Expansion, Purification, and Quantitative Analysis of an APRc-Deficient Rickettsia parkeri Strain
Abstract/Description/Artist Statement
Rickettsia is a genus of Gram-negative, obligate intracellular bacteria comprising many highly pathogenic species known to cause severe illnesses such as epidemic typhus and Rocky Mountain spotted fever. Recent research indicates that the range and incidence of tick-borne disease are increasing due to global temperature change. Being primarily tick-borne, Rickettsia spp. are considered significant emerging pathogens, requiring investigation into their pathogenicity in the interest of public health. In previous work, we discovered a novel, highly conserved retropepsin-like protease (APRc) present in all Rickettsia species. Our results indicate APRc acts as an immune-evasion factor by binding antibodies and cleaving several complement components. To further investigate the multifunctional role of APRc Rickettsia pathogenicity, an R. parkeri mutant with a knockout of the APRc homolog was recently isolated. Using Vero cells as a host system, this work focuses on the expansion, isolation, and titration of this knockout strain. Density gradient purification and immunofluorescence analysis will be used to determine the yield of the purified R. parkeri mutant. The goal of this study is to isolate, expand, and quantify this mutant strain. These quantifications will establish baseline growth, purification, and titration parameters necessary for further studies aimed at elucidating APRc’s role in host immune interactions and the molecular pathogenesis of Rickettsia.
Faculty Advisor/Mentor
Isaura Simões
Faculty Advisor/Mentor Email
isimes@odu.edu
Faculty Advisor/Mentor Department
Department of Biological Science
College/School Affiliation
College of Sciences
Student Level Group
Undergraduate
Presentation Type
Poster
Expansion, Purification, and Quantitative Analysis of an APRc-Deficient Rickettsia parkeri Strain
Rickettsia is a genus of Gram-negative, obligate intracellular bacteria comprising many highly pathogenic species known to cause severe illnesses such as epidemic typhus and Rocky Mountain spotted fever. Recent research indicates that the range and incidence of tick-borne disease are increasing due to global temperature change. Being primarily tick-borne, Rickettsia spp. are considered significant emerging pathogens, requiring investigation into their pathogenicity in the interest of public health. In previous work, we discovered a novel, highly conserved retropepsin-like protease (APRc) present in all Rickettsia species. Our results indicate APRc acts as an immune-evasion factor by binding antibodies and cleaving several complement components. To further investigate the multifunctional role of APRc Rickettsia pathogenicity, an R. parkeri mutant with a knockout of the APRc homolog was recently isolated. Using Vero cells as a host system, this work focuses on the expansion, isolation, and titration of this knockout strain. Density gradient purification and immunofluorescence analysis will be used to determine the yield of the purified R. parkeri mutant. The goal of this study is to isolate, expand, and quantify this mutant strain. These quantifications will establish baseline growth, purification, and titration parameters necessary for further studies aimed at elucidating APRc’s role in host immune interactions and the molecular pathogenesis of Rickettsia.