Mesorhizobium ciceri as biological tool for improving physiological, biochemical and antioxidant state of Cicer aritienum (L.) under fungicide stress.

Fungicides among agrochemicals are consistently used in high throughput agricultural practices to protect plants from damaging impact of phytopathogens and hence to optimize crop production. However, the negative impact of fungicides on composition and functions of soil microbiota, plants and via food chain, on human health is a matter of grave concern. Considering such agrochemical threats, the present study was undertaken to know that how fungicide-tolerant symbiotic bacterium, Mesorhizobium ciceri affects the Cicer arietinum crop while growing in kitazin (KITZ) stressed soils under greenhouse conditions. Both in vitro and soil systems, KITZ imparted deleterious impacts on C. arietinum as a function of dose. The three-time more of normal rate of KITZ dose detrimentally but maximally reduced the germination efficiency, vigor index, dry matter production, symbiotic features, leaf pigments and seed attributes of C. arietinum. KITZ-induced morphological alterations in root tips, oxidative damage and cell death in root cells of C. arietinum were visible under scanning electron microscope (SEM). M. ciceri tolerated up to 2400 µg mL-1 of KITZ, synthesized considerable amounts of bioactive molecules including indole-3-acetic-acid (IAA), 1-aminocyclopropane 1-carboxylate (ACC) deaminase, siderophores, exopolysaccharides (EPS), hydrogen cyanide, ammonia, and solubilised inorganic phosphate even in fungicide-stressed media. Following application to soil, M. ciceri improved performance of C. arietinum and enhanced dry biomass production, yield, symbiosis and leaf pigments even in a fungicide-polluted environment. At 96 µg KITZ kg-1 soil, M. ciceri maximally and significantly (p ≤ 0.05) augmented the length of plants by 41%, total dry matter by 18%, carotenoid content by 9%, LHb content by 21%, root N by 9%, shoot P by 11% and pod yield by 15% over control plants. Additionally, the nodule bacterium M. ciceri efficiently colonized the plant rhizosphere/rhizoplane and considerably decreased the levels of stressor molecules (proline and malondialdehyde) and antioxidant defence enzymes viz. ascorbate peroxidise (APX), guaiacol peroxidise (GPX), catalase (CAT) and peroxidises (POD) of C. arietinum plants when inoculated in soil. The symbiotic strain effectively colonized the plant rhizosphere/rhizoplane. Conclusively, the ability to endure higher fungicide concentrations, capacity to secrete plant growth modulators even under fungicide pressure, and inherent features to lower the level of proline and plant defence enzymes makes this M. ciceri as a superb choice for augmenting the safe production of C. arietinum even under fungicide-contaminated soils.

Isolation, characterization, and selection of heavy metal-resistant and plant growth-promoting endophytic bacteria from root nodules of Robinia pseudoacacia in a Pb/Zn mining area.

Multiple heavy metals (HMs) commonly coexist in mining areas, which highlights the necessity to select multiple HM-resistant plant growth-promoting bacteria for improving phytoremediation efficiency. In this study, we isolated and characterized 82 endophytic bacteria from the root nodules of black locust (Robinia pseudoacacia) grown in a Pb-Zn mining area. There were 80 isolates showing resistance to four HMs, 0.01-18.0 mM/L for Cd, 0.2-40.0 mM/L for Zn, 0.3-2.2 mM/L for Pb, and 0.2-1.4 mM/L for Cu. Indole-3-acetic acid production, siderophore production, and 1-aminocyclopropane-1-carboxylate deaminase activity were detected in 43, 50, and 17 isolates, respectively. Two symbiotic isolates selected with the highest potential for HM resistance and PGP traits, designated Mesorhizobium loti HZ76 and Agrobacterium radiobacter HZ6, were evaluated for promotion of plant growth and metal uptake by R. pseudoacacia seedlings grown in pots containing different levels of Cd, Zn, Pb, or Cu. HZ76 significantly increased plant shoot biomass, while HZ6 did not, compared with non-inoculated controls. The results indicate that inoculation with HZ76 or HZ6 relieved HM stress in the plants, depending on the type and concentration of HM in the treatment. Mesorhizobium loti HZ76 may be a better candidate for application in phytoremediation than A. radiobacter HZ6. The microsymbiosis between HM-resistant rhizobia and R. pseudoacacia is an interesting mutualistic system for phytoremediation in mining areas contaminated with multiple HMs.

Rheological, textural and emulsifying properties of an exopolysaccharide produced by Mesorhizobium loti grown on a crude glycerol-based medium.

In the present study, a new extracellular polysaccharide (EPS-M816) was obtained during the growth of Mesorhizobium loti Semia 816 on a crude glycerol-based medium. EPS-M816 precipitate mainly consisted of carbohydrates (82.54%) and proteins (11.31%), and the weight average molecular weight was estimated at 1.646 × 106 Da. The biopolymer was characterized by FT-IR and NMR spectroscopy, and was found to have typical functional groups of other rhizobial polysaccharides. Furthermore, the rheological and emulsifying properties were investigated. The EPS-M816 solution (1.0% w/v) showed typical pseudoplastic non-Newtonian fluid behavior, and the addition of sodium and potassium chloride (1 mol L-1) increased the apparent viscosity. Regarding its emulsification activity, EPS-M816 formed emulsions with different food-grade vegetable oils (soybean, rice, canola, sunflower and corn oils), showing emulsification index values over 65% in 24 h, indicative of strong emulsion-stabilizing capacity. The biopolymer was able to form gels with texture parameters similar to those reported for xanthan gum and low syneresis. Overall, these results suggest that EPS-M816 is a good candidate for application in the food, cosmetics and pharmaceutical industries as a thickening, gelling, stabilizing and emulsifying agent.

Biotic elicitation of ginsenoside metabolism of mutant adventitious root culture in Panax ginseng.

Biotic elicitation is an important biotechnological strategy for triggering the accumulation of secondary metabolites in adventitious root cultures. These biotic elicitors can be obtained from safe, economically important strains of bacteria found in the rhizosphere and fermented foods. Here, we assayed the effects of filtered cultures of five nitrogen-fixing bacteria and four types of fermentation bacteria on mutant adventitious Panax ginseng root cultures induced in a previous study by colchicine treatment. The biomass, pH, and electrical conductivity (EC) of the culture medium were altered at 5 days after treatment with bacteria. The saponin content was highest in root cultures treated with Mesorhizobium amorphae (GS3037), with a concentration of 105.58 mg g-1 dry weight saponin present in these cultures versus 74.48 mg g-1 dry weight in untreated root cultures. The accumulation of the ginsenosides Rb2 and Rb3 dramatically increased (19.4- and 4.4-fold, and 18.8- and 4.8-fold) 5 days after treatment with M. amorphae (GS3037) and Mesorhizobium amorphae (GS336), respectively. Compound K production increased 1.7-fold after treatment with M. amorphae (GS3037) compared with untreated root cultures. These results suggest that treating mutant adventitious root cultures with biotic elicitors represents an effective strategy for increasing ginsenoside production in Panax ginseng.

Quantitative proteomics of a B12 -dependent alga grown in coculture with bacteria reveals metabolic tradeoffs required for mutualism.

The unicellular green alga Lobomonas rostrata requires an external supply of vitamin B12 (cobalamin) for growth, which it can obtain in stable laboratory cultures from the soil bacterium Mesorhizobium loti in exchange for photosynthate. We investigated changes in protein expression in the alga that allow it to engage in this mutualism. We used quantitative isobaric tagging (iTRAQ) proteomics to determine the L. rostrata proteome grown axenically with B12 supplementation or in coculture with M. loti. Data are available via ProteomeXchange (PXD005046). Using the related Chlamydomonas reinhardtii as a reference genome, 588 algal proteins could be identified. Enzymes of amino acid biosynthesis were higher in coculture than in axenic culture, and this was reflected in increased amounts of total cellular protein and several free amino acids. A number of heat shock proteins were also elevated. Conversely, photosynthetic proteins and those of chloroplast protein synthesis were significantly lower in L. rostrata cells in coculture. These observations were confirmed by measurement of electron transfer rates in cells grown under the two conditions. The results indicate that, despite the stability of the mutualism, L. rostrata experiences stress in coculture with M. loti, and must adjust its metabolism accordingly.

The Phenylalanine Ammonia Lyase Gene LjPAL1 Is Involved in Plant Defense Responses to Pathogens and Plays Diverse Roles in Lotus japonicus-Rhizobium Symbioses.

Phenylalanine ammonia lyase (PAL) is important in the biosynthesis of plant secondary metabolites that regulate growth responses. Although its function is well-established in various plants, the functional significance of PAL genes in nodulation is poorly understood. Here, we demonstrate that the Lotus japonicus PAL (LjPAL1) gene is induced by Mesorhizobium loti infection and methyl-jasmonate (Me-JA) treatment in roots. LjPAL1 altered PAL activity, leading to changes in lignin contents and thicknesses of cell walls in roots and nodules of transgenic plants and, hence, to structural changes in roots and nodules. LjPAL1-knockdown plants (LjPAL1i) exhibited increased infection thread and nodule numbers and the induced upregulation of nodulin gene expression after M. loti infection. Conversely, LjPAL1 overexpression delayed the infection process and reduced infection thread and nodule numbers after M. loti inoculation. LjPAL1i plants also exhibited reduced endogenous salicylic acid (SA) accumulation and expression of the SA-dependent marker gene. Their infection phenotype could be partially restored by exogenous SA or Me-JA application. Our data demonstrate that LjPAL1 plays diverse roles in L. japonicus-rhizobium symbiosis, affecting rhizobial infection progress and nodule structure, likely by inducing lignin modification, regulating endogenous SA biosynthesis, and modulating SA signaling.

Role of exopolysaccharide in salt stress resistance and cell motility of Mesorhizobium alhagi CCNWXJ12-2T.

Mesorhizobium alhagi, a legume-symbiont soil bacterium that forms nodules with the desert plant Alhagi sparsifolia, can produce large amounts of exopolysaccharide (EPS) using mannitol as carbon source. However, the role of EPS in M. alhagi CCNWXJ12-2T, an EPS-producing rhizobium with high salt resistance, remains uncharacterized. Here, we studied the role of EPS in M. alhagi CCNWXJ12-2T using EPS-deficient mutants constructed by transposon mutagenesis. The insertion sites of six EPS-deficient mutants were analyzed using single primer PCR, and two putative gene clusters were found to be involved in EPS synthesis. EPS was extracted and quantified, and EPS production in the EPS-deficient mutants was decreased by approximately 25 times compared with the wild-type strain. Phenotypic analysis revealed reduced salt resistance, antioxidant capacity, and cell motility of the mutants compared with the wild-type strain. In conclusion, our results indicate that EPS can influence cellular Na+ content and antioxidant enzyme activity, as well as play an important role in the stress adaption and cell motility of M. alhagi CCNWXJ12-2T.

Global transcriptional response to salt shock of the plant microsymbiont Mesorhizobium loti MAFF303099.

Soil salinity affects rhizobia both as free-living bacteria and in symbiosis with the host. The aim of this study was to examine the transcriptional response of the Lotus microsymbiont Mesorhizobium loti MAFF303099 to salt shock. Changes in the transcriptome of bacterial cells subjected to a salt shock of 10% NaCl for 30 min were analyzed. From a total of 7231 protein-coding genes, 385 were found to be differentially expressed upon salt shock, among which 272 were overexpressed. Although a large number of overexpressed genes encode hypothetical proteins, the two most frequently represented COG categories are "defense mechanisms" and "nucleotide transport and metabolism". A significant number of transcriptional regulators and ABC transporters genes were upregulated. Chemotaxis and motility genes were not differentially expressed. Moreover, most genes previously reported to be involved in salt tolerance were not differentially expressed. The transcriptional response to salt shock of a rhizobium with low ability to grow under salinity conditions, but enduring a salinity shock, may enlighten us concerning salinity stress response mechanisms.

[Mutant construction and characterization of hfq in Mesorhizobium huakuii 7653R].

Objective: We studied the functions and characteristics of hfq gene in Mesorhizobium huakuii 7653R in adverse environment and symbiotic with its host plant. Methods: The hfq mutant of 7653R was constructed via homologous recombination with small cloned fragments on suicide plasmids pK19mob to insert target gene. We applied 7653RΔhfq to characterize stress tolerance and symbiosis with host plant, in comparison with the complementary strains 7653R △hfq-C and the wild type. Results: Mutant 7653RΔhfq presented lower growth rate, and higher mortality after heat shock-pretreated than that of the wild type, as well as the decreasing adaptability under the stress of 4.5% ethanol and 50 mmol H2O2. The defection of hfq affected the expression of some sRNAs in 7653R. Moreover, the mutant displayed significant reduced nodulation ability and nitrogenase activity compared with the wild type. Conclusion: As a crucial post transcriptional regulatory factor, hfq plays an important role in Mesorhizobium Huakuii 7653R on both processes of stress resistance and symbiosis with the host plant Astragalus sinicus L.

Ancient Heavy Metal Contamination in Soils as a Driver of Tolerant Anthyllis vulneraria Rhizobial Communities.

Anthyllis vulneraria is a legume associated with nitrogen-fixing rhizobia that together offer an adapted biological material for mine-soil phytostabilization by limiting metal pollution. To find rhizobia associated with Anthyllis at a given site, we evaluated the genetic and phenotypic properties of a collection of 137 rhizobia recovered from soils presenting contrasting metal levels. Zn-Pb mine soils largely contained metal-tolerant rhizobia belonging to Mesorhizobium metallidurans or to another sister metal-tolerant species. All of the metal-tolerant isolates harbored the cadA marker gene (encoding a metal-efflux PIB-type ATPase transporter). In contrast, metal-sensitive strains were taxonomically distinct from metal-tolerant populations and consisted of new Mesorhizobium genospecies. Based on the symbiotic nodA marker, the populations comprise two symbiovar assemblages (potentially related to Anthyllis or Lotus host preferences) according to soil geographic locations but independently of metal content. Multivariate analysis showed that soil Pb and Cd concentrations differentially impacted the rhizobial communities and that a rhizobial community found in one geographically distant site was highly divergent from the others. In conclusion, heavy metal levels in soils drive the taxonomic composition of Anthyllis-associated rhizobial populations according to their metal-tolerance phenotype but not their symbiotic nodA diversity. In addition to heavy metals, local soil physicochemical and topoclimatic conditions also impact the rhizobial beta diversity. Mesorhizobium communities were locally adapted and site specific, and their use is recommended for the success of phytostabilization strategies based on Mesorhizobium-legume vegetation. IMPORTANCE: Phytostabilization of toxic mine spoils limits heavy metal dispersion and environmental pollution by establishing a sustainable plant cover. This eco-friendly method is facilitated by the use of selected and adapted cover crop legumes living in symbiosis with rhizobia that can stimulate plant growth naturally through biological nitrogen fixation. We studied microsymbiont partners of a metal-tolerant legume, Anthyllis vulneraria, which is tolerant to very highly metal-polluted soils in mining and nonmining sites. Site-specific rhizobial communities were linked to taxonomic composition and metal tolerance capacity. The rhizobial species Mesorhizobium metallidurans was dominant in all Zn-Pb mines but one. It was not detected in unpolluted sites where other distinct Mesorhizobium species occur. Given the different soil conditions at the respective mining sites, including their heavy-metal contamination, revegetation strategies based on rhizobia adapting to local conditions are more likely to succeed over the long term compared to strategies based on introducing less-well-adapted strains.

Mesorhizobium olivaresii sp. nov. isolated from Lotus corniculatus nodules.

In this study four Mesorhizobium strains isolated from Lotus corniculatus nodules in Granada (Spain) were characterized. Their 16S rRNA gene sequences were closely related to those of M. albiziae LMG 23507T and M. chacoense Pr5T showing 99.4 and 99.2% similarity values, respectively. The analysis of concatenated rpoB, recA, atpD and glnII genes showed they formed a cluster with internal similarities higher than 97%. The closest species also were M. albiziae LMG 23507T and M. chacoense Pr5T showing similarity values lower than 92% in rpoB, recA and glnII genes and lower than 96.5% in the atpD gene. These results indicated that the L. corniculatus strains belong to a new species of genus Mesorhizobium which was confirmed by DNA-DNA hybridization and phenotypic characterization. Therefore a new species with the name Mesorhizobium olivaresii sp. nov. is proposed, and the type strain is CPS13T (LMG 29295T=CECT 9099T).

Low temperature adaptation and the effects of cryoprotectants on mesorhizobia strains.

In this study, the tolerance of Mesorhizobium sp. ACMP18, Mesorhizobium sp. USDA3350, and Mesorhizobium temperatum LMG23931 strains, to cold and freezing were investigated. The ability to withstand freezing at -20 °C and -70 °C for 24 months was different among the studied strains and depended on the cryoprotectant used. The survivability of mesorhizobial strains at -20 °C and -70 °C was significantly improved by some cryoprotectans (glycerol and sucrose/peptone). It is worth noting that the greatest resistance to freezing was detected when stress treatments were performed in glycerol as a cryoprotectant. Using PCR analysis, cspA genes were identified in the studied strains. Their nucleotide sequences were most similar to the sequences of the corresponding genes of the Mesorhizobium species. The expression of the cspA gene in the studied bacteria was analyzed using the RT-PCR technique. The fatty acid composition of the mesorhizobia was determined at 5, 10, 15, and 28 °C. It was noticed that growth temperature significantly affected the fatty acid composition and the amounts of unsaturated fatty acids, especially that of cis-vaccenic acid (18:1É·(11)), increased markedly in bacterial cells cultivated at 5, 10, and 15 °C.

[Characterization of phenotype and expression regulation of an RND-type multidrug efflux pump in Mesorhizobium huakuii 7653R].

Objective: To study the function of an RND family efflux pump encoded by MCHK_0866 and MCHK_0867 in Mesorhizobium huakuii 7653R. Methods: Genetic organization of target genes was analyzed in genome. The change of growth was observed by measuring OD600. Drug sensitivity was detected by minimal inhibitory concentrations; relative transcription level of target genes was measured by RT-PCR. Transcript regulation of the efflux pump was validated by bacterial one-hybrid system. Results: Proteins encoded by MCHK_0866 and MCHK_0867 formed an RND family efflux pump. The OD600 of growth curve reduced and it showed more sensitivity to nalidixic acid, tetracycline and SDS after disrupting the efflux pump. Genes relative transcription level increased in response to nalidixic acid treatment. Meanwhile, the downstream gene MCHK_0869 belongs to TetR family transcription factor and its expression product can interact with the promoter region of MCHK_0867. Conclusion: The efflux pump is possibly associated with the transportation of nalidixic acid and affects rhizobial free-living growth. The pump is putatively regulated by a downstream local transcription factor.

Copper tolerance mechanisms of Mesorhizobium amorphae and its role in aiding phytostabilization by Robinia pseudoacacia in copper contaminated soil.

The legume-rhizobium symbiosis has been proposed as an important system for phytoremediation of heavy metal contaminated soils due to its beneficial activity of symbiotic nitrogen fixation. However, little is known about metal resistant mechanism of rhizobia and the role of metal resistance determinants in phytoremediation. In this study, copper resistance mechanisms were investigated for a multiple metal resistant plant growth promoting rhizobium, Mesorhizobium amorphae 186. Three categories of determinants involved in copper resistance were identified through transposon mutagenesis, including genes encoding a P-type ATPase (CopA), hypothetical proteins, and other proteins (a GTP-binding protein and a ribosomal protein). Among these determinants, copA played the dominant role in copper homeostasis of M. amorphae 186. Mutagenesis of a hypothetical gene lipA in mutant MlipA exhibited pleiotropic phenotypes including sensitivity to copper, blocked symbiotic capacity and inhibited growth. In addition, the expression of cusB encoding part of an RND-type efflux system was induced by copper. To explore the possible role of copper resistance mechanism in phytoremediation of copper contaminated soil, the symbiotic nodulation and nitrogen fixation abilities were compared using a wild-type strain, a copA-defective mutant, and a lipA-defective mutant. Results showed that a copA deletion did not affect the symbiotic capacity of rhizobia under uncontaminated condition, but the protective role of copA in symbiotic processes at high copper concentration is likely concentration-dependent. In contrast, inoculation of a lipA-defective strain led to significant decreases in the functional nodule numbers, total N content, plant biomass and leghemoglobin expression level of Robinia pseudoacacia even under conditions of uncontaminated soil. Moreover, plants inoculated with lipA-defective strain accumulated much less copper than both the wild-type strain and the copA-defective strain, suggesting an important role of a healthy symbiotic relationship between legume and rhizobia in phytostabilization.

Global transcriptome analysis of Mesorhizobium alhagi CCNWXJ12-2 under salt stress.

BACKGROUND: Mesorhizobium alhagi CCNWXJ12-2 is a α-proteobacterium which could be able to fix nitrogen in the nodules formed with Alhagi sparsifolia in northwest of China. Desiccation and high salinity are the two major environmental problems faced by M. alhagi CCNWXJ12-2. In order to identify genes involved in salt-stress adaption, a global transcriptional analysis of M. alhagi CCNWXJ12-2 growing under salt-free and high salt conditions was carried out. The next generation sequencing technology, RNA-Seq, was used to obtain the transcription profiles. RESULTS: We have compared the transcriptome of M. alhagi growing in TY medium under high salt conditions (0.4 M NaCl) with salt free conditions as a control. A total of 1,849 differentially expressed genes (fold change ≧ 2) were identified and 933 genes were downregulated while 916 genes were upregulated under high salt condition. Except for the upregulation of some genes proven to be involved in salt resistance, we found that the expression levels of protein secretion systems were changed under high salt condition and the expression levels of some heat shock proteins were reduced by salt stress. Notably, a gene encoding YadA domain-containing protein (yadA), a gene encoding trimethylamine methyltransferase (mttB) and a gene encoding formate--tetrahydrofolate ligase (fhs) were highly upregulated. Growth analysis of the three gene knockout mutants under salt stress demonstrated that yadA was involved in salt resistance while the other two were not. CONCLUSIONS: To our knowledge, this is the first report about transcriptome analysis of a rhizobia using RNA-Seq to elucidate the salt resistance mechanism. Our results showed the complex mechanism of bacterial adaption to salt stress and it was a systematic work for bacteria to cope with the high salinity environmental problems. Therefore, these results could be helpful for further investigation of the bacterial salt resistance mechanism.

Genes commonly involved in acid tolerance are not overexpressed in the plant microsymbiont Mesorhizobium loti MAFF303099 upon acidic shock.

Rhizobia are legume bacterial symbionts that fix nitrogen in the root nodules of plants. The aim of the present study was to investigate the global transcriptional response of rhizobia upon an acidic shock. Changes in the transcriptome of cells of Mesorhizobium loti strain MAFF303099 upon an acidic shock at pH 3 for 30 min were analysed. From a total of 7,231 protein-coding genes, 433 were found to be differentially expressed upon acidic shock, of which 322 were overexpressed. Although most of the overexpressed genes encode hypothetical proteins, the two most represented Cluster of Orthologous Group (COG) categories are 'defence mechanisms' and 'transcription'. Differentially expressed genes are dispersed throughout the chromosome, with the exception of the symbiosis island, where most genes remain unchanged. A significant number of transcriptional regulators and ABC transporter genes are overexpressed. No overexpression of genes typically associated to acid tolerance in rhizobia, such as act and exo genes, was detected. Overall, this study suggests a transcriptional response to acidic shock of M. loti distinct from other rhizobia. Additional studies are in course to explore the role of some of the highly overexpressed genes and to further elucidate the molecular bases of acid stress response.

Genome-wide transcriptional responses of two metal-tolerant symbiotic Mesorhizobium isolates to zinc and cadmium exposure.

BACKGROUND: Mesorhizobium metallidurans STM 2683T and Mesorhizobium sp. strain STM 4661 were isolated from nodules of the metallicolous legume Anthyllis vulneraria from distant mining spoils. They tolerate unusually high Zinc and Cadmium concentrations as compared to other mesorhizobia. This work aims to study the gene expression profiles associated with Zinc or Cadmium exposure and to identify genes involved in metal tolerance in these two metallicolous Mesorhizobium strains of interest for mine phytostabilization purposes. RESULTS: The draft genomes of the two Mezorhizobium strains were sequenced and used to map RNAseq data obtained after Zinc or Cadmium stresses. Comparative genomics and transcriptomics allowed the rapid discovery of metal-specific or/and strain-specific genes. Respectively 1.05% (72/6,844) and 0.97% (68/6,994) predicted Coding DNA Sequences (CDS) for STM 2683 and STM 4661 were significantly differentially expressed upon metal exposure. Among these, a significant number of CDS involved in transport (13/72 and 13/68 for STM 2683 and STM 4661, respectively) and sequestration (15/72 and 16/68 for STM 2683 and STM 4661, respectively) were identified. Thirteen CDS presented homologs in both strains and were differentially regulated by Zinc and/or Cadmium. For instance, several PIB-type ATPases and genes likely to participate in metal sequestration were identified. Among the conserved CDS that showed differential regulation in the two isolates, we also found znuABC homologs encoding for a high affinity ABC-type Zinc import system probably involved in Zinc homeostasis. Additionally, global analyses suggested that both metals also repressed significantly the translational machinery. CONCLUSIONS: The comparative RNAseq-based approach revealed a relatively low number of genes significantly regulated in the two Mesorhizobium strains. Very few of them were involved in the non-specific metal response, indicating that the approach was well suited for identifying genes that specifically respond to Zinc and Cadmium. Among significantly up-regulated genes, several encode metal efflux and sequestration systems which can be considered as the most widely represented mechanisms of rhizobial metal tolerance. Downstream functional studies will increase successful phytostabilization strategies by selecting appropriate metallicolous rhizobial partners.

Molecular characterization of the Lotus japonicus NRT1(PTR) and NRT2 families.

Nitrate is an essential element for plant growth, both as a primary nutrient in the nitrogen assimilation pathway and as an important signal for plant development. Low- and high-affinity transport systems are involved in the nitrate uptake from the soil and its distribution between different plant tissues. By an in silico search, we identified putative members of both systems in the model legume Lotus japonicus. We investigated, by a time course analysis, the transcripts abundance in root tissues of nine and four genes encoding putative low-affinity (NRT1) and high-affinity (NRT2) nitrate transporters, respectively. The genes were sub-classified as inducible, repressible and constitutive on the basis of their responses to provision of nitrate, auxin or cytokinin. Furthermore, the analysis of the pattern of expression in root and nodule tissues after Mesorhizobium loti inoculation permitted the identification of sequences significantly regulated during the symbiotic interaction. The interpretation of the global regulative networks obtained allowed to postulate roles for nitrate transporters as possible actors in the cross-talks between different signalling pathways triggered by biotic and abiotic factors.

Draft genome sequence of Mesorhizobium alhagi CCNWXJ12-2T, a novel salt-resistant species isolated from the desert of northwestern China.

Mesorhizobium alhagi strain CCNWXJ12-2(T) is a novel species of soil-dwelling, nitrogen-fixing bacteria that can form symbiotic root nodules with Alhagi sparsifolia. Moreover, the strain has high resistance to salt and alkali. Here we report the draft genome sequence of Mesorhizobium alhagi strain CCNWXJ12-2(T). A large number of osmotic regulation-related genes have been identified.