Abstract
The diversity of Bradyrhizobium strains nodulating three cowpea (Vigna unguiculata L. Walp.) cultivars in favorable and water-limited conditions occuring at flowering was analysed. PCR- Restriction Fragment Length Polymorphism (PCR-RFLP) analysis of 16S-23S rDNA intergenic spacer region (IGS) directly applied on 85 crushed nodules distinguished four genetic profiles, IGS types I, II, III and IV. The distribution of these IGS types according to water conditions and cowpea cultivars (B-21, TN 88-63 and Mouride) showed that nodulating strains appeared more diverse in water-limited condition. More than three quarters of prospected nodules presented the IGS type I. They were formed on all three cultivars and in both water conditions. Only a small part of nodules was distributed between the IGS type II, III and IV. Nodules showing the IGS types II and III were found mainly in limited conditions on TN 88-63 and Mouride cultivars, whereas nodules presenting the IGS type IV were collected only from cultivars B-21 and Mouride, in both water conditions. Strains corresponding to the different profiles were isolated. The phylogenetic analysis of 16S rRNA gene sequences showed that they belong to the genus, Bradyrhizobium. The sequence analysis of 16S-23S rDNA IGS revealed that the strains exhibiting IGS types II, III and IV were closely related to some Faidherbia albida isolates from Senegal. IGS type II can be assigned with at least 98% similarity to Bradyrhizobium genospecies IV. IGS types III and IV showed more than 96% similarity with genospecies VII and could belong to the same genospecies. IGS type I, the most frequent, exhibits low IGS similarity with reported sequences in the databases, and could represent a new genospecies. (African Journal of Biotechnology: 2003 2(1): 13-23)
Highlights
Cowpea (Vigna unguiculata (L.) Walp.) is one of the earliest plants cultivated by man
We report here the genetic characterization and phylogeny of indigenous rhizobial populations isolated from cowpea cultivated in Senegal under favorable and water limited conditions occuring at flowering, a situation frequently occuring in Senegal. 85 nodules were analyzed by PCR-Restriction Fragment Length Polymorphism (PCR-RFLP) of 16S-23S rDNA intergenic spacer region (IGS) directly on DNA extracted from crushed nodules
The two RFLP profiles obtained for each nodule allowed for the distribution of the 85 nodules into four distinct restriction patterns named IGS types I, II, III and IV (Table 1; Figures 1A and 1B)
Summary
Cowpea (Vigna unguiculata (L.) Walp.) is one of the earliest plants cultivated by man. Rhizobial populations in tropical soils represent an important reservoir from which superior strains adapted to environmental stresses such as drought can be selected (Aurag and Brhada, 1995; Mpepereki et al, 1997). Cowpea rhizobia were first classified in an heterogeneous group of slow-growing rhizobia nodulating promiscuous tropical and subtropical legume species known as ‘cowpea cross-inoculation group’ (Allen and Allen, 1981). They were later transferred to the genus Bradyrhizobium. We report here the genetic characterization and phylogeny of indigenous rhizobial populations isolated from cowpea cultivated in Senegal under favorable and water limited conditions occuring at flowering, a situation frequently occuring in Senegal. Strains corresponding to the different profiles were isolated, and their phylogenetic positions determined by 16S rDNA and 16S-23S IGS sequence analysis
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