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PubMed2002

Cavitation fatigue and its reversal in sunflower (Helianthus annuus L.).

"Cavitation fatigue" is the increased susceptibility of a xylem conduit to cavitation as a result of its prior cavitation. It was investigated whether cavitation fatigue induced in vivo could be repaired in intact plants. Sunflowers (Helianthus annuus L.) were subjected to soil drought in the greenhouse. Native embolism and vulnerability to cavitation was measured in well-watered controls and after 5 d and 10 d of controlled drought. A dramatic cavitation fatigue was observed where droughted xylem that was refilled in the laboratory developed up to 60 PLC (percentage loss of hydraulic conductivity) at -1 MPa versus only 5.2 PLC in non-droughted controls. Rewatered plants showed the complete reversal of cavitation fatigue over 4 d. Reversal of fatigue was correlated with the refilling of embolized vessels in the intact plants (r(2)=0.91, P<0.01), suggesting that xylem transport to fatigued vessels was required for their repair. The in vivo reversal of fatigue was partially duplicated in excised stem segments by perfusing them with root exudates from droughted (DR) and well-watered (WW) plants. The DR exudate had a greater effect, and this was associated with a greater pH in the DR versus WW saps, but there was no difference in total cation concentration. Perfusions with 2 mM CaCl(2) and KCl solutions also partially reversed cavitation fatigue as opposed to no effect with deionized water, suggesting a role of ions in addition to a pH effect. It is suspected that fatigue is caused by stretching and partial disruption of linkages between cellulose microfibrils in inter-conduit pit membranes during air seeding, and that the reversal of fatigue involves restoring these linkages by ingredients in xylem sap.

Biological Transport↗
PubMed1996

Foliage, fine-root, woody-tissue and stand respiration in Pinus radiata in relation to nitrogen status.

We measured respiration of 20-year-old Pinus radiata D. Don trees growing in control (C), irrigated (I), and irrigated + fertilized (IL) stands in the Biology of Forest Growth experimental plantation near Canberra, Australia. Respiration was measured on fully expanded foliage, live branches, boles, and fine and coarse roots to determine the relationship between CO(2) efflux, tissue temperature, and biomass or nitrogen (N) content of individual tissues. Efflux of CO(2) from foliage (dark respiration at night) and fine roots was linearly related to biomass and N content, but N was a better predictor of CO(2) efflux than biomass. Respiration (assumed to be maintenance) per unit N at 15 degrees C and a CO(2) concentration of 400 micro mol mol(-1) was 1.71 micro mol s(-1) mol(-1) N for foliage and 11.2 micro mol s(-1) mol(-1) N for fine roots. Efflux of CO(2) from stems, coarse roots and branches was linearly related to sapwood volume (stems) or total volume (branches + coarse roots) and growth, with rates for maintenance respiration at 15 degrees C ranging from 18 to 104 micro mol m(-3) s(-1). Among woody components, branches in the upper canopy and small diameter coarse roots had the highest respiration rates. Stem maintenance respiration per unit sapwood volume did not differ among treatments. Annual C flux was estimated by summing (1) dry matter production and respiration of aboveground components, (2) annual soil CO(2) efflux minus aboveground litterfall, and (3) the annual increment in coarse root biomass. Annual C flux was 24.4, 25.3 and 34.4 Mg ha(-1) year(-1) for the C, I and IL treatments, respectively. Total belowground C allocation, estimated as the sum of (2) and (3) above, was equal to the sum of root respiration and estimated root production in the IL treatment, whereas in the nutrient-limited C and I treatments, total belowground C allocation was greater than the sum of root respiration and estimated root production, suggesting higher fine root turnover or increased allocation to mycorrhizae and root exudation. Carbon use efficiency, the ratio of net primary production to assimilation, was similar among treatments for aboveground tissues (0.43-0.50). Therefore, the proportion of assimilation used for construction and maintenance respiration on an annual basis was also similar among treatments.

Journal Article↗
PubMedDate not supplied

Identification and characterization of a Rhizobium leguminosarum bv. phaseoli gene that is important for nodulation competitiveness and shows structural homology to a Rhizobium fredii host-inducible gene.

DNA sequence analysis of a 1.4-kb SalI-HindIII segment located approximately 2 kb upstream of the Rhizobium leguminosarum bv. phaseoli syrM gene revealed the presence of an open reading frame (ORF3) encoding a putative 295-amino acid polypeptide with a molecular mass of 33,401 Da. ORF3 is homologous to a R. fredii host-inducible gene. The proteins encoded by R. l. bv. phaseoli ORF3 and by the R. fredii host-inducible gene share 37% sequence identity. In contrast to the R. fredii host-inducible gene, expression of ORF3 is not induced in the presence of Phaseolus vulgaris root exudates or by specific flavonoids, able to induce nodulation genes in R. l. bv. phaseoli. A R. l. bv. phaseoli ORF3 mutant was constructed by site-directed deletion/replacement mutagenesis. This mutant strain is not affected in symbiotic nitrogen fixation but exhibits a delay in nodulation on Phaseolus vulgaris. Moreover, this mutant was shown to be defective in competition for nodulation.

Amino Acid Sequence↗
PubMedDate not supplied

The enhancement of ammonium assimilation in Rhizobium etli prevents nodulation of Phaseolus vulgaris.

The modification of the ammonium assimilation pathway of Rhizobium etli (GS-GOGAT) by adding an additional ammonium assimilation enzyme, GDH, strongly affects its symbiotic interaction with beans. The plasmid pAM1a, based in the stable vector pTR101 (M. Weinstein, R. C. Roberts, and D. R. Helsinki, J. Bacteriol. 174,7486-7489, 1992), containing the Escherichia coli gdhA gene flanked by two transcription-translation terminators was constructed. The expression of GDH in both, the wild type (CFN42/pAM1a) and a ntrC- mutant (CFN2012/pAM1a) R. etli strains, gave a similar metabolic effect, i.e., high GDH and reduced GOGAT activities, and an increased synthesis and excretion of several amino acids. The total inhibition of bean nodulation was observed when the minimum optimal inoculum of R. etli CFN42/pAM1a was used; however, an effective symbiosis occurred with the CFN2012/pAM1a mutant strain. While a total inhibition of the induction of the nodA gene by bean root exudate or by naringenin was observed in the CFN42/pAM1a strain, at 10 mM ammonium, the CFN2012/pAM1a showed an optimal nodA gene induction. A correlation between nodA gene induction, Nod factor production, and nodulation was observed. We conclude that in R. etli, there is a down-regulation of nod gene expression and nodulation when a high internal nitrogen content is built up by the presence of a functional GDH and that NtrC is involved in such regulation. An instability of the plasmid harboring the gdhA gene was observed during symbiosis, indicating a strong selection against cells containing this plasmid.

Down-Regulation↗
PubMed1999

Comparison of characteristics of the nodX genes from various Rhizobium leguminosarum strains.

We have analyzed the nucleotide sequences of the nodX genes from two strains of Rhizobium leguminosarum bv. viciae able to nodulate Afghan peas (strains A1 and Himalaya) and from two strains of R. leguminosarum bv. trifolii (ANU843 and CSF). The nodX genes of strains A1 and ANU843 were shown to be functional for the induction of nodules on Afghan peas. To analyze the cause of phenotypic differences of strain A1 and strain TOM we have studied the composition of the lipochitin-oligosaccharides (LCOs) produced by strain A1 after induction by the flavonoid naringenin or various pea root exudates. The structural analysis of the LCOs by mass spectrometry revealed that strain A1 synthesizes a family of at least 23 different LCOs. The use of exudates instead of naringenin resulted only in quantitative differences in the ratios of various LCOs produced.

Amino Acid Sequence↗
PubMed2026

Azotobacter vinelandii gene fitness following carbon shift from sucrose to acetate, succinate and glycerol.

Nitrogen-fixing microbes are a primary contributor of this important nutrient to the global nitrogen cycle. Biological nitrogen fixation (BNF) through the enzyme nitrogenase requires extensive energy that in whole cells is generally studied during the oxidation of carbohydrates such as sugars. The nitrogen-fixing bacterium Azotobacter vinelandii is a model diazotroph for the study of aerobic BNF. Much is known about metabolism in A. vinelandii when cultured on a simple medium where energy is provided primarily in the form of sucrose or glucose. Outside of the laboratory, this soil bacterium grows on metabolites primarily derived from plant root exudates or from the degradation of dead plant matter. In this work, we expand on previous studies looking at genes that are essential to BNF in A. vinelandii when grown on sucrose medium using transposon sequencing (Tn-seq). We applied Tn-seq to determine the genes essential to growth when the medium was shifted to acetate, succinate or glycerol as the primary carbon and energy source to fuel both growth and BNF. A global overview of the genes of central metabolism and those directing substrates toward central metabolism, along with a selection of unexpected genes that were essential for specific growth substrates, is provided.

Azotobacter vinelandii↗
PubMed2003

The mid genes of Rhizobium sp strain TAL1145 are required for degradation of mimosine into 3-hydroxy-4-pyridone and are inducible by mimosine.

Mimosine is a toxin present in the tree-legume leucaena (Leucaena leucocephala), including its root nodules and the root exudates. The leucaena-nodulating Rhizobium sp. strain TAL1145 degrades mimosine (Mid(+)) and utilizes it as a source of carbon and nitrogen. Twelve TAL1145 mutants defective in mimosine degradation (Mid(-)) were made through Tn3Hogus, TnphoA or kanamycin-resistance-cassette insertions. A 5.0 kb PstI fragment of TAL1145, subcloned from a cosmid clone containing mid genes for mimosine degradation, complemented most of the Mid(-) mutants. Sequencing this fragment and the adjacent 0.9 kb PstI fragment identified five genes, midA, midB, midC, midD and midR, of which the first three genes encode ABC transporter proteins involved in mimosine uptake, while midD encodes an aminotransferase required for degrading mimosine into 3-hydroxy-4-pyridone, and midR is a regulatory gene encoding a LysR-type transcriptional activator. The location of MidA in the periplasm was shown by making two midA : : phoA fusions, which made active alkaline phosphatase in the periplasm. The various mid : : gus and midA : : phoA fusions were inducible by mimosine, and a midD : : gus fusion mutant showed beta-glucuronidase activity in the leucaena nodules, indicating that midD is expressed in the nodules. Similarly, a midA : : phoA fusion expressed alkaline phosphatase activity in the leucaena nodules, indicating that mimosine induces midA transcription in the bacteroids. mid genes are specific for the Mid(+) strains of leucaena Rhizobium and are absent in strains of other Rhizobium, Sinorhizobium and Bradyrhizobium spp.

Bacterial Proteins↗
PubMed2025

Fast-growing Bacillus sensu lato rhizosphere populations are constrained by antagonistic Pseudomonadota, Actinomycetota and other Bacillus sensu lato.

Copiotrophic Bacillus and related taxa grow rapidly and are commonly isolated from soil. Despite their growth rate, Bacillus sensu lato (BSL) constitute less than one percent of soil bacterial communities, and the nutrient-enriched rhizosphere contains even fewer. Amendment of bulk soil with synthetic root exudate did not lead to increase in Bacillus culturable counts. We hypothesized that BSL populations in soil enriched with growth-supporting carbon are suppressed by various soil microbes. A screen using B. pseudomycoides as tester strain yielded 124 growth inhibiting isolates, aligning by 16S rRNA genes to 3 Alphaproteobacteria, 6 Betaproteobacteria, 5 Gammaproteobacteria, 3 Streptomyces, and 19 Bacillaceae. Most antagonists also suppressed four other BSL, and over 70% of the BSL isolates suppressed each other. The 11 sequenced BSL genomes encoded between 2 and 10 antibiotic biosynthetic gene clusters. Incubation of multiple isolates in artificial soil microcosms resulted in population growth restraint through a high percentage of endospores formed. This indicated that growth suppression by antagonists was due primarily to induction of sporulation. These results support our hypothesis that Bacillus populations in soil enriched with growth-supporting carbon are suppressed by various soil microbes.

Bacillus↗
PubMed1998

Release of flavonoids by the soybean cultivars McCall and peking and their perception as signals by the nitrogen-fixing symbiont sinorhizobium fredii

Sinorhizobium fredii strain USDA191 forms N-fixing nodules on the soybean (Glycine max L. Merr.) cultivars (cvs) McCall and Peking, but S. fredii strain USDA257 nodulates only cv Peking. We wondered whether specificity in this system is conditioned by the release of unique flavonoid signals from one of the cultivars or by differential perception of signals by the strains. We isolated flavonoids and used nodC and nolX, which are nod-box-dependent and -independent nod genes, respectively, to determine how signals activate genes in the microsymbionts. Seeds of cv McCall and cv Peking contain the isoflavones daidzein, genistein, and glycitein, as well as their glucosyl and malonylglucosyl glycosides. Roots exude picomolar concentrations of daidzein, genistein, glycitein, and coumestrol. Amounts are generally higher in cv Peking than in cv McCall, and the presence of rhizobia markedly influences the level of specific signals. Nanomolar concentrations of daidzein, genistein, and coumestrol induce expression of nodC and nolX in strain USDA257, but the relative nolX-inducing activities of these signals differ in strain USDA191. Glycitein and the conjugates are inactive. Strain USDA257 deglycosylates daidzin and genistin into daidzein and genistein, respectively, thereby converting inactive precursors into active inducers. Although neither soybean cultivar contains unique nod-gene-inducing flavonoids, strain- and cultivar-specific interactions are characterized by distinct patterns of signal release and response.

Journal Article↗
PubMed1966

Effects of decenylsuccinic Acid on the permeability and growth of bean roots.

Decenylsuccinic acid (DSA) at 10(-3)m has been reported to increase the permeability of bean root systems to water without seriously injuring the plants. We have confirmed the increase in permeability at 10(-3)m, but have found that 10(-4)m DSA reduces the permeability. Both concentrations cause leakage of salts from the roots and cessation of root pressure exudation. The roots of intact bean plants are killed by 1 hour's immersion in 10(-3)m DSA, but the plants may survive by producing new roots. Up to 4 hours in 10(-4)m DSA causes only temporary cessation of growth. Comparisons are made between the effects of DSA and some metabolic inhibitors. It is suggested that DSA is acting as a metabolic inhibitor, and that increase in water permeability is the result of injury to the roots. Experiments with 3 other species indicated variations in response to 10(-3)m DSA. These could be largely attributed to differences in susceptibility to injury.

Journal Article↗
PubMed1990

Role of the nodD and syrM genes in the activation of the regulatory gene nodD3, and of the common and host-specific nod genes of Rhizobium meliloti.

To analyse the regulation of the nodulation (nod) genes of Rhizobium meliloti RCR2011 we have isolated lacZ gene fusions to a number of common, host-range and regulatory nod genes, using the mini-Mu-lac bacteriophage transposon MudII1734. Common (nodA, nodC, nod region IIa) and host-range (nodE, nodG, nodH) genes were found to be regulated similarly. They were activated (i) by the regulatory nodD1 gene in the presence of flavones such as chrysoeriol, luteolin and 7,3',4'-trihydroxyflavone, (ii) by nodD2 in the presence of alfalfa root exudate but not with the NodD1-activating flavones, and (iii) by the regulatory genes syrM-nodD3 even in the absence of plant inducers. Thus common and host-range nod genes belong to the same regulon. In contrast to the nodD1 gene, the regulatory nodD3 gene was not expressed constitutively and exhibited a complex regulation. It required syrM for expression, was activated by nodD1 in the presence of luteolin and was positively autoregulated.

DNA, Bacterial↗
PubMed1995

Organization of host-inducible transcripts on the symbiotic plasmid of Rhizobium sp. NGR234.

In a systematic approach to identify genes involved in the early steps of the legume-Rhizobium symbiosis, we studied transcription patterns of symbiotic plasmid-borne loci. A competitive hybridization procedure was used to identify DNA restriction fragments carrying genes whose expression is enhanced by plant root exudates or by purified flavonoids. Fragments containing induced genes were then located on the physical map of the 500 kb pNGR234a. New inducible loci as well as previously described genes were identified and their time course of induction determined. After initial induction, transcription of loci such as nodABC and the host-specificity genes nodSU decreased to undetectable levels 24 h after incubation with purified flavonoids. In contrast, expression of other loci is detectable only after several hours of induction. Surprisingly, many genes remained transcribed in the nodD1- mutant suggesting the presence of other flavonoid-dependent activators in NGR234. The hsnl region, which is involved in host specificity, was shown to carry several inducible but independently regulated transcripts. Sequencing analysis revealed several open reading frames whose products, based on sequence similarities, may be involved in L-fucose metabolism and its adjunction to the Nod factors.

Binding, Competitive↗
PubMed2000

Insecticidal toxin from Bacillus thuringiensis is released from roots of transgenic Bt corn in vitro and in situ.

The insecticidal toxin encoded by the cry1Ab gene from Bacillus thuringiensis was released in root exudates from transgenic Bt corn during 40 days of growth in soil amended to 0, 3, 6, 9, or 12% (v/v) with montmorillonite or kaolinite in a plant growth room and from plants grown to maturity in the field. The presence of the toxin in rhizosphere soil was determined by immunological and larvicidal assays. No toxin was detected in any soils from isogenic non-Bt corn or without plants. Persistence of the toxin was apparently the result of its binding on surface-active particles in the soils, which reduced the biodegradation of the toxin. The release of the toxin could enhance the control of insect pests or constitute a hazard to nontarget organisms, including the microbiota of soil, and increase the selection of toxin-resistant target insects.

Journal Article↗
PubMed2000

Seasonal variation in rates of methane production from peat of various botanical origins: effects of temperature and substrate quality.

The methane produced in peat soils can vary over the growing season due to variations in the supply of available substrate, the activity of the microbial community or changes in temperature. Our aim was to study how these factors regulate the methane production over the season from five different peat types of different botanical origin. Peat samples were collected on seven occasions between June and September. After each sampling, the peat soils were incubated at five different temperatures (7, 10, 15, 20 and 25 degrees C) without added substrate, or at 20 degrees C with added substrate (glucose, or H(2)/CO(2), or starch). Rates of methane production averaged over the season differed significantly (P<0.05, R(2)=0.76) among the five peat types, the minerotrophic lawn producing the highest rates, and the hummock peat producing the lowest. The seasonal average Q(10) values for each plant community varied between 4.6 and 9.2, the highest value being associated with the ombrotrophic lawn and the lowest value with the mud-bottom plant community. For the unamended peat samples, the rates of methane production from each plant community varied significantly (P<0.05) over the season. This implies that the quality of organic matter, in combination with changes in temperature, explains the seasonal variation in methane production. However, addition of saturating amounts of glucose, H(2)/CO(2) or starch at 20 degrees C significantly reduced the seasonal variation (P<0.05) in methane production in peat from the minerotrophic lawn, wet carpet and mud-bottom plant communities. This suggests that substrate supply (e.g. root exudates) for the micro-organisms also varied over the season at these sites. Seasonal variation in methane production rates was apparent in peat from the hummock and ombrotrophic lawn plant communities even after addition of substrates, suggesting that the active biomass of the anaerobic microbial populations at these sites was regulated by other factors than the ones studied.

Journal Article↗
PubMed1978

Biological dinitrogen fixation (acetylene reduction) associated with Florida mangroves.

Biological dinitrogen fixation in mangrove communities of the Tampa Bay region of South Florida was investigated using the acetylene reduction technique. Low rates of acetylene reduction (0.01 to 1.84 nmol of C(2)H(4)/g [wet weight] per h) were associated with plant-free sediments, while plant-associated sediments gave rise to slightly higher rates. Activity in sediments increased greatly upon the addition of various carbon sources, indicating an energy limitation for nitrogenase (C(2)H(2)) activity. In situ determinations of dinitrogen fixation in sediments also indicated low rates and exhibited a similar response to glucose amendment. Litter from the green macroalga, Ulva spp., mangrove leaves, and sea grass also gave rise to significant rates of acetylene reduction. Higher rates of nitrogenase activity (15 to 53 nmol of C(2)H(4)/g [wet weight] per h were associated with washed excised roots of three Florida mangrove species [Rhizophora mangle L., Avicennia germinans (L) Stern, and Laguncularia racemosa Gaertn.] as well as with isolated root systems of intact plants (11 to 58 mug of N/g [dry weight] per h). Following a short lag period, root-associated activity was linear and did not exhibit a marked response to glucose amendment. It appears that dinitrogen-fixing bacteria in the mangrove rhizoplane are able to use root exudates and/or sloughed cell debris as energy sources for dinitrogen fixation.

Acetylene↗
PubMed1992

Identification and cloning of Bradyrhizobium japonicum genes expressed strain selectively in soil and rhizosphere.

The growth of Bradyrhizobium japonicum USDA 110 and USDA 438 in soil extract-supplemented medium led to transcription of a large amount of DNA not expressed in basal medium. Strain USDA 438 was more competitive for the nodulation of soybean than strain USDA 110. To identify and isolate DNA regions which were expressed specifically in strain USDA 438 but not in strain USDA 110 in response to soil extract or soybean root exudate, we developed a subtractive RNA hybridization procedure. Several cosmid clones which showed strain-specific gene expression were isolated from a USDA 438 gene library. Two clones enhanced competitive nodulation when mobilized to USDA 110. The method described may be useful for identifying genes expressed in response to environmental stimuli or genes expressed differently in related microbial strains.

Base Sequence↗
PubMed1996

Promotion of nod Gene Inducers and Nodulation in Common Bean (Phaseolus vulgaris) Roots Inoculated with Azospirillum brasilense Cd.

Inoculation of Phaseolus vulgaris with Azospirillum brasilense Cd promoted root hair formation in seedling roots and significantly increased total and upper nodule numbers at different concentrations of Rhizobium inoculum. In experiments carried out in a hydroponic system, A. brasilense caused an increase in the secretion of nod gene-inducing flavonoids, as was observed by nod gene induction assays of root exudates fractionated by high-performance liquid chromatography. Possible mechanisms involved in the influence of A. brasilense on this symbiotic system are discussed.

Journal Article↗