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

Three methods to evaluate the relative ability of different strains of Sino rhizobium meliloti to occupy nodules formed on alfalfa after co-inoculation were compare in this study. Results obtained using the three methods of ev aluation together, provided insight into the relative nodulation competitiv eness between two given sinorhizobial strains. A simple visual phenotypic m arker, i.e,, melanin production was used to distinguish individual strains in a given assay. As such, melanin producing strains were compared with mel anin non-producing strains throughout this study. Method 1 required the use of an ELISA plate, took 35 min fur the analysis of 40 nodules, and allowed strain identification by melanin production 2 days after nodule harvest. M ethod 2 required 3 h for the analysis of 40 nodules, used an ELISA plate, g rowth of bacteria on Petri dishes, acid melanin production was analysed aft er 48 h of cell culture. Finally, method 3 involved the whole nodulated pla nt root, required less material than the above methods, and results were ob tained after 24 h. Only method 2 was useful in determining if both a melani n producing strain and a melanin non-producing strain had occupied an indiv idual nodule. Each of the three methods represented a rapid way of studying strain competition for field studies, using a natural trait as a marl;er. (C) 2000 Elsevier Science B.V. All rights reserved.