Two Major Clades of Bradyrhizobia Dominate Symbiotic Interactions with Pigeonpea in Fields of Côte d'Ivoire

Abstract

In smallholder farms of Côte d'Ivoire, particularly in the northeast of the country, Cajanus cajan (pigeonpea) has become an important crop because of its multiple beneficial facets. Pigeonpea seeds provide food to make ends meet, are sold on local markets, and aerial parts serve as forage for animals. Since it fixes atmospheric nitrogen in symbiosis with soil bacteria collectively known as rhizobia, C. cajan also improves soil fertility and reduces fallow time. Yet, seed yields remain low mostly because farmers cannot afford chemical fertilizers. To identify local rhizobial strains susceptible to be used as bio-inoculants to foster pigeonpea growth, root nodules were collected in six fields of three geographically distant regions of Côte d'Ivoire. Nodule bacteria were isolated and characterized using various molecular techniques including matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry (MS) and DNA sequencing. These molecular analyses showed that 63 out of 85 nodule isolates belonged to two major clades of bradyrhizobia, one of which is known as the Bradyrhizobium elkanii super clade. Phylogenies of housekeeping (16S-ITS-23S, rpoB) and symbiotic (nifH) genes were not always congruent suggesting that lateral transfer of nitrogen fixation genes also contributed to define the genome of these bradyrhizobial isolates. Interestingly, no field-, plant-, or cultivar-specific effect was found to shape the profiles of symbiotic strains. In addition, nodule isolates CI-1B, CI-36E, and CI-41A that belong to distinct species, showed similar symbiotic efficiencies suggesting that any of these strains might serve as a proficient inoculant for C. cajan.

Cowpea (Vigna unguiculata L. Walp) hosts several widespread bradyrhizobial root nodule symbionts across contrasting agro-ecological production areas in Kenya

“…MALDI-TOF MS protein profiling allows for high sample throughput at moderate costs without compromising on resolution. The method was already applied several times on rhizobia ( Ferreira et al, 2011 ; Ziegler et al, 2012 ; Sanchez-Juanes et al, 2013 ; Ziegler et al, 2015 ; Fossou et al, 2016 ). It was for instance used to discriminate and detect Bradyrhizobium strains from nodules of Lupinus in Spain ( Sanchez-Juanes et al, 2013 ) and most recently nodules of pigeonpea in Côte d’Ivoire ( Fossou et al, 2016 ).…”

Sinorhizobium fredii Strains HH103 and NGR234 Form Nitrogen Fixing Nodules With Diverse Wild Soybeans (Glycine soja) From Central China but Are Ineffective on Northern China Accessions

“…Plants were grown in controlled conditions with a 12 h photoperiod, a day temperature of 27°C and a night temperature of 20°C. To confirm the identity of bacteria found inside nodules, the 5′-end of the 23S rRNA genes of nodule bacteria was amplified using total genomic DNA of nodules as templates ( Fossou et al, 2016 ) together with the 23S-For1 (5′-ACGAACTAGTGTCAAAAGGGC-3′) and ITS-Rev2 (5′-TGGTCCGCGTTCGCTCGCC-3′) primers. The resulting amplicons of NGR234 (317 bp) and HH103 (380 bp) were clearly discriminated after migration on a 1% agarose gel.…”

Bradyrhizobium ivorense sp. nov. as a potential local bioinoculant for Cajanus cajan cultures in Côte d’Ivoire

“…Accordingly, a polyphasic approach was applied to determine the taxonomic status of the 19 pigeonpea isolates of Côte d’Ivoire grouped in cluster II. Isolates CI-1B T , CI-14A, CI-19D and CI-41S were selected as representative strains because they originated from different fields and regions of Côte d’Ivoire (Table S1, available in the online version of this article) and possessed protein mass spectra that reflected the genetic diversity existing within cluster II isolates [ 30 ]. For long term preservation, 30 % glycerol stocks were kept at −60 °C and bacteria were routinely grown in/on Rhizobium minimal medium (RMM) supplemented with 12 mM succinate (RMS) [ 37 ], or in/on RMM containing alternative carbon sources when specific metabolic analyses were needed.…”

“…For MLSA studies, the dnaK , glnII , gyrB , recA and rpoB marker genes were amplified using procedures described in Fossou et al . [ 30 ] and primers specifically designed for bradyrhizobia sequences (Table S2). Marker genes were sequenced on both strands using Sanger sequencing.…”

“…A number of recent surveys identified bradyrhizobia in sub-Saharan countries, however, including in Angola and Namibia [ 27 ], Ethiopia [ 14 ], Kenya [ 28 ] and Senegal [ 29 ]. In addition, a sampling of six fields in three geographically distant regions of Côte d’Ivoire has led to the molecular characterisation of >80 pigeonpea isolates by matrix-assisted laser desorption/ionization (MALDI) time of flight (TOF) mass spectrometry (MS) and DNA sequencing [ 30 ]. That data indicated that 63 of the 85 nodule isolates (74 %) were bradyrhizobia, only one of which (strain CI-41A) belonged to the B. japonicum super clade.…”