Phenotypic, genomic and phylogenetic characteristics of rhizobia isolated from root nodules of Robinia pseudoacacia (bla
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ORIGINAL PAPER
Phenotypic, genomic and phylogenetic characteristics of rhizobia isolated from root nodules of Robinia pseudoacacia (black locust) growing in Poland and Japan Borena Mierzwa · Sylwia Wdowiak-Wróbel · Wanda Maiek
Received: 16 February 2009 / Revised: 8 July 2009 / Accepted: 27 July 2009 / Published online: 8 August 2009 © Springer-Verlag 2009
Abstract Rhizobial strains, rescued from the root nodules of Robinia pseudoacacia growing in Japan and Poland, were characterized for the phenotypic properties, genomic diversity as well as phylogeny and compared with the reference strains representing diVerent species and genera of nodule bacteria. They had a moderately slow growth rate, a low tolerance to antibiotics, a moderate resistance to NaCl and produced acid in yeast mannitol agar. Cluster analysis based on the phenotypic features divided all bacteria involved in this study into four phena, comprising: (1) Rhizobium sp. + Sinorhizobium sp., (2) Bradyrhizobium sp., (3) R. pseudoacacia microsymbionts + Mesorhizobium sp., and (4) Rhizobium galegae strains at similarity coeYcient of 74%. R. pseudoacacia nodule isolates and Mesorhizobium species were placed on a single branch clearly distinct from other rhizobium genera lineages. Strains representing R. pseudoacacia microsymbionts shared 98–99% 16S rDNA sequence identity with Mesorhizobium species and in 16S rDNA phylogenetic tree all these bacteria formed common cluster. The rhizobia tested are genomically heterogeneous as indicated by the AFLP (AmpliWed Fragment Length Polymorphism) method. The bacteria studied exhibited high degree of speciWcity for nodulation. Nitrogenase
Communicated by Ursula Priefer. Electronic supplementary material The online version of this article (doi:10.1007/s00203-009-0500-0) contains supplementary material, which is available to authorized users. B. Mierzwa · S. Wdowiak-Wróbel · W. Maiek (&) Department of Genetics and Microbiology, M. Curie-Skiodowska University, Akademicka 19 St., 20-033 Lublin, Poland e-mail: [email protected]
structural genes in these strains were located on 771–961 kb megaplasmids. Keywords Robinia pseudoacacia rhizobia · Taxonomy · Symbiotic plasmid
Introduction Rhizobia are soil bacteria that form nitrogen-Wxing root and/or stem nodules in leguminous plants. Until recently, this capacity has been exclusively attributed to the Rhizobium, Sinorhizobium, Mesorhizobium, and Bradyrhizobium species belonging to alpha proteobacteria. In the last 8 years, new legume-nodulating bacteria, related to those from the genera Blastobacter, Devosia, Methylobacter, Ochrobacterium, Burkholderia, and Ralstonia, have been described (Sawada et al. 2003; Willems 2006). Interestingly, the latter two belong to the beta subclass of proteobacteria. The modern classiWcation of the nodule bacteria is based on phenotypic as well as genotypic characteristics and includes also the 16S rRNA gene sequence analysis (Maiek and Sajnaga 1999; Willems 2006; Young 1996). Sequence comparisons of the small subunit of the ribo
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