Biodiversity of populations of phosphate solubilizing rhizobia that nodulates chickpea in different in Spanish soils

Raúl Rivas, Encarna Velázquez, Pedro F. Mateos and Eustoquio Martínez-Molina

Departamento de Microbiología y Genética, Facultad de Farmacia, Edificio Departamental, Universidad de Salamanca, Salamanca 37007, SPAIN.

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We isolated strains nodulating chickpea in different Spanish soils and their ability to solubilize phosphates in vitro was checked using YED-P plates. To group the isolates a new procedure PCR-based, TP-RAPD, that uses the two primers that amplifies the 16S rDNA molecule to obtain patterns that are characteristic for each subspecies. Using TP-RAPD we found 10 groups. From each group we selected a strain to obtain the complete 16S rDNA sequence. According it, strains from groups I and II were colsely related to Mesorhizobium mediterraneum, strains from groups III, V and VIII were closely related to M. thiansanense, strains from groups IV and X are closely related to M. loti. Groups VI, VII and IX??? are closely related to M. amorpha. These results are in agreement with those obtained using LMW RNA profiles. However, the TP-RAPD patterns did not coincide with any of those of type strain identified using LMW RNA. These results suggest the existence of several subspecies within M. mediterraneum, M. thiansanense, M. amorpha and M. loti. This is in agreement with the data obtained by 16S rDNA sequencing because none of the strains studied shows a 100% similarity with those of the type strains from these species. Currently, a difference of 0.5% in 16S rRNA sequence is accepted as a variation corresponding to different subspecies of the same species. Only strains belong to groups I, II, III and IX were able to solubilize phosphate in plates.