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Phylogenetic diversity of rhizobial strains nodulating Robinia pseudoacacia L.

ZALF – Centre for Agricultural Landscape and Land Use Research, Institute of Primary Production and Microbial Ecology, Eberswalder Str. 84, D-15374 Müncheberg, Germany¹
Federal Research Centre for Forestry and Forest Products, Institute for Forest Genetics and Forest Tree Breeding, Eberswalder Chaussee 3, D-15377 Waldsieversdorf, Germany²

Author for correspondence: Andreas Ulrich. Tel: +49 33432 82345. Fax: +49 33432 82344. e-mail: aulrich{at}zalf.de

Lack of knowledge exists regarding the diversity of rhizobial strains nodulating black locust (Robinia pseudoacacia L.), which is a neophytic tree species widely distributed in Europe. Seventeen rhizobial strains isolated from nodules of black locust at a German location were examined by phenotypic characterization and 16S rDNA analysis. The isolates were classified in nine 16S rDNA genotypes using a set of seven endonucleases. Based on RFLP analysis and sequencing, the strains were shown to belong to the genera Mesorhizobium (76%) and Rhizobium (24%). Five genotypes were identical to the species Mesorhizobium amorphae, Mesorhizobium loti, Mesorhizobium huakuii, Rhizobium leguminosarum and Rhizobium tropici. A strong similarity between the 16S rDNA sequence of another two genotypes and M. amorphae (99·9%) as well as the Mesorhizobium strain R88b (99·8%) was found. The two remaining genotypes were classified in the genus Rhizobium, without a significant relationship at the species level. Comparing isolates nodulating Rob. pseudoacacia and Amorpha fruticosa, a parallel picture of phylogenetic diversity was detected with a range of phylogenetically different rhizobia and M. amorphae dominating. For this study, 18 rhizobial strains which had originally been isolated from a forest in Maryland where black locust is native were additionally analysed. Results revealed seven genotypes all belonging to the genus Mesorhizobium, with four genotypes identical to the isolates from the German sampling location. Whereas the genotype identical to M. amorphae dominated within the strains obtained from the German location, the dominance of a genotype identical to M. huakuii was found among the strains from the native location. Summarizing data from both locations, Rob. pseudoacacia was nodulated with various genomic species, most of which belonged to the genus Mesorhizobium. Concerning phenotypic features such as growth rate, pH tolerance or use of certain carbohydrates, most isolates corresponded to described species and genera. However, there were differences in salt tolerance between these isolates and the corresponding reference strains. Overall, the results demonstrated a high phenotypic and phylogenetic diversity of rhizobial strains nodulating Rob. pseudoacacia. This may be a characteristic of neophytic and other widely spread legumes and may contribute to the success of black locust as a pioneer tree species for the temperate zone.

Keywords: Mesorhizobium, Robinia pseudoacacia, black locust, 16S rDNA sequences, PCR-RFLP

The EMBL accession numbers for the sequences reported in this paper are AJ271898–AJ271902 for the Mesorhizobium strains Rob6, Rob8 and Rob23 and the Rhizobium strains Rob18 and Rob20, respectively.