• Login
    View Item 
    •   UMB Digital Archive
    • School, Graduate
    • Theses and Dissertations All Schools
    • View Item
    •   UMB Digital Archive
    • School, Graduate
    • Theses and Dissertations All Schools
    • View Item
    JavaScript is disabled for your browser. Some features of this site may not work without it.

    Browse

    All of UMB Digital ArchiveCommunitiesPublication DateAuthorsTitlesSubjectsThis CollectionPublication DateAuthorsTitlesSubjects

    My Account

    LoginRegister

    Statistics

    Display statistics

    Detection of species-specific DNA sequences of Borrelia burgdorferi in infected humans, animal reservoirs, and ixodid tick vectors

    • CSV
    • RefMan
    • EndNote
    • BibTex
    • RefWorks
    Find Full text
    Author
    Malloy, Diane Catherine
    Advisor
    Nauman, Robert K.
    Date
    1992
    Type
    dissertation
    
    Metadata
    Show full item record
    Abstract
    Segments of the ospA gene that encode hydrophobic regions of the outer membrane protein, OspA, of Borrelia burgdorferi strain B31 were synthesized for use as oligonucleotide primers in the polymerase chain reaction (PCR). These oligonucleotide primers flank a 309-base-pair segment within the ospA gene. Optimal amplification conditions were achieved in a reaction mixture containing 0.2 uM of each oligonucleotide primer and 2 mMMgCl{dollar}\sb2.{dollar} Dimethyl sulfoxide at a concentration of 10% or higher was found to inhibit amplification and gelatin had no effect at concentrations below 100 ug/ml, and slight inhibition was seen at concentrations higher than 100 ug/ml. After 30 cycles of amplification under optimal conditions, the target fragment could be detected by agarose gel electrophoresis or dot hybridization with a {dollar}\sp{lcub}32{rcub}{dollar}P- or digoxigenin-labeled probe. This segment was amplified in all strains of B. burgdorferi, but it was not detected in other bacterial species. The sensitivity of PCR for the detection of B. burgdorferi in clinical samples was evaluated by seeding blood and urine specimens with B. burgdorferi and subjecting them to amplification. Ten organisms per ml of blood or urine could be detected using PCR with dot hybridization detection. In a blinded study of Lyme disease patients, the OspA PCR was positive in 31% of patients who were early in disease and who had not received oral antibiotic therapy. No patient who had received antibiotics was positive in the PCR. Blood and urine specimens were obtained from canines with clinical and serologic evidence of Lyme disease and subjected to PCR analysis. Of 17 clinical specimens from 15 canines, one blood specimen showed reactivity in the PCR. Two of 32 cerebrospinal fluid specimens from suspected neuroborreliosis patients showed reactivity in the PCR. B. burgdorferi could be detected optimally in tissue only after DNA extraction. Nine of ten mice from a highly endemic Lyme disease area in Wisconsin showed reactivity in the PCR when DNA extracted from heart, kidney, or bladder was used as the target. Two of five punch biopsy tissue samples from skin lesions from suspected Lyme disease patients showed reactivity in the PCR. Of all tissues studied, one yielded a positive spirochete stain and all were negative by immunoperoxidase staining with a polyclonal antibody to B. burgdorferi. The conclusion of this study is that PCR can detect and identify B. burgdorferi in clinical samples from Lyme disease with greater sensitivity than any other currently available method and that this tool can be used to detect the spirochete in tick and animal reservoirs.
    Description
    University of Maryland, Baltimore. Microbiology. Ph.D. 1992
    Keyword
    Biology, Molecular
    Biology, Microbiology
    Health Sciences, Public Health
    species-specific DNA sequences
    Borrelia burgdorferi--genetics
    Disease Reservoirs
    Lyme Disease
    Polymerase Chain Reaction
    Identifier to cite or link to this item
    http://hdl.handle.net/10713/2544
    Collections
    Theses and Dissertations All Schools
    Theses and Dissertations School of Dentistry

    entitlement

     
    DSpace software (copyright © 2002 - 2023)  DuraSpace
    Quick Guide | Policies | Contact Us | UMB Health Sciences & Human Services Library
    Open Repository is a service operated by 
    Atmire NV
     

    Export search results

    The export option will allow you to export the current search results of the entered query to a file. Different formats are available for download. To export the items, click on the button corresponding with the preferred download format.

    By default, clicking on the export buttons will result in a download of the allowed maximum amount of items.

    To select a subset of the search results, click "Selective Export" button and make a selection of the items you want to export. The amount of items that can be exported at once is similarly restricted as the full export.

    After making a selection, click one of the export format buttons. The amount of items that will be exported is indicated in the bubble next to export format.