Showing papers on "Visceral leishmaniasis published in 2022"

Canine Leishmaniasis: Update on Epidemiology, Diagnosis, Treatment, and Prevention

Abstract: Simple Summary Dogs are the main host of Leishmania infantum, a parasite that causes an incurable disease called canine leishmaniasis. This parasite is transmitted through the bite of a sandfly (a small insect related to mosquitoes and flies) in tropical and subtropical countries, but direct transmission between dogs, and from pregnant dogs to their puppies, exists. We reviewed the advances in tools and techniques for the surveillance of the disease, its diagnosis, treatment, and prevention. Canine leishmaniasis is expanding to the Northern Hemisphere, where it is barely known, due to climate change and the importation of dogs. Surveillance is therefore necessary in order to determine the extent of the disease in these areas and to monitor the appearance of the sandflies. Molecular techniques and rapid serological tests are now widespread for diagnosis and epidemiological studies. Several vaccines have been developed in the last decade, and even though their efficacy is limited, these advances will pave the way for the development of better vaccines against Leishmania and other parasites. Although new pharmacological tools are available, we are still waiting for the ideal drug that can eliminate the parasite from target organs and limit transmission to sandflies, without the side effects of current antileishmanials. Abstract Dog are the main reservoir of Leishmania infantum, causing canine leishmaniasis, an incurable multisystemic disease that leads to death in symptomatic dogs, when not treated. This parasite causes visceral, cutaneous, and mucosal leishmaniasis in people in the Mediterranean Basin, North Africa, South America, and West Asia. This disease is mostly unknown by veterinarians outside the endemic areas, but the disease is expanding in the Northern Hemisphere due to travel and climate change. New methodologies to study the epidemiology of the disease have found new hosts of leishmaniasis and drawn a completely new picture of the parasite biological cycle. Canine leishmaniasis diagnosis has evolved over the years through the analysis of new samples using novel molecular techniques. Given the neglected nature of leishmaniasis, progress in drug discovery is slow, and the few drugs that reach clinical stages in humans are unlikely to be commercialised for dogs, but several approaches have been developed to support chemotherapy. New-generation vaccines developed during the last decade are now widely used, along with novel prevention strategies. The implications of the epidemiology, diagnosis, treatment, and prevention of canine leishmaniasis are fundamental to public health.

Formulation of Amphotericin B in PEGylated Liposomes for Improved Treatment of Cutaneous Leishmaniasis by Parenteral and Oral Routes

Abstract: Liposomal amphotericin B (AmB) or AmBisome® is the most effective and safe therapeutic agent for visceral leishmaniasis (VL), but its clinical efficacy is limited in cutaneous leishmaniasis (CL) and HIV/VL co-infection. The aim of this work was to develop a formulation of AmB in PEGylated liposomes and compare its efficacy to AmBisome® in a murine model of CL. Formulations of AmB in conventional and PEGylated liposomes were characterized for particle size and morphology, drug encapsulation efficiency and aggregation state. Those were compared to AmBisome® in Leishmania amazonensis-infected BALB/c mice for their effects on the lesion size growth and parasite load. The conventional and PEGylated formulations showed vesicles with 100–130 nm diameter and low polydispersity, incorporating more than 95% of AmB under the non-aggregated form. Following parenteral administration in the murine model of CL, the PEGylated formulation of AmB significantly reduced the lesion size growth and parasite load, in comparison to control groups, in contrast to conventional liposomal AmB. The PEGylated formulation of AmB was also effective when given by oral route on a 2-day regimen. This work reports for the first time that PEGylated liposomal AmB can improve the treatment of experimental cutaneous leishmaniasis by both parenteral and oral routes.

Host–Pathogen Interaction in Leishmaniasis: Immune Response and Vaccination Strategies

Abstract: Leishmaniasis is a zoonotic and vector-borne infectious disease that is caused by the genus Leishmania belonging to the trypanosomatid family. The protozoan parasite has a digenetic life cycle involving a mammalian host and an insect vector. Leishmaniasisis is a worldwide public health problem falling under the neglected tropical disease category, with over 90 endemic countries, and approximately 1 million new cases and 20,000 deaths annually. Leishmania infection can progress toward the development of species–specific pathologic disorders, ranging in severity from self-healing cutaneous lesions to disseminating muco-cutaneous and fatal visceral manifestations. The severity and the outcome of leishmaniasis is determined by the parasite’s antigenic epitope characteristics, the vector physiology, and most importantly, the immune response and immune status of the host. This review examines the nature of host–pathogen interaction in leishmaniasis, innate and adaptive immune responses, and various strategies that have been employed for vaccine development.

Guideline for the Treatment of Leishmaniasis in the Americas. Second Edition

Abstract: Leishmaniasis is a neglected infectious disease of great importance in the Region of the Americas because of its prevalence, wide geographical distribution, morbidity and mortality. Several species of Leishmania can cause disease, and the resulting presentations differ in their clinical manifestations, diagnostic signs, severity, and treatment responses. The three main forms of leishmaniasis disease are: cutaneous, mucosal or visceral, of which cutaneous leishmaniasis is the most common. Visceral leishmaniasis (caused by L. infantum) is the most severe form and can cause death in up to 90% of untreated people. In 2013, PAHO, with the support of the Spanish Agency for International Development Cooperation, developed recommendations for the treatment of leishmaniasis in the Americas using the Grading of Recommendations Assessment, Development and Evaluation (GRADE) methodology. Since that time, new evidence has accumulated necessitating a revision of these recommendations. This publication, the second edition of the treatment guidelines for the Americas, has been put together with the leadership of PAHO’s Regional Program for Leishmaniasis with the support of the World Health Organization. It presents updated therapeutic recommendations for all three major forms of leishmaniasis disease, detailing the schemes and criteria for indication of treatment in the regional context. There are several notable changes from the first edition. For cutaneous disease, ketoconazole has been removed from the list of treatment options. Evidence for thermotherapy and pentamidine isethionate has been upgraded to “conditional” from “weak” in the previous edition. The number of Leishmania species for which there is strong evidence of the effectiveness of miltefosine has increased from two to four. And the evidence for intralesional antimonials in this disease form is now strong, whereas previously it was considered weak. The evidence for treatments for mucosal leishmaniasis – which is now considered separately to cutaneous disease – has become stronger since the first analysis, with the recommendation for use of pentavalent antimonials with or without oral pentoxifylline now strong. For visceral disease, the evidence has moved in the other direction. Whereas in the first edition, the evidence was considered strong for pentavalent antimonials, amphotericina B deoxycholate, it is now conditional. For miltefosine, there is now strong evidence against its useage. Further changes include the division of recommendations by adult and pediatric populations and the addition of new specific recommendations for immunocompromised patients that were not available in the first edition, including a strong recommendation against the use of pentavalent antimonials.

Visceral Leishmaniasis: Epidemiology, Diagnosis, and Treatment Regimens in Different Geographical Areas with a Focus on Pediatrics

Abstract: Visceral Leishmaniasis (VL) is a vector-borne disease caused by an intracellular protozoa of the genus Leishmania that can be lethal if not treated. VL is caused by Leishmania donovani in Asia and in Eastern Africa, where the pathogens’ reservoir is represented by humans, and by Leishmania infantum in Latin America and in the Mediterranean area, where VL is a zoonotic disease and dog is the main reservoir. A part of the infected individuals become symptomatic, with irregular fever, splenomegaly, anemia or pancytopenia, and weakness, whereas others are asymptomatic. VL treatment has made progress in the last decades with the use of new drugs such as liposomal amphotericin B, and with new therapeutic regimens including monotherapy or a combination of drugs, aiming at shorter treatment duration and avoiding the development of resistance. However, the same treatment protocol may not be effective all over the world, due to differences in the infecting Leishmania species, so depending on the geographical area. This narrative review presents a comprehensive description of the clinical picture of VL, especially in children, the diagnostic approach, and some insight into the most used pharmacological therapies available worldwide.

Surveillance of leishmaniasis cases from 15 European centres, 2014 to 2019: a retrospective analysis

Abstract: BackgroundSurveillance of human leishmaniasis in Europe is mostly limited to country-specific information from autochthonous infections in the southern part. As at the end of 2021, no integrated analysis has been performed for cases seen across centres in different European countries.AimTo provide a broad perspective on autochthonous and imported leishmaniasis cases in endemic and non-endemic countries in Europe.MethodsWe retrospectively collected records from cutaneous, mucosal and visceral leishmaniasis cases diagnosed in 15 centres between 2014 and 2019. Centres were located in 11 countries: Belgium, France, Germany, Italy, the Netherlands, Norway, Portugal, Spain, Sweden, Switzerland and the United Kingdom. Data on country of infection, reason for travelling, infecting species, age and sex were analysed.ResultsWe obtained diagnostic files from 1,142 cases, of which 76%, 21% and 3% had cutaneous, visceral, and mucosal disease, respectively. Of these, 68% were men, and 32% women, with the median age of 37 years (range: 0-90) at diagnosis. Visceral leishmaniasis was mainly acquired in Europe (88%; 167/190), while cutaneous leishmaniasis was primarily imported from outside Europe (77%; 575/749). Sixty-two percent of cutaneous leishmaniasis cases from outside Europe were from the Old World, and 38% from the New World. Geographic species distribution largely confirmed known epidemiology, with notable exceptions.ConclusionsOur study confirms previous reports regarding geographic origin, species, and traveller subgroups importing leishmaniasis into Europe. We demonstrate the importance of pooling species typing data from many centres, even from areas where the aetiology is presumably known, to monitor changing epidemiology.

Long-term hematopoietic stem cells as a parasite niche during treatment failure in visceral leishmaniasis

Abstract: Abstract Given the discontinuation of various first-line drugs for visceral leishmaniasis (VL), large-scale in vivo drug screening, establishment of a relapse model in rodents, immunophenotyping, and transcriptomics were combined to study persistent infections and therapeutic failure. Double bioluminescent/fluorescent Leishmania infantum and L. donovani reporter lines enabled the identification of long-term hematopoietic stem cells (LT-HSC) as a niche in the bone marrow with remarkably high parasite burdens, a feature confirmed for human hematopoietic stem cells (hHSPC). LT-HSC are more tolerant to antileishmanial drug action and serve as source of relapse. A unique transcriptional ’StemLeish’ signature in these cells was defined by upregulated TNF/NF-κB and RGS1/TGF-β/SMAD/SKIL signaling, and a downregulated oxidative burst. Cross-species analyses demonstrated significant overlap with human VL and HIV co-infected blood transcriptomes. In summary, the identification of LT-HSC as a drug- and oxidative stress-resistant niche, undergoing a conserved transcriptional reprogramming underlying Leishmania persistence and treatment failure, may open therapeutic avenues for leishmaniasis.

Updates on Prevalence and Trend Status of Visceral Leishmaniasis at Two Health Facilities in Amhara Regional State, Northwest Ethiopia: A Retrospective Study

Abstract: Ethiopia is one of the countries accounted for over 90% of annual visceral leishmaniasis incidence. Despite this, yet there are no active and passive surveillance activities in the Amhara Region that will give up-to-date information about the disease status at the health facility levels. Therefore, this study aimed to report up-to-date information about visceral leishmaniasis and its trend status at two health facilities and the surrounding areas. A retrospective study from October 2017 to May 2021 was conducted by reviewing patient records at Metema and Addis Zemen Hospitals. Data on Sex, age, occupation, residence, month, year, and rK39 test results were collected using a questionnaire and were analyzed using Statistical Package for Social Sciences (SPSS) version 20. The chi-square test was used to see the association between variables. p < 0.05 was considered as statistically significant. Of the 2,703 visceral leishmaniasis suspected cases diagnosed with the rK39 test, 877 (32.4%) were confirmed (positive) cases. Monthly and yearly trends depicted that the largest number of suspected cases was reported in October and 2018, respectively. Daily laborers were the most affected individuals in Metema areas.

AmBisome Monotherapy and Combination AmBisome–Miltefosine Therapy for the Treatment of Visceral Leishmaniasis in Patients Coinfected With Human Immunodeficiency Virus in India: A Randomized Open-Label, Parallel-Arm, Phase 3 Trial

Abstract: Abstract Background Visceral leishmaniasis (VL) in patients with human immunodeficiency virus (HIV) presents an increasingly important patient cohort in areas where both infections are endemic. Evidence for treatment is sparce, with no high-quality studies from the Indian subcontinent. Methods This is a randomized, open-label, parallel-arm, phase 3 trial conducted within a single hospital in Patna, India. One hundred and fifty patients aged ≥18 years with serologically confirmed HIV and parasitologically confirmed VL were randomly allocated to 1 of 2 treatment arms, either a total 40 mg/kg intravenous liposomal amphotericin B (AmBisome; Gilead Pharmaceuticals) administered in 8 equal doses over 24 days or a total 30 mg/kg intravenous AmBisome administered in 6 equal doses given concomitantly with a total 1.4 g oral miltefosine administered through 2 daily doses of 50 mg over 14 days. The primary outcome was intention-to-treat relapse-free survival at day 210, defined as absence of signs and symptoms of VL or, if symptomatic, negative parasitological investigations. Results Among 243 patients assessed for eligibility, 150 were recruited between 2 January 2017 and 5 April 2018, with no loss to follow-up. Relapse-free survival at day 210 was 85% (64/75; 95% CI, 77–100%) in the monotherapy arm, and 96%, (72/75; 90–100%) in the combination arm. Nineteen percent (28/150) were infected with concurrent tuberculosis, divided equally between arms. Excluding those with concurrent tuberculosis, relapse-free survival at day 210 was 90% (55/61; 82–100%) in the monotherapy and 97% (59/61; 91–100%) in the combination therapy arm. Serious adverse events were uncommon and similar in each arm. Conclusions Combination therapy appears to be safe, well tolerated, and effective, and halves treatment duration of current recommendations. Clinical Trials Registration Clinical Trial Registry India (CTRI/2015/05/005807; the protocol is available online at https://osf.io/avz7r).

Paradoxical immune response in leishmaniasis: The role of toll‐like receptors in disease progression

Abstract: Toll‐like receptors (TLRs), members of pattern recognition receptors, are expressed on many cells of the innate immune system, and their engagements with antigens regulate specific immune responses. TLRs signalling influences species‐specific immune responses during Leishmania infection; thus, TLRs play a decisive role towards elimination or exacerbation of Leishmania infection. To date, there is no single therapeutic or prophylactic approach that is fully effective against leishmaniasis. An in‐depth understanding of the mechanisms by which Leishmania species evade, or exploit host immune machinery could lead to the development of novel therapeutic approaches for the prevention and management of leishmaniasis. In this review, the role of TLRs in the induction of a paradoxical immune response in leishmaniasis was discussed. This review focuses on highlighting the novel interplay of TLR2‐ /TLR9‐driven resistance or susceptibility to 5 clinically important Leishmania species in human. The activation of TLR2/TLR9 can induce diverse anti‐Leishmania activities depending on the species of infecting Leishmania parasite. Infection with L. infantum and L. mexicana initiates TLR2/9 activation leading to host protective immune response, while infection with L. major, L. donovani and L. amazonensis trigger either a TLR2‐ /9‐related protective or non‐protective immune responses. These findings suggest that TLR2 and TLR9 are targets worth pursuing either for modulation or blockage to trigger host protective immune response towards leishmaniasis.

Immunological factors, but not clinical features, predict visceral leishmaniasis relapse in patients co-infected with HIV

Abstract: •Visceral leishmaniasis patients co-infected with HIV suffer from frequent VL relapse•VL relapse is associated with low CD4+ T cell counts and low production of IFN-γ•VL relapse is associated with increased PD1 expression on CD4+ and CD8+ T cells•These three markers can predict whether and when these patients relapse Visceral leishmaniasis (VL) has emerged as a clinically important opportunistic infection in HIV patients, as VL/HIV co-infected patients suffer from frequent VL relapse. Here, we follow cohorts of VL patients with or without HIV in Ethiopia. By the end of the study, 78.1% of VL/HIV—but none of the VL patients—experience VL relapse. Despite a clinically defined cure, VL/HIV patients maintain higher parasite loads, lower BMI, hepatosplenomegaly, and pancytopenia. We identify three immunological markers associated with VL relapse in VL/HIV patients: (1) failure to restore antigen-specific production of IFN-γ, (2) persistently lower CD4+ T cell counts, and (3) higher expression of PD1 on CD4+ and CD8+ T cells. We show that these three markers, which can be measured in primary hospital settings in Ethiopia, combine well in predicting VL relapse. The use of our prediction model has the potential to improve disease management and patient care. Visceral leishmaniasis (VL) has emerged as a clinically important opportunistic infection in HIV patients, as VL/HIV co-infected patients suffer from frequent VL relapse. Here, we follow cohorts of VL patients with or without HIV in Ethiopia. By the end of the study, 78.1% of VL/HIV—but none of the VL patients—experience VL relapse. Despite a clinically defined cure, VL/HIV patients maintain higher parasite loads, lower BMI, hepatosplenomegaly, and pancytopenia. We identify three immunological markers associated with VL relapse in VL/HIV patients: (1) failure to restore antigen-specific production of IFN-γ, (2) persistently lower CD4+ T cell counts, and (3) higher expression of PD1 on CD4+ and CD8+ T cells. We show that these three markers, which can be measured in primary hospital settings in Ethiopia, combine well in predicting VL relapse. The use of our prediction model has the potential to improve disease management and patient care. Visceral leishmaniasis (VL) is one of the most neglected tropical diseases. An estimated 550 million individuals are at risk of VL in high-burden countries, and 17,082 new cases of VL were reported in 2018.1Ruiz-Postigo J.A. Grouta L. Jaina S. Global leishmaniasis surveillance, 2017–2018, and first report on 5 additional indicators.Wkly. Epidemiol. Rec. 2020; 25: 265-280Google Scholar These numbers are widely acknowledged to underestimate the real burden because of the remote location of areas endemic for VL and the lack of surveillance. VL inflicts an immense toll on the developing world and impedes economic development, with an estimated annual loss of 2.3 million disability-adjusted life-years2World Health OrganizationControl of the leishmaniases. Report of a meeting of the WHO Expert Committee on the Control of Leishmaniases, Geneva, 22–26 March 2010.2010http://apps.who.int/iris/bitstream/handle/10665/44412/WHO_TRS_949_eng.pdf;jsessionid=28E4D43831D1121B3EC6F3B6721BCD05?sequence=1Google Scholar In Ethiopia, VL is one of the most significant vector-borne diseases; >3.2 million people are at risk of infection.3Gadisa E. Tsegaw T. Abera A. Elnaiem D.E. den Boer M. Aseffa A. Jorge A. Eco-epidemiology of visceral leishmaniasis in Ethiopia.Parasit. Vectors. 2015; 8: 381Crossref PubMed Scopus (39) Google Scholar VL is caused by infections with parasites of the Leishmania donovani species complex, but the majority of infected individuals control parasite replication and do not progress to disease. Some individuals will progress and develop VL, which is characterized by hepatosplenomegaly, fever, anemia, and wasting; this stage of the disease is generally fatal if left untreated.4Davidson R.N. Visceral leishmaniasis in clinical practice.J. Infect. 1999; 39: 112-116Abstract Full Text PDF PubMed Scopus (24) Google Scholar,5van Griensven J. Diro E. Visceral leishmaniasis.Infect. Dis. Clin. North Am. 2012; 26: 309-322Abstract Full Text Full Text PDF PubMed Scopus (176) Google Scholar Following the HIV-1 pandemic, VL has emerged as an opportunistic infection: VL accelerates the progression of HIV infection to AIDS, and conversely, HIV infection increases the risk of developing symptomatic VL.6Alvar J. Cañavate C. Gutiérrez-Solar B. Jiménez M. Laguna F. López-Vélez R. Molina R. Moreno J. Leishmania and human immunodeficiency virus coinfection: the first 10 years.Clin. Microbiol. Rev. 1997; 10: 298-319Crossref PubMed Google Scholar,7Olivier M. Badaró R. Medrano F.J. Moreno J. The pathogenesis of Leishmania/HIV co-infection: cellular and immunological mechanisms.Ann. Trop. Med. Parasitol. 2003; 97: 79-98Crossref PubMed Scopus (57) Google Scholar Ethiopia has the highest rate of VL/HIV co-infections in Africa, with HIV present in up to 30% of VL cases.8Leta S. Dao T.H. Mesele F. Alemayehu G. Visceral leishmaniasis in Ethiopia: an evolving disease.PLoS Negl. Trop. Dis. 2014; 8: e3131Crossref PubMed Scopus (59) Google Scholar HIV co-infection present a major challenge in the prevention and control of VL9Diro E. Lynen L. Mohammed R. Boelaert M. Hailu A. van Griensven J. High parasitological failure rate of visceral leishmaniasis to sodium stibogluconate among HIV co-infected adults in Ethiopia.PLoS Negl. Trop. Dis. 2014; 8: e2875Crossref PubMed Scopus (47) Google Scholar,10Hurissa Z. Gebre-Silassie S. Hailu W. Tefera T. Lalloo D.G. Cuevas L.E. Hailu A. Clinical characteristics and treatment outcome of patients with visceral leishmaniasis and HIV co-infection in northwest Ethiopia.Trop. Med. Int. Health. 2010; 15: 848-855Crossref PubMed Scopus (68) Google Scholar: VL/HIV co-infected patients experience higher rates of treatment failure, drug toxicity, mortality, and VL relapse rates compared to patients with VL alone.10Hurissa Z. Gebre-Silassie S. Hailu W. Tefera T. Lalloo D.G. Cuevas L.E. Hailu A. Clinical characteristics and treatment outcome of patients with visceral leishmaniasis and HIV co-infection in northwest Ethiopia.Trop. Med. Int. Health. 2010; 15: 848-855Crossref PubMed Scopus (68) Google Scholar,11World Health OrganizationEthiopia.https://www.who.int/leishmaniasis/resources/ETHIOPIA.pdfGoogle Scholar In Ethiopia, >50% of VL/HIV co-infected patients will experience relapse of VL between 3 and 9 months post-antileishmania treatment12Diro E. Edwards T. Ritmeijer K. Fikre H. Abongomera C. Kibret A. Bardonneau C. Soipei P. Mutinda B. Omollo R. et al.Long term outcomes and prognostics of visceral leishmaniasis in HIV infected patients with use of pentamidine as secondary prophylaxis based on CD4 level: a prospective cohort study in Ethiopia.PLoS Negl. Trop. Dis. 2019; 13: e0007132Crossref PubMed Scopus (7) Google Scholar. The mechanisms accounting for the increased rate of VL relapse in VL/HIV co-infected patients are poorly characterized. Markers such as low CD4+ T cell counts, high parasite loads at the time of VL diagnosis and during follow-up, not undergoing antiretroviral therapy (ART) at the time of VL diagnosis, and Leishmania antigenuria have shown variable degrees of prediction accuracy.9Diro E. Lynen L. Mohammed R. Boelaert M. Hailu A. van Griensven J. High parasitological failure rate of visceral leishmaniasis to sodium stibogluconate among HIV co-infected adults in Ethiopia.PLoS Negl. Trop. Dis. 2014; 8: e2875Crossref PubMed Scopus (47) Google Scholar,13Cota G.F. de Sousa M.R. Rabello A. Predictors of visceral leishmaniasis relapse in HIV-infected patients: a systematic review.PLoS Negl. Trop. Dis. 2011; 5: e1153Crossref PubMed Scopus (116) Google Scholar, 14Abongomera C. Diro E. de Lima Pereira A. Buyze J. Stille K. Ahmed F. van Griensven J. Ritmeijer K. The initial effectiveness of liposomal amphotericin B (AmBisome) and miltefosine combination for treatment of visceral leishmaniasis in HIV co-infected patients in Ethiopia: a retrospective cohort study.PLoS Negl. Trop. Dis. 2018; 12: e0006527Crossref PubMed Scopus (14) Google Scholar, 15van Griensven J. Mengesha B. Mekonnen T. Fikre H. Takele Y. Adem E. Mohammed R. Ritmeijer K. Vogt F. Adriaensen W. Diro E. Leishmania Antigenuria to Predict Initial Treatment Failure and Relapse in Visceral Leishmaniasis/HIV Coinfected Patients: An Exploratory Study Nested Within a Clinical Trial in Ethiopia.Front. Cell. Infect. Microbiol. 2018; 8: 94Crossref PubMed Scopus (3) Google Scholar Another predictive marker of VL relapse in VL/HIV co-infected patients is a history of VL relapse.16ter Horst R. Collin S.M. Ritmeijer K. Bogale A. Davidson R.N. Concordant HIV infection and visceral leishmaniasis in Ethiopia: the influence of antiretroviral treatment and other factors on outcome.Clin. Infect. Dis. 2008; 46: 1702-1709Crossref PubMed Scopus (85) Google Scholar One of the main immunological characteristics of VL patients is their profound immunosuppression.17Hailu A. van Baarle D. Knol G.J. Berhe N. Miedema F. Kager P.A. T cell subset and cytokine profiles in human visceral leishmaniasis during active and asymptomatic or sub-clinical infection with Leishmania donovani.Clin. Immunol. 2005; 117: 182-191Crossref PubMed Scopus (60) Google Scholar These patients do not respond to the Leishmanin skin test, their peripheral blood mononuclear cells (PBMCs) have an impaired capacity to produce IFN-γ and to proliferate in response to Leishmania antigen; this dysfunctional response to antigenic challenge is restored following successful chemotherapy18Adem E. Tajebe F. Getahun M. Kiflie A. Diro E. Hailu A. Shkedy Z. Mengesha B. Mulaw T. Atnafu S. et al.Successful Treatment of Human Visceral Leishmaniasis Restores Antigen-Specific IFN-γ, but not IL-10 Production.PLoS Negl. Trop. Dis. 2016; 10: e0004468Crossref PubMed Scopus (19) Google Scholar and reviewed by Nylén and Sacks,19Nylén S. Sacks D. Interleukin-10 and the pathogenesis of human visceral leishmaniasis.Trends Immunol. 2007; 28: 378-384Abstract Full Text Full Text PDF PubMed Scopus (272) Google Scholar Goto and Prianti,20Goto H. Prianti Md. Immunoactivation and immunopathogeny during active visceral leishmaniasis.Rev. Inst. Med. Trop. São Paulo. 2009; 51: 241-246Crossref PubMed Scopus (83) Google Scholar and Kumar and Nylén.21Kumar R. Nylén S. Immunobiology of visceral leishmaniasis.Front. Immunol. 2012; 3: 251Crossref PubMed Scopus (112) Google Scholar The mechanisms leading to impaired T cell responses in VL patients remain to be fully understood. Our knowledge of the immunopathology of VL/HIV co-infections is particularly sparse. Based on the current literature, it appears that the failure to control parasite replication results in chronic inflammation that leads to exhaustion of the immune system and failure to generate efficient T cell responses. Little is known about the immunological parameters associated with successful therapy. At the end of treatment, the discharge of VL/HIV co-infected patients from the hospital is based on clinical and parasitological cure.22World Health OrganizationGuideline for diagnosis, treatment & prevention of leishmaniasis in Ethiopia, 2nd edition.2013https://www.who.int/leishmaniasis/burden/Guideline_for_diagnosis_treatment_and_prevention_of_leishmaniasis_in_Ethiopia.pdfGoogle Scholar However, no clinical sign predicts an increased risk of relapse.22World Health OrganizationGuideline for diagnosis, treatment & prevention of leishmaniasis in Ethiopia, 2nd edition.2013https://www.who.int/leishmaniasis/burden/Guideline_for_diagnosis_treatment_and_prevention_of_leishmaniasis_in_Ethiopia.pdfGoogle Scholar Here, we followed VL and VL/HIV co-infected patients in Ethiopia and collected detailed clinical and immunological data during 12 months of follow-up. Genetic variation between parasites or re-infection of VL/HIV patients could be responsible for the increased rate of relapse. However, genomic data from isolates taken from the same patient cohorts as in the present study show that infections in VL and VL/HIV patients are caused by parasites from the same population and that almost all relapses are caused by recrudescence of the initial infection.23Franssen S.U. Takele Y. Adem E. Sanders M.J. Müller I. Kropf P. Cotton J.A. Diversity and Within-Host Evolution of Leishmania donovani from Visceral Leishmaniasis Patients with and without HIV Coinfection in Northern Ethiopia.MBio. 2021; 12: e0097121PubMed Google Scholar Here, we aimed to generate the most detailed picture to date of the natural history of VL and VL/HIV infections in Ethiopia and to identify clinical and immunological markers associated with VL relapse. These markers need to be suitable for measurement in a primary hospital setting in Ethiopia, so that they could contribute to the improved evaluation of treatment success and ultimately improve poor outcomes for these patients. We followed VL/HIV patients for up to 3 years and compared their rates of VL relapse with those of VL patients. During this study, 3 VL/HIV patients left the treatment center before the end of treatment and 5 died during treatment; 41 were treated successfully (i.e., had a negative test of cure [TOC]). Following antileishmanial treatment, 9 were lost to follow-up and 32 VL/HIV patients were followed for up to 3 years; of these, 25 (78.1%) experienced at least 1 episode of VL relapse. This was in sharp contrast to VL patients; 4 VL patients died during treatment and 10 were lost to follow-up, but none of the successfully treated VL patients experienced VL relapse (Figure 1A). The 46 VL patients who reached the end of the antileishmanial treatment responded well to their first course of treatment. In the VL/HIV group, 25 patients responded well to the first course of antileishmanial treatment, and had a negative TOC at the end of treatment, and were therefore discharged from the hospital. Sixteen VL/HIV patients responded poorly to the first line of treatment and had a positive TOC at end of treatment (EoT); they therefore required a longer course of antileishmanial drugs. We compared the duration of this initial treatment between VL/HIV patients who relapsed and those who did not relapse during follow-up: our results (Figure S1A) show that the duration of treatment was similar. Of the 16 patients who had a longer (>30 days) duration of treatment, 12 came back for follow-up and 4 were lost. As shown in Table S2, these 12 patients received different treatments. Parasite grades in VL/HIV patients at time of diagnosis (ToD) were significantly higher than those in VL patients (p < 0.001; Figure 1B), despite a similar duration of symptoms by patients in both groups (VL: 2.0 ± 0.2 versus VL/HIV: 2.0 ± 0.2 months, p > 0.05; Figure S1B). Since parasite grade is mainly measured at ToD, when the spleen is easily palpable, we used RNA sequencing (RNA-seq) to measure the total expression of L. donovani mRNAs (Ld mRNA) in blood (Figure 1C). In agreement with the parasite grades measured at ToD, there was significantly more Ld mRNA in VL/HIV patients; these levels decreased significantly in both cohorts of patients after antileishmanial treatment at EoT, but stayed significantly higher in VL/HIV patients. VL/HIV patients who relapsed during follow-up displayed significantly higher Ld mRNAs in blood than those without relapse (Figure 1D). Of note, by the end of our study, of the 5 VL/HIV patients who did not relapse but still had detectable Ld mRNA, 3 patients did not relapse and 2 were lost to follow-up (data not shown). There was no association in these 5 patients between the expression level of Ld mRNA and spleen sizes, interferon-γ (IFN-γ) level, and number of white and red blood cells and platelets (data not shown). Measurement of plasma HIV-1 viral load showed that despite undergoing ART, 58.9% of VL/HIV patients still had detectable viral loads (Table 1, viral load [copies/mL]). There were no significant differences in either viral load (Table 1, viral load HIV mRNA) or total expression of HIV mRNA between time points (Table 1, viral load [copies/mL]) or between patients with and without relapse during follow-up (Table 1, HIV mRNA). There was no correlation between Ld mRNAs and viral loads in VL/HIV patients who relapsed during follow-up (p = 0.3356, data not shown); similar results were obtained with the correlation between the total expression of HIV-1 and Ld mRNAs (p = 0.0745, data not shown). Of note, there was no systematic pattern in how viral loads varied through follow-up (Figure S2A).Table 1Viral load, clinical symptoms, and liver and kidney function testsToDEoT3 months6–12 monthspViral load (copies/mL)VL/HIV255 ± 402,369150 ± 115,9510.1 ± 61,3950.1 ± 200,7670.3632Viral load HIV mRNAVL/HIV673 ± 492,848150 ± 62,36675 ± 81,420273 ± 141,0250.4205No relapseRelapsepHIV mRNAVL/HIV0.1 ± 154.1397 ± 97,2020.0575FeverVL37.6 ± 0.236.0 ± 0.136.2 ± 0.136.0 ± 0.1< 0.0001VL/HIV36.6 ± 0.136.4 ± 0.136.3 ± 0.236.3 ± 0.1< 0.0001p0.02250.15020.48720.0233BMIVL16.3 ± 0.216.7 ± 0.318.5 ± 0.219.1 ± 0.3< 0.0001VL/HIV16.4 ± 0.317.0 ± 0.317.9 ± 0.317.1 ± 0.40.0010p0.79750.60940.03520.0050Epistaxis (%)Edema (%)Concomitant infections (%)Other symptomsVL9 (18.0)13 (26.0)15 (30.0)VL/HIV11 (22.4)4 (8.1)14(28.6)VL (%)VL/HIV (%)Concomitant infectionsPneumonia7 (14.0)6 (12.2)TB2 (4.0)5 (10.2)Intestinal parasites2 (4.0)0Malaria1 (2.0)2 (4.1)Herpes zoster1 (2.0)1 (2.0)Sepsis1 (2.0)0Viral hepatitis1 (2.0)0ToDEoTpSGPT (<43 U/L)VL34.0 ± 7.543.5 ± 5.7nsVL/HIV20.0 ± 2.128.5 ± 4.2nsp<0.0010.002SGOT (<38 U/L)VL62.0 ± 13.761.0 ± 5.2nsVL/HIV41.0 ± 4.935.0 ± 5.6nsp<0.001<0.001BUN (4.7–23.5 mg/dL)VL12.0 ± 1.99.0 ± 0.50.0022VL/HIV12.4 ± 1.113.8 ± 1.1nspns<0.001Creatinine (0.6–1.1 mg/dL)VL0.9 ± 0.10.8 ± 0.10.0092VL/HIV0.9 ± 0.21.0 ± 0.1nspns<0.001HIV-1 viral load (copies/mL) in plasma from VL/HIV (ToD: n = 39, EoT: n = 33, 3 months: n = 27, 6–12 months: n = 21) patients. Viral load HIV mRNA: quantification of the total expression of HIV mRNA in blood from VL/HIV patients (ToD: n = 35, EoT: n = 30, 3 months: n = 24, 6–12 months: n = 25). HIV mRNA: quantification of the total HIV mRNA expression in blood from VL/HIV patients who did not relapse (n = 13) and who relapsed (n = 24) after successful antileishmanial treatment (3 and 6–12 months) during the 3- and 6- to 12-month follow-up periods. If a patient did not relapse during the 2 time points of follow-up and if a patient relapsed at both 3 and 6–12 months, this is represented as 2 measurements. Fever: body temperature was measured on VL (ToD: n = 49, EoT: n = 45, 3 months: n = 36, 6–12 months: n = 26) and VL/HIV (ToD: n = 49, EoT: n = 38, 3 months: n = 32, 6–12 months: n = 26) patients and controls (n = 25). Other symptoms: numbers and percentages of VL and VL/HIV patients presenting at ToD with epistaxis, edema, or concomitant infections. Concomitant infections: numbers and percentages of VL and VL/HIV patients presenting with the different concomitant infections. SGPT: serum glutamic oxaloacetic transaminase; SGPT, serum glutamic pyruvic transaminase; BUN, blood urea nitrogen; creatinine: creatinine was measured in the plasma of VL patients as described in Method details. The values in italic and in parentheses represent the normal values. Statistical differences between VL and VL/HIV patients at each time point were determined using a Mann-Whitney test, and statistical differences between the 4 different time points for each cohort of patients were determined by the Kruskal-Wallis test. ToD, time of diagnosis; EoT, end of treatment; ns, not significant; 3 m, 3 months post-EoT; 6–12 m, 6–12 months post-EoT. Open table in a new tab HIV-1 viral load (copies/mL) in plasma from VL/HIV (ToD: n = 39, EoT: n = 33, 3 months: n = 27, 6–12 months: n = 21) patients. Viral load HIV mRNA: quantification of the total expression of HIV mRNA in blood from VL/HIV patients (ToD: n = 35, EoT: n = 30, 3 months: n = 24, 6–12 months: n = 25). HIV mRNA: quantification of the total HIV mRNA expression in blood from VL/HIV patients who did not relapse (n = 13) and who relapsed (n = 24) after successful antileishmanial treatment (3 and 6–12 months) during the 3- and 6- to 12-month follow-up periods. If a patient did not relapse during the 2 time points of follow-up and if a patient relapsed at both 3 and 6–12 months, this is represented as 2 measurements. Fever: body temperature was measured on VL (ToD: n = 49, EoT: n = 45, 3 months: n = 36, 6–12 months: n = 26) and VL/HIV (ToD: n = 49, EoT: n = 38, 3 months: n = 32, 6–12 months: n = 26) patients and controls (n = 25). Other symptoms: numbers and percentages of VL and VL/HIV patients presenting at ToD with epistaxis, edema, or concomitant infections. Concomitant infections: numbers and percentages of VL and VL/HIV patients presenting with the different concomitant infections. SGPT: serum glutamic oxaloacetic transaminase; SGPT, serum glutamic pyruvic transaminase; BUN, blood urea nitrogen; creatinine: creatinine was measured in the plasma of VL patients as described in Method details. The values in italic and in parentheses represent the normal values. Statistical differences between VL and VL/HIV patients at each time point were determined using a Mann-Whitney test, and statistical differences between the 4 different time points for each cohort of patients were determined by the Kruskal-Wallis test. ToD, time of diagnosis; EoT, end of treatment; ns, not significant; 3 m, 3 months post-EoT; 6–12 m, 6–12 months post-EoT. The following clinical and laboratory data were collected from each patient before the start of antileishmanial therapy. Results presented in Table 1 (fever) show that whereas both VL and VL/HIV patients had increased body temperatures at ToD (controls: 36.0°C ± 0.1°C, p < 0.0001), it was significantly lower in VL/HIV patients (p = 0.023) and decreased over time in both groups (p < 0.0001), but at 6–12 months was higher in VL/HIV patients as compared to VL patients (p = 0.023). There was no significant difference in the body temperature of VL/HIV patients with and without relapse during follow-up (data not shown). As shown in Figure 1E, spleen sizes were similarly increased in both cohorts of patients at ToD and decreased at EoT. These continued to decrease in VL patients but stayed higher in VL/HIV patients at 3 and 6–12 months. Spleens were significantly more enlarged in VL/HIV patients who relapsed (Figure 1F). The liver was also palpable at ToD in 40.0% of VL and 63.3% of VL/HIV patients and was significantly more enlarged in VL/HIV as compared to VL patients (p = 0.012); the liver size decreased significantly at EoT (Figure S1C). There was no significant difference in liver size in VL/HIV patients with and without relapse during follow-up (data not shown). The median BMI of patients with VL and VL/HIV was below 18.5 at ToD (VL patients: 16.4 ± 0.3, VL/HIV patients: 16.7 ± 0.3; Table 1, BMI). The BMI of VL patients increased over time and at 3 months, was similar to those of controls (controls: 19.9 ± 0.6, p > 0.05); in contrast, the BMI of the VL/HIV patients stayed significantly lower compared to VL patients and controls over time (Table 1, BMI). Of note, when compared at EoT, these clinical presentations were not significantly different in the VL/HIV groups that went on to relapse and those that did not (Table S3). Epistaxis, edema, and concomitant infections are clinical symptoms that are routinely recorded at ToD in these patients. As shown in Table 1 (other symptoms), there were no differences between the number of patients experiencing epistaxis or co-infections (pneumonia, tuberculosis [TB], intestinal parasites, malaria, herpes zoster, sepsis, or viral hepatitis; Table 1, concomitant infections), but there were more VL patients presenting with edema. Liver and kidney function are also routinely measured at ToD and EoT. Results show significantly lower levels of serum glutamic oxaloacetic transaminase (SGOT) and serum glutamic pyruvic transaminase (SGPT) in the VL/HIV groups as compared to the VL groups at both ToD and EoT (Table 1, SGOT and SGPT). The levels of blood urea nitrogen (BUN) and creatinine decreased significantly in the VL group at EoT but remained similar in the VL/HIV groups (Table 1, BUN and creatinine). White (WBC) and red blood cell (RBC) and platelet (PLT) counts were significantly decreased at ToD as compared to controls (Figures S3A, S3C, and S3E). At ToD, only PLT counts were higher in VL/HIV patients as compared to VL patients (Figure S3E). WBC counts increased in both groups at EoT, but thereafter only continued to increase in VL, but not in VL/HIV patients. Of note, WBC count levels in both VL and VL/HIV patients had not been restored 6–12 m post-EoT. Similarly, RBC and PLT counts increased in VL patients, and, in contrast to the counts in VL/HIV patients, were restored to levels similar to those of controls at EoT. During the follow-up of VL/HIV patients, WBC, RBC, and PLT counts were significantly lower in VL/HIV patients who relapsed as compared to those who did not relapse (Figures S3B, S3D, and S3F). In summary, the results from the clinical data show that over time, VL/HIV patients maintained higher parasite loads, hepatosplenomegaly, and lower BMI, and remained pancytopenic as compared to VL patients. Antigen-specific production of IFN-γ by whole blood cells was low at ToD in both cohorts of patients, but increased significantly in VL patients at EoT and was restored during follow-up (Figure 2A). In contrast, the levels of IFN-γ produced by whole blood (WB) cells from VL/HIV patients remained lower at all time points as compared to VL patients (Figure 2A). This was also true in the longitudinal follow-up of patients (Figures S2B and S2C, p = 0.0038 and p = 0.1682, respectively). We also compared the levels of antigen-specific IFN-γ in VL/HIV patients with and without relapse; in the longitudinal follow-up (Figure S2C), those who did not relapse produced significantly more IFN-γ (p = 0.0022). The median levels of IFN-γ produced by WB cells from patients who did not relapse after treatment were also significantly higher as compared to those who relapsed (Figure 2B) and were similar to those measured at ToD in VL patients (Figure 2A). There was a clear correlation between IFN-γ concentrations and parasite grade at ToD (Figures 2C and 2D). IFN-γ production in response to phytohemagglutinin (PHA) followed the same pattern as antigen-specific stimulation, failing to recover from a lower level in VL/HIV patients as compared to VL patients (Table 2, VL: IFN-γ, VL/HIV: IFN-γ, and PHA: IFN-γ).Table 2Production of IFN-γ and IL-10 in response to SLA and PHAToDEoT3 months6–12 monthspVL: IFN-γSLA28.4 ± 41.3115.4 ± 80.4972.2 ± 205.0898.4 ± 229.6<0.0001PHA30.6 ± 35.6177.8 ± 123.0190.8 ± 186.0389.5 ± 196.70.0001p0.67280.65520.06370.1971LV/HIV: IFN-γSLA0.3 ± 7.80.8 ± 15.50.4 ± 40.53.2 ± 58.40.3411PHA6.4 ± 65.933.1 ± 129.114.5 ± 101.114.2 ± 148.60.5850p0.00010.00020.02490.0524PHA: IFN-γVL30.6 ± 35.6177.8 ± 123.0190.8 ± 186.0389.5 ± 196.7VL/HIV6.4 ± 65.933.1 ± 129.114.5 ± 101.114.2 ± 148.6p0.10890.07640.0075<0.0001VL: IL-10SLA2.9 ± 1.30.1 ± 0.761.7 ± 1.30.3 ± 3.1PHA26.9 ± 19.4142.1 ± 30.2648.9 ± 67.4390.7 ± 38.7p0.0002<0.0001<0.0001<0.0001VL/HIV: IL-10SLA0.3 ± 3.12.6 ± 1.80.7 ± 0.70.2 ± 1.20.1524PHA25.4 ± 16.9227.3 ± 42.0319.2 ± 41.6255.5 ± 56.3<0.0001p<0.0001<0.0001<0.0001<0.0001PHA: IL-10VL26.9 ± 19.4142.1 ± 30.2648.9 ± 67.4390.7 ± 38.7VL/HIV25.4 ± 16.9227.3 ± 42.0319.2 ± 41.6255.5 ± 56.3p0.10890.07640.00290.1076VL: IFN-γ: whole blood cells from VL patients (ToD: n = 43, EoT: n = 44, 3 months: n = 30, 6–12 months: n = 44) were cultured in the presence of SLA and PHA, and IFN-γ levels in the supernatants were measured by ELISA after 24 h.;VL/HIV: IFN-γ: whole blood cells from VL/HIV patients (ToD: n = 39, EoT: n = 40, 3 months: n = 25, 6–12 months: n = 25) were cultured in the presence of SLA and PHA, and IFN-γ levels in the supernatants were measured by ELISA after 24 h; PHA: IFN-γ: comparison of the levels of IFN-γ produced in response to PHA between VL and VL/HIV patients; VL: IL-10: whole blood cells from VL patients (ToD: n = 43, EoT: n = 44, 3 months: n = 30, 6–12 months: n = 44) were cultured in the presence of SLA and PHA, and IL-10 levels in the supernatant were measured by ELISA after 24 h; VL/HIV: IL-10: whole blood cells from VL/HIV patients (ToD: n = 39, EoT: n = 40, 3 months: n = 25, 6–12 months: n = 25) were cultured in the presence of SLA and PHA and IL-10 levels in the supernatant were measured by ELISA after 24 h. PHA: IL-10: comparison of the levels of IL-10 produced in response to PHA between VL and VL/HIV patients. Statistical differences between VL and VL/HIV patients or between SLA and PHA at each time point were determined using a Mann-Whitney test and statistical differences between the 4 different time points for each cohort of patients were determined by the Kruskal-Wallis test. Open table in a new tab VL: IFN-γ: whole blood cells from VL patients (ToD: n = 43, EoT: n = 44, 3 months: n = 30, 6–12 months: n = 44) were cultured in the presence of SLA and PHA, and IFN-γ levels in the supernatants were measured by ELISA after 24 h.;VL/HIV: IFN-γ: whole blood cells from VL/HIV patients (ToD: n = 39, EoT: n = 40, 3 months: n = 25, 6–12 months: n = 25) were cultured in the presence of SLA and PHA, and IFN-γ levels in the supernatants were measured by ELISA after 24 h; PHA: IFN-γ: comparison of the levels of IFN-γ produced in response to PHA between VL and VL/HIV patients; VL: IL-10: whole blood cells from VL patients (ToD: n = 43, EoT: n = 44, 3 months: n = 30, 6–12 months: n = 44) were cultured in the presence of SLA and PHA, and IL-10 levels in the supernatant were measured by ELI

1,2,4-Trioxolane and 1,2,4,5-Tetraoxane Endoperoxides against Old-World Leishmania Parasites: In Vitro Activity and Mode of Action

Abstract: Leishmaniasis remains one of the ten Neglected Tropical Diseases with significant morbidity and mortality in humans. Current treatment of visceral leishmaniasis is difficult due to a lack of effective, non-toxic, and non-extensive medications. This study aimed to evaluate the selectivity of 12 synthetic endoperoxides (1,2,4-trioxolanes; 1,2,4,5-tetraoxanes) and uncover their biochemical effects on Leishmania parasites responsible for visceral leishmaniasis. The compounds were screened for in vitro activity against L. infantum and L. donovani and for cytotoxicity in two monocytic cell lines (J774A.1 and THP-1) using the methyl thiazol tetrazolium assay. Reactive oxygen species formation, apoptosis, and mitochondrial impairment were measured by flow cytometry. The compounds exhibited fair to moderate anti-proliferative activity against promastigotes of the 2 Leishmania species, with IC50 values ranging from 13.0 ± 1.7 µM to 793.0 ± 37.2 µM. Tetraoxanes LC132 and LC138 demonstrated good leishmanicidal activity on L. infantum amastigotes (IC50 13.2 ± 5.2 and 23.9 ± 2.7 µM) with low cytotoxicity in mammalian cells (SIs 22.1 and 118.6), indicating selectivity towards the parasite. Furthermore, LC138 was able to induce late apoptosis and dose-dependent oxidative stress without affecting mithocondria. Compounds LC132 and LC138 can be further explored as potential antileishmanial chemotypes.

Tissue Specific Dual RNA-Seq Defines Host–Parasite Interplay in Murine Visceral Leishmaniasis Caused by Leishmania donovani and Leishmania infantum

Abstract: Visceral leishmaniasis (VL) is caused by two species of Leishmania parasites, L. donovani in the Old World and L. infantum in the New World and countries bordering the Mediterranean. Although cardinal features such as hepato-splenomegaly and alterations in blood and immune function are evident, clinical presentation may vary by geography, with for example severe bleeding often associated with VL in Brazil. ABSTRACT Visceral leishmaniasis is associated with hepato-splenomegaly and altered immune and hematological parameters in both preclinical animal models and humans. We studied mouse experimental visceral leishmaniasis caused by Leishmania infantum and Leishmania donovani in BALB/c mice using dual RNA-seq to investigate the transcriptional response of host and parasite in liver and spleen. We identified only 4 species-specific parasite expressed genes (SSPEGs; log2FC >1, FDR <0.05) in the infected spleen, and none in the infected liver. For the host transcriptome, we found 789 differentially expressed genes (DEGs; log2FC >1, FDR <0.05) in the spleen that were common to both infections, with IFNγ signaling and complement and coagulation cascade pathways highly enriched, and an additional 286 and 186 DEGs that were selective to L. donovani and L. infantum infection, respectively. Among those, there were network interactions between genes of amino acid metabolism and PPAR signaling in L. donovani infection and increased IL1β and positive regulation of fatty acid transport in L. infantum infection, although no pathway enrichment was observed. In the liver, there were 1,939 DEGs in mice infected with either L. infantum or L. donovani in comparison to uninfected mice, and the most enriched pathways were IFNγ signaling, neutrophil mediated immunity, complement and coagulation, cytokine-chemokine responses, and hemostasis. Additionally, 221 DEGs were selective in L. donovani and 429 DEGs in L. infantum infections. These data show that the host response for these two visceral leishmaniasis infection models is broadly similar, and ∼10% of host DEGs vary in infections with either parasite species. IMPORTANCE Visceral leishmaniasis (VL) is caused by two species of Leishmania parasites, L. donovani in the Old World and L. infantum in the New World and countries bordering the Mediterranean. Although cardinal features such as hepato-splenomegaly and alterations in blood and immune function are evident, clinical presentation may vary by geography, with for example severe bleeding often associated with VL in Brazil. Although animal models of both L. donovani and L. infantum have been widely used to study disease pathogenesis, a direct side-by-side comparison of how these parasites species impact the infected host and/or how they might respond to the stresses of mammalian infection has not been previously reported. Identifying common and distinct pathways to pathogenesis will be important to ensure that new therapeutic or prophylactic approaches will be applicable across all forms of VL.

Wnt5A Signaling Blocks Progression of Experimental Visceral Leishmaniasis

Abstract: Visceral leishmaniasis, caused by L. donovani infection is fatal if left untreated. The intrinsic complexity of visceral leishmaniasis complicated further by the increasing emergence of drug resistant L. donovani strains warrants fresh investigations into host defense schemes that counter infections. Accordingly, in a mouse model of experimental visceral leishmaniasis we explored the utility of host Wnt5A in restraining L. donovani infection, using both antimony sensitive and antimony resistant L. donovani strains. We found that Wnt5A heterozygous (Wnt5A +/-) mice are more susceptible to L. donovani infection than their wild type (Wnt5A +/+) counterparts as depicted by the respective Leishman Donovan Units (LDU) enumerated from the liver and spleen harvested from infected mice. Higher LDU in Wnt5A +/- mice correlated with increased plasma gammaglobulin level, incidence of liver granuloma, and disorganization of splenic white pulp. Progression of infection in mice by both antimony sensitive and antimony resistant strains of L. donovani could be prevented by activation of Wnt5A signaling through intravenous administration of rWnt5A prior to L. donovani infection. Wnt5A mediated blockade of L. donovani infection correlated with the preservation of splenic macrophages and activated T cells, and a proinflammatory cytokine bias. Taken together our results indicate that while depletion of Wnt5A promotes susceptibility to visceral leishmaniasis, revamping Wnt5A signaling in the host is able to curb L. donovani infection irrespective of antimony sensitivity or resistance and mitigate the progression of disease.

Parasitological and immunological evaluation of a quinoline derivative salt incorporated into a polymeric micelle formulation against Leishmania infantum infection

Abstract: Leishmaniasis is a parasitic disease caused by Leishmania protozoa, which presents a large spectrum of clinical manifestations. In the present study, a quinoline derivative salt named N-(2-((7-chloroquinolin-4-yl)amino)ethyl)-N-(prop-2-yn-1-yl)prop-2-yn-1-aminium chloride or QDS3 was in vitro and in vivo tested against L. infantum by means of its incorporation in Poloxamer 407-based polymeric micelles (QDS3/M). The in vitro antileishmanial activity of QDS3 and QDS3/M was investigated in L. infantum promastigotes, axenic amastigotes and infected macrophages. BALB/c mice were infected with L. infantum, and parasitological parameters were evaluated 1 and 15 days post-treatment by determining the parasite load by a limiting dilution assay, besides a quantitative PCR (qPCR) method. Immunological response was assessed based on production of cellular cytokines, as well as by quantification of nitrite levels and specific antibodies. In vitro results showed that QDS3 free or in micelles presented effective antileishmanial action against both parasite stages, being more effective in amastigotes. In vivo data showed that treatment using QDS3 or QDS3/M reduced the parasite load in the livers, spleens, draining lymph nodes (dLN) and bone marrows of the treated animals, 1 and 15 days after treatment, when compared to values found in the control groups. Additionally, treated mice developed a polarized Th1-type immune response, with higher levels of IL-12, IFN-γ, GM-CSF and nitrite, besides high production of specific IgG2a antibodies, when compared to the controls. Parasitological and immunological data obtained using the micellar composition were better than the others. In conclusion, QDS3, mainly when applied in a delivery adjuvant system, could be considered for future studies as therapeutic candidate against VL.

A Case Report of Two Kala-azar Cases in China Diagnosed by Metagenomic Next-Generation Sequencing

Abstract: Background Leishmaniasis being a local disease, as kala-azar this particular form is a visceral form. It is transmitted by sandflies, and is a parasitic disease involving the reticuloendothelial system of mononuclear macrophages. Due to its poor prognosis and high fatality rate, the fatality rate of patients without effective treatment can exceed 95%. Thereby, early diagnosis and treatment can significantly improve its prognosis. The metagenomic next-generation sequencing (mNGS) has the advantage of being able to find pathogens that cannot be detected by traditional methods. More importantly, it can conduct nucleic acid detection of pathogens covering a wide range in a short time. For infectious diseases like kala-azar, which is clinically complicated and difficult, mNGS detection provides a basis for accurate etiological diagnosis. Case Report We report 2 cases of kala-azar in West China Hospital, Chengdu, China. The first case is a 47-year-old male patient who had recurrent fever for 4 months, accompanied by reduction of red blood cell, white blood cell, and blood platelet. He was detected by mNGS and clinically diagnosed as kala-azar (Leishmania detection), finally died of multiple organ failure. The second patient was a 15-year-old male who had fever for more than 10 days. He was detected by mNGS and clinically diagnosed as kala-azar (Leishmania detection). He recovered and discharged quickly after treatment with sodium stibogluconate. Conclusion Efforts should be made to improve early etiological diagnosis in order to improve patient prognosis. mNGS detection is beneficial to the diagnosis and treatment of infectious diseases with unknown causes in the early stage of emergency treatment.

Distance-based paper device using combined SYBR safe and gold nanoparticle probe LAMP assay to detect Leishmania among patients with HIV

Abstract: Abstract Asymptomatic visceral leishmaniasis cases increase continuously, particularly among patients with HIV who are at risk to develop further symptoms of leishmaniasis. A simple, sensitive and reliable diagnosis is crucially needed due to risk populations mostly residing in rural communities with limited resources of laboratory equipment. In this study, a highly sensitive and selective determination of Leishmania among asymptomatic patients with Leishmania/ HIV co-infection was achieved to simultaneously interpret and semi-quantify using colorimetric precipitates (gold-nanoparticle probe; AuNP-probe) and fluorescence (SYBR safe dye and distance-based paper device; dPAD) in one-step loop-mediated isothermal amplification (LAMP) assay. The sensitivities and specificities of 3 detection methods were equivalent and had reliable performances achieving as high as 95.5%. Detection limits were 10 2 parasites/mL (0.0147 ng/µL) which were 10 times more sensitive than other related studies. To empower leishmaniasis surveillance as well as prevention and control, this dPAD combined with SYBR safe and gold nanoparticle probe LAMP assay is reliably fast, simple, inexpensive and practical for field diagnostics to point-of-care settings in resource-limited areas which can be set up in all levels of healthcare facilities, especially in low to middle income countries.

Hemophagocytic lymphohistiocytosis and visceral leishmaniasis in children: a series of cases and literature review.

Abstract: Objective: Hemophagocytic lymphohistiocytosis syndrome (HLHS) is characterized by an immunological hyperactivation of cytotoxic T cells, natural killer cells, and macrophages, leading to the secretion of proinflammatory cytokines. HLHS associated with Visceral Leishmaniasis might be difficult to diagnose once symptoms are similar, resulting in the death of untreated patients. Our aim is to describe a series of cases of Visceral Leishmaniasis with HLHS admitted to a referral hospital for infectious diseases. Case description: All 115 cases of Visceral Leishmaniasis referred to a referral center for pediatric infectious diseases were reviewed to identify the cases of HLHS. Five cases (4.5%) were confirmed with HLHS and they presented fever, splenomegaly, cytopenia, hypertriglyceridemia or hypofibrinogenemia, increased ferritin and hemophagocytosis in the bone marrow. Comments: It important to rule out HLHS in children with infectious diseases that do not respond adequately to treatment or in patients with severe symptoms, especially in leishmaniasis endemic areas.%

Utilizing Quantitative Proteomics to Identify Species-Specific Protein Therapeutic Targets for the Treatment of Leishmaniasis

Abstract: Leishmaniasis is a tropical disease caused by Leishmania parasites, which are transmitted through the bites of infected sandflies. We focused on the emergence of leishmaniasis in Thailand caused by a species (Leishmania orientalis). Treatment by chemotherapy is not effective against L. orientalis. Hence, we intended to solve this issue using a proteomics approach to investigate protein profiles and in silico analysis for the identification of antigenic proteins from L. orientalis, Leishmania martiniquensis, and Leishmania donovani. Using principal component analysis (PCA), protein profile comparisons indicated that different species of Leishmania are different at the protein level. Proteomics analysis identified 6099 proteins. Among these proteins, 1065 proteins were used for further analysis. There were 16 proteins that were promising candidates for therapeutic aspects as they were abundantly expressed and common to all species. In silico analysis of protein’s antigenicity revealed that eight proteins had the potential for the development of antigenic molecules. Protein profile information and these antigenic proteins may play key roles in the pathogeny of leishmaniasis and can be used as novel therapeutic targets against leishmaniasis in the future.

Comparative Genomic Analyses of New and Old World Viscerotropic Leishmanine Parasites: Further Insights into the Origins of Visceral Leishmaniasis Agents

Abstract: Visceral leishmaniasis (VL), also known as kala-azar, is an anthropozoonotic disease affecting human populations on five continents. Aetiologic agents belong to the Leishmania (L.) donovani complex. Until the 1990s, three leishmanine parasites comprised this complex: L. (L.) donovani Laveran & Mesnil 1903, L. (L.) infantum Nicolle 1908, and L. (L.) chagasi Lainson & Shaw 1987 (=L. chagasi Cunha & Chagas 1937). The VL causal agent in the New World (NW) was previously identified as L. (L.) chagasi. After the development of molecular characterization, however, comparisons between L. (L.) chagasi and L. (L.) infantum showed high similarity, and L. (L.) chagasi was then regarded as synonymous with L. (L.) infantum. It was, therefore, suggested that L. (L.) chagasi was not native to the NW but had been introduced from the Old World by Iberian colonizers. However, in light of ecological evidence from the NW parasite’s enzootic cycle involving a wild phlebotomine vector (Lutzomyia longipalpis) and a wild mammal reservoir (the fox, Cerdocyon thous), we have recently analyzed by molecular clock comparisons of the DNA polymerase alpha subunit gene the whole-genome sequence of L. (L.) infantum chagasi of the most prevalent clinical form, atypical dermal leishmaniasis (ADL), from Honduras (Central America) with that of the same parasite from Brazil (South America), as well as those of L. (L.) donovani (India) and L. (L.) infantum (Europe), which revealed that the Honduran parasite is older ancestry (382,800 ya) than the parasite from Brazil (143,300 ya), L. (L.) donovani (33,776 ya), or L. (L.) infantum (13,000 ya). In the present work, we have now amplified the genomic comparisons among these leishmanine parasites, exploring mainly the variations in the genome for each chromosome, and the number of genomic SNPs for each chromosome. Although the results of this new analysis have confirmed a high genomic similarity (~99%) among these parasites [except L. (L.) donovani], the Honduran parasite revealed a single structural variation on chromosome 17, and the highest frequency of genomic SNPs (more than twice the number seen in the Brazilian one), which together to its extraordinary ancestry (382,800 ya) represent strong evidence that L. (L.) chagasi/L. (L.) infantum chagasi is, in fact, native to the NW, and therefore with valid taxonomic status. Furthermore, the Honduran parasite, the most ancestral viscerotropic leishmanine parasite, showed genomic and clinical taxonomic characteristics compatible with a new Leishmania species causing ADL in Central America.

Leishmaniasis control in the light of the COVID-19 pandemic in Africa

Abstract: Leishmaniasis is a parasitic disease, endemic to Africa, Asia, and South America due to inadequate access to medication and underreporting of leishmaniasis cases. Leishmaniasis has two forms: cutaneous and visceral. The fight against leishmaniasis has been greatly affected by the coronavirus disease 2019 (COVID-19) pandemic that impacted resource distribution and access to medication. Continuous effort in vaccine development and affordable therapeutics are necessary to eliminate leishmaniasis in low-income countries. Further research is necessary to determine molecular drug resistance markers in leishmaniasis patients. In this analysis, we focus on the effect of COVID-19 on leishmaniasis in Africa.

The current epidemiology of leishmaniasis in Turkey, Azerbaijan and Georgia and implications for disease emergence in European countries

Abstract: Leishmania spp. are sand fly‐borne protozoan parasites causing leishmaniasis in humans and animals. The aim of the study was to analyse the epidemiology of leishmaniasis in Turkey, Azerbaijan and Georgia from 2005 to 2020 and evaluate the associated risk for disease emergence in European countries. It is based on an analysis of WHO and OIE reported cases between 2005 and 2020, a review of scientific articles published in SCOPUS between 2009 and 2020 and a questionnaire survey to public health and veterinary authorities in these countries. Endemic Leishmania spp. include L. infantum in the three countries, L. major in Azerbaijan and Turkey and L. tropica and L. donovani in Turkey. Leishmaniasis is reported in humans, animals and sand flies and incidence is spatially and temporarily variable. In the southern Caucasus and particularly in Georgia, reported incidence of human visceral leishmaniasis by L. infantum remains high. However, whilst Georgia experienced a gradual decrease from >4.0 cases per 100,000 population in 2005–09 to 1.13 cases per 100,000 population in 2020, the period with highest incidence in Azerbaijan, which ranged between 0.40 and 0.61 cases per 100,000 population, was 2016–2019, and no cases have so far been reported for 2020. Visceral leishmaniasis in the Southern Caucasus affects mostly young children from deprived urban areas and its closely associated to canine leishmaniasis. Turkey reported cases of visceral leishmaniasis between 2005 and 2012 and in 2016 only, and incidence ranged between 0.02 and 0.05 per 100,000 population. In contrast, the reported annual incidence of cutaneous leishmaniasis in Turkey was much greater and peaked at 7.02 cases per 100,000 population in 2013, associated to imported cases from cutaneous leishmaniasis endemic Syria. Leishmaniasis by L. infantum in Azerbaijan and Georgia represents a regional public and animal health challenge that requires support to improve diagnosis, treatment and control. The unprecedented rise of cutaneous leishmaniasis and the spread of L. tropica and L. donovani in Turkey is an important risk factor for their emergence in Europe, especially in Mediterranean countries where competent vectors are widespread.