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Visceral leishmaniasis is a potentially devastating neglected tropical disease caused by the protozoan parasites Leishmania donovani and L. infantum (chagasi). These parasites reside in tissue macrophages and survive by deploying a number of mechanisms aimed at subverting the host immune response. CD4+ T cells play an important role in controlling Leishmania parasites by providing help in the form of pro-inflammatory cytokines to activate microbiocidal pathways in infected macrophages. However, because these cytokines can also cause tissue damage if over-produced, regulatory immune responses develop, and the balance between pro-inflammatory and regulatory CD4+ T cells responses determines the outcomes of infection. Past studies have identified important roles for pro-inflammatory cytokines such as IFNg and TNF, as well as regulatory co-inhibitory receptors and the potent antiinflammatory cytokine IL-10. More recently, other immunoregulatory molecules have been identified that play important roles in CD4+ T cell responses during VL. In this review, we will discuss recent findings about two of these molecules; the NK cell granule protein Nkg7 and the anti-inflammatory cytokine TGFβ, and describe how they impact CD4+ T cell functions and immune responses during visceral leishmaniasis. [ABSTRACT FROM AUTHOR]

Background: Leishmania is an intracellular flagellate protozoan parasite that causes a wide range of clinical diseases in humans. The basis of immunological resistance against leishmaniasis depends on Thl reactions and is within the time period of cytokine function. Methods: In this study, human anti-IL17 antibody and IFNγ-producing promastigote were produced to be used in leishmanization. A sequence of light and heavy chains' gene of anti-IL17 antibody and human IFNγ (hIFNγ) was obtained from the NCBI database and synthesized in the ECORV reaction site in the plasmid pGH, which it's called pGH–hIFNγ–antiIL17. The synthesized part using the restriction enzyme ECORV was extracted from the plasmid and after purification by electroporation was transferred to Iranian lizard Leishmania (I.L.L). Evaluation of structural presence in the I.L.L genome at the level of DNA and mRNA was assessed. The expressions of hIFNγ and anti-IL17 were evaluated and confirmed using ELISA and western blot analysis. The hIFNγ secreted from the culture medium was collected at high concentrations of 124.36 ± 6.47 pg/mL. Results: Targeted gene replacement into the I.L.L genome was successfully performed for the first time using the pGH–hIFNγ–antiIL17 plasmid in an identical replacement process. Stabilized recombinant DNA contains a target gene that has no toxicity to the parasite. Conclusions: The effective achievement of producing a recombinant gene was done for the first time by replacing the I.L.L-CPC gene with plasmid pGH-hIFNγ-antiIL17 by targeted gene replacement. This cab can regulate the production of hIFNγ and anti-IL17. This makes it a viable choice for eliminating leishmania. [ABSTRACT FROM AUTHOR]

Background: Ischemic stroke is the most common form of stroke and the second most common cause of death and incapacity worldwide. Its pathogenesis and treatment have been the focus of considerable research. In traditional Chinese medicine, the root of Mongolian astragalus has been important in the treatment of stroke since ancient times. Astragalus polysaccharide (APS) is a key active ingredient of astragalus and offers therapeutic potential for conditions affecting the neurological system, the heart, cancer, and other disorders. However, it is not yet known how APS works to protect against ischemic stroke. Methods: Rats were subjected to middle cerebral artery occlusion (MCAO) to imitate localized cerebral ischemia. Each of four experimental groups (normal, sham, MCAO, and MCAO+APS) contained 12 adult male Sprague-Dawley (SD) rats selected randomly from a total of 48 rats. Following successful establishment of the model, rats in the MCAO+APS group received intraperitoneal injection of APS (50 mg/kg) once daily for 14 days, whereas all other groups received no APS. The Bederson nerve function score and the forelimb placement test were used to detect motor and sensory function defects, while Nissl staining was used to investigate pathological defects in the ventroposterior thalamic nucleus (VPN). Immunohistochemical staining and Western blot were used to evaluate the expression of Neurogenic locus notch homolog protein 1 (Notch1), hairy and enhancer of split 1 (Hes1), phospho-nuclear factor-κB p65 (p-NFκB p65), and nuclear factor-κB p65 (NFκB p65) proteins in the VPN on the ischemic side of MCAO rats. Results: APS promoted the recovery of sensory and motor function, enhanced neuronal morphology, increased the number of neurons, and inhibited the expression of Notch1/NFκB signaling pathway proteins in the VPN of rats with cerebral ischemia. Conclusion: After cerebral ischemia, APS can alleviate symptoms of secondary damage to the VPN, which may be attributed to the suppression of the Notch1/NFκB pathway. [ABSTRACT FROM AUTHOR]

Aim: To compare and evaluate the differences between EGCG and phosphatidylcholine-encapsulated EGCG in terms of their anti-inflammatory and antioxidant capacities., Methods: In this study, transdermal absorption experiments were conducted to compare the absorption capacity of EGCG and phosphatidylcholine-encapsulated EGCG. Subsequently, the disparity in anti-inflammatory and antioxidant efficacy between EGCG and phosphatidylcholine-encapsulated EGCG were evaluated through cytotoxicity experiments, as well as the determination of cellular inflammatory factors and the measurement of ROS content under different treatment conditions., Results: The concentration of EGCG, encapsulated in phosphatidylcholine, in porcine skin is 40.76 ± 1.29 μg/cm 2 , which is significantly higher than the concentration of EGCG alone (31.62 ± 2.01 μg/cm 2 ). Also, the ability of phosphatidylcholine-encapsulated EGCG to suppress inflammatory factors such as tumor necrosis factor-α (TNF-α), cyclooxygenase-2 (COX-2), and prostaglandin E2 (PGE2) was notably superior to that of EGCG alone. Both phosphatidylcholine-encapsulated EGCG and EGCG showed excellent ROS scavenging ability in terms of antioxidant capacity., Conclusion: The percutaneous absorption and anti-inflammatory impact of EGCG encapsulated within phosphatidylcholine were substantially enhanced when compared to EGCG by itself. Additionally, both formulations exhibited enhanced ROS scavenging capacities, albeit the variance between them was not pronounced. These insights furnish a vital theoretical underpinning for the utilization of phosphatidylcholine-encapsulated EGCG in cosmetic applications, specifically for fostering products with anti-inflammatory and antioxidant benefits., (© 2024 The Author(s). Journal of Cosmetic Dermatology published by Wiley Periodicals LLC.)

Purpose: Visceral leishmaniasis (VL) is caused by an intracellular parasite that is transmitted to humans by sandfly bites. It is prevalent throughout Asia, Africa, the Americas, and the Mediterranean area, where 147 million people are at risk of contracting the illness. The manifestation of heterotrophic illness relies on both Leishmania implicated and the host's immunological response, ranging from asymptomatic to severe leishmaniasis with potentially lethal effects., Method: We reviewed the literature (published till 31st December 2023) on the worldwide situation of leishmaniasis, standard and novel detection techniques, and traditional and modern treatment strategies and endeavors to eliminate VL. Moreover, epidemiological data was collected from the World Health Organization's publicly available databases. GraphPad Prism Version 8 was used to analyze and produce figures based on the epidemiological data., Results: Diagnosis of parasites in tissues or serology is commonly employed. Diagnosis by identifying parasite DNA using molecular techniques is becoming more popular. Despite recent findings of L. donovani resistance to pentavalent antimoniate medications, it continues to be the cornerstone in the medical management of VL. Amphotericin B and its lipid formulations, injectable paromomycin, and oral miltefosine are among the new therapy options being researched. The number of reported VL cases has reduced remarkably over the last decade due to human interventions made to eliminate VL. Particularly countries from the South East Asian region have experienced momentous progress in reducing VL cases and eliminating this disease from this region. Owing to the robust elimination programs, countries such as Bangladesh has eliminated VL as a public health concern. India and Nepal are on the verge of its elimination., Conclusion: Rapid diagnosis, effective and inexpensive treatment, simple access to newly discovered medications, appropriate vector control, and a well-designed vaccine are all required for the elimination of this disease burden in impoverished areas of the globe., (© 2024. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Obligate intracellular protozoan parasite, Leishmania donovani, causative agent of visceral leishmaniasis, led to impaired macrophage functions. It is well documented that many of these changes were induced by parasite-mediated reduction in macrophage cholesterol content. Leishmania-mediated alteration in the other lipids has not been explored in detail yet. Here, we found that the expression of key cholesterol biosynthetic genes and total cellular cholesterol were reduced during L. donovani infection. Further, we have also identified that this reduction in the cholesterol led to increased membrane fluidity and inhibition of antigen-presenting potential of macrophages. In addition to this, we studied the relative changes in different lipids in THP-1-derived macrophages during L. donovani infection through liquid chromatography-mass spectrometry. We found that Sphingomyelin (16:0) and ceramide (20:1, 26:0 and 26:1) were significantly reduced in infected macrophages. We further observed that the majority of different sub-classes of phospholipids were downregulated significantly. Overall ratio of phosphatidylcholine versus phosphotidylethanolamine was decreased which indicated the compensatory mechanism of cell in response to cholesterol reduction. The observed Leishmania-mediated alteration in macrophage-lipidome provided the novel insights into mechanism of host-pathogen interactions., (© 2024 John Wiley & Sons Ltd.)

Background: Leishmaniasis is one of the most neglected tropical diseases and is spread mainly in impoverished regions of the world. Although many studies have focused on the host's response to Leishmania invasion, relatively less is known about the complex processes at the metabolic level, especially the metabolic alterations in the infected hosts. Methods: In this study, we conducted metabolomics analysis on the urine of golden hamsters in the presence or absence of visceral leishmaniasis (VL) using the ultra-performance liquid chromatography (UPLC) system tandem high-resolution mass spectrometer (HRMS). The metabolic characteristics of urine samples, along with the histopathological change and the parasite burden of liver and spleen tissues, were detected at 4 and 12 weeks post infection (WPI), respectively. Results: Amino acid metabolism was extensively affected at both stages of VL progression. Meanwhile, there were also distinct metabolic features at different stages. At 4 WPI, the significantly affected metabolic pathways involved alanine, aspartate and glutamate metabolism, the pentose phosphate pathway (PPP), histidine metabolism, tryptophan metabolism and tyrosine metabolism. At 12 WPI, the markedly enriched metabolic pathways were almost concentrated on amino acid metabolism, including tyrosine metabolism, taurine and hypotaurine metabolism and tryptophan metabolism. The dysregulated metabolites and metabolic pathways at 12 WPI were obviously less than those at 4 WPI. In addition, seven metabolites that were dysregulated at both stages through partial least squares-discriminant analysis (PLS-DA) and receiver-operating characteristic (ROC) tests were screened to be of diagnostic potential. The combination of these metabolites as a potential biomarker panel showed satisfactory performance in distinguishing infection groups from control groups as well as among different stages of infection. Conclusion: Our findings could provide valuable information for further understanding of the host response to Leishmania infection from the aspect of the urine metabolome. The proposed urine biomarker panel could help in the development of a novel approach for the diagnosis and prognosis of VL. [ABSTRACT FROM AUTHOR]

Leishmania donovani and Leishmania infantum are closely related species. However, the former is considered the causative agent for anthroponotic visceral leishmaniasis (AVL), while the latter is known to be responsible for zoonotic visceral leishmaniasis (ZVL) with dogs as the main reservoir host. Although molecular detection of L. donovani from naturally infected dogs has been reported in AVL endemic areas, the experimental infection of dogs with this species is very limited. Here, we constructed an experimental canine visceral leishmaniasis (CVL) model with L. donovani infection using beagle dogs. During an observation period of 8 months after parasite inoculation, few clinical symptoms were observed in the three inoculated dogs. The overall hematological and biochemical data of the dogs showed normal levels, and there were no remarkable changes in the peripheral CD4+, CD8+, CD25+, or FoxP3+ T cell populations. Liver biopsy sampling was conducted to monitor the parasite burden in the liver. A similar pattern of the amount of mitochondrial kinetoplast DNA was observed in the peripheral blood and liver by real-time PCR analysis. In addition, parasite antigens were detected from the liver biopsy sections by immunohistochemical analysis, further supporting the existence of parasites in the liver. These results showed a subclinical CVL model for L. donovani in beagle dogs with a similar kinetics of parasite burden in the peripheral blood and liver. [ABSTRACT FROM AUTHOR]

Leishmania is an intracellular parasite with different species pathogenic to humans and causing the disease leishmaniasis. Leishmania donovani causes visceral leishmaniasis (VL) that manifests as hepatosplenomegaly, fever, pancytopenia and hypergammaglobulinemia. If left without treatment, VL can cause death, especially in immunocompromised people. Current treatments have often significant adverse effects, and resistance has been reported in some countries. Determining the metabolites perturbed during VL can lead us to find new treatments targeting disease pathogenesis. We therefore compared metabolic perturbation between L. donovani-infected and uninfected hamsters across organs (spleen, liver, and gut). Metabolites were extracted, analyzed by liquid chromatography-mass spectrometry, and processed with MZmine and molecular networking to annotate metabolites. We found few metabolites commonly impacted by infection across all three sites, including glycerophospholipids, ceramides, acylcarnitines, peptides, purines and amino acids. In accordance with VL symptoms and parasite tropism, we found a greater overlap of perturbed metabolites between spleen and liver compared to spleen and gut, or liver and gut. Targeting pathways related to these metabolite families would be the next focus that can lead us to find more effective treatments for VL. [ABSTRACT FROM AUTHOR]

Background: Recent global evidences showed that asymptomatic blood donor carriers of Leishmania infection will appear as a threat for blood transfusions recipients in endemic areas. As yet, there is no appropriate diagnostic procedure for detecting infection of blood donors in blood banks. Subjects and Methods: The present study was aimed to apply various current diagnostic tests among blood donors in an endemic area of visceral leishmaniasis (VL), Ardabil Province, northwestern Iran. Blood samples were gathered from 860 blood donors in endemic areas of the province between 2017 and 2018, at eight blood donation centers. These samples was assessed using microculture, serological (DAT and rK39-ICT) and molecular based (conventional kDNA-PCR and HRM-PCR) tests. Results: Of 860 eligible donors, 24 (2.8%) were seropositive for VL by DAT, and 388 (45%) were positive by kDNA-PCR. Moreover, 19 (19/860) were positive for both of them. Out of 19 subjects, 5.3% (1/19) was positive by rK39-ICT, 10.5% (2/19), and 79% (15/19) were detected positive in microculture and HRM-PCR methods, respectively. Nineteen donors were followed up for 2 years, of which 16 (84.2%) had a serological conversion, and 4 (21%) were positive by kDNA-PCR. The sensitivity of kDNA-PCR, and HRM-PCR procedures in detecting Leishmania parasite was found to be 98.7%, and 79%, respectively. Conclusions: Our findings justify the use of kDNA-PCR as a convenient and sensitive tool for screening subjects with leishmanial latent infection in blood banks at least in endemic regions. In these areas, however, a PCR-based test should be used to validate Leishmania infection among seropositive donors. [ABSTRACT FROM AUTHOR]

A key point of immunity against protozoan Leishmania parasites is the development of an optimal T cell response, which includes a low apoptotic rate, high proliferative activity and polyfunctionality. During acute infection, antigen-specific T cells recognize the pathogen resulting in pathogen control but not elimination, promoting the development and the maintenance of a population of circulating effector cells that mount rapid response quickly after re-exposure to the parasite. However, in the case of visceral disease, the functionality of specific T cells is lost during chronic infection, resulting in inferior effector functions, poor response to specific restimulation, and suboptimal homeostatic proliferation, a term referred to as T cell exhaustion. Multiple factors, including parasite load, infection duration and host immunity, affect T lymphocyte exhaustion. These factors contribute to antigen persistence by promoting inhibitory receptor expression and sustained production of soluble mediators, influencing suppressive cell function and the release of endogenous molecules into chronically inflamed tissue. Together, these signals encourage several changes, reprogramming cells into a quiescent state, which reflects disease progression to more severe forms, and development of acquired resistance to conventional drugs to treat the disease. These points are discussed in this review. [ABSTRACT FROM AUTHOR]

Structural changes in the spleen have been reported in several infectious diseases. In visceral leishmaniasis (VL), a severe parasitic disease caused by Leishmania spp., the loss of white pulp accompanies a severe clinical presentation. Hamster model reproduces aspects of human VL progression. In the early stages, a transcriptomic signature of leukocyte recruitment was associated with white pulp hyperplasia. Subsequently, impaired leukocyte chemotaxis with loss of T lymphocytes in the periarteriolar lymphoid sheath occurred. This differential gene expression was subsequently corroborated by transcriptomic profiling of spleens in severe human VL. At the latest stage, spleen disorganization was associated with increasing clinical signs of VL. White pulp disruption was accompanied by decreased DLK1 expression. The expression of CXCL13, CCR5, CCL19, CCR6, CCR7 and LTA decreased, likely regulated by CDKN2A overexpression. Our findings enlighten a pathway implying cell cycle arrest and decreased gene expression involved in spleen organization. [ABSTRACT FROM AUTHOR]

The differences in morbidity and mortality patterns and life expectancy between the sexes are well established in different infectious and parasitic conditions, such as in leishmaniases, in which biological, genetic, sexual and hormonal variations can modulate the immune response indicating greater infectivity, prevalence and clinical severity in men. In this regard, in seeking the understanding of factors related to protection and susceptibility to infection, this review aimed to discuss the influence of sex hormones on the immune response to leishmaniases. In the literature, sex hormone variations promote differences in the innate, humoral and cell‐mediated immune response, leading to greater susceptibility, mortality and complications in males. Epidemiological estimates confirm these results, showing a predominance of the disease, in its different clinical forms, in men and suggesting that sexual variations influence immunomodulatory mechanisms since the prevalence of cases comprises the post‐puberty and adulthood period. In this perspective, the action of sex hormones has been investigated in different clinical models, highlighting the potential of testosterone in immunosuppression, given its association with greater susceptibility and poor control of parasite load and the induction of cell apoptosis and attenuation of pro‐inflammatory signalling pathways. Therefore, hormonal variations influence the immune response among males and females against leishmaniases, in which androgens may present immunosuppressive potential, while steroids present immunomodulatory characteristics. [ABSTRACT FROM AUTHOR]

Background: Miltefosine (MIL) is currently the only oral drug available to treat visceral leishmaniasis but its use as first-line monotherapy has been compromised by an increasing treatment failure. Despite the scarce number of resistant clinical isolates, MIL-resistance by mutations in a single aminophospholipid transporter gene can easily be selected in a laboratory environment. These mutations result in a reduced survival in the mammalian host, which can partially be restored by exposure to MIL, suggesting a kind of drug-dependency. Methodology/Principal findings: To enable a combined study of the infection dynamics and underlying immunological events for differential in vivo survival, firefly luciferase (PpyRE9) / red fluorescent protein (DsRed) double-reporter strains were generated of MIL-resistant (MIL-R) and syngeneic MIL-sensitive (MIL-S) Leishmania infantum. Results in C57Bl/6 and BALB/c mice show that MIL-R parasites induce an increased innate immune response that is characterized by enhanced influx and infection of neutrophils, monocytes and dendritic cells in the liver and elevated serum IFN-γ levels, finally resulting in a less efficient establishment in liver macrophages. The elevated IFN-γ levels were shown to originate from an increased response of hepatic NK and NKT cells to the MIL-R parasites. In addition, we demonstrated that MIL could increase the in vivo fitness of MIL-R parasites by lowering NK and NKT cell activation, leading to a reduced IFN-γ production. Conclusions/Significance: Differential induction of innate immune responses in the liver was found to underlie the attenuated phenotype of a MIL-R parasite and its peculiar feature of drug-dependency. The impact of MIL on hepatic NK and NKT activation and IFN-γ production following recognition of a MIL-R strain indicates that this mechanism may sustain infections with resistant parasites and contribute to treatment failure. Author summary: Visceral leishmaniasis is a neglected tropical disease that is fatal if left untreated. Miltefosine is currently the only oral drug available but is increasingly failing to cure patients, resulting in its discontinuation as first-line drug in some endemic areas. To understand these treatment failures, we investigated the complex interplay of the parasite with the host immune system in the presence and absence of miltefosine. Our data indicate that miltefosine-resistant Leishmania parasites become severely hampered in their in vivo infectivity, which could be attributed to the induction of a pronounced innate immune response. Interestingly, the infection deficit was partially restored in the presence of miltefosine. Our results further indicate that miltefosine can exacerbate infections with resistant parasites by reducing innate immune recognition. This study provides new insights into the complex interplay between parasite, drug and host and discloses an immune-related mechanism of treatment failure. [ABSTRACT FROM AUTHOR]

Background: There is a continued need to develop effective and safe treatments for visceral leishmaniasis (VL). Preclinical studies on pharmacokinetics and pharmacodynamics of anti-infective agents, such as anti-bacterials and anti-fungals, have provided valuable information in the development and dosing of these agents. The aim of this study was to characterise the pharmacokinetic and pharmacodynamic properties of the anti-leishmanial drugs AmBisome and miltefosine in a preclinical disease model of VL. Methodology / Principal findings: BALB/c mice were infected with L. donovani (MHOM/ET/67/HU3) amastigotes. Groups of mice were treated with miltefosine (orally, multi-dose regimen) or AmBisome (intravenously, single dose regimen) or left untreated as control groups. At set time points groups of mice were killed and plasma, livers and spleens harvested. For pharmacodynamics the hepatic parasite burden was determined microscopically from tissue impression smears. For pharmacokinetics drug concentrations were measured in plasma and whole tissue homogenates by LC-MS. Unbound drug concentrations were determined by rapid equilibrium dialysis. Doses exerting maximum anti-leishmanial effects were 40 mg/kg for AmBisome and 150 mg/kg (cumulatively) for miltefosine. AmBisome displayed a wider therapeutic range than miltefosine. Dose fractionation at a total dose of 2.5 mg/kg pointed towards concentration-dependent anti-leishmanial activity of AmBisome, favouring the administration of large doses infrequently. Protein binding was >99% for miltefosine and amphotericin B in plasma and tissue homogenates. Conclusion / Significance: Using a PK/PD approach we propose optimal dosing strategies for AmBisome. Additionally, we describe pharmacokinetic and pharmacodynamic properties of miltefosine and compare our findings in a preclinical disease model to available knowledge from studies in humans. This approach also presents a strategy for improved use of animal models in the drug development process for VL. Author summary: Visceral leishmaniasis is a neglected tropical disease, with an estimated 200 000–400 000 cases and 20 000–40 000 deaths per year worldwide. Leishmania parasites proliferate in spleen, liver and bone marrow, and the disease is usually fatal if untreated. Despite progress in the development of new therapies in recent years there is still a need to develop effective and safe treatments. Characterising pharmacokinetic and pharmacodynamic properties and their relationships has provided valuable information in the development and dosing of anti-infective agents, such as anti-bacterials and anti-fungals. The present study characterised these properties of the anti-leishmanial drugs AmBisome and miltefosine in a mouse model of visceral leishmaniasis. We propose optimal dosing strategies for AmBisome and present a new approach for investigating anti-leishmanial drug action, which can be applied to the development of new treatments for visceral leishmaniasis. [ABSTRACT FROM AUTHOR]

The increasing use of immunosuppressants in areas where visceral leishmaniasis (VL) is endemic has increased the number of people susceptible to developing more severe forms of the disease. Few studies have examined the quality of the immune response in immunosuppressed patients or experimental animals with VL. The present work characterises the parasite load developed in, and immune response to, Leishmania infantum-induced VL in C57BL/6 mice that, prior to and during infection, received immunosuppressant treatment with methylprednisolone (MPDN), anti-tumour necrosis factor (anti-TNF) antibodies, or methotrexate (MTX). The latter two treatments induced a significant reduction in the number of CD4+ T lymphocytes over the infection period. The anti-TNF treatment was also associated with a higher parasite load in the liver and a lower parasite load in the spleen. This, plus a possibly treatment-induced reduction in the number of cytokine-producing Th1 cells in the spleen, indicates the development of more severe VL. Interestingly, the MPDN and (especially) MTX treatments provoked a greater presence of soluble Leishmania antigen-specific multi-cytokine-producing T cells in the spleen and a lower liver parasite load than in control animals. These results highlight the need to better understand how immunosuppressant treatments might influence the severity of VL in human patients. Author summary: Patients who are immunosuppressed are at greater risk of developing visceral leishmaniasis (VL) when infected with Leishmania. Prior infection with HIV has been traditionally associated with an increased risk of developing VL, but the use of immunosuppressants in the treatment of autoimmune disease has been linked to a higher incidence of VL in Leishmania-endemic areas. It is important to understand the influence these treatments have on Leishmania infection, paying special attention to how they affect the immune response mediated by IFN-γ-, TNF- and IL-2 T-producing T lymphocytes (such cells are necessary if an infection is to be resolved). Studies in this area require the use of murine models of VL. C57BL/6 mice infected with Leishmania infantum that received immunosuppressant treatment with methylprednisolone, anti-tumour necrosis factor antibodies, or methotrexate prior to and during infection, showed differences in their immune response, and in the parasite load developed in various organs. These results highlight the need to better understand how immunosuppressant treatments might influence the severity of VL in human patients. [ABSTRACT FROM AUTHOR]

Background: During infection with Leishmania donovani, parasite control is linked to the systemic Th1 immune response, but in infected organs (liver, spleen and bone marrow), the response differs according to the micro-environment. The pleiomorphic cytokine interleukin-33 (IL-33) exerts various roles during infection, either protective or detrimental. In this study, we explored the role of IL-33 in the outcome of Leishmania infection in the spleen. Methods: We used several mouse models, on BALB/c and C57BL/6 (B6) backgrounds, infected with L. donovani and sacrificed at 15, 30 or 60 days after infection and characterized mRNA expression of immune markers, immune cell populations, histological response, and parasite loads. Results: During infection IL-33 and ST2 mRNA increased in parallel in the spleen of wild type (wt) animals and paralleled the immunodetection of ST2+ and IL-33+ cells; their expression was twice as high in BALB/c, compared to B6 mice. Mice treated with twice-weekly injections of rIL-33 had higher splenic parasite burdens on D15 (BALB/c) or on D60 (B6). In BALB/c, IL-33 treatment led to immune exhaustion with abolition of Th1 cytokine expression (IFN-γ and IL-12) in the spleen and higher serum levels of Th2 cytokines (IL-4, IL-5 and IL-13). In B6, IL-33 treatment induced the Treg cell pathway with a dramatic increase of FoxP3 mRNA induction and expression on tissue sections. IL-33-KO mice had lower parasite loads and a higher Th1 response than their wt counterparts. Conclusions: IL-33 appears as a factor of aggravation of the disease in the spleen tissue of mice infected with L. donovani. [ABSTRACT FROM AUTHOR]

Background: Visceral leishmaniasis (VL) is caused by Leishmania infantum or L. donovani infection. One of the main problems related to this disease is the emergence of severe clinical forms with a lethality of 5–20%, even while under specific treatment. In humans and other species susceptible to fatal VL, such as dogs and hamsters, the disruption of splenic white pulp (WP) is accompanied by disease progression. Control of VL progression is seen in BALB/c mice, as evidenced by a mild clinical presentation and controlled parasite replication in the liver and spleen. In this study, we investigated the features involved in the morphological remodeling of splenic compartments associated with the control of VL progression to death. Methods: We evaluated cohorts of BALB/c mice after 30, 60, and 90 days of infection by L. infantum. Spleen morphology, cell population subsets and cytokine production were studied in the spleen using flow- and histo-cytometry. Results: Intraperitoneal infection with 108 promastigotes of L. infantum led to progressive increases in spleen size at 60 and 90 days after infection. Splenomegaly was the only clinical sign of disease observed. At 30 days after infection, hyperplasia in the WP and decreased numbers of plasmacytoid dendritic cells were observed. The WP hyperplasia subsided at 60 days post-infection. However, the splenomegaly remained in association with increased numbers of macrophages, B and T lymphocytes and plasma cells. An increased number of lymphoid tissue inducer (LTi) cells was observed; these were distributed around the periarteriolar lymphoid sheath in control mice and scattered throughout the red pulp in the Leishmania -infected mice. After 90 days of infection, increased IL-6 and IFN-γ production was seen in the spleen, as well as higher frequencies of follicular and plasmacytoid dendritic cells. Conclusion: The data presented herein emphasizes the potential role of spleen remodeling in the control of severe forms of VL and highlights features potentially involved in this process. [ABSTRACT FROM AUTHOR]

Leishmania donovani causes visceral leishmaniasis (VL), which is typically fatal without treatment. There is substantial variation between individuals in rates of disease progression, response to treatment and incidence of post-treatment sequelae, specifically post-kala-azar dermal leishmaniasis (PKDL). Nevertheless, the majority of infected people are asymptomatic carriers. Hamsters and mice are commonly used as models of fatal and non-fatal VL, respectively. Host and parasite genetics are likely to be important factors, but in general the reasons for heterogeneous disease presentation in humans and animal models are poorly understood. Host microbiota has become established as a factor in cutaneous forms of leishmaniasis but this has not been studied in VL. We induced intestinal dysbiosis in mice and hamsters by long-term treatment with broad-spectrum antibiotics in their drinking water. There were no significant differences in disease presentation in dysbiotic mice. In contrast, dysbiotic hamsters infected with L. donovani had delayed onset and progression of weight loss. Half of control hamsters had a rapid progression phenotype compared with none of the ABX-treated animals and the nine-month survival rate was significantly improved compared to untreated controls (40% vs. 10%). Antibiotic-treated hamsters also had significantly less severe hepatosplenomegaly, which was accompanied by a distinct cytokine gene expression profile. The protective effect was not explained by differences in parasite loads or haematological profiles. We further found evidence that the gut-liver axis is a key aspect of fatal VL progression in hamsters, including intestinal parasitism, bacterial translocation to the liver, malakoplakia and iron sequestration, none of which occurred in non-progressing murine VL. Diverse bacterial genera were cultured from VL affected livers, of which Rodentibacter was specifically absent from ABX-treated hamsters, indicating this pathobiont may play a role in promoting disease progression. The results provide experimental support for antibiotic prophylaxis against secondary bacterial infections as an adjunct therapy in human VL patients. Author summary: Visceral leishmaniasis (VL) is a potentially fatal neglected infectious disease that is widespread in South Asia, East Africa, South America and the Mediterranean region. VL is caused by infections with the protozoan parasite Leishmania donovani. Infected people may remain asymptomatic or progress, at variable rates, to develop VL disease affecting the liver, spleen and blood. The reasons why clinical outcomes vary so much is not well understood. We decided to investigate whether a host's gut microbiota might be a relevant factor. We treated mice and hamsters with antibiotics to disrupt their gut microbiota and then infected them with L. donovani. No effect was observed in mice, however, treated hamsters had slower onset and progression of VL, less severe enlargement of the liver and spleen, and had a reduced mortality rate. This was accompanied by alterations in the levels of key immune response factors. We also found novel aspects of symptomatic VL in hamsters relating to the gut and its microbiota. Specifically, the gut tissues themselves were parasitized by L. donovani and liver tissues became co-infected with both L. donovani and Gram negative bacteria originating from the gut. Overall, our findings support the inclusion of anti-bacterial therapy as part of VL treatment strategies. [ABSTRACT FROM AUTHOR]

Visceral leishmaniasis (VL) is the most fatal form of leishmaniasis if left untreated and 50,000 to 90,000 new cases of VL occur worldwide each year. Although various vaccines had been studied in animal models, none of them was eligible to prevent human from infections. In this study, according to the silico analysis of Leishmania Amastin, Kmp-11 and Gp63 protein, dominant epitope sequences of these proteins were selected and linked to construct dominant multi-epitopes DNA and protein vaccines (Amastin-Kmp-11, Amastin-Gp63 and Kmp-11-Gp63) against VL. BALB/c mice were immunized with a DNA prime-protein boost immunization strategy and challenged with a new Leishmania parasite strain isolated from a VL patient. After immunization, the results including specific antibody titers, IL-4 and TNF-α levels, and CD4 and CD8 T cell proportion suggested the potent immunogenicity of the three vaccines. After infection, the results of spleen parasite burdens in the three vaccine groups were significantly lower than those of control groups, and the parasite reduction rates of Amastin-Kmp-11, Amastin-Gp63 and Kmp-11-Gp63 groups were 89.38%, 91.01% and 88.42%, respectively. Spleen smear observation and liver histopathological changes showed that all vaccine groups could produce significant immunoprotection against VL and Amastin-Gp63 vaccine was the best. In conclusion, our work demonstrated that the three dominant multi-epitopes Amastin-Kmp-11, Amastin-Gp63 and Kmp-11-Gp63 DNA prime-protein boost vaccines might be new vaccine candidates for VL, and the Amastin-Gp63 vaccine have best efficacy. [ABSTRACT FROM AUTHOR]

Visceral leishmaniasis is an infectious parasitic disease caused by the protozoan parasites Leishmania donovani and Leishmania infantum. The drugs currently used to treat visceral leishmaniasis suffer from toxicity and the emergence of parasite resistance, and so a better solution would be the development of an effective subunit vaccine; however, no approved vaccine currently exists. The comparative testing of a large number of vaccine candidates requires a quantitative and reproducible experimental murine infection model, but the parameters that influence infection pathology have not been systematically determined. To address this, we have established an infection model using a transgenic luciferase-expressing L. donovani parasite and longitudinally quantified the infections using in vivo bioluminescent imaging within individual mice. We examined the effects of varying the infection route, the site of adjuvant formulation administration, and standardised the parasite preparation and dose. We observed that the increase in parasite load within the liver during the first few weeks of infection was directly proportional to the parasite number in the initial inoculum. Finally, we show that immunity can be induced in pre-exposed animals that have resolved an initial infection. This murine infection model provides a platform for systematic subunit vaccine testing against visceral leishmaniasis. [ABSTRACT FROM AUTHOR]

Background: The chemotherapeutic arsenal available to treat visceral leishmaniasis is currently limited, in view of many drawbacks such as high cost, toxicity or emerging resistance. New therapeutic strategies are particularly needed to improve the management and the outcome in immunosuppressed patients. The combination of an immunomodulatory drug to a conventional anti-Leishmania treatment is an emerging concept to reverse the immune bias from Th2 to Th1 response to boost healing and prevent relapses. Methods: Here, immunostimulating and leishmanicidal properties of octyl-β-d-galactofuranose (Galf) were assessed in human monocyte-derived macrophages (HM) and in a murine model, after challenge with Leishmania donovani promastigotes. We recorded parasite loads and expression of various cytokines and immune effectors in HM and mouse organs (liver, spleen, bone marrow), following treatment with free (Galf) and liposomal (L-Galf) formulations. Results: Both treatments significantly reduced parasite proliferation in HM, as well as liver parasite burden in vivo (Galf, P < 0.05). Consistent with in vitro results, we showed that Galf- and L-Galf-treated mice displayed an enhanced Th1 immune response, particularly in the spleen where pro-inflammatory cytokines TNF-α, IL-1β and IL-12 were significantly overexpressed compared to control group. The hepatic recruitment of myeloid cells was also favored by L-Galf treatment as evidenced by the five-fold increase of myeloperoxidase (MPO) induction, which was associated with a higher number of MPO-positive cells within granulomas. By contrast, the systemic level of various cytokines such as IL-1β, IL-6, IL-17A or IL-27 was drastically reduced at the end of treatment. Conclusions: Overall, these results suggest that Galf could be tested as an adjuvant in combination with current anti-parasitic drugs, to restore an efficient immune response against infection in a model of immunosuppressed mice. [ABSTRACT FROM AUTHOR]

Background and Objectives Parasitic infections are one of the most important adverse reactions of blood transfusion. Several strategies are being implemented in blood transfusion centers of the world to prevent the transfusion-transmitted parasitic infections (TTPIs). The objective of this work was to clarify and describe the strategies to minimize the transmission risk of endemic parasitic agents in Iran via blood transfusion. Materials and Methods This study was conducted in the Iranian blood transfusion organization (IBTO). The data were extracted from the latest version of the "medical interview" standard operating procedure (SOP) and then recorded in the prepared sheets. Results The donor selection is the first and only step to reduce the risk of TTPIs in endemic and non-endemic areas of Iran. In all blood transfusion centers of the IBTO, the blood donation volunteers with a previous history of malaria, Chagas disease, visceral leishmaniasis (VL), muco-cutaneous leishmaniasis and babesiosis, as well as those with toxoplasmosis, cutaneous leishmaniasis (CL) and with a history of residence in, or travel to, malaria-endemic areas are permanently or temporarily deferred from the blood donation. Conclusion For some reasons, including the prevalence of malaria, toxoplasmosis and VL in parts of Iran, the donor selection strategy is not sufficient to prevent the TTPIs. Therefore, the changing of donor selection process and the use of other common preventive strategies are recommended to reduce the risk of TTPIs, especially for high-risk groups of toxoplasmosis and VL. [ABSTRACT FROM AUTHOR]

Background: Tumor necrosis factor alpha (TNF-α) blockers are recognized as a risk factor for reactivation of granulomatous infections. Leishmaniasis has been associated with the use of these drugs, although few cases have been reported. Methodology: We performed a retrospective observational study including patients with confirmed leishmaniasis acquired in the Mediterranean basin that were under TNF-α blockers therapy at the moment of the diagnosis. Patients diagnosed in our hospital from 2008 to 2018 were included. Moreover, a systematic review of the literature was performed and cases fulfilling the inclusion criteria were also included. Principal findings: Forty-nine patients were analyzed including nine cases from our series. Twenty-seven (55.1%) cases were male and median age was 55 years. Twenty-five (51%) patients were under infliximab treatment, 20 (40.8%) were receiving adalimumab, 2 (4.1%) etanercept, one (2%) golimumab and one (2%) a non-specified TNF-α blocker. Regarding clinical presentation, 28 (57.1%) presented as cutaneous leishmaniasis (CL), 16 (32.6%) as visceral leishmaniasis (VL) and 5 (10.2%) as mucocutaneous leishmaniasis (MCL). All VL and MCL patients were treated with systemic therapies. Among CL patients, 13 (46.4%) were treated with a systemic drug (11 received L-AmB, one intramuscular antimonials and one miltefosine) while 14 (50%) patients were given local treatment (13 received intralesional pentavalent antimonials, and one excisional surgery). TNF-α blockers were interrupted in 32 patients (65.3%). After treatment 5 patients (10.2%) relapsed. Four patients with a CL (3 initially treated with local therapy maintaining TNF-α blockers and one treated with miltefosine) and one patient with VL treated with L-AmB maintaining TNF-α blockers. Conclusions: This data supports the assumption that the blockage of TNF-α modifies clinical expression of leishmaniasis in endemic population modulating the expression of the disease leading to atypical presentations. According to the cases reported, the best treatment strategy would be a systemic drug and the discontinuation of the TNF-α blockers therapy until clinical resolution. [ABSTRACT FROM AUTHOR]

Purpose. To determine the values of prognostic nutritional and inflammatory markers in chemotherapy outcomes and survival in the patients with advanced nonsmall cell lung cancer (NSCLC) and also in the secondary malnutrition and cachexia. Methods. Twenty-five patients with diagnosis of aNSCLC were registered for the prospective study. Malnutrition was determined by the Subjective Global Assessment (SGA) and performance status by criteria of the Eastern Cooperative Oncology Group (ECOG). Before treatment, serum levels of albumin, prealbumin, vitamin D, zinc (Zn), C-reactive protein (CRP), IL-6, IL-1 β, TNF-α, lipoprotein lipase (LPL), and the Glasgow Prognostic Score (GPS) were recorded. Patients were followed prospectively for treatment outcomes and survival. Results. Due to the deaths of 18 patients during the 4-month follow-up period, no adequate measurements of inflammatory and nutritional markers could be performed. However, seven patients completed the treatment period and evaluations of these markers could be performed during the three periods. Eighty-four percent of patients were male with a mean age of 63.3 ± 8.7 years. Evaluation of the malnutrition by SGA showed that 5 (20%) patients were well nourished (A), 12(48%) were moderately malnourished (B), and 8(32%) were severely malnourished (C). Low levels of serum albumin (<3.5g/dl), prealbumin (<20 mg/ml), 25-hydroxycholecalciferol (<30 ng/ml), and Zn (<70mg/ml) were detected in 15(60%), 17(68%), 24 (96%), and 22 (88%) patients, respectively. Elevated levels of CRP (≥10 mg/L), IL6 (≥18pg/ml), TNF-α (≥24pg/ml), IL-1β (≥10pg/ml), and LPL (<12pg/ml) were found in 24 (96%), 11(44%), 9(36), 13(52%), and 11(44%) patients, respectively. Moderate and severe malnutrition, acute phase response, and reduced survival were determined in patients with NCSLC. In 7 patients that completed the treatment period, there was an association between elevated serum levels of IL-6, IL-1β, TNF-α, CRP, and LPL and also the reduced serum levels of albumin, prealbumin, Zn, vitamin D, and GPS, respectively. Similarly, Friedman analysis indicated that prealbumin significantly increased (p=0.007) in the follow-up period. But the serum levels of CRP (mean 37.3±22.3; Wilcoxon test P=0.368) in the seven patients were lower than those of the 18 patients that expired (mean 75.82±56.2). Conclusion. Malnutrition and cachexia negatively influence oncological outcomes in patients with NSCLC. These nutritional/inflammatory markers may be useful for selection of high risk and reduced survival in patients with aNSCLC undergoing adjuvant chemotherapy. [ABSTRACT FROM AUTHOR]

As the United States military engage with different countries and cultures throughout the world, personnel become exposed to new biospheres as well. There are many infectious pathogens that are not endemic to the US, but two of particular importance are Plasmodium and Leishmania, which respectively cause malaria and leishmaniasis. These parasites are both known to cause significant disease burden in their endemic locales, and thus pose a threat to military travelers. This review introduces readers to basic life cycle and disease mechanisms for each. Local and military epidemiology are described, as are the specific actions taken by the US military for prevention and treatment purposes. Complications of such measures with regard to human health are also discussed, including possible chemical toxicities. Additionally, poor recognition of these diseases upon an individual's return leading to complications and treatment delays in the United States are examined. Information about canine leishmaniasis, poorly studied relative to its human manifestation, but of importance due to the utilization of dogs in military endeavors is presented. Future implications for the American healthcare system regarding malaria and leishmaniasis are also presented. [ABSTRACT FROM AUTHOR]

The aim of this study was to evaluate mannose-anchored thiolated chitosan (MTC) based nanocarriers (NCs) for enhanced permeability, improved oral bioavailability and anti-parasitic potential of amphotericin B (AmB). Transgenic Leishmania donovani parasites expressing red fluorescent protein DsRed2 and imaging-flow cytometry was used to investigate parasitic burdens inside bone marrow-derived macrophages ex vivo. Cytokine estimation revealed that MTC nanocarriers activated the macrophages to impart an explicit immune response by higher production of TNF-α and IL-12 as compared to control. Cells treated with MTC NCs showed a significantly higher magnitude of nitrite and propidium iodide (PI) fluorescence intensity in contrast to cells treated with AmB. Concerning to apparent permeability coefficient (Papp) results, the MTC NCs formulation displayed more specific permeation across the Caco-2 cell monolayer as compared to AmB. The half-life of MTC NCs was about 3.3-fold persistent than oral AmB used as positive control. Also, t oral bioavailability of AmB was increased to 6.4-fold for MTC NCs compared to AmB for positive control. Acute oral evaluation indicated that MTC NCs were significantly less toxic compared to the AmB. Based on these findings, MTC NCs seems to be promising for significant oral absorption and improved oral bioavailability of AmB in leishmaniasis chemotherapy. [ABSTRACT FROM AUTHOR]

Leishmania braziliensis is an intracellular parasite that resides mostly in macrophages. Both the parasite genome and the clinical disease manifestations show considerable polymorphism. Clinical syndromes caused by L. braziliensis include localized cutaneous (CL), mucosal (ML), and disseminated leishmaniasis (DL). Our prior studies showed that genetically distinct L. braziliensis clades associate with different clinical types. Herein, we hypothesized that: (1) L. braziliensis induces changes in macrophage gene expression that facilitates infection; (2) infection of macrophages with strains associated with CL (clade B), ML (clade C), or DL (clade A) will differentially affect host cell gene expression, reflecting their different pathogenic mechanisms; and (3) differences between the strains will be reflected by differences in macrophage gene expression after initial exposure to the parasite. Human monocyte derived macrophages were infected with L. braziliensis isolates from clades A, B, or C. Patterns of gene expression were compared using Affymetrix DNA microarrays. Many transcripts were significantly decreased by infection with all isolates. The most dramatically decreased transcripts encoded proteins involved in signaling pathways, apoptosis, or mitochondrial oxidative phosphorylation. Some transcripts encoding stress response proteins were up-regulated. Differences between L. braziliensis clades were observed in the magnitude of change, rather than the identity of transcripts. Isolates from subjects with metastatic disease (ML and DL) induced a greater magnitude of change than isolates from CL. We conclude that L. braziliensis enhances its intracellular survival by inhibiting macrophage pathways leading to microbicidal activity. Parasite strains destined for dissemination may exert a more profound suppression than less invasive L. braziliensis strains that remain near the cutaneous site of inoculation. [ABSTRACT FROM AUTHOR]