Rhizobium trifolii is a gram-negative bacterium which specifically infects clover root hairs and forms a root nodule symbiosis which fixes atmospheric nitrogen into ammonia. The overall aim of this research is to identify the events leading to successful infection in the Rhizobium trifolii-clover symbiosis as a model for cellular recognition between procaryotic and eucaryotic cells. The focus is on the specific interaction between a clover lectin, called trifoliin A, and polysaccharides on the bacterial surface during the infection process. The lectin-binding capsular polysaccharide (CPS) and lipopolysaccharide (LPS) of R. trifolii are biologically active in the infection process (affect infection thread formation in clover root hairs) at very low concentrations. Recently it has become possible to enzymatically depolymerize the capsular polysaccharide into structurally analyzable oligosaccharides which are still able to bind trifoliin A and influence infection thread formation as a measure of biological activity.
The specific aim of this proposal is to make use of these oligosaccharides to establish the chemical structures of the capsular polysaccharide necessary for specific binding to trifoliin A and enhancement of root hair infection. We also propose to compare the LPS and CPS from wild type R. trifolii strains (0403 & 843) and from a selected collection of mutant derivatives for their ability to interact specifically with trifoliin A and influence clover root hair infection. The collection of mutant strains will include non-infective derivatives of the wild type strains obtained by transposon mutagenesis, and a new class of mutant strains of R. trifolii 0403 which would be selected and screened for the inability to bind to trifoliin A. An analysis of the lectin-binding and invasive properties of these strains should establish whether the ability of the bacteria to produce trifoliin A receptor molecules is necessary for root hair infection. This work would identify the molecular basis for a specific cellular recognition event between a procaryote and an eucaryote. This information is relevant to studies which identify the underlying mechanisms of microbial adhesion and subsequent invasion of eucaryotic host cells as they relate to microbial pathogenesis and the establishment of the normal microbial flora in man.
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