Gonorrhea is a very common bacterial disease in this country and in the developing world. Together with chlamydial disease, it has a major impact on female reproductive health as a cause of chronic pelvic inflammatory disease and its complications, i.e., impaired fertility and ectopic pregnancies. Gonorrhea, as well as other STDs, enhances the transmissibility of HIV virus. A prominent surface antigen of this organism is LPS. This structure is subject to high frequency antigenic variation. Each of the variant LPS structures mimics human glycolipid structures. The biological advantage of these variations is unknown and difficult to study because of the instability of the LPS structure. The applicant's laboratory has characterized a locus encoding 5 glycosyl transferases responsible for the synthesis of the variable portion of the LPS. Three of the genes contain in their coding frames runs of guanosines which render these genes subject to high frequency frame-shift mutations. Inactivation-reactivation of the three genes accounts for the antigenic variation observed. This understanding allows the engineering of gonococcal mutants that express stably any desired LPS phenotype. This permits definitive studies of the biological phenotypes of the different LPS structures. The poly-G regions will be replaced with other codons for glycine by site directed mutagenesis and introduced into gonococci by transformation. The mutants will be tested for their ability to serve as substrates for the addition of sialic acid by the gonococcal sialic acid transferase. They will be assayed for their ability to invade monolayers of human epithelial cells. Transcytosis of these mutants through both epithelial and endothelial cells will be tested using a novel bilayer model. In collaborative studies the ability of the mutants to infect human fallopian tube organ cultures will be determined. Their ability to infect volunteers will be determined in another collaboration. The understanding of LPS genetics will be extended by cloning and characterization of the three glycosyl transferases that add sugar residues to the heptose residues.
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