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Neisseria Gonorrhoeae Defense Against Human Neutrophils and Host-Derived Antimicrobials

Handing, Jonathan
Format
Thesis/Dissertation; Online
Author
Handing, Jonathan
Advisor
Hewlett, Erik
Kendall, Melissa
Casanova, James
Criss, Alison
Brautigan, David
Abstract
Neisseria gonorrhoeae (Gc) is the sole causative agent of the disease gonorrhea. More than 100 million people are infected annually, and reported infection numbers are consistently on the rise. There are clinical isolates of Gc that have developed resistance to the latest third generation cephalosporins, which is the last recommended line of monotherapy for disease treatment, and there is still no available gonococcal vaccine. Following transmission, Gc comes into contact with host mucosal surfaces. Host epithelial cells recognize the pathogen, and respond by releasing both antimicrobials to combat Gc and proinflammatory cytokines to alert the immune system. The cytokine gradient favors recruitment of large numbers of neutrophils to the site of infection. Neutrophils mount a robust immune response characterized by phagocytosis of Gc, production of reactive oxygen species by NADPH oxidase, formation of NETs and release of antimicrobials that are stored in cytoplasmic granules. Despite the potent immune response, Gc resists complete clearance, which suggests that Gc has defensive mechanisms against killing by neutrophils and host-derived antimicrobials at mucosal surfaces. Herein, I investigated the contribution of two important virulence factors, an LOS-modifying enzyme LptA and a clinically relevant antimicrobial efflux pump MtrCDE, for protection of Gc during interactions with human neutrophils. In addition, I explored the antigonococcal activity of a host-derived antimicrobial, β-defensin 22, and characterized its potential for therapeutic application. Gc interactions with neutrophils are not well understood. Though several important observations by our lab and other groups have advanced our understanding, there are still many unknowns. I have identified that LptA and MtrCDE are important for protecting Gc from killing by neutrophils. Previously, LptA was only known to protect Gc from killing by the complement component of innate immunity as well as by the host-derived antimicrobial LL-37. Here, I show that LptA contributes to Gc defense against several neutrophil-derived components including the α-defensin HNP-1, CAP37/azurocidin and the serine protease cathepsin G. I also demonstrate that LptA-expressing Gc manipulates neutrophil phagosome maturation by disrupting primary granule fusion with the phagosomal membrane. Interestingly, the neutrophil-derived antimicrobials that LptA defends Gc against are all stored in primary granules. I also demonstrate that MtrCDE protects Gc against neutrophil killing. Previously, MtrCDE was known to be involved in the export of a variety of structurally diverse toxic substrates, such as antibiotics, fatty acids, nonionic detergents, bile salts and host-derived LL-37, out of the bacterial cell. I observe that MtrCDE improves survival of Gc exposed to a degranulated supernatant containing a mixture of neutrophil-derived antimicrobials. In addition, I show that both MtrCDE and LptA protect Gc from the antimicrobial activity elicited by NETs. Lastly, I identify a β-defensin with antigonococcal activity, though Gc have been reported to be highly resistant to defensins. Defensins are abundant in the reproductive tract, the predominant environmental niche for Gc, and represent an alternative therapeutic option to combat Gc. Taken together, these results expand our understanding of the role of two important Gc virulence factors during interactions with human neutrophils. We are slowly uncovering the inner-workings of the extensive defensive arsenal that Gc has at its disposal to resist killing by neutrophils and epithelium-derived antimicrobials during infection.
Language
English
Published
University of Virginia, Department of Microbiology, PHD (Doctor of Philosophy), 2016
Published Date
2016-07-29
Degree
PHD (Doctor of Philosophy)
Collection
Libra ETD Repository
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