Previous studies have demonstrated that the increased incidence of invasive disease caused by serogroup C Neisseria meningitidis in the United States during the 1990s was attributed primarily to strains belonging to the ST11 clonal complex. Subcapsular genotyping of a subset of isolates from Maryland identified distinct early and late clones defined by antigenic shift at the FetA outer membrane protein. Representational difference analysis (RDA) was used to identify additional genetic differences that may have contributed to the emergence of the late clone. A collection of serogroup C isolates representative of the early and late clone was subjected to pulsed field gel electrophoresis (PFGE) to determine genetic relatedness among the isolates and to identify a candidate tester/driver pair for RDA. RsaI-digested tester genomic DNA (late clone) was ligated to specific adaptors followed by two rounds of subtractive hybridization with RsaI-digested driver genomic DNA (early clone). PCR amplification of subtracted tester DNA with adaptor specific primers generated at least three late clone-specific bands that were absent from the early clone. These products were cloned and sequenced and confirmed by Southern blotting with tester and driver digoxigenin-labeled genomic DNA probes to be tester specific. A BLAST search of late clone-specific sequences identified homology to either IS1301 or pJS-B plasmid N. meningitidis sequences. PCR with primers specific to either IS1301 or pJS-B plasmid sequences amplified these elements from late clone isolates but not from early clone isolates. Thus, RDA successfully identified two unique genetic elements present in an emergent N. meningitidis serogroup C ST-11 clone that had undergone antigenic shift at FetA. Further investigation is required to determine the potential role of these elements in clonal emergence and N. meningitidis pathogenesis. The public health significance of this project stems from increased incidence of meningococcal disease being a major concern: morbidity and mortality increase, outbreaks produce panic and disruption in communities, public health agencies must respond for control and prevention, and mass immunization and antibiotic prophylaxis are often required.
| Identifer | oai:union.ndltd.org:PITT/oai:PITTETD:etd-04122006-165816 |
| Date | 07 June 2006 |
| Creators | Kostelnik, Leah M. |
| Contributors | Jeremy Martinson, PhD, Lee H. Harrison, MD, Yuan Chang, MD |
| Publisher | University of Pittsburgh |
| Source Sets | University of Pittsburgh |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://etd.library.pitt.edu/ETD/available/etd-04122006-165816/ |
| Rights | unrestricted, I hereby certify that, if appropriate, I have obtained and attached hereto a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to University of Pittsburgh or its agents the non-exclusive license to archive and make accessible, under the conditions specified below, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
Page generated in 0.0111 seconds