Phenotypic and genetic characterization of rhizobia associated with alfalfa in the Hokkaido and Ishigaki regions of Japan

Twenty five rhizobial isolates were obtained from root nodules of Medicago sativa inoculated with soil samples collected from the Sapporo region and Ishigaki Island in Japan. To study their diversity and characterize them in relation to the climatic conditions of their soils of origin, a polyphasic approach analyzing stress tolerance, symbiotic and genetic properties was used. Stress tolerance assays revealed marked variations in salinity, pH and temperature tolerance. Isolates originating from a sub-tropical climate in alkaline soil (Ishigaki Island) tolerated high temperature, salinity and pH levels. Moreover, isolates recovered from a temperate climate in acidic soil (Sapporo) were sensitive to high temperature and salinity, and tolerated acidic pH. Phylogenetic analysis of conserved 16S rRNA and recA genes, and symbiotic nodA and nifDK revealed 25 isolates to be closely related to Ensifer meliloti. Furthermore, the branch patterns of phylogenetic trees constructed from different genes revealed the existence of at least two E. meliloti types in the soils studied. These results may be relevant to programs directed towards improving crop productivity through biofertilization with locally adapted and genetically defined strains.     

Phenotypic and genetic diversity in <Emphasis Type="Italic">Sinorhizobium meliloti</Emphasis> and <Emphasis Type="Italic">S. medicae</Emphasis> from drought and salt affected regions of Morocco

Background  

Sinorhizobium meliloti and S. medicae are symbiotic nitrogen fixing bacteria in root nodules of forage legume alfalfa (Medicago sativa L.). In Morocco, alfalfa is usually grown in marginal soils of arid and semi-arid regions frequently affected by drought, extremes of temperature and soil pH, soil salinity and heavy metals, which affect biological nitrogen fixing ability of rhizobia and productivity of the host. This study examines phenotypic diversity for tolerance to the above stresses and genotypic diversity at Repetitive Extragenic Pallindromic DNA regions of Sinorhizobium nodulating alfalfa, sampled from marginal soils of arid and semi-arid regions of Morocco.     

Plant defence and delayed infection of alfalfa pseudonodules induced by an exopolysaccharide (EPS I)-deficient Rhizobium meliloti mutant

Mutants of the symbiotic soil bacterium Rhizobium meliloti that fail to synthesize the acidic exopolysaccharide EPS I were unable to induce infected root nodules on Medicago sativa L. (alfalfa). These strains, however, elicited pseudonodules that contained no infection threads or bacteroids. The cortical cell walls of the pseudonodules were abnormally thick and incrusted with an autofluorescent material. Parts of these cell walls and wall appositions contained callose. Biochemical analysis of nodules induced by the EPS I-deficient R. meliloti mutant revealed an increase of phenolic compounds bound to the nodule cell walls when compared with the wild-type strain. These microscopic and biochemical data indicated that a general plant defence response against the EPS I-deficient mutant of R. meliloti was induced in alfalfa pseudonodules. Following prolonged incubation with the EPS I-deficient R. meliloti mutant, the defence system of the alfalfa plant could be overcome by the rhizobium mutant. In the case of the delayed infections, the mutants colonized lobes of the pseudonodules, but the infection threads in these nodules had an abnormal morphology. They were greatly enlarged and did not contain the typical gum-like matrix inside. The bacteria were tightly packed. Based on the mechanism of phytopathogenic interactions, we propose that EPS I or a related compound may act as a suppressor of the alfalfa plant defence system, enabling R. meliloti to infect the plant.     

Influence of drought on competition between selected Rhizobium meliloti strains and naturalized soil rhizobia in alfalfa

Drought is an important environmental factor that can affect rhizobial competition and N₂ fixation. Three alfalfa (Medicago sativa L. and M. falcata L.) accessions were grown in pots containing soil from an irrigated (Soil 1) and a dryland (Soil 2) alfalfa field in northern Utah, USA. Mutants of three strains of Rhizobium meliloti Dang. from Pakistan (UL 136, UL 210, and UL 222) and a commercial rhizobial strain 102F51a were developed with various levels of resistance to streptomycin. Seeds inoculated with these individual streptomycin-resistant mutants were sown in the two soils containing naturalized rhizobial populations. Soils in the pots were maintained at −0.03, −0.5, and −1.0 MPa. After 10 weeks, plants were harvested and nodule isolates were cultured on agar medium with and without streptomycin to determine nodule occupancy (proportion of the nodules occupied by introduced rhizobial strains). Number of nodules, nodule occupancy, total plant dry weight, and shoot N were higher for Soil 1 than Soil 2. Number of nodules, plant dry weight, and shoot N decreased as drought increased from −0.03 to −1.0 MPa in the three alfalfa accessions. Rhizobial strains UL 136 and UL 222 were competitive with naturalized alfalfa rhizobia and were effective at symbiotic N₂ fixation under drought. These results suggest that nodulation, growth, and N₂ fixation in alfalfa can be improved by inoculation with competitive and drought-tolerant rhizobia and may be one economically feasible way to increase alfalfa production in water-limited environments. Joint contribution from USDA-ARS and the Utah Agric. Exp. Sta., Utah State Univ., Logan, UT 84322-4810, USA. Journal Paper No. 4931. Joint contribution from USDA-ARS and the Utah Agric. Exp. Sta., Utah State Univ., Logan, UT 84322-4810, USA. Journal Paper No. 4931.     

The Symbiotic Requirements of Different Medicago Spp. Suggest the Evolution of Sinorhizobium Meliloti and S. Medicae with Hosts Differentially Adapted to Soil pH

Nitrogen fixing rhizobia associated with the Medicago L. genus belong to two closely related species Sinorhizobium medicae and S. meliloti. To investigate the symbiotic requirements of different Medicago species for the two microsymbionts, 39 bacterial isolates from nodules of eleven Medicago species growing in their natural habitats in the Mediterranean basin plus six historical Australian commercial inocula were symbiotically characterized with Medicago hosts. The bacterial species allocation was first assigned on the basis of symbiotic proficiency with M. polymorpha. PCR primers specific for 16S rDNA were then designed to distinguish S. medicae and S. meliloti. PCR amplification results confirmed the species allocation acquired in the glasshouse. PCR fingerprints generated from ERIC, BOXA1R and nif-directed RPO1 primers revealed that the Mediterranean strains were genetically heterogenous. Moreover PCR fingerprints with ERIC and BOX primers showed that these repetitive DNA elements were specifically distributed and conserved in S. meliloti and S. medicae, clustering the strains into two divergent groups according to their species. Linking the Sinorhizobium species with the plant species of origin we have found that S. medicae was mostly associated with medics well adapted to moderately acid soils such as M. polymorpha, M. arabica and M. murex whereas S. meliloti was predominantly isolated from plants naturally growing on alkaline or neutral pH soils such as M. littoralis and M. tornata. Moreover in glasshouse experiments the S. medicae strains were able to induce well-developed nodules on M. murex whilst S. meliloti was not infective on this species. This feature provides a very distinguishing characteristic for S. medicae. Results from the symbiotic, genotypic and cultural characterization suggest that S. meliloti and S. medicae have adapted to different Medicago species according to the niches these medics usually occupy in their natural habitats.     

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.     

Selection for acid tolerance inRhizobium meliloti

In the selection of acid-tolerantRhizobium meliloti, procedures for the collection and isolation of rhizobia, and the assessment of acid tolerance, have not been critically evaluated. Such procedures form the basis of this study. Root nodules were collected fromMedicago spp. found growing on acid soil in Sardinia. Their encumbent bacteria were isolated directly onto media adjusted over a range of pH values, and then assessed for acid tolerance in both the laboratory and field. Strains ofRhizobium meliloti isolated onto low pH media were, in general, more acid-tolerant than sister isolates from high pH media, when tested in both the laboratory and field. Dilution (10 or 100 fold) of the inocula used in the laboratory assessment did not greatly influence the rating derived, although there was some effect of bacterial colony type on growth rating. The link between polysaccharide production and acid tolerance was not strong. There was a poor correlation between the growth ratings derived from the laboratory screening and acid tolerance as expressed in the field.     

Derivatives of Rhizobium meliloti strains carrying a plasmid of Rhizobium leguminosarum specifying hydrogen uptake and pea-specific symbiotic functions

pIJ1008, a Rhizobium leguminosarum plasmid which determines hydrogen uptake ability and symbiotic functions in pea was transferable to three of seven natural isolates of R. meliloti tested. In these three strains, pIJ1008 was maintained stably with the respective sym megaplasmid indigenous to each R. meliloti strain. These strains carrying both plasmids nodulated alfalfa but not pea. By reisolation and examination of the strains from alfalfa nodule tissue, it was shown that pIJ1008 continued to be maintained but that pea-nodulation ability was suppressed.In one strain of R. meliloti which carries a 200 kb cryptic plasmid (in addition to a megaplasmid), the transfer and selection for pIJ1008 resulted in the loss of the cryptic plasmid.In three separate plant growth experiments, alfalfa nodules induced by each of the R. meliloti strain carrying both sym plasmids were assayed for hydrogen uptake activity. The average activity was 40-, 3.5-and 2-fold higher than with the respective pIJ1008-free strains. However, this higher activity was not accompanied by an increase in plant biomass or nitrogen content of shoots.C.B.R.I. Contribution Number: 1478     

IS<Emphasis Type="Italic">Rm31</Emphasis>, a new insertion sequence of the IS<Emphasis Type="Italic">66</Emphasis> family in<Emphasis Type="Italic"> Sinorhizobium meliloti</Emphasis>

Sinorhizobium meliloti natural populations show a high level of genetic polymorphism possibly due to the presence of mobile genetic elements such as insertion sequences (IS), transposons, and bacterial mobile introns. The analysis of the DNA sequence polymorphism of the nod region of S. meliloti pSymA megaplasmid in an Italian isolate led to the discovery of a new insertion sequence, ISRm31. ISRm31 is 2,803 bp long and has 22-bp-long terminal inverted repeat sequences, 8-bp direct repeat sequences generated by transposition, and three ORFs (A, B, C) coding for proteins of 124, 115, and 541 amino acids, respectively. ORF A and ORF C are significantly similar to members of the transposase family. Amino acid and nucleotide sequences indicate that ISRm31 is a member of the IS66 family. ISRm31 sequences were found in 30.5% of the Italian strains analyzed, and were also present in several collection strains of the Rhizobiaceae family, including S. meliloti strain 1021. Alignment of targets sites in the genome of strains carrying ISRm31 suggested that ISRm31 inserts randomly into S. meliloti genomes. Moreover, analysis of ISRm31 insertion sites revealed DNA sequences not present in the recently sequenced S. meliloti strain 1021 genome. In fact, ISRm31 was in some cases linked to DNA fragments homologous to sequences found in other rhizobia species.     

钒钛磁铁尾矿土壤中水黄皮根瘤菌铁耐受性与钝化能力研究

铁是好氧微生物生长所必需的元素,而铁污染土壤环境中的根瘤菌是否对高浓度铁具有耐受性和钝化能力尚不清楚。以攀枝花钒钛磁铁尾矿土壤作为基质进行水黄皮共生根瘤菌捕获实验,获得水黄皮共生根瘤并从中分离纯化出根瘤菌39株。通过Fe~(2+)/Fe~(3+)耐受性和钝化能力测试筛选出耐受性和钝化能力均强的优势菌株PZHS20、PZHS90、PZHS87,其对Fe~(2+)的最大耐受质量浓度为1 600 mg/L,其中PZHS20在200 mg/L Fe~(2+)溶液中钝化效率最大,为73.54%;PZHS90对Fe~(3+)的最大耐受质量浓度为1 600 mg/L,而PZHS20和PZHS87对Fe~(3+)的最大耐受质量浓度为1 800 mg/L,其在200 mg/L Fe~(3+)溶液中钝化效率分别为84.25%和81.95%。16S rRNA基因系统进化分析将PZHS20鉴定为苍白杆菌(Ochrobactrum),将PZHS90和PZHS87鉴定为慢生根瘤菌(Bradyrhizobium)。研究结果表明,钒钛磁铁尾矿土壤中的水黄皮根瘤菌具有不同程度的Fe~(2+)/Fe~(3+)耐受性和钝化能力,筛选出的优势菌株为进一步利用水黄皮-根瘤菌联合修复高浓度铁污染土壤提供可利用的菌株资源。     

Competition for alfalfa nodulation under metal stress by the metal‐tolerant strain Ochrobactrum cytisi Azn6.2

Legume plants, in association with rhizobia, are gaining increasing interest for heavy metal rhizoremediation. This symbiotic interaction combines the advantages of rhizoremediation and soil nitrogen enrichment. In metal polluted soils, Ochrobactrum cytisi can elicit non‐fixing nodules on legumes, including Medicago sativa. Nodulation kinetics was much slower when M. sativa plants were inoculated with O. cytisi Azn6.2 compared with the natural symbiont Ensifer meliloti 1021 and nodules were ineffective in nitrogen fixation. A competition experiment was performed using alfalfa grown on heavy metals, and co‐inoculated with equal amounts of the metal‐sensitive E. meliloti 1021 and the metal‐resistant O. cytisi Azn6.2. When plants were inoculated in non‐polluted substrates, all nodules were formed by E. meliloti 1021. Nevertheless, under increasing metal concentrations, the number of nodules occupied by O. cytisi grew. At the highest metal concentration, all nodules were elicited by O. cytisi, suggesting that the resistant species can take the place of the natural symbiont. This fact has important ecological and environmental implications when proposing legume–rhizobia symbioses for rhizoremediation and highlights the need of selecting highly resistant rhizobia in order to be competitive in polluted soils.     

Determination of nitrogen fixation effectiveness in selected <Emphasis Type="Italic">Medicago truncatula</Emphasis> isolates by measuring nitrogen isotope incorporation into pheophytin

Effectiveness is a term used to describe the input that a bacterial nitrogen-fixing symbiosis makes to plant nitrogen metabolism. In legumes, effectiveness is considered a polymorphic trait where specific interactions between the plant and symbiotic rhizobia contribute to the success of the interaction. Evaluation of effectiveness using model legumes like Medicago truncatula may open new avenues for genetic studies. In previous work, an isotope dilution mass spectrometry method, which uses the effect of nitrogen fixation on the nitrogen isotope composition of chlorophyll in plants grown on ¹⁵N fertilizer as a measure of effectiveness, was developed for estimating the contribution of symbiotic nitrogen fixation to plant nitrogen content. This ¹⁵N-dilution assay was used to evaluate the level of nitrogen fixation effectiveness in three Medicago truncatula lines that have been used as parents in generating recombinant inbred lines. Three Sinorhizobium meliloti strains, USDA 1600, 102F51 and MK506, differ in this measure of effectiveness on three lines of M. truncatula: Jemalong A17, DZA315.16 and F83005.5. Plant–rhizobia combinations grown in two different conditions showed comparable differences in effectiveness.     

Flavodoxin overexpression reduces cadmium-induced damage in alfalfa root nodules

Flavodoxins are electron carrier flavoproteins that are involved in the response to oxidative stress in bacteria and cyanobacteria. Recently, we obtained Sinorhizobium meliloti bacteria that overexpressed a flavodoxin from the cyanobacterium Anabaena variabilis [Redondo et al. (2009) Plant Physiology 149:1166–1178]. In the present work, tolerance to cadmium was evaluated in free-living transformed S. meliloti and in alfalfa plants nodulated by the flavodoxin-overexpressing rhizobia, in comparison with plants nodulated by wild-type bacteria. Overexpression of flavodoxin protected free-living S. meliloti from cadmium toxicity and had a positive effect on nitrogen fixation of alfalfa plants subjected to cadmium stress. Flavodoxin notably reduced cadmium-induced structural and ultrastructural alterations in alfalfa nodules. Putative protection mechanisms in flavodoxin-overexpressing nodules are discussed. Flavodoxin could have applications as a biotechnological tool to improve the symbiotic performance of alfalfa and other legumes in cadmium polluted soils.     

Inefficient nodulation of chickpea (Cicer arietinum L.) in the arid and Saharan climates in Tunisia bySinorhizobium meliloti biovarmedicaginis

The aim of this work is to study the genetic diversity and the symbiotic effectiveness of the natural populations of rhizobia nodulating chickpea (Cicer arietinum L.) in six locations of South Tunisia, where chickpea had never been cultivated. Nodules were observed only in the two soil samples from Gafsa (0.8 nodules per plant) and Tataouine (2 nodules per plant). PCR-RFLP typing of 165 rRNA genes of 42 isolates indicated that all analysed strains showed the same ribotype as the reference strainSinorhizobium meliloti RCR2011. These isolates induced ineffective nodules on chickpea andMedicago sativa; however nodules onMedicago laciniata were effective. Analysis of the symbiotic diversity by PCR-RFLP, of thenifDK spacer suggested that all chickpea isolates from the South belong to the biovarmedicaginis ofS. meliloti. The present paper is, to our knowledge, the first report showing that chickpea is selectively nodulated under soil conditions by a specific biovar ofS. meliloti showing specificity toM. laciniata. The specificity of this interaction as well the impact of this inefficient nodulation on chickpea cultivation needs to be investigated further.     

The two megaplasmids of Rhizobium meliloti are involved in the effective nodulation of alfalfa

Summary We have shown by physical and genetic means that there are two megaplasmids in all strains of Rhizobium meliloti that we have studied. Megaplasmids from several strains of R. meliloti were mobilized to Agrobacterium tumefaciens and to other Rhizobium strains using the Tn5-Mob system. We were also able to resolve these two megaplasmids in agarose gels for most strains, and to show that only one of them hybridized to nif and nod genes. Transfer of this plasmid, the pSym, to Agrobacterium, R. leguminosarum, and R. trifolii strains conferred on these recipients the ability to nodulate alfalfa ineffectively. The second megaplasmid did not appear to have a direct role in nodule initiation. However, we were able to complement extracellular polysaccharide (EPS^-) mutants of R. meliloti by transferring this second megaplasmid into them. Furthermore, Tn5-induced EPS^- mutants of R. meliloti 2011, which produced ineffective (Fix^-) nodules of abnormal morphology, were shown by hybridization and complementation to carry mutations in this second megaplasmid. This demonstrates that both megaplasmids of R. meliloti are necessary for the effective nodulation of alfalfa.     

Ensifer meliloti bv. lancerottense establishes nitrogen-fixing symbiosis with Lotus endemic to the Canary Islands and shows distinctive symbiotic genotypes and host range

Eleven strains were isolated from root nodules of Lotus endemic to the Canary Islands and they belonged to the genus Ensifer, a genus never previously described as a symbiont of Lotus. According to their 16S rRNA and atpD gene sequences, two isolates represented minority genotypes that could belong to previously undescribed Ensifer species, but most of the isolates were classified within the species Ensifer meliloti. These isolates nodulated Lotus lancerottensis, Lotus corniculatus and Lotus japonicus, whereas Lotus tenuis and Lotus uliginosus were more restrictive hosts. However, effective nitrogen fixation only occurred with the endemic L. lancerottensis. The E. meliloti strains did not nodulate Medicago sativa, Medicago laciniata Glycine max or Glycine soja, but induced non-fixing nodules on Phaseolus vulgaris roots. nodC and nifH symbiotic gene phylogenies showed that the E. meliloti symbionts of Lotus markedly diverged from strains of Mesorhizobium loti, the usual symbionts of Lotus, as well as from the three biovars (bv. meliloti, bv. medicaginis, and bv. mediterranense) so far described within E. meliloti. Indeed, the nodC and nifH genes from the E. meliloti isolates from Lotus represented unique symbiotic genotypes. According to their symbiotic gene sequences and host range, the Lotus symbionts would represent a new biovar of E. meliloti for which bv. lancerottense is proposed.     

Salt-tolerant rhizobia isolated from a Tunisian oasis that are highly effective for symbiotic N2-fixation with Phaseolus vulgaris constitute a novel biovar (bv. mediterranense) of Sinorhizobium meliloti

Nodulation of common bean was explored in six oases in the south of Tunisia. Nineteen isolates were characterized by PCR–RFLP of 16S rDNA. Three species of rhizobia were identified, Rhizobium etli, Rhizobium gallicum and Sinorhizobium meliloti. The diversity of the symbiotic genes was then assessed by PCR–RFLP of nodC and nifH genes. The majority of the symbiotic genotypes were conserved between oases and other soils of the north of the country. Sinorhizobia isolated from bean were then compared with isolates from Medicago truncatula plants grown in the oases soils. All the nodC types except for nodC type p that was specific to common bean isolates were shared by both hosts. The four isolates with nodC type p induced N₂-fixing effective nodules on common bean but did not nodulate M. truncatula and Medicago sativa. The phylogenetic analysis of nifH and nodC genes showed that these isolates carry symbiotic genes different from those previously characterized among Medicago and bean symbionts, but closely related to those of S. fredii Spanish and Tunisian isolates effective in symbiosis with common bean but unable to nodulate soybean. The creation of a novel biovar shared by S. meliloti and S. fredii, bv. mediterranense, was proposed.     

Diversity and symbiotic divergence of endophytic and non-endophytic rhizobia of <Emphasis Type="Italic">Medicago sativa</Emphasis>

Knowledge of rhizobium diversity is helping to enable the utilization of rhizobial resources. To analyze the phenotypic and genetic diversity and the symbiotic divergence of rhizobia of Medicago sativa, 30 endophytic and non-endophytic isolates were collected from different parts of five alfalfa varieties in three geographic locations in Gansu, China. Numerical analyses based on 72 phenotypic properties and restriction fragment length polymorphism (RFLP) fingerprinting indicated the abundant phenotypic and genetic diversity of the tested strains. According to the phylogenetic analysis of 16S RNA, atpD, glnII, and recA gene sequences, Rhizobium and Ensifer were further classified into four different genotypes: Rhizobium radiobacter, Rhizobium sp., Rhizobium rosettiformans, and Ensifer meliloti. The differences in architecture and functioning of the rhizobial genomes and, to a lesser extent, environment diversification helped explain the diversity of tested strains. The tested strains exhibited similar symbiotic feature when inoculated onto M. sativa cvs. Gannong Nos. 3 and 9 and Qingshui plants for the clustering feature of their parameter values. An obvious symbiotic divergence of rhizobial strains was observed in M. sativa cvs. Longzhong and WL168HQ plants because of the scattered parameter values. Their symbiotic divergence differed according to alfalfa varieties, which indicated that the sensitivity of different alfalfa varieties to rhizobial strains may differ. Most of the tested strains exhibited plant growth-promoting traits including phosphate solubilization and production of indole-3-acetic acid (IAA) when colonizing plant tissues and soil.     

苜蓿中华根瘤菌desA基因功能的鉴定

为研究苜蓿中华根瘤菌脂肪酸脱饱和酶desA基因在不饱和脂肪酸合成、共生结瘤固氮以及应对逆境胁迫中的功能,为高效利用苜蓿中华根瘤菌提供理论依据,本文通过异体遗传互补和脂肪酸组成薄层层析,分析SmdesA编码蛋白是否具有脱饱和酶的活性并参与不饱和脂肪酸的合成,构建SmdesA的缺失突变株和互补菌株,比较各菌株在不同逆境胁迫条件下的生长速率以及回接宿主植物后与紫花苜蓿共生结瘤的能力.结果表明SmdesA不能互补大肠杆菌CY57中EcfabA的突变,但具有将饱和脂肪酸脱饱和形成不饱和的棕榈油酸和十八碳烯酸的能力.另外,SmdesA缺失突变对苜蓿中华根瘤菌的脂肪酸组成影响不大,但会显著影响低温和高盐条件下菌株的生长速率以及与紫花苜蓿共生结瘤的能力.我们推测,SmdesA参与的脱饱和途径可能是苜蓿中华根瘤菌不饱和脂肪酸合成的补偿途径,其编码的蛋白DesA不是不饱和脂肪酸合成的关键酶,但在应对逆境胁迫和共生结瘤中具有重要的生物学功能.     

Genetic diversity of indigenous tropical fast-growing rhizobia isolated from soybean nodules

This study characterized genetically 30 fast-growing rhizobial strains isolated from nodules of Asian and modern soybean genotypes that had been inoculated with soils from disparate regions of Brazil. Analyses by rep-PCR (ERIC and REP) and RAPD indicated a high level of genetic diversity among the strains. The RFLP-PCR and sequencing analysis of the 16S rRNA genes indicated that none of the strains was related to Sinorhizobium (Ensifer) fredii, whereas most were related to Rhizobium tropici (although they were unable to nodulate Phaseolus vulgaris) and to Rhizobium genomic species Q. One strain was related to Rhizobium sp. OR 191, while two others were closely related to Agrobacterium (Rhizobium) spp.; furthermore, symbiotic effectiveness with soybean was maintained in those strains. Five strains were related to Bradyrhizobium japonicum and B. elkanii, with four of them being similar to strains carried in Brazilian inoculants, therefore modifications in physiological properties, as a shorter doubling time might have resulted from adaptation to local conditions. Phospholipid fatty acid analysis (PFLA) was less precise in delineating taxonomic relationships. The strains fit into eight Nod-factor profiles that were related to rhizobial species, but not to N₂-fixation capacity or competitiveness. The data obtained highlight the diversity and promiscuity of rhizobia in the tropics, being capable of nodulating exotic legumes and might reflect ecological strategies to survive in N-poor soils; in addition, the diversity could also represent an important source of efficient and competitive rhizobial strains for the tropics. Putative new rhizobial species were detected only in undisturbed soils. Three species (R. tropici, B. japonicum and B. elkanii) were found under the more sustainable management system known as no-till, while the only species isolated from soils under conventional till was R. tropici. Those results emphasize that from the moment that agriculture was introduced into undisturbed soils rhizobial diversity has changed, being drastically reduced when a less sustainable soil management system was adopted.