Mimosine produced by the tree-legume Leucaena provides growth advantages to some Rhizobium strains that utilize it as a source of carbon and nitrogen

Growth of most Rhizobium strains is inhibited by mimosine, a toxin found in large quantities in the seeds, foliage and roots of plants of the genera Leucaena and Mimosa. Some Leucaena-nodulating strains of Rhizobium can degrade mimosine (Mid⁺) and are less inhibited by mimosine in the growth medium than the mimosine-nondegrading (Mid^-) strains. Ten Mid⁺ strains were identified that did not degrade 3-hydroxy-4-pyridone (HP), a toxic intermediate of mimosine degradation. However, mimosine was completely degraded by these strains and HP was not accumulated in the cells when these strains were grown in a medium containing mimosine as the sole source of carbon and nitrogen. The mimosine-degrading ability of rhizobia is not essential for nodulation of Leucaena species, but it provides growth advantages to Rhizobium strains that can utilize mimosine, and it suppresses the growth of other strains that are sensitive to this toxin.     

Phenotypic and genotypic characterisation of root nodule bacteria nodulating Millettia pinnata (L.) Panigrahi, a biodiesel tree

Aims

Milletia pinnata is a leguminous tropical tree that produces seed oil suitable for biodiesel and is targeted to be planted on marginal land associated with nitrogen poor soil. This study aimed to identify effective rhizobia species for M. pinnata.

Methods

Soil samples were collected from M. pinnata grown in Kununurra, Australia. Rhizobia were trapped, characterised and sequenced for 16S rRNA, atpD, dnaK and recA genes.

Results

Forty isolates tolerated pH 7 – 9, temperatures 29 – 37 °C, salinity below 1 % NaCl, and had optimal growth on mannitol, arabinose or glutamate as a single carbon source, a few grew on sucrose and none grew on lactose. Inoculation of isolates increased shoot dry weight of M. pinnata’s seedlings in nitrogen minus media. Slow-growing isolates were closely related to Bradyrhizobium yuanmingense, Bradyrhizobium sp. DOA10, Bradyrhizobium sp. ORS305 and B. liaoningense LMG 18230^T. The fast-growing isolates related to Rhizobium sp. 8211, R. miluonense CCBAU 41251^T, R miluonense CC-B-L1, Rhizobium sp. CCBAU 51330 and Rhizobium sp. 43015

Conclusions

Millettia pinnata was effectively nodulated by slow-growing isolates related to Bradyrhizobium yuanmingense, Bradyrhizobium sp. DOA10 Bradyrhizobium sp. ORS305, B. liaoningense LMG 18230^T and fast-growing isolates related Rhizobium sp. 8211, R. miluonense, Rhizobium sp. CCBAU 51330 and Rhizobium sp. 43015     

Pyruvate carboxylase is involved in metabolism of mimosine by <Emphasis Type="Italic">Rhizobium</Emphasis> sp. strain TAL1145

The objective of this study was to determine the role of midK, which encodes a protein similar to pyruvate carboxylase, in mimosine degradation by Rhizobium sp. strain TAL1145. The midK gene is located downstream of midR in the cluster of genes for mimosine degradation in Rhizobium sp. strain TAL1145. The midK mutants of TAL1145 degraded mimosine slower than the wild-type. These mutants could utilize pyruvate as a source of carbon, indicating that there is another pyruvate carboxylase (pyc) gene in TAL1145. Two classes of clones were isolated from the library of TAL1145 by complementing a pyc mutant of Rhizobium etli, one class contained midK, while the other carried pyc. Both midK and pyc of TAL1145 complemented the midK mutant for mimosine degradation, and also the R. etli pyc mutant for pyruvate utilization. The midK-encoded pyruvate carboxylase was required for an efficient conversion of mimosine into 3-hydroxy-4-pyridone (HP). Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Jonathan D. Awaya and Panlada Tittabutr contributed equally to this work.     

Use of the Chrome Azurol S Agar Plate Technique To Differentiate Strains and Field Isolates of Rhizobium leguminosarum biovar trifolii

Identification of Rhizobium and Bradyrhizobium strains and especially of indigenous isolates continues to be one of the major difficulties associated with competition studies. Because there is no universally accepted method, the method of choice depends on preference, experience, and equipment. Here, an agar plate technique was used to distinguish strains and field isolates of Rhizobium leguminosarum biovar trifolii to provide a basis for identifying nodule occupants in further competition studies. A rapid plate technique, based on differential growth characteristics, complements other techniques such as serological reactions, particularly when antisera cross-react with nonhomologous strains. The technique involves culturing strains and isolates on chrome azurol S agar. Although similar responses were observed among some strains, the response was highly reproducible and was considered an ideal complementary technique used in conjunction with serological procedures. Strains with similar responses could often be differentiated by varying media components, such as the source of carbon.     

Isolation of Insertion Sequence ISRLdTAL1145-1 from a Rhizobium sp. (Leucaena diversifolia) and Distribution of Homologous Sequences Identifying Cross-Inoculation Group Relationships

Insertion sequence (IS) element ISRLdTAL1145-1 from Rhizobium sp. (Leucaena diversifolia) strain TAL 1145 was entrapped in the sacB gene of the positive selection vector pUCD800 by insertional inactivation. A hybridization probe prepared from the whole 2.5-kb element was used to determine the distribution of homologous sequences in a diverse collection of 135 Rhizobium and Bradyrhizobium strains. The IS probe hybridized strongly to Southern blots of genomic DNAs from 10 rhizobial strains that nodulate both Phaseolus vulgaris (beans) and Leucaena leucocephala (leguminous trees), 1 Rhizobium sp. that nodulates Leucaena spp., 9 R. meliloti (alfalfa) strains, 4 Rhizobium spp. that nodulate Sophora chrysophylla (leguminous trees), and 1 nonnodulating bacterium associated with the nodules of Pithecellobium dulce from the Leucaena cross-inoculation group, producing distinguishing IS patterns for each strain. Hybridization analysis revealed that ISRLdTAL1145-1 was strongly homologous with and closely related to a previously isolated element, ISRm USDA1024-1 from R. meliloti, while restriction enzyme analysis found structural similarities and differences between the two IS homologs. Two internal segments of these IS elements were used to construct hybridization probes of 1.2 kb and 380 bp that delineate a structural similarity and a difference, respectively, of the two IS homologs. The internal segment probes were used to analyze the structures of homologous IS elements in other strains. Five types of structural variation in homolog IS elements were found. The predominate IS structural type naturally occurring in a strain can reasonably identify the strain's cross-inoculation group relationships. Three IS structural types were found in Rhizobium species that nodulate beans and Leucaena species, one of which included the designated type IIB strain of R. tropici (CIAT 899). Weak homology to the whole IS probe, but not with the internal segments, was found with two Bradyrhizobium japonicum strains. The taxonomic and ecological implications of the distribution of ISRLdTAL1145-1 are discussed.     

A Study of Mimosine Toxicity in Plants

Mimosine and 3,4-dihydroxypyridine are inhibitory to the growthof mung bean seedlings. Mimosine inhibition could be reversedby tyrosine or nicotinic acid, but it was reversed by pyridoxalphosphate or ferrous ions. Inhibition due to dihydroxypyridinealso could be reversed by ferrous ions, but not by pyridoxalphosphate. The possible mechanisms of growth inhibition arediscussed. An enzyme is present in extracts of Leucaena seedlings thatdegrades mimosine stoichiometrically to 3,4-dihydroxypyridine,pyruvate, and ammonia. An enzymic breakdown of mimosine is alsocatalysed by extracts of mung bean seedlings. The occurencein Leucaena of dichrostachinic acid and a C-S-Iyase which degradesit to a thiol derivative, pyruvate, and ammonia were described;the latter enzyme was shown to be distinct from the enzyme-splittingmimosine. The substrate specificity, pH optimum, co-factor requirements,and inhibitors of this mimosine C-N-Iyase were investigated.     

Growth performance of,and mimosine excretion by,young chicks fed on leucaena leucocephala

Growth performance and mimosine output in excreta have been measured in young chicks fed on diets containing three forms of Leucaena leaf meal (LLM) imported from Malawi. In one of the experiments, the effects of supplementation of LLM with selected metal ions and enzymes were also measured. Growth and efficiency of food utilisation were similar in groups fed on a control soya bean meal diet or a basal diet containing 150 g kg^?1 of the whole leaf (WL) form of LLM. Supplementation of this basal LLM diet with FeSO₄ or Al₂(SO₄)₃ had little effect on growth or efficiency of food utilisation, but the ratio of mimosine output to mimosine ingested (MO/MI) increased from 0.781 to 0.881 and 1.003 on addition of FeSO₄ and Al₂(SO₄)₃, respectively. Growth performance was unaffected by the addition of an enzyme mixture (containing cellulase, acid and neutral proteases and alpha amylase) to the basal LLM diet. In the second experiment, two other forms of LLM were also tested: a ground leaf (GL) form, and a powder produced by grinding pellets of Leucaena leaf (GP). The WL and GP forms were each added at 50 and 100 g kg^?1 diet, and the GL form at 100 g kg^?1 diet. These diets permitted daily intakes of mimosine ranging from 80 to 350 mg per replicate of 4 chicks. In comparison with control groups, growth performance was not impaired significantly in any of the LLM-fed groups. Mimosine excretion was high in all LLM-fed groups, the GP form inducing a maximum MO/MI ratio of 0.924.     

Synergistic interaction of Rhizobium leguminosarum bv. viciae and arbuscular mycorrhizal fungi as a plant growth promoting biofertilizers for faba bean (Vicia faba L.) in alkaline soil

Egyptian soils are generally characterized by slightly alkaline to alkaline pH values (7.5–8.7) which are mainly due to its dry environment. In arid and semi-arid regions, salts are less concentrated and sodium dominates in carbonate and bicarbonate forms, which enhance the formation of alkaline soils. Alkaline soils have fertility problems due to poor physical properties which adversely affect the growth and the yield of crops. Therefore, this study was devoted to investigating the synergistic interaction of Rhizobium and arbuscular mycorrhizal fungi for improving growth of faba bean grown in alkaline soil. A total of 20 rhizobial isolates and 4 species of arbuscular mycorrhizal fungi (AMF) were isolated. The rhizobial isolates were investigated for their ability to grow under alkaline stress. Out of 20 isolates 3 isolates were selected as tolerant isolates. These 3 rhizobial isolates were identified on the bases of the sequences of the gene encoding 16S rRNA and designated as Rhizobium sp. Egypt 16 (HM622137), Rhizobium sp. Egypt 27 (HM622138) and Rhizobium leguminosarum bv. viciae STDF-Egypt 19 (HM587713). The best alkaline tolerant was R. leguminosarum bv. viciae STDF-Egypt 19 (HM587713). The effect of R. leguminosarum bv. viciae STDF-Egypt 19 and mixture of AMF (Acaulospora laevis, Glomus geosporum, Glomus mosseae and Scutellospora armeniaca) both individually and in combination on nodulation, nitrogen fixation and growth of Vicia faba under alkalinity stress were assessed. A significant increase over control in number and mass of nodules, nitrogenase activity, leghaemoglobin content of nodule, mycorrhizal colonization, dry mass of root and shoot was recorded in dual inoculated plants than plants with individual inoculation. The enhancement of nitrogen fixation of faba bean could be attributed to AMF facilitating the mobilization of certain elements such as P, Fe, K and other minerals that involve in synthesis of nitrogenase and leghaemoglobin. Thus it is clear that the dual inoculation with Rhizobium and AMF biofertilizer is more effective for promoting growth of faba bean grown in alkaline soils than the individual treatment, reflecting the existence of synergistic relationships among the inoculants.     

Influence of ε-caprolactam on growth and physiology of environmental bacteria

ε-Caprolactam was found to have an effect on ecologically important soil bacteria. It inhibited the growth of several Bacillus sp. and Rhizobium sp. but cells of Arthrobacter sp. were able to grow in the presence of caprolactam. Sphingomonas sp. lost its inherent capacity to produce extracellular polymer (EPS) if grown in medium containing caprolactam. In the case of raw domestic sewage, the diversity of native bacteria was diminished in presence of caprolactam. Polluted sea water yielded predominantly one type of caprolactam-degrading bacteria of the genus Achromobacter. These cells efficiently utilized up to 10 g caprolactam/L as the sole source of carbon and nitrogen in synthetic medium even in the presence of 20 g NaCl/L. Compared to cells of Arthrobacter sp., cells of Achromobacter sp. accumulated high amount of 6-aminocaproic acid due to degradation of caprolactam. When using caprolactam as sole source of carbon and nitrogen, Achromobacter cells showed unique physiological ability to produce EPS upon prolonged incubation in solid medium and in broth with low phosphate (C:N:P ratio 100:20:0.05). Hydrolyzed cell-free EPS had glucose as its major component though the only substrate provided in the medium for growth was caprolactam.     

Symbiotic effectiveness and plant growh promoting traits in some Rhizobium strains isolated from Phaseolus vulgaris L.

The ability of Rhizobia to colonize roots of certain legumes and promote their growth has been proven previously. In this study the symbiotic efficiency of 47 Rhizobium strains with 6 common bean cultivars was evaluated under greenhouse condition. Fourteen strains showed the best symbiotic efficiency, whereas some isolates could not induce nodules on host plants. The ability of fourteen superior strains to solubilize phosphorus and zinc and to produce auxin, HCN and siderohores was evaluated in the laboratory assays. Rhizobium strain Rb102 produced the highest amount of auxin (14.2?mg?l^?1) in the medium containing l-tryptophan. None of the isolates were able to solubilize ZnO and ZnCO₃ on solid medium but in liquid medium some of them had negligible solubilization. The highest P solubility in liquid and solid medium was observed in strains Rb113 and Rb130, respectively. Strain Rb102 produced the highest amount of siderophores. None of the isolates were able to produce HCN. This study showed that there was a great diversity between the strains of Rhizobium in terms of their plant growth promoting traits symbiotic efficiency which supports the importance of screening rhizobia for selecting the most efficient strains. The genetic diversity of the isolates was analyzed by PCR–RFLP of the 16S rDNA. Our rhizobia were clustered into 10 groups showing high levels of diversity.     

Competition Among Rhizobium spp. for Nodulation of Leucaena leucocephala in Two Tropical Soils

The successful nodulation of legumes by a Rhizobium strain is determined by the competitive ability of that strain against the mixture of other native and inoculant rhizobia. Competition among six Leucaena rhizobial strains in single and multistrain inoculants were studied. Field inoculation trials were conducted in an oxisol and a mollisol soil, both of which contained indigenous Leucaena-nodulating rhizobia. Strain-specific fluorescent antibodies were used for the identification of the strains in Leucaena nodules. Mixtures of three recommended inoculum strains for Leucaena spp. (TAL82, TAL582, and TAL1145) were used in peat-based inocula either alone or with one of the three other strains isolated from the sites, B213, B214, and B215. Each of these latter three strains was also used as single-strain inocula to study their competition with the native rhizobia in the two soil systems. In the oxisol soil, strains B213 and B215, when used as single-strain inocula, outcompeted the native rhizobia and formed 92 and 62% of the nodules, respectively. Strain B214 was the least competitive in oxisol soil, where it formed 30% of the nodules, and the best in mollisol soil, where it formed 70% of the nodules. The most successful competitor for nodulation in multistrain inocula was strain TAL1145, which outcompeted native and other inoculum Leucaena rhizobia in both soils. None of the strains in single or multistrain inoculants was capable of completely overcoming the resident rhizobia, which formed 4 to 70% of the total nodules in oxisol soil and 12 to 72% in mollisol soil. No strong relationship was detected between the size of the rhizosphere population of a strain and its successful occupation of nodules.     

Putative roles of bacteria in the carbon and nitrogen cycles in a tropical peat swamp forest

Tropical peat swamp forests are waterlogged, acidic, anoxic and oligotrophic ecosystems. They are important terrestrial carbon pools that help mitigating global warming through carbon sequestration in peat. This study aimed at investigating putative roles of bacteria in the carbon and nitrogen cycles in North Selangor peat swamp forest, Malaysia. Whole-genome sequencing was performed on four bacterial isolates using Illumina NextSeq 500 to decipher their genetic information while Gen III Microplate (Biolog) was applied to verify carbon source utilization. The isolates were identified as Dyella sp. strain C9, Dyella sp. strain C11, Klebsiella sp. strain C31 and Paraburkholderia sp. strain C35. Both Dyella spp. and Paraburkholderia sp. strain C35 were likely novel species while Klebsiella sp. strain C31 was a different strain of the type species, Klebsiella pneumoniae. Both genomic and bioassay results suggested the involvement of the isolates in the degradation of lignocellulose, carbohydrates, sugar alcohols, organic acids and aromatic compounds. The isolates could potentially perform methanotrophy, which helps to mitigate methane emissions from tropical peatlands. In addition, the isolates also contained genes encoding enzymes for nitrite, nitrate and nitric oxide reduction, as well as dissimilatory nitrate reduction to ammonium, which retains the nitrogen in the ecosystems. The results generated insights into potential functions of bacteria in the energy production and nutrient cycling of tropical peatlands, which are essential for the sustainability of high biomass and biodiversity in these ecosystems.     

Effect of Rhizobium–Azospirillum coinoculation on nitrogen fixation and yield of two contrasting Phaseolus vulgaris L. genotypes cultivated across different environments in Cuba

Azospirillum spp. have shown potential to enhance nodulation and plant growth of legumes when coinoculated with Rhizobium. The effect of Azospirillum on the Rhizobium-legume symbiosis is, however, dependent on the host genotype used. Previous greenhouse experiments identified two genotypes of common bean (Phaseolus vulgaris L.), BAT477 and DOR364, contrasting in nodulation response to Azospirillum when coinoculated with Rhizobium. Genetic analysis revealed a genetic basis (Quantitative Trait Loci) on the bean genome related to the differential responsiveness to Azospirillum between the two bean genotypes. In this study, on-station and on-farm field experiments in different regions in Cuba were conducted to evaluate the agronomic relevance of the differences in response to Azospirillum–Rhizobium coinoculation between the two genotypes BAT477 and DOR364. It was observed that Azospirillum–Rhizobium coinoculation as compared to single Rhizobium inoculation increased the amount of fixed nitrogen and the yield of DOR364 across all sites. For BAT477, on the contrary, a negative effect of Azospirillum–Rhizobium coinoculation on yield and nitrogen fixation was observed on most of the sites as compared to single Rhizobium inoculation. The modified stability regression equations resulting from this study may contribute to predict how a combination of genotype and inoculum will perform at a certain environmental setting. This study highlights the importance of genotype × inocula interactions in agricultural outputs and establishes a link between greenhouse phenotype, genetic background and performance in the field.     

Growth of Octopine-Catabolizing Pseudomonas spp. under Octopine Limitation in Chemostats and Their Potential To Compete with Agrobacterium tumefaciens

The growth characteristics of five octopine-catabolizing pseudomonads have been determined in batch and continuous cultures. All five strains belonged to rRNA homology group I and showed a more psychrotrophic growth pattern than did Agrobacterium tumefaciens B6 and ATCC 15955. In chemostats limited by octopine, either as the source of carbon and nitrogen or the sole source of nitrogen, maximum specific growth rates and substrate affinities were lower than those in chemostats limited by glutamate. These growth dynamics were similar to those observed for Agrobacterium strains B6 and ATCC 15955 even though the catabolic genes and pathways are believed to be different in the two genera. An analysis of the yields in octopine-limited chemostats indicated that the use of octopine as the sole source of carbon and nitrogen was grossly inefficient. Octopine and presumably lysopine and octopinic acid provided a better source of nitrogen than of carbon. One of the Pseudomonas fluorescens strains, E175D, was able to produce its highest yield on octopine as a nitrogen source. Competition models formulated on pure culture parameters indicated that two of the Pseudomonas spp. would dominate A. tumefaciens B6 and ATCC 15955 when in simple competition for octopine as a limiting substrate.     

Isolation and Partial Characterization of Bacterial Strains on Low Organic Carbon Medium from Soils Fertilized with Different Organic Amendments

A total of 720 bacterial strains were isolated from soils with four different organic amendment regimes on a low organic carbon (low-C) agar medium (10 µg C ml^?1) traditionally used for isolation of oligotrophs. Organic amendments in combination with field history resulted in differences in dissolved organic carbon contents in these soils. There were negative correlations between total and dissolved organic carbon content and the number of isolates on low-C agar medium, whereas these correlations were absent for bacterial strains isolated from the same soil on high-C agar medium (1,000 µg C ml^?1). Repeated transfers (up to ten times) of the isolates from low-C agar medium to fresh low- and high-C agar media were done to test for exclusive growth under oligotrophic conditions. The number of isolates exclusively growing under oligotrophic conditions dropped after each subsequent transfer from 241 after the first to 98 after the third transfer step. Identification on the basis of partial 16S rRNA gene sequences revealed that most of the 241 isolates (as well as the subset of 98 isolates) belong to widespread genera such as Streptomyces, Rhizobium, Bradyrhizobium, and Mesorhizobium, and the taxonomic composition of dominant genera changed from the first transfer step to the third. A selected subset of 17 isolates were further identified and characterized for exclusive growth on low-C agar medium. Two isolates continued to grow only on low-C agar medium up to the tenth transfer step and matched most closely with Rhizobium sullae and an uncultured bacterium on the basis of the almost full-length 16S rRNA gene. It was concluded that the vast majority of strains which are isolated on low-C agar media belong to the trophic group of microorganisms adapted to a “broad range” of carbon concentrations, including well-known and widespread bacterial genera. Oligotrophy is a physiological, not a taxonomic property, and can only be identified by cultural means so far. We showed that true oligotrophs that are unable to grow on high carbon media are rare and belong to genera that also contain broad-range and copiotrophic strains.     

midD-encoded ‘rhizomimosinase’ from Rhizobium sp. strain TAL1145 is a C–N lyase that catabolizes L-mimosine into 3-hydroxy-4-pyridone, pyruvate and ammonia

Rhizobium sp. strain TAL1145 catabolizes mimosine, which is a toxic non-protein amino acid present in Leucaena leucocephala (leucaena). The objective of this investigation was to study the biochemical and catalytic properties of the enzyme encoded by midD, one of the TAL1145 genes involved in mimosine degradation. The midD-encoded enzyme, MidD, was expressed in Escherichia coli, purified and used for biochemical and catalytic studies using mimosine as the substrate. The reaction products in the enzyme assay were analyzed by HPLC and mass spectrometry. MidD has a molecular mass of ~45 kDa and its catalytic activity was found to be optimal at 37 °C and pH 8.5. The major product formed in the reaction had the same retention time as that of synthetic 3-hydroxy-4-pyridone (3H4P). It was confirmed to be 3H4P by MS/MS analysis of the HPLC-purified product. The K _m, V _max and K _cat of MidD were 1.27 × 10^?4 mol, 4.96 × 10^?5 mol s^?1 mg^?1, and 2,256.05 s^?1, respectively. Although MidD has sequence similarities with aminotransferases, it is not an aminotransferase because it does not require a keto acid as the co-substrate in the degradation reaction. It is a pyridoxal-5′-phosphate (PLP)-dependent enzyme and the addition of 50 μM hydroxylamine completely inhibited the reaction. However, the supplementation of the reaction with 0.1 μM PLP restored the catalytic activity of MidD in the reaction containing 50 μM hydroxylamine. The catalytic activity of MidD was found to be specific to mimosine, and the presence of its structural analogs including l-tyrosine, l-tryptophan and l-phenylalanine did not show any competitive inhibition. In addition to 3H4P, we also identified pyruvate and ammonia as other degradation products in equimolar quantities of the substrate used. The degradation of mimosine into a ring compound, 3H4P with the release of ammonia indicates that MidD of Rhizobium sp. strain TAL1145 is a C–N lyase.     

Genetic analysis of nitrogen fixation in a tropical fast-growing Rhizobium

The Rhizobium strain ORS571, which is associated with the tropical legume Sesbania rostrata, has the property of growing in the free-living state at the expense of ammonia or N₂ as sole nitrogen source. Five mutants, isolated as unable to form colonies on plates under conditions of nitrogen fixation, were studied. All of them, which appear as Fix^- in planta, are nif mutants. With mutant 5740, nitrogenase activity of the crude extract was restored by addition of pure Mo-Fe protein of Klebsiella pneumoniae. A 13-kb BamHI DNA fragment from the wild-type strain, which hybridized with a probe carrying the nifHDK genes of K. pneumoniae, was cloned in vector pRK290 to yield plasmid pRS1. The extent of homology between the probe and the BamHI fragment was estimated at 4 kb and hybridization with K. pneumoniae nifH, nifK, and possibly nifD was detected. The pRS1 plasmid was introduced into the sesbania rhizobium nif mutants. Genetic complementation was observed with strain 5740(pRS1) both in the free-living state and in planta. It thus appears that biochemistry and genetics of nitrogen fixation in this particular Rhizobium strain can be performed with bacteria grown under non-symbiotic conditions.     

Genes encoding a fructose-1,6-bisphosphate aldolase and a fructose-1,6-bisphosphatase are present within the gene cluster for mimosine degradation in Rhizobium sp. strain TAL1145

Rhizobium sp. strain TAL1145 can catabolize mimosine, a toxic amino acid produced by the tree-legume leucaena. The mid and pyd genes involved in mimosine degradation in TAL1145 are located in two clusters within a 25-kb region in the chromosome, which was cloned in plasmid pUHR263. A 5.5-kb EcoRI fragment, located between the mid and pyd genes in pUHR263, was characterized by sequencing and transposon-insertion mutagenesis and six open reading frames (ORF) were identified. Based on high homologies with other known proteins and conserved signature domains, ORF1 and ORF2 were identified as fba and fbp genes, encoding fructose-1,6-bisphosphate aldolase (FBA) and fructose-1,6-bisphosphatase (FBP), respectively. The fba mutant showed a slightly reduced growth rate compared to TAL1145 while the fbp mutant did not show any growth defects. Both mutants could catabolize mimosine and formed normal nitrogen-fixing nodules on leucaena, suggesting that these genes are not involved in mimosine degradation and symbiosis.     

Rhizobium qilianshanense sp. nov., a novel species isolated from root nodule of Oxytropis ochrocephala Bunge in China

During a study of the diversity and phylogeny of rhizobia isolated from root nodules of Oxytropis ochrocephala grown in the northwest of China, four strains were classified in the genus Rhizobium on the basis of their 16S rRNA gene sequences. These strains have identical 16S rRNA gene sequences, which showed a mean similarity of 94.4 % with the most closely related species, Rhizobium oryzae. Analysis of recA and glnA sequences showed that these strains have less than 88.1 and 88.7 % similarity with the defined species of Rhizobium, respectively. The genetic diversity revealed by ERIC-PCR fingerprinting indicated that the isolates correspond to different strains. Strain CCNWQLS01^T contains Q-10 as the predominant ubiquinone. The major fatty acids were identified as feature 8 (C18: 1ω7c and/or C18: 1ω6c; 67.2 %). Therefore, a novel species Rhizobium qilianshanense sp. nov. is proposed, and CCNWQLS01^T (= ACCC 05747^T = JCM 18337^T) is designated as the type strain.