Loss of effectiveness and infectivity in mutants of Rhizobium resistant to metabolic inhibitors.

Abstract

Association between resistance to metabolic inhibitors and ability for symbiosis was investigated for isolates of Rhizobium blocked at different stages of nodule development. Approximately 530 spontaneous mutants were isolated from symbiotically effective strains (mainly R. leguminosarum and R. trifolii) for resistance to one of 21 inhibitors, mainly structural analogues of L-amino acids. Mutants resistant to fluorophenylalanine, norleucine, norvaline, D-leucine, D-methionine, D-histidine, or D-alanine comprised a group having several characteristics in common. Those derived from R. leguminosarum strains were typically non-nodulating (non-infective) on pea seedlings while those from R. trifolii strains were ineffective on red clover. Loss or non-loss of parental ability to form effective nodules could be predicted with about 85% accuracy on the basis of an additional criterion, namely collateral sensitivity to D-aspartic or D-glutamic acid. Limited bioassays for metabolic excretion showed no consistent change in excretion for any class of mutant. In some of the mutants, partial auxotrophy was associated with resistance to inhibitors; in one strain of R. trifolii the nutritional requirement appeared to be related also to loss of effectiveness. Loss of symbiotic ability was most pronounced for mutants resistant to amino acid antagonists which most readily induced spheroplast formation in culture. Cell wall or membrane defects are considered as one likely basis for ineffectiveness in these mutants.