Mechanisms of Iron Acquisition Employed by NeisseriaGonorrhoeae for Survival within Cervical Epithelial Cells

Abstract

MECHANISMS OF IRON ACQUISITION EMPLOYED BY NEISSERIA GONORRHOEAE FOR SURVIVAL WITHIN CERVICAL EPITHELIAL CELLS By Tracey Ann Hagen A Dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy at Virginia Commonwealth University Virginia Commonwealth University, 2006 Major Director: Cynthia Nau Cornelissen, Ph.D. Associate Professor, Department of Microbiology and Immunology Neisseria gonowhoeae has evolved a repertoire of high-affinity iron acquisition systems to facilitate essential iron uptake in the human host. Acquisition of iron requires both the energy-harnessing cytoplasmic membrane protein, TonB, as well as specific outer membrane TonB-dependent transporters (TdTs). The overall goal of this study was to investigate the extraand intracellular iron acquisition mechanisms of N. gonorrhoeae and determine the role of the TonB and TdTs in this process. The ability of gonococci to acquire potential exogenous iron sources was determined by plate bioassay. Gonococcal growth was promoted by various catecholate and hydromate siderophores; however, growth was not dependent upon TonB expression. As all previously characterized siderophore-iron uptake is dependent upon this protein, a potential TonB-bypass mechanism is suggested. The role of the Ton system and TdTs in gonococcal survival within human cervical epithelial cells was also determined for two gonococcal strains, FA1090 and MS 1 1. We demonstrate that intracellular survival of both strains was dependent upon host cell iron acquisition, yet the expression of the Ton system was only critical to the survival of FA1090. One characterized difference between these two strains is possession of the gonococcal genetic island (GGI) which is present in approximately 80% of gonococcal strains. This study demonstrates that the GGI provides a mechanism to bypass intracellular TonB-dependent iron acquisition. In the strain lacking the genetic island, none of the characterized TdTs provided a benefit to the gonococcus when grown intracellularly. However, expression of one uncharacterized TdT, TdfF, was necessary for successful intracellular survival. To our knowledge, this is the first demonstration of a specific requirement for a single iron transporter in the survival of a bacterial pathogen within host epithelial cells. In the GGI-containing strain, TonB function was not critical to survival within cervical epithelial calls. The presence of the GGI was associated with the ability to bypass TonB-dependant uptake. Specifically, this bypass mechanism was mediated by components of the T4S machinery encoded by the GGI, and replication was directly xvi related to iron acquisition. To our knowledge, this study provides the first direct link between bacterial iron acquisition and a type IV secretion system.