Characteristics of rhizobia isolated from three legumes indigenous to the Canadian high arctic:Astragalus alpinus,Oxytropis maydelliana,andOxytropis arctobia

Summary Forty-eight strains of rhizobia were isolated from the root nodules ofAstragalus alpinus (21),Oxytropis maydelliana (19) andOxytropis arctobia (8), three species of arctic legumes found in the Melville Peninsula, Northwest Territories, Canada. On the basis of 74 characteristics (cultural, physiological, biochemical and host nodulation range) the 48 arctic rhizobia could be divided into 11 distinct groups by numerical analysis techniques. All 48 arctic rhizobia were able to nodulate the three arctic legume species and also sainfoin (Onobrychis viciifolia), however, milkvetch (Astragalus cicer) was only nodulated by 33 strains. In general, the arctic rhizobia showed properties found in both Rhizobium and Bradyrhizobium. The adaptation of the arctic strains to low temperature is indicated by their ability to grow in liquid culture at 5°C. Contribution no 293 of Agriculture Canada Research Station at Sainte-Foy.     

Leguminosae native nodulating bacteria from a gold mine As-contaminated soil: Multi-resistance to trace elements,and possible role in plant growth and mineral nutrition

Efficient N₂-fixing Leguminosae nodulating bacteria resistant to As may facilitate plant growth on As-contaminated sites. In order to identify bacteria holding these features, 24 strains were isolated from nodules of the trap species Crotalaria spectabilis (12) and Stizolobium aterrimum (12) growing on an As-contaminated gold mine site. 16S rRNA gene sequencing revealed that most of the strains belonged to the group of α-Proteobacteria, being representatives of the genera Bradyrhizobium, Rhizobium, Inquilinus, Labrys, Bosea, Starkeya, and Methylobacterium. Strains of the first four genera showed symbiotic efficiency with their original host, and demonstrated in vitro specific plant-growth-promoting (PGP) traits (production of organic acids, indole-3-acetic-acid and siderophores, 1-aminocyclopropane-1-carboxylate deaminase activity, and Ca₃(PO₄)₂ solubilization), and increased resistance to As, Zn, and Cd. In addition, these strains and some type and reference rhizobia strains exhibited a wide resistance spectrum to β-lactam antibiotics. Both intrinsic PGP abilities and multi-element resistance of rhizobia are promising for exploiting the symbiosis with different legume plants on trace-element-contaminated soils.     

Effect of Inoculation with Various Rhizobia Strains on the Fatty Acid Composition of Peribacteroid and Bacteroid Membranes of the Nodule Symbiosomes

The fatty acid (FA) composition of bacteroid and peribacteroid membranes was studied in the symbiotic pairs differing in their nitrogen-fixing efficiency; the results are compared with the FA composition of plasmalemma and free-living rhizobia. The experiments involved lupine plants inoculated with strains of Bradyrhizobium lupini359a (Nod⁺Fix⁺) and 400 (Nod⁺Fix L) manifesting high and low nitrogen-fixing efficiency, respectively, and broad bean plants inoculated with strains of Rhizobium leguminosarum97 (Nod⁺Fix⁺) and 87 (Nod⁺Fix L) of high and low nitrogen-fixing efficiency, respectively. We showed that the rhizobia of the strains 359a and 97 were able to form nodules with peribacteroid membranes containing FA mainly or exclusively of plant origin. These strains were able to develop effective symbiotic pairs with legume plants. The use of strains 400 and 87 resulted in the formation of nodules with peribacteroid membranes containing typical bacterial (branched-chain) FAs; these strains were characterized by an ineffective symbiosis.     

Diversity in symbiotic specificity of cowpea rhizobia indigenous to Zimbabwean soils

Tropical cowpea rhizobia are often presumed to be generally promiscuous but poor N fixers. This study was conducted to evaluate symbiotic interactions of 59 indigenous rhizobia isolates (49 of them from cowpea (Vigna unguiculata)), with up to 13 other (mostly tropical) legume species. Host ranges averaged 2.4 and 2.3 legume species each for fast- and slow-growing isolates respectively compared to 4.3 for slow-growing reference cowpea strains. An average of 22% and 19% of fast- and slow-growing cowpea isolates respectively were effective on each of 12 legume species tested. We conclude that the indigenous cowpea rhizobia studied have relatively narrow host ranges. The ready nodulation of different legumes in tropical soils appears due to the diversity of indigenous symbiotic genotypes, each consisting of subgroups compatible with a limited number of legume species.     

Host plant as an organizer of microbial evolution in the beneficial symbioses

Evolution of beneficial plant–microbe symbioses is presented as a result of selective processes induced by hosts in the associated microbial populations. These processes ensure a success of “genuine mutualists” (which benefit the host, often at the expense of their own fitness) in competition with “symbiotic cheaters” (which consume the resources provided by host without expressing the beneficial traits). Using a mathematical model describing the cyclic microevolution of rhizobia–legume symbiosis, we suggest that the selective pressures in favor of N₂-fixing (Fix⁺) strains operate within the in planta bacterial population due to preferential allocation of C resources into Fix⁺ nodules (positive partners’ feedbacks). Under the clonal infection of nodules, Fix⁺ strains (“genuine mutualists”) are supported by the group (inter-deme, kin) selection while under the mixed infections, this selection is ineffective since the Fix⁺ strains are over-competed by Fix⁻ ones (“symbiotic cheaters”) in the nodular habitats. Nevertheless, under mixed infections, Fix⁺ strains may be supported due to the coevolutionary responses form plant population which induce the mutualism-specific types of natural (group, individual) selection including the frequency dependent selection implemented in rhizobia population during the competition for host infection. Using the model of multi-strain bacterial competition for inoculation of symbiotic (rhizospheric, nodular) habitats, we demonstrate that the individual selection in favor of host-specific mutualist genotypes is more intensive than in favor of non-host-specific genotypes correlating the experimental data on the coordinated increases of symbiotic efficiency and specificity in the rhizobia–legume coevolution. However, an overall efficiency of symbiotic system is maximal when the non-host-specific mutualists are present in rhizobia population, and selection in favor of these mutualists operating at the whole population level is of key importance for improving the symbiosis. Construction of the agronomically valuable plant–microbe systems should provide the optimization of host-specific versus non-host-specific mutualists’ composition in legume inoculants combined with the clonal penetration of these mutualists into the nodules.     

Comparative symbiotic performance of native rhizobia of the Flooding Pampa and strains currently used for inoculating Lotus tenuis in this region

The Flooding Pampa (FP) is the most important area for cattle breeding in Argentina. In this region, persistence and yield of typical forage legumes are strongly limited by soil salinity and alkalinity, which affect around 30% of the total area. Instead, naturalized Lotus tenuis is the main forage legume in this region. Rhizobial strains currently used for inoculating L. tenuis in the FP are exotic or native from non-saline soils of this region, their taxonomic identity being unknown. Assuming that rhizobia native from the most restrictive environments are well adapted to adverse conditions, the use of such isolates could improve the productivity of L. tenuis in the FP. Hence, the goal of this study was to evaluate the symbiotic efficiency of selected L. tenuis rhizobia native from the FP, as compared with strains currently used for field inoculation of this legume. Under non-stressing conditions, the symbiotic performance of native strains of FP exceeded those ones currently used for L. tenuis. Moreover, the symbiotic performance of the native strain ML103 was considerably high under salt stress, compared with strains currently used as inoculants. Analysis of 16S rRNA gene sequencing revealed that unclassified rhizobia currently used for field inoculation of L. tenuis and native strains grouped with the genus Mesorhizobium. As a whole, results obtained demonstrate that soils of the FP are a source of efficient and diverse rhizobia that could be used as a sustainable agronomic tool to formulate inoculants that improve forage yield of L. tenuis in this region.     

Breeding for better symbiosis

The present review gives a critical assessment of the literature dealing with symbiosis between rhizobia and legumes and between AM fungi and most plants. Associative N₂ fixation (even though strictly speaking not a symbiotic relationship) does have some characteristics of symbiosis due to mutualistic dependence and usefulness of the relationship, and is therefore covered in this review. Nodulation in the rhizobia–legume symbiosis may be limited by an insufficient amount of the nod-gene inducers released from seed and/or roots. However, there is genotypic variation in the germplasm of legume species in all components of the signalling pathway, suggesting a prospect for improving nodulation by selecting and/or transforming legume genotypes for increased exudation of flavonoids and other signalling compounds. Deciphering chromosomal location as well as cloning nod, nif and other genes important in nodulation and N₂ fixation will allow manipulation of the presence and expression of these genes to enhance the symbiotic relationship. Increased efficacy of symbiotic N₂ fixation can be achieved by selecting not only the best host genotypes but by selecting the best combination of host genotype and nodule bacteria. As flavonoids exuded by legume seedlings may not only be nod-gene inducers, but also stimulants for hyphal growth of the AM fungi, selecting and/or transforming plants to increase exudation of these flavonoids may result in a double benefit for mycorrhizal legumes. Mutants unable to sustain mycorrhizal colonisation are instrumental in understanding the colonisation process, which may ultimately pay off in breeding for the more effective symbiosis. In conclusion, targeted efforts to breed genotypes for improved N₂ fixation and mycorrhizal symbiosis will bring benefits in increased yields of crops under a wide range of environmental conditions and will contribute toward sustainability of agricultural ecosystems in which soil-plant-microbe interactions will be better exploited.     

Nodulation and nitrogen fixation in extreme environments

Biological nitrogen fixation is a phenomenon occurring in all known ecosystems. Symbiotic nitrogen fixation is dependent on the host plant genotype, theRhizobium strain, and the interaction of these symbionts with the pedoclimatic factors and the environmental conditions. Extremes of pH affect nodulation by reducing the colonization of soil and the legume rhizosphere by rhizobia. Highly acidic soils (pH<4.0) frequently have low levels of phosphorus, calcium, and molybdenum and high concentrations of aluminium and manganese which are often toxic for both partners; nodulation is more affected than host-plant growth and nitrogen fixation. Highly alkaline soils (pH>8.0) tend to be high in sodium chloride, bicarbonate, and borate, and are often associated with high salinity which reduce nitrogen fixation. Nodulation and N-fixation are observed under a wide range of temperatures with optima between 20–30°C. Elevated temperatures may delay nodule initiation and development, and interfere with nodule structure and functioning in temperate Iegumes, whereas in tropical legumes nitrogen fixation efficiency is mainly affected. Furthermore, temperature changes affect the competitive ability ofRhizobium strains. Low temperatures reduce nodule formation and nitrogen fixation in temperate legumes; however, in the extreme environment of the high arctic, native legumes can nodulate and fix nitrogen at rates comparable to those observed with legumes in temperate climates, indicating that both the plants and their rhizobia have successfully adapted to arctic conditions. In addition to low temperatures, arctic legumes are exposed to a short growing season, a long photoperiod, low precipitation and low soil nitrogen levels. In this review, we present results on a number of structural and physiological characteristics which allow arctic legumes to function in extreme environments.     

Diversity of rhizobia and interactions among the host legumes and rhizobial genotypes in an agricultural-forestry ecosystem

To investigate the diversity of rhizobia and interactions among the host legumes and rhizobial genotypes in the same habitat, a total of 97 rhizobial strains isolated from nine legume species grown in an agricultural-forestry ecosystem were identified into seven genomic species and 12 symbiotic genotypes within the genera Bradyrhizobium, Mesorhizobium, Rhizobium and Sinorhizobium based upon analyses of genomic DNA regions and symbiotic genes. The results evidenced that the symbiotic genotypes of rhizobia were consistent with their hosts of origin; revealed that vertical transfer was the main mechanism in rhizobia to maintain the symbiotic genes but lateral transfer of symbiotic genes might have happened between the closely related rhizobial species; suggested the existence of co-distribution and co-evolution among the legume hosts and compatible rhizobia. All of these data demonstrated that the biogeography of rhizobia was a result of interactions among the host legumes, bacterial genomic backgrounds and environments.     

Note on the specificity of the rhizobia of crownvetch and sainfoin

Summary Rhizobium of crownvetch (Coronilla varia L.) was tested in test tubes on agar on several legumes and rhizobia from different cross inoculation groups were tested on crownvetch. Effective nodules were only formed on crownvetch after inoculation with crownvetch rhizobium and with two rhizobium strains of sainfoin (Onobrychis vicifólia). Four rhizobium strains from sainfoin nodules and six strains from crownvetch nodules were tested in quartzsand on sainfoin and crownvetch; all strains produced effective nodules on the roots of both species. Rhizobium of crownvetch was present in 32 of 57 soil samples collected in 7 provinces of the Netherlands. This rhizobium is present in most soils in the Netherlands with a pH of 7.8 to 7.0.     

Influence of the Size of Indigenous Rhizobial Populations on Establishment and Symbiotic Performance of Introduced Rhizobia on Field-Grown Legumes

Indigenous rhizobia in soil present a competition barrier to the establishment of inoculant strains, possibly leading to inoculation failure. In this study, we used the natural diversity of rhizobial species and numbers in our fields to define, in quantitative terms, the relationship between indigenous rhizobial populations and inoculation response. Eight standardized inoculation trials were conducted at five well-characterized field sites on the island of Maui, Hawaii. Soil rhizobial populations ranged from 0 to over 3.5 × 10⁴ g of soil^-1 for the different legumes used. At each site, no less than four but as many as seven legume species were planted from among the following: soybean (Glycine max), lima bean (Phaseolus lunatus), cowpea (Vigna unguiculata), bush bean (Phaseolus vulgaris), peanut (Arachis hypogaea), Leucaena leucocephala, tinga pea (Lathyrus tingeatus), alfalfa (Medicago sativa), and clover (Trifolium repens). Each legume was (i) inoculated with an equal mixture of three effective strains of homologous rhizobia, (ii) fertilized at high rates with urea, or (iii) left uninoculated. For soybeans, a nonnodulating isoline was used in all trials as the rhizobia-negative control. Inoculation increased economic yield for 22 of the 29 (76%) legume species-site combinations. While the yield increase was greater than 100 kg ha^-1 in all cases, in only 11 (38%) of the species-site combinations was the increase statistically significant (P ≤ 0.05). On average, inoculation increased yield by 62%. Soybean (G. max) responded to inoculation most frequently, while cowpea (V. unguiculata) failed to respond in all trials. Inoculation responses in the other legumes were site dependent. The response to inoculation and the competitive success of inoculant rhizobia were inversely related to numbers of indigenous rhizobia. As few as 50 rhizobia g of soil^-1 eliminated inoculation response. When fewer than 10 indigenous rhizobia g of soil^-1 were present, economic yield was significantly increased 85% of the time. Yield was significantly increased in only 6% of the observations when numbers of indigenous rhizobia were greater than 10 cells g of soil^-1. A significant response to N application, significant increases in nodule parameters, and greater than 50% nodule occupancy by inoculant rhizobia did not necessarily coincide with significant inoculation responses. No less than a doubling of nodule mass and 66% nodule occupancy by inoculant rhizobia were required to significantly increase the yield of inoculated crops over that of uninoculated crops. However, lack of an inoculation response was common even when inoculum strains occupied the majority of nodules. In these trials, the symbiotic yield of crops was, on average, only 88% of the maximum yield potential, as defined by the fertilizer N treatment. The difference between the yield of N-fertilized crops and that of N₂-fixing crops indicates a potential for improving inoculation technology, the N₂ fixation capacity of rhizobial strains, and the efficiency of symbiosis. In this study, we show that the probability of enhancing yield with existing inoculation technology decreases dramatically with increasing numbers of indigenous rhizobia.     

Nitrogen contributions from faba bean (Vicia faba L.) reliant on soil rhizobia or inoculation

Background and Aims

Understanding the impact of soil rhizobial populations and inoculant rhizobia in supplying sufficient nodulation is crucial to optimising N₂ fixation by legume crops. This study explored the impact of different rates of inoculant rhizobia and contrasting soil rhizobia on nodulation and N₂ fixation in faba bean (Vicia faba L.).

Methods

Faba beans were inoculated with one of seven rates of rhizobial inoculation, from no inoculant to 100 times the normal rate of inoculation, sown at two field sites, with or without soil rhizobia present, and their nodulation and N₂ fixation assessed.

Results

At the site without soil rhizobia, inoculation increased nodule number and increased N₂ fixation from 21 to 129 kg shoot N ha^?1, while N₂ fixation increased from 132 to 218 kg shoot N ha^?1 at the site with high background soil rhizobia. At the site without soil rhizobia, inoculation increased concentrations of shoot N from 14 to 24 mg g^?1, grain N from 32 to 45 mg g^?1, and grain yields by 1.0 Mg (metric tonne) ha^?1. Differences in nodulation influenced the contributions of fixed N to the system, which varied from the net removal of 20 kg N ha^?1 from the system in the absence of rhizobia, to a net maximum input of 199 kg N ha^?1 from legume shoot and root residues, after accounting for removal of N in grain harvest.

Conclusions

The impact of inoculation and soil rhizobia strongly influenced grain yield, grain N concentration and the potential contributions of legume cropping to soil N fertility. In soil with resident rhizobia, N₂ fixation was improved only with the highest inoculation rate.     

The contributions of nitrogen-fixing crop legumes to the productivity of agricultural systems

Data collated from around the world indicate that, for every tonne of shoot dry matter produced by crop legumes, the symbiotic relationship with rhizobia is responsible for fixing, on average on a whole plant basis (shoots and nodulated roots), the equivalent of 30–40 kg of nitrogen (N). Consequently, factors that directly influence legume growth (e.g. water and nutrient availability, disease incidence and pests) tend to be the main determinants of the amounts of N₂ fixed. However, practices that either limit the presence of effective rhizobia in the soil (no inoculation, poor inoculant quality), increase soil concentrations of nitrate (excessive tillage, extended fallows, fertilizer N), or enhance competition for soil mineral N (intercropping legumes with cereals) can also be critical. Much of the N₂ fixed by the legume is usually removed at harvest in high-protein seed so that the net residual contributions of fixed N to agricultural soils after the harvest of legumegrain may be relatively small.Nonetheless, the inclusion of legumes in a cropping sequence generally improves the productivity of following crops. Whilesome of these rotational effects may be associated with improvements in availability of N in soils, factors unrelated to N also play an important role. Recent results suggest that one such non-N benefit may be due to the impact on soil biology of hydrogenemitted from nodules as a by-product of N₂, fixation.     

Toward More Productive,Efficient, and Competitive Nitrogen-Fixing Symbiotic Bacteria

The prospects of developing strains of legume nodule bacteria that provide higher productivity of leguminous plants are described. The generic, biochemical, physiological, regulatory, and economic constraints that govern the ability of private and public efforts to construct better inoculants for legume nodulation are discussed. Success in constructing better inoculants requires a two-pronged approach. First, strains need to be improved in order to compete successfully with indigenous strains for root nodulation of legumes. Several loci have been identified to date that affect competitiveness for strain nodule occupancy. Usually mutations in these loci affect the ability of a strain to form nodules rapidly and efficiently. Other loci, such as those that confer antibiotic production, can be added to strains to enhance nodulation competitiveness when co-inoculated with antibiotic-sensitive strains. Second, the inoculum strains must be improved with respect to symbiotic nitrogen fixation. Efforts to enhance the symbiotic productivity of legume nodule bacteria either by mutation or genetic engineering are also described. The best characterized example of these is the hydrogenase system. Due to nitrogenase-dependent catalysis of proton reduction, diazotrophs evolve large amounts of H₂. An approach to maximize the efficiency of symbiotic N₂ fixation, and therefore of legume productivity, is to construct strains of Rhizobium with the ability to oxidize this otherwise wasted H₂. The electrons produced by H₂ oxidation are funneled through energy-conserving electron transport chains. Our knowledge of the genetics and biochemistry of H₂ oxidation in Bradyrhizobium japonicum and Rhizobium leguminosarum has developed rapidly in recent years. At least 20 genes are needed for these bacteria to manufacture and efficiently express a nickel-containing H₂-uptake hydrogenase. These genes include those encoding regulatory elements, posttranslational processing enzymes, nickel-sensing and nickel-metabolism proteins, and electron transport components for integrating the electrons from H₂ oxidation into the respiratory chain. Some of the components for oxidizing H₂ in the symbiotic N₂ fixing bacteria are distinct from the analogous components in (nonsymbiotic) H₂ oxidizing bacteria.     

Nodulation and nitrogen fixation by fast- and slow-growing rhizobia strains of soybean on several temperate and tropical legumes

Three slow-growingBradyrhizobium japonicum (G₃, USDA-110 and KUL-150) of diverse origins and two fast-growing strains ofRhizobium fredii (USDA-192 and USDA-193) were tested with a cropped soybean (Glycine max L. Merrill) cultivar, two cowpeas (Vigna unguiculata), one mung-bean (Phaseolus radiata), one winged-bean (Psophocarpus tetragonolobus) and one field bean (Phaseolus vulgaris) varieties.TheR. fredii strains nodulated and fixed Nitrogen as effectively as the strains ofB. japonicum in a modern european soybean cultivar, namely Fiskeby V. The other western bred soybeans tested were not nodulated by theseR. fredii strains. All of the soybean rhizobia produced nodules in both cowpeas and in mung-bean; theR. fredii strains showed effective N₂-fixation in the cowpeas, particularly USDA-193, yielding shoot dry weights greater than those from theB. japonicum. The symbiotic performance of theR. fredii strains with soybean and other legumes indicated that they should be placed in an intermediate group between the slow-growingB. japonicum and cowpearhizobium sp.The hydrogen uptake activites suggested a possible host effect on the expression of such genes in one out of theB. japonicum strains tested. Furthermore, the slow-growing rhizobia showed significantly higher nitrate-reduction than theR. fredii in the nodules.     

Enhancing legume N2 fixation through plant and soil management

Atmospheric N₂ fixed symbiotically by associations between Rhizobium spp. and legumes represents a renewable source of N for agriculture. Contribution of legume N₂ fixation to the N-economy of any ecosystem is mediated by: (i) legume reliance upon N₂ fixation for growth, and (ii) the total amount of legume-N accumulated. Strategies that change the numbers of effective rhizobia present in soil, reduce the inhibitory effects of soil nitrate, or influence legume biomass all have potential to alter net inputs of fixed N. A range of management options can be applied to legumes growing in farming systems to manipulate N₂ fixation and improve the N benefits to agriculture and agroforestry.     

Phylogenetic diversity of rhizobia associated with horsegram [Macrotyloma uniflorum (Lam.) Verdc.] grown in South India based on glnII, recA and 16S-23S intergenic sequence analyses

Horsegram [Macrotyloma uniflorum (Lam.) Verdc.) is an important grain legume and fodder crop in India. Information on root nodule endosymbionts of this legume in India is limited. In the present study, 69 isolates from naturally occurring root nodules of horsegram collected from two agro-eco-climatic regions of South India was analyzed by generation rate, acid/alkali reaction on YMA medium, restriction fragment length polymorphism analysis of 16S-23S rDNA intergenic spacer region (IGS), and sequence analyses of IGS and housekeeping genes glnII and recA. Based on the rDNA IGS RFLP by means of three restriction enzymes rhizobia were grouped in five clusters (I–V). By sequence analysis of 16S-23S rDNA IGS identified genotypes of horsegram rhizobia were distributed into five divergent lineages of Bradyrhizobium genus which comprised (I) the IGS type IV rhizobia and valid species B. yuanmingense, (II) the strains of IGS type I and Bradyrhizobium sp. ORS 3257 isolated from Vigna sp., (III) the strains of the IGS type II and Bradyrhizobium sp. CIRADAc12 from Acacia sp., (IV) the IGS type V strains and Bradyrhizobium sp. genospecies IV, and (V) comprising genetically distinct IGS type III strains which probably represent an uncharacterized new genomic species. Nearly, 87% of indigenous horsegram isolates (IGS types I, II, III, and V) could not be related to any other species within the genus Bradyrhizobium. Phylogeny based on housekeeping glnII and recA genes confirmed those results found by the analysis of the IGS sequence. All the isolated rhizobia nodulated Macrotyloma sp. and Vigna spp., and only some of them formed nodules on Arachis hypogeae. The isolates within each IGS type varied in their ability to fix nitrogen. Selection for high symbiotic effective strains could reward horsegram production in poor soils of South India where this legume is largely cultivated.     

Photoinhibition as a control on photosynthesis and production of Sphagnum mosses

The effect of high light intensity on photosynthesis and growth of Sphagnum moss species from Alaskan arctic tundra was studied under field and laboratory conditions. Field experiments consisted of experimental shading of mosses at sites normally exposed to full ambient irradiance, and removal of the vascular plant canopy from above mosses in tundra water track habitats. Moss growth was then monitored in the experimental plots and in adjacent control areas for 50 days from late June to early August 1988. In shaded plots total moss growth was 2–3 times higher than that measured in control plots, while significant reductions in moss growth were found in canopy removal plots. The possibility that photoinhibition of photosynthesis might occur under high-light conditions and affect growth was studied under controlled laboratory conditions with mosses collected from the arctic study site, as well as from a temperate location in the Sierra Nevada, California. After 2 days of high-light treatment (800 mol photons m^–2 s^–1) in a controlled environmental chamber, moss photosynthetic capacity was significantly lowered in both arctic and temperate samples, and did not recover during the 14-day experimental period. The observed decrease in photosynthetic capacity was correlated (r ²=0.735, P<0.001) with a decrease in the ratio of variable to maximum chlorophyll fluorescence (F _v/F _m) in arctic and temperate mosses. This relationship indicates photoinhibition of photosynthesis in both arctic and temperate mosses at even moderately high light intensities. It is suggested that susceptibility to photoinhibition and failure to photoacclimate to higher light intensities in Sphagnum spp. may be related to low tissue nitrogen levels in these exclusively ombrotrophic plants. Photoinhibition of photosynthesis leading to lowered annual carbon gain in Sphagnum mosses may be an important factor affecting CO₂ flux at the ecosystem level, given the abundance of these plants in Alaskan tussock tundra.     

Genetic diversity of <Emphasis Type="Italic">Sinorhizobium meliloti</Emphasis> associated with alfalfa in Chilean volcanic soils and their symbiotic effectiveness under acidic conditions

A study was conducted with the aim of evaluating the genetic diversity of alfalfa rhizobia isolated from volcanic soils in southern Chile and their ability to establish an effective symbiosis with alfalfa. Rhizobial strains isolated from nodules were identified and selected based on PCR analyses and acid tolerance. Symbiotic effectiveness (nodulation and shoot dry weight) of acid-tolerant rhizobia was evaluated in glasshouse experiments under acidic conditions. The results revealed that Sinorhizobium meliloti is the dominant species in alfalfa nodules with a high genetic diversity at strain level grouped in three major clusters. There was a close relationship (r ² = 0.895, P ≤ 0.001, n = 40) between soil pH and the size of rhizobial populations. Representative isolates from major cluster groups showed wide variation in acid tolerance expressed on buffered agar plates (pH 4.5–7.0) and symbiotic effectiveness with alfalfa. One isolate (NS11) appears to be suitable as an inoculant for alfalfa according to its acid tolerance and symbiotic effectiveness at low pH (5.5). The isolation and selection of naturalized S. meliloti strains with high symbiotic effectiveness under acidic conditions is an alternative approach to improving the productivity of alfalfa and for reducing the application of synthetic fertilizers in Chile.     

Symbiotic properties of a chimeric Nod-independent photosynthetic Bradyrhizobium strain obtained by conjugative transfer of a symbiotic plasmid

The lateral transfer of symbiotic genes converting a predisposed soil bacteria into a legume symbiont has occurred repeatedly and independently during the evolution of rhizobia. We experimented the transfer of a symbiotic plasmid between Bradyrhizobium strains. The originality of the DOA9 donor is that it harbours a symbiotic mega-plasmid (pDOA9) containing nod, nif and T3SS genes while the ORS278 recipient has the unique property of inducing nodules on some Aeschynomene species in the absence of Nod factors (NFs). We observed that the chimeric strain ORS278-pDOA9* lost its ability to develop a functional symbiosis with Aeschynomene. indica and Aeschynomene evenia. The mutation of rhcN and nodB led to partial restoration of nodule efficiency, indicating that T3SS effectors and NFs block the establishment of the NF-independent symbiosis. Conversely, ORS278-pDOA9* strain acquired the ability to form nodules on Crotalaria juncea and Macroptillium artropurpureum but not on NF-dependent Aeschynomene (A. afraspera and A. americana), suggesting that the ORS278 strain also harbours incompatible factors that block the interaction with these species. These data indicate that the symbiotic properties of a chimeric rhizobia cannot be anticipated due to new combination of symbiotic and non-symbiotic determinants that may interfere during the interaction with the host plant.