Enhancing the early performance of the leguminous shrub Retama sphaerocarpa (L.) Boiss.: fertilisation versus Rhizobium inoculation

We have investigated the effect on growth of fertilisation versus biological nitrogen fixation by rhizobial nodules in Retama sphaerocarpa(L.) Boiss, a leafless leguminous shrub native to the Iberian Peninsula and North-West Africa that has generated interest for revegation of dry Mediterranean habitats. Our main objective was to optimise the formation of root nodules under nursery conditions and to evaluate their influence on the first year of seedling growth in comparison with standard fertilisation. Seedlings of R. sphaerocarpa from two Spanish localities were grown under two levels of fertilisation, and half of each were inoculated with rhizobia isolated from adult Retama, Cytisus and Adenocarpusplants in the field. Although some promiscuity was observed, nodulation was significantly successful with specific rhizobia. At the end of the experiment, highly fertilised plants were taller and heavier and exhibited larger photosynthetic rates than either nodulated or non-nodulated plants under low fertilisation. High fertilisation enhanced seedling growth but inhibited both the nodulation and the nitrogenase activity of the nodules. Thus, physiological differences between nodulated and non-nodulated plants were observed in the low but not in the high fertilisation treatment. Nitrogen uptake and use was enhanced by root nodules, which translated into enhanced photosynthesis and growth. Since inoculation is simple, environmentally friendly and cheap, and nodulated plants are more likely to overcome transplant stress than non-nodulated ones, our results suggest that inoculation together with low, background fertilisation (instead of high fertilisation) should be used when producing high quality seedlings of this autochthonous Mediterranean shrub.     

Two effects of combined nitrogen on the adhesion of Rhizobium etli to bean roots

Combined forms of nitrogen negatively influence rhizobia-legume symbiosis. The effects of combined nitrogen are known for nodulation and dinitrogen (N₂) fixation, but little is known about the effect on preinfection events. Here, we studied the effects of combined nitrogen on the adhesion of Rhizobium etli to common bean (Phaseolus vulgaris L.) roots. When potassium nitrate (KNO₃) or sodium glutamate was added to an incubation mixture of rhizobia and plants that were previously grown in nitrogen-free solution, rhizobial adhesion to roots was stimulated. However, the rhizobial adhesion to bean roots that were previously grown with 10 mM KNO₃ was reduced by half. A fraction of the bean root exudates, which is thermolabile and has molecular mass larger than 12 kDa stimulated rhizobial adhesion, but this stimulatory activity was lost in root exudates obtained with 10 mM KNO₃. Thus, the inhibition of symbiosis in response to combined nitrogen may be controlled by the plant at the preinfection stage as well.     

Saprophytic Actinomycetes Promote Nodulation in <Emphasis Type="Italic">Medicago sativa</Emphasis>-<Emphasis Type="Italic">Sinorhizobium meliloti</Emphasis> Symbiosis in the Presence of High N

Saprophytic rhizoactinomycetes isolated from the root nodule surface of the nitrogen-fixing actinorhizal plant Discaria trinervis, Streptomyces MM40, Actinoplanes ME3, and Micromonospora MM18, previously shown to stimulate nodulation in Frankia-Discaria trinervis symbiosis, were assayed as co-inoculants with Sinorhizobium meliloti 2011 on Medicago sativa. When plants were fertilized with a low level of N (0.07 mM), the inoculation of the actinomycetes alone did not show any effect on plant growth. Meanwhile, when actinomycetes were co-inoculated with S. meliloti, nodulation and plant growth were significantly stimulated compared to plants inoculated with only S. meliloti. The analysis of nodulation kinetics of simultaneously or delayed co-inoculations suggests that the effect of the actinomycetes operates in early infection and nodule development counteracting the autoregulation of nodulation by the plant. Because the actinomycete effect was found in the symbiotic nitrogen-fixing state of the plant, we investigated the effects of the actinomycetes, in single inoculation or co-inoculation with S. meliloti, on plants grown under a high level of N (7 mM) that was inhibitory for nodulation by S. meliloti. The inoculation of the actinomycetes alone did not show any effect on plant growth although high N was available. Unexpectedly, the co-inoculation of actinomycetes with S. meliloti on plants grown with high N (7 mM) significantly stimulates nodulation, clearly counteracting the inhibition of nodulation by high N. These results corroborate that the interaction of rhizoactinomycetes would interfere with the autoregulation of nodulation in alfalfa mediated by high N, opening new research lines of potential agronomical applications.     

The Biological Activity of the Sinorhizobium meliloti Glucan

The study of the effect of periplasmic glucan isolated from the root-nodule bacterium Sinorhizobium meliloti CXM1-188 on the symbiosis of another strain (441) of the same root-nodule bacterium with alfalfa plants showed that this effect depends on the treatment procedure. The pretreatment of alfalfa seedlings with glucan followed by their bacterization with S. meliloti 441 insignificantly influenced the nodulation parameters of symbiosis (the number of root nodules and their nitrogen-fixing activity) but induced a statistically significant increase in the efficiency of symbiosis (expressed as the masses of the alfalfa overground parts and roots). At the same time, the pretreatment of S. meliloti 441 cells with glucan brought about a considerable decrease in the nodulation parameters of symbiosis (the number of root nodules and their nitrogen-fixing activity decreased by 2.5–11 and 7 times, respectively). These data suggest that the stimulating effect of rhizobia on host plants may be due not only to symbiotrophic nitrogen fixation but also to other factors. Depending on the experimental conditions, the treatment of alfalfa plants with glucan and their bacterization with rhizobial cells enhanced the activity of peroxidase in the alfalfa roots and leaves by 10–39 and 12–27%, respectively.     

Agrobacterium rhizogenes transformed soybean roots differ in their nodulation and nitrogen fixation response to genistein and salt stress

We evaluated response differences of normal and transformed (so-called ‘hairy’) roots of soybean (Glycine max L. (Merr.), cv L17) to the Nod-factor inducing isoflavone genistein and salinity by quantifying growth, nodulation, nitrogen fixation and biochemical changes. Composite soybean plants were generated using Agrobacterium rhizogenes-mediated transformation of non-nodulating mutant nod139 (GmNFR5α minus) with complementing A. rhizogenes K599 carrying the wild-type GmNFR5α gene under control of the constitutive CaMV 35S promoter. We used genetic complementation for nodulation ability as only nodulated roots were scored. After hairy root emergence, primary roots were removed and composite plants were inoculated with Bradyrhizobium japonicum (strain CB1809) pre-induced with 10 μM genistein and watered with NaCl (0, 25, 50 and 100 mM). There were significant differences between hairy roots and natural roots in their responses to salt stress and genistein application. In addition, there were noticeable nodulation and nitrogen fixation differences. Composite plants had better growth, more root volume and chlorophyll as well as more nodules and higher nitrogenase activity (acetylene reduction) compared with natural roots. Decreased lipid peroxidation, proline accumulation and catalase/peroxidase activities were found in ‘hairy’ roots under salinity stress. Genistein significantly increased nodulation and nitrogen fixation and improved roots and shoot growth. Although genistein alleviated lipid peroxidation under salinity stress, it had no significant effect on the activity of antioxidant enzymes. In general, composite plants were more competitive in growth, nodulation and nitrogen fixation than normal non-transgenic even under salinity stress conditions.     

Effects of calcium in the nitrogen-fixing symbiosis between actinorhizal <Emphasis Type="Italic">Discaria trinervis</Emphasis> (Rhamnaceae) and <Emphasis Type="Italic">Frankia</Emphasis>

We have analysed the growth and symbiotic performance of actinorhizal Discaria trinervis at various Ca supply regimes. We aimed at discriminating between specific, if any, effects on nodulation and general growth stimulation by Ca. The hypothesis that a high Ca supply would interfere with nodulation by Frankia was also tested. Results showed that plant growth increased with Ca supply. Nodulation was stimulated by moderate levels of Ca, but inhibited by Ca higher than 0.77 mM. Growth of nodules was less affected by Ca than shoot and root growth. Ca concentration of symbiotic plants increased with Ca supply, but nitrogen concentration was independent of Ca at concentrations which did not impair plant growth. All together, these results show that Ca has a positive effect on the establishment and functioning of the symbiosis between Discaria trinervis and Frankia. However, the positive influence of Ca was more likely due to a promotion of plant growth rather than a direct effect on nodule growth and nitrogen fixation itself. At high levels of Ca supply nodulation was impaired. Given the intercellular infection pathway in Discaria trinervis, we suggest that the increment of Ca availability would strengthen its root cell walls, thus decreasing Frankia penetration of the root.     

The biological activity of the Sinorhizobium meliloti glucan

The study of the effect of the periplasmic glucan isolated from the root-nodule bacterium S. meliloti CXM1-188 on the symbiosis of another strain (441) of the same root-nodule bacterium with alfalfa plants showed that this effect depends on the treatment procedure. The pretreatment of alfalfa seedlings with the glucan followed by their bacterization with S. meliloti 441 insignificantly influenced the nodulation parameters of symbiosis (the number of root nodules and their nitrogen-fixing activity) but induced a statistically significant increase in the efficiency of symbiosis (expressed as the masses of the alfalfa overground parts and roots). At the same time, the pretreatment of S. meliloti 441 cells with the glucan brought about a considerable decrease in the nodulation parameters of symbiosis (the number of the root nodules and their nitrogen-fixing activity decreased by 2.5-11 and 7 times, respectively). These data suggest that the stimulating effect of rhizobia on host plants may be due not only to symbiotrophic nitrogen fixation but also to other factors. Depending on the experimental conditions, the treatment of alfalfa plants with the glucan and their bacterization with rhizobial cells enhanced the activity of peroxidase in the alfalfa roots and leaves by 10-39 and 12-27%, respectively.     

Charcoal and shrubs modify soil processes in ponderosa pine forests of western Montana

In split-root systems of alfalfa (Medicago sativa L.), already existing nodules or arbuscular mycorrhizal roots suppress further establishment of symbiosis in other root parts, a phenomenon named autoregulation. Roots treated with rhizobial nodulation signals (Nod factors) induce a similar systemic suppression of symbiosis.In order to test the hypothesis that flavonoids play a role in this systemic suppression, split-root systems of alfalfa plants were inoculated on one side of the split-root system with Sinorhizobium meliloti or Glomus mosseae or were treated with Nod factor. HPLC-analysis of alfalfa root extracts from both sides of the split-root system revealed a persistent local and systemic accumulation pattern of some flavonoids associated with the different treatments. The two flavonoids, formononetin and ononin, could be identified to be similarily altered after rhizobial or mycorrhizal inoculation or when treated with Nod factor.Exogenous application of formononetin and ononin partially restored nodulation and mycorrhization pointing towards the involvement of these two secondary compounds in the autoregulation of both symbioses.     

Effects of nickel on Frankia and its symbiosis with Alnus glutinosa (L.) Gaertn

The tolerance of nickel by Frankia in culture and in symbiosis with Alnus was determined. Yield of three Frankia strains was not affected significantly by 2.25 mM nickel when cultured in propionate medium containing hydolysed casein as nitrogen source. Yield of two strains in medium without combined nitrogen, and thus reliant on fixed nitrogen, was stimulated markedly by the same nickel concentration. Utilisation of nickel for synthesis of uptake hydrogenases is presumed to be the cause of enhanced nitrogenase activity.Although growth was reduced, treatment of 2-month-old seedlings with 0.025 mM nickel for 4 weeks did not affect nodulation significantly while nitrogenase activity was doubled. Nodulation and nitrogenase activity of seedlings receiving 0.075 mM nickel were inhibited markedly, while 0.5 mM nickel was lethal to all seedlings after 4 weeks of treatment. A few small, ineffective nodules were initiated early on some of the latter seedlings, suggesting that effects of nickel on host plant processes rather than Frankia are the primary cause of inhibition of nodulation. This interpretation is supported by the retention of substantial nitrogenase activity in 10-month-old plants 1 day after the treatment with 0.59 mM nickel, when the nickel content of roots and nodules was already maximal. No nitrogenase activity was detected after 3 days, by which time the leaves were almost completely necrotic. Over a 4 day period, most nickel was retained in the roots and nodules. Supplying histidine simultaneously at concentrations equal to, or in excess of, nickel prevented wilting and leaf necrosis, but did not increase translocation of nickel to the shoot.     

The Sesbania Root Symbionts Sinorhizobium saheli and S. teranga bv. sesbaniae Can Form Stem Nodules on Sesbania rostrata, although They Are Less Adapted to Stem Nodulation than Azorhizobium caulinodans

Sesbania species can establish symbiotic interactions with rhizobia from two taxonomically distant genera, including the Sesbania rostrata stem-nodulating Azorhizobium sp. and Azorhizobium caulinodans and the newly described Sinorhizobium saheli and Sinorhizobium teranga bv. sesbaniae, isolated from the roots of various Sesbania species. A collection of strains from both groups were analyzed for their symbiotic properties with different Sesbania species. S. saheli and S. teranga bv. sesbaniae strains were found to effectively stem nodulate Sesbania rostrata, showing that stem nodulation is not restricted to Azorhizobium. Sinorhizobia and azorhizobia, however, exhibited clear differences in other aspects of symbiosis. Unlike Azorhizobium, S. teranga bv. sesbaniae and S. saheli did not induce effective stem nodules on plants previously inoculated on the roots, although stem nodulation was arrested at different stages. For Sesbania rostrata root nodulation, Sinorhizobium appeared more sensitive than Azorhizobium to the presence of combined nitrogen. S. saheli and S. teranga bv. sesbaniae were effective symbionts with all Sesbania species tested, while Azorhizobium strains fixed nitrogen only in symbiosis with Sesbania rostrata. In a simple screening test, S. saheli and S. teranga bv. sesbaniae were incapable of asymbiotic nitrogenase activity. Thus, Azorhizobium can easily be distinguished from Sinorhizobium among Sesbania symbionts on the basis of symbiotic and free-living nitrogen fixation. The ability of Azorhizobium to overcome the systemic plant control appears to be a stem adaptation function. This last property, together with its host-specific symbiotic nitrogen fixation, makes Azorhizobium highly specialized for stem nodulation of the aquatic legume Sesbania rostrata.     

Soil moisture and potassium affect the performance of symbiotic nitrogen fixation in faba bean and common bean

Potassium (K) is reported to improve plant's resistance against environmental stress. A frequently experienced stress for plants in the tropics is water shortage. It is not known if sufficient K supply would help plants to partially overcome the effects of water stress, especially that of symbiotic nitrogen fixation which is often rather low in the tropics when compared to that of temperate regions. Thus, the impact of three levels of fertilizer potassium (0.1, 0.8 and 3.0 mM K) on symbiotic nitrogen fixation was evaluated with two legumes under high (field capacity to 25% depletion) and low (less than 50% of field capacity) water regimes. Plants were grown in single pots in silica sand under controlled conditions with 1.5 mM N (¹⁵N enriched NH₄NO₃). The species were faba bean (Vicia faba L.), a temperate, amide producing legume and common bean (Phaseolus vulgaris L.), a tropical, ureide producing species. In both species, 0.1 mM K was insufficient for nodulation at both moisture regimes, although plant growth was observed. The supply of 0.8 or 3.0 mM K allowed nodulation and subsequent nitrogen fixation which appeared to be adequate for respective plant growth. High potassium supply had a positive effect on nitrogen fixation, on shoot and root growth and on water potential in both water regimes. Where nodulation occurred, variations caused by either K or water supply had no consequences on the percentage of nitrogen derived from the symbiosis. The present data indicate that K can apparently alleviate water shortage to a certain extent. Moreover it is shown that the symbiotic system in both faba bean and common bean is less tolerant to limiting K supply than plants themselves. However, as long as nodulation occurs, N assimilation from the symbiotic source is not selectively affected by K as opposed to N assimilation from fertilizer.     

氮肥和水分条件对长喙田菁生长、结瘤和固氮的影响

 盆栽试验研究了长喙田菁(Sesbania rostrata) 茎瘤固氮根瘤菌（Azorhizobium caulinodans）共生体系在不同水分和无机氮肥条件下的生长、结瘤和氮积累。水分处理为: 不浸水、浅浸水(土壤水分为田间持水量)和深浸水, 氮肥处理分别为每千克土施加无机氮肥10 mg、20 mg和40 mg。结果表明,水分条件对共生体系有较大影响, 浸水环境虽然抑制了根瘤的形成, 却促进了茎瘤的结瘤作用和长喙田菁根系的生长, 田间持水量状态下共生体系能获得最大的生物量和氮累积量; 无机氮肥对共生体系的影响受水分供应的影响较大,实验氮肥施用浓度范围内,长喙田菁的结瘤作用对复合态氮无负敏感反应,明显有别于一般根瘤体系的复合态氮反应。     

Proline accumulation has prevalence over polyamines in nodules of Medicago sativa in symbiosis with Sinorhizobium meliloti during the initial response to salinity

Background and aims

Polyamines are cationic molecules that play an important role in the plant response to environmental stresses. The aim of this work is to determine the role of these compounds in the response to salinity of Medicago sativa plants in symbiosis with the soil bacteria Sinorhizobium meliloti.

Methods

M. sativa plants inoculated with S. meliloti were subjected to 100 and 150 mM NaCl treatments. The concentration of nodular polyamines was determined in relation to the nitrogen fixation parameters, proline accumulation, and oxidative damage. In addition, polyamines concentrations were analyzed in different nodular fractions as well as the effect of exogenous polyamines in the nodulation response.

Results

The concentration of nodular polyamines decreased by the salinity in correlation with the nitrogenase activity after 2 and 4 weeks of salt treatment while spermine accumulated after 6 weeks. On the contrary, proline accumulation was induced by the salinity at all time points. The analysis of different nodular fractions showed the highest polyamines concentration in bacteroids being homospermidine the most abundant.

Conclusion

Proline accumulation had prevalence over polyamines at the earliest response to salinity probably due to nitrogen limitation under salt stress conditions and the existence of a common precursor for both compounds in the nodule. Nevertheless, after long salt exposure, spermine was also accumulated. The analysis of different nodular fractions indicated the bacteroidal origin of polyamines in nodules being homoespermidine, one of the most abundant.     

Effects of plant quality and ant defence on herbivory rates in a neotropical ant‐plant

1. Understanding the degree to which populations and communities are limited by both bottom‐up and top‐down effects is still a major challenge for ecologists, and manipulation of plant quality, for example, can alter herbivory rates in plants. In addition, biotic defence by ants can directly influence the populations of herbivores, as demonstrated by increased rates of herbivory or increased herbivore density after ant exclusion. The aim of this study was to evaluate bottom‐up and top‐down effects on herbivory rates in a mutualistic ant‐plant. 2. In this study, the role of Azteca alfari ants as biotic defence in individuals of Cecropia pachystachya was investigated experimentally with a simultaneous manipulation of both bottom‐up (fertilisation) and top‐down (ant exclusion) factors. Four treatments were used in a fully factorial design, with 15 replicates for each treatment: (i) control plants, without manipulation; (ii) fertilised plants, ants not manipulated; (iii) unfertilised plants and excluded ants and (iv) fertilised plants and ants excluded. 3. Fertilisation increased the availability of foliar nitrogen in C. pachystachya, and herbivory rates by chewing insects were significantly higher in fertilised plants with ants excluded. 4. Herbivory, however, was more influenced by bottom‐up effects – such as the quality of the host plant – than by top‐down effects caused by ants as biotic defences, reinforcing the crucial role of leaf nutritional quality for herbivory levels experienced by plants. Conditionality in ant defence under increased nutritional quality of leaves through fertilisation might explain increased levels of herbivory in plants with higher leaf nitrogen.     

Effects of Zn<Superscript>2+</Superscript> on nodulation and growth of a South American actinorhizal plant, <Emphasis Type="Italic">Discaria americana</Emphasis> (Rhamnaceae)

Discaria americana is a xerophytic shrub which lives in symbiosis with an actinomycete of the genus Frankia. The objective of this paper was to investigate the effects of high soil Zn²⁺ concentrations on growth and nodulation on the association Discaria americana–Frankia with the aim of determining if this association is suitable for improving contaminated soils. Two experiments were performed in 1 dm³ pots containing soil and different Zn additions, from 0 to 2,000 mg Zn²⁺ kg⁻¹ dry soil, with or without N fertilization. Zn additions strongly delayed shoot and root growth, but once growth was initiated, the biomass production of the plants supplied with moderate Zn amounts did not differ from the control plants. Zn reduced the final nodule number, but not the total nodule biomass. At the end of the experiment only the highest Zn treatments showed a lower nodule weight than the control plants, while N addition completely inhibited nodulation. It is concluded than Zn reduces the number of Frankia infections, but once the actinomycete is inside the roots, nodules can continue growing according to plant demand for N, compensating the reduced nodule number with more biomass. On the other hand, there is a toxic effect of Zn itself on plants when present in very high concentrations.     

Soil nematodes mediate positive interactions between legume plants and rhizobium bacteria

Symbiosis between legume species and rhizobia results in the sequestration of atmospheric nitrogen into ammonium, and the early mechanisms involved in this symbiosis have become a model for plant-microbe interactions and thus highly amenable for agricultural applications. The working model for this interaction states that the symbiosis is the outcome of a chemical/molecular dialogue initiated by flavonoids produced by the roots of legumes and released into the soil as exudates, which specifically induce the synthesis of nodulation factors in rhizobia that initiate the nodulation process. Here, we argue that other organisms, such as the soil nematode Caenorhabditis elegans, also mediate the interaction between roots and rhizobia in a positive way, leading to nodulation. We report that C. elegans transfers the rhizobium species Sinorhizobium meliloti to the roots of the legume Medicago truncatula in response to plant-released volatiles that attract the nematode. These findings reveal a biologically-relevant and largely unknown interaction in the rhizosphere that is multitrophic and may control the initiation of the symbiosis.     

5-Aminolevulinic acid improves photosynthetic gas exchange capacity and ion uptake under salinity stress in oilseed rape (Brassica napus L.)

Salinity is one of the major constraints in oilseed rape (Brassica napus L.) production. One of the means to overcome this constraint is the use of plant growth regulators to induce plant tolerance. To study the plant response to salinity in combination with a growth regulator, 5-aminolevulinic acid (ALA), oilseed rape plants were grown hydroponically in greenhouse conditions under three levels of salinity (0, 100, and 200 mM NaCl) and foliar application of ALA (30 mg/l). Salinity depressed the growth of shoots and roots, and decreased leaf water potential and chlorophyll concentration. Addition of ALA partially improved the growth of shoots and roots, and increased the leaf chlorophyll concentrations of stressed plants. Foliar application of ALA also maintained leaf water potential of plants growing in 100 mM salinity at the same level as that of the control plants, and there was also an improvement in the water relations of ALA-treated plants growing in 200 mM. Net photosynthetic rate and gas exchange parameters were also reduced significantly with increasing salinity; these effects were partially reversed upon foliar application with ALA. Sodium accumulation increased with increasing NaCl concentration which induced a complex response in the macro-and micronutrients uptake and accumulation in both roots and leaves. Generally, analyses of macro- (N, P, K, S, Ca, and Mg) and micronutrients (Mn, Zn, Fe, and Cu) showed no increased accumulation of these ions in the leaves and roots (on dry weight basis) under increasing salinity except for zinc (Zn). Foliar application of ALA enhanced the concentrations of all nutrients other than Mn and Cu. These results suggest that under short-term salinity-induced stress (10 days), exogenous application of ALA helped the plants improve growth, photosynthetic gas exchange capacity, water potential, chlorophyll content, and mineral nutrition by manipulating the uptake of Na⁺.     

The ethylene-inhibitor aminoethoxyvinylglycine restores normal nodulation by Rhizobium leguminosarum biovar. viciae on Vicia sativa subsp. nigra by suppressing the ‘Thick and short roots’ phenotype

Nodulation of Vicia sativa subsp. nigra L. by Rhizobium bacteria is coupled to the development of thick and short roots (Tsr). This root phenotype as well as root-hair induction (Hai) and root-hair deformation (Had) are caused by a factor(s) produced by the bacteria in response to plant flavonoids. When very low inoculum concentrations (0.5–5 bacteria·ml^-1) were used, V. sativa plants did not develop the Tsr phenotype and became nodulated earlier than plants with Tsr roots. Furthermore, the nodules of these plants were located on the primary root in contrast to nodules on Tsr roots, which were all located at sites of lateral-root emergence. The average numbers of nodules per plant were not significantly different for these two types of nodulation. Root-growth inhibition and Hai, but not Had, could be mimicked by ethephon, and inhibited by aminoethoxyvinylglycine (AVG). Addition of AVG to co-cultures of Vicia sativa and the standard inoculum concentration of 5·10⁵ bacteria·ml^-1 suppressed the development of the Tsr phenotype and restored nodulation to the pattern that was observed with very low concentrations of bacteria (0.5–5 bacteria·ml^-1). The delay in nodulation on Tsr roots appeared to be caused by the fact that nodule meristems did not develop on the primary root, but only on the emerging laterals. The relationship between Tsr, Hai, Had, and nodulation is discussed.Abbreviations AVG aminoethoxyvinylglycine - cfu colonyforming units - Had root-hair deformation - Hai root-hair induction - NB naringenin-bacteria filtrate - Tsr Thick and short roots     

Functional Activity of Exoglycans from Rhizobium leguminosarum bv. viciae 250a and Its Nitrogen-Resistant Mutant M-71 during the Formation of Legume–Rhizobia Symbiosis against a High-Nitrogen Background

The functional activity of the exoglycan complex (EGC) polysaccharides from Rhizobium leguminosarum bv. viciae 250a and its nitrogen-resistant mutant M-71, capable of inducing the formation of nitrogen-fixing nodules on pea roots against a high-nitrogen background (4.8 mM NO₃ ^–), was studied in vegetation tests. For this purpose, the bacterial inoculum washed free of its own exoglycans was supplemented with EGC of the same or another strain grown in the presence of 6 or 20 mM nitrate. The best symbiotic characteristics (nodule number and nitrogenase activity, mass of the roots and aerial parts of plants) were recorded when the inoculum cells and exoglycans were obtained from strain M-71 grown in the presence of 20 mM nitrate. When the plants were inoculated with the cells (grown at 6 mM nitrate) + EGC (obtained at 6 mM nitrate) of this strain, the nodulation characteristics and the effectiveness of symbiosis decreased 1.5- to 2-fold. Partial recovery of the symbiotic potential of strain M-71 was observed when EGC (obtained at 20 mM nitrate) was substituted for its exoglycans (obtained at 6 mM nitrate). In the presence of exoglycans of the parent strain 250a (obtained at 6 or 20 mM nitrate), the mutant formed a substantially lesser number of nodules with a very low nitrogen-fixing activity. In turn, the mutant exoglycans synthesized in medium with either high or low nitrate nitrogen concentration did not recover the fix⁺ phenotype of strain 250a, capable of forming symbiosis with pea plants only against a low-nitrogen background. In study of the relative content of high-molecular-weight exopolysaccharide components and low-molecular-weight glycans in the exoglycan complex, it was established that, in strain 250a (grown at 6 and 20 mM nitrate), as well as in its mutant M-71 (grown at 6 mM nitrate), exopolysaccharides prevailed, accounting for 72–75% of the sum of both types of glycopolymers, while low-molecular-weight glycans accounted for 25–28%. In contrast, in the EGC of strain M-71 obtained at 20 mM nitrate, which was the most active inducer of the formation of the symbiotrophic system by strain M-71 in the presence of a high mineral nitrogen concentration, low-molecular-weight glycans were the main component, accounting for 61% of total glycopolymers, while the polysaccharide content was 39%. Low-molecular-weight exoglycans are supposed to be involved in maintaining the physiological activity and the symbiotic status of rhizobia under unfavorable environmental conditions.     

Ecological roles of arbuscular mycorrhizal fungi in two wild legume plants

Two wild legume plants,Glycine soja andCassia mimosoides var.nomame, and a cultivated plant, soybean (Glycine max), were employed for a study of triple symbiosis with an inoculum ofScutellispora heterogama harvested from natural soils and an inoculum of their own rhizobial cells. The dry weight, colonization of arbuscular mycorrhizal fungus, nodule formation and N₂-fixation activity were estimated as the parameters of triple symbiosis. The two wild legume plants showed greater growth with colonization of arbuscular mycorrhizae than with nodulation, whereas the cultivated legume showed more nodulation than colonization of arbuscular mycorrhizae. Moreover,S. heterogama appeared to stimulate the triple symbiosis for the wild legume plants. The results suggested that spores ofS. heterogama are important in disturbed soils in Korea.