Sinorhizobium fredii HH103 nolR and nodD2 mutants gain capacity for infection thread invasion of Lotus japonicus Gifu and Lotus burttii.

Sinorhizobium fredii HH103 Rif , a broad-host-range rhizobial strain, forms ineffective nodules with Lotus japonicus but induces nitrogen-fixing nodules in Lotus burttii roots that are infected by intercellular entry. Here we show that HH103 Rif nolR or nodD2 mutants gain the ability to induce infection thread formation and to form nitrogen-fixing nodules in L. japonicus Gifu.

Opening the "black box" of nodD3, nodD4 and nodD5 genes of Rhizobium tropici strain CIAT 899.

Background: Transcription of nodulation genes in rhizobial species is orchestrated by the regulatory nodD gene. Rhizobium tropici strain CIAT 899 is an intriguing species in possessing features such as broad host range, high tolerance of abiotic stresses and, especially, by carrying the highest known number of nodD genes--five--and the greatest diversity of Nod factors (lipochitooligosaccharides, LCOs). Here we shed light on the roles of the multiple nodD genes of CIAT 899 by reporting, for the first time, results obtained with nodD3, nodD4 and nodD5 mutants.

CYN determination in tissues from freshwater fish by LC-MS/MS: validation and application in tissues from subchronically exposed tilapia (Oreochromis niloticus).

Harmful cyanobacterial blooms are occurring in eutrophic freshwater lakes and reservoirs throughout the world and, because of the production of toxins such as cylindrospermopsin (CYN), they can present a public safety hazard through contamination of seafood and fish for human consumption. Therefore it is important to develop methods to determine CYN at trace levels in those organisms. A new method for unconjugated CYN determination in tissues (liver and muscle) of tilapia (Oreochromis niloticus) is herein described and discussed; it is based on solvent extraction and purification with C18 and graphitized carbon cartridges, and quantification by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS).

Comparison of Microcystis aeruginosa (PCC7820 and PCC7806) growth and intracellular microcystins content determined by liquid chromatography-mass spectrometry, enzyme-linked immunosorbent assay anti-Adda and phosphatase bioassay.

Cyanobacteria are able to produce several metabolites that have toxic effects on humans and animals. Among these cyanotoxins, the hepatotoxic microcystins (MC) occur frequently. The intracellular MC content produced by two strains of Microcystis aeruginosa, PCC7806 and PCC7820, and its production kinetics during the culture time were studied in order to elucidate the conditions that favour the growth and proliferation of these toxic strains.

Analysis of MC-LR and MC-RR in tissue from freshwater fish (Tinca tinca) and crayfish (Procambarus clarkii) in tench ponds (Cáceres, Spain) by liquid chromatography-mass spectrometry (LC-MS).

In the present study a new method has been developed and validated for detecting free microcystins (MCs) (MC-RR, MC-LR and MC-YR) by liquid chromatography-mass spectrometry (LC-MS) in the cyprinid Tinca tinca and in the crayfish Procambarus clarkii collected from three ponds in Extremadura (Spain) where the presence of the cyanobacteria species Microcystis aeruginosa and Anabaena spiroides has been confirmed. Once the method had been validated, free MCs were determined in fish (tench, T. tinca) and crayfish from different ponds in order to understand how they are bioaccumulated through the food web.

High NaCl concentrations induce the nod genes of Rhizobium tropici CIAT899 in the absence of flavonoid inducers.

The nodulation (nod) genes of Rhizobium tropici CIAT899 can be induced by very low concentrations (micromolar to nanomolar range) of several flavonoid molecules secreted by the roots of leguminous plants under a number of different conditions. Some of these conditions have been investigated and appear to have a great influence on the concentration and the number of different Nod factors, which can induce root nodule primordia and pseudonodules in several leguminous plant roots. In one such condition, we added up to 300 mM NaCl to the induction medium of R.

Development and optimization of a method for the determination of Cylindrospermopsin from strains of Aphanizomenon cultures: intra-laboratory assessment of its accuracy by using validation standards.

The occurrence of cyanobacterial blooms in aquatic environments is increasing in many regions of the world due to progressive eutrophication of water bodies. Because of the production of toxins such as Cylindrospermopsin (CYN), contamination of water with cyanobacteria is a serious health problem around the world. Therefore it is necessary to develop and validate analytical methods that allow us to quantify CYN in real samples in order to alert the public of this toxin.

Cylindrospermopsin determination in water by LC-MS/MS: optimization and validation of the method and application to real samples.

A new method for determining dissolved cylindrospermopsin (CYN) in waters using solid-phase extraction (SPE) with graphitized carbon cartridges and quantification by liquid chromatography coupled with tandem mass spectrometry is described and discussed. The method has been suitably validated: the linear range covered is from 0.900 to 125 µg CYN/L.

The nodulation of alfalfa by the acid-tolerant Rhizobium sp. strain LPU83 does not require sulfated forms of lipochitooligosaccharide nodulation signals.

The induction of root nodules by the majority of rhizobia has a strict requirement for the secretion of symbiosis-specific lipochitooligosaccharides (nodulation factors [NFs]). The nature of the chemical substitution on the NFs depends on the particular rhizobium and contributes to the host specificity imparted by the NFs. We present here a description of the genetic organization of the nod gene cluster and the characterization of the chemical structure of the NFs associated with the broad-host-range Rhizobium sp.

Sinorhizobium fredii HH103 cgs mutants are unable to nodulate determinate- and indeterminate nodule-forming legumes and overproduce an altered EPS.

Sinorhizobium fredii HH103 produces cyclic beta glucans (CG) composed of 18 to 24 glucose residues without or with 1-phosphoglycerol as the only substituent. The S. fredii HH103-Rifr cgs gene (formerly known as ndvB) was sequenced and mutated with the lacZ-gentamicin resistance cassette.

Different and new Nod factors produced by Rhizobium tropici CIAT899 following Na+ stress.

The root nodule bacterium Rhizobium tropici strain CIAT899 is highly stress resistant. It grows under acid conditions, in large amounts of salt, and at high osmotic pressure. An earlier study reported a substantial qualitative and quantitative effect of acid stress on the biosynthesis of Nod factors.

Signal molecules in the peanut-bradyrhizobia interaction.

Main nodulation signal molecules in the peanut-bradyrhizobia interaction were examined. Flavonoids exuded by Arachis hypogaea L. cultivar Tegua were genistein, daidzein and chrysin, the latest being released in lower quantities.

Growth and exopolysaccharide (EPS) production by Oenococcus oeni I4 and structural characterization of their EPSs.

Aims: To study the influence of medium constituents on growth, and exopolysaccharide (EPS) production by a strain of Oenococcus oeni. The structure of one of the EPSs has also been characterized.

Methods And Results: EPS concentration was estimated by the phenol/sulfuric acid method.

Structural determination of the Nod factors produced by Rhizobium gallicum bv. gallicum R602.

Rhizobium gallicum is a fast-growing bacterium found in European, Australian and African soils; it was first isolated in France. It is a microsymbiont which is able to nodulate plants of the genus Phaseolus. Rhizobium gallicum bv.

Low pH changes the profile of nodulation factors produced by Rhizobium tropici CIAT899.

Rhizobium tropici CIAT899 has been cataloged as a nodulator of bean, a plant often growing in areas characterized by highly acidic soils. The purpose of this work was to explore the effects of acidity on the production of Nod factors by this strain and their impact on the establishment of effective symbioses. We report that acidity increases rhizobial Nod factors production, and we exhaustively study the nodulation factor structures produced under abiotic stress.

Structural analysis of the capsular polysaccharide from Sinorhizobium fredii HWG35.

We have determined the structure of a capsular polysaccharide from Sinorhizobium fredii HWG35. This polysaccharide was isolated following the standard protocols applied for lipopolysaccharide isolation. On the basis of monosaccharide analysis, methylation analysis, mass spectrometric analysis, one-dimensional (1)H and (13)C NMR, and two-dimensional NMR experiments, the structure was shown to consist of a polymer having the following disaccharide repeating unit: -->6)-2,4-di-O-methyl-alpha-d-Galp-(1-->4)-beta-d-GlcpA-(1-->.

NolR regulates diverse symbiotic signals of Sinorhizobium fredii HH103.

We have investigated in Sinorhizobium fredii HH103-1 (=HH103 Str(r)) the influence of the nolR gene on the production of three different bacterial symbiotic signals: Nod factors, signal responsive (SR) proteins, and exopolysaccharide (EPS). The presence of multiple copies of nolR (in plasmid pMUS675) repressed the transcription of all the flavonoid-inducible genes analyzed: nodA, nodD1, nolO, nolX, noeL, rhcJ, hesB, and y4pF. Inactivation of nolR (mutant SVQ517) or its overexpression (presence of pMUS675) altered the amount of Nod factors detected.

Structural determination of the lipo-chitin oligosaccharide nodulation signals produced by Rhizobium giardinii bv. giardinii H152.

Rhizobium giardinii bv. giardinii is a microsymbiont of plants of the genus Phaseolus and produces extracellular signal molecules that are able to induce deformation of root hairs and nodule organogenesis. We report here the structures of seven lipochitooligosaccharide (LCO) signal molecules secreted by R.

Alfalfa nodulation by Sinorhizobium fredii does not require sulfated Nod-factors.

Rhizobium strain 042B(s) is able to nodulate both soybean and alfalfa cultivars. We have demonstrated, by mass spectrometry, that the nodulation (Nod) factors produced by this strain are characteristic of those produced by Sinorhizobium fredii, which typically nodulates soybean; they have 3-5 N-acetylglucosamine (GlcNAc) residues, a mono-unsaturated or saturated C16, C18 or C20 fatty-acyl chain, and a (methyl)fucosyl residue on C of the reducing-terminal GlcNAc. In order to study Rhizobium strain 042B(s) and its nodulation behaviour further, we introduced an insertion mutation in the noeL gene, which is responsible for the presence of the (methyl)fucose residue on the reducing terminal GlcNAc of the Nod-factors, yielding mutant strain SVQ523.

Sinorhizobium fredii HH103 has a truncated nolO gene due to a -1 frameshift mutation that is conserved among other geographically distant S. fredii strains.

Strain SVQ121 is a mutant derivative of Sinorhizobium fredii HH103 carrying a transposon Tn5-lacZ insertion into the nolO-coding region. Sequence analysis of the wild-type gene revealed that it is homologous to that of Rhizobium sp. NGR234, which is involved in the 3 (or 4)-O-carbamoylation of the nonreducing terminus of Nod factors.

Regulation of nod factor sulphation genes in Rhizobium tropici CIAT899.

Rhizobium tropici CIAT899 is a tropical symbiont able to nodulate various legumes such as Leucaena, Phaseolus, and Macroptilium. Broad host range of this species is related to its Nod factors wide spectrum. R.

Extracellular polysaccharide composition of Azospirillum brasilense and its relation with cell aggregation.

The exopolysaccharide (EPS) and capsular polysaccharide (CPS) composition of four Azospirillum brasilense strains differing in their aggregation capacity was analyzed by high performance anion exchange chromatography. When growing the different strains in an aggregation inducing medium containing a high carbon:nitrogen (C:N) ratio, both EPS and CPS showed a positive correlation between aggregation and the relative amount of arabinose. Arabinose was not detected in polysaccharides from Sp72002, a pleiotrophic Tn5 mutant strain impaired in aggregation.