Deteccion de portadores asintomaticos de quistes hidatidicos: aumento de la especificidad del ensayo immunoenzimatico

Una de las actividades que realizan los programas de control de hidatidosis causada por Echinococcus granulosus en la Republica Argentina es la busqueda de portadores asintomaticos de quistes hidatidicos entre la poblacion general que habita las areas de risco, mediante un ensayo inmunoenzimatico (EIE) con antigeno de liquido hidatidico total (EIE-ALHT) que selecciona los posibles portadores. En base a la experiencia recogida se ha observado que dependiendo de la prevalencia del area, entre el 10 por cento y el 30 por cento de las personas seleccionadas por el EIE con valores de densidad optica (DO * DO + 4S no presentan imagens compatibles con quistes hidatidicos. El proposito del estudio fue mejorar la especificidad de la prueba. Con ese fin, se evaluo la influencia de la modificacion de la oferta antigenica y el efecto de la absorcion de los anticuerpos anti-componentes sericos ovinos y anti-fosforilcolina de los sueros en estudios, 114 sueros no hidatidicos seleccionados por su alto nivel de reacciones cruzadas en EIE-ALHTy 118 sueros hidatidicos se estudiaron frente a 4 fracciones antigenicas de liquido hidatidico ovino. El EIE que empleo la fraccion antigenica S2B conjuntamente con la absorcion previa de los sueros (EIE-S2B/A) fue el sistema que mejor discrimino los sueros hidatidicos de los no hidatidicos

[Detection of asymptomatic carriers of hydatid cysts: specificity increase of the immunoenzyme assay]. / Detección de portadores asintomáticos de quistes hidatídicos: aumento de la especificidad del ensayo inmunoenzimático.

An enzymoimmunoassay (EIE) as a screening test to select potential asymptomatic cyst carriers among the general population of areas under risk is being used in programs for the control of hydatic diseases caused by Echinococcus granulosus in Argentina. The experience obtained up to date, applying this assay in population surveys, indicates that depending on the prevalence in the area 10% to 30% of the individuals selected did not show images compatible with hydatic cysts. The purpose of the present study was to improve the specificity of the test. To this purpose, the influence of the modification of the antigenic availability and the effect of the absorption from the serum samples of antibodies anti-normal ovine sera and anti-phosphorylcholine was evaluated. One hundred and fourteen non hydatic sera selected because of their high cross reactivity in EIE using the whole hydatid antigen (WHA) and 118 hydatid sera, were studied with four fractions of ovine hydatid cyst fluid. The EIE employing the S2B antigenic fraction with previous absorption of the sera (EIE-S2B/A) was the system that discriminated better hydatid sera from non hydatid sera with high levels of cross reactivity. The replacement of the EIE employing WHA by the EIE-S2B/A system, for the active search of asymptomatic cyst carriers in field conditions, is proposed. The four antigenic fractions were analyzed by double diffusion and SDS-PAGE. The S2B fraction revealed a high content of parasitic components of less than 30 Kd which probably includes antigen B and subunits or fragments of antigen 5.

Standardization and evaluation of an enzyme immunoassay as a screening test for the seroepidemiology of human hydatidosis.

The immunodiagnosis of human hydatidosis caused by Echinococcus granulosus is based on the detection of antibodies against arc 5 antigens by the double diffusion (DD5) or counterimmunoelectrophoresis (CIE5) tests. However, neither of those tests is practical for seroepidemiological surveys in which a great number of samples have to be processed. The present study was carried out to evaluate if a solid phase enzyme immunoassay (EIA) standardized to detect total antibodies against E. granulosus cyst fluid antigens would permit the selection of sera with antibody activity against arc 5 antigens detectable by DD5. Groups of sera representing 5 possible results with DD5 were studied by the EIA proposed as a screening test. Using the EIA all the sera with antibody activity against arc 5 antigens were differentiated from all those which did not produce any precipitation band in DD5. It was also possible to differentiate those with no antibodies from all those with antibodies against 3 non-arc 5 antigens, most with antibodies against 2 non-arc 5 antigens and about a fifth with antibodies against 1 non-arc 5 antigen. It is concluded that the EIA, standardized as described, is capable of detecting with a high degree of efficiency all the sera that should be tested by DD5 and excludes most of those in which results with DD5 have no diagnostic significance.

Application of antigens from Taenia hydatigena cyst fluid for the immunodiagnosis of human hydatidosis.

Fluid was collected from cysts of Taenia hydatigena in 60 adult sheep and fluid from each animal pooled separately. By double diffusion antigen 5 was demonstrated in all pools but one. The criteria are described for selection and standardization of these preparations for use as antigens for the immunodiagnosis of human hydatid disease. Sera from 50 persons harbouring hydatid cysts and from 50 patients with other disease conditions were examined by the arc-5 double-diffusion test, using two antigens prepared from Echinococcus granulosus and T. hydatigena cyst fluids, respectively. The results showed that a higher diagnostic sensitivity was obtained with the hydatid antigen. The significance of the findings is discussed in terms of their application to human immunodiagnosis in areas where hydatidosis, but not cysticercosis, is rare in livestock.

La Hidatidosis en el Perú: importancia del inmunodiagnóstico y de los estudios seroepidemiológicos / Hydatidosis in Peru: importance of the inmunodiagnosis and the seroepidemiologycal studies

En el presente estudio se describe la aplicación de pruebas estandarizadas para el diagnóstico inmunológico de la hidatidosis humana en el Instituto de Medicina Tropical "Daniel A. Carrión", Universidad Nacional Mayor de San Marcos, Lima, Perú. Estas se emplearon para obtener la conformación prequirúrgica de la parasitosis en situaciones clínicas, y para la detección de portadores asintomáticos de quistes hidatídicos mediante encuestas. Los resultados se utilizaron como fuente de datos para evaluar la calidad de la información que sobre el número de casos de la afección en distintas áreas ecológicas, se obtiene a partir de los casos hospitalarios notificados a los niveles oficiales. Los resultados demuestran la utilidad que puede brindar la extensión de la cobertura de los servicios para el inmunodiagnóstico de la hidatidosis a las áreas endémicas del país con fines de atención primaria y vigilancia epidemiológica. Esto permitiría obtener mejor información sobre el impacto que la contaminación ambiental por los huevos de Echinococcus granulosus produce en la población humana y medir los cambios que resultarían de la efectiva aplicación de medidas de control.

Immunodiagnosis of human hydatid disease: applications and contributions to a control program in Argentina.

The sequential development of the hydatid immunodiagnostic activities of the control program of the Province of Neuquén, Argentina is described. Test results were used to obtain immunological confirmation of clinical cases and to detect asymptomatic cyst carriers amongst residents of rural endemic areas. The information was also valuable for improving the accuracy of prevalence estimates of human hydatidosis and the quality of surveillance data in different areas of the Province, characterized by varying degrees of environmental contamination by Echinococcus granulosus. In population groups examined by radiologic and immunologic methods, the latter detected more cases. When only immunodiagnostic surveys were carried out, mostly liver but also pulmonary hydatidosis cases were detected. This experience illustrates the advantages which may be obtained in endemic areas through the local application of hydatid immunodiagnosis based on arc 5 positivity.

Significance of hydatid immunodiagnostic surveys to health care and estimation of prevalence in the Argentine Province of Chubut.

More hydatidosis cases were detected in an immunodiagnostic survey of rural schoolchildren from an endemic area in Argentina by the arc 5 double diffusion (DD5) test than by indirect hemagglutination (IHA) and latex agglutination (LA) tests. Evidence of infection was obtained by clinical, radiologic, echographic, and/or computerized axial tomography examinations in only one of three DD5-negative individuals, and in 2 of 4 students showing a questionable reaction of identity with arc 5 in DD5, who were positive in LA and/or IHA. In contrast, cysts could be demonstrated in all DD5-positive cases, whether positive or negative in the agglutination procedures. These cysts in DD5-positive cases included the smallest cysts (1-1.2 cm wide) detected in surveys to date. Comparison of data obtained in the immunodiagnostic survey and in a review of hospital records was used to assess the relative contribution of each method in estimating the prevalence of infection. The findings are discussed in terms of the advantages and limitations of carrying out immunodiagnostic surveys of residents of endemic areas for purposes of primary medical care and surveillance.

Broadly reacting precipitating and agglutinating antigen of leptospirae.

A saprophytic Leptospira biflexa strain of equine origin was found which cross-reacts with immune rabbit antisera to 14 pathogenic Leptospira interrogans serotypes. Sera from goats experimentally inoculated with the saprophyte showed multiple low-level cross-agglutination reactions against a battery of live L. interrogans serotypes. Sonically treated and saline-extracted suspensions of the L. biflexa strain and serotypes canicola, icterohaemorrhagiae, and pomona yielded a common precipitating protein antigen that was detected by immunodiffusion and immunoelectrophoresis with all of the antileptospiral sera examined. In cross-absorption and gel diffusion tests, the precipitinogen from each of the strains was shown to be identical. Formaldehyde treatments and heating at 100 degree C suggest that the cellular location of the common antigen is either somatic or subsurface, and Sephadex G-200 gel filtration enabled the isolation of the active fraction of the L. biflexa antigen. Monoprecipitin sera against the common antigen of L. biflexa were produced by immunizing rabbits with specific precipitates in agar. In gel diffusion and immunoelectrophoresis tests the antisera with each of the soluble antigens developed a single precipitin formation, and the antisera agglutinated formolized and heated whole-cell suspensions of serotypes canicola, icterohaemorrhagiae, and pomona at low dilutions. The soluble L. biflexa antigen was evaluated as an immunogen and in passive immunity tests for protection against death and kidney infection in hamsters. No cross-protection occurred when the hamsters were challenged with virulent leptospires. In contrast, the animals vaccinated or administered hamster immune serum before challenge died earlier than the control animals.

Immunoelectrophoresis tests showing Echinococcus granulosus arc 5 in human cases of Echinococcus vogeli and cysticercosis-multiple myeloma.

Human sera from one case of polycystic hydatidosis due to Echnincoccus vogeli and from a case of cysticercosis and multiple myeloma were positive to the immunoelectrophoresis (IEP) test for hydatidosis based on the E. granulosus arc 5 positivity criterion. Arc 5 can therefore be elicited in IEP tests of human sera not only by E. granulosus and E. multilocularis but also by E. vogeli antigens. Whether the cross reaction observed in the second serum was due to the multiple myeloma or to the cysticerci remains to be determined. Although arc 5 antigens are known to be present in Taenia hydatigena cyst fluid this is the first report of an arc 5 due to antigens other than Echinococcus in IEP of human sera.

Detection of antibodies against Echinococcus granulosus arc 5 antigens by double diffusion test.

This report deals with an evaluation of a double diffusion (DD5) test which employs a control antiserum against Echinococcus granulosus arc 5 antigens to recognize arc 5-positive sera by a reaction of identity. The DD5 test was more sensitive than the immuno-electrophoresis (IEP5) test based on the same positivity criterion and was equally specific for the immunodiagnosis of human hydatidosis. Its greater simplicity suggest its application as a substitute for the IEP5 test in diagnosis, particularly areas. The application of both DD5 and latex agglutination tests is recommended for hydatid immunodiagnosis on the basis of the present findings.

Detection of the Echinococcus granulosus diagnostic arc 5 in sera from patients with surgically-confirmed E. multilocularis infection.

The Echinococcus granulosus diagnostic arc 5 was revealed by sera from an Alaskan and a Swiss patient with surgically confirmed E. multilocularis infections. The possibility of each patient harboring a concurrent infection with both parasites may be disregarded on the basis of radiologic, scintillographic and surgical data. This conclusion is compatible with ecologic and epidemiologic considerations. These observations suggest that the immunoelectrophoresis test based on arc 5 positivity is not E. granulosus-specific as originally described.

Comparative antigenic characterisation of Echinococcus granulosus and Taenia hydatigena cyst fluids by immunoelectrophoresis.

Hydatid cyst fluid from Echinococcus granulosus (HCF) and cyst fluid from Taenia hydatigena (TCF) cysts were compared in reciprocal immunoelectrophoresis (IEP) tests using homologous and heterologous antisera which were free of antibodies to host serum contaminants. The antigens for the E granulosus arc 5 were demonstrated in TCF. Antibody activity to these and other antigens common to HCF and TCF was removed from homologous antisera by absorptions with the heterologous antigenic preparation. Antigens not shared by the two metacestodes fluids were then demonstrated by IEP tests. These findings are discussed in terms of their significance to phylogenetic and immunodiagnostic studies of these parasites in their immediate hosts.