Human infection with Trypanosoma cruzi induces parasite antigen-specific cytotoxic T lymphocyte responses.

Experimental models of Chagas' disease, an infection caused by the intracellular protozoan Trypanosoma cruzi, have demonstrated the crucial immunoprotective role played by CD8(+) T lymphocytes. These cells dominate inflammatory foci in parasitized tissues and their elimination from mice leads to uncontrolled parasite replication and subsequent death of the infected host. A trypomastigote surface antigen, TSA-1, and two amastigote surface molecules, ASP-1 and ASP-2, were recently identified as targets of CD8(+) cytotoxic T lymphocytes (CTL) in T. cruzi-infected mice. Until now, however, there was no evidence for the development of parasite-specific CTL in T. cruzi-infected humans. In this study, human CTL specific for TSA-1-, ASP-1-, and ASP-2-derived peptides were detected in the peripheral blood mononuclear cells from 21 of 24 HLA-A2(+) T. cruzi-infected patients. CTL recognition was antigen specific, A2-restricted, and CD8(+) T cell-dependent. Demonstration of human CTL against T. cruzi and against target molecules identified using the murine model provides important information for the optimal design and evaluation of vaccines to prevent or ameliorate Chagas' disease.

Lung function and symptoms among indigenous Mayan women exposed to high levels of indoor air pollution.

SETTING: Indoor air pollution from burning of biomass fuel in open fires is a known risk factor for chronic obstructive pulmonary disease (COPD) in developing countries. OBJECTIVE: To estimate the prevalence of respiratory symptoms and lung function among women in rural Guatemala and to describe the methods and practical issues associated with the assessment of respiratory health. DESIGN: Information about respiratory symptoms, lung function and individual measurement of exposure was collected cross-sectionally among 350 Mayan-Indian women aged 15-50 years who used traditional open fires. RESULTS: These women, exposed to indoor air pollution since birth, had a relatively high prevalence of cough (22.6%), phlegm (15.1%), wheeze (25.1%) and tightness in the chest (31.4%). Respiratory symptoms were positively associated with exposure levels. Lung function was higher than the most feasible reference population (average above predicted forced expiratory volume in 1 s [FEV(1)] +4.5% and forced vital capacity [FVC] +4.2%). Only one woman had a FEV(1)/FVC ratio lower than 70%. CONCLUSIONS: According to the Global Initiative for Chronic Obstructive Lung Disease (GOLD) guidelines, almost one third of these young non-smoking women were at risk (stage 0) of developing COPD. The methodological issues encountered during the study highlight the importance of standardising approaches to local adaptation of established questionnaires to study respiratory health in rural areas of developing countries.

Inorganic particles in the skin of inhabitants of volcanic areas of Central America: their possible immunomodulatory influence in leishmaniasis and leprosy.

We have evaluated biopsies from patients with atypical nodular and typical ulcerated lesions of cutaneous leishmaniasis, from leishmanin reactions and skin from normal individuals from Nicaragua, Honduras and Guatemala for the presence of inorganic particles using confocal microscopy with a polarised light source and conventional histopathological techniques. Analysis by semiquantitative confocal microscopy permitted the demonstration of significantly larger numbers of particles in atypical lesions. Silica and aluminium, important components of these particles, were less abundant in particles from normal skin. The histology of these atypical lesions, characterised by 'naked' sarcoidal granulomas with epithelioid differentiation but very few lymphocytes, was very similar to the histological reaction observed after 14 days in persisting inflammation at leishmanin skin test sites. The presence of these unusual lesions in areas of Central American countries characterised by the presence of large amounts of volcanic ash, as well the unexpectedly low prevalence of leprosy in Central America, suggest that environmental factors may contribute significantly to the frequency and clinical manifestations of these infections. Among possible environmental features, the presence of inorganic particles with immunomodulatory properties in the skin may be a significant factor.

Miltefosine for new world cutaneous leishmaniasis.

The oral agent miltefosine has demonstrated a >95% cure rate in Indian visceral leishmaniasis. We performed a large, placebo-controlled study of miltefosine therapy (2.5 mg/kg per day orally for 28 days) against cutaneous leishmaniasis in Colombia and Guatemala. In regions in Colombia where Leishmania vianna panamensis is common, the per-protocol cure rates for miltefosine and placebo were 91% (40 of 44 patients) and 38% (9 of 24). These values are similar to historic values for the antimony standard of care and placebo. In regions in Guatemala where L. v. braziliensis and L. mexicana mexicana are common, the per-protocol cure rates were 53% (20 of 38) for miltefosine and 21% (4 of 19) for placebo. The miltefosine rate was lower than historic antimony cure rates of >90%. Miltefosine was well tolerated. Miltefosine is a useful oral agent against cutaneous leishmaniasis due to L. v. panamensis in Colombia but not against leishmaniasis due to L. v. braziliensis in Guatemala.

Comparison of active and passive case detection of cutaneous leishmaniasis in Guatemala.

To estimate the degree to which passive case detection underestimates the true incidence of cutaneous leishmaniasis in Guatemala, we compared data from the passive surveillance system of the Guatemalan Ministry of Health with a cross-sectional population-based survey of cutaneous leishmaniasis in Guatemala. Of the 2,938 persons interviewed, 143 (5%) reported having had cutaneous leishmaniasis at some time in the past, 37 (1.3%) reported the onset of infection in the 12 months before the survey, 31 (1.1%) had active infections, and 16 (0.5%) had parasitologically confirmed infections. Calculated on the basis of these reports and the estimated population of the endemic area, the total number of new cases in the leishmaniasis-endemic area in the 12 months before the survey was approximately 2,574; during the same 12 month period, Ministry of Health data based on passive surveillance listed 64 cases of cutaneous leishmaniasis. In Guatemala, incidence estimates based on passive surveillance may underestimate the occurrence of cutaneous leishmaniasis by as much as a factor of 40.

Randomized, controlled, double-blind trial of topical treatment of cutaneous leishmaniasis with paromomycin plus methylbenzethonium chloride ointment in Guatemala.

A double-blind, randomized trial was undertaken in Guatemala to determine the therapeutic efficacy of an ointment for the treatment of cutaneous leishmaniasis that contained 15% paromomycin and 12% methylbenzethonium chloride and that was applied twice a day for 20 days. The treatment group included 35 patients, and the placebo group included 33 patients. The initial clinical response rate (13 weeks after completing the treatment) was 91.4% in the treatment group and 39.4% in the placebo group. The final clinical response rate at the 12-month follow-up examination was 85.7% (31 of 35) in the treatment group and 39.4% (13 of 33) in the placebo group (P < or = 0.001). In general, the treatment was well tolerated and was never interrupted because of adverse effects. The number of adverse effects reported in the placebo group was lower than in the treatment group (16 events versus 30 events). All adverse effects reported by patients disappeared within 1 week of completing the treatment. Our findings show that the combination of paromomycin with methylbenzethonium chloride for 20 days is a good alternative for antimonial treatments of cutaneous leishmaniasis in Guatemala.

Cutaneous leishmaniasis in Guatemala: comparison of diagnostic methods.

A comparison was made of methods used to diagnose suspected cutaneous leishmaniasis in Guatemala. The most sensitive method was a combination of thin smears made from superficial scrapings of the ulcers and inoculation of culture medium with either aspirates or scrapings. The diagnosis was confirmed in 252 (70%) of 362 patients. Ability to cultivate Leishmania was correlated with the concentration of amastigotes seen on thin smears. Leishmania were cultured in 42 (27%) of 153 patients with no amastigotes found in 400 oil-immersion fields and in 174 (83%) of 209 patients with at least 1 amastigote. No difference in diagnostic outcome was found when we compared smears or cultures taken from the center or the border of the ulcer or from an incision made tangential from the ulcer. We found no difference when we compared smears obtained with scalpels, capillary tubes, or dental broaches. The use of scrub brushes soaked in iodine neither decreased the rate of culturing parasites nor decreased contamination rates.

Placebo-controlled clinical trial of meglumine antimonate (glucantime) vs. localized controlled heat in the treatment of cutaneous leishmaniasis in Guatemala.

Sixty-six Guatemalans with parasitologically proven cutaneous leishmaniasis were randomly and equally divided into 3 treatment groups: those receiving meglumine antimonate (Glucantime), 850 mg antimony/day im for 15 days; those receiving localized controlled heat from a radio-frequency generator, 50 degrees C for 30 sec, 3 treatments at 7 day intervals; and those receiving treatment with a placebo. Of 53 isolates identified, 40 were Leishmania braziliensis braziliensis and 13 were L. mexicana mexicana. Thirteen weeks after beginning treatment, the number of patients from each group with completely healed and parasitologically negative lesions were as follows: meglumine antimonate, 16 (73%); localized heat, 16 (73%); and placebo, 6 (27%). The cure rate for those with infections due to L. b. braziliensis in each group was as follows: meglumine antimonate, 11 out of 14 (79%); controlled heat, 9 out of 14 (64%); and placebo, 0 out of 11.

Evaluation of a standardized leishmanin skin test in Guatemala.

Before recommending the skin-test use at national level in Guatemala of an antigen prepared from Leishmania major (a Leishmania species not found in the New World), we conducted a study in 100 Guatemalans to determine its sensitivity and specificity. The antigen consisted of 0.1 mL of a solution that contained 5 x 10(6) promastigotes of L. major (MRHO/IR/75/VAX). Positive leishmanin skin test (LST) reactions at 48 h were observed in 16 (80%) of 20 patients with proven active cutaneous leishmaniasis (CL), 18 (90%) of 20 with previously treated proven CL, and in 18 (90%) of 20 with a history and compatible scan of previously suspected but unconfirmed CL. None of 20 healthy controls or 20 patients with skin lesions due to causes other than CL had positive reactions to the LST, giving a sensitivity of 85% and specificity of 100%. There were no statistically significant differences in ethnic group, age, duration of the lesion, lesion size or Leishmania species between the 34 persons with true positive reactions. Even though it will be necessary to test this antigen on a larger number of patients, these preliminary results show that this antigen is specific and reasonably sensitive in identifying current or past CL and that it is a reasonable choice for epidemiological studies on CL in Guatemala.

The characterization of Leishmania parasites and their vectors from Central America using molecular techniques.

The disfiguring cutaneous and mucocutaneous and often fatal visceral forms of leishmaniasis are an increasing public health problem in Central America. There are some evidences of a highest prevalence of L. braziliensis and L. mexicana in Guatemala; L. braziliensis, L. panamensis and L. infantum (L. chagasi) in Honduras and Nicaragua, and L. panamensis in Costa Rica and Panamá. Our research project, just starting, attempts to use molecular approaches for a more rapid and accurate diagnosis and to identify the parasites. The techniques include non-radioactive DNA probes, PCR, schizodeme, isoenzyme analysis and monoclonal antibodies; and specific recombinant peptides will be used to improve immunodiagnosis of the visceral form. Intra-specific heterogeneity in Leishmania isolates will be examined by pulsed field electrophoresis and random amplified polymorphic DNA. Using selected strains, the techniques will be compared on a regional basis leading to a better knowledge of parasite distribution and the related clinical entities. Furthermore, DNA probes for the identification of parasites in vectors and for sibling species of vectors, are also to be applied; chemical identification of sandfly male pheromones will be attempted possibly leading to potential novel control measures.

Chemotherapy of cutaneous leishmaniasis: a review.

A wide variety of treatment modalities have been employed for cutaneous leishmaniasis (CL); however, none has been demonstrated to be good enough to be used as the first-line drug to treat patients in all the CL epidemiological scenarios. Although pentavalent antimonials remain a drug widely used in the treatment of all forms of leishmaniasis, they are far from being satisfactory. A great variety of topical treatments have been used to treat CL. However, the majority of these modalities have been tested in non-controlled studies, with only few subjects and the interpretation of results is usually difficult due to the lack of a standard and well-accepted cure definition. Several oral drugs, such as ketoconazole and itraconazole, have also been tested for CL, but results obtained were not completely satisfactory. As of today, miltefosine, originally developed as an oral antineoplastic agent, seems, from the preliminary results obtained in different phase II clinical studies, to be the most promising oral drug developed during the last decade.

The natural history of cutaneous leishmaniasis in Guatemala.

The natural history of American cutaneous leishmaniasis was studied in Guatemala by analyzing the characteristics of 355 untreated leishmanial lesions, observing the evolution of 57 lesions on persons who received a placebo in treatment trials, and analyzing data from a population-based survey concerning the duration of 82 untreated lesions. Of 25 lesions caused by Leishmania mexicana that were followed prospectively, 22 (88%) completely reepithelialized by a median lesion age of 14 weeks, and 17 (68%) were classified as cured (no residual wound inflammation or reactivation during at least 6 months of follow-up). In contrast, 7 (22%) of 32 lesions caused by Leishmania braziliensis reepithelialized by a median lesion age of 13 weeks, and only 2 (6%) cured. These data demonstrate that the species of Leishmania is the primary determinant of the clinical course and outcome of untreated lesions and underscore the need for field-applicable diagnostic techniques that provide rapid species identification.

Cutaneous leishmaniasis in Guatemala: people's knowledge, concepts and practices.

Ten rural communities in the northern area of Guatemala where cutaneous leishmaniasis (CL) is endemic were investigated to determine the residents' knowledge of the disease, their related concepts and practices, and their treatment preferences, and to identify the communication channels they use to acquire information. Of 425 heads of household interviewed, 96.7% could accurately describe a typical CL lesion. CL was found to be the fourth most frequently mentioned disease (in studies based on a free list format) and to be considered the sixth most serious (in studies based on paired comparisons). A series of three-way comparisons, used to analyse the subjects' concepts about the similarities of various discases, indicated that CL was considered to be most closely related to skin problems and to be different from any other group of diseases. All interviewees believed that it was necessary to receive treatment for CL, because without treatment the disease would progress, reach the bone, and take years to heal. More than half (55%) of the respondents knew about meglumine antimonate (Glucantime), the most commonly prescribed drug for treating CL in Guatemala. Only a few communication channels that were used by respondents to receive information were identified; the use of radio broadcasts and direct communication via the community leaders appeared to be the most effective.

Efficacy of a short course (10 days) of high-dose meglumine antimonate with or without interferon-gamma in treating cutaneous leishmaniasis in Guatemala.

Sixty-six Guatemalan men with parasitologically confirmed cutaneous leishmaniasis, due most commonly to Leishmania braziliensis, were randomly assigned to receive one of three treatment regimens: meglumine antimonate (meglumine) for 20 days; meglumine for 10 days; and meglumine for 10 days plus alternate-day injections of interferon-gamma. In each group, meglumine was given intravenously as 20 mg of antimony/(kg of body weight.d). All treatment regimens were associated with similar response rates: the lesions of 19 (90%) of 21 patients who received meglumine for 20 days, 18 (90%) of 20 patients who received meglumine for 10 days, and all 22 patients who received meglumine plus interferon-gamma were completely reepithelialized by 13 weeks. In addition, for patients receiving all treatment regimens, test-of-cure cultures for Leishmania were negative and reactivation of lesions did not occur during 12 months of follow-up. The high efficacy of our 10-day course of meglumine indicates that the currently recommended duration of 20 days may be unnecessary for infections caused by L. braziliensis and suggests that a 10-day course of high-dose antimony should be tested as therapy for cutaneous leishmaniasis in other geographic areas.

The Leishmania hertigi (Kinetoplastida; Trypanosomatidae) complex and the lizard Leishmania: their classification and evidence for a neotropical origin of the Leishmania-Endotrypanum clade.

The relationships of the Leishmania hertigi complex and the lizard Leishmania species to the main groups of mammalian Leishmania and Endotrypanum parasites were examined. Restriction fragment length polymorphisms and sequences of small subunit ribosomal RNA genes and hybridization studies of kinetoplast DNA indicated that the L. hertigi complex was more closely related to the genus Endotrypanum than to the genus Leishmania. The lizard Leishmania species were found to be at the crown of the Leishmania tree. The data provides strong evidence for a Neotropical origin of the Endotrypanum/Leishmania clade since the parasites closest to the root of the tree are all found exclusively in the Neotropics. The evolution of the Leishmania/Endotrypanum clade in relation to the evolution of the known hosts of these parasites is discussed.