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ABSTRACT: A highly supported maximum-likelihood species phylogeny for the genus Bradyrhizobium was inferred from a supermatrix obtained from the concatenation of partial atpD, recA, glnII, and rpoB sequences corresponding to 33 reference strains and 76 bradyrhizobia isolated from the nodules of Glycine max (soybean) trap plants inoculated with soil samples from Myanmar, India, Nepal, and Vietnam. The power of the multigene approach using multiple strains per species was evaluated in terms of overall tree resolution and phylogenetic congruence, representing a practical and portable option for bacterial molecular systematics. Potential pitfalls of the approach are highlighted. Seventy-five of the isolates could be classified as B. japonicum type Ia (USDA110/USDA122-like), B. liaoningense, B. yuanmingense, or B. elkanii, whereas one represented a novel Bradyrhizobium lineage. Most Nepalese B. japonicum Ia isolates belong to a highly epidemic clone closely related to strain USDA110. Significant phylogenetic evidence against the monophyly of the of B. japonicum I and Ia lineages was found. Analysis of their DNA polymorphisms revealed high population distances, significant genetic differentiation, and contrasting population genetic structures, suggesting that the strains in the Ia lineage are misclassified as B. japonicum. The DNA polymorphism patterns of all species conformed to the expectations of the neutral mutation and population equilibrium models and, excluding the B. japonicum Ia lineage, were consistent with intermediate recombination levels. All species displayed epidemic clones and had broad geographic and environmental distribution ranges, as revealed by mapping climate types and geographic origins of the isolates on the species tree.
Applied and environmental microbiology 10/2008; 74(22):6987-96. · 3.69 Impact Factor
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ABSTRACT: A genotype effect on associative (rhizosphere) N2-fixation was observed with two cultivars of Sorghum bicolor (nutans) with a maximum rate of 8 μmol C2H4 · h−1 · plant−1 in one genotype compared to 0.9 μmol in the other. Characteristics of the high fixing genotype were a reduced transpiration rate, a lower number of stomata and increased root exudate production per gram root dry weight with higher concentration of dicarboxylic acids. The bacterial rhizosphere composition revealed a three times higher number of N2-fixing bacteria, a tenfold reduction of actinomycetes and a threefold reduction of Arthrobacter associated with the high fixing cultivar compared to the low fixing genotype. From these and other plant rhizospheres two new nitrogen fixing bacteria, Pseudomonas stutzeri and Erwinia herbicola, were characterized. With the N2-fixing bacteria Azospirillum brasilense and Klebsiella pneumoniae an enhancement of specific nitrogenase activity by aromatic compounds, for example phenolics, the herbicide alachlor and the insecticide carbofuran was demonstrated. An oscillating nitrogenase activity in Azospirillum brasilense under microaerobic conditions was found, resulting from an encystation and deencystation under those conditions. Experiments with wheat roots demonstrated that reduced oxygen tensions, essential for a maximum rhizosphere N2-fixation, reduced root growth significantly and altered the N-metabolism of the roots.Einfluß von pflanzlichen, mikrobiellen und Boden-Faktoren auf die N2-Fixierung in der RhizosphreEin Genotypen-Effekt auf die assoziative (Rhizosphre) N2-Fixierung wurde an zwei Sorten von Sorghum bicolor (nutans) mit einer maximalen Fixierungsrate von 8 μmol C2H4 · h−1 · Pflanze−1 in einem Genotyp verglichen mit 0.9 μmol in dem anderen Genotyp nachgewiesen. Merkmale des hoch N2-fixierenden Genotyps waren eine reduzierte Transpirationsrate und eine niedrigere Zahl von Stomata, eine erhöhte Wurzelexsudation pro Gramm Wurzeltrockengewicht mit einem erhöhten Anteil von Dicarbon-suren. Die Untersuchung der bakteriellen Rhizosphrenzusammensetzung zeigte eine dreifach erhöhte Zahl von N2-fixierenden Bakterien bei der hochfixierenden Sorte, eine Reduktion auf ein Zehntel bei den Actinomyceten und auf ein Drittel bei Arthrobacter-Anea. Aus diesen und anderen Rhizosphrenböden wurden als neue N2-fixierende Bakterienarten Pseudomonas stutzeri und Erwinia herbicola charakterisiert. Bei den diazotrophen Bakterien Azospirillum brasilense und Klebsiella pneumoniae wurde eine spezifische Steigerung der Nitrogenaseaktivitt durch aromatische Verbindungen wie Phenole, das Herbizid'Alachlor und das Insektizid Carbofuran nachgewiesen. Als weitere Variable, die die Rhizosphren-N2-Fixierung verndert, wurde eine oszillierende Nitrogenase-aktivitt in Azospirillum brasilense unter mikroaeroben Bedingungen gefunden, die aufgrund von aufeinander folgenden Encystierungen und De-encystierungen erklrt wird. Untersuchungen an Getreidewurzeln zeigten, daß niedrige Sauerstoffpartialdrücke eine wichtige Voraussetzung für eine maximale Rhizosphren-N2-Fixierung sind, aber gleichzeitig das Wurzelwachstum erheblich reduzierten und den N-Metabolismus in den Wurzeln vernderten.
Journal of Plant Nutrition and Soil Science 01/2007; 152(2):231 - 236. · 1.60 Impact Factor
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ABSTRACT: The legume root nodules are the site of biological nitrogen fixation in the legume—rhizobium symbiosis. They are a structure unique to this symbiosis and morphologically as well as physiologically distinct from other plant organs. We will present evidence that the formation of nodules (nodulation) may serve for ecotoxicological evaluation of contaminated soil. Substances affecting the macro- and/or microsymbionts vitality, such as certain heavy metals or polycyclic aromatic hydrocarbons (PAHs), reduce nodulation before visible damage of the plant occurs. Tests were performed in petri dishes that were planted with alfalfa seedlings and inoculated with Rhizobium meliloti. Mineral agar served as substrate. Test substances were added to the mineral agar before pouring it into petri dishes (application of heavy metals), or were sprayed as etheral solutions onto the surface of solidified medium (application of PAHs). For investigation of contaminated soil, the soil material was first mixed with mineral agar and then poured into petri dishes. Appropriate controls were made by extracting an aliquot of soil with methylene chloride to remove hazardous substances. Data from stressed plants were taken after 15 days of cultivation on contaminated substrate. A dose-responsive decrease in nodulation was found after application of CdCl2, NaAsO2, fluoranthene, and other PAHs. PAH-contaminated soil (75 mg/kg) caused only a slight reduction in nodulation of alfalfa. But when PAHs were extracted and restored into the extracted soil material, toxicity and uptake of PAHs into roots and shoots dramatically increased. Toxicity and uptake of PAHs were also enhanced by reducing the amount of humic matter added to the test system. © by John Wiley & Sons, Inc.
Environmental Toxicology and Water Quality 05/2006; 10(2):127 - 133.
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ABSTRACT: Rhizobium tropici CIAT899 is highly tolerant to several environmental stresses and is a good competitor for nodule occupancy of common bean plants in acid soils. Random transposon mutagenesis was performed to identify novel genes of this strain involved in symbiosis and stress tolerance. Here, we present a genetic analysis of the locus disrupted by the Tn5 insertion in mutant 899-PV9, which lead to the discovery of sycA, a homolog of the ClC family of chloride channels and Cl-/H+ exchange transporters. A nonpolar deletion in this gene caused serious deficiencies in nodule development, nodulation competitiveness, and N2 fixation on Phaseolus vulgaris plants, probably due to its reduced ability to invade plant cells and to form stable symbiosomes, as judged by electron transmission microscopy. A second gene (olsC), found downstream of sycA, is homologous to aspartyl/asparaginyl beta-hydroxylases and modifies two species of ornithine-containing lipids in vivo, presumably by hydroxylation at a still-unknown position. A mutant carrying a nonpolar deletion in olsC is symbiotically defective, whereas overexpressed OlsC in the complemented strain provokes an acid-sensitive phenotype. This is the first report of a ClC homolog being essential for the establishment of a fully developed N2-fixing root nodule symbiosis and of a putative beta-hydroxylase that modifies ornithine-containing membrane lipids of R. tropici CIAT899, which, in turn, are contributing to symbiotic performance and acid tolerance.
Molecular Plant-Microbe Interactions 12/2005; 18(11):1175-85. · 4.43 Impact Factor
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ABSTRACT: Highly diverse Bradyrhizobium strains nodulate genistoid legumes (brooms) in the Canary Islands, Morocco, Spain and the Americas. Phylogenetic analyses of ITS, atpD, glnII and recA sequences revealed that these isolates represent at least four distinct evolutionary lineages within the genus, namely Bradyrhizobium japonicum and three unnamed genospecies. DNA-DNA hybridization experiments confirmed that one of the latter represents a new taxonomic species for which the name Bradyrhizobium canariense is proposed. B. canariense populations experience homologous recombination at housekeeping loci, but are sexually isolated from sympatric B. japonicum bv. genistearum strains in soils of the Canary Islands. B. canariense strains are highly acid-tolerant, nodulate diverse legumes in the tribes Genisteae and Loteae, but not Glycine species, whereas acid-sensitive B. japonicum soybean isolates such as USDA 6(T) and USDA 110 do not nodulate genistoid legumes. Based on host-range experiments and phylogenetic analyses of symbiotic nifH and nodC sequences, the biovarieties genistearum and glycinearum for the genistoid legume and soybean isolates, respectively, were proposed. B. canariense bv. genistearum strains display an overlapped host range with B. japonicum bv. genistearum isolates, both sharing monophyletic nifH and nodC alleles, possibly due to the lateral transfer of a conjugative chromosomal symbiotic island across species. B. canariense is the sister species of B. japonicum, as inferred from a maximum-likelihood Bradyrhizobium species phylogeny estimated from congruent glnII+recA sequence partitions, which resolves eight species clades. In addition to the currently described species, this phylogeny uncovered the novel Bradyrhizobium genospecies alpha and beta and the photosynthetic strains as independent evolutionary lineages. The type strain for B. canariense is BTA-1(T) (=ATCC BAA-1002(T)=LMG 22265(T)=CFNE 1008(T)).
International journal of systematic and evolutionary microbiology 04/2005; 55(Pt 2):569-75. · 2.27 Impact Factor
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ABSTRACT: A combination of population genetics and phylogenetic inference methods was used to delineate Bradyrhizobium species and to uncover the evolutionary forces acting at the population-species interface of this bacterial genus. Maximum-likelihood gene trees for atpD, glnII, recA, and nifH loci were estimated for diverse strains from all but one of the named Bradyrhizobium species, and three unnamed "genospecies," including photosynthetic isolates. Topological congruence and split decomposition analyses of the three housekeeping loci are consistent with a model of frequent homologous recombination within but not across lineages, whereas strong evidence was found for the consistent lateral gene transfer across lineages of the symbiotic (auxiliary) nifH locus, which grouped strains according to their hosts and not by their species assignation. A well resolved Bayesian species phylogeny was estimated from partially congruent glnII+recA sequences, which is highly consistent with the actual taxonomic scheme of the genus. Population-level analyses of isolates from endemic Canarian genistoid legumes based on REP-PCR genomic fingerprints, allozyme and DNA polymorphism analyses revealed a non-clonal and slightly epidemic population structure for B. canariense isolates of Canarian and Moroccan origin, uncovered recombination and migration as significant evolutionary forces providing the species with internal cohesiveness, and demonstrated its significant genetic differentiation from B. japonicum, its sister species, despite their sympatry and partially overlapped ecological niches. This finding provides strong evidence for the existence of well delineated species in the bacterial world. The results and approaches used herein are discussed in the context of bacterial species concepts and the evolutionary ecology of (brady)rhizobia.
Molecular Phylogenetics and Evolution 02/2005; 34(1):29-54. · 3.61 Impact Factor
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ABSTRACT: Rhizobium tropici CIAT899 is highly acid tolerant and a good competitor for Phaseolus vulgaris nodule occupancy at low pH values. Using Tn5 mutagenesis, we identified an operon required for acid tolerance and nodulation competitiveness. The insertion was mapped to the 5' end of atvA, encoding a product with high sequence identity to the agro-bacterial AcvB virulence protein. Complementation analyses indicated that atvA is an ortholog of acvB, both genes being required for acid tolerance. A Ser/Ala substitution in the LIPASE_SER motif of AtvA resulted in an acid sensitive Fix+ but very poorly competing strain, demonstrating that Ser-313 is essential for AtvA function. atvA is the second gene in an operon that is transcriptionally upregulated by acid shock. The acid-responsive promoter was mapped to a 469-bp intergenic region located upstream of lpiA, the first gene in the operon. lpiA-like genes are found in several alpha, beta, and gamma Proteobacteria that interact with eukaryotic host cells, and they are predicted to encode membrane proteins related to the FmtC/MprF family from low G+C Firmicutes. The latter proteins are involved in resistance to cationic antimicrobial peptides. A nonpolar deletion in lpiA caused a sevenfold decrease in relative nodulation competitiveness.
Molecular Plant-Microbe Interactions 03/2003; 16(2):159-68. · 4.43 Impact Factor
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ABSTRACT: The novel lipochitin oligosaccharide (LCOs) structures produced by Rhizobium etli KIM5s were characterized using a nanoHPLC reverse-phase system coupled to an ion-trap mass spectrometer. This technique was shown to be more sensitive for structural elucidation of LCOs than previously used mass spectrometric methods. The structures of the LCOs of R. etli KIM5s, the majority containing six monosaccharide residues, differed from those synthesized by all other rhizobia analyzed to date. In addition, novel structures in which the chitin backbone was deacetylated at one or more GlcNAc moieties were found as minor compounds. The difference in host range of this strain compared to that of other known bean microsymbionts is discussed.
Carbohydrate Research 08/2002; 337(13):1193-202. · 2.33 Impact Factor
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ABSTRACT: Viele chemalige Rüstungsstandorte sind mit dem Sprengstoff 2,4,6-Trinitrotoluol sowie dessen Derivaten und Abbauprodukten
kontaminiert. Solche Nitroaromaten besitzen ein hohes human- und ökotoxisches Potential. Ein Transfer dieser Schadstoffe in
Pflanzen ist grundsätzlich möglich und auch unter Freilandbedingungen nachweisbar.
In dieser Arbeit wurde eine Schadstoffanalyse des Rhizosphärenbodens — des unmittelbaren Einflußbereichs der Wurzel — vorgenommen,
um die Wirkung von Pflanzen auf TNT-kontaminierte Böden näher zu charakterisieren. Als Versuchspflanzen dienten Buschbohnen
(Phaseolus vulgaris).
Für zwei unterschiedlich hoch belastete Freilandflächen konnte in den jeweiligen Rhizosphärenböden eine Abreicherung des extrahierbaren
TNT um den Faktor 5 bis 6 nachgewiesen werden. In der Wurzel kam es demgegenüber auf der hoch belasteten Fläche zu einer Anreicherung
von Nitroaromaten, wobei sich Aminodinitrotoluole als nachweisbare Hauptschadstoffe bestätigen ließen.
Der Abreicherungsbefund verweist auf einen potentiellen Einsatz von Pflanzen im Rahmen der biologischen Sanierung rüstungsspezifisch
belasteter Böden.
Many sites of former ammunition plants are contaminated with the explosive 2,4,6-trinitrotoluene, its derivatives and reduction
products. Such nitroaromatic compounds have a high human and ecotoxic potential. Plant uptake of these organic pollutants
is known and could be confirmed under field conditions.
In this paper rhizospheric soil was analyzed to characterize the effect of vegetation on TNT-contaminated soil. Common beans
(Phaseolus vulgaris) were used as test plants. The experiments were performed on two areas with different TNT concentrations.
The data reveal a significant decrease of extractable TNT in rhizospheric soil by a factor of 5 to 6 during the vegetation
period. On the area with high TNT concentration, an enrichment of nitroaromatic compounds in root tissue was found compared
to the rhizospheric soil. Aminodinitrotoluene could be confirmed as the primary pollutant in the root.
These results refer to a potential application of plants to biological remediation of soil contaminated with ammunition-specific
pollutants.
Umweltwissenschaften und Schadstoff-Forschung 05/1996; 8(5):243-247.
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ABSTRACT: The fate of the explosive 2,4,6-TNT in plants is of major interest. Therefore, a method was developed to analyse TNT and derivatives
in plant tissue. The method was utilized to investigate the uptake and metabolism of TNT inMedicago sativa andAllium schoenoprasum grown in hydroponic cultures containing TNT levels of 0.1 to 10 mg/1. Detectable concentrations of nitrotoluenes were significantly
higher inAllium schoenoprasum than inMedicago sativa. The uptake of TNT in plants was directly related to the initial TNT level. The principal nitroaromatic components in roots
and shoots of both plant species were identified as 4-ADNT and 2-ADNT in equal amounts, with substantially less TNT.
Environmental Science and Pollution Research 04/1994; 1(4):229-233. · 2.65 Impact Factor
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ABSTRACT: Flavins in different compartments of effective nodules fromGlycine max cv Maple Arrow xBradyrhizobium japonicum strains were studied by spectrophotometry and chromatographic techniques. Flavins in the peribacteroid space were riboflavin (80%) and FMN (20%), as identified by TLC and HPLC. Flavin concentrations in the soybean root nodule cytoplasm, in the symbiosome space (PBS) and in the cytosol of bacteroids were monitored between 20 and 40 days post infection (d.p.i.) Between the 20th and 29th d.p.i. an at least four times higher flavin/protein ratio was found in PBS of effective nodules compared with the nodule cytoplasm. Between nitrogenase activity in the free-living state and bacterial flavin accumulation, no correlation could be observed. Flavin accumulation is not restricted to an effective symbiosis, as indicated by the analysis of ineffective nodules with strainB. japonicum RH-31 Marburg. Flavin accumulation is absent in uninfected soybean root tissue and in free-living rhizobia, thus indicating that flavin accumulation is a result of symbiotic interaction. Flavin accumulation is also missing in nodules with a hypersensitive response against the bacteria.
Current Microbiology 01/1992; 25(5):269-273. · 1.82 Impact Factor
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ABSTRACT: A new type of dialysis culture system was used to cocultivate different bacterial species under oligotrophic conditions. In a mineral medium with no added carbon source, the testedRhizobium leguminosarum bv.phaseoli strains outgrew anArthrobacter strain, but not anEnterobacter strain. TheRhizobium strains had generation times of approximately 4.3 h,Enterobacter agglomerans 1.5 h, andArthrobacter globiformis more than 30 h. After adding glutamate to the system to a final concentration of either 2 or 10 M, we found significant (P < 0.05)="" differences="" in="" growth="" rates="">Rhizobium leguminosarum bv.phaseoli strains. Strains Ciat 899 and Kim5s showed maximal growth at the 10 M glutamate concentration with generation times of 2.8 and 3.1 h, respectively. In contrast, strains Ciat 895 and CE3 reacted with increased generation times of 3.8 and 4.3 h. However, when immunofluorescence techniques were used to follow populations of the strains in an acid and a neutral tropical soil, no significant strain differences in decline were observed. Thus, strain survival in unplanted soils was not related to growth rates under oligotrophic conditions. In contrast to this, the ability to grow more or less rapidly under carbon-limited conditions was positively correlated with the known competitiveness ofRhizobium leguminosarum bv.phaseoli strains in legume host nodulation.
Current Microbiology 08/1991; 23(3):159-163. · 1.82 Impact Factor
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ABSTRACT: For Bradyrhizobium japonicum, the chemotactic and the nod gene-inducing effects of hydroxycinnamic acids and two of their derivatives were compared with those of isoflavonoids. Only the hydroxycinnamic acids were strong chemoattractants, while the other substances tested were chemotactically inactive. Besides the known nod gene induction by isoflavonoids, a weak nod gene induction by coniferyl alcohol, chlorogenic acid, and ferulic acid was found.
Applied and Environmental Microbiology 02/1991; 57(1):316-9. · 3.83 Impact Factor
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ABSTRACT: Wyerone is the predominant phytoalexin produced byVicia faba. At a concentration of 100 M, wyerone prolonged the lag phase before the onset of exponential growth of different strains ofRhizobium leguminosarum andBradyrhizobium japonicum. The response to wyerone was dose dependent, with a shorter lag phase occurring at lower concentrations. Wyerone was only moderately inhibitory towards the phytopathogenic bacteriumErwinia carotovora cv. atroseptica. HPLC analysis of the medium during bacterial growth indicated thatRhizobium leguminosarum was able to metabolize wyerone. The identification of the product as hydroxyester wyerol was confirmed by GC/MS analysis.
Current Microbiology 01/1991; 23(3):153-157. · 1.82 Impact Factor
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Bulletin of Environmental Contamination and Toxicology 09/1990; 45(4):619-626. · 1.02 Impact Factor
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ABSTRACT: Host-cell cytoplasm from soybean plants infected with the peribacteroid membrane (PBM)-building Rhizobium japonicum strain 61-A-101 (effective, N(2)-fixing) had much higher choline kinase activity than cytoplasm from either uninfected tissue or tissue infected with the non-PBM-building (ineffective, non-N(2)-fixing) strain 61-A-24. Ion-exchange chromatography showed that both types of nodule and root tissue possessed constitutive choline kinase I activity that had a K(m) for choline of approximately 150 muM. The nodules of the effective symbiosis had another activity, choline kinase II (K(m) = 81 muM). Nondenaturing and NaDodSO(4) electrophoresis revealed no multimeric subunit structure of the two enzyme forms but did show the molecular sizes for choline kinase I, 58-59 kDa, and choline kinase II, 60 kDa. Choline kinase I and II and pI values of 8.1 and 8.5, respectively, and two-dimensional gel electrophoresis of whole cytoplasm from control and infected tissue showed a spot corresponding to choline kinase II only in the case of the effective symbiosis, whereas both tissue types had spots corresponding to choline kinase I. Choline kinase II is presumed to be encoded by the plant as neither free-living nor symbiotic (bacteroid) forms of the prokaryote showed any choline kinase activity.
Proceedings of the National Academy of Sciences 03/1986; 83(3):659-63. · 9.68 Impact Factor
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ABSTRACT: A cultivation system with simultaneous growth of six bacterial cultures in separate bags in dialysis culture was developed. In a medium with no added carbon source (one half concentrated Hoagland solution, water deionized and distilled), cell number ofRhizobium japonicum increased during a 7 day period by a factor of 35, whereas the number ofEnterobacter aerogenes cells decreased to one half. With a concentration of 100 nM succinate as an additional carbon source in the inflow,Rhizobium japonicum 61-A-101 cell number increased by a factor of 50 during an 8 day period, whereas cell number ofEnterobacter cloacae NCTC 10005 only doubled and ofEnterobacter aerogenes NCTC 10006 decreased. At 10 mM concentration of succinate in the inflow, doubling time the twoEnterobacter strains was about 12 h, compared to about 24 h for theRhizobium japonicum strain. Varying the succinate concentration from 10 mM to 100 nM in the inflow,Rhizobium japonicum 61-A-101 surpassed theEnterobacter aerogenes strains in the growth rate between 1 mM and 100 M succinate in the inflowing medium. Three otherRhizobium japonicum strains (fix+ and fix-) did grow with a similar rate as strain 61-A-101 at very low concentrations of substrate. Growth rates for the strains were confirmed by protein data per culture. Growing in competition with twoPseudomonas strains,Rhizobium japonicum RH 31 Marburg (fix-) did overgrow alsoPseudomonas fluorescens, was however outgrown byPseudomonas putida. In utilizing low concentrations of a14C labelled organic acid (malonate), three strains ofRhizobium japonicum left 2–4 times smaller amounts of14C in the medium than two species ofPseudomonas and two species ofArthrobacter.
Archives of Microbiology 07/1985; 142(3):223-228. · 1.43 Impact Factor
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ABSTRACT: Bacteroids in ineffective (nitrogenase negative) nodules of Glycine max, infected with Rhizobium japonicum 61-A-24, as compared to those in effective nodules are characterized by reduced specific activities of alanine dehydrogenase to 15%, of 3-hydroxybutyrate dehydrogenase to 50%, and an increase of glutamine synthetase to 400%. In the plant cytoplasm of ineffective nodules, glutamine synthetase activity is reduced to 10–30%, glutamate dehydrogenase to 50–70%, and the aspartate aminotransferase and alanine aminotransferase are enhanced to 120–200%, depending on the age of the nodules. The total pool of soluble amino acids is reduced to 52 mol per g nodule fresh weight, as compared to 186 mol in effective nodules, with a replacement of asparagine (42 mol% of the amino acids) by an unknown amino compound. This compound is absent in nitrogenase, repressed and derepressed, free-living Rhizobium japonicum cells and in the uninfected root tissue. In nitrogenase derepressed, as compared to the repressed free-living cells of Rhizobium japonicum 61-A-101, arginine shows the most obvious change with a reduction to less than one tenth. The ultrastructure of the ineffective nodule is different from the effective organ even in the early stages. The membrane envelopes of the infection vacuoles are decomposing in heavily infected cells within 18 to 20 d after infection. In lightly infected cells very large vacuoles develop with only a few bacteroids inside. No close associations of cristae-rich mitochondria with amyloplasts are observed as in effective nodules. The uninfected cells keep their large starch granules even 40 d after infection. Some poly--hydroxybutyrate accumulation in the bacteroids is observed but only in the early stages, and it is almost absent in old nodules (40 d). At this age the infected cells are obviously compressed by uninfected cells, whereas in effective nodules with nitrogenase activity and leghaemoglobin formation, the infected cells have a much higher osmotic pressure than the neighbouring uninfected cells.
Planta 12/1979; 147(4):320-329. · 3.00 Impact Factor
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ABSTRACT: Plants of Glycine max var. Caloria, infected as 14 d old seedlings with a defined titre of Rhizobium japonicum 3Il b85 in a 10 min inoculation test, develop a sharp maximum of nitrogenase activity between 17 and 25 d after infection. This maximum (143 nmol C2H4 h-1 mg nodule fresh weight-1), expressed as per mg nodule or per plant is followed by a 15 d period of reduced nitrogen fixation (20–30% of peak activity). 11 d after infection the first bacteroids develop as single cells inside infection vacuoles in the plant cells, close to the cell wall and infection threads. As a cytological marker for peak multiplication of bacteroids and for peak N2-fixation a few days later the association of a special type of nodule mitochondria with amyloplasts is described. 20 d after inoculation, more than 80% of the volume of infected plant cells is occupied by infection vacuoles, mostly containing only one bacteroid. The storage of poly--hydroxybutyrate starts to accumulate at both ends of the bacteroids. Non infected plant cells are squeezed between infected cells (25d), with infection vacuoles containing now more than two (up to five) bacteroids per section. Bacteroid development including a membrane envelope is also observed in the intercellular space between plant cells. 35 d after infection, more than 50% of the bacteroid volume is occupied by poly--hydroxybutyrate. The ultrastructural differentiation is discussed in relation to some enzymatic data in bacteroids and plant cell cytoplasm during nodule development.
Planta 12/1977; 141(2):169-177. · 3.00 Impact Factor
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ABSTRACT: Components of the peribacteroid space (PBS) and the bacteroid periplasm (PPS) of soybean root nodules infected with Bradyrhizobium japonicum 61-A-101 were isolated by sucrose density gradient centrifugation. Non activity of either glucose-6 phosphate (glucose-6P)-dehydrogenase or alanine dehydrogenase, marker enzymes for the nodule and bacteroid cytoplasms, could be detected in the PBS and only minor activity of the bacteroid marker β-hydroxybutyrate dehydrogenase (0.06% of bacteroid activity). One dimensional SDS-PAGE of the PBS and the PPS revealed similar patterns for these two fractions. In the PBS fraction some proteins were quantitatively enhanced. The same enzyme activities were present in the PBS and the PPS but several enzymes had much higher specific activities in the PBS than in the PPS. The specific activities of α-glucosidase and α-trehalase in the PBS were 34-fold, 25-fold and 8-fold higher, respectively, than in the nodule cytoplsm. No activity was founds in root cytoplasm. The enzymes are considered as symbiosis specific proteins, enriched in the peribacteroid space. The estimated isoelectric point for α-glucosidase was 7.2 and for α-trehalase, 5.6 α-glucosidase and α-trehalase were not present in root cytoplasm. The possible functions of these PBS enzymes in symbiotic nitrogen fixation are discussed.
Plant Science.
