Evaluation of a wet reagent based loop isothermal amplification test for the detection of Mycobacterium ulcerans in clinical sample

Abstract

Background: Buruli ulcer is a disease of poverty globally reported in >37 countries across the world. The disease affects people of all age group with >65% of cases usually examined in the advanced stage, a situation which has been shown to increase the cost of treatment. Enumerated among six research priority areas of the world health organization (WHO) is the development of simple and sensitive diagnostic tools which holds relevant keys to unlocking the other 5 research areas. In meeting the diagnostic gap, various assays have been developed including five Loop isothermal amplification test (LAMP) for detecting M. ulcerans. Exceptional among these LAMP test is a wet reagent base-LAMP test designed by Ablordey and co-workers which generates real-time results; a deviation from the earlier colourimetric based LAMP designs. The study aimed at evaluating the newly improved LAMP assay for the detection of Mycobacterium ulcerans in clinical samples. Methods and materials: Protocol for the wet reagent based LAMP (WRB-LAMP) test was optimized. Using deoxyribonucleic acid (DNA) samples of M. ulcerans prepared from culture and 75 archived clinical swab samples, technical and clinical characteristics of the test was determined. Results: WRB-LAMP detected as low as 31.4 ag/uL and was shown to be highly specific with 97% (95% CI: 84%–100%) clinical sensitivity and 100% (95% CI: 92%–100%) clinical specificity yielding an overall accuracy of 99% (95% CI: 95%–99%). Amplification properties such as constant incubation temperature and short test duration are an added advantage for field use. The assay gave an almost perfect cohen kappa agreement (p < 0.05) with the gold standard polymerase chain reaction method. The assay is rapid and meets the recommended 90% sensitivity requirement expected by WHO for new developed tools for the diagnosis of Buruli ulcer and also have similar performance as the PCR test. Conclusion: WRB-LAMP has the potential for consideration as a test deployable in district health facilities for diagnosis of Buruli ulcer disease.