First Serological Study Revealing High Humoral Response and Evidence for Antigenic Heterogeneity in Leishmania donovani Induced CL in Sri Lanka

Abstract

Posing a threat to the ongoing leishmaniasis elimination efforts in the Indian subcontinent, L. donovani-induced cutaneous leishmaniasis (CL) has been recently reported in many countries. Sri Lanka reports a large focus of human cutaneous leishmaniasis (CL) caused by Leishmania donovani, a usually visceralizing parasite. Enhanced case detection, early treatment, and in-depth understanding of sequalae are required to contain the spread of disease. Visceralizing potential of dermotropic strains has not been fully ruled out. Sri Lankan strains have shown a poor response to established serological assays. The present concern was to develop an in-house serological assay and to determine the seroprevalence of CL for identifying visceralizing potential and its usefulness in enhancing case detection. Crude cell lysate of dermotropic L. donovani promastigotes-based indirect enzyme-linked immunosorbent assay (ELISA) was previously optimized. Assay was evaluated using sera from 200 CL patients, 50 endemic and 50 nonendemic healthy controls, 50 patients with other skin diseases, and 50 patients with other systemic diseases. Seroprevalence and clinicoepidemiological associations were analyzed. Assay was compared with light microscopy (LM) and in vitro culturing (IVC). Cost comparison was carried out. Seroprevalence of CL was 82.0%. The assay had 99.5% specificity, and all healthy controls were negative at 0.189 cut-off. Positive and negative predictive values were 99.4% and 84.7%, respectively. Positivity obtained in ELISA was comparable to LM and higher than that of IVC. Cost per patient was 3.0 USD for both ELISA and LM and 6.0 USD for IVC. Infections occurring in all age groups and both genders demonstrated >75.0% of seropositivity. Patients had lesions with different durations/types/sizes showed >70.0% of seropositivity. Study identified a high seroprevalence of L. donovani-induced CL for the first time, indicating potential for visceralization or transient serological response. This can be used as a second line test in LM-negative CL cases to enhance clinical case detection. Further studies are warranted to examine in-depth correlations, antigen profiles, comparison with other established serological tools, and usefulness in the detection of asymptomatic cases. (National patent LK/P/1/19697).

An Insight Into Systemic Immune Response in Leishmania donovani Mediated Atypical Cutaneous Leishmaniasis in the New Endemic State of Himachal Pradesh, India

“…Further, with Leishmania being an intracellular pathogen, the total of T-cell immunity comprising heterogeneous T-cell subsets with discrete effector cytokines direct the course of the disease ranging from non-symptomatic to mild-to-severe VL and CL phenotypes ( 14 , 16 , 17 , 21 , 22 ). With substantial insight into the host immune correlates that underline the classical visceral and cutaneous manifestation, scarce reports are available on host immune correlates in L. donovani atypical CL disease emerging in known and newer endemic sites in the Indian subcontinent ( 10 , 30 – 32 , 35 , 36 ). Knowledge on comparative host immune response in LdCL versus typical VL is important to define the key immune checkpoints that can modulate the disease towards atypical phenotype ( 9 – 12 , 37 ).…”

ELISA‐based evaluation of antibody response to Leishmania in a region endemic for cutaneous leishmaniasis

“…Promastigote forms of L. donovani grown in complete medium 199 at late log phase were harvested. 27 The culture was subjected to centrifugation at 15,000 rpm for 10 min and the isolated pellet was freeze-thawed (frozen in liquid nitrogen and thawed at 37 C) five times. The suspension was disrupted with a syringe (21G needle) a couple of times and the cell debris was pelleted down at 1000 rpm for 5 min.…”

“…Previous studies on CL patients which evaluated the rK39 antibody response 24 and expression of peroxidoxin antigen of Leishmania amastigotes using commercially available immunodiagnostic assays showed low sensitivity in the local setting. 25 Demonstration of a high seroprevalence in L. donovani-induced CL in recent local studies 26,27 warrants further evaluation of these humoral immune responses at different clinical stages.…”

Validation of an In-House ELISA Method in the Diagnosis of Cutaneous Leishmaniasis Caused by Leishmania donovani in Hambantota District, Sri Lanka

“…Recent studies in Sri Lanka have estimated a seroprevalence rate of 34% among CL cases [ 20 ]. A follow-up study revealed a higher seroprevalence of 82% using crude protein lysate of the local dermatropic strain [ 21 ]. Even though CL in Sri Lanka is caused by L. donovani , a normally visceralizing parasite, the rK39 antibody detection has been consistently low in Sri Lanka [ 20 , 22 ].…”