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Improved method of detection and molecular typing of Borrelia burgdorferi sensu lato in clinical samples by polymerase chain reaction without DNA purification

Folia microbiologica. 2005 ; 50 (1) : 31-9.

Folia Microbiol (Praha)
ISSN 0015-5632
Source

Language English Country United States Media print

Document type Journal Article, Research Support, Non-U.S. Gov't

Persistent link https://www.medvik.cz/link/pmid15954531

PubMed 15954531
DOI 10.1007/bf02931291
Knihovny.cz E-resources

MeSH
Antigens, Surface genetics MeSH
Bacterial Vaccines MeSH
Benzothiazoles MeSH
Borrelia burgdorferi Group classification genetics immunology isolation & purification MeSH
Quinolines MeSH
Diamines MeSH
DNA, Bacterial analysis isolation & purification MeSH
Electrophoresis, Agar Gel MeSH
Enzyme-Linked Immunosorbent Assay MeSH
Genotype MeSH
Immunoglobulin G blood MeSH
Immunoglobulin M blood MeSH
Humans MeSH
Lipoproteins genetics MeSH
Lyme Disease diagnosis MeSH
Cerebrospinal Fluid microbiology MeSH
Organic Chemicals metabolism MeSH
Polymerase Chain Reaction methods MeSH
Bacterial Outer Membrane Proteins genetics MeSH
Antibodies, Bacterial blood MeSH
Sensitivity and Specificity MeSH
Serum microbiology MeSH
Bacterial Typing Techniques MeSH
Blotting, Western MeSH
Check Tag
Humans MeSH
Publication type
Journal Article MeSH
Research Support, Non-U.S. Gov't MeSH
Names of Substances
Antigens, Surface MeSH
Bacterial Vaccines MeSH
Benzothiazoles MeSH
Quinolines MeSH
Diamines MeSH
DNA, Bacterial MeSH
Immunoglobulin G MeSH
Immunoglobulin M MeSH
Lipoproteins MeSH
Organic Chemicals MeSH
OspA protein MeSH Browser
Bacterial Outer Membrane Proteins MeSH
Antibodies, Bacterial MeSH
SYBR Green I MeSH Browser

A simple assay by polymerase chain reaction was used for the of detection of Borrelia burgdorferi, causative agent of Lyme borreliosis (LB). It involves no DNA purification and is based on the amplification of a specific region of ospA gene of B. burgdorferi, followed by direct detection of the PCR product with SYBR Green I by agarose gel electrophoresis. The method was used to analyze samples from patients with LB diagnosis, with presumable infection with the LB spirochete, those with unclear clinical symptoms and after the course of an antibiotic treatment. Spirochetal DNA was detected by PCR even in contaminated samples in which B. burgdorferi was overgrown by fungi and other bacteria. Spirochetal DNA was detected and borrelia species was identified in cerebrospinal fluid of two patients hospitalized with the diagnosis "fever of unknown origin". Western blot and ELISA were negative in both cases. Total analysis of 94 samples from the hospital in Ceské Budejovice (South Bohemia, Czechia) showed infection with B. burgdorferi sensu stricto in 11% and B. garinii in 15% of cases. The highest prevalence was found for B. afzelii (43%). Co-infection was confirmed in 24 % of the analyzed symplex; 7% of samples that were B. burgdorferi sensu lato positive gave no results in DNA amplification with B. burgdorferi sensu stricto-, B. garinii- and B. afzelii-specific primers. The proposed reliable, rapid, unexpensive and specific technique could form the basis of laboratory tests for routine detection and identification of Lyme-disease spirochete in different samples.

Faculty of Biological Sciences University of South Bohemia 370 05 Ceské Budejovice Czechia

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