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We studied a collection of 126 rhizobial isolates from eight species of Crotalaria (C. comosa, C. glaucoides, C. goreensis, C. hyssopifolia, C. lathyroides, C. perrottetii, C. podocarpa, and C. retusa) growing in Senegal. Nodulation and nitrogen-fixation tests on nine Crotalaria species revealed two specificity groups within the genus Crotalaria. Group I consists of plants solely nodulated by very specific fast-growing strains. Group II plants are nodulated by slow-growing strains similar to promiscuous Bradyrhizobium spp. strains already reported to nodulate many tropical legumes. SDS-PAGE studies showed that slow-growing strains grouped with Bradyrhizobium while fast-growing strains constituted a homogeneous group distinct from all known rhizobia. Amplified ribosomal DNA restriction analysis (ARDRA) of 10 representative strains of this group using four restriction enzymes showed a single pattern for each enzyme confirming the high homogeneity of group I. The 16S rDNA sequence analysis revealed that this specific group belonged to the genus Methylobacterium, thus constituting a new branch of nodulating bacteria.

We studied a collection of 126 rhizobial isolates from eight species of Crotalaria (C. comosa, C. glaucoides, C. goreensis, C. hyssopifolia, C. lathyroides, C. perrottetii, C. podocarpa, and C. retusa) growing in Senegal. Nodulation and nitrogen-fixation tests on nine Crotalaria species revealed two specificity groups within the genus Crotalaria. Group I consists of plants solely nodulated by very specific fast-growing strains. Group II plants are nodulated by slow-growing strains similar to promiscuous Bradyrhizobium spp. strains already reported to nodulate many tropical legumes. SDS-PAGE studies showed that slow-growing strains grouped with Bradyrhizobium while fast-growing strains constituted a homogeneous group distinct from all known rhizobia. Amplified ribosomal DNA restriction analysis (ARDRA) of 10 representative strains of this group using four restriction enzymes showed a single pattern for each enzyme confirming the high homogeneity of group I. The 16S rDNA sequence analysis revealed that this specific group belonged to the genus Methylobacterium, thus constituting a new branch of nodulating bacteria.

Polymerase Chain Reaction/Restriction Fragment Length Polymorphism (PCR/RFLP) of the InterGenic Spacer (IGS) between rDNA 16S and 23S was used to identify indigenous strains nodulating four clones of Acacia mangium-Acacia auriculiformis hybrids cultivated in non-sterilized sandy soil from Sangalkam (Senegal) under greenhouse conditions. The experiment was for 4 months. The analysis of restriction fragment length polymorphism obtained with MspI and HaeIII restriction enzymes allowed the identification of 15 different IGS Groups with a distribution which significantly differed according to the clone of the hybrid (strains of one clone can belong to three and five different IGS Groups). Three large multi-lobed nodules were obtained on the root system of clone 3.26 within 5 months. Also, the nature of the rhizobia contained in each lobe was determined. The results showed that the lobes of large nodules can be occupied by one or two strains and the nodules analysed were mainly occupied by those belonging to IGS Group 12.

No abstract

A greenhouse experiment was conducted to measure nitrogen fixation in three Crotalaria species : C. ochroleuca, C. perrottetii and C. retusa growing in Senegal by using 15 N direct isotope dilution technique. Two non-fixing plants, Senna obtusifolia and Senna occidentalis served as reference plants. The amount of nitrogen fixed two months after planting was obtained using the average of the two reference plants. The atom % 15 N excess in the Crotalaria species was significantly lower than that of the reference plants, indicating that significant nitrogen fixation occurred in the three plants. Significant differences were observed between the Crotalaria species; C. ochroleuca yielded more dry matter weight and total nitrogen than did C. perrottetti and C. retusa. The % nitrogen derived from atmosphere (%Ndfa) in leaves and stems was also higher in C. ochroleuca. There was no significant difference in %Ndfa in the whole plant between the three Crotalaria species (47% to 53%). In contrast, interspecific variability was observed based on the %Ndfa. C. ochroleuca significantly exhibited the higher amount of total nitrogen fixed, equivalent to 83 kg of nitrogen fixed per hectare. Based on these data, it was concluded that C. ochroleuca could be used in multiple cropping systems in Senegal for making more nitrogen available to other plants.

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We have isolated 17 strains of rhizobiums from nodules collected on roots of Pterocarpus officinalis in different sites of swamp forest in Guadeloupe and Martinique. The genotypic diversity of these strains were characterized by PCR-RFLP analysis of the IGS (intergenic spacer) region between 16S and 23S rRNA genes, by using two 4-base restriction endonucleases Hae III and Msp I. Results of IGS PCR-RFLP analysis showed that restriction patterns were similar for DNA extract from root nodules and cultures of rhizobium strains. The diversity of these strains varied from one site to another. They were grouped into 5 ribotypes by IGS PCR-RFLP analysis. Results of IGS sequence analysis showed that the 5 ribotypes are very close to the genus Bradyrhizobium. The symbiotic properties of the 5 ribotypes were evaluated on Acacia seyal, a plant test that is able to form nodules with several Bradyrhizobium species. One month and nineteen days after inoculation, the green colour of the leaves and the best growth of inoculated seedlings contrasting with the yellow colour of the leaves and the lowest growth of uninoculated seedlings, were the parameters indicating the level of nitrogen-fixing activity of the three Bradyrhizobium strains. Concurrently, the vesicular and arbuscular mycorrhizal status of P. officinalis was established. The two types of symbiotic microorganisms could contribute to mobilize available phosphorus and nitrogen for improving the growth of P. officinalis in degraded swamp forests. RÉSUMÉNous avons isolé 17 souches de rhizobiums à partir de nodules récoltés sur des racines de Pterocarpus officinalis dans différents sites de forêt marécageuse en Guadeloupe et Martinique. La diversité de ces souches a été caractérisée par PCR-RFLP de l'intergène IGS situé entre les gènes nucléaires codant pour les ARNr 16S et 23S en utilisant deux enzymes de restriction Hae III et Msp I. Des profils RFLP obtenus par digestion de l'IGS d'extraits d'ADN de nodules ou de cultures bactériennes sont comparables. L'analyse de ces profils RFLP a révélé une hétérogénéité génétique des souches qui varie suivant les sites. Les 17 souches de P. officinalis ont été groupées en 5 ribotypes par comparaison de leur profil RFLP. L'analyse phylogénétique de l'IGS a montré que ces ribotypes sont très proches du genre Bradyrhizobium. Les propriétés symbiotiques d'un représentant de chaque groupe de souches ont été testées sur Acacia seyal, espèce qui nodule avec un grand nombre d'espèces de Bradyrhizobium.