Co-Inoculation of Mesorhizobium ciceri with Either Bacillus sp. or Enterobacter aerogenes on Chickpea Improves Growth and Productivity in Phosphate-Deficient Soils in Dry Areas of a Mediterranean Region.

Biological nitrogen fixation requires a large amount of phosphorus (P). However, most of the soils are P-deficient and the extensive use of P- chemical fertilizers constitute a serious threat to the environment. In this context, two field experiments were carried out to investigate the effect of co-inoculation of Mesorhizobium ciceri with phosphate solubilizing bacteria (PSB), Bacillus sp., and Enterobacter aerogenes, on chickpea as an alternative to chemical nitrogen (N) and phosphorous fertilizers in P-deficient soils in dry areas of Morocco. The results revealed that combined inoculation of chickpea with rhizobia and PSB showed a significant enhancement of chickpea nodulation, biomass production, yields and N, P, and protein content in grains as compared to single inoculation or single application of N or P. A significantly higher increase was obtained by inoculating chickpea with Mesorhizobium sp. MA72 combined with E. aerogenes P1S6. This combination allowed an enhancement of more than 270% in nodulation, 192% in shoot dry weight and 242% in grain yield. The effect of this combination was equivalent to the effect of combined application of N and P fertilizers. Formulation of biofertilizers based on tasted strains could be used for chickpea co-inoculation in P-deficient soils for an eco-friendly sustainable production of chickpea.

Development and Proof-of-Concept Application of Genome-Enabled Selection for Pea Grain Yield under Severe Terminal Drought.

Terminal drought is the main stress limiting pea (Pisum sativum L.) grain yield in Mediterranean environments. This study aimed to investigate genotype × environment (GE) interaction patterns, define a genomic selection (GS) model for yield under severe drought based on single nucleotide polymorphism (SNP) markers from genotyping-by-sequencing, and compare GS with phenotypic selection (PS) and marker-assisted selection (MAS). Some 288 lines belonging to three connected RIL populations were evaluated in a managed-stress (MS) environment of Northern Italy, Marchouch (Morocco), and Alger (Algeria). Intra-environment, cross-environment, and cross-population predictive ability were assessed by Ridge Regression best linear unbiased prediction (rrBLUP) and Bayesian Lasso models. GE interaction was particularly large across moderate-stress and severe-stress environments. In proof-of-concept experiments performed in a MS environment, GS models constructed from MS environment and Marchouch data applied to independent material separated top-performing lines from mid- and bottom-performing ones, and produced actual yield gains similar to PS. The latter result would imply somewhat greater GS efficiency when considering same selection costs, in partial agreement with predicted efficiency results. GS, which exploited drought escape and intrinsic drought tolerance, exhibited 18% greater selection efficiency than MAS (albeit with non-significant difference between selections) and moderate to high cross-population predictive ability. GS can be cost-efficient to raise yields under severe drought.

Phenotypic and Genotypic Diversity Among Symbiotic and Non-symbiotic Bacteria Present in Chickpea Nodules in Morocco.

Environmental pollution problems and increased demand for green technologies in production are forcing farmers to introduce agricultural practices with a lower impact on the environment. Chickpea (Cicer arietinum) in arid and semi-arid environments is frequently affected by harsh environmental stresses such as heat, drought and salinity, which limit its growth and productivity and affect biological nitrogen fixation ability of rhizobia. Climate change had further aggravated these stresses. Inoculation with appropriate stress tolerant rhizobia is necessary for an environmentally friendly and sustainable agricultural production. In this study, endophytic bacteria isolated from chickpea nodules from different soil types and regions in Morocco, were evaluated for their phenotypic and genotypic diversity in order to select the most tolerant ones for further inoculation of this crop. Phenotypic characterization of 135 endophytic bacteria from chickpea nodules showed a wide variability for tolerance to heavy metals and antibiotics, variable response to extreme temperatures, salinity, pH and water stress. 56% of isolates were able to nodulate chickpea. Numerical analysis of rep-PCR results showed that nodulating strains fell into 22 genotypes. Sequencing of 16S rRNA gene of endophytic bacteria from chickpea nodules revealed that 55% of isolated bacteria belong to Mesorhizobium genus. Based on MLSA of core genes (recA, atpD, glnII and dnaK), tasted strains were distributed into six clades and were closely related to Mesorhizobium ciceri, Mesorhizobium opportunistum, Mesorhizobium qingshengii, and Mesorhizobium plurifarium. Most of nodulating strains were belonging to a group genetically distinct from reference Mesorhizobium species. Three isolates belong to genus Burkholderia of the class ß- proteobacteria, and 55 other strains belong to the class γ- proteobacteria. Some of the stress tolerant isolates have great potential for further inoculation of chickpea in the arid and semiarid environments to enhance biological nitrogen fixation and productivity in the context of climate change adaptation and mitigation.

Global-level population genomics reveals differential effects of geography and phylogeny on horizontal gene transfer in soil bacteria.

Although microorganisms are known to dominate Earth's biospheres and drive biogeochemical cycling, little is known about the geographic distributions of microbial populations or the environmental factors that pattern those distributions. We used a global-level hierarchical sampling scheme to comprehensively characterize the evolutionary relationships and distributional limitations of the nitrogen-fixing bacterial symbionts of the crop chickpea, generating 1,027 draft whole-genome sequences at the level of bacterial populations, including 14 high-quality PacBio genomes from a phylogenetically representative subset. We find that diverse Mesorhizobium taxa perform symbiosis with chickpea and have largely overlapping global distributions. However, sampled locations cluster based on the phylogenetic diversity of Mesorhizobium populations, and diversity clusters correspond to edaphic and environmental factors, primarily soil type and latitude. Despite long-standing evolutionary divergence and geographic isolation, the diverse taxa observed to nodulate chickpea share a set of integrative conjugative elements (ICEs) that encode the major functions of the symbiosis. This symbiosis ICE takes 2 forms in the bacterial chromosome-tripartite and monopartite-with tripartite ICEs confined to a broadly distributed superspecies clade. The pairwise evolutionary relatedness of these elements is controlled as much by geographic distance as by the evolutionary relatedness of the background genome. In contrast, diversity in the broader gene content of Mesorhizobium genomes follows a tight linear relationship with core genome phylogenetic distance, with little detectable effect of geography. These results illustrate how geography and demography can operate differentially on the evolution of bacterial genomes and offer useful insights for the development of improved technologies for sustainable agriculture.

Elucidation of functional chemical groups responsible of compost phytotoxicity using solid-state 13C NMR spectroscopy under different initial C/N ratios.

More than 1 million tons of fresh organic wastes is produced in the Souss-Massa region in Morocco. Tomato organic residues represent more than 25% of the total organic wastes and are deposited in uncontrolled landfills. Thus, composting can represent a valuable and pertinent solution to this environmental problem. The objectives of this experiment are to identify the potential functional groups responsible for compost phytotoxicity and to determine the optimum initial carbon to nitrogen ratio (C/N) for maximum recovery of tomato residues. The experiment consisted of the variation of the initial C/N ratios (25, 30, 35, and 40) using mixtures of different raw materials (tomato residues, melon residues, olive mill pomace, and sheep manure). Physicochemical parameters (pH, electrical conductivity, C/N ratio, and humic acid/fulvic acid ratio) were determined and spectroscopic analyses (UV-vis and NMR-13C) were performed during the composting process along with quality parameters (germination and phytotoxicity tests) at the end. The results showed that the compost with the initial C/N ratio of 35 is the most humified with the least phytotoxic effect. The germination and phytotoxicity tests were negatively correlated with the methoxyl/N-alkyl-C ratio and O-alkyl-C. These two functional groups are probably the origin of phytotoxicity expression in compost quality tests. Thus, a simple and precise quality test could be performed to evaluate directly the phytotoxicity and maturity of compost.

Phenotypic and genetic diversity in Sinorhizobium meliloti and S. medicae from drought and salt affected regions of Morocco.

BACKGROUND: Sinorhizobium meliloti and S. medicae are symbiotic nitrogen fixing bacteria in root nodules of forage legume alfalfa (Medicago sativa L.). In Morocco, alfalfa is usually grown in marginal soils of arid and semi-arid regions frequently affected by drought, extremes of temperature and soil pH, soil salinity and heavy metals, which affect biological nitrogen fixing ability of rhizobia and productivity of the host. This study examines phenotypic diversity for tolerance to the above stresses and genotypic diversity at Repetitive Extragenic Pallindromic DNA regions of Sinorhizobium nodulating alfalfa, sampled from marginal soils of arid and semi-arid regions of Morocco. RESULTS: RsaI digestion of PCR amplified 16S rDNA of the 157 sampled isolates, assigned 136 isolates as S. meliloti and the rest as S. medicae. Further phenotyping of these alfalfa rhizobia for tolerance to the environmental stresses revealed a large degree of variation: 55.41%, 82.16%, 57.96% and 3.18% of the total isolates were tolerant to NaCl (>513 mM), water stress (-1.5 MPa), high temperature (40 degrees C) and low pH (3.5), respectively. Sixty-seven isolates of S. meliloti and thirteen isolates of S. medicae that were tolerant to salinity were also tolerant to water stress. Most of the isolates of the two species showed tolerance to heavy metals (Cd, Mn and Zn) and antibiotics (chloramphenicol, spectinomycin, streptomycin and tetracycline). The phenotypic clusters observed by the cluster analysis clearly showed adaptations of the S. meliloti and S. medicae strains to the multiple stresses. Genotyping with rep-PCR revealed higher genetic diversity within these phenotypic clusters and classified all the 157 isolates into 148 genotypes. No relationship between genotypic profiles and the phenotypes was observed. The Analysis of Molecular Variance revealed that largest proportion of significant (P < 0.01) genetic variation was distributed within regions (89%) than among regions (11%). CONCLUSION: High degree of phenotypic and genotypic diversity is present in S. meliloti and S. medicae populations from marginal soils affected by salt and drought, in arid and semi-arid regions of Morocco. Some of the tolerant strains have a potential for exploitation in salt and drought affected areas for biological nitrogen fixation in alfalfa.