Towards a paradigm shift in the treatment of chronic Chagas disease.

Treatment for Chagas disease with currently available medications is recommended universally only for acute cases (all ages) and for children up to 14 years old. The World Health Organization, however, also recommends specific antiparasite treatment for all chronic-phase Trypanosoma cruzi-infected individuals, even though in current medical practice this remains controversial, and most physicians only prescribe palliative treatment for adult Chagas patients with dilated cardiomyopathy. The present opinion, prepared by members of the NHEPACHA network (Nuevas Herramientas para el Diagnóstico y la Evaluación del Paciente con Enfermedad de Chagas/New Tools for the Diagnosis and Evaluation of Chagas Disease Patients), reviews the paradigm shift based on clinical and immunological evidence and argues in favor of antiparasitic treatment for all chronic patients. We review the tools needed to monitor therapeutic efficacy and the potential criteria for evaluation of treatment efficacy beyond parasitological cure. Etiological treatment should now be mandatory for all adult chronic Chagas disease patients.

Presence of Rhodnius ecuadoriensis in sylvatic habitats in the southern highlands (Loja Province) of Ecuador.

The main vectors of Chagas disease in Ecuador are Triatoma dimidiata and Rhodnius ecuadoriensis. The latter species occupies domestic and peridomestic habitats, as well as sylvatic ecotopes--particularly associated with Phytelephas aequatorialis palm trees--in the western coastal region of Ecuador. In the southern highlands, however, such palm tree habitats are uncommon, and sylvatic populations of R. ecuadoriensis have not previously been reported to date. This study was carried out in five rural communities in Loja Province in southern Ecuador, where manual triatomine searches were conducted in various sylvatic habitats. A total of 81 squirrel nests (Sciurus stramineus) and > 200 bird nests and other habitats were searched. One hundred three R. ecuadoriensis individuals were found in 11 squirrel nests (infestation index = 13.6%, density = 2 bugs per nest searched, crowding = 9.5 bugs per infested nest, colonization index = 72.7% of infested nests with nymphs). No triatomines were found in bird nests or other sylvatic habitats. The presence of sylvatic R. ecuadoriensis in the southern highlands of Ecuador has important implications for the long-term control of Chagas disease in the region because of the possibility of reinfestation of dwellings after insecticide-based control interventions.

High household infestation rates by synanthropic vectors of Chagas disease in southern Ecuador.

Entomological surveys were conducted in five rural communities (138 domiciliary units [DUs]) in the southern Andes of Ecuador. Adobe walls and ceramic tile roofs were predominant construction materials. A 35% house infestation rate with Panstrongylus chinai (Del Ponte, 1929) (0.7%), Panstrongylus rufotuberculatus (Champion, 1899) (0.7%), Rhodnius ecuadoriensis (Lent & León, 1958) (27%), and/or Triatoma carrioni (Larrousse, 1926) (7%) was found. Adults and nymphs of R. ecuadoriensis and T. carrioni were found in intradomiciliary and peridomiciliary areas. Breeding triatomine colonies were present in 85% of infested DUs, and the average insect crowding was 52+/-113 triatomine bugs per infested house. T. cruzi-like organisms were found by microscopic examination in the feces or hindgut but not the salivary glands of 4% of examined R. ecuadoriensis and 12% T. carrioni. Serological tests detected a general anti-T. cruzi antibody seroprevalence of 3.9% (n = 1136). Only 2% of individuals had heard of Chagas disease, and although triatomines were reported as a major nuisance by the population they were not considered vectors of disease. Additional baseline field research is needed for the design and implementation of a Chagas disease control program in the region.

PCR-RFLP detection and species identification of fungal pathogens in patients with febrile neutropenia.

OBJECTIVE: To assess the usefulness of polymerase chain reaction (PCR) assays in the diagnosis of fungal infections in immunocompromised patients. METHODS: A rapid and sensitive PCR-based assay for the detection and identification of fungal pathogens was designed and applicability of this method was investigated in a group of children with cancer and febrile neutropenia (FN). RESULTS: The ITS2 sequences and adjacent regions of 40 fungal pathogens were analyzed and primers for detection of all analyzed fungal species were designed. Amplification product length polymorphism (APLP) and restriction fragment length polymorphism (RFLP) generated genus- or species-specific patterns. The sensitivity of the method was approximately three cells of Candida albicans per 1 mL of blood. The results were available within 8 h after sample collection. The method was tested on 53 blood samples and one lung biopsy sample from 24 children with cancer and febrile neutropenia (FN). The PCR assay detected fungal DNA in 25 clinical samples from ten patients. Blood cultures were positive in only five samples, while another two blood-culture negative patients had positive cultures from throat swabs. The remaining 14 patients were both culture- and PCR-negative. Culture-isolated strains matched completely those obtained by PCR-APLP-RFLP identification. The identity of fungal species was confirmed by direct sequencing of amplified products. CONCLUSION: Our results suggest that PCR-APLP-RFLP assays can be useful in the diagnosis of fungal infections in immunocompromised patients.

[Molecular diagnosis of infections]. / Molekulární diagnostika infekcních stavu.

Molecular diagnostics (detection of nucleic acids by molecular genetics techniques) become more valuable in clinical diagnosis of disease. Apart from the already long-time used genetic techniques for detection of congenital anomalies, current use of molecular techniques includes detection of microbial pathogens. The character of these techniques increases the possibility of achieving diagnosis in cases where classical cultivation is not possible, is not reliable or is not fast enough. As with every new approach, molecular diagnostics have faced encountered reactions from the scientific community. Some scientists tend to overestimate the value of molecular diagnostic techniques, while sceptics, sometimes influenced by a biased or incomplete knowledge of the technology, think it is of little value. In this work, on the basis of literature and our own data from more than 5 years of experience with these methods, we have assessed the pros and cons of the use of molecular diagnostics of infectious diseases in the light of their potential use in clinical practice.

Molecular diagnosis of culture negative infective endocarditis: clinical validation in a group of surgically treated patients.

OBJECTIVE: To assess the clinical validity of polymerase chain reaction (PCR) based molecular methods in the microbiological diagnosis of culture negative infective endocarditis in a group of surgically treated patients. DESIGN: Retrospective case-control study. SETTING: Reference cardiovascular surgical centre. PATIENTS AND SAMPLES: 15 culture negative patients with infective endocarditis classified according to Duke criteria, with 17 heart valve samples; 13 age and sex matched control patients without infective endocarditis, with 13 valve samples. INTERVENTIONS: Medical records were reviewed and clinical, demographic, and microbiological data collected, including results of molecular detection of bacteria and fungi from valve samples. The clinical validity of molecular diagnosis was assessed, along with the sensitivity and speed of the systems. RESULTS: In the study group, 14 patients were PCR positive (93%). Organisms detected were streptococci (3), staphylococci (2), enterobacter (1), Tropheryma whippelii (1), Borrelia burgdorferi (1), Candida albicans (1), and Aspergillus species (2). Three cases were positive on universal bacterial detection but the pathogen could not be identified because of contaminating background. One case was negative. All but two positive cases showed clinical correlations. These two cases had no symptoms of infective endocarditis but there was agreement with the surgical findings. All control cases were PCR negative. Results were available within eight hours, and if sequencing was necessary, within 48 hours. CONCLUSIONS: PCR based molecular detection of pathogens in valve samples from surgically treated culture negative infective endocarditis patients is fast, sensitive, and reliable. The technology, combined with thorough validation and clinical interpretation, may be a promising tool for routine testing of infective endocarditis.

[Evaluation of clinical competence in the postgraduate family medicine course, by means of the Structured Objective Clinical Test]. / Evaluación de la competencia clínica en el posgrado de medicina familiar mediante el Examen Clínico Objetivo Estructurado.

OBJECTIVE: To evaluate the clinical competence of first-year residents on the course specialising in family medicine, by applying a structured objective clinical test (SOCT) for simultaneous assessment of cognitive, affective and psychomotor areas. DESIGN: Observational and descriptive. SETTING: Family medicine clinics in the metropolitan area of Mexico City.Participants. 89 doctors on the specialist course in Family Medicine at the Autonomous University of Mexico (UNAM). MAIN MEASUREMENTS: After prior design of comparison lists, expert validation of content, and design of support material for the evaluation and pilot study, a SOCT with ten themes or sections, eight dynamic and two static, was administered. The cut-off point for competent performance was 60 out of 100, both in each section and in the overall average. The statistical analysis was univariate, using descriptive statistics. RESULTS: The overall average of scores in all sections was under 60. The highest average was 73, in the section for monitoring healthy children. In the five highest-scoring sections, the scattering was lower than in the other five. There were low scores in the family study section. CONCLUSIONS: Academic performance, as evaluated by a SOCT measuring clinical competence, was low. This kind of test enabled clinical competence to be assessed more objectively. Interval evaluation scales need to be tested so as to evaluate better the quality of the performance of clinical activities.

Mouse model for Chagas disease: immunohistochemical distribution of different stages of Trypanosoma cruzi in tissues throughout infection.

Different stages of Trypanosoma cruzi are seen during mammalian infection. Histologic sections of infected hearts have shown amastigotes and, when using immunohistochemistry (IHC), parasite antigens; however, demonstration of trypomastigotes in these tissues has proven elusive. Using a mouse strain that develops chagasic cardiomyopathy (histologically similar to human infection) 70 days after injecting T. cruzi-Brazil strain, we studied the distribution of parasite stages and the extent of inflammation. All organs had varying amounts of mononuclear inflammation by day 10, which peaked between day 20 and day 30, and decreased by day 50. Amastigotes were detected in myocytes, histiocytes, acinar pancreatic cells, astrocytes and ependymal cells by day 10, and the number of amastigotes peaked on day 30. Immunohistochemistry demonstrated trypomastigotes in sinusoids, vessels and interstitial tissues of several organs between day 15 and 50. Abundant parasite antigens (granular staining) were detected in connective tissues throughout the infection. The burden of amastigotes and trypomastigotes during the acute phase seems to correlate with the degree of inflammation and granular staining in the chronic stage.

Use of polymerase chain reaction to diagnose the fifth reported US case of autochthonous transmission of Trypanosoma cruzi, in Tennessee, 1998.

In July 1998, the mother of an 18-month-old boy in rural Tennessee found a triatomine bug in his crib, which she saved because it resembled a bug shown on a television program about insects that prey on mammals. The gut contents of the Triatoma sanguisuga were found, by light microscopy and polymerase chain reaction (PCR), to be infected with Trypanosoma cruzi; PCR products hybridized with T. cruzi-specific oligonucleotide probes. Whole-blood specimens obtained from the child in July and August were negative by buffy-coat examination and hemoculture but positive by PCR and DNA hybridization, suggesting that he had low-level parasitemia. Specimens obtained after treatment with benznidazole were negative. He did not develop anti-T. cruzi antibody; 19 relatives and neighbors also were seronegative. Two of 3 raccoons trapped in the vicinity had positive hemocultures for T. cruzi. The child's case of T. cruzi infection-the fifth reported US autochthonous case-would have been missed without his mother's attentiveness and the availability of sensitive molecular techniques.

Immunological characterization of antigens released by Trypanosoma cruzi-infected cells.

Chagas' disease, caused by Trypanosorna cruzi, is characterized by the appearance of pathological lesions in the heart and other tissues during the chronic phase. The mechanisms responsible for such damage are still unclear. In the vertebrate host, T. cruzi replicates intracellularly before transforming from amastigotes into trypomastigotes. The infected host cell then lyses, releasing the cytoplasmic contents and the parasites that shed membrane glycoproteins soon after release. The sum of all these components we have termed released antigen (Rag). We characterized antigens, released in vitro by fibroblasts infected with T. cruzi, obtained by concentrating conditioned serum-free culture media. The results demonstrate that Rag contains a complex protein mixture including stage-specific T. cruzi antigens (Ssp-1, -2, -4), glucose-regulated protein (Grp) 78h, and peptides recognized by the monoclonal antibody 2B10. These peptides exhibit neuraminidase activity and are expressed by intracellular and 10-20% of released trypomastigotes. Additionally, Rag is recognized by sera from T. cruzi-infected mice and human chagasic patients. Rag also stimulates in vitro production of interferon-gamma by splenocytes from resistant C57B1/6 and susceptible BALB/c infected mice and interleukin-4 by splenocytes from BALB/c infected mice. Altogether these results indicate that Rag is immunologically relevant and could contribute to pathogenesis of T. cruzi infection.

Vitamin A deficiency and low prevalence of anemia in Yaqui Indian children in northwest Mexico.

A study of 296 school-age Yaqui Indian children (6-10 y) was conducted in 26 rural communities. Vitamin A status was determined by retinol and carotenoid serum levels according to a method described previously (IVACG, 1982). Serum retinol and carotenoids in children were analyzed according to community size. Vitamin A intake was assessed in a sub-sample by means of a 24 h recall questionnaire. Serum retinol distribution showed that 6.3% of the children were below 10 microg/100 mL (0.35 micromol/L) and 40% were in the range of 10-20 microg/100 mL (0.35-0.70 micromol/L). Differences (p < 0.02) were found between small and large communities (Median, 95% CI): 19.2 (17.1, 20.9) microg/100 mL and 22.9 (20.3, 24.1) microg/100 mL. Serum carotenoid levels were significantly higher in large than in small and medium communities: 72 (68.2, 77.8) microg/100 mL versus 62.4 (53.3, 68.2) and 62.4 (55.7, 69.6) microg/100 mL, respectively. Food staples were wheat flour tortillas, pinto beans, corn tortillas, few animal products and scarce fresh vegetables. Mean vitamin A consumption was 244+/-29 microg RE (34.9% of the US RDA). Iron status showed that only 4 children were classified as anemic, with two of them having iron deficiency anemia. Iron deficient erythropoiesis was observed in 7.8% of the children and iron depletion only in 4.4%. The Yaqui diet seems to provide adequate amounts of iron but not of vitamin A or its precursors, which renders a vitamin A status of sub-clinical deficiency that could be considered a public health problem.

Incidence of heavy metal contamination in water supplies in northern Mexico.

Contaminants in drinking water present public health risks. The objective of this study was to analyze water samples taken from wells or storage tanks, direct sources for domestic water in Northern Mexico, for the presence of lead (Pb), copper (Cu), cadmium (Cd), arsenic (As), and mercury (Hg). The samples were analyzed by atomic absorption coupled with a hydride generator or a graphite furnace. High levels of Pb (0.05-0.12 ppm) were found in Hermosillo, Guaymas, and Nacozari. Forty-three percent of the samples in Sonora exceeded the action level (0.015 ppm) established by the EPA for Pb. For As, 8.92% exceeded the limit with a range of 0.002-0.305 ppm. Several studies have indicated a possible link between As and fluoride (F) in drinking water. This study showed a positive correlation between F and As (r = 0.53, P = 0.01, and n = 116). One location in Hermosillo had 7.36 ppm of F and 0.117 ppm of As, 3.5 times the recommended F levels in drinking water and 2 times higher than the level permitted for As. Hg contamination was found in 42% of the samples. Based on the results of this study, it appears that As, Hg, and Pb contamination in the drinking water for some areas of the state of Sonora is a major concern.

Short report: screening for Trypanosoma cruzi in the blood supply by the Red Cross blood bank in Quito, Ecuador.

The status of Chagas' disease in Ecuador is not clear. In response to reports suggesting the possibility of transfusion-associated transmission of Chagas' disease in the blood bank in Quito, the Ecuadorian Red Cross in collaboration with the Instituto Nacional de Higiene, Zona Norte and the Tropical Disease Institute of Ohio University implemented a pilot Chagas' disease screening of the donated blood in the Quito blood bank. The results of the screening showed a low incidence of seropositivity among the donors (0.01% in 1994, 0.04% in 1995, and 0.02% in 1996) to the Quito blood bank and a higher seropositivity in samples donated to smaller blood banks (0.4% in 1994, 0.28% in 1995, and 0.13% in 1996) located in areas considered endemic, as well as from at least two areas previously considered nonendemic for Chagas' disease. This report highlights the need for a comprehensive evaluation of the prevalence and distribution of Chagas' disease in Ecuador.

Blood donors in a vector-free zone of Ecuador potentially infected with Trypanosoma cruzi.

Chagas' disease is a serious health problem for the population of South and Central America. Blood transfusion is the second most common way in which this disease is transmitted. Several studies have reported finding Trypanosoma cruzi-infected blood in blood banks in endemic areas. Serum samples were taken from the Red Cross blood bank in Quito, a nonendemic and vector free zone of Ecuador, in December 1992 and May 1993 and analyzed by enzyme-linked immunosorbent assay using crude epimastigote extract from the Brazil strain of T. cruzi. Of 162 samples examined in December 1992, 12.1%, 13.9%, and 74% were seropositive, indeterminate, and seronegative, respectively. Of 173 samples taken in May 1993, 6.2%, 17.9%, 75.9% were seropositive, indeterminate, and seronegative, respectively. Western blot analysis of these sera using sodium dodecyl sulfate-polyacrylamide gel electrophoresis with 7.5% gels separated T. cruzi epimastigote antigen proteins, and revealed a reaction with a 205-kD doublet antigen with most of the seropositive samples. These results indicate the necessity for long-term screening of blood bank donors to reduce the risk of transfusion transmission of the disease even in areas of endemic countries where the vector is not present.

Suplementación con hierro en escolares anémicos

Se estudiaron 60 niños, con un promedio de edad de 6 años a 7 meses en una localidad subtropical del Ecuador. Se obtuvieron valores de hemoglobina (Hb) antes del tratamiento y luego de 30 días de administración de hierro oral 3 mg/kg/día. el cumplimiento de las indicaciones terapéuticas se evaluó siguiendo la metodología vigente en unidades de salud y por profesinales médicos. Se analizó el patrón de rendimiento escolar por calificaciones globales y se obtuvo datos antropométricos (edad, peso, talla). Se encontró una prevalencia de anemia de 33.3 por ciento, más acentuada en hombre y en el grupo de niños nutricionalmnente normales (36.3 por ciento). La prevalencia de desnutrición global, crónica y aguda fue de 42.5 por ciento, 38.3 por ciento y 20.07 por ciento respectivamente. Llama la atención este último dato, el cual es mayor que las cifras de referencia. El cumplimiento del tratamiento fue del 80 por ciento y la corrección de anemia fue únicamente del 18 por ciento. No se encontró cambios en el rendimiento escolar luego de la suplementación con hierro...

[Elaboration, by linear programming, of new products from cereals and legumes]. / Elaboración, por programación lineal, de nuevos productos a partir de cereales y leguminosas.

The differing contents of essential amino acids in cereals and legumes bring about an overall increase in protein quality when these foods are consumed together. This study describes a least cost formulation method for preparing products based on cereals and legumes using linear programming. The mixture was formulated under different constraints; from a nutritional standpoint, a given amino acid pattern, and another one on a technological feasibility constraint, which depends on the type of product to be elaborated. From the formulation based on wheat, chick-pea, sorghum, and soybean flours, three products were developed: bread, tortillas and cookies; from these, bread was selected for further evaluation. The product was chemically evaluated by proximate analysis composition, and amino acids were determined by HPLC. Biological evaluation was performed by the PER and RPV methods, obtaining a PER of 1.69 for the developed bread product, and of 0.68 for the control bread. The RPV for the developed product was 64.31% of lactoalbumin and 23% for the control bread, which represents an increase of 41%. The sensory evaluation results did not indicate significant differences in taste, texture, color or overall acceptability of the developed bread product as compared to the control.