Liposomal amphotericin B is more effective in polymorphic lesions of post kala-azar dermal leishmaniasis.

BACKGROUND: Post kala-azar dermal leishmaniasis (PKDL) is thought to be the reservoir of infection for visceral leishmaniasis in South Asia. The development of strategies for the diagnosis and treatment of PKDL are important for the implementation of the visceral leishmaniasis elimination program. AIMS: Liposomal amphotericin B (L-AMB) has been an overwhelming success in the treatment of visceral leishmaniasis. However, the empirical three-week regimen of L-AMB proposed for PKDL was shown to be inadequate, especially in the macular variant. This study aimed to delineate response of the different variants of PKDL to L-AMB. METHODS: Skin biopsies were collected from PKDL cases at disease presentation and upon completion of treatment with L-AMB. Parasite DNA was detected by Internal Transcribed Spacer-1 PCR (ITS-1 PCR) and quantified by amplification of parasite kDNA. CD68 + macrophages were estimated in tissue sections by immunohistochemistry. RESULTS: Treatment with L-AMB decreased the parasite load by 97% in polymorphic cases but only by 45% in macular cases. The median parasite load (89965 vs 5445 parasites/µg of genomic DNA) as well as infiltration by CD68+ cells before treatment was much greater in the polymorphic cases. LIMITATIONS: Although monitoring of the parasite load for 12 months post-treatment would have been ideal, this was not possible owing to logistical issues as well as the invasive nature of biopsy collection procedure. CONCLUSION: A dramatic decrease in the parasite burden was noted in patients with polymorphic lesions. Although patients with macular disease also had a decrease in parasite burden, this was not as marked as in the polymorphic cases. There was also a significantly greater infiltration of CD68 + macrophages in polymorphic PKDL before therapy.

Evaluation of anti-gal enzyme-linked immunosorbent assay for the diagnosis of Indian post-kala-azar dermal leishmaniasis.

BACKGROUND: Elimination of kala azar from India is challenging as there are potential reservoirs of Leishmania donovani in patients with post-kala-azar dermal leishmaniasis (PKDL). The vast repertoire of carbohydrate moieties on L. donovani is known to elicit specific and strong humoral responses in patients with kala azar. AIM: The present study was undertaken to evaluate the diagnostic performances of anti-gal antibodies using enzyme-linked immunosorbent assay for successful serological diagnosis of PKDL in Indian patients and to differentiate cases of past cured visceral leishmaniasis infections. METHODS: We developed Gal enzyme-linked immunosorbent assay to measure specific anti-gal IgG isotype in the sera of 71 Indian patients with PKDL. The diagnostic efficacy of the newly developed assay was evaluated for precision, sensitivity and accuracy. RESULTS: Gal2 enzyme-linked immunosorbent assay revealed three-fold increased anti-gal titers in 71 patients with active PKDL compared to controls. Subclass enzyme-linked immunosorbent assay analysis further revealed enhanced IgG2 and IgG3 anti-gal titers in patients with PKDL compared to control subjects. The rank order for specificity and sensitivity for IgG subclasses was IgG3>IgG2>IgG4>IgG1. The area under the curve values of 0.98 and 0.99 were obtained for IgG and IgG3 Gal2 enzyme-linked immunosorbent assays respectively. Overall sensitivity and specificity were 95.7% (95% CI: 88.1-99.1) and 98.1% (95% confidence interval: 90.1-99.9), and 98.5% (95% CI: 92.4-99.9) and 98.1% (95% CI: 90.1-99.9), respectively. Intra-assay coefficient of variation was 1.5% and inter-assay coefficient of variation was 11.7%. LIMITATIONS: The Gal2 enzyme-linked immunosorbent assay needs to be further investigated in mass surveys. CONCLUSION: Taken together, anti-gal titers detected through Gal2 enzyme-linked immunosorbent assay can serve as an effective diagnostic tool in disease elimination setting and help in better case management in endemic districts.

Glycoproteins in circulating immune complexes are biomarkers of patients with Indian PKDL: A study from endemic districts of West Bengal, India.

BACKGROUND: Post Kala Azar Dermal Leishmaniasis (PKDL) occurs as dermal consequence of previous Visceral Leishmaniasis (VL) infection and serves as an important reservoir for transmission of VL. Diagnosis of PKDL is often challenging for its symptomatic resemblance to other co-endemic diseases like Leprosy or Vitiligo. Parasitological examination by slit-skin smear and culture are the standard methods but lack high sensitivity. Thus, for efficient control of VL, reliable diagnostic and prognostic assay of PKDL are required. OBJECTIVE: Previously, glycoproteins (9-OAcSA) have been reported as promising biomarkers of Indian VL patients. However, till date, the status of glycans in Indian PKDL patients remains unexplored. Accordingly, in this study, the glyco-profile of PKDL Circulating Immune Complexes (CICs) as compared to other cross diseases like Vitiligo and Leprosyhas been investigated. Further, a novel Glyco CIC assay has been developed for efficient Indian PKDL patient diagnosis. METHODS/PRINCIPAL FINDING: In the present study, 90 PKDL patients were enrolled from 3 VL endemic districts of West Bengal during 2015-16. Glycosylation profile of isolated CICs from sera of PKDL patients were initially analyzed through gradient SDS gel electrophoresis followed by PAS silver double staining, which revealed the presence of several glycan rich PKDL specific proteins of varying molecular weights. To further characterize the glyco-profile of acid dissociated affinity purified immuno-reactive antigens present in the CICs, glycosylation was demonstrated in these purified CIC antigens by DIG glycan differentiation kit with or without glycosidase as well as neuraminidase treatment. Diagnostic evaluation of the newly developed colorimetric Glyco CIC assay through Receiver Operating Characteristic (ROC) curve analysis revealed excellent (0.99) AUC value as compared to other conventional serodiagnostic assays like PEG CIC, Parasite ELISA (IgG and IgM). Additionally, longitudinal monitoring of 18 PKDL patients further revealed its good prognostic utility. CONCLUSION: These results highlight the glycosylation status of CICs among Indian PKDL patients present in all the studied endemic districts of West Bengal. These PKDL biomarkers were completely absent in cross diseases like Vitiligo and Leprosy. Further, the newly developed Glyco CIC assay had an improved sensitivity of 95.6%, specificity of 99.3%, NPV of 97.1% and PPV of 98.9%.

Tuberculoid leprosy and cytomegalovirus retinitis as immune restoration disease in a patient with AIDS.

Here we report a unique case of tuberculoid leprosy and cytomegalovirus retinitis in a 27-year-old female patient with AIDS, suggestive of highly active antiretroviral therapy (HAART)-induced immune restoration disease. After initiation of HAART, the patient presented with decreased visual acuity, hypoesthetic patch with local nerve thickening, and an increase in her CD4+ T cell count. On further investigations cytomegalovirus retinitis and tuberculoid leprosy were confirmed. To our knowledge no case with such a co-existence has previously been reported.