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The volatile compounds of twelve legume cultivars were evaluated by headspace-solid phase micro-extraction (HS-SPME) coupled to gas chromatography–mass spectrometry (GC-MS) from both whole seeds and their flours. The total identification percentages of the extracted volatiles were ranged between 94.2% and 99.7%. The number of aroma compounds detected was 104. Among them, 60 compounds were determined in the emission of whole legume seeds and 69 from legume flours. The volatiles were classified into seven chemical classes, i.e. monoterpene hydrocarbons, oxygenated monoterpenes, sesquiterpene hydrocarbons, phenylpropanoids, apocarotenes, nitrogen/sulfur derivatives, and non-terpene derivatives. Aldehydes were considered one of the most abundant constituents in non-terpene derivatives, followed by esters, alkanes, alcohols, ketones, lactones, phenols, phenones, alkenes, and hydrocarbons. A wide difference in term of volatiles was observed between whole and crushed seeds. Multivariate data analysis (PCA) was assessed to compare among legume cultivars, which leads to the identification of relevant constituents to specific cultivar. Overall, this is the first time a multivariate analysis was used to characterize the volatile profiles of twelve legume cultivars with their flours. This study can provide useful information on the conception of legume–based ingredients combined with specific volatile characteristics in order to reduce unwanted odors and definitely for pertinent breeding programs.

In this study, the effect of the phyto-pathogenic fungus Fusarium oxysporum on Medicago truncatula-Sinorhizobium symbiosis performance was investigated. Fusarium oxysporum (KLR13) was inoculated on two genotypes of M. truncatula, one tolerant (Jemalong A17) and one susceptible (TN1.11), in symbiosis with Sinorhizobium meliloti (TII7), Sinorhizobium medicae (SII4) or fertilized with KNO3. Fungus infection reduced significantly biomass production and photosynthetic related parameters of non-symbiotic M. truncatula A17 and TN1.11. Reduction was less pronounced in A17, where a lower accumulation of H2O2 and lipid peroxidation (MDA) in concomitance to high activities of antioxidant enzymes, catalase (CAT), peroxidase (POD) and superoxide dismutase (SOD) was detected. In symbiosis, the fungus affected the nodule number and weight and inhibited the nitrogen fixing capacity (NFC), as well as the fructose and glucose contents. Nevertheless, inoculation with S. meliloti and S. medicae alleviated the effect of F. oxysporum on plant biomass production compared to the KNO3 fertilized plants. Moreover, the Sinorhizobium inoculation decreased H2O2 and MDA over-production caused by the fungal infection in leaves of both genotypes. This tolerance enhancing effect was more pronounced in S. medicae-TN1.11 symbiosis, where the relative maintenance of photosynthesis related parameters, growth capacity and NFC performance was concomitant to the induction of SOD and CAT activities and an increase of the sucrose content in leaves. For A17 genotype, both Sinorhizobium strains induced an increase of the proline and sucrose accumulation. In conclusion, F. oxysporum effect on M. truncatula was mitigated when plants were in symbiosis with Sinorhizobium species, mainly Sinorhizobium medicae that alleviated the oxidative stress and enhanced the accumulation of sucrose as energy source in TN1.11 genotype, which attenuates its susceptibility to F. oxysporum infection.

Alleviation of cadmium-induced root genotoxicity and cytotoxicity by calcium chloride (CaCl(2)) in faba bean (Vicia faba L. var. minor) seedlings were studied. Faba bean seeds were treated with H(2)O or 2% CaCl(2) for 6 h before germination. Seeds were then exposed to 0 and 50 µM CdCl(2) concentrations for 7 days. Genotoxic damaging effects of Cd was examined through the determination of the mitotic index (MI), chromosomal aberrations (CA) and micronucleus (MN) in the meristem cells of faba bean roots. Similarly, effects of Cd stress on metal accumulation, total membrane lipid contents, total fatty acid composition (TFA), lipid peroxidation as indicated by malondialdehyde production, soluble protein and non-protein thiols (NP-SH) contents, hydrogen peroxide production and the activities of superoxide dismutase (SOD), catalase (CAT) and guaiacol peroxidase (GPX) were evaluated after 7 days of Cd stress in the seedling roots. Cd stress resulted in the reduction of MI, in addition to MN formation and CA induction in the roots of non-primed seeds (treated with H(2)O). Moreover, Cd induced lipid peroxidation, H(2)O(2) overproduction and loss of membrane lipid amount and soluble protein content, and changes in the TFA composition in roots of faba bean seedlings. SOD activity declined, but CAT and GPX activities increased. However, seed pre-treatment with CaCl(2) attenuated the genotoxic and cytotoxic effects of Cd on Vicia faba roots. The results showed that CaCl(2) induced reduction of Cd accumulation, improved cell membrane stability and increased the antioxidant defence systems, thus reducing and alleviating Cd genotoxicity and oxidative damage.

In this work, we have compared the physiological responses of alfalfa plants inoculated with either Sinorhizobium meliloti strain S412 (Cu-tolerant) or S112 (Cu-sensitive) in the presence or absence of 0.5 mM of CuSO4. The addition of copper (Cu) introduced a decrease of nodule number and their dry weight (DW) in both symbioses. The interaction established with the Cu-sensitive strain is more affected by Cu than that with the tolerant one. In fact, plants inoculated with the sensitive strain revealed a decrease of shoot and root DW, larger than that found in plants inoculated with the tolerant strain. However, under copper supply, Medicago sativa with the Cu-tolerant strain did not show any significant changes in both shoot and root biomass production. Under Cu excess, high levels of Cu were detected in different parts of the plant in the two symbioses and a high translocation of Cu to aerial parts was shown with the strain S412. Plants with S412 were able to accumulate large quantities of calcium (Ca) in...

The aim of this study was to assess the effect of symbiotic bacteria inoculation on the response ofMedicago truncatulagenotypes to iron deficiency. The present work was conducted on threeMedicago truncatulagenotypes: A17, TN8.20, and TN1.11. Three treatments were performed: control (C), direct Fe deficiency (DD), and induced Fe deficiency by bicarbonate (ID). Plants were nitrogen-fertilized (T) or inoculated with two bacterial strains:Sinorhizobium melilotiTII7 andSinorhizobium medicaeSII4. Biometric, physiological, and biochemical parameters were analyzed. Iron deficiency had a significant lowering effect on plant biomass and chlorophyll content in allMedicago truncatulagenotypes. TN1.11 showed the highest lipid peroxidation and leakage of electrolyte under iron deficiency conditions, which suggest that TN1.11 was more affected than A17 and TN8.20 by Fe starvation. Iron deficiency affected symbiotic performance indices of allMedicago truncatulagenotypes inoculated with bothSinorhizobiumstrains, mainly nodules number and biomass as well as nitrogen-fixing capacity. Nevertheless, inoculation withSinorhizobiumstrains mitigates the negative effect of Fe deficiency on plant growth and oxidative stress compared to nitrogen-fertilized plants. The highest auxin producing strain, TII7, preserves relatively high growth and root biomass and length when inoculated to TN8.20 and A17. On the other hand, both TII7 and SII4 strains improve the performance of sensitive genotype TN1.11 through reduction of the negative effect of iron deficiency on chlorophyll and plant Fe content. The bacterial inoculation improved Fe-deficient plant response to oxidative stress via the induction of the activities of antioxidant enzymes.

The aim of the present study was to evaluate abiotic stress tolerance of rhizobial strains belonging to Mesorhizobium, Sinorhizobium, and Rhizobium genera, as well as to investigate specie specific stress response mechanisms. Effect of NaCl and PEG on growth capacity, protein, lipid peroxydation (MDA), membrane fatty acid composition and antioxidant enzymes were investigated. Growth capacity and viability of overall rhizobia strains decreased proportionally to the increase of NaCl and PEG levels in the medium. Sinorhizobium strains appeared the most tolerant, where 4H41strain was able to grow at 800 mM NaCl and 40% PEG. On the other hand, growth of R. gallicum and M. mediterraneum was inhibited by 200 mM NaCl. The content of MDA was unchanged in Sinorhizobium strains under both stresses. For Mesorhizobium, only PEG treatment increased the content of MDA. Amount of the C19:0 cyclo fatty-acid was increased in both Sinorhizobium and Mesorhizobium tolerant strains. NaCl stress increased Superoxide dismutase (SOD) activity of overall species; especially the most tolerant strain 4H41. Both treatments increased catalase (CAT) activity in 4H41, TII7, and 835 strains. Obtained results suggest that major response of tolerant Sinorhizobium and Mesorhizobium strains to NaCl and PEG stresses is a preferential accumulation of the C19:0 cyclo fatty acid within bacterial membrane as mechanism to reduce fluidity and maintain integrity. Cell integrity and functioning is also assured by maintaining and/or increasing activity of SOD and CAT antioxidant enzymes for tolerant strains to omit structural and functional damages related to reactive oxygen species overproduced under stressful conditions. Abbreviations: CAT – catalase; CFA – cellular fatty acid; FAME – fatty acid methyl esters; MDA – malondialdehyde; ROS – reactive oxygen species; PEG – polyethylene glycol; PUFA – poly unsaturated fatty acid; SOD – superoxide dismutase; SDS PAGE – sodium dodecyl sulfate polyacrylamide gel electrophoresis

The ability of Phoma medicaginis to infect legume nodules and the subsequent effect on their functioning was investigated in Medicago truncatula lines, TN.8.20 and Jemalong6, inoculated with an efficient Sinorhizobium meliloti strain. Colonization of nodules by P. medicaginis strain Pm8 was confirmed by morphological and molecular analyses. P. medicaginis effect was more relevant on the most susceptible line, TN.8.20, where it decreased the nitrogen fixation capacity (NFC) significantly in nodules from 62 days after sowing (das) till the end of nodule life cycle; however in Jemalong6 nodules, the NFC inhibition was significant only at the flowering stage. Analyses of stress indices as structure, lipid peroxidation and leghemoglobin content did not show significant structural or metabolic alterations in infected nodules. Superoxide dismutase (SOD) activity was increased in nodules of both M. truncatula lines up to 62 das. After 78 das, increase of SOD activity continued only in Jemalong6 nodules. Changes in peroxidase (POX) activity were correlated to those revealed on NFC. Indeed, POX was decreased in nodules of TN.8.20 line but remained almost unchanged in Jemalong6. These results, suggest that POX represents a strong component of the defence mechanisms in nodules of resistant M. truncatula lines, where it could be involved in reinforcing the cell wall barrier. The stimulation of SOD activity is also a key mechanism in maintaining nodule integrity via controlling ROS overproduction when counteracting fungal attacks.

Biofilm formation by the pathogenic strain Candida albicans (C. albicans) is a crucial virulence factor on biotic and abiotic surfaces especially in medical devices. The present study investigated the effects of Cinnamomum verum and Thymus capitatus essential oils (EOs) and their combination with antifungal drugs on the development of biofilm and mature biofilm. The molecular mechanism of action was also investigated. Results showed that C. verum and T. capitatus EOs were broadly effective against C. albicans biofilm formation exhibiting 85.57% and 80.6% inhibition at half MIC values, respectively. Furthermore, they showed 92.79 and 94.46% eradication of C. albicans mature biofilm at 2xMIC values. Moreover, these EOs were able to reduce the production of secreted aspartic proteases by >90% at 0.75xMIC. The antibiofilm potency was improved when EOs were combined with AMB, effectively eradicating C. albicans mature biofilm on central venous catheter (CVC). Transcriptomic analyses indicated that EOs, especially when combined with AMB, down-regulate Ras-cAMP-Efg and MAPK signaling genes. In addition, EOs and their combination with AMB significantly inhibited the expression of adhesion and secreted aspartyl proteinase virulence factor genes. Our results demonstrated that C. verum and T. capitatus EOs and their combination with AMB could provide an effective treatment against C. albicans biofilm on medical devices.

The biochemical processes underlying the expression of resistance in the roots of Medicago truncatula against Aphanomyces euteiches infection was investigated, with emphasis on oxidative stress. The levels of H2O2, superoxide dismutase, peroxidase, ascorbate peroxidase, catalase, soluble phenolics and lignin were measured in the roots of two lines, A17 partially resistant and F83005.5 susceptible to A. euteiches at three infection stages; penetration of the epidermis (1 dpi), colonization of the cortex (3 dpi) and invasion of the root stele (6 dpi). A rapid and large decrease of the H2O2 levels in A17 roots occurred. However, in F83005.5 roots, the decrease in H2O2 levels was delayed until 3 dpi. In A17 roots, the activities of ascorbate peroxidase, peroxidase and catalase were induced as early as 1 dpi, whereas a general decrease in the activity of the four antioxidant enzymes was observed in F83005.5 roots. The levels of soluble phenolics and lignin were increased in A17 roots at 3 and 6 dpi, respectively. The H2O2 levels were negatively correlated to ascorbate peroxidase, catalase and lignin production at 1, 3 and 6 dpi, respectively in A17 roots. Physiological concentrations of H2O2 found in M. truncatula infected roots had no detrimental effect on the in vitro growth of this oomycete. Our data suggest that H2O2 does not have a direct antimicrobial effect on M. truncatula resistance to A. euteiches, but is involved in cell wall strengthening around the root stele, preventing pathogen invasion of the vascular tissues.

Antioxidant responses and nodule function of Medicago truncatula genotypes differing in salt tolerance were studied. Salinity effects on nodules were analysed on key nitrogen fixation proteins such as nitrogenase and leghaemoglobin as well as estimating lipid peroxidation levels, and were found more dramatic in the salt-sensitive genotype. Antioxidant enzyme assays for catalase (CAT, EC 1.11.1.6), superoxide dismutase (EC 1.15.1.1), ascorbate peroxidase (EC 1.11.1.11) and guaiacol peroxidase (EC 1.11.1.7) were analysed in nodules, roots and leaves treated with increasing concentrations of NaCl for 24 and 48 h. Symbiosis tolerance level, depending essentially on plant genotype, was closely correlated with differences of enzyme activities, which increased in response to salt stress in nodules (except CAT) and roots, whereas a complex pattern was observed in leaves. Gene expression responses were generally correlated with enzymatic activities in 24-h treated roots in all genotypes. This correlation was lost after 48 h of treatment for the sensitive and the reference genotypes, but it remained positively significant for the tolerant one that manifested a high induction for all tested genes after 48 h of treatment. Indeed, tolerance behaviour could be related to the induction of antioxidant genes in plant roots, leading to more efficient enzyme stimulation and protection. High induction of CAT gene was also distinct in roots of the tolerant genotype and merits further consideration. Thus, part of the salinity tolerance in M. truncatula is related to induction and sustained expression of highly regulated antioxidant mechanisms.

Sinorhizobium meliloti strain A5, isolated from root nodules of Medicago truncatula growing spontaneously in Tunisian dry lands, showed better growth performance under different sodium chloride concentrations than the standard reference strain RCR2011. Strain A5 has the ability to grow even at 684 mM NaCl. Two-dimensional (2D) electrophoretic analysis of the salt effect on proteome profiles revealed the greatest change on the 4th day of exposure. The halotolerant strain A5 showed more upregulated protein spots than the standard RCR2011 strain (23% of the analyzed protein spots vs only 10% in strain RCR2011) and fewer downregulated protein spots (28% vs 69%). Proteins over-expressed in salty medium as determined by Matrix-assisted laser desorption ionization time-of-flight mass spectrometry for both strains were identified as an outer-membrane protein, an l-amino-acid binding periplasmic transporter protein, a 50S ribosomal protein (L25), a heat shock protein, and a putative oxido-reductase protein. Down-regulated proteins were identified as a putative iron-binding periplasmic transporter protein, a putative lipoprotein precursor, a putative spermidine/putrescine-binding periplasmic transporter protein, and a putative amino-acid binding ABC transporter protein. The profile of strain A5 showed higher over-expression of an oxidoreductase and a heat-shock protein, and specifically an induction of a putative oligopeptide ABC transporter ATP-binding protein.

Thirty-six symbiotic associations involving six chickpea cultivars against six rhizobial strains were evaluated for symbiotic performance and responses to osmotic stress applied by mannitol (50 mM) in aerated hydroponic cultures. Analyses in different symbioses were focused on biomass production, nodulation, nitrogen fixation, and their modulation under osmotic stress conditions, as well as expression of nodular antioxidant enzymes. Mesorhizobium ciceri reference (835) and local (CMG6) strains, as well as the local (C11) M. mediterraneum allowed the best symbiotic efficiency for all chickpea cultivars. The osmotic stress induces severe decrease ranging 30–50% in aerial biomass and 50–70% for nitrogen fixation. Nevertheless, plants inoculated with M. ciceri (835) and M. mediterraneum (C11) preserve a relatively high growth (4 g plant−1) with nitrogen-fixing activity (25 μmols h−1 plant−1). The bacterial partner was the most important factor of variance of the analysed parameters in osmotic stress or physiological conditions where it gets to 60–85%. The strains allowing the best competent symbioses were proposed for field assays. Under osmotic stress, nodular peroxidase (POX) and ascorbate peroxidase (APX) activities were significantly enhanced. The increase of POX and APX was inversely correlated with the inhibition of aerial biomass production (p = 0.05) and nitrogen-fixing capacity (p = 0.01), suggesting a protective role of these enzymes in nodules. Superoxide dismutase (SOD) was also activated in stressed nodules. However, the spectacular decrease in catalase (CAT) activity discounts its involvement in osmotic stress response.

Agrobacterium sp. 10C2 is a nonpathogenic and non-symbiotic nodule-endophyte strain isolated from root nodules of Phaseolus vulgaris. The effect of this strain on nodulation, plant growth and rhizosphere bacterial communities of P. vulgaris is investigated under seminatural conditions. Inoculation with strain 10C2 induced an increase in nodule number (+54 %) and plant biomass (+16 %). Grains also showed a significant increase in phosphorus (+53 %), polyphenols (+217 %), flavonoids (+62 %) and total antioxidant capacity (+82 %). The effect of strain 10C2 on bacterial communities was monitored using terminal restriction fragment length polymorphism of PCR-amplified 16S rRNA genes. When the initial soil was inoculated with strain 10C2 and left 15 days, the Agrobacterium strain did not affect TRF richness but changed structure. When common bean was sown in these soils and cultivated during 75 days, both TRF richness and structure were affected by strain 10C2. TRF richness increased in the rhizosphere soil, while it decreased in the bulk soil (root free). The taxonomic assignation of TRFs induced by strain 10C2 in the bean rhizosphere revealed the presence of four phyla (Firmicutes, Actinobacteria, Bacteroidetes and Proteobacteria) with a relative preponderance of Firmicutes, represented mainly by Bacillus species. Some of these taxa (i.e., Bacillus licheniformis, Bacillus pumilus, Bacillus senegalensis, Bacillus subtilis, Bacillus firmus and Paenibacillus koreensis) are particularly known for their plant growth-promoting potentialities. These results suggest that the beneficial effects of strain 10C2 observed on plant growth and grain quality are explained at least in part by the indirect effect through the promotion of beneficial microorganisms.

The relationships between symbiotic performance and nodular antioxidant enzymes were studied for the associations between three Medicago truncatula lines and three Sinorhizobium meliloti strains. The results showed that the variability in symbiotic efficiency was dependent on the bacterial partner, host plant and their interaction. The contribution of each factor to the total amount of variance differed with the measured parameter. The aerial biomass production and nitrogen-fixing capacity were affected similarly by the three factors, whereas root and nodule biomass and catalase (CAT, E.C. 1.11.1.6), guaiacol peroxidase (POX, E.C. 1.11.1.7) and ascorbate peroxidase (APX, E.C. 1.11.1.11) antioxidant activities were mainly influenced by the M. truncatula line. The nodule number was dependent on the bacterial strain, and superoxide dismutase (SOD, E.C. 1.15.1.1) was dependent mainly on the plant-rhizobium interaction. A highly significant correlation was found between nitrogen-fixing activity, shoot biomass production, total nodule protein content and catalase activity. The other nodular antioxidant enzymes were differentially expressed between associations and showed no clear correlation with symbiotic efficiency.

Mesorhizobium ciceri, Mesorhizobium mediterraneum and Sinorhizobium medicae strains showed different symbiotic performances when inoculated to chickpea (Cicer arietinum L., cv. chetoui) at unstressed conditions and under salt stress. The analysis of nodular proteic composition and antioxidant enzyme activities revealed a polymorphism of patterns on SDS and native PAGE suggesting a potential dependence on the bacterial partner. Salt effect was analysed on plant growth, nitrogen fixation and antioxidant enzymes. M. ciceri, the most efficient strain, seemed to allow a best tolerance to chickpea plants under salt stress. This constraint did not affect the nodular superoxide dismutase (SOD, E.C. 1.15.1.1) activity of the symbiosis implicating the latter strain. This symbiosis showed the least decrease for the nodule protein level and the catalase (CAT, E.C. 1.11.1.6) activity, and the highest increase of peroxidase (POX, E.C. 1.11.1.7) activity that seemed to be related with the tolerance to salt.

In this study we investigated effects of Zn supply on germination, growth, inorganic solutes (Zn, Ca, Fe, and Mg) partitioning and nodulation of Medicago sativa This plant was cultivated with and without Zn (2 mM). Treatments were plants without (control) and with Zn tolerant strain (S532), Zn intolerant strain (S112) and 2 mM urea nitrogen fertilisation. Results showed that M. sativa germinates at rates of 50% at 2 mM Zn. For plants given nitrogen fertilisation, Zn increased plant biomass production. When grown with symbionts, Zn supply had no effect on nodulation. Moreover, plants with S112 showed a decrease of shoot and roots biomasses. However, in symbiosis with S532, an increase of roots biomass was observed. Plants in symbiosis with S. meliloti accumulated more Zn in their roots than nitrogen fertilised plants. Zn supply results in an increase of Ca concentration in roots of fertilised nitrogen plants. However, under Zn supply, Fe concentration decreased in roots and increased in nodules of plants with S112. Zn supply showed contrasting effects on Mg concentrations for plants with nitrogen fertilisation (increase) and plants with S112 (decrease). The capacity of M. sativa to accumulate Zn in their nodulated roots encouraged its use in phytostabilisation processes.

The aim of the present study was to evaluate abiotic stress tolerance of rhizobial strains belonging to Mesorhizobium, Sinorhizobium, and Rhizobium genera, as well as to investigate specie specific stress response mechanisms. Effect of NaCl and PEG on growth capacity, protein, lipid peroxydation (MDA), membrane fatty acid composition and antioxidant enzymes were investigated. Growth capacity and viability of overall rhizobia strains decreased proportionally to the increase of NaCl and PEG levels in the medium. Sinorhizobium strains appeared the most tolerant, where 4H41strain was able to grow at 800 mM NaCl and 40% PEG. On the other hand, growth of R. gallicum and M. mediterraneum was inhibited by 200 mM NaCl. The content of MDA was unchanged in Sinorhizobium strains under both stresses. For Mesorhizobium, only PEG treatment increased the content of MDA. Amount of the C19:0 cyclo fatty-acid was increased in both Sinorhizobium and Mesorhizobium tolerant strains. NaCl stress increased Superoxide dismutase (SOD) activity of overall species; especially the most tolerant strain 4H41. Both treatments increased catalase (CAT) activity in 4H41, TII7, and 835 strains. Obtained results suggest that major response of tolerant Sinorhizobium and Mesorhizobium strains to NaCl and PEG stresses is a preferential accumulation of the C19:0 cyclo fatty acid within bacterial membrane as mechanism to reduce fluidity and maintain integrity. Cell integrity and functioning is also assured by maintaining and/or increasing activity of SOD and CAT antioxidant enzymes for tolerant strains to omit structural and functional damages related to reactive oxygen species overproduced under stressful conditions.

Alternative oxidase (AOX) is the central component of the non-phosphorylating alternative respiratory pathway in plants and may be important for mitochondrial function during environmental stresses. Recently it has been proposed that Aox can be used as a functional marker for breeding stress tolerant plant varieties. This requires characterization of Aox alleles in plants with different degree of tolerance in a certain stress, affecting plant phenotype in a recognizable way. In this study we examined Aox1 gene expression levels in Medicago truncatula genotypes differing in salt stress tolerance, in order to uncover any correlation between Aox expression and tolerance to salt stress. Results demonstrated a specific induction of Aox1 gene expression in roots of the tolerant genotype that presented the lowest modulation in phenotypic and biochemical stress indices such as morphologic changes, protein level, lipid peroxidation and ROS generation. Similarly, in a previous study we reported that induction of antioxidant gene expression in the tolerant genotype contributed to the support of the antioxidant cellular machinery and stress tolerance. Correlation between expression patterns of the two groups of genes was revealed mainly in 48 h treated roots. Taken together, results from both experiments suggest that M. truncatula tolerance to salt stress may in part due to an efficient control of oxidative balance thanks to (i) induction of antioxidant systems and (ii) involvement of the AOX pathway. This reinforces the conclusion that differences in antioxidant mechanisms can be essential for salt stress tolerance in M. truncatula and possibly the corresponding genes, especially Aox, could be utilized as functional marker.

Plant production is essentially dependant on culture medium and conditions. Vegetable crops are known to grow in soil which, in natural conditions, acts as a mineral nutrient reservoir, but the soil itself is not essential to plant growth. Plant roots are able to absorb the mineral nutrients of the soil dissolved in water. When the required mineral nutrients are introduced into artificially synthesized medium (case of hydroponic cultivation), soil is no longer required for the plant growth. Hydroponic cultivation is a soil-free technology of vegetable growing. First reports about hydroponic culture date from the seventeenth century. Later, it was reported that in the Second World War, American soldiers grew their own food hydroponically while stationed on barren Pacific islands. Hydroponics was a necessity on Wake Island because there was no soil, and it was prohibitively expensive to airlift in fresh vegetables. Hydroponic cultivation is associated to the most revolutionary future technological projects. Indeed, for “future”, NASA Scientists are researching optimization of some factor as light, temperature and carbon dioxide, along with plant species for cultivation on planets like Mars.

The purpose of this study was to identify strains of Sinorhizobium meliloti that formed either an effective or completely ineffective symbiosis with Medicago truncatula L. ‘Jemalong A17’ and, subsequently, to determine whether differences existed between their exoH genes. Sinorhizobium meliloti TII7 and A5 formed an effective and ineffective symbiosis with M. truncatula ‘Jemalong A17’, respectively. Using a multilocus sequence typing method, both strains were shown to have chromosomes identical with S. meliloti Rm1021 and RCR2011. The 2260-bp segments of DNA stretching from the 3′ end of exoI through open reading frames of hypothetical proteins SM_b20952 and SM_b20953 through exoH into the 5′ end of exoK in strains TII7 and Rm1021 differed by a single nucleotide at base 127 of the hypothetical protein SM_b20953. However, the derived amino acid sequences of the exoH genes of effective TII7, ineffective A5, and strain Rm1021 were shown to be identical with each other. Therefore, it would seem unlikely that the gene product of exoH is directly involved with the low efficiency of a symbiosis of strain Rm1021 with M. truncatula ‘Jemalong A17’. Complementation or complete genome sequence analyses involving strains TII7 and A5 might be useful approaches to investigate the molecular bases for the differential symbiotic response with M. truncatula ‘Jemalong A17’.

Three genotypes of the model legume Medicago truncatula were assessed for symbiotic effectiveness in cross inoculation with two strains of Sinorhizobium meliloti under mannitol-mediated osmotic stress. Symbioses showed different tolerance levels revealed on plant growth, nitrogen-fixing capacity and indices of nodule functioning and protection. The variability of stress response was essentially correlated with performance at non-stressful conditions. Symbiosis attitude depended on bacterial partner, host-plant genotype and their interaction. Plant genotype manifested the highest contribution to symbiotic efficiency indices under osmotic stress, even for nodulation and nitrogen fixation where the bacterial strain effect is highly pronounced. Contrasting (tolerant/sensitive) associations were identified for tolerance behaviours, involving the same plant genotype with different rhizobial strains and vice versa. In nodules, osmotic stress leads to accumulation of oxidized lipids and decrease in total protein and leghaemoglobin contents. Antioxidant responses were manifested as induction of guaiacol peroxidase (POX, E.C. 1.11.1.7) and superoxide dismutase (E.C. 1.15.1.1). POX induction was higher in tolerant symbioses and both enzymes were suggested as contributors to the protection of nodule integrity and functioning under osmotic stress. In conclusion, symbiotic efficiency in M. truncatula–S. meliloti combinations under osmotic stress is determined by each symbiont’s input as well as the plant‐microbe genotype interaction, and POX induction could prove a sensitive marker of tolerant symbioses.

The effect of salt (NaCl) stress on plant growth and nitrogen-fixing apparatus was studied for symbiotic associations established between three Medicago truncatula lines inoculated and two Sinorhizobium meliloti strains. Salinity modulated all parameters analyzed as indices of growth performance, nitrogen-fixing capacity (acetylene-reduction assay) and nodule antioxidant system. Under stressful conditions, symbioses showed variability of response to salt application. Contrasting symbiotic associations were identified for nodulation, nitrogen-fixing capacity and salt tolerance. The total variance of analyzed performance indices under stressful conditions depended essentially on plant genotype factor. In nodules, the NaCl application had a decreasing effect on the rates of protein content and some antioxidant enzymes mainly catalase. Other enzymes such as guaiacol peroxidase increased in stressed nodules.