Detection and quantification of Lyme spirochetes using sensitive and specific molecular beacon probes
<p>Abstract</p> <p>Background</p> <p>Lyme disease, caused by <it>Borrelia burgdorferi</it>, affects a large number of people in both the USA and Europe. The mouse is a natural host for this spirochete and is widely used as a model system to study Lyme pathog...
| Main Authors: | , , |
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| Format: | Article |
| Language: | English |
| Published: |
BMC
2009-02-01
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| Series: | BMC Microbiology |
| Online Access: | http://www.biomedcentral.com/1471-2180/9/43 |
| _version_ | 1818491457081704448 |
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| author | Parveen Nikhat Marras Salvatore AE Saidac Diana S |
| author_facet | Parveen Nikhat Marras Salvatore AE Saidac Diana S |
| author_sort | Parveen Nikhat |
| collection | DOAJ |
| description | <p>Abstract</p> <p>Background</p> <p>Lyme disease, caused by <it>Borrelia burgdorferi</it>, affects a large number of people in both the USA and Europe. The mouse is a natural host for this spirochete and is widely used as a model system to study Lyme pathogenesis mechanisms. Since disease manifestations often depend upon the spirochete burden in a particular tissue, it is critical to accurately measure the bacterial number in infected tissues. The current methods either lack sensitivity and specificity (SYBR Green), or require independent analysis of samples in parallel to quantitate host and bacterial DNA (TaqMan). We have developed a novel molecular beacon-based convenient multiplex real-time quantitative PCR assay to identify and detect small numbers of <it>B. burgdorferi </it>in infected mouse tissues.</p> <p>Results</p> <p>We show here that molecular beacons are effective, sensitive and specific probes for detecting and estimating wide-ranging numbers of <it>B. burgdorferi </it>in the presence of mouse DNA. In our assays, the spirochete <it>recA </it>and the mouse <it>nidogen </it>gene amplicons were detected simultaneously using molecular beacons labeled with different fluorophores. We further validated the application of these probes by quantifying the wild-type strain and <it>bgp</it>-defective mutant of <it>B. burgdorferi</it>. The <it>bgp</it>-defective mutant shows a ten-fold reduction in the level of spirochetes present in various tissues.</p> <p>Conclusion</p> <p>The high sensitivity and specificity of molecular beacons makes them superior probes for the detection of small numbers of <it>B. burgdorferi</it>. Furthermore, the use of molecular beacons can be expanded for the simultaneous detection and quantification of multiple pathogens in the infected hosts, including humans, and in the arthropod vectors.</p> |
| first_indexed | 2024-12-10T17:31:05Z |
| format | Article |
| id | doaj.art-ae445b7c206945c2a72b468b5c2760b7 |
| institution | Directory Open Access Journal |
| issn | 1471-2180 |
| language | English |
| last_indexed | 2024-12-10T17:31:05Z |
| publishDate | 2009-02-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Microbiology |
| spelling | doaj.art-ae445b7c206945c2a72b468b5c2760b72022-12-22T01:39:42ZengBMCBMC Microbiology1471-21802009-02-01914310.1186/1471-2180-9-43Detection and quantification of Lyme spirochetes using sensitive and specific molecular beacon probesParveen NikhatMarras Salvatore AESaidac Diana S<p>Abstract</p> <p>Background</p> <p>Lyme disease, caused by <it>Borrelia burgdorferi</it>, affects a large number of people in both the USA and Europe. The mouse is a natural host for this spirochete and is widely used as a model system to study Lyme pathogenesis mechanisms. Since disease manifestations often depend upon the spirochete burden in a particular tissue, it is critical to accurately measure the bacterial number in infected tissues. The current methods either lack sensitivity and specificity (SYBR Green), or require independent analysis of samples in parallel to quantitate host and bacterial DNA (TaqMan). We have developed a novel molecular beacon-based convenient multiplex real-time quantitative PCR assay to identify and detect small numbers of <it>B. burgdorferi </it>in infected mouse tissues.</p> <p>Results</p> <p>We show here that molecular beacons are effective, sensitive and specific probes for detecting and estimating wide-ranging numbers of <it>B. burgdorferi </it>in the presence of mouse DNA. In our assays, the spirochete <it>recA </it>and the mouse <it>nidogen </it>gene amplicons were detected simultaneously using molecular beacons labeled with different fluorophores. We further validated the application of these probes by quantifying the wild-type strain and <it>bgp</it>-defective mutant of <it>B. burgdorferi</it>. The <it>bgp</it>-defective mutant shows a ten-fold reduction in the level of spirochetes present in various tissues.</p> <p>Conclusion</p> <p>The high sensitivity and specificity of molecular beacons makes them superior probes for the detection of small numbers of <it>B. burgdorferi</it>. Furthermore, the use of molecular beacons can be expanded for the simultaneous detection and quantification of multiple pathogens in the infected hosts, including humans, and in the arthropod vectors.</p>http://www.biomedcentral.com/1471-2180/9/43 |
| spellingShingle | Parveen Nikhat Marras Salvatore AE Saidac Diana S Detection and quantification of Lyme spirochetes using sensitive and specific molecular beacon probes BMC Microbiology |
| title | Detection and quantification of Lyme spirochetes using sensitive and specific molecular beacon probes |
| title_full | Detection and quantification of Lyme spirochetes using sensitive and specific molecular beacon probes |
| title_fullStr | Detection and quantification of Lyme spirochetes using sensitive and specific molecular beacon probes |
| title_full_unstemmed | Detection and quantification of Lyme spirochetes using sensitive and specific molecular beacon probes |
| title_short | Detection and quantification of Lyme spirochetes using sensitive and specific molecular beacon probes |
| title_sort | detection and quantification of lyme spirochetes using sensitive and specific molecular beacon probes |
| url | http://www.biomedcentral.com/1471-2180/9/43 |
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