Detection of with IS loop-mediated isothermal amplification and a fluorescent reporter probe.

An exquisitely sensitive quantitative PCR (qPCR) assay targeting the high-copy number insertion sequence, IS, is the gold standard diagnostic test for , the agent of the neglected tropical skin disease Buruli ulcer. Here, we designed and tested an alternative diagnostic test, a fluorescent probe-based, loop-mediated isothermal amplification (P-LAMP) assay that also targets IS. Benchmarked against IS qPCR, P-LAMP was equally specific and nearly as sensitive (analytical sensitivity of four vs two genome copies). Clinical and environmental specimen validation against IS qPCR showed P-LAMP had 100% sensitivity and specificity. P-LAMP was twice as fast as qPCR with an average time-to-positive at the limit-of-detection of 19 minutes. P-LAMP targeting IS is a versatile assay that addresses the performance issues of previously described IS LAMP formats. This study tackles a key research priority for Buruli ulcer and represents another avenue for the development of rapid and accessible molecular diagnostics for this neglected tropical disease.IMPORTANCEBuruli ulcer is a neglected tropical disease caused by infection with . Correct diagnosis is essential before appropriate treatment for Buruli ulcer can be started. Development of a portable, easy-to-use diagnostic test for has been identified by the World Health Organization as a research priority. Buruli ulcer most commonly occurs in remote, rural areas; therefore, an ideal test is one that can be used at (or near) the point of care (community health centres) without the need for specialized laboratories. Here, we describe a molecular test using loop-mediated isothermal amplification (LAMP) to detect DNA specific to and show that this new test has equivalent performance to the gold standard PCR test currently used worldwide. Our new test is rapid (30 minutes to run), simple to perform, and could be further developed into a robust, portable format to provide accessible and affordable diagnostics anywhere.