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Blood transfusion remains an important aspect of patient management in visceral leishmaniasis (VL). However, transfusion triggers considered are poorly understood. This review summarises the transfusion practices adopted in VL efficacy studies using the Infectious Diseases Data Observatory VL clinical trials library. Of the 160 studies (1980-2021) indexed in the IDDO VL library, description of blood transfusion was presented in 16 (10.0%) (n=3459 patients) studies. Transfusion was initiated solely based on haemoglobin (Hb) measurement in nine studies, combining Hb measurement with an additional condition (epistaxis/poor health/clinical instability) in three studies and the criteria was not mentioned in four studies. The Hb threshold range for triggering transfusion was 3-8 g/dL. The number of patients receiving transfusion was explicitly reported in 10 studies (2421 patients enrolled, 217 underwent transfusion). The median proportion of patients who received transfusion in a study was 8.0% (Interquartile range: 4.7% to 47.2%; range: 0-100%; n=10 studies). Of the 217 patients requiring transfusion, 58 occurred before VL treatment initiation, 46 during the treatment/follow-up phase and the time was not mentioned in 113. This review describes the variation in clinical practice and is an important initial step in policy/guideline development, where both the patient's Hb concentration and clinical status must be considered., (© The Author(s) 2024. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene.)

Treatment of visceral leishmaniasis (VL) is compromised by drug toxicity, high cost and/or the emergence of resistant strains. Though canine vaccines are available, there are no licensed prophylactic human vaccines. One strategy to improve clinical outcome for infected patients is immunotherapy, which associates a chemotherapy that acts directly to reduce parasitism and the administration of an immunogen-adjuvant that activates the host protective Th1-type immune response. In this study, we evaluated an immunotherapy protocol in a murine model by combining recombinant (r)LiHyp1 (a hypothetical amastigote-specific Leishmania protein protective against Leishmania infantum infection), with monophosphoryl-lipid A (MPLA) as adjuvant and amphotericin B (AmpB) as reference antileishmanial drug. We used this protocol to treat L. infantum infected-BALB/c mice, and parasitological, immunological and toxicological evaluations were performed at 1 and 30 days after treatment. Results showed that mice treated with rLiHyp1/MPLA/AmpB presented the lowest parasite burden in all organs evaluated, when both a limiting dilution technique and qPCR were used. In addition, these animals produced higher levels of IFN-γ and IL-12 cytokines and IgG2a isotype antibody, which were associated with lower production of IL-4 and IL-10 and IgG1 isotype. Furthermore, low levels of renal and hepatic damage markers were found in animals treated with rLiHyp1/MPLA/AmpB possibly reflecting the lower parasite load, as compared to the other groups. We conclude that the rLiHyp1/MPLA/AmpB combination could be considered in future studies as an immunotherapy protocol to treat against VL., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

The protozoan parasite Leishmania donovani, residing and replicating within the cells of the monocyte-macrophage (mono-mac) lineage, causes visceral leishmaniasis (VL) in humans. While, Leishmania infantum, is the main causative agent for zoonotic VL, where dogs are the main reservoirs of the disease. The chemotherapy is a serious problem because of restricted repertoire of drugs, drug-resistant parasites, drug-toxicity and the requirement for parenteral administration, which is a problem in resource-starved countries. Moreover, immunocompromised individuals, particularly HIV-1 infected are at higher risk of VL due to impairment in T-helper cell and regulatory cell responses. Furthermore, HIV-VL co-infected patients report poor response to conventional chemotherapy. Recent efforts are therefore directed towards devising both prophylactic and therapeutic immunomodulation. As far as prophylaxis is concerned, although canine vaccines for the disease caused by Leishmania infantum or Leishmania chagasi are available, no vaccine is available for use in humans till date. Therefore, anti-leishmanial immunotherapy triggering or manipulating the host's immune response is gaining momentum during the last two decades. Immunomodulators comprised of small molecules, anti-leishmanial peptides, complex ligands for host receptors, cytokines or their agonists and antibodies have been given trials both in experimental models and in humans. However, the success of immunotherapy in humans remains a far-off target. We, therefore, propose that devising a successful immunotherapy is an act of balancing enhanced beneficial Leishmania-specific responses and deleterious immune activation/hyperinflammation just as the swings in a trapeze., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Immunotherapy, a treatment based on host immune system activation, has been shown to provide a substitute for marginally effective conventional chemotherapy in controlling visceral leishmaniasis (VL), the deadliest form of leishmaniasis. As the majority of endemic inhabitants exhibit either subclinical or asymptomatic infection which often develops into the active disease state, therapeutic intervention seems to be an important avenue for combating infections by stimulating the natural defense system of infected individuals. With this perspective, the present study focuses on two immunodominant Leishmania ( L. ) donovani antigens (triosephosphate isomerase and enolase) previously proved to be potent prophylactic VL vaccine candidates, for generating a recombinant chimeric antigen. This is based on the premise that in a heterogeneous population, a multivalent antigen vaccine would be required for an effective response against leishmaniasis (a complex parasitic disease). The resulting molecule rLdT-E chimeric protein was evaluated for its immunogenicity and immunotherapeutic efficacy. A Th1 stimulating adjuvant BCG was employed with the protein which showed a remarkable 70% inhibition of splenic parasitic multiplication positively correlated with boosted Th1 dominant immune response against lethal L. donovani challenge in hamsters as evidenced by high IFN- γ and TNF- α and low IL-10. In addition, immunological analysis of antibody subclass presented IgG2-based humoral response besides considerable delayed-type hypersensitivity and lymphocyte proliferative responses in rLdT-E/BCG-treated animals. Our observations indicate the potential of the chimera towards its candidature for an effective vaccine against Leishmania donovani infection., Competing Interests: The authors declare no conflict of interest., (Copyright © 2021 Sneha Ratnapriya et al.)

Visceral Leishmaniasis (VL) is the most fatal form of disease leishmaniasis. To date, there are no effective prophylactic measures and therapeutics available against VL. Recently, new immunotherapy-based approaches have been established for the management of VL. Cytokines, which are predominantly produced by helper T cells (Th) and macrophages, have received great attention that could be an effective immunotherapeutic approach for the treatment of human VL. Cytokines play a key role in forming the host immune response and in managing the formation of protective and non-protective immunities during infection. Furthermore, immune response mediated through different cytokines varies from different host or animal models. Various cytokines viz. IFN-γ, IL-2, IL-12, and TNF-α play an important role during protection, while some other cytokines viz. IL-10, IL-6, IL-17, TGF-β, and others are associated with disease progression. Therefore, comprehensive knowledge of cytokine response and their interaction with various immune cells is very crucial to determine appropriate immunotherapies for VL. Here, we have discussed the role of cytokines involved in VL disease progression or host protection in different animal models and humans that will determine the clinical outcome of VL and open the path for the development of rapid and accurate diagnostic tools as well as therapeutic interventions against VL., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Samant, Sahu, Pandey and Khare.)

Background: Zoonotic visceral leishmaniasis by Leishmania infantum is a first-order pathology in canine veterinary clinics in endemic areas. Moreover, canine infections are considered the main reservoir for human disease; despite their importance in the control of the disease within a One Health approach, no scientometric study has been published. Aims of the study included analyzing the impact of canine leishmaniasis (CanL) on the scientific literature, drugs or combinations used, trends in the period from 2000 to 2020 and efficacy criteria employed., Methods: A Web of Science (WOS)-based analysis of publications on CanL and chemotherapy of the disease in the period 2000-2020 was carried out using a stepwise methodology. Data were analyzed by year, geographical origin, chemical groups, drugs and combinations, and efficacy criteria., Results: Reports on CanL (n = 3324) represented < 16% of all publications on leishmaniasis (n = 20,968), and of these around 18% (n = 596) were related to chemotherapy. Publication records on CanL followed the distribution of the infection by L. infantum in endemic areas although Mediterranean countries were overrepresented in the reports on chemotherapy of CanL. Publications on the main antileishmanial drugs used in clinical practice showed a sustained tendency in the period analyzed. Pentavalent antimonials (Sb V ), alone or in combination with allopurinol, represented > 50% of all publications on chemotherapy of CanL despite the availability of more recently marketed drugs., Conclusions: Chemotherapy of CanL still relies on Sb V and combinations and to a lesser extent on miltefosine (MIL). Reports on chemotherapy are scarce and mostly publicly funded, and the variability of experimental conditions hampers the direct comparison of the efficacy of drugs, combinations and schedules. The vast majority of reports on efficacy do not include any information on supportive therapy; this reduces the actual value of the studies if intended for the practical management of the disease. Complete reports on the chemotherapy (etiological + symptomatic) would add value to the trials performed.

Aims: Given the involvement of IL-9 in the immune responses to parasitic infections, we aimed to determine alterations in the levels of IL-9+CD4+ T cells and the cytokines influencing their differentiations and functions following treatment in paediatric visceral leishmaniasis (VL)., Methods and Results: Eighteen VL and 20 healthy children were included. The levels of IL-9+CD4+ T cells and cytokines influencing their differentiations and functions were measured in the blood and PBMC culture supernatant at the onset of diagnosis and 1 and 2 weeks and 2 months after treatment, using flow cytometer. IL-9+CD4+ T cells, IL-2 and TNF-α were significantly higher in the blood of VL patients than those in the controls; however, following treatment, IL-9+CD4+ T cells down-regulated and IL-33 and IFN-γ significantly up-regulated. After ex vivo stimulation, although the released cytokines were not significantly different between the study groups, the levels of IL-2, IL-9 and IFN-γ significantly decreased., Conclusions: The higher frequency of IL-9+CD4+ T cells and its decline following treatment implies their roles in the immunopathogenesis of VL; however, at the diagnosis onset, lower levels of serum IL-9 and its higher level in the culture supernatant may confer in vivo dysfunction of IL-9+CD4+ T cells in the acute phase of human VL., (© 2020 John Wiley & Sons Ltd.)

Background: Visceral leishmaniasis (VL), caused by Leishmania infantum, is a systemic parasitic disease and presents a global health problem which can be fatal if not diagnosed and treated. Dogs are the main hosts and provide reservoirs for the transmission of the disease to humans., Methods: In this study, the gene encoding a 21-kDa protein was cloned and expressed as a fusion protein in Escherichia coli strain BL21 (DE3) for developing a rapid immunochromatographic test (ICT) to identify infected dogs. The expression of the recombinant 21-kDa protein (r21) was investigated using SDS-PAGE and Western blot methods. The purified r21-kDa protein was spotted onto ICT strips and tested by sera from experimentally infected, naturally infected and uninfected dogs., Results: The SDS-PAGE and Western blot methods showed the successful expression of r21-kDa protein. The ICT strip test revealed that the r21-kDa protein was detected by the sera of experimentally and naturally infected dogs. The specificity tests also confirmed no cross-reactivity with animals infected with Trypanosoma cruzi, Toxoplasma gondii and Ehrlichia canis., Conclusions: Based on these findings, the new r21-kDa protein may be a suitable target for developing a new simple, specific and rapid serological method to detect VL in infected dogs., (© 2020 John Wiley & Sons Ltd.)

The aims of this study were to describe a smartphone app aimed at healthcare professionals who work in areas endemic for visceral and tegumentary leishmaniases, and to report the user's perception of the app in these areas. The software, called LeishCare ® , has the following features: data registration, image filter to record the evolution of skin lesions using photos, calculation of a score set to identify the risk of death from visceral leishmaniasis, and guides to the diseases. LeishCare ® was made available to healthcare professionals in endemic municipalities in Brazil, and the perception of potential users was evaluated at baseline and after 6 and 12 months. In the first meeting, 96 (94.1%) of the 102 professionals who knew the app reported positive expectations for its use. The installation of LeishCare ® on the individual device and the evaluation of user perception were completed at 6 months with 16 users and at 12 months with 20 users. More than 90% of the professionals evaluated in both assessments found the information of the app useful. The features related to the calculation of visceral leishmaniasis severity score, and the guides to leishmaniases were the most frequently accessed. Users reported competence gain attributed to the app for all items evaluated. In conclusion, LeishCare ® was found to be a promising tool to help healthcare professionals in endemic areas with leishmaniasis management.

The major issues in available therapeutic modalities against leishmaniasis are cost, toxicity, and the emergence of drug resistance. The aim of this work was to develop a successful therapeutic adjuvant against drug-resistant Leishmania donovani infection by means of combining Mycobacterium indicus pranii with heat-induced promastigotes (HIP). One-month postinfected BALB/c mice were administered subcutaneously with M. indicus pranii (10 8 cells) and HIP (100 μg) for 5 days. Spleens were harvested for flow cytometric and reverse transcriptase PCR analysis. The antileishmanial effect of the combination strategy was associated with induction of a disease-resolving Th1 and Th17 response with simultaneous downregulation of CD4 + CD25 + Foxp3 + (nTreg) cells and CD4 + CD25 - Foxp3 - (Tr1) cells in the spleen. The significant expansion of CD4 + T CM (CD4 + CD44 hi CD11a hi CD62L hi ) cells was a further interesting outcome of this therapeutic strategy in the context of long-term protection of hosts against secondary infection. Toll-like receptor 2 (TLR2) was also found instrumental in this antiparasitic therapy. Induced interleukin-6 (IL-6) production from expanded CD11c + CD8α + (cDC1) and CD11c + CD11b + (cDC2) dendritic cells (DCs) but not from the CD11b + Ly6c + inflammatory monocytes (iMOs), was found critical in the protective expansion of Th17 as evidenced by an in vivo IL-6 neutralization assay. It also promoted the hematopoietic conversion toward DC progenitors (pre-DCs) from common dendritic cell progenitors (CDPs), the immediate precursors, in bone marrow. This novel combinational strategy demonstrated that expansion of Th17 by IL-6 released from CD11c + classical DCs is crucial, together with the conventional Th1 response, to control drug-resistant infection., (Copyright © 2020 American Society for Microbiology.)

The 5th Post-Kala-Azar Dermal Leishmaniasis (PKDL) Consortium meeting brought together PKDL experts from all endemic areas to review and discuss existing and new data on PKDL. This report summarizes the presentations and discussions and provides the overall conclusions and recommendations.

Background: The disturbance of host metabolic pathways by Leishmania parasites has crucial consequences for the activation status of immune cells and the outcome of infection. Glutamine has been described as an immunomodulatory amino acid, yet its role during Leishmania infection is still unknown., Methods: We performed transcriptomics in uninfected and L. donovani-infected macrophages 6 hours post-infection. Glutamine quantification by HPLC was assessed in the supernatant of macrophages throughout the infection course. For experimental L. donovani infections, mice were infected with 1.0 x 108 stationary L. donovani promastigotes. Glutaminase (GLS) chemical inhibition was performed using BPTES and glutamine was administered throughout infection. For combined therapy experiment, a daily administration of miltefosine and glutamine was performed by oral gavage. Parasite burden was determined using a Taqman-based assay. Immune cell phenotyping and cytotoxicity were performed in splenic cells using flow cytometry., Findings: We show that glutamine is essential for the control of L. donovani infection. Transcriptomic analysis of L. donovani-infected macrophages demonstrated an upregulation of genes involved in glutamine metabolism. Pharmacological inhibition of glutaminolysis significantly increased the susceptibility to infection, accompanied by an increased recruitment of anti-inflammatory myeloid cells and impaired T cell responses. Remarkably, the supplementation of glutamine to mice infected with L. donovani during miltefosine treatment potentiates parasite clearance through the development of a more effective anti-Leishmania adaptive immune response., Conclusions: Our data indicates that dietary glutamine supplementation may act as a promising adjuvant for the treatment of visceral leishmaniasis., Competing Interests: The authors have declared that no competing interests exist.

Background: A visceral leishmaniasis outbreak was reported from a village in a low-endemic district of Bihar, India., Methods: Outbreak investigation with house-to-house search and rapid test of kala-azar suspects and contacts was carried out. Sandfly collection and cone bio-assay was done as part of entomological study., Results: A spatially and temporally clustered kala-azar outbreak was found at Kosra village in Sheikhpura district with 70 cases reported till December 2018. Delay of more than a year was found between diagnosis and treatment of the index case. The southern hamlet with socio-economically disadvantaged migrant population was several times more affected than rest of the village (attack rate of 19.0% vs 0.5% respectively, OR MH  = 39.2, 95% CI 18.2-84.4). The median durations between onset of fever to first contact with any health services, onset to kala-azar diagnosis, diagnosis to treatment were 10 days (IQR 4-18), 30 days (IQR 17-73) and 1 day (IQR 0.5 to 3), respectively, for 50 kala-azar cases assessed till June 2017. Three-fourths of these kala-azar cases had out-of-pocket medical expenditure for their condition. Known risk factors for kala-azar such as illiteracy, poverty, belonging to socially disadvantaged community, migration, residing in kutcha houses, sleeping in rooms with unplastered walls and non-use of mosquito nets were present in majority of these cases. Only half the dwellings of the kala-azar cases were fully sprayed. Fully gravid female P. argentipes collected post indoor residual spraying (IRS) and low sandfly mortality on cone-bioassay indicated poor effectiveness of vector control., Conclusions: There is need to focus on low-endemic areas of kala-azar. The elimination programme should implement a routine framework for kala-azar outbreak response. Complete case-finding, use of quality-compliant insecticide and coverage of all sprayable surfaces in IRS could help interrupt transmission during outbreaks.

Visceral leishmaniasis (VL), caused by digenetic protozoa of the genus Leishmania , is the most severe form of leishmaniasis. Leishmania infantum is one of the species responsible for VL and the disease caused is considered a zoonosis whose main reservoir is the dog. Canine visceral leishmaniasis (CVL) can lead to the death of the animal if left untreated. Furthermore, the available pharmocologial treatment for CVL presents numerous disadvantages, such as relapses, toxicity, drug resistance, and the fact treated animals continue to be reservoirs when treatment fails to achieve parasitological cure. Moreover, the available VL control methods have not been adequate when it comes to controlling parasite transmission. Advances in immune response knowledge in recent years have led to a better understanding of VL pathogenesis, allowing new treatments to be developed based on immune system activation, often referred to as immunotherapy. In fact, well-defined protocols have been described, ranging from the use of immunomodulators to the use of vaccines. This treatment, which can also be associated with chemotherapy, has been shown to be effective in restoring or inducing an adequate immune response to reduce parasitic burden, leading to clinical improvement. This review focuses on immunotherapy directed at dogs infected by L. infantum , including a literature review of what has already been done in dogs. We also introduce a promising strategy to improve the efficacy of immunotherapy., (Copyright © 2019 Gonçalves, Leite, Resende, Mariano, Silveira, Melo-Júnior, Ribeiro, de Oliveira, Soares, Santos, Marques, Galdino, Martins-Filho, Dutra, da Silveira-Lemos and Giunchetti.)

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.

Leishmaniasis is a parasitic disease of humans, highly prevalent in parts of the tropics, subtropics, and southern Europe. The disease mainly occurs in three different clinical forms namely cutaneous, mucocutaneous, and visceral leishmaniasis (VL). The VL affects several internal organs and is the deadliest form of the disease. Epidemiology and clinical manifestations of VL are variable based on the vector, parasite (e.g., species, strains, and antigen diversity), host (e.g., genetic background, nutrition, diversity in antigen presentation and immunity) and the environment (e.g., temperature, humidity, and hygiene). Chemotherapy of VL is limited to a few drugs which is expensive and associated with profound toxicity, and could become ineffective due to the parasites developing resistance. Till date, there are no licensed vaccines for humans against leishmaniasis. Recently, immunotherapy has become an attractive strategy as it is cost-effective, causes limited side-effects and do not suffer from the downside of pathogens developing resistance. Among various immunotherapeutic approaches, cytokines (produced by helper T-lymphocytes) based immunotherapy has received great attention especially for drug refractive cases of human VL. Therefore, a comprehensive knowledge on the molecular interactions of immune cells or components and on cytokines interplay in the host defense or pathogenesis is important to determine appropriate immunotherapies for leishmaniasis. Here, we summarized the current understanding of a wide-spectrum of cytokines and their interaction with immune cells that determine the clinical outcome of leishmaniasis. We have also highlighted opportunities for the development of novel diagnostics and intervention therapies for VL.

Background: This study was aimed to assess the impact of quality of life using WHOQOL-BREF in patients with Visceral leishmaniasis (VL)., Methods: A total of 95 VL cases and 95 healthy participants filled out the questionnaires. Data on socio-demographic aspects along with disease duration were collected. Data were compared using a t-test, analysis of variance and chi-square test., Results: VL patients experienced very high impact on their quality of life. Study cohort had male preponderance (72.63%). Majority (64.21%) were aged < 40 years. Longer disease duration was found to have significantly poor quality of life (p < 0.05). The physical domain was found to be most affected domains of quality of life (QOL). QOL was affected most in illiterate, married, housewife, rural population and patients with longer disease duration (p < 0.05). The psychological and environmental domains were significantly affected in > 40 years of age group married patients (p < 0.05)., Conclusions: VL significantly impaired the patients' (QOL) in all four domains (physical, psychological, social relationship and environmental). Physical domain was significantly the most affected domain.

Human visceral leishmaniasis is a growing anthropozoonosis in Brazil, and particularly in the southern region of the country. It is an infectious disease transmitted to humans, dogs and other animals in urban and rural areas of the Americas, mainly due to the bite of Lutzomya longipalpis infected with Leishmania infantum. This article aims to portray the current epidemiological situation of the human visceral leishmaniasis arrival in Porto Alegre city, located in the southern region of Brazil. It is a descriptive study, a case series and a critical review. Six human cases with human visceral leishmaniasis were notified by the date of conclusion of the study, all human visceral leishmaniasis cases were diagnosed at late stage, leading to four deaths., (Copyright © 2019 Sociedade Brasileira de Infectologia. Published by Elsevier España, S.L.U. All rights reserved.)

Introduction: Leishmaniasis is a poverty-related disease that causes a significant socioeconomic burden to affected households. Visceral leishmaniasis is fatal if untreated, yet illness costs may lead to delays in accessing care. Skin manifestations of leishmaniasis cause a psychological burden and even longer treatment trajectories. The objective of this review is to evaluate illness costs associated with leishmaniasis across different settings (Asia, Africa, and Latin America) and the consequences to households. Areas covered: Through a systematic review of cost-of-illness studies, we documented the distribution of costs, the health-seeking behavior, and the consequences of leishmaniasis. We discuss the value of cost-of-illness studies for leishmaniasis. Expert commentary: Despite the free provision of diagnostics and treatment in the public health care sector, out-of-pocket payments remain substantial. There has been progress in addressing the economic burden of leishmaniasis, particularly through the elimination initiative in the Indian subcontinent. Though the illness cost is decreasing due to shorter treatment regimens and better access to care, the situation remains challenging in Africa. Improvement of control tools is critical. There is a need to update cost estimates to inform policy-making and ensure sustainable solutions to reduce financial barriers to leishmaniasis care, especially in pursuing universal health coverage.

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.

Background: Leishmaniasis is a neglected and poorly reported parasitic infection transmitted by sand flies in tropical and subtropical regions. Knowledge about leishmaniasis has become important in non-endemic countries due to increased migration and travel. Few studies of the clinical management of cutaneous, mucocutaneous and visceral leishmaniasis in non-endemic regions have been published to date. In this study, we aimed to evaluate patient characteristics, clinical manifestations and treatments of leishmaniasis in Sweden, over a 20-year period., Methods: A retrospective observational nationwide study was performed using medical records of patients diagnosed with leishmaniasis in Sweden from 1996 to 2016. Cases with culture and polymerase chain reaction verified leishmaniasis were identified at the Public Health Agency of Sweden., Results: In total, 165 cases of leishmaniasis were diagnosed from 1996 to 2016. Medical records from 156 patients (95%) were available for review and included in the study. Cutaneous leishmaniasis was the dominant manifestation (n = 149, 96%), and in 66 patients (44%) cutaneous leishmaniasis was due to Leishmania tropica. Other manifestations were mucocutaneous (n = 4, 3%), visceral (n = 2, 1%) and post-kala-azar dermal leishmaniasis (n = 1, 1%). During this time period, the number of cases increased, especially after 2013. Most patients (n = 81, 52%) were migrants who were infected in their countries of origin (from 2013 to 2016, mainly Syria or Afghanistan). Other groups were Swedish tourists (25%) and returning workers (13%). The time from collection of the diagnostic sample to the start of treatment was less than one month in 81 (66%) patients and under three months in 124 patients (96%). Among the 149 patients with cutaneous leishmaniasis, 125 patients received antileishmanial treatment, and in 88 of these patients (70%) cure was achieved, regardless of treatment., Conclusions: The number of leishmaniasis cases diagnosed in Sweden increased between 1996 and 2016, mainly in migrants from endemic countries. Although leishmaniasis is a rare disease in Sweden, patients appear to be diagnosed early and treated according to current European guidelines, resulting in an overall high cure rate.

Background: Ethiopia bears a high burden of visceral leishmaniasis (VL). Early access to VL diagnosis and care improves clinical prognosis and reduces transmission from infected humans; however, significant obstacles exist. The approximate 250,000 seasonal mobile workers (MW) employed annually in northwestern Ethiopia may be particularly disadvantaged and at risk of VL acquisition and death. Our study aimed to assess barriers, and recommend interventions to increase access, to VL diagnosis and care among MWs., Methodology/principal Findings: In 2017, 50 interviews and 11 focus group discussions were conducted with MWs, mobile residents, VL patients and caretakers, community leaders and healthcare workers in Kafta Humera District, Tigray. Participants reported high vulnerability to VL among MWs and residents engaged in transitory work. Multiple visits to health facilities were consistently needed to access VL diagnosis. Inadequate healthcare worker training, diagnostic test kit unavailability at the primary healthcare level, lack of VL awareness, insufficient finances for care-seeking and prioritization of income-generating activities were significant barriers to diagnosis and care. Social (decision-making and financial) support strongly and positively influenced care-seeking; workers unable to receive salary advances, compensation for partial work, or peer assistance for contract completion were particularly disadvantaged. Participants recommended the government/stakeholders intervene to ensure: MWs access to bed-nets, food, shelter, water, and healthcare at farms or sick leave; decentralization of diagnostic tests to primary healthcare facilities; surplus medications/staff during the peak season; improved referral/feedback/reporting/training within the health system; free comprehensive healthcare for all VL-related services; and community health education., Conclusions/significance: Contrary to what health policy for VL dictates in this endemic setting, study participants reported very poor access to diagnosis and, consequently, significantly delayed access to treatment. Interventions tailored to the socio-economic and health needs of MWs (and other persons suffering from VL) are urgently needed to reduce health disparities and the VL burden., Competing Interests: The authors have declared that no competing interests exist.

Better tools are necessary to eliminate visceral leishmaniasis (VL). Modeling studies for regional Leishmania elimination indicate that an effective vaccine is a critical tool. Dogs are the reservoir host of L. infantum in Brazil and the Mediterranean basin, and therefore are an important target for public health interventions as well as a relevant disease model for human VL. No vaccine has been efficacious as an immunotherapy to prevent progression of already diagnostically positive individuals to symptomatic leishmaniasis. We performed a double-blinded, block-randomized, placebo-controlled, vaccine immunotherapy trial testing the efficacy of a recombinant Leishmania A2 protein, saponin-adjuvanted, vaccine, LeishTec®, in owned hunting dogs infected with L. infantum. The primary outcome was reduction of clinical progression, with reduction of mortality as a secondary outcome. Vaccination as an immunotherapy reduced the risk of progression to clinically overt leishmaniasis by 25% in asymptomatic dogs (RR: 1.33 95% C.I. 1.009-1.786 p-value: 0.0450). Receiving vaccine vs. placebo reduced all-cause mortality in younger asymptomatic dogs by 70% (RR: 3.19 95% C.I.: 1.185-8.502 p-value = 0.0245). Vaccination of infected-healthy animals with an anti-Leishmania vaccine significantly reduced clinical progression and decreased all-cause mortality. Use of vaccination in infected-healthy dogs can be a tool for Leishmania control., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Visceral leishmaniasis (VL), one of the deadliest parasitic diseases in the world, causes more than 50,000 human deaths each year and afflicts millions of people throughout South America, East Africa, South Asia, and Mediterranean Region. In 2015 the World Health Organization classified VL as a neglected tropical disease (NTD), prompting concentrated study of the VL epidemic using mathematical and simulation models. This paper reviews literature related to prevalence and prevention control strategies. More than thirty current research works were reviewed and classified based on VL epidemic study methods, including modeling approaches, control strategies, and simulation techniques since 2013. A summarization of these technical methods, major findings, and contributions from existing works revealed that VL epidemic research efforts must improve in the areas of validating and verifying VL mathematical models with real-world epidemic data. In addition, more dynamic disease control strategies must be explored and advanced simulation techniques must be used to predict VL pandemics.

Leishmania donovani is a causative pathogen of potentially fatal visceral leishmaniasis (VL). Therapeutic agents are available; however, their use is limited because of high cost, serious side effects, and development of antimicrobial resistance. Protective immunity against VL depends on CD4 + Th1 cell-mediated immunity. Studies have shown that progression of VL is due to exhaustion of T cells; however, the mechanism involved is not clearly understood. Here, we examined the role of PD1/PDL-1 in the pathogenesis of VL by using a murine model of VL. Our data indicate that L. donovani is able to elicit initial expansion of gamma interferon-producing CD4 + Th1 and CD8 + T cells at day 7 postinfection (p.i.); however, the frequency of those cells and inflammatory response decreased at day 21 p.i., despite persistence of parasites. Persistent infection-induced expansion of interleukin-10 + FOXP3 + Treg and CD4 + and CD8 + T cells expressing PD1. Blocking of PDL-1 signaling in vivo resulted in restoration of protective type 1 responses by both CD4 + and CD8 + T cells, which resulted in a significant decrease in the parasite burden. Mechanistically, PDL-1 blocking inhibited autophagy, a cellular degradation process hijacked by Leishmania to acquire host cell nutrients for their survival. Inhibition of autophagy was marked by decreased lipidation of microtubule-associated protein 1 light chain 3, a marker of autophagosome formation, and P62 accumulation. Together, our findings show for the first time that anti-PDL-1 antibody is an effective therapeutic approach for restoration of effector arms of protective immunity against VL and subsequent parasite clearance., (Copyright © 2018 Habib et al.)

Physical contact between dendritic cells (DCs) and T cell lymphocytes is necessary to trigger the immune cell response. CCL19 and CCL21 chemokines bind to the CCR7 receptor of mature DCs, and of T cells and regulate DCs migration to the white pulp (wp) of the spleen, where they encounter lymphocytes. In visceral leishmaniasis (VL), cellular immunosuppression is mediated by impaired DC migration due to the decreased chemokine secretion by endothelium and to the reduced DCs CCR7 expression. The Leishmania (L.) donovani nucleoside hydrolase NH36 and its C-terminal domain, the F3 peptide are prominent antigens in the generation of preventive immunity to VL. We assessed whether these vaccines could prevent the migrating defect of DCs by restoring the expression of CCR7 receptors. C57Bl6 mice were vaccinated with NH36 and F3 and challenged with L. (L.) infantum chagasi . The F3 vaccine induced a 100% of survival and a long-lasting immune protection with an earlier CD4 + Th1 response, with secretion of higher IFN-γ and TNF-α/IL-10 ratios, and higher frequencies of CD4 + T cells secreting IL-2 + , TNF-α + , or IFN-γ + , or a combination of two or the three cytokines (IL-2 + TNF-α + IFN-γ + ). The CD8 + T cell response was promoted earlier by the NH36-vaccine, and later by the F3-vaccine. Maximal number of F3-primed DCs migrated in vitro in response to CCL19 and showed a high expression of CCR7 receptors (26.06%). Anti-CCR7 antibody treatment inhibited DCs migration in vitro (90%) and increased parasite load in vivo . When transferred into 28-day-infected mice, only 8% of DCs from infected, 59% of DCs from NH36-vaccinated, and 84% of DCs from F3-vaccinated mice migrated to the wp. Consequently, immunotherapy of infected mice with F3-primed DCs only, promoted increases in corporal weight and reductions of spleen and liver parasite loads and relative weights. Our findings indicate that vaccination with F3-vaccine preserves the maturation, migration properties and CCR7 expression of DCs, which are essential processes for the generation of cell-mediated immunity. The F3 vaccine is more potent in reversing the migration defect that occurs in VL and, therefore, more efficient in immunotherapy of VL.

Early diagnosis and treatment is the principal strategy to control visceral leishmaniasis (VL), or kala-azar in East Africa. As VL strikes remote rural, sparsely populated areas, kala-azar care might not be accessed optimally or timely. We conducted a qualitative study to explore access barriers in a longstanding kala-azar endemic area in southern Gadarif, Sudan. Former kala-azar patients or caretakers, community leaders, and health-care providers were purposively sampled and thematic data analysis was used. Our study participants revealed the multitude of difficulties faced when seeking care. The disease is well known in the area, yet misconceptions about causes and transmission persist. The care-seeking itineraries were not always straightforward: "shopping around" for treatments are common, partly linked to difficulties in diagnosing kala-azar. Kala-azar is perceived to be "hiding," requiring multiple tests and other diseases must be treated first. Negative perceptions on quality of care in the public hospitals prevail, with the unavailability of drugs or staff as the main concern. Delay to seek care remains predominantly linked to economic constraint: albeit treatment is for free, patients have to pay out of pocket for everything else, pushing families further into poverty. Despite increased efforts to tackle the disease over the years, access to quality kala-azar care in this rural Sudanese context remains problematic. The barriers explored in this study are a compelling reminder of the need to boost efforts to address these barriers.

Visceral leishmaniasis (VL) is a potentially fatal disease, in which the treatment based on chemotherapy is considered toxic. The cure of disease is associated with the life-long Th1-type immunity against the infection. The Th1-related cytokines production by peripheral blood mononuclear cells (PBMCs) seems to be crucial for host control of parasite load and clinical cure. In the current study, we used five proteins (IgE-dependent histamine-releasing factor [HRF], LiHyD, LiHyV, LiHyT and LiHyp6) recently shown to be antigenic and/or immunogenic in the canine VL, aiming to evaluate the antigen-specific antibody levels and cytokine production in PBMCs culture supernatants collected from VL patients before and after anti-VL treatment. In the results, when PBMCs were exposed to rHRF, rLiHyD and rLiHyT, higher IFN-γ and lower IL-10 levels were observed in all patients that were treated and clinically cured. Analysis of specific antibody subclasses was in line with in vitro cellular response, since a higher IgG2 production was found in the treated and cured patients, when compared to the IgG1 subclass levels. In addition, evaluating the diagnostic efficacy of the recombinant molecules, the rHRF, rLiHyD and rLiHyT proteins showed the best results in the serology assays to identify all VL patients, as well as these antigens were not recognized by antibodies in sera from non-infected subjects or those with leishmaniasis-related diseases. Our results corroborate the view that clinical cure of VL is associated with a sustained Th1-related response, and indicate the potential use of rHRF, rLiHyD and rLiHyT as immune biomarkers of VL treatment., (Copyright © 2017 Elsevier GmbH. All rights reserved.)

Objective: To estimate the Brazilian direct and indirect costs of human visceral leishmaniasis (VL) in 2014., Methods: Cost-of-illness study on the Brazilian public health system and societal perspective. VL cases registered in the Notifiable Diseases Information System in the year of 2014 were considered. Direct medical costs regarding diagnostic, treatment and care provided to patients with VL were estimated through the top-down approach. The indirect costs related to productivity loss due to premature mortality and morbidity were estimated by means of the human-capital method., Results: In 2014, 9895 suspected cases of VL were reported in the Notifiable Diseases Information System, and 3453 were later confirmed. There were 234 patients with Leishmania-HIV coinfection underwent a secondary prophylaxis. The total cost of VL in Brazil was US$ 14 190 701.50 (US$ 14 189 150.10 to 14 199 940.53) that varied according to the sensitivity analysis. The total of direct medical costs corresponded to US$ 1 873 681.96 (US$1 872 130.55 to 1 882 920.99), and the majority of costs was associated with hospitalisation (40%), followed by treatment (22%), and secondary prophylaxis (18%). Productivity loss corresponded to US$ 11 421 683.37 for premature mortality and US$ 895 336.18 for work absence due to hospitalisation by the illness., Conclusions: VL represents an expensive health problem for the Brazilian public health system and society, mainly because of its productivity loss due to premature mortality. Interventions to reduce VL lethality could have a great impact on decreasing the cost of illness., (© 2017 John Wiley & Sons Ltd.)

Leishmania donovani causes cutaneous leishmaniasis (CL) in Sri Lanka. Standard treatment is multiple, painful doses of intralesional sodium stibogluconate (IL-SSG). Treatment failures are increasingly reported, hence the need to investigate alternatives. Efficacy, safety, and cost-effectiveness of thermotherapy were assessed for the first time for L . donovani CL. A single blinded noninferiority randomized controlled trial was conducted on new laboratory-confirmed CL patients with single lesions ( N = 213). Selected patients were randomly assigned to 1) test group ( N = 98; single session of radiofrequency-induced heat therapy (RFHT) given at 50°C for 30 seconds) and 2) control group ( N = 115; 1-3 mL IL-SSG given weekly, until cure/10 doses). Patients were followed-up fortnightly for 12 weeks to assess clinical cure. Cost of treatment was assessed using scenario building technique. Cure rates by 8, 10, and 12 weeks in RFHT group were 46.5%, 56.5%, and 65.9% as opposed to 28%, 40.8%, and 59.4% in IL-SSG group, with no major adverse events. Cure rate by RFHT was significantly higher at 8 weeks ( P = 0.009, odds ratio [OR]: 2.236, confidence interval [CI]: 1.217-4.108) and 10 weeks ( P = 0.035, OR: 1.881, CI: 1.044-3.388), but comparable thereafter. Cost of RFHT was 7 times less (USD = 1.54/patient) than IL-SSG (USD = 11.09/patient). A single application of RFHT is safe, cost-effective, and convenient, compared with multiple doses of IL-SSG in the treatment of L. donovani CL. Therefore, RFHT would be considered noninferior as per trial outcome when compared with standard IL-SSG therapy with multiple benefits for the patient and the national health care system.

Leishmaniasis is a disease caused by obligate intracellular parasites of the genus Leishmania. There are three main clinical forms of leishmaniasis: cutaneous, visceral and mucocutaneous. It belongs to the diverse group of the tropical “neglected diseases” and is the only tropical vector-borne disease that has remained endemic to southern Europe for decades. Visceral leishmaniasis represents the most serious clinical syndrome. It is characterized by fever, weight loss, anemia and enlargement of the spleen and liver. Incubation period is usually from 2 weeks to 18 months. Leishmaniasis is considered a worldwide reemerging disease. Some of the factors that favor this situation are the changes in the climatic conditions, migrations and urbanizations deficient in environmental sanitation. We report an interesting case of an European child who was spending holidays in Cordoba and presented to our hospital with fever and pancytopenia, motivating a multidisciplinary approach with favorable clinical outcome., (Sociedad Argentina de Pediatría.)

The prevalence of Th1/Th2 response, spleen changes and megakaryocytes were investigated in BALB/c mice (n=138) infected with Leishmania infantum, and treated with Leishmania infantum 30× (10 -30 ) biotherapy - BioLi30×. We performed controlled experiments using 8-to-12-week-old mice, infected with 5×10 7 L. infantum promastigotes, divided into eight groups: G1 (healthy), G2 (infected with L. infantum), G3 (BioLi30× pre-treated), G4 (BioLi30× pre/post-treated), G5 (BioLi30× post-treated), G6 (Water 30× post-treated), G7 (Antimonium crudum 30× post-treated) and G8 (Glucantime® post-treated). G3-G7 groups were orally treated with their respective drugs diluted in filtered water (1:10), and G8 received Glucantime® (0.6mg/100µl of PBS), intraperitoneally. Spleen fragments were submitted to double blind histopathological evaluation and the number of megakaryocytes was counted. Besides, animals' serum was measured after 49days of infection, and cytokines (IFN-γ, IL-4, IL-10, IL-12), as well as the Th1/Th2 correlation (IFN-γ/IL-4 and IFN-γ/IL-10), were analyzed. Spleen histological parameters were classified as: healthy appearance (G1); discreet (G3-G7), moderate (G2) and moderate to severe (G8) white pulp hyperplasia; proliferation of megakaryocytes (G2-G8), and intense disruption (G2-G8). All groups, except for G7, showed higher percentages of megakaryocytes per field ranging from 87% to 15%, when compared to healthy animals (G1). Th1 predominance in IFN-γ/IL-4 ratio (comparing to G2) was detected in G4, G5, G6 and G7. Finally, pre/post (BioLi30x) and post-treatment (Antimonium crudum 30x) presented reduction of megakaryocytes/spleen changes due to immunomodulation animal process, controlling the infection process, probably by the Th1 cytokine predominance., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Leishmaniases are vector-borne diseases for which no vaccine exists. These diseases are caused by the Leishmania species complex. Activation of the CD8 + T cell is crucial for protection against intracellular pathogens, and peptide antigens are attractive strategies for the precise activation of CD8 + T in vaccine development against intracellular infections. The traditional approach to mine the epitopes is an arduous task. However, with the advent of immunoinformatics, in silico epitope prediction tools are available to expedite epitope identification. In this study, we employ different immunoinformatics tools to predict CD8 + T cell specific 9 mer epitopes presented by HLA-A*02 and HLA-B40 within the highly conserved 3'-ectonucleotidase of Leishmania donovani. We identify five promiscuous epitopes, which have no homologs in humans, theoretically cover 85% of the world's population and are highly conserved (100%) among Leishmania species. Presentation of selected peptides was confirmed by T2 cell line based HLA-stabilization assay, and three of them were found to be strong binders. The in vitro peptide stimulation of peripheral blood mononuclear cells (PBMC) from cured HLA-A02 + visceral leishmaniasis (VL) subjects produced significantly higher IFN-γ, IL-2 and IL-12 compared to no peptide control healthy subjects. Further, CD8 + cells from treated VL subjects produced significantly higher intracellular IFN-γ, lymphocyte proliferation and cytotoxic activity against selected peptides from the PBMCs of treated HLA-A02 + VL subjects. Thus, the CD8 + T cell specific epitopes shown in this study will speed up the development of polytope vaccines for leishmaniasis., (Copyright © 2017. Published by Elsevier Masson SAS.)

New biomarkers are needed for monitoring the effectiveness of treatment for visceral leishmaniasis (VL). They might also improve the detection of the asymptomatic population in Leishmania- endemic areas. This paper examines the IL-2, IFN-γ, IFN-γ-induced protein 10 (IP-10), and monokine-induced-by-IFN-γ (MIG) levels in whole blood-stimulated in vitro with soluble Leishmania antigen (SLA)-taken from asymptomatic individuals and patients treated for VL living in a post-outbreak ( Leishmania infantum ) area in Spain, and in an endemic ( Leishmania donovani ) area of Bangladesh. IP-10 was found to be an accurate global marker of asymptomatic subjects with positive cellular/humoral tests, while MIG was found to be a better marker of contact with L. donovani than IL-2 but no for those with L. infantum . Determining IP-10, MIG, and IFN-γ levels proved useful in monitoring the cellular immune response following treatment for active disease caused by L. infantum .

Background: Visceral leishmaniasis (VL or kala azar) is the most serious form of human leishmaniasis, responsible for over 20,000 deaths annually, and post kala azar dermal leishmaniasis (PKDL) is a stigmatizing skin condition that often occurs in patients after successful treatment for VL. Lack of effective or appropriately targeted cell mediated immunity, including CD8+ T cell responses, underlies the progression of VL and progression to PKDL, and can limit the therapeutic efficacy of anti-leishmanial drugs. Hence, in addition to the need for prophylactic vaccines against leishmaniasis, the development of therapeutic vaccines for use alone or in combined immuno-chemotherapy has been identified as an unmet clinical need. Here, we report the first clinical trial of a third-generation leishmaniasis vaccine, developed intentionally to induce Leishmania-specific CD8+ T cells., Methods: We conducted a first-in-human dose escalation Phase I trial in 20 healthy volunteers to assess the safety, tolerability and immunogenicity of a prime-only adenoviral vaccine for human VL and PKDL. ChAd63-KH is a replication defective simian adenovirus expressing a novel synthetic gene (KH) encoding two Leishmania proteins KMP-11 and HASPB. Uniquely, the latter was engineered to reflect repeat domain polymorphisms and arrangements identified from clinical isolates. We monitored innate immune responses by whole blood RNA-Seq and antigen specific CD8+ T cell responses by IFNγ ELISPOT and intracellular flow cytometry., Findings: ChAd63-KH was safe at intramuscular doses of 1x1010 and 7.5x1010 vp. Whole blood transcriptomic profiling indicated that ChAd63-KH induced innate immune responses characterized by an interferon signature and the presence of activated dendritic cells. Broad and quantitatively robust CD8+ T cell responses were induced by vaccination in 100% (20/20) of vaccinated subjects., Conclusion: The results of this study support the further development of ChAd63-KH as a novel third generation vaccine for VL and PKDL., Trial Registration: This clinical trial (LEISH1) was registered at EudraCT (2012-005596-14) and ISRCTN (07766359).

This tutorial promotes good practice for exploring the rationale of systems pharmacology models. A safety systems engineering inspired notation approach provides much needed rigor and transparency in development and application of models for therapeutic discovery and design of intervention strategies. Structured arguments over a model's development, underpinning biological knowledge, and analyses of model behaviors are constructed to determine the confidence that a model is fit for the purpose for which it will be applied., (© 2016 The Authors CPT: Pharmacometrics & Systems Pharmacology published by Wiley Periodicals, Inc. on behalf of American Society for Clinical Pharmacology and Therapeutics.)

The recent adaptation of CRISPR Cas9 genome editing to Leishmania spp. has opened a new era in deciphering Leishmania biology. The method was recently improved using a PCR-based CRISPR Cas9 approach, which eliminated the need for cloning. This new approach, which allows high-throughput gene deletion, was successfully validated in L. mexicana and L. major . In this study, we validated the toolkit in Leishmania donovani targeting the flagellar protein PF16, confirming that the tagged protein localizes to the flagellum and that null mutants lose their motility. We then used the technique to characterise CK1.1, a member of the casein kinase 1 family, which is involved in the regulation of many cellular processes. We showed that CK1.1 is a low-abundance protein present in promastigotes and in amastigotes. We demonstrated that CK1.1 is not essential for promastigote and axenic amastigote survival or for axenic amastigote differentiation, although it may have a role during stationary phase. Altogether, our data validate the use of PCR-based CRISPR Cas9 toolkit in L. donovani , which will be crucial for genetic modification of hamster-derived, disease-relevant parasites.

The present works reviewed the existing information on leishmaniasis in Sri Lanka and in other countries, focusing on challenges of controlling leishmaniasis in the country, in an outbreak. Evidence from recent studies suggests that there is a possibility of a leishmaniasis outbreak in Sri Lanka in the near future. Difficulty of early diagnosis due to lack of awareness and unavailability or inadequacy of sensitive tests are two of the main challenges for effective case management. Furthermore, the absence of a proper drug for treatment and lack of knowledge about vector biology, distribution, taxonomy and bionomics, and reservoir hosts make the problem serious. The evident potential for visceralization in the cutaneous variant of L . donovani in Sri Lanka may also complicate the issue. Lack of knowledge among local communities also reduces the effectiveness of vector and reservoir host control programs. Immediate actions need to be taken in order to increase scientific knowledge about the disease and a higher effectiveness of the patient management and control programs must be achieved through increased awareness about the disease among general public and active participation of local community in control activities.

Objectives: Poor treatment outcomes of visceral leishmaniasis (VL) are responsible for the high mortality rate of this condition in resource-limited settings such as Ethiopia. This study aimed to identify the proportion of poor VL treatment outcomes in northwest Ethiopia and to evaluate the determinants associated with poor outcomes., Methods: A hospital-based retrospective study was conducted among 595 VL patients who were admitted to Kahsay Abera Hospital in northwest Ethiopia from October 2010 to April 2013. Data were entered into Epi Info version 7.0 and exported to SPSS version 20 for analysis. Bivariate and multivariate logistic regression models were fitted to identify the determinants of VL treatment outcomes. Adjusted odds ratio (aORs) with 95% confidence intervals (CIs) were used, and p -values <0.05 were considered to indicate statistical significance., Results: The proportion of poor treatment outcomes was 23.7%. Late diagnosis (≥29 days) (aOR, 4.34; 95% CI, 2.22 to 8.46), severe illness at admission (inability to walk) (aOR, 1.63; 95% CI, 1.06 to 2.40) and coinfection with VL and human immunodeficiency virus (HIV) (aOR, 2.72; 95% CI, 1.40 to 5.20) were found to be determinants of poor VL treatment outcomes., Conclusions: Poor treatment outcomes, such as death, treatment failure, and non-adherence, were found to be common. Special attention must be paid to severely ill and VL/HIV-coinfected patients. To improve VL treatment outcomes, the early diagnosis and treatment of VL patients is recommended.

Background: To describe an unusual clinical presentation of visceral leishmaniasis affecting the colon., Case Report: We report the case of an HIV-positive patient with visceral leishmaniasis. We describe the clinical case, the procedures performed, the treatment provided and the patient's evolution. A comparative table of previously reported similar cases is shown., Discussion: Visceral leishmaniasis with intestinal involvement is an uncommon process. Nevertheless, this possibility should be taken into consideration in the differential diagnosis of immunosuppressed patients with symptoms of diarrhea, as a favorable prognosis depends on early diagnosis and appropriate treatment.

Travel to developing countries is associated with possibility to acquire different, also parasitic, diseases which have become a diagnostic and therapeutic problem. The ENT specialist is one of the medical officers who may make initial contact with a patient with a suspected parasitic disease. One of them is leishmaniosis: a parasitic disease classified as Neglected Tropical Diseases (NTD), which occurs in the tropics, subtropics and southern Europe. While asymptomatic invasion is observed in some cases, three forms of symptomatic leishmaniosis can be distinguished: cutaneous leishmaniosis (CL), visceral leishmaniosis (VL), and mucosal leishmaniosis (ML). Signs and symptoms of these forms, as well as the differential diagnosis, diagnostic procedures and treatment are presented in the paper.

Background: The therapy of visceral leishmaniasis (VL) is limited by resistance, toxicity and decreased bioavailability of the existing drugs coupled with dramatic increase in HIV-co-infection, non-availability of vaccines and down regulation of cell-mediated immunity (CMI). Thus, we envisaged combating the problem with plant-derived antileishmanial drug that could concomitantly mitigate the immune suppression of the infected hosts. Several plant-derived compounds have been found to exert leishmanicidal activity via immunomodulation. In this direction, we investigated the antileishmanial activity of eugenol emulsion (EE), complemented with its immunomodulatory and therapeutic efficacy in murine model of VL., Methodology/principal Findings: Oil-in-water emulsion of eugenol (EE) was prepared and size measured by dynamic light scattering (DLS). EE exhibited significant leishmanicidal activity with 50% inhibitory concentration of 8.43±0.96 μg ml-1 and 5.05±1.72 μg ml─1, respectively against the promastigotes and intracellular amastigotes of Leishmania donovani. For in vivo effectiveness, EE was administered intraperitoneally (25, 50 and 75 mg/kg b.w./day for 10 days) to 8 week-infected BALB/c mice. The cytotoxicity of EE was assessed in RAW 264.7 macrophages as well as in naive mice. EE induced a significant drop in hepatic and splenic parasite burdens as well as diminution in spleen and liver weights 10 days post-treatment, with augmentation of 24h-delayed type hypersensitivity (DTH) response and high IgG2a:IgG1, mirroring induction of CMI. Enhanced IFN-γ and IL-2 levels, with fall in disease-associated Th2 cytokines (IL-4 and IL-10) detected by flow cytometric bead-based array, substantiated the Th1 immune signature. Lymphoproliferation and nitric oxide release were significantly elevated upon antigen revoke in vitro. The immune-stimulatory activity of EE was further corroborated by expansion of IFN-γ producing CD4+ and CD8+ splenic T lymphocytes and up-regulation of CD80 and CD86 on peritoneal macrophages. EE treated groups exhibited induction of CD8+ central memory T cells as evidenced from CD62L and CD44 expression. No biochemical alterations in hepatic and renal enzymes were observed., Conclusions: Our results demonstrate antileishmanial activity of EE, potentiated by Th1 immunostimulation without adverse side effects. The Th1 immune polarizing effect may help to alleviate the depressed CMI and hence complement the leishmanicidal activity., Competing Interests: The authors have declared that no competing interests exist.

Visceral leishmaniasis is a neglected tropical disease endemic in East Africa where improved patient-adapted treatments are needed. The Leishmaniasis East Africa Platform (LEAP) was created in 2003 to strengthen clinical research capacity, serve as a base for training, and evaluate and facilitate implementation of new treatments. Major infrastructure upgrades and personnel training have been carried out. A short course of Sodium Stibogluconate and Paramomycin (SSG&PM) was evaluated and is now first-line treatment in the region; alternative treatments have also been assessed. LEAP can serve as a successful model of collaboration between different partners and countries when conducting clinical research in endemic countries to international standards., (© The Author 2016. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene.)

Background: Previously we reported that Kala-azar patients show progressive decrease in serum cholesterol as a function of splenic parasite burden. Splenic macrophages (MΦ) of Leishmania donovani (LD) infected mice show decrease in membrane cholesterol, while LD infected macrophages (I-MΦ) show defective T cell stimulating ability that could be corrected by liposomal delivery of cholesterol. T helper cells recognize peptide antigen in the context of class II MHC molecule. It is known that the conformation of a large number of membrane proteins is dependent on membrane cholesterol. In this investigation we tried to understand the influence of decreased membrane cholesterol in I-MΦ on the conformation of MHC-II protein and peptide-MHC-II stability, and its bearing on the antigen specific T-cell activation., Methodology/principal Findings: MΦ of CBA/j mice were infected with Leishmania donovani (I-MΦ). Two different anti-Aκ mAbs were used to monitor the status of MHC-II protein under parasitized condition. One of them (11.5-2) was conformation specific, whereas the other one (10.2.16) was not. Under parasitized condition, the binding of 11.5-2 decreased significantly with respect to the normal counterpart, whereas that of 10.2.16 remained unaltered. The binding of 11.5-2 was restored to normal upon liposomal delivery of cholesterol in I-MΦ. By molecular dynamics (MD) simulation studies we found that there was considerable conformational fluctuation in the transmembrane domain of the MHC-II protein in the presence of membrane cholesterol than in its absence, which possibly influenced the distal peptide binding groove. This was evident from the faster dissociation of the cognate peptide from peptide-MHC complex under parasitized condition, which could be corrected by liposomal delivery of cholesterol in I-MΦ., Conclusion: The decrease in membrane cholesterol in I-MΦ may lead to altered conformation of MHC II, and this may contribute to a faster dissociation of the peptide. Furthermore, liposomal delivery of cholesterol in I-MΦ restored its normal antigen presenting function. This observation brings strength to our previous observation on host directed therapeutic application of liposomal cholesterol in experimental visceral leishmaniasis.

Chronic disease caused by infections, cancer or autoimmunity can result in profound immune suppression. Immunoregulatory networks are established to prevent tissue damage caused by inflammation. Although these immune checkpoints preserve tissue function, they allow pathogens and tumors to persist, and even expand. Immune checkpoint blockade has recently been successfully employed to treat cancer. This strategy modulates immunoregulatory mechanisms to allow host immune cells to kill or control tumors. However, the utility of this approach for controlling established infections has not been extensively investigated. Here, we examined the potential of modulating glucocorticoid-induced TNF receptor-related protein (GITR) on T cells to improve anti-parasitic immunity in blood and spleen tissue from visceral leishmaniasis (VL) patients infected with Leishmania donovani. We found little effect on parasite growth or parasite-specific IFNγ production. However, this treatment reversed the improved anti-parasitic immunity achieved by IL-10 signaling blockade. Further investigations using an experimental VL model caused by infection of C57BL/6 mice with L. donovani revealed that this negative effect was prominent in the liver, dependent on parasite burden and associated with an accumulation of Th1 cells expressing high levels of KLRG-1. Nevertheless, combined anti-IL-10 and anti-GITR mAb treatment could improve anti-parasitic immunity when used with sub-optimal doses of anti-parasitic drug. However, additional studies with VL patient samples indicated that targeting GITR had no overall benefit over IL-10 signaling blockade alone at improving anti-parasitic immune responses, even with drug treatment cover. These findings identify several important factors that influence the effectiveness of immune modulation, including parasite burden, target tissue and the use of anti-parasitic drug. Critically, these results also highlight potential negative effects of combining different immune modulation strategies.

The chemotherapy of visceral leishmaniasis (VL) has several limitations including resistance and toxicity of the existing drugs. Down regulation of immune system further aggravates the problems. To combat this situation we evaluated the leishmanicidal efficacy of Boerhaavia diffusa and Ocimum sanctum through oral route in L. donovani infection in BALB/c mice. Results have demonstrated maximum clearance of the parasites from infected animals treated with combination of B. diffusa and O. sanctum (@ 100 and 400 mg/kg body wt., respectively 5 days) as depicted through Leishman Donovan Units in liver. Up-regulation of cell-mediated immunity was also observed in animals of this group as heightened delayed type hypersensitivity responses and increased IgG2a levels were observed. Moreover, increased levels of SGOT, SGPT, serum urea, blood urea nitrogen and serum creatinine were brought down to normal levels. Since VL is associated immunosuppression, the above treatment is a good option as it helps in the up-regulation of Th1 responses and reduction in parasite load in L. donovani infected mice. These findings suggest a new option for antileishmanial chemotherapy at lower cost and nil toxicity.

Visceral leishmaniasis (VL) is a severe and potentially fatal disease caused by different Leishmania species, Leishmania chagasi prevailing in Brazil. Main symptoms include fever, malaise, anorexia, weight loss and abdominal enlargement with typically occurring hepatosplenomegaly Currently, VL is considered an opportunistic infection in immunocompromised hosts, including solid organ transplanted patients. The present study reports a case of VL associated to pregnancy after renal transplantation.

Background: To eliminate visceral leishmaniasis (VL) in India and Nepal, challenges of VL diagnosis, treatment and reporting need to be identified. Recent data indicate that VL is underreported and patients face delays when seeking treatment. Moreover, VL surveillance data might not reach health authorities on time. This study quantifies delays for VL diagnosis and treatment, and analyses the duration of VL reporting from district to central health authorities in India and Nepal., Methods: A cross-sectional study conducted in 12 districts of Terai region, Nepal, and 9 districts of Bihar State, India, in 2012. Patients were interviewed in hospitals or at home using a structured questionnaire, health managers were interviewed at their work place using a semi-structured questionnaire and in-depth interviews were conducted with central level health managers. Reporting formats were evaluated. Data was analyzed using two-tailed Mann-Whitney U or Fisher's exact test., Results: 92 VL patients having experienced 103 VL episodes and 49 district health managers were interviewed. Patients waited in Nepal 30 days (CI 18-42) before seeking health care, 3.75 times longer than in Bihar (8d; CI 4-12). Conversely, the lag time from seeking health care to receiving a VL diagnosis was 3.6x longer in Bihar (90d; CI 68-113) compared to Nepal (25d; CI 13-38). The time span between diagnosis and treatment was short in both countries. VL reporting time was in Nepal 19 days for sentinel sites and 76 days for "District Public Health Offices (DPHOs)". In Bihar it was 28 days for "District Malaria Offices". In Nepal, 73% of health managers entered data into computers compared to 16% in Bihar. In both countries reporting was mainly paper based and standardized formats were rarely used., Conclusions: To decrease the delay between onset of symptoms and getting a proper diagnosis and treatment the approaches in the two countries vary: In Nepal health education for seeking early treatment are needed while in Bihar the use of private and non-formal practitioners has to be discouraged. Reinforcement of VL sentinel reporting in Bihar, reorganization of DPHOs in Nepal, introduction of standardized reporting formats and electronic reporting should be conducted in both countries.