Detection of borrelia microti in blood samples spotted on dna banking card by using loop-mediated isothermal amplification (lamp)

Faezeh Houman sadr,1,* Mohammad soleimani,2 Gholamreza javadi,3 Saied reza naddaf,4

1. Department of Biology Cellular and Molecular, Science and Research branch, Islamic Azad University, Tehran, Iran
2. Department of micribiology , Faculty of Medicine, AJA University of Medical Sciences, Tehran, Iran
3. Department of Biology Cellular and Molecular, Science and Research branch, Islamic Azad University, Tehran, Iran
4. Department of Parasitology, Pasteur Institute of Iran, Tehran, Iran

Abstract

Introduction

Tick-borne relapsing fever (tbrf) is an endemic disease in iran, with more 100 annual cases. the glycerophosphodiester phosphodiesterase (glpq) is an enzyme in spirochetes causing tbrf borreliae such as borrelia microti. we successfully established a detection method which exhibited a markedly higher sensitivity than previously developed detection methods for borrelia microti by combining dna banking card (dbc), and loop-mediated isothermal amplification (lamp).

Methods

The specific primers were designed for amplification of a unique fragment of glpq gene. thirty blood samples of spirochete infected mice and thirty blood samples of non-infected mice were collected and kept on dbc card. the dbc card pieces were used to extract dna for glpq-lamp reactions and the result was compared with microscopic inspection (gold standard).

Results

In this study, of 30 positive samples analyzed by the glpq-lamp, 30 (100%) showed the expected amplification products in turbidity inspection with naked eye. none of 30 negative samples (100%) showed amplification products. the results showed that the sensitivity and specificity of glpq-lamp test were found to be 100%.

Conclusion

We have demonstrated that the glpq-lamp assay has high sensitivity and specificity for detecting tbrf borreliae. in contrast to pcr, lamp has duel advantages of being simple to carry out and cost-effective. in addition, combining the glpq-lamp and dna dbc could be useful to detect b.microtti infection in endemic regions for tbrf disease.

Keywords

Borrelia microtti - dbc card - lamp - glpq