An Investigation on the Ecology of Triatoma vitticeps (Stal, 1859) and its Possible Role in the Transmission of Trypanosoma cruzi, in the Locality of Triunfo, Santa Maria Madalena Municipal District, State of Rio de Janeiro, Brazil

From January 1989 to April 1995, 465 specimens of Triatoma vitticeps were collected in the locality of Triunfo, 2nd District of Santa Maria Madalena municipal district, State of Rio de Janeiro. The bugs were found indoors by local residents with predominance of adults. The flight activity was high in hot months when the incidence in the domicile also increased. Two hundred and two bugs (111 alive and 91 dead) were examined for Trypanosoma cruzi infection. This was detected in 31 of the dead bugs (34%) and 88 (79%) of the live bugs examined. With a view to investigate the possible vertebrate hosts of the T. cruzi isolates, the blood of 122 mammals was examined through Giemsa-stained smears, hemocultures and xenodiagnosis. T. cruzi was detected in three specimens of Didelphis marsupialis and T. (M.) theileri was detected in one specimen of Bos taurus. The parasites were isolated from triatomine feces, xenoculture and hemoculture. No evidence of human infection was detected in 58 inhabitants examined, as evaluated by indirect imunofluorescence technique using T. cruzi epimastigotes as antigens. These results show that T. vitticeps is still a sylvatic species although nymphs have been found inside the domicile. Thus, an epidemiological vigilance is necessary to know the behaviour of this species following the continuous modifications promoted by the presence of man.

Trypanosoma cruzi: Maintenance in Culture Modify Gene and Antigenic Expression of Metacyclic Trypomastigotes

In this study we examined whether the maintenance of Trypanosoma cruzi by long-time in axenic culture produces changes in gene expression and antigenic profiles. The studies were made with a Dm30L-clone from a low-virulent strain and a non-cloned virulent EP-strain of T. cruzi. Both parasites were maintained, for at least seven years, by successive alternate passage triatomine/mouse (triatomine condition), or by serial passage in axenic medium (culture condition). The comparison of the [35S]methionine metabolic labeling products of virulent and non-virulent parasites by 2D-SDS-PAGE, clearly indicates that the expression of metacyclic trypomastigotes (but not of epimastigotes) proteins have been altered by laboratory maintenance conditions. Western blot analysis of EP and Dm30L-epimastigotes using a serum anti-epimastigotes revealed that although most of antigens are conserved, four antigens are characteristics of triatomine condition parasites and three other are characteristics of culture condition parasites. Anti-metacyclics serum revealed significative differences in EP- and Dm30L-metacyclic trypomastigotes from triatomine condition. However, avirulent metacyclic forms were antigenically very similar. These results suggest that besides a possible selection of avirulent subpopulation from T. cruzi strains genetically heterogeneous when maintained by long time in axenic culture, changes in virulence might be due to post-translational modifications of the antigens induced by the absence of the natural alternability (vertebrate-invertebrate) in the life-cycle of T. cruzi

Schistosomiasis vaccine development: progress and prospects

The undisputed, worldwide success of chemotherapy notwithstanding, schistosomiasis continues to defy control efforts in as much rapid reinfection demands repeated treatment, sometimes as often as once a year. There is thus a need for a complementary tool with effect for the longer term, notably a vaccine. International efforts in this direction have been ongoing for several decades but, until the recombinant DNA techniques were introduced, antigen production remained an unsurmountable bottleneck. Although animal experiments have been highly productive and are still much needed, they probably do not reflect the human situation adequately and real progress can not be expected until more is known about human immune responses to schistosome infection. It is well-known that irradiated cercariae consistently produce high levels of protection in experimental animals but, for various reasons, this proof of principle cannot be directly exploited. Research has instead been focussed on the identification and testing of specific schistosome antigens. This work has been quite successful and is already at the stage where clinical trials are called for. Preliminary results from coordinated in vitro laboratory and field epidemiological studies regarding the protective potential of several antigens support the initiation of such trials. A series of meetings, organized earlier this year in Cairo, Egypt, reviewed recent progress, selecteded suitable vaccine candidates and made firm recommendations for future action including pledging support for large-scale production according to good manufacturing practice (GMP) and Phase I trials. Scientists at the American Centers for Disease Control and Prevention (CDC) have drawn up a detailed research plan. The major financial support will come from USAID, Cairo, which has established a scientific advisory group of Egyptian scientists and representatives from current and previous international donors such as WHO, NIAID, the European Union and the Edna McConnell Clark Foundation.

Immunization with PIII, a fraction of Schistosoma mansoni soluble adult worm antigenic preparation, affects nitric oxide production by murine spleen cells

Nitric oxide (NO) is an important effector molecule involved in immune regulation and defense. NO produced by cytokine-activated macrophages was reported to be cytotoxic against the helminth Schistosoma mansoni. Identification and characterization of S. mansoni antigens that can provide protective immunity is crucial for understanding the complex immunoregulatory events that modulate the immune response in schistosomiasis. It is, then, essential to have available defined, purified parasite antigens. Previous work by our laboratory identified a fraction of S. mansoni soluble adult worm antigenic preparation (SWAP), named PIII, able to elicit significant in vitro cell proliferation and at the same time lower in vitro and in vivo granuloma formation when compared either to SEA (soluble egg antigen) or to SWAP. In the present work we report the effect of different in vivo trials with mice on their spleen cells ability to produce NO. We demonstrate that PIII-immunization is able to significantly increase NO production by spleen cells after in vitro stimulation with LPS. These data suggest a possible role for NO on the protective immunity induced by PIII.

A comparative study on IgG-ELISA, IgM-IFT and Kato-Katz methods for epidemiological purposes in a low endemic area for schistosomiasis

The high sensitivity and the possibility of automation of the enzyme-linked-immunosorbent-assay (ELISA) has indicated this technique as one of the most useful serological test for epidemiological studies. In the present study, an ELISA for detection of IgG antibodies against adult worm antigens (IgG-ELISA) was investigated for epidemiological purposes, in a rural area of the municipality of Itariri (São Paulo, Brazil). Blood on filter paper (1,180 samples) from about 650 schoolchildren were submitted to ELISA and the data compared to the results of the parasitological method of Kato-Katz and also to the IgM-IFT (immunofluorescence test for IgM antibodies to gut associated antigens). The prevalence rates respectively of 8.5%, 43.0%, and 56.2% by the Kato-Katz, IgG-ELISA, and IgM-IFT methods suggest the poor sensitivity of the parasitological method for detection of Schistosoma mansoni eggs in individuals with low worm burden, situation commonly observed in low endemic areas. These results can partially explain the poor degree of agreement between the IgG-ELISA and the Kato-Katz, as suggested by the Kappa index of 0.170. Otherwise, the Kappa index of 0.675 showed substantial agreement between the two serological tests. Some discrepancy of results between the two serological techniques must be better investigated.

Immunofluorescence test on Schistosoma mansoni worm paraffin sections (IgM-IFT) for the study of schistosomiasis transmission in Campinas, São Paulo, Brazil

The detection of IgM antibodies for Schistosoma mansoni using gut-associated antigens (IgM-IFT) was compared to the parasitological Kato-Katz method for study of the transmission of schistosomiasis in an urban area in Campinas. About 400 schoolchildren whose ages ranged from 6 to 18 years, were observed for a period of two years. Blood samples on filter paper and fecal samples were collected, at intervals of six months. Serological (IgM-IFT) prevalence rates of 1.2%, 4.3%, 3.6%, 2.9% and 3.4% were obtained in five surveys carried out. S. mansoni eggs were detected in only one child out of the 225 children (0.4%) who were submitted to the Kato-Katz method (three slides for each fecal sample) in the 1st survey. Sixty eight children who submitted five blood samples, one for each survey, were found IFT negative throughout the study. No child was found to be IFT positive in all five surveys, and only four children showed IFT positive results in at least four surveys. Seroconversion from IFT negative to positive was observed from the 1st to the 2nd survey in six chidren, from the 2nd to the 3rd survey in three children, from the 3rd to the 4th survey in four children, and from the 4th to the 5th survey in two cases. However, confirmation of S. mansoni infection using the fecal examination was not possible in any of the cases. Also, in most of them the IFT result oscillated from negative to positive and vice versa. Our data implied that recent transmission of schistosomiasis in the study area was not possible.

Immunotherapy for Visceral Leishmaniasis: Ability of Factors Produced during Anti-leishmania Responses of Skin Test Positive Adults to Inhibit Peripheral Blood Mononuclear Cell Activities Associated with Visceral Leishmaniasis

The course of human Leishmania chagasi infections appears to be determined by the balance between type 1 (T1) CD4+ and CD8+ T suppressor (Ts) cell activities. Skin test positive adults living in hyperendemic areas who have no history of visceral leishmaniasis (VL) have T1 CD4+ T cell immunodominant responses against L. chagasi. The cytokines they secrete during anti-leishmania responses are a probable source of cytokines which inhibit the CD8+ Ts cells associated with VL. The ability of supernatants generated from peripheral blood mononuclear cells derived from skin test positive adults to reverse immune responses which appear to be mediated by CD8+ Ts cells was assessed in three sets of screening assays. The supernatants displayed three candidate factors. One, which could be explained by Leishmania antigens in the supernatant, decreased high endogenous IL-10 secretion characteristic of one class of VL patients. A second activity decreased high endogenous proliferation characteristic of the same class of patients without decreasing antigen specific proliferation. The third activity inhibited or killed CD8+ T cells but not CD4+ T cells. These activities might be useful in treating VL.

The Last Fifteen Years of Schistosomiasis in Venezuela: Features and Evolution

Control of schistosomiasis in Venezuela has been a topic of major interest and controversy among the metaxenic parasitosis. A small area of transmission of approximately 15,000 km2 was thought to be eradicated some years ago. However, some epidemiological characteristics of our transmission area have limited the success on the way toward eradication. Since 1945, when the Schistosomiasis Control Program started, the prevalence in the endemic area has decreased from 14% in 1943 to 1.4% in 1996. Until 1982, the surveillance of active cases was based on massive stool examination. Since then, the Schistosomiasis Research Group (SRG) recommended the additional use of serologic tests in the Control Program and the selective or massive chemotherapy depending on serological and parasitological prevalence of each community. At present, the real prevalence is underestimated due to the fact that approximately 80% of the individuals eliminate less than 100 eggs/g of feces. Those persons could be responsible for the maintenance of the foci going on and therefore limiting the impact of the control measures. Efforts of the SRG are being oriented toward improvement of immunodiagnostic tests by using defined antigens (enzymes) and chemically synthesized peptides, derived from relevant molecules of the parasite, either for antibodies or antigens search. On the other hand, introduction of snail competitors has been a biological weapon in the control of the intermediate host in certain areas. However, the recent reinfestation of water courses by Biomphalaria glabrata, the increased prevalence in some areas, together with important administrative changes at the Control Program of the Minister of Health, have arisen new questions and doubts, challenging the eradication strategy proposed during the last decade.

Characterization of Trypanosoma rangeli Strains Isolated in Central and South America: an Overview

Trypanosoma rangeli is a hemoflagelate parasite that infects domestic and sylvatic animals, as well as man, in Central and South America. T. rangeli has an overlapping distribution with T. cruzi, the etiological agent of Chagas disease, sharing several animal reservoirs and triatomine vectors. We have isolated T. rangeli strains in the State of Santa Catarina, in southern Brazil, which dramatically increased the distribution area of this parasite. This brief review summarizes several studies comparing T. rangeli strains isolated in Santa Catarina with others isolated in Colombia, Honduras and Venezuela. The different methods used include indirect immunofluorescence and western blot assays, lectin agglutination, isoenzyme electrophoresis and random amplified polymorphic DNA analysis, triatomine susceptibility, in vitro cell infection assays, and mini-exon gene analysis.

Improved Methods for the Diagnosis of African Trypanosomosis

The diagnosis of trypanosomosis in animals with low parasitaemia is hampered by low diagnostic sensitivity of traditional detection methods. An immunodiagnostic method based on a direct sandwich enzyme-linked immunosorbent assay (ELISA), using monoclonal antibodies, has been examined in a number of African laboratories for its suitability for monitoring tsetse control and eradication programmes. Generally, the direct sandwich ELISAs for the detection of trypanosomal antigens in serum samples have proved to be unsatisfactory with respect to diagnostic sensitivity when compared with traditional parasitological methods such as the dark ground/phase contrast buffy-coat technique. Consequently, antigen-detection systems exploiting various other direct, indirect and sandwich ELISA systems and sets of reagents are being developed to improve diagnosis. In addition, an existing indirect ELISA for the detection of antibodies has been improved and is being evaluated in the field in order to detect cattle that are or have been recently infected with trypanosomes. Developments and advantages of other diagnostic techniques, such as dip-stick assay and tests based on the polymerase chain reaction are also considered.

Humoral Immune Response Kinetics in Philander opossum and Didelphis marsupialis Infected and Immunized by Trypanosoma cruzi Employing an Immunofluorescence Antibody Test

Philander opossum and Didelphis marsupialis considered the most ancient mammals and an evolutionary success, maintain parasitism by Trypanosoma cruzi without developing any apparent disease or important tissue lesion. In order to elucidate this well-balanced interaction, we decided to compare the humoral immune response kinetics of the two didelphids naturally and experimentally infected with T. cruzi and immunized by different schedules of parasite antigens, employing an indirect fluorescence antibody test (IFAT). Both didelphids responded with high serological titers to different immunization routes, while the earliest response occurred with the intradermic route. Serological titers of naturally infected P. opossum showed a significant individual variation, while those of D. marsupialis remained stable during the entire follow-up period. The serological titers of the experimentally infected animals varied according to the inoculated strain. Our data suggest that (1) IFAT was sensitive for follow-up of P. opossum in natural and experimental T. cruzi infections; (2) both P. opossum and D. marsupialis are able to mount an efficient humoral immune response as compared to placental mammals; (3) experimentally infected P. opossum and D. marsupialis present distinct patterns of infection, depending on the subpopulation of T. cruzi, (4) the differences observed in the humoral immune responses between P. opossum and D. marsupialis, probably, reflect distinct strategies selected by these animals during their coevolution with T. cruzi.

Atypical Mucocutaneous Leishmaniasis Caused by Leishmania braziliensis in an Acquired Immunodeficiency Syndrome Patient: T-cell Responses and Remission of Lesions Associated with Antigen Immunotherapy

An atypical case of acquired immunodeficiency syndrome-associated mucocutaneous lesions due to Leishmania braziliensis is described. Many vacuolated macrophages laden with amastigote forms of the parasite were found in the lesions. Leishmanin skin test and serology for leishmaniasis were both negative. The patient was resistant to therapy with conventional drugs (antimonial and amphotericin B). Interestingly, remission of lesions was achieved after an alternative combined therapy of antimonial associated with immunotherapy (whole promastigote antigens). Peripheral blood mononuclear cells were separated and stimulated in vitro with Leishmania antigens to test the lymphoproliferative responses (LPR). Before the combined immunochemotherapy, the LPR to leishmanial antigens was negligible (stimulation index - SI=1.4). After the first course of combined therapy it became positive (SI=4.17). The antigen responding cells were predominantly T-cells (47.5%) most of them with CD8+ phenotype (33%). Very low CD4+ cells (2.2%) percentages were detected. The increased T-cell responsiveness to leishmanial antigens after combined therapy was accompanied by interferon-g (IFN-g) production as observed in the cell culture supernatants. In this patient, healing of the leishmaniasis lesions was associated with the induction of a specific T-cell immune response, characterized by the production of IFN-g and the predominance of the CD8+ phenotype among the Leishmania-reactive T-cells.