Evaluation of different DNA extraction methods and loop-mediated isothermal amplification primers for the detection of Mycobacterium ulcerans in clinical specimens

BMC Infectious Diseases

Joseph M. Ndung’u¹  Isra Cruz²  Sandra A. King³  Charles A. Narh³  Evans Ahotor³  Anthony Ablordey³  Dziedzom K. de Souza⁴ 

[1] Foundation for Innovative New Diagnostics, Geneva, Switzerland;Foundation for Innovative New Diagnostics, Geneva, Switzerland;National School of Public Health, Instituto de Salud Carlos III, Madrid, Spain;Noguchi Memorial Institute for Medical Research, College of Health Sciences, University of Ghana, Legon, Accra, Ghana;Noguchi Memorial Institute for Medical Research, College of Health Sciences, University of Ghana, Legon, Accra, Ghana;Foundation for Innovative New Diagnostics, Geneva, Switzerland;

关键词: Loop mediated isothermal amplification;    DNA extraction;    Sensitivity;    Specificity;   

DOI  :  10.1186/s12879-021-06308-z

来源: Springer

【 摘 要 】

BackgroundEarly diagnosis and treatment of Buruli ulcer is critical in order to avoid the debilitating effects of the disease. In this regard, the development of new diagnostic and point of care tools is encouraged. The loop-mediated isothermal amplification for the detection of Mycobacterium ulcerans represents one of the new tools with a good potential of being developed into a point of care test. There is however the need to standardize the assays, reduce sample preparation times, improve the detection/visualization system and optimize them for high-throughput screening, adaptable to low resourced laboratories.MethodsIn this study, we assessed two DNA extraction protocols (modified Boom and EasyNAT methods), three previously published LAMP primer sets (BURULI, MU 2404 and BU-LAMP), and compared the sensitivity and specificity of LAMP assays on three DNA amplification platforms.ResultsOur results show that Buruli ulcer diagnosis using primers targeting IS2404 for the LAMP method is sensitive (73.75–91.49%), depending on the DNA extraction method used. Even though the modified Boom DNA extraction method provided the best results, its instrumentation requirement prevent it from being field applicable. The EasyNAT method on the other hand is simpler and may represent the best method for DNA extraction in less resourced settings.ConclusionsFor further work on the development and use of LAMP tests for Buruli diagnosis, it is recommended that the BURULI sets of primers be used, as these yielded the best results in terms of sensitivity (87.50–91.49%) and specificity (89.23–100%), depending on the DNA extraction methods used.

【 授权许可】

CC BY