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ABSTRACT: Schistosomiasis is a neglected tropical disease that remains a considerable public health problem worldwide. Since the mainstay of schistosomiasis control is chemotherapy with a single drug, praziquantel, drug resistance is a concern. Here, we examined the in vitro effects of dermaseptin 01 (DS 01), an antimicrobial peptide found in the skin secretion of frogs of the genus Phyllomedusa, on Schistosoma mansoni adult worms. DS 01 at a concentration of 100 μg/ml reduced the worm motor activity and caused the death of all worms within 48 h in RPMI 1640 medium. At the highest sublethal concentration of antimicrobial peptide (75 μg/ml), a 100% reduction in egg output of paired female worms was observed. Additionally, DS 01 induced morphological alterations on the tegument of S. mansoni, and a quantitative analysis carried out by confocal microscopy revealed extensive destruction of the tubercles in a dose-dependent manner over the concentration range of 50-200 μg/ml. It was the first time that an anthelmintic activity towards schistosomes has been reported for a dermaseptin.
Chemistry & Biodiversity 03/2011; 8(3):548-58. · 1.80 Impact Factor
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ABSTRACT: Schistosomiasis is one of the world's greatly neglected tropical diseases, and its control is largely dependent on a single drug, praziquantel. Here, we report the in vitro effect of piplartine, an amide isolated from Piper tuberculatum (Piperaceae), on Schistosoma mansoni adult worms. A piplartine concentration of 15.8 μM reduced the motor activity of worms and caused their death within 24h in a RPMI 1640 medium. Similarly, the highest sub-lethal concentration of piplartine (6.3 μM) caused a 75% reduction in egg production in spite of coupling. Additionally, piplartine induced morphological changes on the tegument, and a quantitative analysis carried out by confocal microscopy revealed an extensive tegumental destruction and damage in the tubercles. This damage was dose-dependent in the range of 15.8-630.2 μM. At doses higher than 157.6 μM, piplartine induced morphological changes in the oral and ventral sucker regions of the worms. It is the first time that the schistosomicidal activity has been reported for piplartine.
Experimental Parasitology 02/2011; 127(2):357-64. · 2.12 Impact Factor
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ABSTRACT: Schistosomiasis affects more than 200 million people worldwide; another 600 million are at risk of infection. The schistosomulum stage is believed to be the target of protective immunity in the attenuated cercaria vaccine model. In an attempt to identify genes up-regulated in the schistosomulum stage in relation to cercaria, we explored the Schistosoma mansoni transcriptome by looking at the relative frequency of reads in EST libraries from both stages. The 400 genes potentially up-regulated in schistosomula were analyzed as to their Gene Ontology categorization, and we have focused on those encoding-predicted proteins with no similarity to proteins of other organisms, assuming they could be parasite-specific proteins important for survival in the host. Up-regulation in schistosomulum relative to cercaria was validated with real-time reverse transcription polymerase chain reaction (RT-PCR) for five out of nine selected genes (56%). We tested their protective potential in mice through immunization with DNA vaccines followed by a parasite challenge. Worm burden reductions of 16-17% were observed for one of them, indicating its protective potential. Our results demonstrate the value and caveats of using stage-associated frequency of ESTs as an indication of differential expression coupled to DNA vaccine screening in the identification of novel proteins to be further investigated as potential vaccine candidates.
Parasitology Research 01/2011; 108(1):123-35. · 2.15 Impact Factor
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ABSTRACT: To observe the effects of the parasitic infection on the biology of B. tenagophila, field and laboratory populations of this mollusk from Itariri, in Vale do Ribeira, Brazil, were experimentally infected. Each mollusk received 10 miracidia of Schistosoma mansoni (SJ lineage) and was observed throughout the parasite's development. The biological variables were compared according to the criteria "group" and "infectious phase". The main damage caused by the parasitic infection manifested itself in reproduction, longevity and lesions on the shell of the mollusks in the patent phase. An infection rate of 58.8% was observed. Microanatomical study of the mollusk's digestive gland and ovotestis revealed the presence of evolving larval forms and cercariae. It was concluded that the effects of the parasitic infection on both populations were moderate, despite the low survival rate of the infected mollusks, the damage did not prevent either reproduction or the elimination of cercariae, which continued for a long time.
Revista do Instituto de Medicina Tropical de São Paulo 04/2010; 52(2):101-5. · 1.00 Impact Factor
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Leonardo P Farias,
Fernanda C Cardoso,
Patricia A Miyasato,
Bogar O Montoya,
Cibele A Tararam,
Henrique K Roffato, Toshie Kawano,
Andrea Gazzinelli,
Rodrigo Correa-Oliveira,
Patricia S Coulson,
R Alan Wilson,
Sérgio C Oliveira,
Luciana C C Leite
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ABSTRACT: Schistosomiasis affects more than 200 million individuals worldwide, with a further 650 million living at risk of infection, constituting a severe health problem in developing countries. Even though an effective treatment exists, it does not prevent re-infection, and the development of an effective vaccine still remains the most desirable means of control for this disease.
Herein, we report the cloning and characterization of a S. mansoni Stomatin-like protein 2 (SmStoLP-2). In silico analysis predicts three putative sites for palmitoylation (Cys11, Cys61 and Cys330), which could contribute to protein membrane association; and a putative mitochondrial targeting sequence, similar to that described for human Stomatin-like protein 2 (HuSLP-2). The protein was detected by Western blot with comparable levels in all stages across the parasite life cycle. Fractionation by differential centrifugation of schistosome tegument suggested that SmStoLP-2 displays a dual targeting to the tegument membranes and mitochondria; additionally, immunolocalization experiments confirm its localization in the tegument of the adult worms and, more importantly, in 7-day-old schistosomula. Analysis of the antibody isotype profile to rSmStoLP-2 in the sera of patients living in endemic areas for schistosomiasis revealed that IgG1, IgG2, IgG3 and IgA antibodies were predominant in sera of individuals resistant to reinfection as compared to those susceptible. Next, immunization of mice with rSmStoLP-2 engendered a 30%-32% reduction in adult worm burden. Protective immunity in mice was associated with specific anti-rSmStoLP-2 IgG1 and IgG2a antibodies and elevated production of IFN-gamma and TNF-alpha, while no IL-4 production was detected, suggesting a Th1-predominant immune response.
Data presented here demonstrate that SmStoLP-2 is a novel tegument protein located in the host-parasite interface. It is recognized by different subclasses of antibodies in patients resistant and susceptible to reinfection and, based on the data from murine studies, shows protective potential against schistosomiasis. These results indicate that SmStoLP-2 could be useful in a combination vaccine.
PLoS Neglected Tropical Diseases 01/2010; 4(2):e597. · 4.69 Impact Factor
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ABSTRACT: Schistosoma mansoni is the major causative agent of schistosomiasis. The parasite takes advantage of host signals to complete its development in the human body. Tumor necrosis factor-alpha (TNF-alpha) is a human cytokine involved in skin inflammatory responses, and although its effect on the adult parasite's metabolism and egg-laying process has been previously described, a comprehensive assessment of the TNF-alpha pathway and its downstream molecular effects is lacking.
In the present work we describe a possible TNF-alpha receptor (TNFR) homolog gene in S. mansoni (SmTNFR). SmTNFR encodes a complete receptor sequence composed of 599 amino acids, and contains four cysteine-rich domains as described for TNFR members. Real-time RT-PCR experiments revealed that SmTNFR highest expression level is in cercariae, 3.5 (+/-0.7) times higher than in adult worms. Downstream members of the known human TNF-alpha pathway were identified by an in silico analysis, revealing a possible TNF-alpha signaling pathway in the parasite. In order to simulate parasite's exposure to human cytokine during penetration of the skin, schistosomula were exposed to human TNF-alpha just 3 h after cercariae-to-schistosomula in vitro transformation, and large-scale gene expression measurements were performed with microarrays. A total of 548 genes with significantly altered expression were detected, when compared to control parasites. In addition, treatment of adult worms with TNF-alpha caused a significantly altered expression of 1857 genes. Interestingly, the set of genes altered in adults is different from that of schistosomula, with 58 genes in common, representing 3% of altered genes in adults and 11% in 3 h-old early schistosomula.
We describe the possible molecular elements and targets involved in human TNF-alpha effect on S. mansoni, highlighting the mechanism by which recently transformed schistosomula may sense and respond to this host mediator at the site of cercarial penetration into the skin.
PLoS Neglected Tropical Diseases 01/2009; 3(12):e556. · 4.69 Impact Factor
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ABSTRACT: The single cell gel electrophoresis or the comet assay was established in the freshwater snail Biomphalaria glabrata. For detecting DNA damage in circulating hemocytes, adult snails were irradiated with single doses of 2.5, 5, 10 and 20 Gy of (60)Co gamma radiation. Genotoxic effect of ionizing radiation was detected at all doses as a dose-related increase in DNA migration. Comet assay in B. glabrata demonstrated to be a simple, fast and reliable tool in the evaluation of genotoxic effects of environmental mutagens.
Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis 07/2008; 654(1):58-63. · 2.85 Impact Factor
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ABSTRACT: Dominant lethal effects of the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) were evaluated in the freshwater snail Biomphalaria glabrata. Wild-type snails were exposed during 10 days to 50, 75 and 100ppm of 2,4-D dimethylamine salt (2,4-D DMA) and paired with non-exposed albino snails 1, 11, 25 and 40 days after the exposure. The offspring of the non-exposed albino snails was scored for lethal malformations. One day after the exposure, a significant effect was observed at 75 and 100ppm without a dose-response relationship. After 11 days, the effect was observed only at the highest dose. After 25 days, significant increases in the dominant lethal effects occurred at 50 and 75ppm; effects were directly related to the doses. Background levels of lethal malformations were resumed after 40 days. Although the major and direct measure of dominant lethal mutations is the rate of lethal malformations in the heterozygous offspring of the albino snails, the sensitivity of the assay was substantially increased with the evaluation of all non-viable embryos, that are the sum of those with lethal malformations, identified or not as wild-type.
Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis 01/2007; 611(1-2):83-8. · 2.85 Impact Factor
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ABSTRACT: The dominant lethal effects of gamma radiation of 60Co in the snail Biomphalaria glabrata were studied. Three groups of 13 wild-type snails were irradiated with single doses of 2.5; 10 and 20 Gy. Crossings were carried out at intervals of 7, 17, 23, 30 and 36 days after irradiation. The dominant lethal effect was observed only at the first crossing occurring 7 days after irradiation with 2.5 Gy. With 10 and 20 Gy, the induction of lethal mutations was detected at 7, 17 and 23 days after irradiation; a dose-response effect was observed. The effect was stronger 7 days after irradiation, decreasing in the succeeding crossings up to 30 days. Cell-killing effects on germ cells were detected in the crossings at 23 days and 30 days after irradiation with 20 Gy. After 36 days, frequencies of malformations resumed background levels; crossing rates partially recovered. These results show that gamma radiation affected all the stages of spermatogenesis. Germ cells at later phases were more sensitive to the mutagenic effect of radiation and the cell killing effects were observed on the youngest cells. This response was similar to the highly homogeneous pattern observed in widely different species and allowed us to estimate some parameters of spermatogenesis in B. glabrata.
Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis 08/2004; 561(1-2):139-45. · 2.85 Impact Factor
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Ricardo DeMarco,
Andre T Kowaltowski,
Abimael A Machado,
M Bento Soares,
Cybele Gargioni, Toshie Kawano,
Vanderlei Rodrigues,
Alda M B N Madeira,
R Alan Wilson,
Carlos F M Menck,
João C Setubal,
Emmanuel Dias-Neto,
Luciana C C Leite,
Sergio Verjovski-Almeida
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ABSTRACT: Using the data set of 180,000 expressed sequence tags (ESTs) of the blood fluke Schistosoma mansoni generated recently by our group, we identified three novel long-terminal-repeat (LTR)- and one novel non-LTR-expressed retrotransposon, named Saci-1, -2, and -3 and Perere, respectively. Full-length sequences were reconstructed from ESTs and have deduced open reading frames (ORFs) with several uncorrupted features, characterizing them as possible active retrotransposons of different known transposon families. Alignment of reconstructed sequences to available preliminary genome sequence data confirmed the overall structure of the transposons. The frequency of sequenced transposon transcripts in cercariae was 14% of all transcripts from that stage, twofold higher than that in schistosomula and three- to fourfold higher than that in adults, eggs, miracidia, and germ balls. We show by Southern blot analysis, by EST annotation and tallying, and by counting transposon tags from a Serial Analysis of Gene Expression library, that the four novel retrotransposons exhibit a 10- to 30-fold lower copy number in the genome and a 4- to 200-fold-higher transcriptional rate per copy than the four previously described S. mansoni retrotransposons [corrected]. Such differences lead us to hypothesize that there are two different populations of retrotransposons in S. mansoni genome, occupying different niches in its ecology. Examples of retrotransposon fragment inserts were found into the 5' and 3' untranslated regions of four different S. mansoni target gene transcripts. The data presented here suggest a role for these elements in the dynamics of this complex human parasite genome.
Journal of Virology 04/2004; 78(6):2967-78. · 5.40 Impact Factor
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Invertebrate Reproduction and Development 01/2004; 46(2-3):85-91. · 0.48 Impact Factor
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Sergio Verjovski-Almeida,
Ricardo DeMarco,
Elizabeth A L Martins,
Pedro E M Guimarães,
Elida P B Ojopi,
Apuã C M Paquola,
João P Piazza,
Milton Y Nishiyama,
João P Kitajima,
Rachel E Adamson, [......],
M Bento Soares,
Cybele Gargioni, Toshie Kawano,
Vanderlei Rodrigues,
Alda M B N Madeira,
R Alan Wilson,
Carlos F M Menck,
João C Setubal,
Luciana C C Leite,
Emmanuel Dias-Neto
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ABSTRACT: Schistosoma mansoni is the primary causative agent of schistosomiasis, which affects 200 million individuals in 74 countries. We generated 163,000 expressed-sequence tags (ESTs) from normalized cDNA libraries from six selected developmental stages of the parasite, resulting in 31,000 assembled sequences and 92% sampling of an estimated 14,000 gene complement. By analyzing automated Gene Ontology assignments, we provide a detailed view of important S. mansoni biological systems, including characterization of metazoa-specific and eukarya-conserved genes. Phylogenetic analysis suggests an early divergence from other metazoa. The data set provides insights into the molecular mechanisms of tissue organization, development, signaling, sexual dimorphism, host interactions and immune evasion and identifies novel proteins to be investigated as vaccine candidates and potential drug targets.
Nature Genetics 11/2003; 35(2):148-57. · 35.53 Impact Factor
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ABSTRACT: Mutagens in the environment may represent a long-term risk for ecosystems. The reproductive potential of populations can be affected by alterations in the fecundity and offspring viability caused by germ cell mutations. Despite the ecological relevance of these effects, there are few studies on germ cell mutagenicity in natural populations. Biomphalaria glabrata was chosen for this study because of the scarcity of data on freshwater invertebrates and the ecological importance of this group. The aim of this study was to establish a germ cell mutagenicity test in B. glabrata by using a similar approach to that used in the dominant lethal test in rodents. Mitomycin C was used as a direct mutagen and cyclophosphamide as a mutagen that requires metabolic activation. Wild-type snails were exposed for 10 days to three concentrations of each agent and crossed with non-exposed albino snails at the end of the treatment. The total frequencies of malformations were analyzed in the offspring of wild-type snails; among the offspring of albino snails, only the heterozygous wild-type embryos were analyzed for malformations. Both agents induced germ cell mutations. The analysis of the offspring of the wild-type snails showed an effect of the exposure up to approximately 5 days after the end of the treatment with cyclophosphamide; the effect of mitomycin C was observed until 45 days after the end of the exposure. There was an increase in the frequencies of malformations in the wild-type offspring of the non-exposed albino snails crossed with the wild-type snails exposed to both agents. The dominant lethal test in B. glabrata proposed in this work is easy to perform, efficient, specific and sensitive in the evaluation of germ cell mutations induced by reference mutagens. The possibility of expanding its use to environmental biomonitoring studies seems very promising and worth trying.
Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis 05/2003; 536(1-2):145-54. · 2.85 Impact Factor
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ABSTRACT: Biomphalaria tenagophila é um hermafrodita simultâneo cujo investimento na função masculina foi avaliado pela contagem de espermatozóides do ovoteste e da vesícula seminal, sob diferentes condições experimentais, durante o processo de cópula. Foi criada uma nova técnica para contar espermatozóides separados em uma solução de alquilbenzeno sulfonato de sódio. Em todas as condições experimentais, a vesícula seminal teve mais do dobro de espermatozóides do ovoteste: da ordem de uma centena de milhar para três ou quatro centenas de milhares. Há evidências de que os espermatozóides transferidos no processo de cópula procederam da vesícula seminal do doador e que o equilíbrio do número de espermatozóides na vesícula seminal era restabelecido em menos de 24 horas.
Revista Brasileira de Biologia. 01/1998;
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ABSTRACT: A total of 909 Biomphalaria tenagophila were collected from two areas in Guarulhos (Metropolitan area of São Paulo, São Paulo State, Brazil) to assess larval trematode infections. In all collection sites, only this species was found and 183 (20.13%) harbored trematode infections. In these collections, four morphologically distinguishable types of cercariae were identified by confocal microscopy. Xiphidiocercaria (Cercaria lutzi) was the most common type of cercaria recovered, contributing 76.5% of all infections. Schistosoma mansoni cercariae were recovered and comprised the total of 13.11%. Strigea cercaria (Cercaria caratinguensis) and Brevifurcate pharyngeate Clinostomatoide cercaria (Cercaria ocellifera) contributed 8.33% and 2.22% of all infections, respectively. Double infections (S. mansoni and C. lutzi) were found in twelve snails, contributing 6.55% of all infections. In all sites studied, small vertebrates were found in snail habitats and it was observed human contact with the water. The presence of trematode infected snails in large cities has public health implications. It further provides a starting point for some comprehensive studies on snail-related aspects of transmission and biology of trematode of medical and veterinary importance.
Revista do Instituto de Medicina Tropical de São Paulo 51(2):77-82. · 1.00 Impact Factor
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ABSTRACT: Experiments were carried out to test the susceptibility of Biomphalaria tenagophila to the infection with strain SJ of Schistosoma mansoni in the F1, F2 and non-selected parental generation. The potential adaptation of B. tenagophila to desiccation, in healthy mollusks and those exposed to the larvae of S. mansoni of the F1, F2 and non-selected parental generations was also studied. The presence of mucus and soil, at the shell opening, protected the snails against desiccation, favoring survival. The healthy mollusks performed more attempts against desiccation than those exposed to the larvae of the parasite. The mortality rate, during desiccation, was higher among mollusks that remained buried and with the shell opening unobstructed. During the desiccation period the stage of development of the parasite was influenced by the weight loss and the survival of the snails. The longer the period of desiccation, the greater was the weight loss observed, abbreviating survival. The non-selected parental generation was more sensitive to desiccation than the F1 and F2 generations, both in healthy mollusks and in those exposed to S. mansoni larvae. Healthy mollusks were more resistant to desiccation than those exposed to the larvae of the S. mansoni. Desiccation did not interrupt the development of S. mansoni larvae in mollusks, causing a delay in the cercariae elimination. The susceptibility of B. tenagophila to the SJ strain of S. mansoni, in mollusks maintained in water during the larvae incubation period, was similar in all three generations.
Revista do Instituto de Medicina Tropical de São Paulo 44(4):191-201. · 1.00 Impact Factor
