Direct monitoring of Rhizobium sp. (Cicer) in nodules and soil using lac Z fusions

We constructed lacZ fusions in Rhizobium sp. (Cicer) by random Tn5-lacZ mutagenesis. The lacZ⁺ fusants formed blue colonies on a Rhizobial minimal medium containing 5-bromo-4-chloro-3-indolyl--D-galactopyranoside (X-gal). Rhizobium sp. (Cicer) fusant HSL-2 was identified in nodules and soil in a mixed population on the basis of the lacZ⁺ phenotype. Nodule occupancy of inoculated Rhizobium sp. (Cicer) HSL-2 (lacZ⁺) was assessed by directly streaking the nodule sap on X-gal plates. This method revealed differences between rhizobia carrying identical antibiotic markers. The rhizobial population in soil was estimated by direct plate counts using a medium containing X-gal. Introduction of lacZ into the Rhizobium sp. thus provided a simple and direct method for identifying strains from nodules and soil.