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PubMedDate not supplied

Activation of nif gene expression in Azotobacter by the nifA gene product of Klebsiella pneumoniae.

Structural similarity of nitrogenase, the enzyme responsible for biological nitrogen fixation, from various diazotrophic bacteria has been shown by intergeneric biochemical complementation of component proteins in vitro, DNA and protein sequence analysis, and DNA hybridization between nif (nitrogen fixation) structural genes from Klebsiella pneumoniae and genomic sequences from other nitrogen-fixing bacteria. Despite nitrogenase homologies, little is known about similarities among nif regulatory mechanisms although repression of nitrogenase synthesis by NH4+ and O2 occurs in most diazotrophs. In K. pneumoniae, the ntr (gln) genes concerned with regulation of nitrogen metabolism control expression of the nifLA operon whose products act as repressor (nifL) and activator (nifA) of the seven other nif transcriptional units. Here we report that the nifA gene product of K. pneumoniae can activate expression of nif genes in both Azotobacter vinelandii and Azotobacter chroococcum, organisms whose aerobic physiology contrasts with that of the facultative K. pneumoniae.

Azotobacter↗

PubMed1999

Site-directed mutagenesis analysis of draTG genes and their downstream region from Azospirillum brasilense Yu62.

draT and draG genes are involved in posttranslational regulation of nitrogenase activity of Azospirillum brasilense Yu62. Both genes and their downstream region were mutagenized by Kmr cassette insertions. Analysis of mutations introduced into the draTG region on the A. brasilense Yu62 chromosome showed that mutants affected in draT were incapable of regulating nitrogenase activity in response to ammonium. In contrast, a mutant with an insertion in draG was still capable of ADP-ribosylating dinitrogenase reductase in response to ammonium but was no longer able to recover activity after ammonium depletion. Analysis of mutations introduced into the draTG downstream region (the mutagenized site is about 2 kb downstream from draG) showed that the mutant had higher nitrogenase activity than the wild strain while growing in nitrogen-free medium and medium with 2 mmol/L ammonium. These results reveal that there is no gene required for nitrogen fixation in this mutagenized region, but it is possible that there are genes which play a role in regulating nitrogen fixation. The results of monitoring the expression of transcriptional nifH-lacZ gene fusion in the mutant YZ4 showed that the transcriptional regulation of nif gene in the mutant YZ4 was the same as that in the wild type.

Azospirillum brasilense↗

PubMed2001

Isolation and 16S rRNA sequence analysis of the beneficial bacteria from the rhizosphere of rice.

The present study deals with the isolation of plant growth promoting rhizobacteria (PGPR) from rice (variety NIAB IRRI-9) and the beneficial effects of these inoculants on two Basmati rice varieties. Nitrogen-fixing activity (acetylene-reduction activity) was detected in the roots and submerged shoots of field-grown rice variety NIAB IRRI-9. Estimation of the population size of diazotrophic bacteria by ARA-based MPN (acetylene reduction assay-based most probable number) in roots and shoots indicated about 10(5)-10(6) counts/g dry weight at panicle initiation and grain filling stages. Four bacterial isolates from rice roots and shoots were obtained in pure culture which produced phytohormone indoleacetic acid (IAA) in the growth medium. Among these, three isolates S1, S4, and R3 reduced acetylene to ethylene in nitrogen-free semi-solid medium. Morphological and physiological characteristics of the isolates indicated that three nitrogen-fixing isolates S1, S4, and R3 belonged to the genus Enterobacter, while the non-fixing isolate R8 belonged to the genus Aeromonas. 16S rRNA sequence of one isolate from root (R8) and one isolate from shoot (S1) was obtained which confirmed identification of the isolates as Aeromonas veronii and Enterobacter cloacae, respectively. The 1517-nucleotide-long sequence of the isolate R8 showed 99% similarity with Aeromonas veronii (accession No. AF099023) while partial 16S rRNA sequence (two stretches of total 1271 nucleotide length) of S1 showed 97% similarity with the sequence of Enterobacter cloacae (accession No. AJ251469). The seedlings of two rice varieties Basmati 385 and Super Basmati were inoculated with the four bacterial isolates from rice and one Azospirillum brasilense strain Wb3, which was isolated from wheat. In the rice variety Basmati 385, maximum increase in root area and plant biomass was obtained in plants inoculated with Enterobacter S1 and Azospirillum Wb3, whereas in the rice variety Super Basmati, inoculation with Enterobacter R3 resulted in maximum increase of root area and plant biomass. Nitrogen fixation was quantified by using 15N isotopic dilution method. Maximum fixation was observed in Basmati 385 with the inoculants Azospirillum Wb3 and Enterobacter S1 where nearly 46% and 41% of the nitrogen was derived from atmosphere (%Ndfa), respectively. In general, higher nitrogen fixation was observed in variety Basmati 385 than in Super Basmati, and different bacterial strains were found more effective as inoculants for the rice varieties Basmati 385 and Super Basmati.

Aeromonas↗

PubMed1999

Studies on the roles of GlnK and GlnB in regulating Klebsiella pneumoniae NifL-dependent nitrogen control.

In Klebsiella pneumoniae, nitrogen fixation (nif) genes are regulated in response to fixed nitrogen and oxygen. The activity of the nif-specific transcriptional activator NifA is modulated by NifL, which mediates both oxygen and nitrogen control. The signal transduction protein GlnK is required to relieve the inhibitory effect of NifL on NifA that occurs when the intracellular N status is high and in a wild-type cell, the action of GlnK cannot be substituted by the structurally related protein PII. We have studied the modulation of NifA activity by NifL in an heterologous system in which the host organism is Escherichia coli. Using a DeltaglnB, DeltaglnK mutant, we have shown that the modulation of NifA activity by NifL is dependent on the concentration of GlnK in the cell and that when overproduced, PII can substitute for GlnK. Furthermore, our data suggest that PII can counteract the positive action of GlnK in relieving NifL-dependent inhibition of NifA activity. This negative effect of PII may be physiologically important in establishing repression of nif gene expression when the intracellular nitrogen status rises.

Bacterial Proteins↗

PubMed1996

Oxygen taxis and proton motive force in Azospirillum brasilense.

The microaerophilic nitrogen-fixing bacterium Azospirillum brasilense formed a sharply defined band in a spatial gradient of oxygen. As a result of aerotaxis, the bacteria were attracted to a specific low concentration of oxygen (3 to 5 microM). Bacteria swimming away from the aerotactic band were repelled by the higher or lower concentration of oxygen that they encountered and returned to the band. This behavior was confirmed by using temporal gradients of oxygen. The cellular energy level in A. brasilense, monitored by measuring the proton motive force, was maximal at 3 to 5 microM oxygen. The proton motive force was lower at oxygen concentrations that were higher or lower than the preferred oxygen concentration. Bacteria swimming toward the aerotactic band would experience an increase in the proton motive force, and bacteria swimming away from the band would experience a decrease in the proton motive force. It is proposed that the change in the proton motive force is the signal that regulates positive and negative aerotaxis. The preferred oxygen concentration for aerotaxis was similar to the preferred oxygen concentration for nitrogen fixation. Aerotaxis is an important adaptive behavioral response that can guide these free-living diazotrophs to the optimal niche for nitrogen fixation in the rhizosphere.

Azospirillum brasilense↗

PubMed2002

Redundancy in periplasmic binding protein-dependent transport systems for trehalose, sucrose, and maltose in Sinorhizobium meliloti.

We have identified a cluster of six genes involved in trehalose transport and utilization (thu) in Sinorhizobium meliloti. Four of these genes, thuE, -F, -G, and -K, were found to encode components of a binding protein-dependent trehalose/maltose/sucrose ABC transporter. Their deduced gene products comprise a trehalose/maltose-binding protein (ThuE), two integral membrane proteins (ThuF and ThuG), and an ATP-binding protein (ThuK). In addition, a putative regulatory protein (ThuR) was found divergently transcribed from the thuEFGK operon. When the thuE locus was inactivated by gene replacement, the resulting S. meliloti strain was impaired in its ability to grow on trehalose, and a significant retardation in growth was seen on maltose as well. The wild type and the thuE mutant were indistinguishable for growth on glucose and sucrose. This suggested a possible overlap in function of the thuEFGK operon with the aglEFGAK operon, which was identified as a binding protein-dependent ATP-binding transport system for sucrose, maltose, and trehalose. The K(m)s for trehalose transport were 8 +/- 1 nM and 55 +/- 5 nM in the uninduced and induced cultures, respectively. Transport and growth experiments using mutants impaired in either or both of these transport systems show that these systems form the major transport systems for trehalose, maltose, and sucrose. By using a thuE'-lacZ fusion, we show that thuE is induced only by trehalose and not by cellobiose, glucose, maltopentaose, maltose, mannitol, or sucrose, suggesting that the thuEFGK system is primarily targeted toward trehalose. The aglEFGAK operon, on the other hand, is induced primarily by sucrose and to a lesser extent by trehalose. Tests for root colonization, nodulation, and nitrogen fixation suggest that uptake of disaccharides can be critical for colonization of alfalfa roots but is not important for nodulation and nitrogen fixation per se.

ATP-Binding Cassette Transporters↗

PubMed1992

Role of integration host factor in stimulating transcription from the sigma 54-dependent nifH promoter.

In a wide variety of nitrogen-fixing organisms among the Purple Bacteria (large division of Gram-negative bacteria) the nitrogen fixation (nif) operons are transcribed by an alternative holoenzyme form of RNA polymerase, sigma 54-holoenzyme. Transcription depends on the activator protein NIFA (nitrogen fixation protein A), which catalyzes isomerization of closed complexes between this polymerase and a promoter to transcriptionally productive open complexes. NIFA-mediated activation of transcription from the nifH promoter of Klebsiella pneumoniae is greatly stimulated by the integration host factor IHF, which binds to a site between the upstream binding site for NIFA and the promoter, and bends the DNA. IHF fails to stimulate activation of transcription from this promoter by another activator of sigma 54-holoenzyme, NTRC (nitrogen regulatory protein C), which lacks a specific binding site in the nifH promoter region. As predicted, if the IHF-induced bend facilitates interaction between NIFA and sigma 54-holoenzyme, substitution of an NTRC-binding site for the NIFA-binding site allowed IHF to stimulate NTRC-mediated activation of transcription from the nifH promoter. The stimulation was of the same order of magnitude as that for NIFA in the native configuration of the promoter-regulatory region (up to 20-fold). With purified NTRC and the substitution construct we could demonstrate that stimulation by IHF in a purified transcription system was comparable to that in a crude coupled transcription-translation system, indicating that the stimulation in the crude system could be accounted for by IHF. The IHF stimulation was observed on linear as well as supercoiled templates, indicating that the geometric requirements are relatively simple. We have attempted to visualize the arrangement of proteins on DNA fragments carrying the nifH promoter-regulatory region of K. pneumoniae by electron microscopy. IHF stimulated NIFA-mediated activation of transcription from the nifH and nifD promoters of Bradyrhizobium japonicum and less so from the nifH promoters of Rhizobium meliloti and Thiobacillus ferrooxidans, consistent with previous observations that stimulation is greatest at promoters that are weak binding sites for sigma 54-holoenzyme in closed complexes.

Bacterial Proteins↗

PubMed1980

[Applied gene technology using biological N2-fixation for an example (author's transl)].

About biological nitrogen fixation most information on the genetic and enzymatic level is known from investigations with Klebsiella pneumoniae. The gene region for N2-fixation (nif) was transferred from K.pneumoniae to the enterobacterium Escherichia coli. After acquisition of nif E.coli is able to fix nitrogen. Plasmid pRD1 was constructed which carries the nif-region. Using genetic engineering techniques the exact location of nif on pRD1 was determined. Subsequently the nif-region comprising 7 operons with 14 known genes was cloned on a multicopy plasmid vector. The use of the Klebsiella-nif-region in biotechnology is discussed. Difficulties arising in experiments designed to construct N2-fixing plants are mentioned.

Cloning, Molecular↗

PubMed2000

Altered exopolysaccharides of Bradyrhizobium japonicum mutants correlate with impaired soybean lectin binding, but not with effective nodule formation.

The exact mechanism(s) of infection and symbiotic development between rhizobia and legumes is not yet known, but changes in rhizobial exopolysaccharides (EPSs) affect both infection and nodule development of the legume host. Early events in the symbiotic process between Bradyrhizobium japonicum and soybean (Glycine max [L.] Merr.) were studied using two mutants, defective in soybean lectin (SBL) binding, which had been generated from B. japonicum 2143 (USDA 3I-1b-143 derivative) by Tn5 mutagenesis. In addition to their SBL-binding deficiency, these mutants produced less EPS than the parental strain. The composition of EPS varied with the genotype and with the carbon source used for growth. When grown on arabinose, gluconate, or mannitol, the wild-type parental strain, B. japonicum 2143, produced EPS typical of DNA homology group I Bradyrhizobium, designated EPS I. When grown on malate, strain 2143 produced a different EPS composed only of galactose and its acetylated derivative and designated EPS II. Mutant 1252 produced EPS II when grown on arabinose or malate, but when grown on gluconate or mannitol, mutant 1252 produced a different EPS comprised of glucose, galactose, xylose and glucuronic acid (1:5:1:1) and designated EPS III. Mutant 1251, grown on any of these carbon sources, produced EPS III. The EPS of strain 2143 and mutant 1252 contained SBL-binding polysaccharide. The amount of the SBL-binding polysaccharide produced by mutant 1252 varied with the carbon source used for growth. The capsular polysaccharide (CPS) produced by strain 2143 during growth on arabinose, gluconate or mannitol, showed a high level of SBL binding, whereas CPS produced during growth of strain 2143 on malate showed a low level of SBL binding. However, the change in EPS composition and SBL binding of strain 2143 grown on malate did not affect the wild-type nodulation and nitrogen fixation phenotype of 2143. Mutant 1251, which produced EPS III, nodulated 2 d later than parental strain 2143, but formed effective, nitrogen-fixing tap root nodules. Mutant 1252, which produced either EPS II or III, however nodulated 5-6 d later and formed few and ineffective tap root nodules. Restoration of EPS I production in mutant 1252 correlated with restored SBL binding, but not with wild-type nodulation and nitrogen fixation.

Arabinose↗

PubMed1980

Effect of rice plants on nitrogenase activity of flooded soils.

In samples of flooded soil containing blue-green algae (cyanobacteria), the presence of rice plants did not influence the nitrogenase activity of the algae. Nitrogenase activity of heterotrophic bacteria was enhanced by the presence of rice plants, but this activity was not affected by changes in plant density. The rate of nitrogen fixation in the rhizosphere, however, varied significantly among the 16 rice varieties tested. A simple method was devised to test the nitrogen-fixing activity in the root zone of rice varieties, and data were obtained showing marked differences in the activities of the 16 varieties. In tests of two varieties with dissimilar rates of nitrogen fixation in their rhizospheres, the variety which had the greater root weight and lesser shoot weight and which supported greater methane formation had the greater nitrogenase activity.

Journal Article↗

PubMed2005

Isolation and identification of nitrogen-fixing bacilli from plant rhizospheres in Beijing region.

AIMS: To isolate and identify nitrogen-fixing bacilli from the plant rhizospheres in Beijing region of China. METHODS AND RESULTS: A total of 29 isolates were selectively obtained from the rhizospheres of wheat, maize, ryegrass and willow based on their growth on nitrogen-free medium and their resistance to 100 degrees C for 10 min. Of the 29 isolates, seven had nifH gene determined by PCR amplification. The seven isolates were found to belong to the genera Bacillus and Paenibacillus based on phenotypic characterization, 16S rDNA sequence, G+C content and DNA-DNA hybridization. Isolates T1 and W5 were identified as Bacillus cereus and Bacillus marisflavi respectively. Isolates G1, C4 and C5 were identified as Bacillus megaterium. Isolate G2 was identified as Paenibacillus polymyxa and isolate T7 as Paenibacillus massiliensis. CONCLUSIONS: This study suggests that nifH gene could be detected in the both genera Bacillus and Paenibacillus. These degenerate primers for nifH gene fragment used in this study were shown to be useful for identifying nitrogen-fixing bacilli. SIGNIFICANCE AND IMPACT OF THE STUDY: It is the first demonstration that nitrogen fixation exists in B. marisflavi and P. massiliensis and the first report of the sequences of the nifH gene from B. megaterium and B. cereus. The nitrogen-fixing bacilli obtained in this study will be used in our future research for investigating the mechanisms of nitrogen fixation in bacilli.

Bacillus↗

PubMed2004

Phytoestrogen signaling and symbiotic gene activation are disrupted by endocrine-disrupting chemicals.

Some organochlorine pesticides and other synthetic chemicals mimic hormones in representatives of each vertebrate class, including mammals, reptiles, amphibians, birds, and fish. These compounds are called endocrine-disrupting chemicals (EDCs). Similarly, hormonelike signaling has also been observed when vertebrates are exposed to plant chemicals called phytoestrogens. Previous research has shown the mechanism of action for EDCs and phytoestrogens is as unintended ligands for the estrogen receptor (ER). Although pesticides have been synthesized to deter insects and weeds, plants produce phytoestrogens to deter herbivores, as attractant cues for insects, and as recruitment signals for symbiotic soil bacteria. Our data present the first evidence that some of the same organochlorine pesticides and EDCs known to disrupt endocrine signaling through ERs in exposed wildlife and humans also disrupt the phytoestrogen signaling that leguminous plants use to recruit Sinorhizobium meliloti soil bacteria for symbiotic nitrogen fixation. Here we report that a variety of EDCs and pesticides commonly found in agricultural soils interfere with the symbiotic signaling necessary for nitrogen fixation, suggesting that the principles underlying endocrine disruption may have more widespread biological and ecological importance than had once been thought.

Cell Communication↗

PubMed1983

Effect of temperature on nitrogenase functioning in cowpea nodules.

Nitrogenase (EC 1.7.99.2) activity of a cowpea (Vigna unguiculata (L.) Walp cv Caloona) symbiosis formed with a Rhizobium strain (176A27) lacking uptake hydrogenase and maintained under conditions of a 12-hour day at an air temperature of 30 degrees C (800-1000 microeinsteins per square meter per second) and a 12-hour night at an air temperature of 20 degrees C showed a marked diurnal variation in ratio of nitrogen fixed to hydrogen evolved. As little as 0.3 micromole nitrogen was fixed per micromole hydrogen evolved in the photoperiod versus up to 0.6 in the dark period. In plants maintained under the same diurnal illumination regime but at constant (day and night) air temperature (30 degrees C), this difference was abolished and a relatively constant ratio of nitrogen fixed to hydrogen evolved (around 0.3 micromole per micromole) was observed day and night. Exposure of nodulated roots to a range of temperatures maintained for 2 hours in a single photoperiod indicated that, whereas hydrogen evolution increased with increasing temperature from 15 degrees C to a maximum around 35 degrees C, nitrogen fixation was largely unaffected over this temperature range. Both functions of the enzyme declined sharply at temperatures above 38 degrees C. A similar general response of nitrogen fixation to root temperature was observed in glasshouse-grown, sand-cultured plants maintained under a range of temperatures (from 15 to 35 degrees C) for a 14-day period in mid vegetative growth. The effect of temperature on the proportion of electrons allocated to proton reduction compared with nitrogen reduction showed a linearly increasing relationship (correlation coefficient = 0.96) between 15 degrees C and 47 degrees C.

Journal Article↗

PubMed2005

Quantitative analysis of nifH genes and transcripts from aquatic environments.

The availability of fixed inorganic nitrogen often plays a fundamental role in regulating primary production in both aquatic and terrestrial ecosystems. Because biological nitrogen fixation is an important source of nitrogen in marine environments, the study of N2-fixing microorganisms is of fundamental importance to our understanding of global nitrogen and carbon cycles. Quantitative molecular tools have made it possible to examine uncultivated N2-fixing microorganisms directly in the environment. Currently, we are using quantitative polymerase chain reaction (PCR; Q-PCR) and quantitative reverse transcriptase PCR (Q-RT-PCR) to study the ecology and gene expression of N2-fixing bacteria in aquatic environments. Using these methods, we discovered that specific estuarine diazotrophs have distinct nonrandom distributions and that some diazotrophs in the open ocean have different diel patterns of nifH gene expression. This chapter describes briefly our 5' nuclease assay protocols for Q-PCR and Q-RT-PCR of nifH gene fragments in environmental samples and discusses some important methodological considerations for the quantitative molecular examination of microbes in aquatic environments.

Base Sequence↗

PubMed1998

Circadian rhythm of nitrogenase gene expression in the diazotrophic filamentous nonheterocystous cyanobacterium Trichodesmium sp. strain IMS 101.

Recent studies suggested that the daily cycle of nitrogen fixation activity in the marine filamentous nonheterocystous cyanobacterium Trichodesmium sp. is controlled by a circadian rhythm. In this study, we evaluated the rhythm of nitrogen fixation in Trichodesmium sp. strain IMS 101 by using the three criteria for an endogenous rhythm. Nitrogenase transcript abundance oscillated with a period of approximately 24 h, and the cycle was maintained even under constant light conditions. The cyclic pattern of transcript abundance was maintained when the culture was grown at 24 and 28.5 degrees C, although the period was slightly longer (26 h) at the higher temperature. The cycle of gene expression could be entrained with light-dark cues. Results of inhibitor experiments indicated that transcript abundance was regulated primarily by transcription initiation, rather than by degradation. The circadian rhythm, the first conclusively demonstrated endogenous rhythm in a filamentous cyanobacterium, was also reflected in nitrogenase MoFe protein abundance and patterns of Fe protein posttranslational modification-demodification.

Bacterial Proteins↗

PubMed2004

Vertical distribution of nitrogen-fixing phylotypes in a meromictic, hypersaline lake.

We investigated the diversity of nitrogenase genes in the alkaline, moderately hypersaline Mono Lake, California to determine (1) whether nitrogen-fixing (diazotrophic) populations were similar to those in other aquatic environments and (2) if there was a pattern of distribution of phylotypes that reflected redox conditions, as well as (3) to identify populations that could be important in N dynamics in this nitrogen-limited lake. Mono Lake has been meromictic for almost a decade and has steep gradients in oxygen and reduced compounds that provide a wide range of aerobic and anaerobic habitats. We amplified a fragment of the nitrogenase gene (nifH) from planktonic DNA samples collected at three depths representing oxygenated surface waters, the oxycline, and anoxic, ammonium-rich deep waters. Forty-three percent of the 90 sequences grouped in nifH Cluster I. The majority of clones (57%) grouped in Cluster III, which contains many known anaerobic bacteria. Cluster I and Cluster III sequences were retrieved at every depth indicating little vertical zonation in sequence types related to the prominent gradients in oxygen and ammonia. One group in Cluster I was found most often at every depth and accounted for 29% of all the clones. These sequences formed a subcluster that contained other environmental clones, but no cultivated representatives. No significant nitrogen fixation was detected by the 15N2 method after 48 h of incubation of surface, oxycline, or deep waters, suggesting that pelagic diazotrophs were contributing little to nitrogen fluxes in the lake. The failure to measure any significant nitrogen fixation, despite the detection of diverse and novel nitrogenase genes throughout the water column, raises interesting questions about the ecological controls on diazotrophy in Mono Lake and the distribution of functional genes in the environment.

Bacteria↗

PubMed1982

Regulation of glutamine synthetase activity and synthesis in free-living and symbiotic Anabaena spp.

Regulation of the synthesis and activity of glutamine synthetase (GS) in the cyanobacterium Anabaena sp. strain 7120 was studied by determining GS transferase activity and GS antigen concentration under a variety of conditions. Extracts prepared from cells growing exponentially on a medium supplemented with combined nitrogen had a GS activity of 17 mumol of gamma-glutamyl transferase activity per min per mg of protein at 37 degrees C. This activity doubled in 12 h after transfer of cells to a nitrogen-free medium, corresponding to the time required for heterocyst differentiation and the start of nitrogen fixation. Addition of NH3 to a culture 11 h after an inducing transfer immediately blocked the increase in GS activity. In the Enterobacteriaceae, addition of NH3 after induction results in the covalent modification of GS by adenylylation. The GS of Anabaena is not adenylylated by such a protocol, as shown by the resistance of the transferase activity of the enzyme to inhibition by Mg2+ and by the failure of the enzyme to incorporate 32P after NH3 upshift. Methionine sulfoximine inhibited Anabaena GS activity rapidly and irreversibly in vivo. After the addition of methionine sulfoximine to Anabaena, the level of GS antigen neither increased nor decreased, indicating that Glutamine cannot be the only small molecule capable of regulating GS synthesis. Methionine sulfoximine permitted heterocyst differentiation and nitrogenase induction to escape repression by NH3. Nitrogen-fixing cultures treated with methionine sulfoximine excreted NH3. The fern Azolla caroliniana contains an Anabaena species living in symbiotic association. The Anabaena species carries out nitrogen fixation sufficient to satisfy all of the combined nitrogen requirements of the host fern. Experiments by other workers have shown that the activity of GS in the symbiont is significantly lower than the activity of GS in free-living Anabaena. Using a sensitive radioimmune assay and a normalization procedure based on the content of diaminopimelic acid, a component unique to the symbiont, we found that the level of GS antigen in the symbiont was about 5% of the level in free-living Anabaena cells. Thus, the host fern appears to repress synthesis of Anabaena GS in the symbiotic association.

Cyanobacteria↗

PubMed2000

Enhanced attachment of Bradyrhizobium japonicum to soybean through reduced root colonization of internally seedborne microorganisms.

Internally seedborne microorganisms are those surviving common surface sterilization procedures. Such microbes often colonize the radicle surface of a germinating soybean (Glycine max) seed, introducing an undefined parameter into studies on attachment and infection by Bradyrhizobium japonicum. Bacterial isolates from surface-sterilized soybean seed, cv. Williams 82 and cv. Maverick, used in our studies, were identified as Agrobacterium radiobacter, Aeromonas sp., Bacillus spp., Chryseomonas luteola, Flavimonas oryzihabitans, and Sphingomonas paucimobilis. Growth of these microbes during seed germination was reduced by treating germinating seeds with 500 micrograms/mL penicillin G. The effects of this antibiotic on seedling development and on B. japonicum 2143 attachment, nodulation, and nitrogen fixation are reported here. Penicillin G treatment of seeds did not reduce seed germination or root tip growth, or affect seedling development. No differences in nodulation kinetics, nitrogen fixation onset or rates were observed. However, the number of B. japonicum attached to treated intact seedlings was enhanced 200-325%, demonstrating that other root-colonizing bacteria can interfere with rhizobial attachment. Penicillin G treatment of soybean seedlings can be used to reduce the root colonizing microbes, which introduce an undefined parameter into studies of attachment of B. japonicum to the soybean root, without affecting plant development.

Bacterial Adhesion↗