Methods to evaluate nodulation competitiveness between Sinorhizobium meliloti strains using melanin production as a...

Three methods to evaluate the relative ability of different strains of Sinorhizobium meliloti to occupy nodules formed on alfalfa after co-inoculation were compare in this study. Results obtained using the three methods of evaluation together, provided insight into the relative nodulation competitiveness between two given sinorhizobial strains. A simple visual phenotypic marker, i.e., melanin production was used to distinguish individual strains in a given assay. As such, melanin producing strains were compared with melanin non-producing strains throughout this study. Method 1 required the use of an ELISA plate, took 35 min for the analysis of 40 nodules, and allowed strain identification by melanin production 2 days after nodule harvest. Method 2 required 3 h for the analysis of 40 nodules, used an ELISA plate, growth of bacteria on Petri dishes, and melanin production was analysed after 48 h of cell culture. Finally, method 3 involved the whole nodulated plant root, required less material than the above methods, and results were obtained after 24 h. Only method 2 was useful in determining if both a melanin producing strain and a melanin non-producing strain had occupied an individual nodule. Each of the three methods represented a rapid way of studying strain competition for field studies, using a natural trait as a marker.