ABSTRACT
Background Increasing urbanisation has led to the occurrence of cutaneous leishmaniasis (CL) in new areas, which was otherwise localised to endemic areas. Healthcare workers should be made aware of this entity to ensure clinical suspicion of CL and investigations needed to confirm CL. The article describes patients seen at a tertiary hospital in Delhi. Aims To establish the utility of the CL Detect Rapid test as a diagnostic tool and the efficacy of Liposomal Amphotericin B (LAmB) for the complete cure of CL patients. Methods Data of patients of CL (n = 16) was retrospectively analysed concerning diagnosis and treatment. Diagnosis rested on histopathology, real-time PCR, and CL Detect Rapid Test. Speciation of the parasite was based on the Internal transcribed spacer-I gene. Patients were treated with LAmB (i.v., 5 mg/kg up to three doses, five days apart). Results A positivity of 81.3% (95%CI, 54.4-96) was observed for CL Detect Rapid test in comparison with 100% (95%CI, 79.4-100.0) for real-time PCR and 43.8% (95%CI, 19.8-70.1) for microscopy/histopathological examination. L. tropica was the infective species in all cases. All the patients treated with LAmB responded to treatment, and 9/10 patients demonstrated complete regression of lesions, while one was lost to follow-up. Limitations It is a retrospective study, and the data includes only confirmed cases of CL at a single centre. Conclusion This study highlights the utility of CL Detect as a promising diagnostic tool and the efficacy of LAmB for the complete cure of CL.
Subject(s)
Amphotericin B , Antiprotozoal Agents , Leishmaniasis, Cutaneous , Humans , Retrospective Studies , Antiprotozoal Agents/therapeutic use , Tertiary Care Centers , Leishmaniasis, Cutaneous/diagnosis , Leishmaniasis, Cutaneous/drug therapy , Leishmaniasis, Cutaneous/epidemiology , India/epidemiologyABSTRACT
BACKGROUND: Sensitive and definitive diagnostic tests are required for timely treatment of leprosy and to control its transmission. AIM: In the present study, we report the development of loop-mediated isothermal amplification assay using six primers targeting the RLEP gene sequence uniquely present in Mycobacterium leprae. METHODS: Tissue punch samples (n = 50) and slit aspirates (n = 50) from confirmed cases of leprosy (M. leprae positive by quantitative polymerase chain reaction), reporting at the Department of Dermatology, Safdarjung Hospital, New Delhi, were analyzed using newly developed closed tube loop-mediated isothermal amplification assay. The sensitivity and specificity; positive predictive value, negative predictive value and accuracy were calculated using MedCalc statistical software. RESULTS: The loop-mediated isothermal amplification assay specifically amplified M. leprae genomic DNA with an analytical sensitivity of 100 fg. About 47 Out of the 50 quantitative polymerase chain reactions confirmed M. leprae positive tissue samples, 47 were positive by loop-mediated isothermal amplification assay (sensitivity 94%; 95% confidence interval 83.5%-98.8%) while only 31/50 were positive by histopathology (sensitivity 62%; 95% confidence interval 47.2%-75.4%) . Using slit aspirate samples of these 50 patients, 42 were positive by both quantitative polymerase chain reaction and loop-mediated isothermal amplification assay (sensitivity 84%; 95% confidence interval 70.9%-92.8%) while only 23/50 (sensitivity 46%; 95% confidence interval 31.8%-60.7%) were positive by microscopy. LIMITATIONS: In the present study, the leprosy patient cohort was not uniform, as it comprised a lower number of paucibacillary cases (22%) compared to multibacillary (78%) cases. CONCLUSION: Loop-mediated isothermal amplification assay established here provides a rapid and accurate diagnostic test for leprosy in terms of sensitivity and specificity. The assay is simple to perform in comparison with other molecular techniques (polymerase chain reaction/quantitative polymerase chain reaction) and has potential for field applicability.
Subject(s)
Leprosy/diagnosis , Molecular Diagnostic Techniques/methods , Mycobacterium leprae/genetics , Mycobacterium leprae/isolation & purification , Nucleic Acid Amplification Techniques/methods , Adolescent , Adult , Aged , DNA, Bacterial/genetics , Female , Humans , Male , Middle Aged , Real-Time Polymerase Chain Reaction , Sensitivity and Specificity , Young AdultABSTRACT
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.