Form Of Visceral Leishmaniasis Research Articles

A novel outlook in the delivery of artemisinin: production and efficacy in experimental visceral leishmaniasis

ABSTRACT The visceral form of leishmaniasis (VL), due to infection by Leishmania infantum, is a neglected tropical disease. The accessible therapeutic options are limited. Artemisinin is an efficient antileishmanial product with poor biological availability that requires high repetition of therapeutic doses in VL. Solid lipid nanoparticles (SLNs) provide targeted delivery, increase bioavailability and reduce toxicity of the traditional therapeutic strategy. The spherical shape artemisinin-loaded SLNs were prepared in a particle diameter of 222.0 ± 14.0 nm. The SLNs showed no particular toxic effect on the parasites, whereas the native artemisinin demonstrated a significant toxicity rate of 31% in viability of the promastigotes at the 250 µg/ml concentration. The therapeutic efficacy of the artemisinin-loaded SLNs was demonstrated in the experimental VL, using the L. infantum-infected BALB/c mice, in the present study. The 10 and 20 mg/kg doses of artemisinin-loaded SLNs showed higher level of antileishmanial efficacy compared with the free artemisinin. There was a significant diminishing of the parasite burden in liver (84.7 ± 4.9%) and spleen (85.0 ± 3.1%) and hepatosplenomegaly by the artemisinin-loaded SLNs treated at 20 mg/kg compared to the free artemisinin. Therefore, the present study supports the superior efficacy of artemisinin-loaded SLNs over the free artemisinin and could be considered as a new therapeutic strategy in the treatment of leishmaniasis.

Combined Cutaneous & Visceral Forms of Leishmaniasis; Case Report

This is a three years old child from Himreen rural areas the patient had fever, pallor, & hepatosplenomegaly and two ulcers on the face. The patient responded to Pentostam. While we used in Iraq to see leishmaniasis as either pure cutaneous or pure visceral forms of leishmaniasis, this is a case with combined cutaneous & visceral forms of leishmaniasis in a patient with normal immune system. Which may explained by the leishmaniae species that primarily cause cutaneous disease & rarely cause visceral disease or by primarily viscerotropic leishma'nia that primarily cause visceral disease which may also cause skin lesions or by mixed infections by two different species of leishmaniae, one of them is dermatotropic, & the other one is viscerotropic.

Monitoring of leishmaniasis of dogs in the republic of Armenia

The authors analyzed the available literature data and statistics, and results can be found for leishmaniasis in Armenia. Between June 2019 and December 2021, 375 different breeds were deployed. 135 animals were subjected to a comprehensive study with laboratory diagnostics of blood serum for the detection of Leishmania infantum antigen. In 68 dogs, antigens were detected using the VetExpert Leishmania Ab immunochromatographic instrumentless test system. In the publication of the authors Yerevan and other cities of the republic. In the studies in animals affected by leishmaniasis, the cutaneous form of leishmaniasis (CL) prevailed, which was recorded in 85.3% of cases, and the visceral form of leishmaniasis (VL) was recorded in 14.7% of cases. Sexual and breed predisposition of the studied dogs to the disease with leishmaniasis was not found by the authors. The visceral form of leishmaniasis (VL) in the studies was recorded in dogs over the age of 4 years. The authors believe that in the near future it is necessary to conduct epidemiological, epizootological and entomological studies to continuously monitor the situation of leishmaniasis in the Republic of Armenia.

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

ABSTRACTVisceral 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.

Leishmania Vaccines: A Narrative Review on Last Decade Developments

Leishmaniasis is a vector-borne protozoan disease transferred by the sting of a female sandfly. Currently, both cutaneous and visceral forms of leishmaniasis (VL) are observed in Iran. Cutaneous leishmaniasis (CL) is reported in 18 provinces in the country in rural (zoonotic) and urban (anthroponotic) forms. VL is endemic in some areas of Iran. Various drugs are used in the treatment of leishmaniasis, but due to created challenges such as high toxicity, side effects, prolongation, and drug resistance, the process of curing will face problems. The prevention of infection is also costly. One of the best ways to control the disease is to find an effective vaccine. The efficient vaccine will cause long-term immunity and simultaneously breaks the disease transmission chain. This study aimed to review different types of existing vaccines, along with those in trials. It was found that multi anti-gene vaccines created the best immunity against the parasite and could be more successful. Developments in genetic engineering and release systems have raised hope to achieve the vaccines soon.

Adjuvant effects of TLR agonist gardiquimod admixed with Leishmania vaccine in mice model of visceral leishmaniasis.

Tropical and subtropical areas of the world are affected by leishmaniasis, which is caused by Leishmania spp. It has been categorized as an NTD (neglected tropical disease) because of its negligence. The sand fly of genus Phlebotomus acts as the vector for the transmission of the promastigote form of this protozoan parasite to the mammalian host where it converts to amastigote form in the macrophages. Visceral form of leishmaniasis (VL) is a deadly infection in the endothelial system of the human and other mammals. Only a few chemotherapeutic agents are available for the treatment of this infectious disease whereas no vaccine is available for the control of leishmanial infection. Therefore in the current study, we have tested the effects of gardiquimod (a TLR agonist) as an adjuvant in combination with the formalin-killed antigen of L. donovani as a vaccine. The mice were vaccinated thrice at an interval of 2weeks and challenged with L. donovani promastigotes after 2weeks of the last vaccination. We assessed the parasite load, delayed-type hypersensitivity (DTH) responses, humoral and cell-mediated immune response in BALB/c mice before and after challenge infection with L. donovani. Immunized mice were found to have the least parasite load, high DTH response, elevated levels of Th1 cytokines, IgG2a, and nitric oxide than non-immunized and infected control mice. The efficacy of the vaccine was boosted with the use of adjuvant gardiquimod that depicts its potential as an adjuvant in this study. Our study is reporting the adjuvant effects of gardiquimod for the first time. Further studies using other Leishmania species can be performed to signify its role.

Identification of potential Leishmania chagasi superoxide dismutase allosteric modulators by structure-based computational approaches: homology modelling, molecular dynamics and pharmacophore-based virtual screening

The visceral form of Leishmaniasis, also known as kala-azar, caused by Leishmania chagasi is the main etiological agent of this form in Brazil responsible for 30,000 annual deaths. Despite its epidemiological impact, treatment of the disease is limited by resistance, species-dependent efficacy and serious adverse effects. The application of computational tools to prioritize potential bioactive molecules based on 3D structural of biological target is a viable alternative. Among the L. chagasi validated targets, Fe + 2 superoxide dismutase B2 (LcFeSODB2) is the first parasite enzyme against oxidative stress and it is involved in essential metabolic processes for its survival. Due to substrate binding-site volume (superoxide ion) and consequent difficulty in its active site modulation for small molecules, the search for allosteric sites at LcFeSODB2 3D structure is a promising strategy. As there are no 3D structures of LcFeSODB2, comparative modeling was applied to build 3D models by SWISS-MODEL and MODELLER version 9.19. Next, the best 3D model was used in molecular dynamics (MD) routines with multiple probes on GROMACS version 5.1.2. In addition, potential allosteric sites predicted by FTMap and Metapocket web servers were used with probe occupancy maps from MD to select an allosteric binding site and propose a pharmacophore model. Next, it was used as a template in virtual screening by UNITY® module available on SYBYL-X version 2.1.1 at Sigma-Aldrich CPR™ subset of ZINC12 database. The pharmacophore-based virtual screening resulted in the selection of two potential allosteric LcFeSOD compounds with partial pharmacophoric requirements, drug-like properties and commercial availability for enzymatic assays. Communicated by Ramaswamy H. Sarma

PECULIARITIES OF MANIFESTATION OF THE SKIN FORM OF LEISHMANIASIS OF MAXILLOFACIAL LOCALIZATION

Leishmaniasis is a vector-borne disease in humans. Simple intracellular parasites cause the disease. Risk factors: socio-economic conditions, poverty. The disease is associated with malnutrition, with population displacement, with poor housing conditions, with a weak immune system, poor sanitary conditions, and lack of waste disposal. Up to 1 million new infections and up to 65,000 deaths from this disease occur annually. About 95% of cases of skin form disease occur in the countries of the American continent. Leishmaniasis occurs in Uzbekistan and Turkmenistan.&#x0D; The carriers of this disease are female mosquitoes. In the mosquito, parasites are in flagellate form. The natural reservoirs of Leishmania can be around 70 species of animals and humans. Infection occurs through mosquito saliva. After a bite, the parasite invades human mononuclear phagocytes. There may be infection of a person in contact with ulcers and other types of damage. Veterinarians have noted a dramatically increasing number of cases of disease in domestic animals.&#x0D; The following forms are clinically distinguished: cutaneous, mucocutaneous, diffuse cutaneous and visceral forms of leishmaniasis. After the disease develops, a stable immunity to this particular type of leishmaniasis develops.&#x0D; Cutaneous leishmaniasis is the most common form. Ulcers form in open areas of the body. Typical places of localization: face, ears, knees, elbows. Some nodules may have a warty surface or resemble xanthomas, keloids. After healing of these ulcers, scars remain for life.&#x0D; The nasopharynx, oral cavity, or nasal mucosa can be affected without destroying the nasal septum. For cutaneous leishmaniasis, the formation of an infectious granuloma is also characteristic.&#x0D; Treatment of cutaneous leishmaniasis can be local or systemic, depending on the damage and pathogen. Local treatment is suitable for minor and uncomplicated lesions. Local treatment options: heat therapy, cryotherapy. Systemic therapy is used in patients with multiple extensive rashes.&#x0D; Patient K. turned to the maxillofacial department. The patient came home to Ukraine for the purpose of diagnosis and treatment. The patient works in Poland at a construction site. According to the patient, several courses of treatment. The treatment had no result. The pharmacotherapy of our Polish colleagues is not known to us. Clinically: superficial skin lesions on the face. On the face are three ulcers of different sizes. Two ulcers on the cheeks and one on the nose. Palpation of the edges of the ulcers is very painful.&#x0D; Diagnostic search for the etiology of this process. Consultation of a rheumatologist ̶ the goal of eliminating Wegener's disease, rheumatological diseases. Hematologist consultation ̶ exclude hematology. Infectionist consultation ̶ rule out parasitic diseases. Laboratory examination: biochemical blood test, immunogram, rheumatic tests. CT scan of the abdomen.&#x0D; Consultation of a parasitologist. The diagnosis was not in doubt.The patient refused examination and treatment in the infectious diseases hospital. The further fate of this patient is not known to us. We want to draw the attention of doctors to the need for a thorough history taking. An epidemiological history is crucial in such cases.&#x0D; Treatment of leishmaniasis is long and toxic. No method of treatment gives 100% of the result. The choice of treatment method will depend on the type of pathogen and the geographical location of the infection.This disease can be brought by tourists, students from relevant countries of the world to Ukraine.

Review of Leishmaniasis in the Middle East and North Africa.

Cutaneous and visceral forms of leishmaniasis are the most important protozoan infection in the Middle East and North Africa (MENA). Review the current knowledge on leishmaniasis in the MENA. The data presented in this review are gathered primarily from WHO reports and from an extensive literature search on PubMed. There are four cycles of transmission of leishmaniasis: zoonotic cutaneous leishmaniasis (ZCL), induce by Leishmania (L.) major, transmitted by Phlebotomus (P.) papatasi, with rodent species of Psammomys obesus, Meriones libycus, Nesokia indica, and Rhombomys opimus are considered as host reservoirs. Zoonotic visceral leishmaniasis (ZVL) is inducing by L. infantum, transmitted by several Phlebotomus spp. of the sub-genus Larroussius and mainly P. perniciosus in more than one-half of the MENA countries and the dog species of Canis familiaris are considered as the main reservoirs. Anthroponotic cutaneous leishmaniasis (ACL), induce by L. tropica and transmitted by P. sergenti, without any non-human reservoir in most cases. Anthroponotic visceral leishmaniasis (AVL) induces by L. donovani spreads through P. alexandri, circulates exclusively in humans. There are many challenges facing the successful control of leishmaniasis. However, there is continuing research into the treatment of leishmaniasis and potentially vaccinations for the disease.

Visceral leishmaniasis (Kala-azar) in Qom Province, Iran: Report of two cases

Visceral leishmaniasis (VL) is a fatal parasitic zoonotic worldwide disease, which transmits to humans by the infected Phlebotomine sand fly bite. The common form of VL in Iran is the Mediterranean type with the causative agent of Leishmania infantum, whose main reservoirs are stray and domesticated dogs. The disease has several endemic foci in Iran, mostly seen among children under the age of 10, living in rural areas and nomadic tribes. The first cases of Kala-Azar in Qom province, central Iran, were reported in the year 2001, from the villages of Ghahan district. After conducting VL control strategies in the area, no new cases of the disease had been reported until recently. The cases described here are two 2-year-old girls, living in the urban parts of Qom province, one of whom did not have a history of traveling to known endemic areas of the disease. The patients were admitted to hospital in 2016-2017, complaining from recurrent fever with unrecognized reason, associated with decreased appetite and weight loss. Disease follow-up demonstrated anemia and splenomegaly, which led to diagnosis of VL, and both patients are now fully recovered. VL was presumed to be controlled in Qom province but the present cases indicate that possible VL existence remains in the region. Therefore, urgent studies and periodic monitoring are needed to identify potential reservoirs of VL in the area.

Regulation of immunity during visceral Leishmania infection.

Unicellular eukaryotes of the genus Leishmania are collectively responsible for a heterogeneous group of diseases known as leishmaniasis. The visceral form of leishmaniasis, caused by L. donovani or L. infantum, is a devastating condition, claiming 20,000 to 40,000 lives annually, with particular incidence in some of the poorest regions of the world. Immunity to Leishmania depends on the development of protective type I immune responses capable of activating infected phagocytes to kill intracellular amastigotes. However, despite the induction of protective responses, disease progresses due to a multitude of factors that impede an optimal response. These include the action of suppressive cytokines, exhaustion of specific T cells, loss of lymphoid tissue architecture and a defective humoral response. We will review how these responses are orchestrated during the course of infection, including both early and chronic stages, focusing on the spleen and the liver, which are the main target organs of visceral Leishmania in the host. A comprehensive understanding of the immune events that occur during visceral Leishmania infection is crucial for the implementation of immunotherapeutic approaches that complement the current anti-Leishmania chemotherapy and the development of effective vaccines to prevent disease.

Leishmaniasis and IFN-γ dependent chemokines.

Leishmaniasis is a disease caused by Leishmania and spread by the bite of certain types of sandflies. Leishmaniasis affects as many as 12 million people worldwide, with 2 million new cases each year. Leishmania infection, and Leishmaniasis, following occupational exposure has been also reported. Three forms of Leishmaniasis are known: a- Cutaneous Leishmaniasis (CL); b-Mucocutaneous Leishmaniasis; c- Visceral Leishmaniasis (VL). The visceral form of Leishmaniasis has an estimated incidence of 500,000 new cases. Cytokines and chemokines primarily orchestrate leukocyte recruitment to the areas of leishmania infection, and they are critical mediators of immune and inflammatory responses, in particular of the induction of Interferon (IFN)-γ and IFN-γ dependent chemokines. Skin biopsies from patients with CL showed higher expression of interferon-γ-induced protein (IP)-10, in recent lesions than in late lesions. Following L. braziliensis infection, enhanced expression of IP-10 and its receptor, chemokine C-X-C receptor (CXCR) 3, was predominantly detected in CD14(+) monocyte; this may contribute to disease severity by increasing cellular recruitment. It has been shown that IP-10 renders protection against VL, and it is associated with a strong host-protective T helper cell (Th)1 immune response. High clinical scores were positively correlated with IP-10 expression. Furthermore IP-10 is critical for rendering a protective cellular immunity during soluble leishmanial antigen (SLA) pulsed-CpG-ODN stimulated dendritic cells (SLA-CpG-DCs) vaccination that confers protection against L. donovani infection. Further studies are needed to evaluate IP-10 in Leishmaniosis, and to evaluate it as a potential therapeutic target.

Developing imidazole analogues as potential inhibitor for Leishmania donovani trypanothione reductase: virtual screening, molecular docking, dynamics and ADMET approach

Visceral leishmaniasis (VL) affects Indian subcontinent, African and South American continent, and it covers 70 countries worldwide. Visceral form of leishmaniasis is caused by Leishmania donovani in Indian subcontinent which is lethal if left untreated. Extensive resistance to antileishmanial drugs such as sodium stibogluconate, pentamidine and miltefosine and their decreased efficacy has been reported in the endemic region. Amphotericin B drug has shown good antileishmanial activity with significant toxicity, but its cost of treatment has limited the outreach of this treatment to affected people living in endemic zone. So, there is an urgent need to identify new antileishmanial drugs with excellent activity and minimal toxicity issues. Trypanothione reductase, a component of antioxidant system, is necessary for parasite growth and survival to raise infection. To develop potential inhibitor, we docked nine hundred and eighty-four 5-nitroimidazole analogues along with clomipramine which is a well-known inhibitor for TR. Total one hundred and forty-seven 5-nitroimidazole analogues with better docking score than clomipramine were chosen for ADMET and QikProp studies. Among these imidazole analogues, total twenty-four imidazole analogues and clomipramine were chosen on the basis of their ADMET, QikProp, and prime MM-GBSA study. Later on, two analogues with best MM-GBSA dG bind were undergone molecular dynamic simulation to ensure protein–ligand interactions. Using above approach, we confirm that ethyl 2-acetyl-5-[4-butyl-2-(3-hydroxypentyl)-5-nitro-1H-imidazol-1-yl]pent-2-enoate can be a drug candidate against L. donovani for the treatment of VL in the Indian subcontinent.

Concurrent visual diagnosis and susceptibility profiling of the first line drug against visceral leishmaniasis by plasmonic detection of PCR amplified genetic biomarker

Visceral form of leishmaniasis (also known as Kala-azar) is a fatal neglected tropical disease affecting 95 countries worldwide. Recently, substantial proportion of resistance related treatment failure cases have been reported against its first line drug, sodium-antimony gluconate (SAG). We report an easy, fast, sensitive and cheap visual diagnosis and SAG susceptibility profiling for this disease based on recently recognized genetic biomarker and gold nanoparticle based plasmonic detection phenomenon. This is a non-gel, non-culture based detection technique, which can be used as simultaneous high throughput detection and SAG-susceptibility profiling in Leishmania endemic resource stringent countries.

Genetic Structure and Evolution of the Leishmania Genus in Africa and Eurasia: What Does MLSA Tell Us

Leishmaniasis is a complex parasitic disease from a taxonomic, clinical and epidemiological point of view. The role of genetic exchanges has been questioned for over twenty years and their recent experimental demonstration along with the identification of interspecific hybrids in natura has revived this debate. After arguing that genetic exchanges were exceptional and did not contribute to Leishmania evolution, it is currently proposed that interspecific exchanges could be a major driving force for rapid adaptation to new reservoirs and vectors, expansion into new parasitic cycles and adaptation to new life conditions.To assess the existence of gene flows between species during evolution we used MLSA-based (MultiLocus Sequence Analysis) approach to analyze 222 Leishmania strains from Africa and Eurasia to accurately represent the genetic diversity of this genus. We observed a remarkable congruence of the phylogenetic signal and identified seven genetic clusters that include mainly independent lineages which are accumulating divergences without any sign of recent interspecific recombination. From a taxonomic point of view, the strong genetic structuration of the different species does not question the current classification, except for species that cause visceral forms of leishmaniasis (L. donovani, L. infantum and L. archibaldi). Although these taxa cause specific clinical forms of the disease and are maintained through different parasitic cycles, they are not clearly distinct and form a continuum, in line with the concept of species complex already suggested for this group thirty years ago. These results should have practical consequences concerning the molecular identification of parasites and the subsequent therapeutic management of the disease.

CXCL10 Is Critical for the Generation of Protective CD8 T Cell Response Induced by Antigen Pulsed CpG-ODN Activated Dendritic Cells

The visceral form of leishmaniasis is the most severe form of the disease and of particular concern due to the emerging problem of HIV/visceral leishmaniasis (VL) co-infection in the tropics. Till date miltefosine, amphotericin B and pentavalent antimony compounds remain the main treatment regimens for leishmaniasis. However, because of severe side effects, there is an urgent need for alternative improved therapies to combat this dreaded disease. In the present study, we have used the murine model of leishmaniasis to evaluate the potential role played by soluble leishmanial antigen (SLA) pulsed-CpG-ODN stimulated dendritic cells (SLA-CpG-DCs) in restricting the intracellular leishmanial growth. We found that mice vaccinated with a single dose of SLA-pulsed DC stimulated by CpG-ODN were protected against a subsequent leishmanial challenge and had a dramatic reduction in parasite burden along with the generation of parasite specific cytotoxic T lymphocytes. Moreover, we demonstrate that the induction of protective immunity conferred by SLA-CpG-DCs depends entirely on the CXC chemokine IFN-γ-inducible protein 10 (CXCL10; IP-10). CXCL10 is directly involved in the generation of a parasite specific CD8+ T cell-mediated immune response. We observed significant reduction of CD8+ T cells in mice depleted of CXCL10 suggesting a direct role of CXCL10 in the generation of CD8+ T cells in SLA-CpG-DCs vaccinated mice. CXCL10 also contributed towards the generation of perforin and granzyme B, two important cytolytic mediators of CD8+ T cells, following SLA-CpG-DCs vaccination. Together, these findings strongly demonstrate that CXCL10 is critical for rendering a protective cellular immunity during SLA-CpG-DC vaccination that confers protection against Leishmania donovani infection.

Overwhelming bone marrow Leishmaniasis.

doi:10.5152/tjh.2011.97 Visceral Leishmaniasis (kala-azar) is caused by the intracellular parasitic organism Leishmania donovani and is transmitted by the bite of an infected sandfly Phlebotomus argentipes. Clinical manifestations of advanced disease usually include constitutional symptoms, such as prolonged fever, anorexia, weight loss, and marked enlargement of the spleen and liver, with varying degrees of pancytopenia [1]. Pakistan is a tropical country situated in the north-west region of South Asia; all of its neighboring countries are endemic for Leishmaniasis, including China, Afghanistan, Iran, and India [2-4]. Leishmaniasis was first reported from Pakistan in 1960 [5]. Initially, it was limited to the northern mountainous region, but currently it is widespread throughout the country with several endemic foci of varying climatic and geographic conditions [6]. Herein we present a 22-year-old female from Hyderabad, Sindh Province, Pakistan that was referred to our laboratory for bone marrow exanimation due to pyrexia of unknown origin. She had fever with chills and rigors that began 2 months earlier, which did not respond to treatment with antimalarials and multiple antibiotics. At presentation she had massive splenomegaly and marked pallor. Complete blood count showed the following: pancytopenia with hypochromic microcytic anemia; hemoglobin: 7.1 g/dL; hematocrit: 21.8%; MCV: 72.5 fL; MCH: 23.8 pg; MCHC: 29.7%; total white cell count: 1.5×109/L; absolute neutrophil count: 0.4×109/L; platelet count: 48×109/L. Peripheral blood smear did not show a leukoerythroblastic reaction or the presence of abnormal cells. Bone marrow examination showed hypercellular marrow and massive infestation with the amastigote form of Leishmania donovani in the bone marrow macrophages and in some granulocytes. Written informed consent was obtained from the patient. The visceral form of Leishmaniasis can be fatal if not treated in a timely fashion and stibogluconate (sodium antimony gluconate) is the drug of choice. Recently, other agents have become available for the treatment of new and resistant cases of visceral Leishmaniasis, including pentamidine isethionate, amphotericin B, miltefosine, and paramomycin [1]. The presented patient received anti-leishmanial treatment with sodium antimony gluconate 20 mg/ kg/d for 14 d. She began to exhibit improvement within 3 d and was clinically cured after the full course of therapy. She was last followed-up 1 month after completion of the treatment and was asymptomatic and healthy.

Comparison of culture media in the isolation and diagnosis of cutaneous leishmaniasis.

Leishmaniasis is a common and significant parasitic disease among the Middle East and Mediterranean countries. Cutaneous and visceral forms of leishmaniasis are seen in Turkey. Direct microscopic examination of Giemsa-stained smears and inoculation into a culture medium are gold standards in diagnosis. In the present study, three diffferent culture media (pepton-yeast extract, brain heart and nutrient broth) were used in comparison with Novy-MacNeal-Nicolle and RPMI 1640 as control media to assess the effectiveness of new culture media both in diagnosis in patients and after isolation from the mice. In addition, smears prepared from the infectious material were stained with Giemsa and examined under a light microscope. Samples obtained from the lesions of cutaneous leishmaniasis produced on the sole of mice were inoculated into all culture media; growth were detected in pepton-yeast extract and nutrient broth media. Smears prepared from the lesions of these mice were stained with Giemsa and amastigotes were detected on them. Growth was also seen in control media. Five of 12 cultivations of the human samples suspected of having cutaneous leishmaniasis revealed growth in pepton-yeast extract medium and nutrient broth medium. Microscopic examination of these five patients showed the presence of amastigotes and growth in control media, while none was observed in the remaining seven cases. These results indicated that pepton-yeast extract and nutrient broth media could be preferred in both isolation of the parasites and diagnosis of cutaneous leishmaniasis. Key words: Leishmaniasis, culture medium, diagnosis, isolation.

Disseminated leishmaniasis caused by Leishmania tropica in HIV-positive patients in the Islamic Republic of Iran

Both cutaneous and visceral forms of leishmaniasis are prevalent in some parts of the Islamic Republic of Iran [1,2]. The Mediterranean type of visceral leishmaniasis (VL) is also reported sporadically throughout the country [3] with canines being the main animal reservoir hosts [4]. The cutaneous form of leishmaniasis is seen in 2 forms: anthroponotic and zoonotic. Anthroponotic cutaneous leishmaniasis (ACL) is caused by Leishmania tropica and is prevalent in many largeand medium-sized cities in the country [1,5]. Zoonotic cutaneous leishmaniasis (ZCL) is caused by L. major and is endemic in many foci in the north, east and south of the country [6]. Some studies have implicated L. tropica as another agent of VL in humans and dogs in reports from the north-west and south of the country [4,7,8]. Few data are available, however, about host immunological response and parasite destruction when leishmaniasis is associated with immunosuppressants. At present, the majority of cases of HIV– leishmaniasis co-infection reported in the Mediterranean basin were caused by L. infantum [9]. Disseminated leishmaniasis caused by L. tropica in patients with HIV infection is uncommon. Here, we report on 2 patients with HIV–leishmaniasis co-infection caused by L. tropica with generalized and multiple skin lesions and visceral involvement. This is the first such report from the Islamic Republic of Iran.

Vector Transmission of Leishmania Abrogates Vaccine-Induced Protective Immunity

Numerous experimental vaccines have been developed to protect against the cutaneous and visceral forms of leishmaniasis caused by infection with the obligate intracellular protozoan Leishmania, but a human vaccine still does not exist. Remarkably, the efficacy of anti-Leishmania vaccines has never been fully evaluated under experimental conditions following natural vector transmission by infected sand fly bite. The only immunization strategy known to protect humans against natural exposure is “leishmanization,” in which viable L. major parasites are intentionally inoculated into a selected site in the skin. We employed mice with healed L. major infections to mimic leishmanization, and found tissue-seeking, cytokine-producing CD4+ T cells specific for Leishmania at the site of challenge by infected sand fly bite within 24 hours, and these mice were highly resistant to sand fly transmitted infection. In contrast, mice vaccinated with a killed vaccine comprised of autoclaved L. major antigen (ALM)+CpG oligodeoxynucleotides that protected against needle inoculation of parasites, showed delayed expression of protective immunity and failed to protect against infected sand fly challenge. Two-photon intra-vital microscopy and flow cytometric analysis revealed that sand fly, but not needle challenge, resulted in the maintenance of a localized neutrophilic response at the inoculation site, and removal of neutrophils following vector transmission led to increased parasite-specific immune responses and promoted the efficacy of the killed vaccine. These observations identify the critical immunological factors influencing vaccine efficacy following natural transmission of Leishmania.