The capacity of N. gonorrhoeae to evade innate defenses in the female genital tract is hypothesized to be multifactorial and complex. Antioxidant factors may protect gonococci from killing by reactive oxygen species produced by phagocytes. Sialyltranferase (Lst) and other factors promote evasion of complement-mediated defenses. Evidence that these factors protect gonococci against evasion of innate host defenses is based primarily on in vitro assays. With the support of the first award, we developed the first reproducible small animal model of gonococcal genital tract infection. This model provides us with a valuable and unique research tool to test gonococcal interactions with host innate defenses. To satisfy the need for in vivo studies on factors hypothesized to contribute to evasion of PMN and complement-mediated killing, here we will i.) measure the relative contribution of the known antioxidant defenses of N. gonorrhoeae (catalase, cytochrome C peroxidase, manganese uptake, methionine sulfoxide reductase) in protection from killing by human PMNs and in survival during experimental murine genital tract infection. We will construct single and double mutants in genes hypothesized to directly defend against oxidative stress (kat, ccp, mntC, msrA), and test their capacity to survive opsonophagocytic killing by human and murine PMNs, and to infect normal mice and NADPH oxidase-deficient mice; ii.) define the role of gonococcal sialyltransferase in conferring resistance to opsonophagocytic killing by murine PMNs and in enhancing survival of N. gonorrhoeae in the murine lower genital tract We will determine if Lst-deficient gonococci are more senstive to PMN killing due to increased uptake or the induction of a stronger respiratory burst. We will utilize C3 and C4-deficient mice and NADPH-deficient mice to test predictions made from PMN killing assays, iii.) Determine the basis for the observed increased infectivity of anaerobically grown N. gonorrhoeae for estradiol-treated mice and for increased resistance to the bactericidal activity of normal human serum. We will test mutants in genes that may confer increased survival in vivo as identified by DNA microarray technology to see if they are responsible for anaerobically-induced increased infectivity. We will assess the role of anaerobically induced nitrite reductase (AniA) in conferring an advantage in vivo and increased resistance to serum by testing the infectivity of an aniA mutant in mice, and by utilizing AniA-specific antiserum to block interactions between anaerobically grown gonococci and complement. ? ?
Showing the most recent 10 out of 25 publications
