Proof of specific DNA using nested-PCR in patiens with different clinical form of Lyme borreliosis
Abstract number: 1733_956
Picha D., Moravcova L., Holeckova D., Sedivy K., Zdarsky E., Maresova V.
Objectives: This study was aimed to the direct proof of borrelial DNA with polymerase chain reaction (PCR) in patients with lyme borreliosis (LB) and to estimate the specifity of individual primers in the diagnostic of its clinical forms.
Methods: Into the prospective study 71 patients with LB were enrolled (35 patients with neuroborreliosis, 14 with joint involvement and 22 with skin form). The criteria for inclusion were recent clinical symptoms of LB and the presence of specific antibodies in the blood, CSF, joint fluid (confirmed by Western blotting) or proof of DNA in body fluids. Before the treatment the presence of borrelial DNA was tested in plasma, CSF, joint fluid and urine, after the treatment in plasma, urine and joint fluid. A system with five newly derived primers has been used, with three primers specific for chromozomal genes coding 16SrDNA, flagellin and p66 and two for plasmid genes which are coding the outer membrane's protein OspA, OspC. The presence of borrelial DNA was tested by nested-PCR in parallel with all the primers.
Results: From the total number of 71 examined patients the borrelial DNA was proved in 42 patients (59.2%) before the treatment; 23 positive patients (65.7%) were found in group of neuroborreliosis, 10 positives (45.5%) showed signs of skin involvement and 9 patients (64.3%) were positive in arthritis. After the treatment the borrelial DNA was proved in half of the patients with neuroborreliosis (13×) and in the skin form patients (4×). In patients with the joint form the proof was the same as before the treatment. From the 105 positive amplifications the most frequently positive primer was 16SrDNA 54× (51.4%). Less sensitive were primers OspA (20×;19%), OspC (14×;13.3%) and flagellin (14×;13.3%). In 14 patients (31.1%) with skin and joint form of LB only borrelial DNA without specific antibodies was proved.
Conclusion: This study has shown that in the period before treatment the sensitivity of PCR was 59.2% using 5 primers and including all the clinical forms of borreliosis. The most positive result has been found by using the primer 16SrDNA (51.4%). The same reactivity was being shown in plasmid sequences coding genes OspA and OspC and chromosomal sequence for flagellin. Examination of the borrelial DNA in urine displayed the same sensitivity as in CSF.
The study was supported by grants of Ministry of Health and Education (IGA-8293, MSM 0021620812)
|Session name:||European Society of Clinical Microbiology and Infectious Diseases|
|Location:||ICC, Munich, Germany|
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