Rapid Multiplex Loop-Mediated Isothermal Amplification (m-LAMP) Assay for Differential Diagnosis of Leprosy and Post–Kala-Azar Dermal Leishmaniasis

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  • 1 Molecular Parasitology Laboratory, ICMR-National Institute of Pathology, Safdarjung Hospital Campus, New Delhi, India;
  • 2 Department of Dermatology, Safdarjung Hospital, New Delhi, India

Leprosy and post–kala-azar dermal leishmaniasis (PKDL) are co-endemic neglected tropical diseases often misdiagnosed because of close resemblance in their clinical manifestations. The test that aids in differential diagnosis of leprosy and PKDL would be useful in endemic areas. Here, we report development of a multiplex loop-mediated isothermal amplification (m-LAMP) assay for differential detection of Mycobacterium leprae and Leishmania donovani using a real-time fluorometer. The m-LAMP assay was rapid with a mean amplification time of 15 minutes, and analytical sensitivity of 1 fg for L. donovani and 100 fg for M. leprae. The distinct mean Tm values for M. leprae and L. donovani allowed differentiation of the two organisms in the m-LAMP assay. Diagnostic sensitivity of the assay was evaluated by using confirmed cases of leprosy (n = 40) and PKDL (n = 40) (tissue and slit aspirate samples). All the leprosy and PKDL samples used in this study were positive by organism-specific QPCR and loop-mediated isothermal amplification assays. The diagnostic sensitivity of the m-LAMP assay was 100% (95% CI: 91.2–100.0%) for detecting PKDL and 95% for leprosy (95% CI: 83.1–99.4%). Our m-LAMP assay was successfully used to detect both M. leprae and L. donovani in a patient coinfected with leprosy and macular PKDL. The m-LAMP assay is rapid, accurate, and applicable for differential diagnosis of leprosy versus PKDL, especially in endemic areas.

Author Notes

Address correspondence to Ruchi Singh or Poonam Salotra, ICMR-National Institute of Pathology, Safdarjung Hospital Campus, New Delhi 110029, India. E-mails: ruchisp@gmail.com or poonamsalotra@hotmail.com

Financial suppot: The research was funded by the Indian Council of Medical Research (ICMR), New Delhi, India (intramural project). S. J. received grant from the Science and Engineering Research Board (SERB), Department of Science and Technology, Government of India, in the form of SERB-National Post-Doctoral Fellowship (Grant no. PDF/2016/003200).

Authors’ addresses: Shweta Joshi, Keerti K. Dixit, Vanila Sharma, Ruchi Singh, and Poonam Salotra, National Institute of Pathology, Molecular Parasitology Laboratory, New Delhi, India, E-mails: shweta26jan83@gmail.com, keerti.dixit@gmail.com, vanilasharma81@gmail.com, ruchisp@gmail.com, and poonamsalotra@hotmail.com. V. Ramesh, Dermatology and Venereology, Vardhman Mahavir Medical College and Safdarjung Hospital, New Delhi, India, E-mail: weramesh@gmail.com.