The long-term goal of this research is to identify virulence factors that may be targeted for the design of effective vaccines and therapeutics for Neisseria gonorrhoeae (gonococci, GC), which causes gonorrhoeae and pelvic inflammatory disease, and Neisseria meningitidis (meningococci, MC), which causes septicemia and meningitis. The objective of this R03 application is to examine the role of the meningococcal disease-associated (MDA) prophage in the pathogenesis of GC and MC. The MDA prophage, which is found at multiple loci on the GC and MC genomes, encodes a potential toxin that is a homologue of the Vibrio cholera CTX prophage zonular occulens toxin (Zot) that alters epithelial and endothelial tight junctions. Thus, the MDA prophage Zot proteins in GC and MC have the potential to be involved in inflammation, sepsis, and/or invasion of the blood-brain barrier. The insertion (and excision) of the MDA phage DNA into the host chromosome to establish lysogeny and possible excision to initiation replication may be mediated by the transposase of the ISNgo2/3 elements, which are part of the phage genome. Defining the mechanism for the movement of the phage DNA is essential to understanding the role of the phage in GC and MC pathogenesis.
The specific aims are to: 1. Determine the activities of the meningococcal and gonococcal Zot proteins, and 2. Define the role of the ISNgo2/3 transposase, Irg, in directing insertion and/or excision of MDAphi DNA and controlling zot expression in MC and GC. Experimental approaches will include assays for zonular occludens toxin activity for Zot from MC and GC on epithelial and brain microvascular endothelial cell membranes;quantitative reverse transcriptase PCR to measure expression of zot and irg under different growth conditions;and quantitative PCR to detect movement and replication of MDAphi. This work is relevant to public health because there is no effective vaccine for Neisseria gonnorrhoeae or for the most prevalent serogroup (B) of Neisseria meningitidis in the U.S. The annual U.S. incidence of gonococcal disease is 130 in 100,000, while the incidence of meningococcal disease is 1 in 100,000. Despite the use of chemotherapeutics, 10-40% of invasive meningococcal infections are fatal and 10-15% of survivors have serious sequelae, including mental retardation and deafness.
