Dermal Leishmaniasis Research Articles

Prevalence of post kala-azar dermal leishmaniasis (PKDL) and treatment seeking behavior of PKDL patients in Nepal.

In Nepal, the burden of post kala-azar dermal leishmaniasis (PKDL) is not known since there is no active case detection of PKDL by the national programme. PKDL patients could pose a challenge to sustain visceral leishmaniasis (VL) elimination. The objective of this study was to determine the prevalence of PKDL and assess PKDL patients' knowledge on VL and PKDL, and stigma associated with PKDL. Household surveys were conducted in 98 VL endemic villages of five districts that reported the highest number of VL cases within 2018-2021. A total of 6,821 households with 40373 individuals were screened for PKDL. Cases with skin lesions were referred to hospitals and examined by dermatologists. Suspected PKDL cases were tested with rK39 and smear microscopy from skin lesions. An integrated diagnostic approach was implemented in two hospitals with a focus on management of leprosy cases where cases with non-leprosy skin lesions were tested for PKDL with rK39. Confirmed PKDL patients were interviewed to assess knowledge and stigma associated with PKDL, using explanatory model interview catalogue (EMIC) with maximum score of 36. Among 147 cases with skin lesions in the survey, 9 (6.12%) were confirmed as PKDL by dermatologists at the hospital. The prevalence of PKDL was 2.23 per 10,000 population. Among these 9 PKDL cases, 5 had a past history of VL and 4 did not. PKDL cases without a past history of VL were detected among the "new foci", Surkhet but none in Palpa. None of the cases negative for leprosy were positive for PKDL. There was very limited knowledge of PKDL and VL among PKDL cases. PKDL patients suffered to some degree from social and psychological stigma (mean ± s.d. score = 17.89 ± 12.84). Strengthening the programme in PKDL case detection and management would probably contribute to sustenance of VL elimination. Awareness raising activities to promote knowledge and reduce social stigma should be conducted in VL endemic areas.

Revisiting the diagnosis and treatment of Para Kala-azar Dermal Leishmaniasis in the endemic foci of Bangladesh.

Para Kala-azar Dermal Leishmaniasis (Para-KDL) manifests the concomitant presence of Post Kala-azar Dermal Leishmaniasis and Visceral Leishmaniasis and works as a reservoir of infection. The study discusses the cases and their management and aims to address the gaps within existing methods of diagnosis and treatment. This retrospective cross-sectional study discusses 16 Para-KDL cases with one-year follow-up data, treated between 2012-2021 at the Surya Kanta Kala-azar Research Center, Bangladesh. We collected data from hospital records and used STATA 16 to analyze and see the frequency distribution and variable means. We found five patients without any history of kala-azar infection. All the patients were treated with 20 mg/kg Liposomal Amphotericin B in 4 divided doses except one with a history of AmBisome hypersensitivity. One year after treatment, all patients were free from skin lesions, with no hepatosplenomegaly, and observed significant improvement in BMI and hemoglobin levels. The Para-KDL patients are challenging to diagnose, and the relapse and treatment failure leishmania patients might have belonged to this rare group, contributing to their poor prognosis. Therefore, developing an appropriate diagnostic workflow and a new drug regimen is essential to sustain the success of our elimination efforts.

Gauging the skin resident Leishmania parasites through a loop mediated isothermal amplification (LAMP) assay in post-kala-azar dermal leishmaniasis

Despite the availability of highly sensitive polymerase chain reaction (PCR)-based methods, the dearth of remotely deployable diagnostic tools circumvents the early and accurate detection of individuals with post-kala-azar dermal leishmaniasis (PKDL). Here, we evaluate a design-locked loop-mediated isothermal amplification (LAMP) assay to diagnose PKDL. A total of 76 snip-skin samples collected from individuals with probable PKDL (clinical presentation and a positive rK39 rapid diagnostic test (RDT)) were assessed by microscopy, qPCR, and LAMP. An equal number of age and sex-matched healthy controls were included to determine the specificity of the LAMP assay. The LAMP assay with a Qiagen DNA extraction (Q-LAMP) showed a promising sensitivity of 72.37% (95% CI: 60.91–82.01%) for identifying the PKDL cases. LAMP assay sensitivity declined when the DNA was extracted using a boil-spin method. Q-qPCR showed 68.42% (56.75–78.61%) sensitivity, comparable to LAMP and with an excellent agreement, whereas the microscopy exhibited a weak sensitivity of 39.47% (28.44–51.35%). When microscopy and/or qPCR were considered the gold standard, Q-LAMP exhibited an elevated sensitivity of 89.7% (95% CI: 78.83–96.11%) for detection of PKDL cases and Bayesian latent class modeling substantiated the excellent sensitivity of the assay. All healthy controls were found to be negative. Notwithstanding the optimum efficiency of the LAMP assay towards the detection of PKDL cases, further optimization of the boil-spin method is warranted to permit remote use of the assay.

Development and validation of an ultra-high performance liquid chromatography coupled to tandem mass spectrometry method for the quantification of the antileishmanial drug paromomycin in human skin tissue

Bioanalytical assay development and validation procedures were performed to quantify antiprotozoal drug paromomycin in human skin tissue by ultra-high performance liquid chromatography coupled to tandem mass spectrometry. Paromomycin, an aminoglycoside drug, is administered intra-muscularly and used in the treatment of multiple clinical presentations of the neglected tropical disease leishmaniasis. It is currently studied in the treatment of post-kala-azar dermal leishmaniasis, a disease where the Leishmania parasites divide and reside in the skin. We present a target-site bioanalytical method to accurately quantify paromomycin in human skin tissue, with the clinical purpose of quantifying paromomycin in skin biopsies from post-kala-azar dermal leishmaniasis patients originating from Sudan. Enzymatic digestion using collagenase A incubated at 37 °C overnight was employed as homogenization method to produce skin tissue homogenates. Further sample preparation was performed by protein precipitation using trichloroacetic acid and a dilution step. Final extracts were injected onto a C18 analytical column and isocratic heptafluorobutyric acid ion-pair separation and elution were employed. The chromatography system was coupled to a triple quadrupole mass spectrometer for detection. The method was validated in digestion solution over a linear range from 5 to 1000 ng/mL (r2 ≥ 0.9967) with the assay performance of accuracy and precision within acceptable criteria values as stated by the EMA guidelines. Furthermore, matrix effects were observed in human skin tissue and were corrected by the multiple deuterated paromomycin internal standard. No substantial IS-normalized matrix effect was detected along with relatively high sample preparation recovery. Consequently, digestion solution matrix serving as the preparation of calibration standards can be used as surrogate matrix for human skin tissue, which is convenient given the limited availability of control matrix. Finally, paromomycin was accurately quantified in skin of post-kala-azar dermal leishmaniasis patients originating from clinical trials in Sudan.

Prevalence and determinants of asymptomatic Leishmania infection in HIV-infected individuals living within visceral leishmaniasis endemic areas of Bihar, India

People living with HIV (PLHIV) have an increased risk of developing visceral leishmaniasis (VL) and poor outcomes compared to HIV negative individuals. Here, we aim to establish the prevalence and determinants of asymptomatic Leishmania infection (ALI) in a cohort of PLHIV in Bihar, India. We hoped to evaluate optimal diagnostic algorithms to detect ALI in PLHIV. We conducted a cross-sectional survey of PLHIV ≥18 years of age with no history or current diagnosis of VL or post kala-azar dermal leishmaniasis (PKDL) at anti-retroviral therapy centres within VL endemic districts of Bihar. ALI was defined as a positive rK39 enzyme-linked immunosorbent assay (ELISA), rK39 rapid diagnostic test (RDT) and/or quantitative polymerase chain reaction (qPCR). Additionally, the urinary Leishmania antigen ELISA was evaluated. Determinants for ALI were established using logistic regression and agreement between diagnostic tests calculated using Cohen’s Kappa. A total of 1,296 PLHIV enrolled in HIV care, 694 (53.6%) of whom were female and a median age of 39 years (interquartile range 33–46), were included in the analysis. Baseline prevalence of ALI was 7.4% (n = 96). All 96 individuals were positive by rK39 ELISA, while 0.5% (n = 6) and 0.4% (n = 5) were positive by qPCR and rK39 RDT, respectively. Negligible or weak agreement was seen between assays. Independent risk factors for ALI were CD4 counts <100 (OR 3.1; 95% CI 1.2–7.6) and CD4 counts 100–199 (OR = 2.1;95% CI:1.1–4.0) compared to CD4 counts ≥300, and a household size ≥5 (OR = 1.9;95% CI:1.1–3.1). A total of 2.2% (n = 28) participants were positive by Leishmania antigen ELISA, detecting 20 additional participants to the asymptomatic cohort. Prevalence of ALI in PLHIV in VL endemic villages in Bihar was relatively high. Using the Leishmania antigen ELISA, prevalence increased to 9.0%. Patients with low CD4 counts and larger household size were found to have significantly higher risk of ALI.Trial Registration: Clinical Trial Registration CTRI/2017/03/008120.

LEISH2b - A phase 2b study to assess the safety, efficacy, and immunogenicity of the Leishmania vaccine ChAd63-KH in post-kala azar dermal leishmaniasis.

Background: The leishmaniases are neglected tropical diseases caused by various Leishmania parasite species transmitted by sand flies. They comprise a number of systemic and cutaneous syndromes including kala-azar (visceral leishmaniasis, VL), cutaneous leishmaniasis (CL), and post-kala-azar dermal leishmaniasis (PKDL). The leishmaniases cause significant mortality (estimated 20 - 50,000 deaths annually), morbidity, psychological sequelae, and healthcare and societal costs. Treatment modalities remain difficult. E.g., East African PKDL requires 20 days of intravenous therapy, andfrequently relapsing VL is seen in the setting of HIV and immunodeficiency. We developed a new therapeutic vaccine, ChAd63-KH for VL / CL / PKDL and showed it to be safe and immunogenic in a phase 1 trial in the UK, and in a phase 2a trial in PKDL patients in Sudan. Methods: This is a randomised double-blind placebo-controlled phase 2b trial to assess the therapeutic efficacy and safety of ChAd63-KH in patients with persistent PKDL in Sudan. 100 participants will be randomly assigned 1:1 to receive placebo or ChAd63-KH (7.5 x10 10vp i.m.) at a single time point. Follow up is for 120 days after dosing and we will compare the clinical evolution of PKDL, as well as the humoral and cellular immune responses between the two arms. Discussion: Successful development of a therapeutic vaccine for leishmaniasis would have wide-ranging direct and indirect healthcare benefits that could be realized rapidly. For PKDL patients, an effective therapeutic vaccination used alone would have very significant clinical value, reducing the need for extensive hospitalization and chemotherapy. Combining vaccine with drug (immuno-chemotherapy) might significantly increase the effective life of new drugs, with lower dose / abbreviated regimens helping to limit the emergence of drug resistance. If therapeutic benefit of ChAd63-KH can be shown in PKDL further evaluation of the vaccine in other forms of leishmaniasis should be considered. Clinicaltrials.gov registration: NCT03969134.

Immune recovery-related patterns of post kala-azar dermal and ocular leishmaniasis in people living with HIV.

Post kala-azar dermal leishmaniasis (PKDL) is a rare complication of visceral leishmaniasis. We aimed at reporting PKDL cases in people living with HIV (PLHIV) and compare their characteristics based on whether PKDL occurred in the context of immune recovery under antiretroviral therapy (ART) or not. National survey and literature review. We called for observations in France in October 2020 and performed a literature review from PubMed (Medline) and Web of Science up to December 2020. Two groups of patients were defined based on whether PKDL occurred in the context of immune recovery under ART (group 1) or not (group 2), and compared. Three PLHIV with PKDL identified in France in the last decade were described and added to 33 cases from the literature. Compared with group 2 (16/36, 44.4%), patients from group 1 (20/36, 55.6%) originated more frequently from Europe (12/20, 60% vs. 2/16, 12.5%; P = 0.0038), had higher median blood CD4 + cell counts (221/μl vs. 61/μl; P = 0.0005) and increase under ART (122/μl, interquartile range 73-243 vs. 33/μl, interquartile range 0-53; P = 0.0044), had less frequently concomitant visceral leishmaniasis (3/20, 15% vs. 8/12, 66.7%; P = 0.006), and a trend to more frequent ocular involvement (7/20, 35% vs. 1/16, 6.25%; P = 0.0531). In PLHIV, PKDL occurs after a cured episode of visceral leishmaniasis as part of an immune restoration disease under ART, or concomitant to a visceral leishmaniasis relapse in a context of AIDS. For the latter, the denomination 'disseminated cutaneous lesions associated with visceral leishmaniasis' seems more accurate than PKDL.

Development and validation of a high-performance liquid chromatography tandem mass spectrometry method for the quantification of the antiparasitic and antifungal drug amphotericin B in human skin tissue

Amphotericin B is an antifungal and antiparasitic drug used in first-line treatment of the parasitic neglected tropical disease leishmaniasis. Liposomal amphotericin B is currently studied for the treatment of cutaneous and post-kala-azar dermal leishmaniasis, where the dermis of the skin is infected with Leishmania parasites. For the optimization of known treatment regimens, accurate target-site concentrations of the drug are required. To date, no assay was available to assess human skin concentrations of amphotericin B. We here present a bioanalytical assay for the quantification of amphotericin B in 4-mm human skin biopsies. Human skin biopsies were homogenized by overnight digestion using collagenase A and were processed afterwards by simple protein precipitation using methanol. Separation and detection were achieved using a Gemini C18 column with slightly acidic chromatographic conditions and a quadrupole – linear ion trap mass spectrometer, respectively. The method was validated in digestion solution over a range of 10–2,000 ng/mL using natamycin as internal standard, with a correlation coefficient (r2) of at least 0.9974. The assay performance, accuracy and precision, were acceptable over the validated range, using international (EMA and FDA) acceptance criteria. In the skin tissue extracts, amphotericin B ion enhancement was observed, however, the internal standard (IS) corrected for this effect hence calibration standards in digestion solvent could be used as a surrogate matrix for the quantification in skin tissue. Sample preparation recoveries were low (around 27%) because of degradation of amphotericin B during digestion and sample preparation processes, albeit highly reproducible, without compromising the accuracy and precision of the method. Using this assay, amphotericin B could be detected and quantified in skin biopsies originating from treated Indian post-kala-azar dermal leishmaniasis patients.

TREPs-A New Class of Functional tRNA-Encoded Peptides.

We asked if transfer RNA (tRNA) ever got an opportunity of translating its own sequence during evolution, what would have been the function of such tRNA-encoded peptides (tREPs)? If not, could one artificially synthesize tREPs to study the corresponding functional outcomes? Here, we report a novel, first-in-the-class, chemically synthesized tREP-18 molecule originating from the Escherichia coli tRNA sequence showing potent antileishmanial property. As a first step, E. coli tRNAs were computationally translated into peptide sequence equivalents and a database of full-length hypothetical tREPs was created. The tREP sequences were sent into sequence, structure, and energy filters to narrow down potential peptides for experimental validation. Based on the functional predictions, tREPs were screened against antiparasitic targets, leading to the identification of tREP-18 as a potential antiparasitic peptide. The in vitro assay of chemically synthesized tREP-18 on the Ag83 strain of Leishmania donovani showed its potent antileishmanial property (IC50 value of 22.13 nM). The atomic force microscopy and scanning electron microscopy images indicated significant alteration in the cytoskeletal architecture of tREP-18-treated parasites. Also, tREP-18 seems to destabilize the mitochondrial membrane potential of parasites, disrupting their cellular integrity and leading to parasitic death. The cellular assays of the tREP-18 peptide on the BS12 strain, a clinical isolate of post-kala azar dermal leishmaniasis, demonstrated its significant efficacy at an IC50 value of 15 nM. The tREP-18 peptide showed a toxic effect on the amastigote stage of the parasite, showing macrophage pathogen clearance at a concentration of 22.5 nM. This study provides the proof of the concept of making a new class of functional peptides from tRNA sequences. It also opens a huge untapped tRNA-peptide space toward novel discoveries and applications. In the future, it would be interesting to perform tREP edits and redesign tREPs toward specific applications.

Quantitative monitoring of experimental and human leishmaniasis employing amastigote-specific genes.

The gold standard for diagnosis of leishmaniasis is the microscopic detection of amastigotes/Leishman Donovan (LD) bodies, but its moderate sensitivity necessitates the development of molecular approaches. This study aimed to quantify in experimental animal models and human leishmaniasis the expression of amastigote-specific virulence genes, A2 and amastin by droplet digital polymerase chain reaction (ddPCR). Total RNA was isolated from L. donovani-infected hamsters or murine peritoneal macrophages and lesional biopsies from patients with post kala-azar dermal leishmaniasis (PKDL). Following cDNA conversion, EvaGreen-based ddPCR was performed using specific primers for A2 or amastin and parasite load expressed in copies per μL. Assay was optimized and the specificity of amastigote-specific A2 and amastin was confirmed. In hepatic and splenic tissues of L. donovani-infected hamsters and peritoneal macrophages, ddPCR demonstrated a greater abundance of A2 than amastin. Treatment of L. donovani-infected peritoneal macrophages with conventional anti-leishmanials, miltefosine and amphotericin B translated into a dose-dependent reduction in copies per μL of A2 and amastin, and the extrapolated IC50 was comparable with results obtained by counting LD bodies in Giemsa-stained macrophages. Similarly, in dermal biopsies of patients with PKDL, A2 and amastin were detected. Overall, monitoring of A2 by ddPCR can be an objective measure of parasite burden and potentially adaptable into a high throughput approach necessary for drug development and monitoring disease progression when the causative species is L. donovani.

Metalloprotease Gp63-Targeting Novel Glycoside Exhibits Potential Antileishmanial Activity.

Visceral leishmaniasis (VL) and post kala-azar dermal leishmaniasis (PKDL) affect most of the poor populations worldwide. The current treatment modalities include liposomal formulation or deoxycholate salt of amphotericin B, which has been associated with various complications and severe side effects. Encouraged from the recent marked antimalarial effects from plant-derived glycosides, in this study, we have exploited a green chemistry-based approach to chemically synthesize a library of diverse glycoside derivatives (Gly1–12) and evaluated their inhibitory efficacy against the AG83 strain of Leishmania donovani. Among the synthesized glycosides, the in vitro inhibitory activity of Glycoside-2 (Gly2) (1.13 µM IC50 value) on L. donovani promastigote demonstrated maximum cytotoxicity with ~94% promastigote death as compared to amphotericin B that was taken as a positive control. The antiproliferative effect of Gly2 on promastigote encouraged us to analyze the structure–activity relationship of Gly2 with Gp63, a zinc metalloprotease that majorly localizes at the surface of the promastigote and has a role in its development and multiplication. The result demonstrated the exceptional binding affinity of Gly2 toward the catalytic domain of Gp63. These data were thereafter validated through cellular thermal shift assay in a physiologically relevant cellular environment. Mechanistically, reduced multiplication of promastigotes on treatment with Gly2 induces the destabilization of redox homeostasis in promastigotes by enhancing reactive oxygen species (ROS), coupled with depolarization of the mitochondrial membrane. Additionally, Gly2 displayed strong lethal effects on infectivity and multiplication of amastigote inside the macrophage in the amastigote–macrophage infection model in vitro as compared to amphotericin B treatment. Gp63 is also known to bestow protection against complement-mediated lysis of parasites. Interestingly, Gly2 treatment enhances the complement-mediated lysis of L. donovani promastigotes in serum physiological conditions. In addition, Gly2 was found to be equally effective against the clinical promastigote forms of PKDL strain (IC50 value of 1.97 µM); hence, it could target both VL and PKDL simultaneously. Taken together, this study reports the serendipitous discovery of Gly2 with potent antileishmanial activity and proves to be a novel chemotherapeutic prototype against VL and PKDL.

Screening and identification of potential inhibitor for visceral leishmaniasis (VL) through computational analysis

AimThe aim of this investigation is to detect potential inhibitor for visceral leishmaniasis through computational analysis. BackgroundLeishmaniasis is categorized as a vector born pathogenic infection prevalent in tropical, subtropical, and in Mediterranean zones spread by intra-macrophage protozoa. The clinical syndrome of leishmaniasis is divided into the following type’s namely cutaneous leishmaniasis, mucocutaneous leishmaniasis, visceral leishmaniasis, and dermal leishmaniasis. Trypanothione synthetase is a key enzyme involving in glutathione biosynthesis as well as hydrolysis. Trypanothione is one of the promising drug targets for parasites. Parasites are inimitable with concern to their dependence on trypanothione to regulate intracellular thiol-redox balance in fighting against oxidative stress and biochemical anxiety. However, trypanothione synthetase was presumed as the target therapeutic alternate in VL therapy. ObjectiveThe important objective of this current investigation is to identify or analyze the potential inhibitor for V. leishmaniasis through computational approaches which include virtual screening, molecular docking, ADME prediction, and molecular dynamic simulation. MethodsAn investigation was performed to develop a 3D protein structure, using computational screening among associated similar structured proteins from popular compound database banks such as Specs, Maybridge, and Enamine, to detect novel staging with a series of validation for emerging innovative drugs molecules. Modeled protein ligand complex was further analyzed to know the binding ability of the complex. Molecular dynamics were performed to ascertain its stability at 50 ns. ResultsTrypanothione synthetase overall ability in the outcome of series of analysis. Among three database compounds screened, the compound from the Specs database exhibited the better protein-ligand docking scores and fulfilled the drug-like properties through ADMET analysis, and the docked complexes had better stability throughout the simulation. Besides, the other two database leads fulfilled the pharmacological properties, and the complexes were stable in the simulation. ConclusionBy analyzing the various compounds from different databases, we concluded that the Specs database compound exhibits potential activity against the target protein and is considered a promising inhibitor for trypanothione synthetase.

CURRENT STRATEGIES AND RECENT ADVANCES IN NANOSTRUCTURED DELIVERY SYSTEMS FOR MANAGING LEISHMANIASIS-A REVIEW

Neglected tropical diseases (NTDs) comprise of a group of 17th infectious conditions endemic in many developing countries. Among these diseases, deadly, prevalent parasitic infection such as leishmaniasis, presents a significant global burden which is responsible for high morbidity/mortality rate especially in low-and middle-income/developed countries transmitted by phlebotomines sandflies. As per the WHO report, a total of 0.7-1.0 million new leishmaniasis cases, which are spread by 23 leishmania species in more than 98 countries are estimated with an alarming 26,000-65,000 death toll every year. The disease can be characterized by at least four syndromes: cutaneous leishmaniasis (CL), muco-cutaneous leishmaniasis (MCL), visceral leishmaniasis (VL) also known as kala-azar in the Indian sub-continent or black fever, which is the most firm form of the disease being lethal if untreated and post-kala-azar dermal leishmaniasis (PKDL). The lack of cheap, portable, easy to use diagnostic tools exhibiting high efficiency and specificity impede the early diagnosis of the disease. Its intracellular nature and disseminated locations of parasite, limited number of chemotherapeutic agents, increasing incidences of resistance to first line drugs, along with the cost, toxicities, low patient compliance and require long-term regimen and usually hospitalization, pose a great challenge to formulation scientists that have necessitated effective management of leishmaniasis infection by modulating the delivery of existing drugs. Over the past decade, research on development of alternative treatments such as nanotechnology-based drug delivery systems (nanoparticles, nanotubes, nanosuspensions, liposomes, nanovaccines, etc.), use of natural products as well as development of antileishmanial vaccine has been extensively investigated due to their unique properties, such as bioavailability, lowered toxicity, targeted drug delivery, high biocomatibilty and biodegradability. To identify all relevant literature, we searched Web of Sciences, Scopus, PubMed, NCBI, Scielo, Google Scholar, and profiled studies published between 1986 and 2020. The aim of this review is to summarize new advances and new strategies used on leishmaniasis therapy addressing alternative and innovative treatment paths such as combination or multi-drug uses, immunomodulation, drug repurposing and the nanotechnology-based drug delivery systems have been used to improve the therapeutic aspects of existing antileishmanial drugs, presenting a critical review and some suggestions for improving drug delivery.

A report on occurrence of Phlebotomine sand flies (Diptera: Psychodidae) and two new country records from Andaman amp; Nicobar Islands, a Union territory of India.

A cross-sectional survey was carried out to determine sand fly fauna of the Andaman and Nicobar Islands that are known for tourist attraction from different countries, due to their scenic beauty. The study area includes 40 localities, stretching from south and middle to north Andaman Islands. Sand fly collections were made from day time resting shelters such as human dwellings, cattle-shed, and bushes in fringe of villages. A total of 1055 sand flies, comprising of six species were collected. Se. (Neo.) barraudi was found to be the predominant species, comprising of 57.7% of the total. Other species found to occur were Se. (Par.) babu (27. 4%), Se. (bailyi) 8. 1%, Se. (Neo.) gemmea (4. 9%), Se. (Neo.) quatei (2. 1%) and Phlebotomus (Ana.) stantoni (0.3%). Se. (Neo.) gemmea is reported to be a vector transmitting visceral leishmaniasis and dermal leishmaniasis in southern Thailand. Se. (Neo.) gemmea and Se (Neo.) quatei are new country records adding to the existing list of sand fly fauna of India. This is the first report on sand flies from Andaman and Nicobar Islands.

Assessment of Kala-Azar in Dumka District of Jharkhand, India

Kala-azar is a vector-borne disease and a serious public health problem in India. Highly affected states in India are Bihar, Jharkhand, West Bengal and Uttar Pradesh. In 2010, 14.87% cases were contributed from Jharkhand. The study was done to know the trend of disease in five year duration (2016-2020) in Dumka District, Jharkhand. Method – This was a cross-sectional study. It was executed in 10 Blocks of District Dumka Jharkhand. Convenience sampling technique was used in this study. Data analysis was done from the line-list of District Vector Borne Disease Control Office, Dumka. Result – During the 5 year study period there was annual reduction in the trend of visceral leishmaniasis. Male cases were more predominant. Age group &gt;21 years were maximum affected. VL had affected all the 10 Blocks. PKDL cases were seen maximum in Blocks like Ramgarh, Dumka Sada, Kathikund and Gopikander. Conclusion- Annual reduction of incidence of Kala-azar reveals that the current Kala-azar elimination strategy is highly effective and elimination of disease from endemic blocks is possible. Further improvement in treatment, elimination strategies and accessibility of primary health care services in endemic rural areas can gear up to get over last-mile obstacles. Keywords: Visceral Leishmaniasis, Indoor Residual Spraying, Post Kala-azar Dermal Leishmaniasis, Kala-azar.

An In-depth Proteomic Map of Leishmania donovani Isolate from Post Kala-azar Dermal Leishmaniasis (PKDL) Patient.

The trypanosomatid protozoan parasite Leishmania donovani is the etiological agent of visceral leishmaniasis (VL) or kala-azar. The patients that have undergone treatment may still harbor the parasite and in a small fraction of the patients the disease re-erupts in the form of post kala-azar dermal leishmaniasis (PKDL). PKDL is a pathological condition found to be intermediate between VL and complete cure of VL. The PKDL disease progression is determined by the host immune response to L. donovani. The majority of the proteomic studies on L. donovani till date have been undertaken on parasites either isolated from kala-azar patients or on established laboratory strains of L. donovani. However, no proteomic information is available on the cutaneous localized isolates of L. donovani from PKDL patients. The promastigote stage of L. donovani isolate from PKDL patient was cultured and harvested. The cell lysates were trypsin digested, followed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. The LC-MS/MS raw data were analyzed on Proteome Discoverer. Further bioinformatics analysis was carried out. In the present, we have used high-resolution mass spectrometry to map the global proteome of a L. donovani isolate from PKDL patient. This in-depth study resulted in the identification of 5537 unique proteins from PKDL isolate of L. donovani which covered 64% of its proteome. This study also identified proteins previously shown to be upregulated in PKDL L. donovani. This is the most in-depth proteome of Leishmania donovani parasite till date.

Utility of smear examination, culture, and serological tests in the diagnosis of visceral leishmaniasis/post-kala-azar dermal leishmaniasis at National Centre for Disease Control, Delhi.

Introduction:A range of assays have been developed to detect specific antileishmanial antibody, such as rK 39 immunochromatographic test (ICT), KE 16 ICT, ELISA test, and indirect immunofluorescent antibody test (IFAT), which play a crucial role in serological diagnosis of visceral leishmaniasis (VL). However, limited published reports are available on the utility of serological test (IFAT test/rk 39), smear examination, and culture in the diagnosis of VL and post-kala-azar dermal leishmaniasis (PKDL) in our country.Materials and Methods:We present utility of serological test (IFAT test/rK 39), smear examination for Leishmania donovani (LD) bodies, and culture in 2589 samples from 2294 VL/PKDL suspected patients (January 2009–December 2019) tested in Centre for Arboviral and Zoonotic diseases, National Centre for Disease Control, New Delhi, India, for laboratory diagnosis of VL/PKDL.Results:A total of 80/553 (14.4%) cases were confirmed of VL (74/522 cases by demonstration of LD bodies in bone marrow smear examination, 5/12 in splenic smear examination 1/19 by culture) and 4/21 (19.0%) cases were confirmed of PKDL (demonstration of LD bodies in slit skin smear examination. In our study 197/1368 (14.4%) cases were diagnosed positive by IFAT, 34/646 (5.2%) cases by rk 39 ICT for VL/PKDL by demonstration of specific antileishmanial antibodies.Conclusion:As the goal of elimination of VL as a public health problem is approaching, apart from serological tests such as rk 39 and IFAT, direct methods of detection such as (parasitic demonstration in BM smear, culture, and molecular tests) for Leishmania may play a crucial role for achieving a correct diagnosis and treatment. We also concluded that IFAT though not field-friendly, its optimal use as an adjunct test with BM smear in all stages of infections may be required. Further rk39 is a simple, reliable, noninvasive, and field-friendly test for diagnosis VL, especially in endemic areas.

Diagnosis of Post Kala Azar Dermal Leishmaniasis in Nonendemic Areas

Leishmaniasis is a heterogeneous group of disease caused by protozoans of the genus Leishmania, transmitted by Phlebotomus sandflies. Human leishmaniasis is usually classified as visceral, cutaneous (localized, diffuse, disseminated), mucocutaneous, and post-kala-azar leishmaniasis. Post-kala-azar leishmaniasis is a sequel of visceral leishmaniasis (VL) caused by Leishmania donovani (LD) and seen in India, Bangladesh, Nepal, and Sudan. In India, it develops in 5%–15% of treated VL cases within 2–5 years. It is characterized by multiple papular, nodular, and pleomorphic lesions over the sun-exposed areas such as face, neck, and arms, without systemic manifestations. It mimics a wide range of dermatoses, particularly leprosy in endemic areas for both the diseases. Inadequate immune response (Th1 and Th2) to infection during and after the treatment of VL plays a role in the pathogenesis. It is thought to be a good reservoir of LD and plays an important role in the treatment of VL. Post-kala-azar dermal leishmaniasis (PKDL) is usually diagnosed by skin smears, immunological tests such as rK39, cutaneous leishmaniasis detection tests, and polymerase chain reaction. A combination of liposomal amphotericin B and miltefosine is the first line of treatment. PKDL is a forgotten clinical entity in the nonendemic areas. We report a case of PKDL in a 52-year-old male from Andhra Pradesh, a nonendemic area for leishmaniasis. This case was treated successfully with combination therapy, liposomal amphotericin B, and miltefosine.

Road‐map of pre‐clinical treatment for Visceral Leishmaniasis

Visceral leishmaniasis (VL) or Kala-azar, is the most lethal form of leishmaniasis, is still prevalent in many countries where it is endemic. It is a threat to human life caused by protozoan parasite Leishmania donovani. The severity of the disease is further increased as the treated individuals might have a chance of developing Post Kala-azar Dermal Leishmaniasis (PKDL) in the long run. Moreover, several countries have reported high number of HIV-VL co-infected patients. Therefore, there is a dire need for the development of efficient diagnostic methods and drugs in order to combat the disease and to control the spread of disease. At present, the treatment for VL entirely relies on therapeutic drugs as no vaccine is available yet. Ever since 1900s a series of drugs have been invented and used for treatment of VL; but the need for one such cost-effective treatment that would completely cure the disease with minimal side-effects, low relapse rate with high efficacy and less toxicity remains yet to be fulfilled. Therefore, identifying novel compounds is very crucial to develop potent antileishmanial agents. Thus, this review enlists several instances of drug development, including the pharmacokinetic and pharmacodynamic properties of antileishmanial drugs, different experimental animal models used to investigate the disease progression and to analyze treatment dosage and pharmacological aspect of drugs. Furthermore, the existing gap in drug development and future measures to improve the process are also discussed in this review.

Generalized old world leishmaniasis: first Moroccan case in an immunocompetent adult?

RésuméIntroductionLa leishmaniose dermique post kala-azar (LDPK) est un syndrome cutané observé après un traitement de leishmaniose viscérale (LV). Nous décrivons un cas probable chez un adulte immunocompétent.ObservationIl s'agit d'un homme âgé de 36 ans, originaire du sud du Maroc, avec antécédent de LV deux ans auparavant, traitée par antimoniate de méglumine et amphotéricine B avec une bonne évolution, hospitalisé en dermatologie pour un placard papulo-nodulaire érythémateux de la face. Le tableau dermatologique remontait à 6 mois avec une atteinte inaugurale de la muqueuse buccale. L'examen de la muqueuse buccale retrouvait une ulcération du tiers postérieur de la langue et un aspect papillomateux du voile du palais. La biopsie et le frottis cutanés retrouvaient des amastigotes de Leishmania. La PCR ITS1 était positive (genre Leishmania). Le diagnostic de LDPK était évoqué. Le patient a bénéficié d'injections intra-musculaires d'antimoniate de méglumine avec une bonne évolution.ConclusionÀ notre connaissance, il s'agit du premier cas de leishmaniose généralisée évoquant une LDPK décrit au Maroc chez un adulte immunocompétent.