The presence of cyanotoxins, mainly microcystins (MCs), in surface freshwater represents a serious health risk to aquatic organisms living in the water body, as well as terrestrial animals and plants that are in contact with contaminated water. Consequently, the use of MCs contaminated water for irrigation represents a hazard for cultivated plants and could induce severe economical losses due to crops' yield reduction. The experimental approach undertaken in this work was exposing Vicia faba seedlings (inoculated with a Rhizobium strain resistant to MCs), to water supplemented with cyanobacterial crude extract containing total microcystins at a concentration of 50 and 100 μg/L (environmental relevant concentrations of MCs dissolved in the raw irrigation water from Lalla Takerkoust Lake-Marrakesh region). After chronic MCs exposure (2 months), biological and physiological parameters (plant growth, nitrogen uptake, mineral assimilation, and oxidative defense mechanisms) were evaluated. The results obtained showed evidence that chronic exposure to cyanobacterial bloom extract containing MCs strongly affected the physiological and biological plants activities; reduction of dry matter, photosynthetic activity, nodule number, and nitrogen assimilation. At the same time, an increase of oxidative stress was observed, as deduced from a significant increase of the activities of peroxidase, catalase, polyphenoloxidase, and phenylalanine ammonia lyase in leaves, roots, and nodules of faba bean plants exposed to cyanotoxins, especially at 100 μg/L of MCs. This experimentation constitutes a simulation of the situation related to cyanotoxins chronic exposure of seedlings-plants via the contaminated irrigation water. For this reason, once should take into consideration the possibility of contamination of agricultural crops and the quality of irrigation water should be by the way monitored for cyanotoxins biohazard.

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A previously undescribed mutation of hereditary γ γ-glutamylcysteine synthetase (GCS) deficiency was found in a 5 year old boy of Moroccan origin. He presented with chronic haemolytic anaemia, delayed psychomotor development and progressive motor sensitive neuropathy of lower extremities. The parents were third degree relatives. The activity of glycolytic enzymes were found to be normal in the propositus, his parents and a sister, but and a complete lack of GSH was found in the propositus. Accordingly, the measurement of de novo GSH synthetic enzymes was undertaken, and severe GCS deficiency was found in the propositus. Both parents and his sister presented GCS activity ranging from 69% to 90% of normal. GCS gene sequencing showed that the propositus was homozygous for a 1241C>T mutation in exon 11 and both parents and his sister were heterozygous. This mutation predicts a Pro414Leu amino acid substitution. Even though the homology between GCS and crystallographically solved, functionally related proteins is not very high, a three-dimensional model of GCS was derived using Modeller Software. GCS deficiency is a very rare autosomal recessive disorder reported so far in only 8 unrelated probands with severe haemolytic anaemia. In only 3 of these was the anaemia associated with severe neurological dysfunction. We report here the fourth case of GCS deficiency presenting neuropathy, giving further support to the eventual relationship between this enzymopathy and neurological damage Haematologica 2007; 92:e102-e105

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SummaryThe genetic regulation underlying the effect of arsenic (As(III)) on the model symbiosis Medicago-Ensifer was investigated using a combination of physiological (split-roots), microscopy and genetic (microarrays, qRT-PCR and composite plants) tools.Nodulation was very sensitive to As(III) (median inhibitory dose (ID50) = 20 lM). The effect on root elongation and on nodulation was local (nonsystemic). A battery of stress (salt, drought, heat shock, metals, etc.)-related genes were induced. Glutathione played a pivotal role in tolerance/detoxification, together with secondary metabolites ((iso)flavonoids and phenylpropanoids). However, antioxidant enzymes were not activated.Concerning the symbiotic interaction, molecular evidence suggesting that rhizobia alleviate As stress is for the first time provided. Chalcone synthase (which is involved in the first step of the legume-rhizobia cross-talk) was strongly enhanced, suggesting that the plants are biased to establish symbiotic interactions under As(III) stress. In contrast, 13 subsequent nodulation genes (involved in nodulation factors (Nod factors) perception, infection, thread initiation and progression, and nodule morphogenesis) were repressed.Overexpression of the ethylene responsive factor ERN in composite plants reduced root stress and partially restored nodulation, whereas overexpression of the early nodulin ENOD12 enhanced nodulation both in the presence and, particularly, in the absence of As, without affecting root elongation. Several transcription factors were identified, which could be additional targets for genetic engineering aiming to improve nodulation and/or alleviate root stress induced by this toxic.

The rhizobia-legume interaction has been proposed as an interesting and appropriate tool for rhizostabilization of soils contaminated with heavy metals. One of the main requirements to use this symbiosis is the availability of tolerant and symbiotically effective rhizobia. The aim of this work was to improve the symbiotic properties of the arsenic-resistant wild-type strain Ensifer medicae MA11 in Cu-contaminated substrates. The copAB genes from a Cu-resistant Pseudomonas fluorescens strain were expressed in E. medicae MA11 under the control of the nifH promoter. The resulting strain E. medicae MA11-copAB was able to alleviate the toxic effect of Cu in Medicago truncatula. At 300 µM Cu, root and shoot dry matter production, nitrogen content, number of nodules and photosynthetic rate were significantly reduced in plants inoculated with the wild-type strain. However, these parameters were not altered in plants inoculated with the genetically modified strain. Moreover, nodules elicited by this strain were able to accumulate twofold the Cu measured in nodules formed by the wild-type strain. In addition, the engineered E. medicae strain increased Cu accumulation in roots and decreased the content in shoots. Thus, E. medicae MA11-copAB increased the capacity of M. truncatula to rhizostabilize Cu, decreasing the translocation factor and avoiding metal entry into the food chain. The plasmid containing the nifH promoter-copAB construct could be a useful biotool for Cu rhizostabilization using legumes, since it can be transferred to different rhizobia microsymbionts of authoctonous legumes growing in Cu-contaminated soils.

A collection of rhizobia isolated from Acacia tortilis subsp. raddiana nodules from various arid soils in Tunisia was analyzed for their diversity at both taxonomic and symbiotic levels. The isolates were found to be phenotypically diverse. The majority of the isolates tolerated 3% NaCl and grew at 40 °C. Genetic characterization emphasized that most of the strains (42/50) belong to the genus Ensifer, particularly the species Ensifer meliloti, Ensifer garamanticus, and Ensifer numidicus. Symbiotic properties of isolates showed diversity in their capacity to nodulate their host plant and to fix atmospheric nitrogen. The most effective isolates were closely related to E. garamanticus. Nodulation tests showed that 3 strains belonging to Mesorhizobium genus failed to renodulate their host plant, which is surprising for symbiotic rhizobia. Furthermore, our results support the presence of non-nodulating endophytic bacteria belonging to the Acinetobacter genus in legume nodules.