Biological behavior of plasmid in Rhizobium sp. strain S25 from Tephrosia candida

Rhizobium sp. strain S25 was isolated from the nodule on Tephrosia candida in Hainan Province, China. The strain showed high stress tolerance. The plasmid profile of strain S25, examined by the Eckhardt procedure, indicated that the strain harbors only one plasmid with an estimated size of 150 kb. The plasmid was shown to carry nod and nif genes by hybridization with probes of nodABC and nifHDK genes. Plasmid curing was carried out using the Bacillus subtilis sacB to generate derivatives of strain S25. In comparison with the parent strain S25, the cured derivative lost its ability to nodulate the host plant. Loss of the plasmid reduced significantly the strain's tolerance to acid, nitrous, and multiple antibiotics. The properties of the cured strain also indicated that the plasmid was involved in carbon and nitrogen metabolism. Reintroduction of the plasmid from S25 in the cured derivative restored its original biological phenotypes.