in the dry and the wet seasons, under flooded conditions. Soil previously labeled with ¹⁵N at 0.26 atom % ¹⁵N excess in mineralizable N was further labeled by ammonium sulfate with 3 and 6 atom % ¹⁵N excess. The results showed that ¹⁵N enrichment of soil NH ₄ ⁺ -N dropped exponentially in the first crop to half the original level in 50 days while in the second crop, it declined gradually to half the level in 130 days. The decline in ¹⁵N enrichment, in both N₂-fixing and non-fixing species, was also steeper in the first crop than in the second crop. Variations in ¹⁵N enrichment among non-fixing species were smaller in the second crop. The ratio of the uptake of soil N to that of fertilizer N in N₂-fixing and non-fixing species was estimated by the technique of varying the ¹⁵N level. In the second crop, this ratio in non-fixing species was higher than that in N₂-fixing species. Comparable estimates of % Ndfa were obtained by using ¹⁵N enrichment of various non-fixing species. There was also good agreement between the estimates obtained by using ¹⁵N enrichment of non-fixing species and those by using soil NH ₄ ⁺ -N, particularly in the second crop. By 25 days after sowing, the first crop of both Sesbania spp. had obtained 50% of total N from the atmosphere and the second crop had obtained 75%. The contribution from air increased with the age of the plant and ranged from 70% to 95% in 45–55 days. S. rostrata fixed substantially higher amounts of N₂ due to its higher biomass production compared with S. cannabina. Mathematical considerations in applying the ¹⁵N dilution method are discussed with reference to these results.     

Seasonal changes in periphyton nitrogen fixation in a protected tropical wetland

Cyanobacteria are important for global nitrogen cycle and often form complex associations referred to as cyanobacterial mats or periphyton that are common in tropical, limestone-based wetlands. The objective of this study was to monitor the nitrogen fixation rate using the acetylene reduction assay of these cyanobacterial mats in a tropical, unfertilized, and protected wetland. To account for temporal and spatial variation of nitrogenase activity, we were interested in seasons in a hydrological cycle (dry, rains, and end of rains), sites with different vascular vegetation, and rates of nitrogenase activity in a 24-h cycle. The annual average of nitrogenase activity was 22 nmol C₂H₄ cm⁻² h⁻¹, with a range of <6 to 35 nmol C₂H₄ cm⁻² h⁻¹, and the annual nitrogen fixation rate of our study site (9.0 g N m⁻² year⁻¹) is higher than similar estimates from other freshwater wetlands. There was a clear temporal pattern in nitrogenase activity with a maximum rate occurring during the rainy season (August) and a maximum nitrogenase activity occurring between 0600 and 1200 hours. We found spatial differences in nitrogenase activity among the four sites that could be attributed to variations in species composition within the periphyton.     

Effects of root-associated N2-fixing cyanobacteria on the growth and nitrogen content of wheat (Triticum vulgare L.) seedlings

Summary Following co-cultivation of wheat with N₂-fixing cyanobacterial isolates capable of forming associations, Nostoc 2S6B, 2S9B or Anabaena C5, for 15 days in the presence or absence of combined N a large stimulation of root length was observed without any increase in root dry weight. Increases in the N concentrations of both roots and shoots occurred following co-cultivation with most cyanobacteria tested. The increase in plant N concentrations appeared to be dependent on the wheat cultivar and the cyanobacterial isolate used. Nostoc isolates had similar nitrogenase activities when associated with roots and when grown in shake-flask cultures. The nitrogenase activity of roots colonized by Anabaena C5 or Nostoc 2S6B was higher following removal of loosely associated cyanobacteria.     

Soil fertility effects on growth,yield, nodulation and nitrogenase activity of Austrian winter pea

Austrian winter pea (Pisum sativum subspecies arvense (L.) Poir) is grown as a cool season annual to produce high protein seed and forage as well as for soil fertility improvement. This legume is grown on a wide range of soil types with many different cropping systems. The objective of these studies was to determine the influence of K levels, with and without P and Ca fertilization, for increased growth, yield, nodulation and nitrogenase activity. Results were from 3 years’ field and greenhouse experiments with a Psammentic Paleustalf (Eufaula series) utilizing Rhizobium leguminosarum (Frank), ATCC 10314 as inoculum. Soil fertility effects on composition and histology of field‐grown nodules are presented.

Available soil P was a limiting plant nutrient in field studies with significant response to K resulting with PK combinations for top growth, tillers, pods, seed yield, nodule mass, and nitrogenase activity levels (C₂H₂, red.). Multiple regression for nitrogenase (umol C₂H₄ h^‐1) = 1.09 tiller number + 3.37 nodule weight + 2.29 pod number, R² = 0.837, C.V. = 29.9%. Results from the greenhouse experiments indicated significant responses with increased K application levels when combined with P and Ca fertilization for top growth, nodule weight, number of nodules and nitro‐genase activity. Highly significant correlations resulted with nitrogenase x nodule weight (r=0.538) and nitrogenase x top growth (r=0.359) with multiple regression of treatment effects for nitrogenase (μmol C₂H₄ h^‐1) = 2.73 P + 1.04 K + 4.92 Ca, R² = 0.797 and C.V. = 48.8%. Soil addition of plant nutrients resulted in significantly increased concentrations of those elements within nodules. Magnesium content was not consistently influenced by P, Ca, and K amendments. Sodium decreased with increased K fertilization. Multiple regression of elemental composition (mg g^‐1 nodule) for nitrogenase (pmol C₂H₄ h^‐1) = 0.21 P + 0.86 K + 2.35 Ca ‐ 2.01 Na, R² = 0.772, C.V. = 55.6%. The proportion of plant nutrients in nodules contained within the nodule cytosol was highest for K (56.2%) and lowest for Ca (21.4%) with intermediate levels of Mg (50.2%), P (45.4%), and Na (37.2%).

Practical application from these data include the requirement of adequate available soil K for increased yield and nitrogen fixation with favorable P and Ca soil levels in Austrian winter pea production.     

The 15N-isotope dilution technique applied to the estimation of biological nitrogen fixation associated with Paspalum notatum cv. batatais in the field

The contribution of associated biological nitrogen fixation to the nitrogen nutrition of Paspaulum notatum cv. batatais was estimated using the ¹⁵N-isotope dilution technique. The plants were grown in the field in concrete cylinders (60 cm dia) filled with soil, to which were added small quantities of ¹⁵N-labelled fertilizer at frequent intervals over 12 months. The pensacola cultivar of P. notatum was used as a non-N₂-fixing control plant. This was justified by the observation that nitrogenase (intact core C₂H₂ reduction) activity associated with the pensacola cultivar was consistently much lower than that of the batatais cultivar.At the first harvest, no evidence for N₂ fixation associated with the batatais cultivar was obtained, probably because of slow establishment of the N₂-fixing association. However, at the subsequent three harvests the batatais cultivar exhibited a lower ¹⁵N-enrichment and yielded more N than the pensacola cultivar. These data together suggested that 8–25% of the N in the batatais cultivar originated from N₂ fixation.The grass Paspalum maritimum was also included in the experiment and exhibited low nitrogenase activity similar to that of the pensacola cultivar of P. notatum. However, the total N and ¹⁵N data of these two grasses were not in good agreement indicating that it is important for the use of the isotope dilution technique that control plants are of very similar physiology and growth habit.     

Indications for passive rather than active release of natural nitrification inhibitors in Brachiaria humidicola root exudates

Plants have the ability to suppress microbial nitrification process through secondary metabolites released from their root exudates or/and leaf litter. For decades, grasses were suggested to control nitrification process, and recently, Brachiaria humidicola accession 26159 (BH) as a tropical and subtropical grass has been shown to reduce nitrification rates under laboratory and soil conditions. In this study, experiments were conducted under controlled conditions in nutrient solution culture to investigate whether the reported release of natural nitrification inhibitors from root exudates of BH is an active or passive phenomenon. So different variables such as N-form (nitrate vs. ammonium), collecting medium (distilled water vs. 1 mM NH₄Cl) and collecting period (6 vs. 24 hrs) were included to study the hypothesis. Results showed when root exudates were collected in distilled water there was no nitrification inhibition activity for all ammonium and nitrate grown plants. However, when collection was done in a medium containing 1 mM NH₄Cl, root exudates showed significant nitrification inhibition activity similar to results obtained by Subbarao et al. The observed nitrification inhibition activity had a positive correlation to ammonium treatment particularly in collection medium, probably due to root cells damage induced by low pH and membrane depolarization under ammonium nutrition. This was more supported by application of shoot homogenates of NH₄⁺, NO₃^? or NH₄NO₃ grown plants that showed significant nitrification inhibition activity compared to distilled water and DMPP controls in a bioassay test, independent of N-form. Potassium concentrations in root exudates (as a result of potassium leakage) were found to increase in root washings of plants, which were grown with ammonium, particularly when root exudates were collected in 1 mM NH₄Cl solution. In addition, higher electric conductivity of root washings after collection of root exudates in ammonium containing medium (low pH) and also in nitrate containing medium which adjusted to pH 3 by applying H₂SO₄, strongly suggest that release of natural nitrification inhibitors from root exudates of B. humidicola may not be an active process, but instead it is rather a passive phenomenon by ammonium induced root physicochemical damages.     

Foliar application of molybdenum in common bean. II. Nitrogenase and nitrate reductase activities in a soil of low fertility

Foliar application of molybdenum (Mo) at 40 g ha^‐1 25 days after plant emergence greatly enhanced nitrogenase and nitrate reductase activities of common bean (Phaseolus vulgaris L.), resulting in an increase in total nitrogen (N) accumulation in shoots. Application of 20 kg N ha^‐1 as ammonium sulfate [(NH₄)₂SO₄] at sowing decreased nodulation and nitrogenase activity. Rhizobium inoculation did not affect nitrogenase activity which demonstrated that Mo application increased the efficiency of native Rhizobia strains. Nitrogen amendment, either at planting (20 kg N ha^‐1) or as a side dressing (30 kg N ha^‐1) 25 days after plant emergence, did not affect the foliar nitrate reductase activity. Molybdenum foliar spray as ammonium molybdate [(NH₄)₆Mo₇O₂₄2H₂O] and N applied as a side dressing increased equally the total amount of N in the pods. A 10% increase in the seed N concentration was obtained with foliar application of Mo, while N applied as a side dressing had no effect on seed N concentration. An average increase of 41% in N export to the seeds was obtained by either Mo or N as side dressing. Nitrogen applied at sowing or Rhizobia inoculation had no effect on the characteristics evaluated 74 days after plant emergence. Plants that received either Mo as foliar spray or as side dressed N had similar yields. This demonstrated that, in certain soils, N fertilization may be replaced by a small amount of Mo as a foliar application.