Myriam Lorca

Prevention and control of cystic echinococcosis

Human cystic echinococcosis (hydatid disease) continues to be a substantial cause of morbidity and mortality in many parts of the world. Elimination is difficult to obtain and it is estimated that, using current control options, achieving such a goal will take around 20 years of sustained efforts. Since the introduction of current (and past) hydatid control campaigns, there have been clear technological improvements made in the diagnosis and treatment of human and animal cystic echinococcosis, the diagnosis of canine echinococcosis, and the genetic characterisation of strains and vaccination against Echinococcus granulosus in animals. Incorporation of these new measures could increase the efficiency of hydatid control programmes, potentially reducing the time required to achieve effective prevention of disease transmission to as little as 5-10 years.

Comparative Analysis of the Diagnostic Performance of Six Major Echinococcus granulosus Antigens Assessed in a Double-Blind, Randomized Multicenter Study

ABSTRACT The serodiagnosis of hydatid disease is a valuable instrument for clinical diagnosis and epidemiological surveillance of high-risk populations. In the past decade a wealth of reports on the diagnostic performance of numerous antigens have been produced. However, their diagnostic value has been estimated under different conditions, using different serum collection, therefore precluding their direct comparison. Here we report an unbiased comparison of the same batch of six major E. granulosus antigens, namely, hydatid cyst fluid (HCF), native antigen B (AgB), two recombinant AgB subunits, an AgB-derived synthetic peptide, and recombinant cytosolic malate dehydrogenase from E. granulosus (EgMDH), against the same serum collection. The double-blind analysis was performed using a standardized protocol and receiver operating characteristic (ROC) data analysis by a network of six South American laboratories. High intercenter reproducibility was attained, and the intralaboratory analysis allowed the comparative ranking of the antigen panel. HCF, AgB, and its AgB8/1 subunit exhibited equivalent diagnostic efficiencies, 81.4% ± 0.5%, 81.3% ± 0.6%, and 81.9% ± 2.0%, respectively; with a more favorable balance toward specificity in the case of the last antigen. The diagnostic efficiencies for the other three antigens were 76.8% ± 6.8%, 69.1% ± 2.7%, and 66.8% ± 2.1%, for the peptide, the AgB8/2 subunit, and the EgMDH, respectively. The study also included an analysis of batch-to-batch variation in the diagnostic performance of different HCF regional preparations. Based on these results, a suggested recommendation on the use of these antigens was drawn.

SEROPREVALENCE OF TOXOPLASMA GONDII INFECTION IN SHEEP AND ALPACAS (LLAMA PACOS) IN CHILE

Serum samples from 408 sheep from different regions of Chile and 447 alpacas (Llama pacos) from the north of the country were tested for Toxoplasma gondii antibodies. The indirect haemagglutination test (IHAT) was used in both species and the indirect immunofluorescence test (IIFT) was also used on the sheep samples in order to compare the performance of the tests in that species. In both tests, titers > or = 1:16 were considered diagnostically significant. Sera from 49 sheep (12%) were positive to T. gondii antibodies by the IHAT. When using the IIFT, 114 sheep sera (28%) were positive. The different results obtained in sheep sera between the tests were significant (p < 0.0001). No differences were observed between geographical locations or sex of the sampled sheep regarding serological detection of T. gondii antibodies in sheep. As expected, adult sheep showed higher T. gondii reactivity than young sheep (p = 0.0008). The corrected prevalence of toxoplasmosis in alpaca was 16.3% (32 positive out of 447). The rather low prevalence in alpacas may be associated with their extensive management as well as the extreme climatic conditions of The Andes which apparently would not be favorable for the transmission of the parasite.

Development of an immunoenzymatic assay for the detection of human antibodies against Trypanosoma cruzi calreticulin, an immunodominant antigen.

We have developed an indirect immunoenzymatic assay (ELISA) for the detection of human antibodies against calreticulin (formerly known as Tc45), a dimorphic Trypanosoma cruzi antigen, described in our laboratory. PVC microtitration plates were sensitized with the monoclonal anti-calreticulin antibody (MoAb) and reacted with calreticulin present in a partially purified preparation. The presence of anti-T. cruzi calreticulin IgG in sera from infected individuals was tested. The data generated with this assay were validated by correlation, in a regression analysis, with those obtained by an indirect immunoradiometric assay (IRMA). From the 12 seropositive sera (as defined by a commercial test), eight came out positive and four negative in both assays. The 12 human sera were also analyzed in direct immunometric assays (ELISA and IRMA), where the solid phase was sensitized with a whole parasite extract. The direct ELISA and IRMA correlated positively (P<0.01). Further validation of this ELISA was achieved with an indirect immunofluorescense assay. The high degree of significance obtained when the indirect IRMA and ELISA systems were compared, indicated that the relatively small sample number used (12) was statistically satisfactory for the purposes of this investigation. Thus, the IRMA can be replaced by the ELISA, with advantages mainly derived from the cumbersome manipulation of radioactive wastes. The MoAb used as an antigen capture agent in the ELISA proposed here, recognizes a homologous protein in Trypanosoma rangeli, suggesting that individuals infected with this parasite might have crossreactive antibodies. However, the system retains its diagnostic interest, given the facts that the MoAb does not recognize a homologous protein in Leishmania mexicana, Leishmania donovani, or Crithidia fasciculata.

Frecuencia de anticuerpos anti toxoplasma gondii en gatos de la ciudad de Valdivia, Chile

Toxoplasmosis is a parasitic zoonosis. The importance of the cat as related to this parasitosis lies in the fact that it is not only the definite host of the parasite, but responsible for its dissemination through the release of oocysts, which subsequently infect both humans and other animals. The objective of this study was to determine the serological prevalence of Toxoplasma gondii in the feline population of the city of Valdivia (Chile) and to establish the possible epidemiological implications of this prevalence. With these goals in mind the technique of indirect immunofluorescence was implemented to detect anti- T. gondii species--specific IgG. Blood samples from 97 cats (selected using a directed sampling process) from different sectors of Valdivia were collected. The sample included 46 males and 51 females of different ages, Positive and negative control sera obtained from de United States were used to verify the results, observed by UV microscopy. The anti-feline IgG antibody was used as directed by the manufacturer (Sigma). Of the 97 selected cats, 32 were found to be positive with a titre higher than 1:4, a prevalence of 33.0%. The number of infected males as compared to the number of infected females was found to be statistically insignificant, using the chi-square analysis with p less than 0.1. In contrast, a definite correlation between age and seropositivity was found; infection levels were higher in older animals. These results are consistent with those obtained in different studies on this topic that have been performed in different parts of the world. However, they would seem to be in conflict with other, similar studies that have been done recently in Chile; this may be due to the fact that the cats selected for this study were exposed to different climatic conditions than those examined in previous works. In conclusion, it has been determined that in Valdivia exist cats infected by T. gondii, which indicate the presence of the necessary epidemiological conditions for the persistence of this parasitic cycle and the source of infection for humans and other animals.

A putative pyruvate dehydrogenase α subunit gene fromTrypanosoma cruzi

A full-length DNA clone encoding a putative pyruvate dehydrogenase alpha subunit (E1 alpha) gene was isolated from a Trypanosoma cruzi (RA strain) DNA library. Sequencing of this clone revealed it to encode a 378 amino acid protein (M(r) 42774) with high sequence similarity to E1 alpha obtained from different sources. The highest score is obtained with human E1 alpha: 43,3% similarity. Southern blot analysis is consistent with the existence of a single copy of this putative T. cruzi E1 alpha gene per haploid genome in different parasite strains. Expression of this gene was demonstrated by Northern blot analysis and its trans-splicing acceptor site was identified by Polymerase Chain Reaction-mediated amplification of its cDNA.

[Chagas disease in the rural area of Metropolitan Region (Santiago) and V Region (Aconcagua), Chile].

Sr Editor: En 1999 se produjo en Chile un hecho inedito para la Salud Publica del pais: la certificacion de la interrupcion de la transmision vectorial de la enfermedad de Chagas1. Este paso intermedio al proceso de la transmision de la enfermedad de Chagas en Chile a traves del vector Triatoma infestans, fue mal entendido por la comunidad medica y cientifica en general, la cual considero interrumpida la enfermedad de Chagas en Chile, situacion muy lejana a la realidad. Esta situacion se ha visto reflejada en cierta medida en una disminucion de las actividades vectoriales y por otra parte los medicos no consideran el diagnostico diferencial de enfermedad de Chagas en nuestro medio. Para establecer cual es la situacion actual de la infeccion por Trypanosoma cruzi en poblaciones de riesgo de zonas rurales de Santiago y de la V region Cordillera (SEREMI Aconcagua), entre los meses de noviembre y diciembre de 2007, se estudiaron 1.162 pacientes del programa cardiovascular y atencion primaria, 510 procedentes de los consultorios Jose Bauza, Batuco y Juan Pablo II de Lampa (Santiago) y 652 de los consultorios de Putaendo y Panquehue (San Felipe) en los cuales nunca se habia buscado antecedentes de infeccion por el parasito. Para ello no debio efectuarse ninguna intervencion especial en los individuos, siguiendo las normas bioeticas sino las correspondientes a las propias de sus examenes regulares de salud. Los analisis para determinar la infeccion por Trypanosoma cruzi se efectuaron en las muestras de sangre enviadas para los controles del Programa Cardiovascular de cada Comuna y se practicaron con el test rapido aplicado en sangre total y recomendado para estos fines: Trypanosoma detect rapid test INBIOS, Seattle WA, USA, todos aquellos que resultaron positivos y 10% de los negativos fueron confirmados con la reaccion de inmunofluorescencia indirecta para enfermedad de Chagas (RIFI). Todos los analisis fueron realizados en el Laboratorio Campvs Ltda. De las 1.162 muestras de sangre analizadas, 53 (4,6%) resultaron positivas en ambos examenes. La distribucion de los infectados chagasicos fue de 30 en Lampa, 19 para los individuos procedentes de Putaendo y 4 en Panquehue. Es decir en ambas zonas rurales se encontro un numero importante de pacientes infectados. En los 510 pacientes de Lampa se demostro una prevalencia de infectados chagasicos de 5,9% y en los 437 procedentes de Putaendo y 215 de Panquehue una prevalencia de 4,3% y 1,9%, respectivamente. Las edades de los infectados oscilaron entre 17 y 79 anos, con una mayor frecuencia de individuos con edades superiores a los 50 anos. La frecuencia de infectados chagasicos en esas zonas y en los mismos grupos etarios en las decadas de 1980-89 y 1990-99, es decir antes de que se estableciera la interrupcion de la transmision vectorial de la infeccion oscilaba entre 0,5% y 2,5% en la Region Metropolitana y 1,0% y 5,3% en la V Region2, por lo tanto los resultados actuales indican que, a pesar del control vectorial, la infeccion humana persiste y es inclusive superior en aquellos grupos etarios que la habian adquirido antes de la aplicacion de los programas de control vectorial. Del total de positivos, 53 pacientes, ocho casos (15,1%) presentaban edades menores a 18 anos, lo que podria sugerir una posible transmision transplacentaria de la infeccion3. Consideramos importante dar a conocer esta realidad, que pasa frecuentemente desapercibida y requiere sin lugar a dudas la aplicacion de las actividades de vigilancia y control vectorial, por una parte, pero por otra, la preocupacion por la atencion y tratamiento del infectado chagasico, especialmente aquel procedente de zonas rurales y periurbanas de la zona endemica de Chile. Ello exige la capacitacion y entrenamiento de los profesionales de la salud en el tema, tanto para sospechar como para actuar frente a un paciente chagasico. Finalmente, deseamos recalcar que los resultados fueron confirmados e informados a las autoridades de salud pertinente (SEREMI) y a los pacientes involucrados, los cuales en la actualidad estan bajo supervision medica y epidemiologica, como corresponde al Programa de atencion medica del paciente chagasico de acuerdo a las normas ministeriales.

Effect of three coccidiostatics against toxoplasmosis in mice.

SummaryDuocoxin, nicrazine and amprol plus showed no therapeutic effect in 50 mice each of which was infected with 25,000 organisms of a virulentToxoplasma strain.ZusammenfassungDuocoxin, Nicrazin und Amprol-Plus zeigten keine therapeutische Wirkung auf 50 Mäuse, die je mit einer Dosis von 25 000 Organismen eines virulentenToxoplasma-Stammes infiziert wurden.

Comparación de la infección por Trypanosoma cruzi en niños menores de 10 años de la V Región, Chile. 1982-1995

Evaluation of desinsectation programs carried out in the two Health Services from the V Region, was undertaken by comparing serologic prevalences of T. cruzi in age groups exposed to the risk of infection in rural areas during the antivectorial campaign activities. (1982 vs 1995). Thus, were studied 2,193 blood samples from children under 10 years of age, proceeding rfrom six chagasic endemic provinces in which antitriatomic domiciliary insecticide sprayings had been performed. Indirect hemagglutination and ELISA tests were carried out to each of the blood samples. A total de 42 (1.9%) children resulted positive. As in five counties no positive case were detected in the last five years it is possible to assume that vector trasmission of T. cruzi should have been interrupted in them. When comparing prevalences existing in 1982 with the present ones, it is possible to observe a 63.5% of reduction of transmission in the V Region, been this reduction higher in the Aconcagua Health Service with 79.6% and lower in the Vina del Mar - Quillota Health Service with 55.6%. According to these results, comparison of prevalence of T. cruzi infection in children less than 10-year-old in diverse periods, allows evaluate the vectorial control of Triatoma infestans programs