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

Abstract

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 β- 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.

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

“…which can limit the development of rhizobia that prefer neutral or alkaline conditions during nodulation [ 63 ]. In addition, analysis of soil pH in Merchouch indicated that it is an acidic soil, and, therefore, we used acid tolerant rhizobia (MA72 and MA100; [ 9 ]) for inoculation to alleviate the effect of these produced acids during P solubilization.…”

“…The strains Mesorhizobium sp. MA72 and Mesorhizobium ciceri MA100 used in this study were isolated from chickpea nodules collected from chickpea fields in Morocco and characterized in our previous study [ 9 ]. They were tolerant to acidic pH 5.5–6.0 and other abiotic stresses (cold temperature of 4 °C, high temperature 36–40 °C, 513–684 mM of NaCl) and showed better symbiotic performance with chickpea [ 9 ].…”

“…Chickpea can fix 20–80% of its nitrogen requirement [ 5 , 6 ], which enhances its productivity and improves soil N content [ 6 , 7 , 8 ]. However, the establishment of this symbiosis is limited by environmental conditions such as drought, salinity, extreme temperatures, soil acidity [ 9 , 10 , 11 , 12 ] and nutrient availability such as nitrogen (N) and phosphate (P) [ 13 , 14 ]. These abiotic stresses may cause damage to the host plant and reduce rhizobial survival [ 15 , 16 ], especially that chickpea in Mediterranean regions is grown as rainfed crop in semi-arid areas prone to water stress and exposed to increasingly variable and extreme weather conditions [ 17 ].…”

“…To alleviate the effect of these stresses, it is necessary to combine stress tolerant cultivars and stress tolerant rhizobacteria [ 9 ]. Moreover, the co-inoculation of rhizobia with other plant growth promoting rhizobacteria (PGPR) such as Pseudomonas, Azotobacter, Bacillus, Erwinia, Serratia , etc.…”

“…PGPRs might be found either in the rhizosphere, in the phyllosphere or inside nodules in the endosphere [ 9 , 21 , 22 , 23 , 24 ]. They mediate plant growth promotion directly through phosphate solubilization, nitrogen fixation, and phytohormones synthesis [ 14 , 19 , 25 ], or indirectly by acting as biocontrol agents either by competing for niches and nutrients or by producing antimicrobial metabolites, siderophores, etc.…”