development of a loop-mediated isothermal amplification (lamp) assay for detection of relapsing fever borreliae

Background: this study aimed to develop a loop-mediated isothermal amplification (lamp) assay for the rapid detection of tick-borne relapsing fever in resource-limited areas. methods: a set of six primers were designed based on the conserved regions of the glycerophosphodiester phos-phodiesterase (glpq) gene of borrelia species. for sensitivity assay, serial dilutions of a recombinant plasmid contain-ing a 219bp sequence of the glpq were prepared and used as the template dna. the lamp reactions containing the six primers and the reagents required for amplification were incubated at 60–65 °c for 60min in a loopamp real-time tur-bidimeter. for the specificity test, dna from 14 other bacteria were included in the assays, and double-distilled water was used as the negative control. also, dna from dried blood spots (dbss) of spirochetemic mice, and blood samples from relapsing fever-suspected patients were examined by the lamp along a borrelia-specific nested pcr that targets the rrs-rrl-igs region. results: the lamp detected as low as 90glpq copies in reactions. the primers reacted with dna from dbs of spi-rochetemic mice showing spirochete concentrations of ≤ one per a 1000x microscopic field. in clinical samples, the lamp assay showed a higher sensitivity compared to nested-pcr. the lamp specificity was 100%, as the primers did not react with other bacteria dna. conclusion: the high sensitivity and specificity of the test, along with the simplicity of the dna extraction procedure, make the lamp a reliable and adaptable tool for the diagnosis of tick-borne relapsing fever in rural endemic areas.