RESUMEN
Taenia solium is a helminth parasite that causes 2 diseases in humans: cysticercosis and taeniasis. The establishment of T. solium metacestodes in the central nervous system causes neurocysticercosis, while development of the adult tapeworm in the small intestine causes taeniasis. Serological diagnosis of neurocysticercosis is performed by Western blot with an enriched fraction of glycoproteins that has been extensively used for clinical diagnosis and epidemiological surveys. The lectin-bound fraction that is used for this assay contains 7 antigenic glycoproteins. These antigenic proteins are considered to be highly specific for cysticercosis when tested with heterologous parasitic diseases. However, recent studies show that people with taeniasis have cross-reactive antibodies against the neurocysticercosis diagnostic glycoproteins and vice versa. Nevertheless, it is not known if these diagnostic proteins are expressed in the adult stage of the parasite. In this paper, we describe the location of 3 of these glycoproteins in T. solium adults and cysticerci using polyclonal antibodies raised against a synthetic peptide based on the amino acid sequence of TS14, a recombinant protein T24H, and the native GP50. The glycoproteins' distribution was different in invaginated and evaginated cysticerci as well as in adult tapeworms. Specifically, the 3 glycoproteins studied were differentially expressed during embryogenesis. Our findings indicate that expression of the diagnostic glycoproteins is developmentally regulated; this is noteworthy since these glycoproteins are considered specific for the diagnosis of neurocysticercosis but nevertheless are present in different structures throughout the development of T. solium. Here we describe the glycoprotein expression and localization, which can be important in understanding their biological functions. In addition, our results help clarify the cross-reaction observed between people with neurocysticercosis and taeniasis to TS14, T24H, and GP50, which are used as diagnostic antigens for neurocysticercosis.
Asunto(s)
Glicoproteínas/análisis , Neurocisticercosis/diagnóstico , Taenia solium/química , Teniasis/diagnóstico , Animales , Anticuerpos Antihelmínticos/inmunología , Antígenos Helmínticos/análisis , Antígenos Helmínticos/inmunología , Antígenos Helmínticos/metabolismo , Western Blotting , Reacciones Cruzadas , Cysticercus/anatomía & histología , Cysticercus/química , Cysticercus/aislamiento & purificación , Glicoproteínas/inmunología , Glicoproteínas/metabolismo , Cabras , Humanos , Sueros Inmunes/inmunología , Inmunohistoquímica , Neurocisticercosis/inmunología , Conejos , Taenia solium/crecimiento & desarrollo , Taenia solium/aislamiento & purificación , Teniasis/inmunologíaRESUMEN
Taeniasis/cysticercosis caused by the tapeworm Taenia solium is a parasite disease transmitted among humans and pigs, the main intermediate host. The larvae/cysts can lodge in several tissues of the pig, i.e. skeletal muscles and different locations of the central nervous system. The molecular mechanisms associated to tissue preferences of the cysts remain poorly understood. The major public health concern about this zoonosis is due to the human infections by the larval form in the central nervous system, causing a highly pleomorphic and debilitating disease known as neurocysticercosis. This study was aimed to explore the 2DE protein maps of T. solium cysts obtained from skeletal muscles and central nervous system of naturally infected pigs. The gel images were analyzed through a combination of PDQuest™ and multivariate analysis. Results showed that differences in the protein patterns of cysts obtained from both tissues were remarkably discrete. Only 7 protein spots were found specifically associated to the skeletal muscle localization of the cysts; none was found significantly associated to the central nervous system. The use of distinct protein fractions of cysts allowed preliminary identification of several tissue-specific antigenic bands. The implications of these findings are discussed, as well as several strategies directed to achieve the complete characterization of this parasite's proteome, in order to extend our understanding of the molecular mechanisms underlying tissue localization of the cysts and to open avenues for the development of immunological tissue-specific diagnosis of the disease.
Asunto(s)
Encéfalo/parasitología , Cisticercosis/veterinaria , Cysticercus/química , Proteínas del Helminto/análisis , Músculo Esquelético/parasitología , Enfermedades de los Porcinos/parasitología , Taenia solium/química , Animales , Cisticercosis/parasitología , Cysticercus/aislamiento & purificación , Electroforesis en Gel Bidimensional , Sus scrofa , Porcinos , Taenia solium/aislamiento & purificaciónRESUMEN
Previously, we have studied the effect of the gold-compound auranofin (AF) on both thioredoxin-glutathione reductasa (TGR) activity and viability of Taenia crassiceps cysticerci. It was demonstrated that micromolar concentrations of AF were high enough to fully inhibit TGR and kill the parasites. In this work, the dynamics of changes in the glutathione pool of T. crassiceps cysticerci following the addition of AF, was analyzed. A dose-dependent decrease in the internal glutathione concentration, concomitant with an increase in ROS production was observed. These changes were simultaneous with the formation of glutathione-protein complexes and the export of glutathione disulfide (GSSG) to the culture medium. Incubation of cysticerci in the presence of both AF and N-acetyl cysteine (NAC) prevents all the above changes, maintaining cysticerci viability. By contrast, the presence of both AF and buthionine sulfoximine (BSO) resulted in a potentiation of the effects of the gold compound, jeopardizing cysticerci viability. These results suggest the lethal effect of AF on T. crassiceps cysticerci, observed at micromolar concentrations, can be explained as a consequence of major changes in the glutathione status, which results in a significant increase in the oxidative stress of the parasites.
Asunto(s)
Auranofina/toxicidad , Glutatión/análisis , Oxidantes/toxicidad , Estrés Oxidativo , Taenia/química , Taenia/efectos de los fármacos , Acetilcisteína/metabolismo , Animales , Antioxidantes/metabolismo , Butionina Sulfoximina/metabolismo , Cysticercus/química , Cysticercus/efectos de los fármacos , Cysticercus/fisiología , Especies Reactivas de Oxígeno/análisis , Análisis de Supervivencia , Taenia/fisiologíaRESUMEN
The host-parasite relationship in cestode infections is complex. One feature of this bidirectional molecular communication is the uptake of host proteins by the parasite. Here we describe the presence of several host proteins in the vesicular fluid of Taenia solium cysticerci dissected from the central nervous system and the skeletal muscle of naturally infected pigs. Using two-dimensional electrophoresis we compared the protein patterns of vesicular fluids of cysticerci vs. the sera of cysticercotic pigs. We found that the vesicular fluids of both groups of cysts showed 17 protein spots matching with the pig's sera spots. After mass spectrometry sequencing of these spots, five host proteins were identified: hemoglobin, albumin, serpin A3-8, haptoglobin, rho GTPase-activating protein 36-like. Three of the 17 spots corresponded to host protein fragments: hemoglobin, albumin and serpin A3-8. IgG heavy and light chains were also identified by Western blot using a specific antibody. Quantitative estimations indicated that the host proteins represented 11-13% of the protein content in the vesicular fluids. We also calculated the relative abundance of these host proteins in the vesicular fluids; all were represented in similar relative abundances as in host sera. This suggests that uptake of host proteins by cysticerci proceeds through an unspecific mechanism such as non-specific fluid pinocytosis.
Asunto(s)
Cisticercosis/veterinaria , Proteínas/análisis , Enfermedades de los Porcinos/parasitología , Porcinos/sangre , Taenia solium/química , Vesículas Transportadoras/química , Secuencia de Aminoácidos , Análisis de Varianza , Animales , Western Blotting , Encéfalo/parasitología , Cisticercosis/sangre , Cisticercosis/parasitología , Cysticercus/química , Electroforesis en Gel Bidimensional , Interacciones Huésped-Parásitos , Espectrometría de Masas , Músculo Esquelético/parasitología , Proteínas/química , Enfermedades de los Porcinos/sangreRESUMEN
Neurocysticercosis is an endemic parasitic disease caused by Taenia solium larva. Although the mechanism of infection is not completely understood, it is likely driven by proteolytic activity that degrades the intestinal wall to facilitate oncosphere penetration and further infection. We analyzed the publicly available T. solium EST/DNA library and identified two contigs comprising a full-length cDNA fragment very similar to Echinococcus granulosus Ag5 protein. The T. solium cDNA sequence included a proteolytic trypsin-like-domain in the C-terminal region, and a thrombospondin type-1 adherence-domain in the N-terminal region. Both the trypsin-like and adherence domains were expressed independently as recombinant proteins in bacterial systems. TsAg5 showed marginal trypsin-like activity and high sequence similarity to Ag5. The purified antigens were tested in a Western immunoblot assay to diagnose human neurocysticercosis. The sensitivity of the trypsin-like-domain was 96.36% in patients infected with extraparenchymal cysts, 75.44% in patients infected with multiple cysts, and 39.62% in patients with a single cyst. Specificity was 76.70%. The thrombospondin type-1 adherence-domain was not specific for neurocysticercosis.
Asunto(s)
Antígenos Helmínticos , Cysticercus/metabolismo , Proteínas del Helminto , Neurocisticercosis/diagnóstico , Taenia solium/metabolismo , Teniasis/diagnóstico , Tripsina , Animales , Antígenos Helmínticos/química , Antígenos Helmínticos/genética , Antígenos Helmínticos/metabolismo , Cysticercus/química , Cysticercus/genética , Cysticercus/crecimiento & desarrollo , Proteínas del Helminto/química , Proteínas del Helminto/genética , Proteínas del Helminto/metabolismo , Humanos , Datos de Secuencia Molecular , Neurocisticercosis/parasitología , Estructura Terciaria de Proteína , Porcinos , Taenia solium/química , Taenia solium/genética , Taenia solium/crecimiento & desarrollo , Teniasis/parasitología , Tripsina/química , Tripsina/genética , Tripsina/metabolismoRESUMEN
A scolex protein antigen (SPA) was prepared from cysticerci of Taenia solium obtained from naturally infected pigs. Yorkshire pigs were vaccinated with SPA plus incomplete Freund's adjuvant (IFA) or with SPA plus Corynebacterium parvum (CP). Controls were given IFA plus phosphate-buffered saline (PBS) or CP plus PBS. All animals were given three subcutaneous injections at 20-day intervals. Ten days after the third injection, the pigs were fed with 10(4) viable eggs of T. solium. All pigs developed a delayed type hypersensitivity, and a transient eosinophilia after the first dose of vaccine. High titers of specific antibodies were detected in the sera of vaccinated animals and in infected controls. A protection level of 71.43% was recorded in animals vaccinated with SPA plus IFA and of 75.00% in those vaccinated with SPA plus CP.