Combining Cationic Liposomal Delivery with MPL-TDM for Cysteine Protease Cocktail Vaccination against Leishmania donovani : Evidence for Antigen Synergy and Protection

BackgroundWith the paucity of new drugs and HIV co-infection, vaccination remains an unmet research priority to combat visceral leishmaniasis (VL) requiring strong cellular immunity. Protein vaccination often suffers from low immunogenicity and poor generation of memory T cells for long-lasting protection. Cysteine proteases (CPs) are immunogenic proteins and key mediators of cellular functions in Leishmania. Here, we evaluated the vaccine efficacies of CPs against VL, using cationic liposomes with Toll like receptor agonists for stimulating host immunity against L. donovani in a hamster model.Conclusion/SignificanceThe present study is the first report of a comparative efficacy of leishmanial CPs and their cocktail using liposomal formulation with MPL-TDM against L. donovani. The level of protection attained has not been reported for any other subcutaneous single or polyprotein vaccination against VL.     

Leishmania Specific CD4 T Cells Release IFNγ That Limits Parasite Replication in Patients with Visceral Leishmaniasis

Visceral leishmaniasis (VL) is associated with increased circulating levels of multiple pro-inflammatory cytokines and chemokines, including IL-12, IFNγ, and TNFα, and elevated expression of IFNγ mRNA in lesional tissue such as the spleen and bone marrow. However, an immunological feature of VL patients is that their peripheral blood mononuclear cells (PBMCs) typically fail to respond to stimulation with leishmanial antigen. Unexpectedly, it was recently shown that Leishmania specific IFNγ, can readily be detected when a whole blood stimulation assay (WBA) is used. We sought to define the conditions that permit whole blood cells to respond to antigen stimulation, and clarify the biological role of the IFNγ found to be released by cells from VL patients. CD4+ T cells were found to be crucial for and the main source of the IFNγ production in Leishmania stimulated whole blood (WB) cultures. Complement, antibodies and red blood cells present in whole blood do not play a significant role in the IFNγ response. The IFNγ production was reduced by blockade of human leukocyte antigen (HLA)-DR, indicating that the response to leishmanial antigens observed in WB of active VL patients is a classical HLA- T cell receptor (TCR) driven reaction. Most importantly, blockade of IFNγ in ex-vivo splenic aspirate cultures demonstrated that despite the progressive nature of their disease, the endogenous IFNγ produced in patients with active VL serves to limit parasite growth.     

Host peroxisomal properties are not restored to normal after treatment of visceral leishmaniasis with sodium antimony gluconate

Reason for post-kala-azar dermal leishmaniasis (PKDL) is yet to be established. Earlier it was observed that morphology and biochemical properties of host peroxisomes were impaired during Leishmania infection. As peroxisome is known to be involved in various metabolic pathways to monitor normal function of the host cells, it is essential that Leishmania-induced dysfunction of this organelle should totally be repaired during treatment of visceral leishmaniasis (VL). In this paper it has been shown that resumption of normal peroxisomal function could not be attained when one of the existing drugs sodium antimony gluconate (SAG) was used for chemotherapy against VL. Although Leishmania parasite was found to be completely eliminated from host liver and spleen after SAG treatment, normal activities of peroxisomal catalase and superoxide dismutase could not be restored. Also unusual peptides were found to be present due to abnormal proteolytic cleavage of proteins. It is proposed that peroxisomal disorder which exists even after successful chemotherapy of VL may be figured out as one of the possible reasons to develop PKDL. It may also be pointed out that continued effect of peroxisomal disorder even after complete treatment of this parasitic disease may also lead to genetic disorders not yet been explored in post-kala-azar patients.     

Cytokine Release Assays as Tests for Exposure to Leishmania,and for Confirming Cure from Leishmaniasis,in Solid Organ Transplant Recipients

Spain has one of the world’s largest pools of organ donors and is a global leader in terms of the number of transplants it performs. The current outbreak of leishmaniasis in Fuenlabrada (in the southwest of the region of Madrid, Spain) has involved 600 clinical cases since late 2009 (prevalence 0.2%). It may therefore be wise to monitor the town’s transplanted population for Leishmania infantum; its members are immunosuppressed and at greater risk of infection and relapse following treatment. The present work examines the use of cytokine release assays to determine the prevalence of Leishmania infection in this population, and to confirm recovery following treatment for visceral leishmaniasis (VL). The humoral and cellular immune responses to L. infantum were characterized in 63 solid organ transplant (SOT) recipients from Fuenlabrada, 57 of whom reported no previous episode of VL (NVL subjects), and six of whom had been cured of VL (CVL subjects). Seventeen subjects (12 NVL and 5 CVL) showed a patent lymphoproliferative response to soluble Leishmania antigen (SLA). Stimulation of peripheral blood mononuclear cell cultures and of whole blood with SLA led to the production of different combinations of cytokines that might serve to confirm Leishmania infection or recovery from VL and help prevent cured patients from relapsing into this serious condition.     

Species-Specific Antimonial Sensitivity in Leishmania Is Driven by Post-Transcriptional Regulation of AQP1

Leishmania is a digenetic protozoan parasite causing leishmaniasis in humans. The different clinical forms of leishmaniasis are caused by more than twenty species of Leishmania that are transmitted by nearly thirty species of phlebotomine sand flies. Pentavalent antimonials (such as Pentostam or Glucantime) are the first line drugs for treating leishmaniasis. Recent studies suggest that pentavalent antimony (Sb(V)) acts as a pro-drug, which is converted to the more active trivalent form (Sb(III)). However, sensitivity to trivalent antimony varies among different Leishmania species. In general, Leishmania species causing cutaneous leishmaniasis (CL) are more sensitive to Sb(III) than the species responsible for visceral leishmaniasis (VL). Leishmania aquaglyceroporin (AQP1) facilitates the adventitious passage of antimonite down a concentration gradient. In this study, we show that Leishmania species causing CL accumulate more antimonite, and therefore exhibit higher sensitivity to antimonials, than the species responsible for VL. This species-specific differential sensitivity to antimonite is directly proportional to the expression levels of AQP1 mRNA. We show that the stability of AQP1 mRNA in different Leishmania species is regulated by their respective 3’-untranslated regions. The differential regulation of AQP1 mRNA explains the distinct antimonial sensitivity of each species.     

Leishmania donovani vs immunity: T-cells sensitized from Leishmania of one donor may modulate their cytokines pattern on re-stimulation with Leishmania from different donor in visceral leishmaniasis

Antimony resistance is frequently encountered during treatment of visceral leishmaniasis (VL) and the differences are well characterized by inadequate IFN-γ dominant type-1 protection mechanisms. The part played by Leishmania parasites derived from antimony treated patients in the outcome of an immune response largely remains to be investigated. In the present study we observed that macrophages of BALB/c mice infected with antimony non-responder (SAG-NR) isolates had a greater amastigote burden than antimony responder (SAG-R) isolates. Later it was observed that antigen from SAG-NR and R L. donovani isolates elicit different cytokine responses in peritoneal blood mononuclear cells (PBMCs) from patients with VL. The production of IFN-γ by T-cells in VL patients increased in response to Leishmania derived from responder patients but this response within same T-cells was lower when sensitized from Leishmania from a non-responder VL patient. On the other hand, IL-4 and IL-10 expression was increased when primed with parasites from non-responder VL source. Such a differential pattern of cytokine expression by the same T-cell population produced to Leishmania from different donors, needs further exploration.     

Development of an Immunochromatographic Test for Diagnosis of Visceral Leishmaniasis Based on Detection of a Circulating Antigen

BackgroundVisceral leishmaniasis (VL) is a life-threatening disease caused by protozoan parasites of the Leishmania donovani complex. Early case detection followed by adequate treatment is essential to the control of VL. However, the available diagnostic tests are either invasive and require considerable expertise (parasitological demonstration of the parasite in tissue smears) or unable to distinguish between past and active infection (serological methods). Therefore, we aimed to develop a lateral flow assay in the form of an immunochromatographic test (ICT) device based on the detection of a circulating Leishmania antigen using monoclonal antibodies (mAbs).Conclusion/SignificanceThe newly developed ICT is an easy to use and more accurate diagnostic tool which fulfils the performance and operational characteristics required for VL case detection under field and laboratory conditions. As our ICT detects a circulating antigen, it will also be useful in monitoring treatment success and diagnosing VL in immunocompromised patients.     

Comparative in vivo expression of amastigote up regulated Leishmania genes in three different forms of Leishmaniasis

In Old World Leishmania infections in India, Leishmania donovani is responsible for visceral leishmaniasis (VL) and post kala-azar dermal leishmaniasis (PKDL) while L. tropica is responsible for cutaneous leishmaniasis (CL) in humans. The molecular differences between the two species of Leishmania and within the same species causing distinct pathologies that govern the outcome of infection and pathogenesis in the human host are unknown. Quantitative expression of selected genes was evaluated directly in lesion tissues of VL, PKDL and CL patients. Assessment of in vivo mRNA level highlighted substantial differences in gene expression patterns, providing an indication of the genes involved in pathogenesis in the three different forms of Leishmaniasis.     

Toll-like receptor signaling: a perspective to develop vaccine against leishmaniasis

The toll-like receptors (TLRs) are the sentinel factor of the innate immunity, which are essential for host defense. These receptors detect the presence of conserved molecular patterns of potentially pathogenic microorganisms and contribute in both, cellular as well as humoral immune responses. Leishmania is an intracellular pathogen that silently invades host immune system. After phagocytosis, it divides and proliferates in the harmful environment of host macrophages by down-regulating its vital effector functions. In leishmaniasis, the outcome of the infection basically relies on the skewed balance between Th1/Th2 immune responses. Lots of work have been done and on progress but still characterization of either preventive or prophylactic candidate antigen/s is far from satisfactory. How does Leishmania regulate host innate immune system? Still it is unanswered. TLRs play very important role during inflammatory process of various diseases such as cancer, bacterial and viral infections but TLR signaling is comparatively less explained in leishmanial infection. In the context to Th1/Th2 dichotomy, identification of leishmanial antigens that modulate toll-like receptor signaling will certainly help in the development of future vaccine. This review will initially describe global properties of TLRs, and later will discuss their role in the pathogenesis of leishmaniasis.     

Leishmaniasis: A new method for confirming cure and detecting asymptomatic infection in patients receiving immunosuppressive treatment for autoimmune disease

Visceral leishmaniasis (VL) in patients receiving immunosuppressant drugs for autoimmune disease has been on the rise. It is important—but difficult—to know when cure has been achieved in these patients since the withdrawal of immunosuppressants during antileishmania treatment is commonly required, and there is a risk of relapse when immunosuppression is restored. The prevalence of asymptomatic infection among those immunosuppressed for autoimmune disease is also uncertain. The present work describes how cytokine release assays can be used to confirm the cure of VL, and to determine the prevalence of asymptomatic infection, in such patients. After collection of blood from volunteers (n = 108), SLA-stimulation of peripheral blood mononuclear cell cultures and of whole blood was found to induce the production of different combinations of cytokines that served to confirm recovery from VL, and asymptomatic Leishmania infection. Indeed, cure was confirmed in 14 patients, all of whom showed a specific Th1 immune response against Leishmania, and the prevalence of asymptomatic infection was determined as 21.27%. Cytokine profiles could be used to manage VL in patients with autoimmune disease, and to identify and better protect those with asymptomatic infection who are at risk of developing this disease.     

Detection and Identification of Old World Leishmania by High Resolution Melt Analysis

Background

Three major forms of human disease, cutaneous leishmaniasis, visceral leishmaniasis and mucocutaneous leishmaniasis, are caused by several leishmanial species whose geographic distribution frequently overlaps. These Leishmania species have diverse reservoir hosts, sand fly vectors and transmission patterns. In the Old World, the main parasite species responsible for leishmaniasis are Leishmania infantum, L. donovani, L. tropica, L. aethiopica and L. major. Accurate, rapid and sensitive diagnostic and identification procedures are crucial for the detection of infection and characterization of the causative leishmanial species, in order to provide accurate treatment, precise prognosis and appropriate public health control measures.

Methods/Principal Findings

High resolution melt analysis of a real time PCR product from the Internal Transcribed Spacer-1 rRNA region was used to identify and quantify Old World Leishmania in 300 samples from human patients, reservoir hosts and sand flies. Different characteristic high resolution melt analysis patterns were exhibited by L. major, L. tropica, L. aethiopica, and L. infantum. Genotyping by high resolution melt analysis was verified by DNA sequencing or restriction fragment length polymorphism. This new assay was able to detect as little as 2-4 ITS1 gene copies in a 5 µl DNA sample, i.e., less than a single parasite per reaction.

Conclusions/Significance

This new technique is useful for rapid diagnosis of leishmaniasis and simultaneous identification and quantification of the infecting Leishmania species. It can be used for diagnostic purposes directly from clinical samples, as well as epidemiological studies, reservoir host investigations and vector surveys.     

Survey of Sand Flies (Diptera: Psychodidae) in an Environmentally Protected Area in Brazil

Brazil is one of the most important endemic areas for leishmaniasis worldwide. Protected areas that are tourist attractions likely present an important risk of transmission of cutaneous leishmaniasis (CL). Furthermore, with the geographical expansion of visceral leishmaniasis (VL), several studies have recorded the occurrence of its vector, Lutzomyia longipalpis, and cases of human and canine VL in such tourist areas. The Parque Estadual do Sumidouro is an environmentally protected area located in the Brazilian Cerrado biome and in an important area endemic for leishmaniasis in the state of Minas Gerais. The purpose of this study was to monitor the sand fly fauna in areas of tourist activity in the park. Sampling was performed every month, from September 2011 to August 2013, using CDC light traps at six sites of differing environmental characteristics. Sampled specimens were identified following Galati (2003), and females were submitted to molecular techniques for the detection and identification of Leishmania DNA. A total of 4,675 sand fly specimens of 25 species belonging to nine genera were collected. The most abundant species were Micropygomyia quinquefer, Lutzomyia renei and Pintomyia pessoai, although only Pi. pessoai is implicated in the transmission of Leishmania braziliensis. The species accumulation curve reached saturation on the 16th sampling event. Species richness, diversity and evenness differed among the sampled areas. The seasonal curve was not determined by a single unique species, and no single species was the most abundant in all environments sampled. The main vector of Leishmania (Leishmania) infantum, Lutzomyia longipalpis, accounted for only 5.35% of the specimens collected. Proven or suspected vectors of Leishmania (Viannia) braziliensis were recorded, and one female of the cortellezzii complex tested positive for Le. braziliensis DNA. Even with a low infection rate (0.62%), these data indicate the circulation of the parasite and reinforce the need for entomological and epidemiological surveillance in the park and its surroundings.     

Integrity of the Actin Cytoskeleton of Host Macrophages is Essential for Leishmania donovani Infection

Visceral leishmaniasis is a vector-borne disease caused by an obligate intracellular protozoan parasite Leishmania donovani. The molecular mechanism involved in internalization of Leishmania is poorly understood. The entry of Leishmania involves interaction with the plasma membrane of host cells. We have previously demonstrated the requirement of host membrane cholesterol in the binding and internalization of L. donovani into macrophages. In the present work, we explored the role of the host actin cytoskeleton in leishmanial infection. We observed a dose-dependent reduction in the attachment of Leishmania promastigotes to host macrophages upon destabilization of the actin cytoskeleton by cytochalasin D. This is accompanied by a concomitant reduction in the intracellular amastigote load. We utilized a recently developed high resolution microscopy-based method to quantitate cellular F-actin content upon treatment with cytochalasin D. A striking feature of our results is that binding of Leishmania promastigotes and intracellular amastigote load show close correlation with cellular F-actin level. Importantly, the binding of Escherichia coli remained invariant upon actin destabilization of host cells, thereby implying specific involvement of the actin cytoskeleton in Leishmania infection. To the best of our knowledge, these novel results constitute the first comprehensive demonstration on the specific role of the host actin cytoskeleton in Leishmania infection. Our results could be significant in developing future therapeutic strategies to tackle leishmaniasis.     

Dual Immune Modulatory Effect of Vitamin A in Human Visceral Leishmaniasis

Vitamin A supplementation has shown to prevent mortality by diarrheal and respiratory diseases in several countries. Nevertheless, there are few studies investigating the effect of vitamin A in visceral leishmaniasis (VL), although there are reports of its deficiency in children with symptomatic VL in Brazil and Bangladesh. This study analyzed the effect of vitamin A on a subset of Treg cells and monocytes isolated from symptomatic VL and from healthy children residing in an endemic area for VL in Northeast Brazil. Serum retinol concentrations correlated inversely with IL-10 and TGF-β productions in CD4⁺CD25^highFoxp3⁺ T cells isolated from children with VL stimulated with leishmanial antigens. All-trans retinoic acid in vitro induced IL-10 in CD4⁺CD25^highFoxp3⁺ T cells; IL-10 and TGF-β production in CD4⁺CD25⁻Foxp3⁻ T cells, and IL-10 in monocytes isolated from healthy children. However, the use of all-trans retinoic acid together with leishmanial antigens in vitro prevented increases in IL-10 production in Treg cells and monocytes isolated from VL children. Strikingly, those results show a potential dual role of vitamin A in the immune system: improvement of a regulatory profile in cells from healthy children after leishmanial stimulation and down modulation of IL-10 in Treg cells and monocytes during symptomatic VL. Therefore, the use of vitamin A concomitant to VL therapy might be useful in improving recovery from disease status caused by Leishmania infantum infection and warrants additional study.     

Asymptomatic Leishmania infection in HIV-positive outpatients on antiretroviral therapy in Pernambuco,Brazil

BackgroundVisceral leishmaniasis (VL) in HIV-positive individuals is a global health problem. HIV-Leishmania coinfection worsens prognosis and mortality risk, and HIV-Leishmania coinfected individuals are more susceptible to VL relapses. Early initiation of antiretroviral therapy can protect against Leishmania infection in individuals living in VL-endemic areas, and regular use of antiretrovirals might prevent VL relapses in these individuals. We conducted a cross-sectional study in Petrolina, Brazil, an VL-endemic area, to estimate the prevalence of asymptomatic Leishmania cases among HIV-positive outpatients.MethodsWe invited any HIV-positive patients, aged ≥ 18-years-old, under antiretroviral therapy, and who were asymptomatic for VL. Patients were tested for Leishmania with enzyme-linked immunosorbent assays (ELISA)-rK39, immunochromatographic test (ICT)-rK39, direct agglutination test (DAT), latex agglutination test (KAtex), and conventional polymerase chain reaction (PCR). HIV-Leishmania coinfection was diagnosed when at least one VL test was positive.ResultsA total of 483 patients were included. The sample was predominantly composed of single, < 48-years-old, black/pardo, heterosexual males, with fewer than 8 years of schooling. The prevalence of asymptomatic HIV-Leishmania coinfection was 9.11% (44/483). HIV mono-infected and HIV-Leishmania coinfected groups differed statistically significantly in terms of race (p = 0.045), marital status (p = 0.030), and HIV viral load (p = 0.046). Black/pardo patients, married patients, and those with an HIV viral load up to 100,000 copies/ml presented higher odds for HIV-Leishmania coinfection.ConclusionsA considerable number of asymptomatic Leishmania cases were observed among HIV-positive individuals in a VL-endemic area. Given the potential impact on transmission and health costs, as well as the impact on these coinfected individuals, studies of asymptomatic Leishmania carriers can be useful for guiding public health policies in VL-endemic areas aiming to control and eliminate the disease.     

Clinical and laboratory characteristics of hemophagocytic lymphohistiocytosis induced by Leishmania infantum infection

BackgroundVisceral leishmaniasis (VL) could progress to secondary hemophagocytic lymphohistiocytosis (HLH), which is a rare but life-threatening condition with poor prognosis. So far, the clinical and laboratory characteristics of VL associated HLH have not been well elucidated.Method and findingsIn this study, we retrospectively analyzed the clinical and laboratory profiles between 17 patients with VL associated HLH and 27 patients with VL alone admitted at the Beijing Friendship Hospital, Capital Medical University from May 2016 to March 2021. In addition to the identification of Leishmania infection, hemophagocytosis was identified in bone marrow in the most cases of VL associated HLH (15/17). The patients with VL associated HLH had higher chances of bleeding, hepatomegaly, thrombocytopenia, hypertriglyceridemia, hyperferritinemia, hypofibrinogenemia, elevated secretion of soluble IL-2 receptor or lower NK cell activity compared to patients with VL only. Furthermore, patients with VL associated HLH had higher inflammation status associated with higher levels of Th1 (TNF-α, IFN-γ, IL-1beta, IL-6, IL-8, IL-12p70), Th2 (IL-4) and Th17 cytokines (IL-17, IL-23) in the peripheral blood, and higher parasite load (qPCR and parasite culture). All 27 VL cases were totally recovered after being treated with Sodium Stibogluconate, five of the 17 patients with VL associated HLH died even after timely treatment with anti-parasite and immunosuppressive chemotherapy.ConclusionWithout appropriate treatment, visceral leishmaniosis could develop to secondary HLH. The parasite culturing and qPCR detection of bone marrow samples facilitates the diagnosis of VL associated HLH in addition to other findings of HLH. Prompt treatment with anti-Leishmania and immunosuppressive chemotherapy is critical to reduce the mortality of VL associated HLH.     

Heterogeneity of Leishmania donovani Parasites Complicates Diagnosis of Visceral Leishmaniasis: Comparison of Different Serological Tests in Three Endemic Regions

Diagnostic tests for visceral leishmaniasis that are based on antigens of a single Leishmania strain can have low diagnostic performance in regions where heterologous parasites predominate. The aim of this study was to investigate and compare the performance of five serological tests, based on different Leishmania antigens, in three endemic countries for visceral leishmaniasis. A total number of 231 sera of symptomatic and asymptomatic cases and controls from three endemic regions of visceral leishmaniasis in East Sudan, North India and South France were evaluated by following serological tests: rKLO8- and rK39 ELISA, DAT (ITMA-DAT) and two rapid tests of rK39 (IT LEISH) and rKE16 (Signal-KA). Overall, rKLO8- and rK39 ELISA were most sensitive in immunocompetent patients from all endemic regions (96–100%) and the sensitivity was reduced to 81.8% in HIV co-infected patients from France. Sera of patients from India demonstrated significantly higher antibody responses to rKLO8 and rK39 compared with sera from Sudan (p<0.0001) and France (p<0.0037). Further, some Indian and Sudanese patients reacted better with rKLO8 than rK39. Sensitivity of DAT (ITMA-DAT) was high in Sudan (94%) and India (92.3%) but low in France being 88.5% and 54.5% for VL and VL/HIV patients, respectively. In contrast, rapid tests displayed high sensitivity only in patients from India (96.2%) but not Sudan (64–88%) and France (73.1–88.5% and 63.6–81.8% in VL and VL/HIV patients, respectively). While the sensitivity varied, all tests showed high specificity in Sudan (96.7–100%) and India (96.6%).Heterogeneity of Leishmania parasites which is common in many endemic regions complicates the diagnosis of visceral leishmaniasis. Therefore, tests based on homologous Leishmania antigens are required for particular endemic regions to detect cases which are difficult to be diagnosed with currently available tests.     

FRB domain of human TOR protein induces compromised proliferation and mitochondrial dysfunction in Leishmaniadonovani promastigotes

Visceral leishmaniasis (VL) or Kala-azar, the second-largest parasitic killer worldwide, is caused by Leishmania donovani. The drugs to treat VL are toxic and expensive. Moreover, their indiscriminate use gave rise to resistant strains. The high rate of parasite proliferation within the host macrophage cells causes pathogenesis. In the proliferative pathway, FRB domain of TOR protein is ubiquitously essential. Although orthologues of mTOR protein are reported in trypanosomatids and Leishmania but therein depth molecular characterization is yet to be done. Considerable protein sequence homology exists between the TOR of kinetoplastidas and mammals. Interestingly, exogenous human FRB domain was shown to block G1 to S transition in mammalian cancer cells. Thus, we hypothesized that expression of human FRB domain would inhibit the proliferation of Leishmaniadonovani. Indeed, promastigotes stably expressing wild type human FRB domain show 4.7 and 1.5 folds less intra- and extra-cellular proliferations than that of untransfected controls. They also manifested 2.65 times lower rate of glucose stimulated oxygen consumption. The activities of all respiratory complexes were compromised in the hFRB expressing promastigotes. In these cells, depolarized mitochondria were 2-fold more than control cells. However, promastigotes expressing its mutant version (Trp²⁰²⁷-Phe) has shown similar characteristics like untransfected cells. Thus, this study reveals greater insights on the conserved role of TOR in the regulation of the respiratory complexes in L. donovani. The slow growing variant of FRB expressing promastigotes will have great potential to be exploited as a prophylactic agent against leishmaniasis.     

Predictors of visceral leishmaniasis relapse in HIV-infected patients: a systematic review

Background and Objectives

Visceral leishmaniasis (VL) is a common complication in AIDS patients living in Leishmania-endemic areas. Although antiretroviral therapy has changed the clinical course of HIV infection and its associated illnesses, the prevention of VL relapses remains a challenge for the care of HIV and Leishmania co-infected patients. This work is a systematic review of previous studies that have described predictors of VL relapse in HIV-infected patients.

Review Methods

We searched the electronic databases of MEDLINE, LILACS, and the Cochrane Central Register of Controlled Trials. Studies were selected if they included HIV-infected individuals with a VL diagnosis and patient follow-up after the leishmaniasis treatment with an analysis of the clearly defined outcome of prediction of relapse.

Results

Eighteen out 178 studies satisfied the specified inclusion criteria. Most patients were males between 30 and 40 years of age, and HIV transmission was primarily via intravenous drug use. Previous VL episodes were identified as risk factors for relapse in 3 studies. Two studies found that baseline CD4+ T cell count above 100 cells/mL was associated with a decreased relapse rate. The observation of an increase in CD4+ T cells at patient follow-up was associated with protection from relapse in 5 of 7 studies. Meta-analysis of all studies assessing secondary prophylaxis showed significant reduction of VL relapse rate following prophylaxis. None of the five observational studies evaluating the impact of highly active antiretroviral therapy use found a reduction in the risk of VL relapse upon patient follow-up.

Conclusion

Some predictors of VL relapse could be identified: a) the absence of an increase in CD4+ cells at follow-up; b) lack of secondary prophylaxis; and c) previous history of VL relapse. CD4+ counts below 100 cells/mL at the time of primary VL diagnosis may also be a predictive factor for VL relapse.     

Seropositivity of Visceral leishmaniasis on people of VL endemic three districts of Nepal

Visceral leishmaniasis (VL) is a life-threatening vector borne disease caused by the Leishmania donovani species complex. In Nepal, it is transmitted to humans by L donovani infected Phlebotomus argentipes sand flies [12]. The pathogenesis of VL is complex, and the clinical presentation ranges from asymptomatic infection to severe and fatal disease. Asymptomatic infection may act as potential reservoirs for sustained transmission of VL in endemic areas. We investigated the sero-prevalence of symptomatic and asymptomatic infection of VL in people of three endemic districts of Nepal by serology targeting family members and neighbors of VL patients. Sero-survey was conducted among 189 people of villages endemic to VL from Palpa, Sarlahi and Saptari districts during 2016 to 2018 using the rK39 rapid diagnostic test (InBios International, Seattle, WA) to detect anti-Leishmania antibodies. Sero-positivity was 35.7% (10/28) in people tested from Sarlahi districts, 6% (3/50) in Saptari district and 1.7% (1/59) from the Palpa district. In Sarlahi, sero-positivity was found to be highest among the age group below 15 years (44.5%). All family members of diagnosed VL cases in Saptari and Palpa districts were found to be rK39 test negative. In Sarlahi district, among the ten sero-positive cases, nine were febrile and became symptomatic VL cases after few days and one case remained asymptomatic during the six month follow up. Asymptomatic cases in VL endemic districts of Nepal were found to be sero-positive, screening of people in VL endemic districts would be important for prevention of VL transmission.