Different Forms Of Leishmaniasis Research Articles

Use of FTA Cards for Direct Sampling of Patients' Lesions in the Ecological Study of Cutaneous Leishmaniasis

The FTA card (Whatman) was assessed for its utility as a molecular epidemiological tool in collecting samples from patients with leishmaniasis in Peru because the card has a variety of merits; it is less invasive for patients and easy to handle for both physicians and other medical personnel for sample collection or diagnosis, in addition to its simplicity and easy countrywide and/or intercountry transportation for analysis. Samples were collected from 132 patients suspected of having leishmaniasis, and Leishmania species were successfully identified in samples from 81 patients in 15 departments of Peru by cytochrome b and mannose phosphate isomerase gene analyses. Of these, 61.7% were identified as Leishmania (Viannia) peruviana, 22.2% as L. (V.) braziliensis, 12.3% as L. (V.) guyanensis, 2.5% as L. (V.) shawi, and 1.2% as L. (V.) lainsoni. The three predominant species, L. (V.) peruviana, L. (V.) braziliensis, and L. (V.) guyanensis, were mainly found in the Andean highlands, in the tropical rainforest, and in northern and central rainforest regions, respectively. This is the first time L. (V.) shawi has been identified outside Brazil. The present study showed that the FTA card will be a useful tool for the ecological study of different forms of leishmaniasis. Furthermore, collecting samples directly from patients' lesions by using the FTA card eliminates (i) the possibility of contamination of Leishmania isolates during short- and/or long-term passages of culture in vitro in each laboratory and (ii) pain and suffering of patients from taking samples by skin biopsy.

Contributions of Ultrastructural Studies to the Cell Biology of Trypanosmatids: Targets for Anti-Parasitic Drugs

Protozoan parasites cause disease in humans worldwide, and many fall into the genera Trypanosoma and Leishmania; these parasites are responsible for African trypanosomiasis, Chagas disease and the different forms of Leishmaniasis. Strategies for the development of new drugs against these protozoans have been based on their cell biol- ogy and biochemistry complemented by the use of electron microscopy. Trypanosoma and Leishmania have special orga- nelles that are involved in metabolic pathways, which are very distinct from those in mammalian cells; these organelles are potential drug targets. Scanning and transmission electron microscopy can identify not only the target organelles but also alterations to the cell surface and ultrastructural changes that characterize distinct forms of programmed cell death.

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

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

Recent developments leading toward a paradigm switch in the diagnostic and therapeutic approach to human leishmaniasis

To identify recent papers showing how human and parasite genetics influence leishmaniasis, and how understanding of the immunopathology may be utilized in immunotherapy for these diseases. Progress has been made in recent years showing the complexity within populations of Leishmania spp. and indicating that different strains lead to diverse clinical pictures and responses to treatment. Thus detection of parasite genetic tags for the precise identification of infecting strains, and for predictive diagnosis of clinical and therapeutic fates seems now possible. Host genetic loci involved in disease outcome have been detected, which may also be explored for better case management. These developments in diagnosis will demand expanding the therapeutic arsenal to take their expected effect. This is starting to be fulfilled by immunotherapies successfully employed to treat cases refractory to standard first line drugs, as the result of a more profound comprehension of the immunopathology of the leishmaniases. The knowledge mounting has already helped explain why different patients present different forms of leishmaniasis and respond differently to treatment, and may be on the verge of catalyzing a major change in the already over a century old paradigm of diagnosing and managing these patients.

Antileishmanial and Antibacterial Activity of a New Pyrazole Derivative Designated 4‐[2‐(1‐(Ethylamino)‐2‐methyl‐ propyl)phenyl]‐3‐(4‐methyphenyl)‐1‐phenylpyrazole

Here, we report for the first time the synthesis and the antileishmanial activity of a new pyrazole derivative, namely 4-[2-(1-(ethylamino)-2-methylpropyl)phenyl]-3-(4-methyphenyl)-1-phenylpyrazole). Micromolar concentrations of this compound were found to inhibit the in vitro multiplication of Leishmania tropica, Leishmania major, and Leishmania infantum, three species causing different forms of leishmaniasis. Furthermore, the 50% inhibitory concentration (IC50) values for the compound are only slightly higher than those of amphotericin B, one of the most active antileishmanial agents used as a satisfactory substitute in cases not responding to pentostam. The IC50 values after 48 h for L. tropica, L. major, and L. infantum promastigote growth were 0.48 microg/mL, 0.63 microg/mL and 0.40 microg/mL, respectively for the compound, while they were 0.23 microg/mL, 0.29 microg/mL and 0.24 microg/mL, respectively for amphotericin B. We also tested this compound for its antibacterial activity against several bacteria. The strongest antibacterial activity was observed against Entrococcus feacalis and Staphylococcus aureus with a minimal inhibitory concentration (MIC) of 60 microg/mL.

Substrate Specificity, Localization, and Essential Role of the Glutathione Peroxidase-type Tryparedoxin Peroxidases in Trypanosoma brucei

Trypanosoma brucei, the causative agent of African sleeping sickness, encodes three nearly identical cysteine homologues of the classical selenocysteine-containing glutathione peroxidases. Although one of the sequences, peroxidase III, carries both putative mitochondrial and glycosomal targeting signals, the proteins are detectable only in the cytosol and mitochondrion of mammalian bloodstream and insect procyclic T. brucei. The enzyme is a trypanothione/tryparedoxin peroxidase as are the 2 Cys-peroxiredoxins of the parasite. Hydrogen peroxide, thymine hydroperoxide, and linoleic acid hydroperoxide are reduced with second order rate constants of 8.7 x 10(4), 7.6 x 10(4), and 4 x 10(4) m(-1) s(-1), respectively, and represent probable physiological substrates. Phosphatidylcholine hydroperoxide is a very weak substrate and, in the absence of Triton X-100, even an inhibitor of the enzyme. The substrate preference clearly contrasts with that of the closely related T. cruzi enzyme, which reduces phosphatidylcholine hydroperoxides but not H(2)O(2). RNA interference causes severe growth defects in bloodstream and procyclic cells in accordance with the peroxidases being essential in both developmental stages. Thus, the cellular functions of the glutathione peroxidase-type enzymes cannot be taken over by the 2 Cys-peroxiredoxins that also occur in the cytosol and mitochondrion of the parasite.

Trypanosoma cruzi ubiquitin as an antigen in the differential diagnosis of Chagas disease and leishmaniasis

In the present report we describe Trypanosoma cruzi ubiquitin as an antigen to be utilized in the differential diagnosis of Chagas disease and leishmaniasis. Initially, recombinant T. cruzi ubiquitin was evaluated against a panel of sera by phage dot immunoassay, showing a good performance against chagasic sera. However, the presence of a carboxy-terminal tail region encoding a ribosomal protein homologous to a related protein present in the genome of Leishmania sp. gave significant cross-reactivity with leishmanial sera. Therefore, ubiquitin was purified by a simple biochemical protocol and its immunoreactivity was studied by enzyme-linked immunosorbent assay. Analysis of 104 sera indicates that the response to ubiquitin is very sensitive towards chronic chagasic sera (98%) and, more important, highly species-specific, presenting better performance compared to the use of the recombinant protein or the total epimastigote extracts when tested against a panel of leishmanial sera, where out of a total of 70 sera tested, only five sera from the mucocutaneous form of the disease reacted with T. cruzi ubiquitin. On the other hand, Leishmania ubiquitin was not recognized by chagasic sera, but was recognized by sera from different forms of leishmaniasis. These results make ubiquitin an excellent candidate to be used in the differential diagnosis of these two parasitic diseases. The molecular basis for this highly species-specific response is discussed.

The parasite-specific trypanothione metabolism of trypanosoma and leishmania.

The bis(glutathionyl)spermidine trypanothione exclusively occurs in parasitic protozoa of the order Kinetoplastida, such as trypanosomes and leishmania, some of which are the causative agents of several tropical diseases. The dithiol is kept reduced by the flavoenzyme trypanothione reductase and the trypanothione system replaces in these parasites the nearly ubiquitous glutathione/glutathione reductase couple. Trypanothione is a reductant of thioredoxin and tryparedoxin, small dithiol proteins, which in turn deliver reducing equivalents for the synthesis of deoxyribonucleotides as well as for the detoxification of hydroperoxides by different peroxidases. Depending on the individual organism and the developmental state, the parasites also contain significant amounts of glutathione, mono-glutathionylspermidine and ovothiol, whereby all four low molecular mass thiols are directly (trypanothione and mono-glutathionylspermidine) or indirectly (glutathione and ovothiol) maintained in the reduced state by trypanothione reductase. Thus the trypanothione system is central for any thiol regeneration and trypanothione reductase has been shown to be an essential enzyme in these parasites. The absence of this pathway from the mammalian host and the sensitivity of trypanosomatids toward oxidative stress render the enzymes of the trypanothione metabolism attractive target molecules for the rational development of new drugs against African sleeping sickness, Chagas' disease and the different forms of leishmaniasis.

Present status of antileishmanial vaccines.

The term leishmaniasis refers collectively to various clinical syndromes that are caused by obligate intracellular protozoa of the genus Leishmania. Approximately 350 million people in 8 countries are estimated to be threatened by the disease. The World Health Organization estimated that there are 12 million cases of all forms of leishmaniasis worldwide, with over 500,000 new cases of visceral disease occurring each year. Most of the drugs commonly used to treat different forms of leishmaniasis are toxic and have unacceptable side effects. Moreover, cases of drug resistant leishmaniasis are on the rise. Due to non-existence of effective vaccine to date, improved immunoprophylactic approaches still remain desirable to combat leishmaniasis. Antileishmanial vaccines developed around the globe are discussed.

In Vivo Evaluation of Immune Responses in Leishmaniasis: The Use of Cross-Species Leishmanin Preparations for Skin Testing

Skin test reactivity to two commercial preparations of Leishmania major leishmanin was evaluated in leishmaniasis patents from Ethiopia (L. aethiopica) and Nicaragua (probably L. braziliensis complex). The purpose of using different preparations of L. major was to evaluate whether L. major skin test antigens could generally be used to detect leishmaniasis due to L. aethiopica and L. braziliensis. One preparation was superior in identifying the majority (83-90%) of confirmed cases of local cutaneous leishmaniasis (LCL) from Ethiopia. In vitro responses to promastigotes of Leishmania spp. correlated well with leishmanin positivity to this preparation. The skin test antigen, which performed less well (showing a positive result in only 50% of the LCL patients), showed promise when used to test active and cured cases of leishmaniasis in Nicaragua (positive result in 92% of the active and cured patents). Cross-reacting Leishmania spp. may be considered for use in the preparation of standardized leishmanin antigen; however, differences in the commercial preparation or the diluent in which the parasites are prepared may affect antigenicity and thus its efficacy in detecting different forms of leishmaniasis in different geographic areas.

Characterization of a membrane antigen of Leishmania amazonensis that stimulates human immune responses.

To investigate human immune responses to defined leishmania Ag we have begun to characterize biochemically and immunologically, an abundant 42-kDa surface Ag of Leishmania amazonensis, a causative agent of human leishmaniasis. We have shown that this Ag, La gp42, is expressed on the surface of L. amazonensis promastigotes, being anchored to the membrane by a glycosyl-phosphatidylinositol moiety. As demonstrated by lectin blotting studies, La gp42 is glycosylated, binding both Con A and wheat germ agglutinin. Immunologically, La gp42 is strongly recognized by sera from patients with different forms of leishmaniasis as well as by patients with Chagas' disease. In addition, we show that purified La gp42 stimulates the proliferation of human T lymphocytes obtained from several leishmaniasis patients. Finally, the N-terminal sequence of La gp42 was obtained and a serologically cross-reactive 42-kDa protein with a homologous sequence was identified in Leishmania major.

A common major surface antigen on amastigotes and promastigotes of Leishmania species.

Enzymatic surface iodination and biosynthetic labeling with [35S]methionine, combined with immunoprecipitation by sera from patients with different forms of Leishmaniasis revealed a 65,000 Mr glycoprotein as the immunodominant moiety in promastigotes and amastigotes of the nine Leishmania species and isolates examined. Sera from patients with one form of Leishmaniasis recognized this component strongly, not only in the homologous, but also in the heterologous species. In addition to the crossreactivity displayed by immune sera, the 65,000 Mr glycoprotein (gp) common to all Leishmania species presented a characteristic shift to Mr 50,000 when samples were run in sodium dodecyl sulfate-polyacrylamide gel electrophoresis under nonreducing conditions. These results are in agreement with our previous studies (7), where a simple and similar profile was obtained for several geographic isolates of L. donovani, with a major surface glycoprotein of 65,000 Mr displaying the same characteristics described here. The structural similarity of the major 65,000 Mr gp of the six Leishmania species was demonstrated by Cleveland mapping. It is suggested that immunological specificities may be contributed by minor differences in glycosylation of this molecule. In keeping with recent data (13-15), where strong cross protection among different Leishmania species has been obtained by prophylactic immunization with irradiated whole promastigotes, this glycoprotein may be a good candidate for an antigen to be used for immunoprophylaxis of all forms of Leishmaniasis.