All isolates of cluster II showed negative nitrate reduction besides urease production. Isolates of cluster 1 (PRNB 16, 28, 29) and cluster II (PRNB-34) also failed to produce urease. Clusters I, II, and III did not produce IAA and failed to grow in Bringer’s TY medium; in contrast clusters IV and V produced IAA and showed growth in TY medium. Further, clusters I, II, and III cross nodulated Vigna unguiculata, Cajanus cajan, Macrotyloma uniflorum, Dolichos lablab, and Arachis hypogaea, whereas clusters IV and V in addition nodulated in V. radiata. Vigna mungo, however, failed to nodulate

Selleck BMS-354825 in M. uniflorum. In contrast to isolates under all the clusters, isolates under cluster V produced acid to utilized carbon source, assimilated disaccharides (sucrose, lactose and maltose), and grew well at pH 10 and 3.0% NaCl concentration. They cross nodulated Vigna unguiculata, Cajanus cajan, and Macrotyloma uniflorum, and they were sensitive to tetracycline, chloramphenicol, and rifampicin. Amplification of the 16S rRNA gene of the isolated strains yielded a single

band of about 1450 base pairs, which corresponded to the expected size of the 16S rRNA gene. A preliminary blast search against the databases revealed buy Olaparib a high similarity between the 16S rRNA gene of strains, and three groups of rhizobia were identified

in Millettia pinnata nodules. Groups 1, 2 and 3 showed 99% similarities to Bradyrhizobium sp. GX5, Bradyrhizobium elkanii SEMIA5002, and Rhizobium sp. TANU14, respectively. However, subsequent alignment of all determined 16S rRNA gene sequences together with those of a number of rhizobial reference type strains was used to generate a phylogenetic tree, as described in Materials and methods. The phylogenetic analysis clustered the representative strains of 6-phosphogluconolactonase 16S rRNA gene with the type strains of B. yuanmingense, B. elkanii, and R. undicola, respectively (Fig. 3). The sequences of all the M. pinnata rhizobial isolates were submitted to the NCBI databank under different accession numbers (Table 3). As M. pinnata was introduced as the most important multipurpose tree for biodiesel production, it has become the most widespread legume in India and other parts of the world. This predominance has resulted from the massive planting of the species for multipurpose use in a broad edaphic range including urban and social forestry. As for reports on nodulation from different parts of the world (Allen & Allen, 1981; Ather, 2005), we also found that soils collected from different regions of Andhra Pradesh, Karnataka, and Maharashtra of India contained rhizobial isolates able to nodulate Millettia pinnata.