The original specific aims of the project were: (i) Examination of the kinetics of carbohydrate transport. The kinetics of glucose, fructose and sucrose transport will be compared; (ii) identification and characterization of transport mechanisms for glucose, fructose and sucrose; (iii) isolation and characterization of transport defective mutants; (iv) characterization of complex polysaccharides as growth substrates; (v) development of a continuous culture system designed to examine carbohydrate transport. Conceptually there is no change in these specific aims and in the goals of the project; that is to elucidate factors important to the colonization of the oral cavity by Fusobacterium species and to develop strategies designed to specifically inhibit colonization of the oral cavity by this bacterium. However, increased emphasis will be placed on the genetics of carbohydrate utilization. This is primarily the result of two developments: (i) the aquisition of an anaerobe chamber, which greatly facilitates the genetic manipulation of the fusobacteria and (ii) the acquisition of a conjugative, sucrose metabolic plasmid compatible with Escherichia coli K-12 strains. The availability of this plasmid will provide a possible hybridization probe which will be used to identify sucrose genes in E. nucleatum as well as allow the cloning of these genes E. coli. Escherichia coli does not have the ability to metabolize sucrose without this plasmid.
