Post-kala-azar Dermal Leishmaniasis Research Articles

Insights from mathematical modelling and quantitative analysis on the proposed WHO 2030 targets for visceral leishmaniasis on the Indian subcontinent.

Visceral leishmaniasis (VL) is a neglected tropical disease (NTD) caused by Leishmania protozoa that are transmitted by female sand flies. On the Indian subcontinent (ISC), VL is targeted by the World Health Organization (WHO) for elimination as a public health problem by 2020, which is defined as <1 VL case (new and relapse) per 10,000 population at district level in Nepal and sub-district level in Bangladesh and India. WHO is currently in the process of formulating 2030 targets, asking whether to maintain the 2020 target or to modify it, while adding a target of zero mortality among detected cases. The NTD Modelling Consortium has developed various mathematical VL transmission models to gain insight into the transmission dynamics of VL, identify the main knowledge gaps, and predict the feasibility of achieving and sustaining the targets by simulating the impact of varying intervention strategies. According to the models, the current target is feasible at the appropriate district/sub-district level in settings with medium VL endemicities (up to 5 reported VL cases per 10,000 population per year) prior to the start of the interventions. However, in settings with higher pre-control endemicities, additional efforts may be required. We also highlight the risk that those with post-kala-azar dermal leishmaniasis (PKDL) may pose to reaching and sustaining the VL targets, and therefore advocate adding control of PKDL cases to the new 2030 targets. Spatial analyses revealed that local hotspots with high VL incidence remain. We warn that the current target provides a perverse incentive to not detect/report cases as the target is approached, posing a risk for truly achieving elimination as a public health problem although this is taken into consideration by the WHO procedures for validation. Ongoing modelling work focuses on the risk of recrudescence when interventions are relaxed after the elimination target has been achieved.

The impact of leishmaniasis on mental health and psychosocial well-being: A systematic review.

BackgroundLeishmaniasis is a neglected tropical parasitic disease endemic in South Asia, East Africa, Latin America and the Middle East. It is associated with low socioeconomic status (SES) and responsible for considerable mortality and morbidity. Reports suggest that patients with leishmaniasis may have a higher risk of mental illness (MI), psychosocial morbidity (PM) and reduced quality of life (QoL), but this is not well characterised. The aim of this study was to conduct a systematic review to assess the reported impact of leishmaniasis on mental health and psychosocial wellbeing.MethodsA systematic review of the literature was carried out. Pre-specified criteria were applied to identify publications including observational quantitative studies or systematic reviews. Two reviewers screened all of the titles, abstracts and full-studies and a third reviewer was consulted for disagreements. Data was extracted from papers meeting the criteria and quality appraisal of the methods was performed using the Newcastle-Ottawa Scale or the Risk of Bias in Systematic Review tool.ResultsA total of 14 studies were identified from 12,517 records. Nine cross-sectional, three case-control, one cohort study and one systematic review were included. Eleven assessed MI outcomes and were measured with tools specifically designed for this; nine measured PM and 12 measured QoL using validated measurement tools. Quality appraisal of the studies showed that six were of good quality. Cutaneous leishmaniasis and post kala-azar dermal leishmaniasis showed evidence of associated MI and PM including depression, anxiety and stigma, while all forms of disease showed decreased QoL. The findings were used to inform a proposed model and conceptual framework to show the possible links between leishmaniasis and mental health outcomes.ConclusionThere is evidence that leishmaniasis has an impact on MI, PM or QoL of patients and their families and this occurs in all the main subtypes of the disease. There are however large gaps in the evidence. Further research is required to understand the full extent of this problem and its mechanistic basis.

Relationship of Serum Antileishmanial Antibody With Development of Visceral Leishmaniasis, Post-kala-azar Dermal Leishmaniasis and Visceral Leishmaniasis Relapse.

IntroductionTo sustain the achievement of kala-azar elimination program (KEP), early detection and treatment of the visceral leishmaniasis (VL) cases and associated modalities such as treatment failure (TF), relapse VL (RVL), and Post-kala-azar dermal leishmaniasis (PKDL) is the cornerstone. A predictive biomarker for VL development and related complications could also play a crucial role in curtailing disease incidence and transmission. Investigations to find a biomarker with prospective capabilities are, however, scarce. Using samples and known clinical outcomes generated within two previous longitudinal cohort studies, we aimed to determine if fluctuations in serum anti-rK39 antibody levels could provide such predictive value.Materials and MethodsSerum samples collected at four different time points (Baseline, 12, 18, and 24 months) from 16 patients who had developed VL within the monitoring period and 15 of their asymptomatic healthy controls counterparts were investigated. To investigate potential prediction of VL related complications, serum samples of 32 PKDL, 10 RVL, 07 TF, and 38 cured VL from a single dose AmBisome trial were analyzed. Of this second panel, all patients were monitored for 5 years and sera were collected at four time points (Baseline then 1, 6, and 12 months after treatment). The level of anti-rK39 antibodies in archived samples was measured by a semi-quantitative ELISA.ResultsThe mean antibody level was significantly higher in VL patients compared to their asymptomatic healthy counterparts at each time point. Likewise, we observed a trend toward elevations in antibody levels for PKDL, RVL, TF relative to the reducing levels observed in cured VL. Receiver operating characteristic (ROC) analysis found a promising predictive power of rK39 antibody levels to reveal progression from asymptomatic Leishmania donovani infection stage to VL, defined as 87.5% sensitive and 95% specific. Following treatment, rk39 antibody notably showed 100% sensitivity and 95% specificity in predicting TF.ConclusionOur data indicate that the relative quantity of serum anti-rK39 antibody has promise within either a predictive or prognostic algorithm for VL and VL-related modalities. These could enable VL control programs to implement more effective measures to eliminate the disease. Further research is, however, imperative to standardize the rK39 antibody ELISA between sites prior to broader use.

In-situ immune profile of polymorphic vs. macular Indian Post Kala-azar dermal leishmaniasis

Post Kala-azar Dermal Leishmaniasis (PKDL), a sequel of apparently cured Visceral Leishmaniasis presents in South Asia with papulonodular (polymorphic) or hypomelanotic lesions (macular). Till date, the polymorphic variant was considered predominant, constituting 85–90%. However, following active-case surveillance, the proportion of macular PKDL has increased substantially to nearly 50%, necessitating an in-depth analysis of this variant. Accordingly, this study aimed to delineate the cellular infiltrate in macular vis-à-vis polymorphic PKDL. To study the overall histopathology, hematoxylin and eosin staining was performed on lesional sections and phenotyping by immunohistochemistry done in terms of dendritic cells (CD1a), macrophages (CD68), HLA-DR, T-cells (CD8, CD4), B-cells (CD20) and Ki67 along with assessment of the status of circulating homing markers CCL2, CCL7 and CXCL13. In polymorphic cases (n = 20), the cellular infiltration was substantial, whereas in macular lesions (n = 20) it was mild and patchy with relative sparing of the reticular dermis. Although parasite DNA was identified in both variants by ITS-1 PCR, the parasite load was significantly higher in the polymorphic variant and Leishman-Donovan bodies were notably minimally present in macular cases. Both variants demonstrated a decrease in CD1a+ dendritic cells, HLA-DR expression and CD4+ T-cells. In macular cases, the proportion of CD68+ macrophages, CD8+ T-cells and CD20+ B-cells was 4.6 fold, 17.0 fold and 1.6 fold lower than polymorphic cases. The absence of Ki67 positivity and increased levels of chemoattractants suggested dermal homing of these cellular subsets. Taken together, as compared to the polymorphic variant, patients with macular PKDL demonstrated a lower parasite load along with a lesser degree of cellular infiltration, suggesting differences in host-pathogen interactions, which in turn can impact on their disease transmitting potential and responses to chemotherapy.

Relationship between treatment regimens for visceral leishmaniasis and development of post-kala-azar dermal leishmaniasis and visceral leishmaniasis relapse: A cohort study from Bangladesh.

BackgroundWe investigated the relationship of treatment regimens for visceral leishmaniasis (VL) with post-kala-azar dermal leishmaniasis (PKDL) and visceral leishmaniasis relapse (VLR) development.MethodsStudy subjects included cohorts of patients cured of VL since treatment with monotherapy by sodium stibogluconate (SSG), miltefosine (MF), paromomycin intramuscular injection (PMIM), liposomal amphotericin B [AmBisome (AMB)] in a single dose (SDAMB) and in multidose (MDAMB), and combination therapies by AMB+PMIM, AMB+MF, and PMIM+MF. Follow up period was 4 years after treatment. Cohorts were prospective except SSG (retrospective) and MF (partially retrospective). We compared incidence proportion and rate in 100-person-4year of PKDL and VLR by treatment regimens using univariate and multivariate analysis.Findings974 of 984 enrolled participants completed the study. Overall incidence proportion for PKDL and VLR was 12.3% (95% CI, 10.4%–14.5%) and 7.0% (95% CI, 5.6%–8.8%) respectively. The incidence rate (95% CI) of PKDL and VLR was 14.0 (8.6–22.7) and 7.6 (4.1–14.7) accordingly. SSG cohort had the lowest incidence rate of PKDL at 3.0 (1.3–7.3) and VLR at 1.8 (0.6–5.6), followed by MDAMB at 8.2 (4.3–15.7) for PKDL and at 2.7 (0.9–8.4) for VLR.InterpretationDevelopment of PKDL and VLR is related with treatment regimens for VL. SSG and MDAMB resulted in less incidence of PKDL and VLR compared to other treatment regimens. MDAMB should be considered for VL as a first step for prevention of PKDL and VLR since SSG is highly toxic and not recommended for VL.

Impact of sequelae of visceral leishmaniasis and their contribution to ongoing transmission of Leishmania donovani.

ABSTRACTVisceral leishmaniasis (VL) in the Old World is caused by infection with Leishmania donovani. Although the numbers of new reported cases of VL in Africa have been relatively stable for several years, the low numbers currently reported on the Indian subcontinent suggest a positive impact of new treatments and intervention strategies. In both regions, however, VL relapse and post-kala-azar dermal leishmaniasis (PKDL) maintain infectious reservoirs and therefore present a threat to control programs. In this review, we outline the evolving appreciation of PKDL as an impactful disease in its own right and discuss the various diagnostic methods that can be applied for the detection and characterization of PKDL cases. We also highlight the data that indicate the potential, and likely contribution, of PKDL cases to ongoing transmission of L. donovani.

Biomarkers in Post-kala-azar Dermal Leishmaniasis.

Post-kala-azar dermal leishmaniasis (PKDL) follows visceral leishmaniasis (VL, kala-azar) in 10–60% of cases. It is characterized by an asymptomatic skin rash, usually starting in the face and consisting of macules, papules, or nodules. Diagnosis is difficult in the field and is often made clinically. There is an extensive differential diagnosis, and parasitological confirmation is preferred particularly when drug treatment is considered. The response to treatment is difficult to assess as this may be slow and lesions take long to heal, thus possibly exposing patients unnecessarily to prolonged drug treatment. Biomarkers are needed; these may be parasitological (from microscopy, PCR), serological (from blood, or from the lesion), immunological (from blood, tissue), pathological (from cytology in a smear, histology in a biopsy), repeated clinical assessment (grading, photography), or combinations. In this paper, we will review evidence for currently used biomarkers and discuss promising developments.

Application of kDNA Minicircle PCR-RFLP to Characterize Leishmania donovani Clinical Isolates Obtained from Post-Kala-Azar Dermal Leishmaniasis in Eastern Nepal.

Post-kala-azar dermal leishmaniasis (PKDL) is a skin manifestation of visceral leishmaniasis (VL) which develops after apparent cure in some patients. PKDL is considered as the potential reservoir for the VL infection. Molecular epidemiological characterization of L. donovani isolates obtained from VL and PKDL isolates is essentially required in order to understand the transmission dynamics of the VL infection. To date, genetic variation among the VL and PKDL L. donovani isolates was not fully elucidated. Therefore, 14 clinical isolates from VL and 4 clinical isolates from PKDL were speciated by hsp70 and rDNA genes. Further characterization of L. donovani by haspB PCR demonstrates two different genotypes. All PKDL isolates have the same genetic structure. kDNA PCR-RFLP assay revealed 18 different genotypes; however, structural analysis showed the two distinct kDNA genotype population (k = 2). The kDNA fingerprint patterns of parasites from hilly districts were clustered separately from low-land districts. Therefore, further study with a large number of samples is urgently required for systematic characterization of the clinical isolates to track the molecular epidemiology of the Leishmania donovani causing VL and the role of PKDL as a reservoir.

Post-Kala-Azar Dermal Leishmaniasis as a Reservoir for Visceral Leishmaniasis Transmission

Post-kala-azar dermal leishmaniasis (PKDL) is a parasitic skin infection which can occur after visceral leishmaniasis (VL). Recent xenodiagnosis studies (Mondal et al., Clin. Infect. Dis., 2018) have uncovered the infectiousness of PKDL. When including this in a transmission model, PKDL cases appear as an important reservoir of infection, likely frustrating the VL elimination efforts on the Indian subcontinent.

OC 8489 CLINICAL DEVELOPMENT OF A THERAPEUTIC VACCINE FOR PREVENTION OF POST KALA AZAR DERMAL LEISHMANIASIS

BackgroundThe leishmaniases represent a complex of human diseases, with 350 million people at risk of infection worldwide. Although the potential benefits of vaccination have been well-recognised, no human vaccine is registered. Post-kala azar dermal leishmaniasis (PKDL) is a chronic skin disease often following treatment for visceral leishmaniasis (VL). In addition to affecting quality of life, evidence suggests that PKDL patients may also act as reservoirs for VL transmission. Hence, PKDL vaccines may have a significant impact on disease burden. We recently developed a third-generation adenoviral vaccine for leishmaniasis (ChAd63-KH) that has been evaluated for safety and immunogenicity in healthy volunteers (Osman et al,2017). ChAd63-KH is currently being evaluated for safety as a therapeutic in Sudanese PKDL patients, with a phase IIb RCT starting in late 2018. With EDCTP funding, we are initiating a new phase IIa/IIb study (PREV_PKDL) to’determine whether ChAd63-KH can prevent PKDL development.MethodsIn PREV_PKDL, we will conduct an open-label phase IIa safety study, followed by a placebo blinded, phase IIb RCT. Safety and clinical response represent primary outcome measures, and immunogenicity is a secondary outcome measure. In addition, working across the four countries of Leishmaniasis East Africa Platform (LEAP), we will use deep phenotyping methods to study the immune status of patients before and after treatment for VL to understand why PKDL development is limited to specific geographic regions. This work, and other research in the region, will be supported by the creation of a new flow cytometry ‘centre of excellence’ within LEAP.ResultsAn update on the progress of our current therapeutic trial in PKDL patients will be provided.ConclusionPREV_PKDL represents an important step in the clinical development of ChAd63-KH and will help develop capacity to support future vaccine and drug trials for leishmaniasis in the East Africa Region.

Lipsosomal amphotericin B: a review of its properties, function, and use for treatment of cutaneous leishmaniasis.

The genus Leishmania includes a number of protozoan parasites that cause a wide range of infections named leishmaniasis. Leishmaniasis may be appear in three clinical forms — cutaneous (CL), visceral, and mucocutaneous (MCL) — with variation in their presentation and severity: diffuse CL and post–kala-azar dermal leishmaniasis). The prevalent signs of CL are nonhealing ulcers on exposed skin, but infected patients may have other dermatologic symptoms. In the 1960s, amphotericin B deoxycholate was introduced as a second-line therapy for CL and MCL. However, widespread administration of the agent was prevented, due to its renal and systemic toxicity, high price, and obstacles to intravenous use in leishmaniasis-endemic regions. Amphotericin B binds to ergosterol in the photogenic cell membranes and causes changes in membrane permeability, leakage of ions, and finally cell death. Compared to amphotericin B deoxycholate, a higher dose of liposomal amphotericin B should be administered to show the treatment effect. A high percentage of liposomal amphotericin B is “fastened” in the liposome and not biologically effective. Amphotericin B deoxycholate has some toxic effects, and liposomal amphotericin B is meaningfully less toxic compared to it. Treatment options for CL are limited, due to variation in species causing CL and pharmacokinetic issues. Amphotericin B is effective against some particular forms of CL.

The usefulness of trained field workers in diagnosis of post-kala-azar dermal leishmaniasis (PKDL) and clinico-epidemiological profile in highly endemic areas of Bihar.

Surveillance of post-kala-azar dermal leishmaniasis (PKDL) is critical to the elimination of visceral leishmaniasis (VL). In this study we assessed the feasibility of using trained field workers for detecting suspected PKDL cases. A cross-sectional study using a multistage sampling technique was conducted in the Araria district of Bihar. Trained field workers were utilized for identification of suspected PKDL case. We investigated 57 099 individuals from 11 300 households. The trained field workers were useful in identifying 107 (18%) probable PKDL cases. The calculated PKDL prevalences were 18.7/10 000 and 9.7/10 000 for probable and confirmed PKDL cases, respectively. The median duration of onset of PKDL was 23 months (interquartile range 16.5-56.5). The younger age group developed PKDL significantly more often compared with the older age group (p=0.007). Of the 107 patients, 25 (55.5%) were positive by microscopy of slit skin smear and 42 (93.3%) by polymerase chain reaction. Of 45 patients, 33 (73%) PKDL cases were cured after full treatment. The risk of not being cured with incomplete treatment was three times higher than with complete treatment (relative risk 3.12 [95% confidence interval 1.23 to 8.67], p=0.004). We conclude that the prevalence of PKDL is high and the use of trained field workers may be feasible to actively detect PKDL cases in VL-endemic areas of Bihar, India.

Histopathological study of post kala-azar dermal leishmaniasis and its importance for diagnosis in a peripheral tertiary institution

Histopathological study of post kala-azar dermal leishmaniasis and its importance for diagnosis in a peripheral tertiary institution - IJCED- Print ISSN No: - 2581-4710 Online ISSN No:- 2581-4729 Article DOI No:- 10.18231/2581-4729.2019.0003, IP Indian Journal of Clinical and Experimental Dermatology-IP Indian J Clin E

Cyto-Pathological Diagnosis of Post Kala-Azar Dermal Leishmaniasis (PKDL) in Tertiary Care Hospital, At Patna, Bihar

Cyto-Pathological Diagnosis of Post Kala-Azar Dermal Leishmaniasis (PKDL) in Tertiary Care Hospital, At Patna, Bihar

Active surveillance identified a neglected burden of macular cases of Post Kala-azar Dermal Leishmaniasis in West Bengal.

BackgroundPost Kala-azar Dermal Leishmaniasis (PKDL) develops in patients apparently cured of Visceral Leishmaniasis (VL), and is the strongest contender for being the disease reservoir. Therefore, existence of a few cases is sufficient to trigger an epidemic of VL in a given community, emphasizing the need for its active detection and in turn ensuring success of the current elimination program. This study explored the impact of active surveillance on the demographic profile of PKDL patients in West Bengal.Methodology/Principal findingsPatients with PKDL were recruited through passive (2003-date, n = 100) and active surveillance (2015-date, n = 202), the former from outpatient departments of dermatology in medical colleges in West Bengal and the latter through an active door-to-door survey in four VL hyper-endemic districts of West Bengal. Passive surveillance indicated a male preponderance and a predominance of polymorphic lesions, whereas active surveillance indicated absence of any gender bias and more importantly, macular PKDL constituted almost 50% of the population burden. In terms of polymorphic vs. macular PKDL, the former appeared at a later age, their disease duration was longer and had a higher parasite burden. In the polymorphic variant, the lesional distribution was asymmetrical, comprised of papules/nodules/macules that were present mainly in sun-exposed areas whereas in macular cases, the hypopigmented patches were diffusely present all over the body.Conclusions/SignificanceActive surveillance unraveled a disease component whose demographic profile showed important differences with PKDL cases who sought treatment in government hospitals. Detection of a higher proportion of macular cases indicates that this variant is not an uncommon presentation as conventionally stated in text books, and should be studied in greater detail to ensure success of the ongoing Leishmaniasis elimination programme.

Target Product Profile for a point-of-care diagnostic test for dermal leishmaniases

ObjectivesLocalized cutaneous leishmaniasis and its evolving forms diffuse cutaneous leishmaniasis, mucosal leishmaniasis and cutaneous leishmaniasis recidivans, together with the visceral leishmaniasis sequelae post-kala azar dermal leishmaniasis account for about one million dermal leishmaniases cases per year worldwide. Although not lethal, the dermal leishmaniases cause chronic and disfiguring skin lesions, which are an important cause of morbidity and stigma.Microscopy remains the reference test for diagnosis of dermal leishmaniasis; however, it has low and variable sensitivity and requires well trained personnel. The technical complexity and cost of the more sensitive molecular techniques (e.g. PCR) limits their application in routine diagnosis in endemic areas. Point-of-care (POC) tests for early diagnosis are much needed in order to benefit both patients and communities, by reducing the risk of both sequelae and Leishmania transmission. To this end we developed a Target Product Profile (TPP) for a POC test for dermal leishmaniases. MethodsThe TPP was defined through several rounds of discussions and by consensus with stakeholders and experts in dermal leishmaniases from different type of organizations and endemic regions. Results and conclusionsA rapid, simple and robust test that can be implemented in resource-limited settings, enabling decentralized diagnosis and treatment of dermal leishmaniasis should be developed. Ideally it should enable the diagnosis of all forms of dermal leishmaniasis, but the minimally accepted target would be localized cutaneous leishmaniasis. A minimum sensitivity of 95% and specificity of 90% would be required. The consensus was that the POC test should target Leishmania antigens.

The safety and efficacy of miltefosine in the long-term treatment of post-kala-azar dermal leishmaniasis in South Asia - A review and meta-analysis.

BackgroundMiltefosine (MF) is the only oral drug available for treatment of visceral leishmaniasis (VL) and post-kala-azar dermal leishmaniasis (PKDL). Although the drug is effective and well tolerated in treatment of VL, the efficacy and safety of MF for longer treatment durations (>28 days) in PKDL remains unclear. This study provides an overview of the current knowledge about safety and efficacy of long treatment courses with MF in PKDL, as a strategy in the VL elimination in South Asia.Methodology/Principal findingsLiterature was searched systematically for articles investigating MF treatment in PKDL. A meta-analysis included eight studies (total 324 PKDL patients) to estimate the efficacy of MF in longer treatment regimens (range 6–16 weeks). We found a per-protocol (PP) initial cure rate of 95.2% (95%CI 89.6–100.8) and a PP definite cure rate of 90% (95%CI 81.6–96.3). Descriptive analysis showed that 20% of patients experienced adverse events, which mostly had an onset in the first week of treatment and were likely to get more severe after four weeks of treatment. Gastrointestinal (GI) side effects such as vomiting, nausea, diarrhoea, and abdominal pain were most common.Conclusions/SignificanceLonger treatment regimens with MF are effective in PKDL patients in India, however with the caveat that the efficacy has recently been observed to decline. GI side effects are frequent, although mostly mild or moderate. However, on the basis of limited data, we cannot conclude that longer MF treatment regimens are safe. Moreover, VL and PKDL pharmacovigilance studies indicate a risk for serious adverse events, questioning the safety of MF. The provision of safer treatment regimens for PKDL patients are therefore recommended. Until these regimens are identified, it should be considered to halt the use of MF monotherapy for PKDL in order to preserve the drug’s efficacy.

Impaired activation of lesional CD8+ T-cells is associated with enhanced expression of Programmed Death-1 in Indian Post Kala-azar Dermal Leishmaniasis

Post Kala-azar dermal leishmaniasis (PKDL), caused by Leishmania donovani is the dermal sequel of Visceral Leishmaniasis and importantly, is the proposed disease reservoir. The survival of Leishmania parasites within monocytes/macrophages hinges on its ability to effectively nullify immune activation mechanisms. Thus, delineating the disease-promoting immune mechanisms can facilitate development of immunotherapeutic strategies. Accordingly, in the absence of an animal model, this study aimed to delineate the status of CD8+ T-cells in patients with PKDL. At disease presentation, the absence of CD4+ T-cells at lesional sites was concomitant with an overwhelming infiltration of CD8+ T-cells that demonstrated an absence of Perforin, Granzyme and Zap-70, along with an enhanced expression of Programmed Death-1 (PD-1) and the skin-homing CCL17. Additionally, the lesional CCR4+CD8+ population was associated with an enhanced expression of IL-10 and IL-5. In circulation, the enhanced CD8+CCR4+ T-cell population and raised levels of CCL17/22 was associated with an increased frequency of PD-1, while CD127 was decreased. Taken together, in PKDL, the enhanced plasma and lesional CCL17 accounted for the dermal homing of CD8+CCR4+ T-cells, that along with a concomitant upregulation of PD-1 and IL-10 mediated immune inactivation, emphasizing the need for designing immunotherapies capable of reinvigorating T-cell potency.

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

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. 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. 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. 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%. The Gal2 enzyme-linked immunosorbent assay needs to be further investigated in mass surveys. 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.

Visceral Leishmaniasis IgG1 Rapid Monitoring of Cure vs. Relapse, and Potential for Diagnosis of Post Kala-Azar Dermal Leishmaniasis.

Background: There is a recognized need for an improved diagnostic test to assess post-chemotherapeutic treatment outcome in visceral leishmaniasis (VL) and to diagnose post kala-azar dermal leishmaniasis (PKDL). We previously demonstrated by ELISA and a prototype novel rapid diagnostic test (RDT), that high anti-Leishmania IgG1 is associated with post-treatment relapse versus cure in VL.Methodology: Here, we further evaluate this novel, low-cost RDT, named VL Sero K-SeT, and ELISA for monitoring IgG1 levels in VL patients after treatment. IgG1 levels against L. donovani lysate were determined. We applied these assays to Indian sera from cured VL at 6 months post treatment as well as to relapse and PKDL patients. Sudanese sera from pre- and post-treatment and relapse were also tested.Results: Of 104 paired Indian sera taken before and after treatment for VL, when deemed clinically cured, 81 (77.9%) were positive by VL Sero K-SeT before treatment; by 6 months, 68 of these 81 (84.0%) had a negative or reduced RDT test line intensity. ELISAs differed in positivity rate between pre- and post-treatment (p = 0.0162). Twenty eight of 33 (84.8%) Indian samples taken at diagnosis of relapse were RDT positive. A comparison of Indian VL Sero K-SeT data from patients deemed cured and relapsed confirmed that there was a significant difference (p < 0.0001) in positivity rate for the two groups using this RDT. Ten of 17 (58.8%) Sudanese sera went from positive to negative or decreased VL Sero K-SeT at the end of 11–30 days of treatment. Forty nine of 63 (77.8%) PKDL samples from India were positive by VL Sero K-SeT.Conclusion: We have further shown the relevance of IgG1 in determining clinical status in VL patients. A positive VL Sero K-SeT may also be helpful in supporting diagnosis of PKDL. With further refinement, such as the use of specific antigens, the VL Sero K-SeT and/or IgG1 ELISA may be adjuncts to current VL control programmes.