Biological N2 fixation represents the major source of N input in agricultural soils including those in arid regions. The major N2-fixing systems are the symbiotic systems, which can play a significant role in improving the fertility and productivity of low-N soils. The Rhizobium-legume symbioses have received most attention and have been examined extensively. The behavior of some N2-fixing systems under severe environmental conditions such as salt stress, drought stress, acidity, alkalinity, nutrient deficiency, fertilizers, heavy metals, and pesticides is reviewed. These major stress factors suppress the growth and symbiotic characteristics of most rhizobia; however, several strains, distributed among various species of rhizobia, are tolerant to stress effects. Some strains of rhizobia form effective (N2-fixing) symbioses with their host legumes under salt, heat, and acid stresses, and can sometimes do so under the effect of heavy metals. Reclamation and improvement of the fertility of arid lands by application of organic (manure and sewage sludge) and inorganic (synthetic) fertilizers are expensive and can be a source of pollution. The Rhizobium-legume (herb or tree) symbiosis is suggested to be the ideal solution to the improvement of soil fertility and the rehabilitation of arid lands and is an important direction for future research.

Rhizobium-Legume Symbiosis and Nitrogen Fixation under Severe Conditions and in an Arid Climate

The nodC and nifH genes were characterized in a collection of 83 rhizobial strains which represented 23 recognized species distributed in the genera Rhizobium, Sinorhizobium, Mesorhizobium and Bradyrhizobium, as well as unclassified rhizobia from various host legumes. Conserved primers were designed from available nucleotide sequences and were able to amplify nodC and nifH fragments of about 930 bp and 780 bp, respectively, from most of the strains investigated. RFLP analysis of the PCR products resulted in a classification of these rhizobia which was in general well-correlated with their known host range and independent of their taxonomic status. The nodC and nifH fragments were sequenced for representative strains belonging to different genera and species, most of which originated from Phaseolus vulgaris nodules. Phylogenetic trees were constructed and revealed close relationships among symbiotic genes of the Phaseolus symbionts, irrespective of their 16S-rDNA-based classification. The nodC and nifH phylogenies were generally similar, but cases of incongruence were detected, suggesting that genetic rearrangements have occurred in the course of evolution. The results support the view that lateral genetic transfer across rhizobial species and, in some instances, across Rhizobium and Sinorhizobium genera plays a role in diversification and in structuring the natural populations of rhizobia.

Classification of rhizobia based on nodC and nifH gene analysis reveals a close phylogenetic relationship among Phaseolus vulgaris symbionts.

https://biblio.ugent.be/publication/1019751/file/1019788.pdf

The biodiversity of beneficial microbe-host mutualism: the case of rhizobia.

Legume nodules, specialized structures for nitrogen fixation, are probably the result of coevolution of plants and ancestral rhizobia. Among the evolutionary processes leading to legume radiation and divergence, coevolution with rhizobia might have occurred. Alternatively, bacteria could have been constantly selected by plants, with bacteria slightly influencing plant evolution (required to fulfill the criteria for a coevolutionary hypothesis). Evidence of bacterial effects on plant evolution is scarce but being searched for. Bacterial genetic plasticity may be indicative of the large capacity of Rhizobium to adapt to legumes. Events such as symbiotic replacement, easy recruitment of symbiotic bacteria by legume plants, and lateral transfer of symbiotic genes seem to erase the coevolutionary or selected relationships in rhizobial-legume symbiosis. In particular, the hypotheses proposed are (1) Rhizobium replaced Bradyrhizobium in a few hosts of the Phaseoleae tribe, Phaseolus vulgaris and P. coccineus; (2) Rhizobium etli as a species did not coevolve with bean; and (3) beta-Proteobacteria replaced alpha-Proteobacteria in South American mimosas. Novel results on symbiosis suggest a more complex evolutionary process for nodulation that may include multiple organisms, such as mycorrhiza, nematodes, and other bacteria in addition to rhizobia.

Coevolution in Rhizobium-Legume Symbiosis?

A group of four diverse rhizobial isolates and two soil isolates that are highly related to Ensifer adhaerens were characterized by a polyphasic approach. On the basis of DNA–DNA hybridizations and phenotypic features, these strains cannot be distinguished clearly form Ensifer adhaerens, a soil bacterium that was described in 1982, mainly on the basis of phenotypic characteristics. Phylogenetically, Ensifer and Sinorhizobium form a single group in the 16S rDNA dendrogram of the α-Proteobacteria, as well as in an analysis of partial recA gene sequences. They may therefore be regarded as a single genus. Because Sinorhizobium was proposed in 1988, according to the Bacteriological Code (1990 Revision) the older name, Ensifer, has priority. However, there are several reasons why a change from Sinorhizobium to Ensifer may not be the best solution and making an exception to Rule 38 may be more appropriate. We therefore propose the species Sinorhizobium adhaerens comb. nov. and put forward a Request for an Opinion to the Judicial Commission regarding the conservation of Sinorhizobium adhaerens over Ensifer adhaerens.

Description of new Ensifer strains from nodules and proposal to transfer Ensifer adhaerens Casida 1982 to Sinorhizobium as Sinorhizobium adhaerens comb. nov. Request for an opinion

Over 4 million ha of land throughout Argentina and Uruguay are used for the production of alfalfa (Medicago sativa L.) (38). Therefore, it is important to manage the N2-fixing symbiosis to maximize the production of this crop. An important constraint to this aim results from the moderately low soil pH that affects the establishment of an effective symbiosis with indigenous and inoculated rhizobia. Large areas of arable lands in the central region of Argentina have progressively acidified over the last 10 to 20 years (21, 34), where the continuous cultivation over time without crop rotation has been identified as one of the main factors that favored the acidification of soils (21, 34).

It has been shown that the poor symbiosis at low pH results from a variety of influences upon the host plant (24, 27), the population of rhizobia (31), and the symbiotic interaction itself (11, 36, 41). Early studies by Munns (36, 37) compared the progress of symbiosis under neutral and acid conditions, concluding that early steps during preinfection are the more acid-sensitive events (36). This observation is in agreement with results reported by Caetano Anolles et al. in 1989, who showed a negative influence of low pH on the bacterial attachment to roots. Most of the fundamental research has sought to characterize the physiology of the interaction (11, 23, 25, 27, 28) and the effects of acidity in laboratory and in field experiments (1a, 39) and only more recently to address the identification of the bacterial determinants of acid tolerance (18, 43, 44). Particularly, Sinorhizobium meliloti strains are among the more acid-sensitive rhizobia (6, 19, 20). Most S. meliloti isolates tolerate acidity in the range between pH 5.5 and 6.0 (25). Although there is no basis to support that a higher acid tolerance of the bacteria will correspond to a better symbiotic performance under acidic conditions, it was found that acid-tolerant (AT) S. meliloti strains isolated from nodulated Medicago spp. collected in Sardinia enhanced the establishment of medic pastures in mildly acidic soils from Western Australia (23, 25). In any case, although symbiotic proficiency and acid tolerance of rhizobia are both desirable bacterial traits, they are not necessarily linked (17, 22, 25, 30, 31). While basic aspects of symbiosis have been extensively characterized, further work is still needed in order to increase our knowledge on the rhizobial ecology under suboptimal environmental conditions such as acidity.

The characterization of the populations of alfalfa-nodulating rhizobia from acid soils had shown the presence of alfalfa-specific nodulating S. meliloti and another lineage represented by strain Or191 isolated from Oregon which also nodulates the common bean (13, 15). Strain Or191 was also shown to be more tolerant to acidity on agar plates (pH 5.2). Results of the genetic characterization indicate that strain Or191 is related to a previously unrecognized taxonomic group that includes strains of Rhizobium phaseoli type I (15), since renamed Rhizobium etli (32, 45). The AT strain Or191 was ineffective in alfalfa (13) but had measurable levels of symbiotic nitrogen fixation (15).

So far, a detailed examination of the composition of native populations of alfalfa-nodulating rhizobia in soils from Argentina and Uruguay has not yet been carried out. In this work, we present results on the isolation and characterization of alfalfa-nodulating rhizobia from local acid soils and demonstrate the presence of two rhizobial populations with marked differences in their acid tolerance and symbiotic properties.

Francisco Martínez Abarca

Papers

Isolation and characterization of alfalfa-nodulating rhizobia present in acidic soils of central argentina and uruguay