Design, Optimization and Evaluation of a Polymerase Chain Reaction for Detection of Borrelia spp.
Journal Title: Advances in Clinical and Experimental Medicine - Year 2013, Vol 22, Issue 5
Abstract
<strong>Background</strong>. Lyme borreliosis and relapsing fever are important zoonotic diseases worldwide and the improvement of diagnostic strategies is a prioritized task considering the morbidity of these diseases in some areas. PCR based methods appear to be of utmost importance because of the high sensitivity and specificity of these assays.<br /><strong>Objectives. </strong>To obtain a molecular method based on PCR for the detection of the genus <em>Borrelia </em>infection in different specimens.<br /><strong>Results. </strong>Sets of reported primers were evaluated “in silico” and they did not fulfill the proposal parameters. On the other hand, the two new, designed sets of primers were theoretically efficient for <em>Borrelia </em>DNA amplification. PCR procedures with these primers were standardized with borrelial DNA and optimum annealing temperatures, primer concentrations and reaction cycle numbers were determined. The PCR analytical sensitivity was 10 genomes per reaction for each technique. Both PCR were highly specific to different <em>Borrelia </em>species DNA and to samples (sera, cerebrospinal liquids and hard ticks) infected artificially with a <em>Borrelia </em>strain, visualizing the amplification of the expected DNA fragment. No amplification was obtained when other microorganisms were used. 36 human clinical samples were negatives in a preliminary study.<br /><strong>Conclusions. </strong>Both sets of primers with their respective PCR protocols showed similar results, which suggest that each one can be used indistinctly in detecting <em>Borrelia </em>spp., mainly in countries where the situation of these diseases are unknown.
Authors and Affiliations
Angel Noda, Islay Rodríguez, Brian Mondeja, Carman Fernándeze
Keywords
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