Desafíos de la Bioquímica en la era hipertecnológica / Challenges of Biochemistry in the hyper technological era / Desafios da Bioquímica na era da hipertecnologia

En la era hipertecnológica, ¿somos asistidos por la Tecnología o asistentes de la Tecnología? La máquina inteligente, el robot ideal, ¿puede realizar todas las tareas del laboratorio, con igual o mayor eficacia que el hombre? Creemos que no, ya que la máquina puede tener solamente "pensamiento calculador". En cambio, el hombre posee "pensamiento reflexivo" y juicio ético, que le permiten un discernimiento crítico fundamental. La máquina debe asistirnos, pero es el profesional quien debe decidir lo atinente al pre y post análisis, al control del instrumental en la fase analítica, y a la atención al paciente. La simplificación de los métodos analíticos conlleva a la realización de los análisis deslocalizados, "al lado del paciente". Los progresos técnicos como la miniaturización, la automatización y la informática favorecen el proceso de deslocalización, que puede tener lugar en unidades críticas, guardias, quirófanos o, incluso, fuera del ámbito hospitalario. La rapidez de respuesta en casos críticos puede justificar los análisis deslocalizados, pero existen claras limitaciones: Aseguramiento de la Calidad, Bioseguridad y aspectos bioéticos, respeto de la confidencialidad y capacidad para brindar información adecuada al paciente. En este contexto, es imperativo repensar las Ciencias de la Salud en la importancia de la tecnología pero, también, y sobre todo, en el factor humano y en el rol social del Bioquímico integrante del equipo de salud.

In the hyper technological era, are we assisted by technology or assistants of technology? The intelligent machine, the ideal robot can perform all tasks in the laboratory, with equal or greater efficacy than man? We think not, because the machine can only have "calculating thinking". Instead, man has "reflexive thinking" and ethical judgment, which allow a fundamental critical discernment. The machine must assist professionals, but is the professional who must decide about questions relative to pre and post analysis, control of instruments in the analytical phase, and patient care. Simplification of analytical methods leads to the realization of delocalized analysis, or "point of care". Technical advances such as miniaturization, automation and computing favour delocalized analysis, which can take place in critical units, emergencies, surgeries or even outside the hospital setting. The speed of response in critical cases can justify the delocalized analysis, but there are clear limitations: Quality Assurance, biosafety and bioethical aspects: confidentiality and ability to provide adequate information to the patient. In this context, in the Health Sciences, it is imperative to rethink the importance of technology, but also, and especially, the human factor and the social role of the Biochemist as health team member.

Na era hipertecnológica, somos assistidos pela Tecnologia ou assistentes da Tecnologia? A máquina inteligente, o robô ideal: pode realizar todas as tarefas de laboratório com igual ou maior eficiência do que o homem? Acreditamos que não, visto que a máquina só pode ter "pensamento calculador". Contudo, o homem possui "pensamento reflexivo" e juízo ético que lhe permitem um discernimento crítico fundamental. A máquina deve assistir-nos, porém, é o profissional quem deve tomar as decisões a respeito das pré e pós-análises, do controle do instrumental na fase analítica e do atendimento do paciente. A simplificação dos métodos analíticos conduz à realização das análises deslocadas, "ao lado do paciente". Os avanços técnicos como a miniaturização, a automação e a informática favorecem o processo de deslocação que pode ter lugar em unidades críticas, plantões, salas de cirurgia, ou até mesmo, fora do ambiente hospitalar. A velocidade de resposta em casos críticos pode justificar as análises deslocadas, porém existem claras limitações: segurança na Qualidade, Biossegurança e aspectos bioéticos; a respeito da confidencialidade e capacidade para fornecer ao paciente informação adequada. Nesse contexto, é imperativo repensar as Ciências da Saúde não só na importância da Tecnologia, mas também, e principalmente, no fator humano e no papel social do Bioquímico como integrante da equipe de saúde.

Diagnostic reliability of an immunochromatographic test for Chagas disease screening at a primary health care centre in a rural endemic area

Many patients with Chagas disease live in remote communities that lack both equipment and trained personnel to perform a diagnosis by conventional serology (CS). Thus, reliable tests suitable for use under difficult conditions are required. In this study, we evaluated the ability of personnel with and without laboratory skills to perform immunochromatographic (IC) tests to detect Chagas disease at a primary health care centre (PHCC). We examined whole blood samples from 241 patients and serum samples from 238 patients. Then, we calculated the percentage of overall agreement (POA) between the two groups of operators for the sensitivity (S), specificity (Sp) and positive (PPV) and negative (NPV) predictive values of IC tests compared to CS tests. We also evaluated the level of agreement between ELISAs and indirect haemagglutination (IHA) tests. The readings of the IC test results showed 100% agreement (POA = 1). The IC test on whole blood showed the following values: S = 87.3%; Sp = 98.8%; PPV = 96.9% and NPV = 95.9%. Additionally, the IC test on serum displayed the following results: S = 95.7%; Sp = 100%; PPV = 100% and NPV = 98.2%. Using whole blood, the agreement with ELISA was 96.3% and the agreement with IHA was 94.1%. Using serum, the agreement with ELISA was 97.8% and the agreement with IHA was 96.6%. The IC test performance with serum samples was excellent and demonstrated its usefulness in a PHCC with minimal equipment. If the IC test S value and NPV with whole blood are improved, then this test could also be used in areas lacking laboratories or specialised personnel.

Estudio comparativo de dos métodos para la cuantificación del antígeno carcinoembrionario / Comparative study of two methods for carcinoembryonic antigen quantification / Estudo comparativo de dois métodos para a quantificação do antígeno carcinoembrionário

El antígeno carcinoembrionario (CEA) es una glicoproteína ampliamente utilizada como complemento del diagnóstico, monitoreo de tratamiento y evolución del cáncer colorrectal. El objetivo del presente trabajo fue realizar un análisis comparativo entre dos métodos para la determinación de CEA: electroquimioluminiscencia y quimioluminiscencia, en muestras de suero de 57 pacientes con diagnóstico de cáncer, principalmente colorrectal. Cuando se analizaron los datos totales se obtuvo una elevada correlación (r=0,9135, p<0,00001). Al realizar un corte de los resultados tomando como límite el valor de 4 ng/mL se observó que las mayores discrepancias entre métodos estuvieron en los valores considerados dentro del rango normal (r=0,5716, p<0,0014, n=29). Por el contrario, en concentraciones mayores al límite de corte, la correlación fue elevada (r=0,9453, p<0,00001, n=28). Estos resultados sugieren que, a diferencia de lo descripto por los fabricantes, los valores de CEA obtenidos por ambos métodos son comparables. La menor correlación observada en concentraciones inferiores a 4 ng/mL no sería tan relevante debido a que estos niveles se consideran dentro del rango de normalidad y, por lo tanto, su importancia desde el punto de vista clínico es relativa. Sin embargo, debido a que pueden detectarse con baja frecuencia diferencias individuales (atribuidas probablemente a diferencias en los epitopes detectados por cada método), para los casos con fuerte presunción clínica y un valor de CEA incongruente, se sugiere repetir la determinación por medio de otra metodología.

The carcinoembryonic antigen (CEA) is a glycoprotein widely employed in colorectal cancer, mainly as evolutive marker and as measure of therapy's ef-ficacy. The goal of this work was to perform a comparative study between two analytical methods to measure serum CEA levels: electrochemiluminescence (ECL) and chemilumines-cence (CL) in serum samples of 57 patients with diagnosis of cancer, mainly colorectal. On the whole, an elevated correlation between ECL and CL (r=0.9135; p<0.00001) was obtained. When data was analyzed with a cut-off value of 4 ng/mL, the main discrepancy between methods occurred in the range of normal values (r=0.5716; p<0.0014; n=29). On the contrary, in concentrations higher than the cut-off, the cor-relation was very high (r=0.9453; p<0.00001; n=28). These results suggest that, in spite of the reports of manufacturers, the CEA values obtained by both methods are comparable. The lower correlation observed in values below 4 ng/mL would not be significant because those values are in the normal range and, for that reason, their clinical importance is minor. However, due to the individual differences that could be detected in some patients (probably resulting from the differences in epitopes detected by each method), in cases with strong clinical evidence without concordance with the CEA result, it could be necessary to repeat the determination using another methodology.

O antígeno carcinoembrionário (CEA) é uma glicoproteína amplamente usada como complemento do diagnóstico, monitoração de tratamento e evolução do câncer colorretal. O objetivo deste trabalho foi realizar uma análise comparativa entre dois métodos para a detecção do CEA: eletroquimioluminescência e quimio-luminescência em amostras de soro de 57 pacientes com diagnóstico de câncer, principalmente colorretal. Quando analisados os dados totais, houve uma correlação elevada (r=0,9135, p<0,00001). Quando realizado um corte dos resultados tomando como valor limite 4 ng/mL, observou-se que as maiores diferenças entre ambos os métodos estiveram nos valores considerados dentro da faixa dos valores normais (r=0,5716, p<0,0014, n=29). No entanto, nas concentrações superiores respeito do limite de corte, a correlação foi elevada (r=0,9453, p<0,00001, n=28). Estes resultados sugerem que, comparado com o descrito pelos fabricantes, os valores de CEA obtidos por ambos os métodos são comparáveis. A menor correlação observada nas concentrações inferiores a 4 ng/mL não seria tão relevante devido a que estes níveis consideram-se dentro da faixa de normalidade e, portanto, sua importância, do ponto de vista clínico, é relativa. Contudo, devido a que podem ser detectados com baixa frequência diferenças individuais (atribuídas provavelmente a diferenças nos epitopos detectados por cada método), para os casos com forte suspeita clínica e um valor de CEA incongruente, sugere-se repetir a determinação através de outra metodologia.

Chagas en comunidades originarias del Monte Impenetrable chaqueño, Argentina / Chagas disease in indigenous communities of the Impenetrable Chaqueño Forest

Se demuestra la elevada prevalencia de Chagas en comunidades wichi y criolla de una zona del Gran Chaco argentino, resaltando la necesidad de un control sustentable y continuo

Vaccination with Trypanosoma rangeli modulates the profiles of immunoglobulins and IL-6 at local and systemic levels in the early phase of Trypanosoma cruzi experimental infection

In America, there are two species of Trypanosoma that can infect humans: Trypanosoma cruzi, which is responsible for Chagas disease and Trypanosoma rangeli, which is not pathogenic. We have developed a model of vaccination in mice with T. rangeli epimastigotes that protects against T. cruzi infection. The goal of this work was to study the pattern of specific immunoglobulins in the peritoneum (the site of infection) and in the sera of mice immunized with T. rangeli before and after challenge with T. cruzi. Additionally, we studied the effects triggered by antigen-antibodies binding and the levels of key cytokines involved in the humoral response, such as IL-4, IL-5 and IL-6. The immunization triggered the production of antibodies reactive with T. cruzi in peritoneal fluid (PF) and in serum, mainly IgG1 and, to a lesser magnitude, IgG2. Only immunized mice developed specific IgG3 antibodies in their peritoneal cavities. Antibodies were able to bind to the surface of the parasites and agglutinate them. Among the cytokines studied, IL-6 was elevated in PF during early infection, with higher levels in non-immunized-infected mice. The results indicate that T. rangeli vaccination against T. cruzi infection triggers a high production of specific IgG isotypes in PF and sera before infection and modulates the levels of IL-6 in PF in the early periods of infection.

Chagas disease: serological and electrocardiographic studies in Wichi and Creole communities of Misión Nueva Pompeya, Chaco, Argentina

Chagas disease, which is caused by Trypanosoma cruzi, affects nearly 16 million people in Latin America and causes 75-90 million people to be at risk of infection. The disease is urbanizing and globalizing due to frequent migrations. There are regions of high prevalence of infection, including the north-eastern provinces of Argentina and the entire phytogeographic region known as the Gran Chaco. In the province of Chaco, Argentina, there are places inhabited by native populations such as the Wichi and Toba communities, among others. Many Creole populations resulting from miscegenation with European colonists and immigrants coexist within these communities. It has been widely accepted that in the chronic phase of the disease, between 25-30 percent of individuals develop some form of cardiac disease, with the right bundle-branch block being the most typical condition described so far. The aim of this work was to study the prevalence of Chagas infection and its electrocardiographic profile in the Wichi and Creole populations of Misión Nueva Pompeya, in the area known as Monte Impenetrable in Chaco, to determine the prevalence and the pattern of heart diseases produced by Chagas disease in this region.

American tripanosomiasis: a study on the prevalence of Trypanosoma cruzi and Trypanosoma cruzi-like organisms in wild rodents in San Luis province, Argentina / Tripanosomiasis americana: um estudo sobre a prevalência do Trypanosoma cruzi e Trypanosoma cruzi-like em roedores silvestres da provincia de San Luis, Argentina

INTRODUCTION: Chagas disease is caused by Trypanosoma cruzi. Wild and perianthropic mammals maintain the infection/transmission cycle, both in their natural habitat and in the peridomestic area. The aim of this paper was to present the results from a study on wild rodents in the central and northern regions of San Luis province, Argentina, in order to evaluate the prevalence of this infection. METHODS: Sherman traps were set up in capture areas located between latitudes 32º and 33º S, and longitudes 65º and 66º W. The captured rodents were taxonomically identified and hemoflagellates were isolated. Morphological, biometric and molecular studies and in vitro cultures were performed. Infection of laboratory animals and histological examination of the cardiac muscle and inoculation area were also carried out. Parasites were detected in circulating blood in Calomys musculinus, Graomys griseoflavus, Phyllotis darwini and Akodon molinae. The parasites were identified using biological criteria. Molecular PCR studies were performed on some isolates, which confirmed the characterization of these hemoflagellates as Trypanosoma cruzi. RESULTS AND CONCLUSIONS: Forty-four percent of the 25 isolates were identified as Trypanosoma cruzi, and the remaining 56 percent as Trypanosoma cruzi-like. These findings provide evidence that wild rats infected with Trypanosoma cruzi and Trypanosoma cruzi-like organisms are important in areas of low endemicity.

INTRODUÇÃO: A doença de Chagas é causada pelo Trypanosoma cruzi e os mamíferos periantrópicos e silvestres mantêm o ciclo de infecção/transmissão, tanto no ambiente natural, como no peridomicílio. O objetivo deste trabalho foi mostrar os resultados de um estudo de roedores silvestres do centro e norte da Província de San Luis, Argentina, para avaliar a prevalência da infecção. MÉTODOS: Estabeleceram-se lugares de caça com armadilhas tipo Sherman entre os 32º - 33º de latitude S e 65º - 66º de longitude W. Identificou-se taxonomicamente os roedores, isolou-se os hemoflagelados e fizeram-se estudos morfológicos, biométricos, moleculares, cultivo in vitro, infecção a animais de laboratório, histologia de músculo cardíaco e de zona de inoculação. Observou-se parasitas em sangue circulante: Calomys musculinus, Graomys griseoflavus, Phyllotis darwini e Akodon molinae. A identificação dos parasitas foi feita utilizando critérios biológicos e, em alguns, realizou estudos moleculares por PCR que confirmaram a caracterização desses hemoflagelados como Trypanosoma cruzi. RESULTADOS E CONCLUSÕES: Dos 25 isolados, 44 por cento foram identificados como Trypanosoma cruzi e 56 por cento como Trypanosoma cruzi like. Este achado nos induz a considerar a importância dos ratos do mato infectados com Trypanosoma cruzi y Trypanosoma cruzi like, em área de baixa endemicidade.

La sepsis como estresor: asociación con los niveles séricos de cortisol, proteína C reactiva e interleuquina1beta / Sepsis as a stressor: association with serum levels of cortisol, C reactive protein and interleukin 1beta

El estrés tiene alta prevalencia en el mundo; una de sus causas es la sepsis. Durante la misma, se liberan citoquinas que activan el eje hipotálamo-hipofiso suprarrenal (HHS) elevándose el cortisol y proteínas de fase aguda. El objetivo de este trabajo fue analizar el efecto de la sepsis sobre el eje HHS a través del cortisol, y asociarlo con interleuquina 1 beta (IL1-beta) y proteína C reactiva (PCR). Se dosó cortisol por electroquimioluminscencia, PCR por inmunoturbidimetría e IL1-beta por ELISA en sueros de pacientes sépticos (S, n=40) y no sépticos (NS, n=21). El cortisol fue significativamente mayor al valor de corte (VC), establecido mediante curva ROC, en el 63% de pacientes S, y sólo en el 14% de NS (OR: 10,0; IC: 2,5 - 39,7; p<0,05). La PCR estuvo elevada en el 55% de S, y en el 43% de NS (p>0,05). En algunos pacientes S se evidenciaron niveles muy aumentados de IL1-beta en el día de admisión. La elevación conjunta de PCR y cortisol fue del 40% en S y del 5% en NS, mientras que el aumento de los tres parámetros sólo se vio en S (8%). En conclusión, en el grupo de pacientes estudiado la sepsis actuó como activador del eje HHS, evidenciado por el incremento en el cortisol.

Stress has a high prevalence in the world; one of its causes is sepsis. During this syndrome, cytokines are released, which activate the hypothalamic - pituitary - adrenal (HPA) axis. This raises cortisol levels and acute phase proteins. The goal of this work was to analize the effects of sepsis on the HPA axis, through cortisol measurements, and to associate it with interleukin 1 beta (IL1-beta) and C reactive protein (CRP). Cortisol was measured by electrochemoluminiscence, CRP by immunoturbidimetric method and IL1-beta by ELISA in sera of septic (S, n=40) and non septic (NS, n=21) patients. Cortisol was significantly higher than its cut-off (CO), established by ROC curves, in 63% of S patients, and only in 14% of NS (OR: 10,0; CI: 2.5 - 39.7; p<0.05). CRP was elevated in 55% of S, and in 43% of NS (p>0.05). In some S patients, very high levels of IL1-beta were observed on the day of admission. The joint elevation of CRP and cortisol was of 40% in S and 5% in NS, while rises in all parameters were only seen in S (8%). It can be concluded that in the group of patients studied sepsis acted as an activator of the HPA axis, as evidenced by the elevated cortisol levels.

Vaccination with epimastigotes of different strains of Trypanosoma rangeli protects mice against Trypanosoma cruzi infection

In our laboratory, we have developed a model of vaccination in mice with Trypanosoma rangeli, a non-pathogenic parasite that shares many antigens with Trypanosoma cruzi. The vaccinated mice were protected against infection with virulent T. cruzi. The goal of the present work was to study the protective activity of strains of T. rangeli of different origin, with the aim of analysing whether this protective capacity is a common feature of T. rangeli. BALB/c mice were vaccinated with live or fixed epimastigotes of two T. rangeli strains, Choachi and SC-58. Vaccinated (VM) and control mice (CM) were infected with virulent T. cruzi, Tulahuen strain. The results showed that the levels of parasitemia of VM, vaccinated with the two strains of T. rangeli were significantly lower than those developed in CM. The survival rate of VM was higher than that CM. Histological studies revealed many amastigote nests and severe inflammatory infiltrates in the heart and skeletal muscles of CM, whereas in the VM only moderate lymphomonocytic infiltrates were detected. Altogether, the results of the present work as well as previous studies show that the antigens involved in the protection induced by T. rangeli are expressed in different strains of this parasite. These findings could prove useful in vaccine preparation.

Chagas' disease: study of congenital transmission in cases of acute maternal infection

Se descrevem 3 gestantes com a doença de Chagas aguda. Duas gestantes infetadas no 3º trimestre de gestação não tiveram crianças infetadas. O 3º filho, doquella madre foi infetada no 1º trimestre, nasceu com doença de Chagas congénita. Estes resultados inducem a investigação sobre os fatores de riscos da transmição e sobre as desições médicas na conducção dos casos de gestantes com a doença de Chagas aguda.

Detección de anticuerpos para Chagas y Toxoplasmosis en trasudado mucoso oral / Detection of antibodies to Trypanosoma cruzi andToxoplasma gondii in oral mucosal transudate

El Trasudado Mucoso Oral (TMO) es un fluido biológico que puede obtenerse mediante una almohadilla absorbente colocada entre la encía y la mejilla inferior y que contiene 20% de IgG, 40% de IgA y 10% de IgM en relación al suero. El objetivo del trabajo fue analizar la confiabilidad del TMO como muestra biológica para la detección de anticuerpos en Chagas y Toxoplasmosis. Sueros de pacientes ambulatorios, embarazadas y voluntarios sanos fueron estudiados para Chagas y Toxoplasmosis empleando Inmunofluorescencia, ELISA y Hemaglutinación. Las muestras de TMO fueron estudiadas por ELISA y los resultados comparados con los métodos de referencia para determinar sensibilidad (S), especificidad (E), valor predictivo positivo (VPP) y valor predictivo negativo (VPN). En Chagas, la sensibilidad osciló entre 91% y 100% con tres diferentes equipos ensayados, mientras la especificidad varió entre 90 y 100%, el VPP entre 95% y 96% y el VPN entre 97% y 99%. En Toxoplasmosis no se detectaron resultados falsos positivos (S 95%, E 100%, VPP 100% y VPN 98%). Estos resultados sugieren que el TMO puede ser un fluido biológico alternativo adecuado para estudios inmunoepidemiológicos y también servir como screening en el diagnóstico y prevención de la transmisión vertical de enfermedades infecciosas.

Oral mucosal transudate (OMT) is a biological fluid that can be obtained by an absorbent pad placed between lower cheek and gum, and contains20% IgG, 40% IgA and 10% IgM comparing with serum. The aim of this work was to analyse the performance of OMT as biological material to detect antibodies in Chagas' disease and Toxoplasmosis. Sera of ambulatory patients, pregnant women and healthyvolunteers were tested for Chagas and Toxoplasmosis employingImmunofluorescence, ELISA andHemagglutination.OMT of the same patients were assayed by ELISA, and the results compared to determinate sensibility, specificity and predictive value. In Chagas serology, three different commercial kits were assayed. The sensibility ranged from 91 to 100%,specificitybetween 90 and 100%. The predictive values oscillate between 95% and 99%. The studies in Toxoplasmosis did not shown false positive results. The sensibility was 95%, specificity 100% and the predictive values between 98% and100%. Sera from neonates born from Toxoplasmosis infected mothers were also studied, and the results were in agreement with reference tests. These results suggest that OMT could be a suitable alternative biological fluid in immunoepidemio-logical surveys, and also as screening test in the diagnosis and prevention of materno-fetal transmission of infectious diseases.

Vaccination with live epimastigotes of Trypanosoma rangeli protects mice against Trypanosoma cruzi lethal infection

Trypanosoma cruzi and Trypanosoma rangeli share geographical areas, vectors and hosts. Although both parasites are antigenically similar, T. rangeli is not pathogenic for humans. In consequence, T. rangeli have been experimentally employed as immunogen to protect against T. cruzi infection. The aim of this work was to analyze the evolution of T. cruzi infection in mice previously vaccinated with live epimastigotes of T. rangeli obtained from cultures, and to measure TNF-alfa, IL12 and IL-18 productions. The evolution of the T. cruzi (Tulahuen strain) infection was evaluated by parasitemia levels, survival of Balb/c mice, tissue lesions and/or the presence of parasites. Cytokine levels were measured by an immuno-enzyme assay technique. The mice that were not vaccinated, died in the acute stage of infection with high parasitemias, nests of amastigotes and inflamatory foci in heart and skeletal muscle tissues, associated with high TNF-alfa levels. On the order hand, mice that were previously infected with T. rangeli, survived the acute stage of T. cruzi infection with low TNF-alfa level and high IL-18 level. In conclusion, this work describes a new model of immunization with T. rangeli associated with resistance to T. cruzi infection with modulation of proinflammatory TNF-alfa and increased IL-18 serum level.

Interleuquina-6, receptor soluble de interleuquina-2 y proteína C reactiva como marcadores de sepsis neonatal / Interleukin-6, soluble interleukin-2 receptor and Creactive proteins markers of neonatal sepsis

La sepsis bacteriana neonatal constituye una de las principales causas de muerte, o de secuelas muchos veces irreversibles, después de las primeras 72 horas de vida. Debido a que el diagnóstico precoz es crucial para mejorar el pronóstico de los neonatos infectados, y que los métodos actuales no presentan la necesaria sensibilidad y rapidez, el objetivo del presente trabajo fue analizar la posible utilidad de la determinación de Interleuquina 6 (IL6) y del Receptor soluble de Interleuquina 2 (sIL-2R) como marcadores diagnósticos y pronósticos de sepsis neonatal y, asimismo, efectuar la comparación con un marcador clásico de infección bacteriana como es la Proteína C Reactiva (PCR). Se estudiaron cuatro grupos de recién nacidos, comprendidos en la siguiente clasificación clínico-epidemiológica: sepsis confirmada (SC), n=22, sepsis muy probable (SMP), n=18, sepsis probable (SP), n=11 y sin infección bacteriana (SIB), n=13. Mediante inmunoensayo enzimático se realizó el dosaje de los niveles séricos de IL6 y sIL-2R y por inmunodifusión la determinación semicuantitativa de PCR. Se calculó el límite de corte para cada determinación con los valores obtenidos en los neonatos no infectados y para el cálculo de sensibilidad, especificidad y valor predictivo se tomó como referencia el resultado del análisis microbiológico. Los siguientes resultados fueron obtenidos en las muestras tomadas en el momento de la admisión: sensibilidad: IL6 90 por ciento, sIL-2R 60 por ciento, PCR 41 por ciento; especificidad: IL6 50 por ciento, sIL-2R 100 por ciento, PCR 100 por ciento; valor predictivo negativo: IL6 75 por ciento, sIL-2R 58 por ciento, PCR 59 por ciento. Para evaluar el comportamiento como marcadores evolutivos, se tomó una muestra a las 48 horas, cuando los pacientes de los grupos SC, SMP y SP estaban bajo tratamiento antibiótico. Los resultados mostraron que IL6 descendió en los neonatos con evolución favorable e incrementó marcadamente en los que tuvieron desenlace fatal; sIL-2R mostró un comportamiento inverso y PCR no sufrió variaciones detectables en el período estudiado. Los resultados obtenidos en el presente trabajo sugieren que IL6 puede ser un buen marcador de sepsis neonatal, fundamentalmente por su elevada sensibilidad, y que la especificidad puede incrementarse mediante el estudio de un perfil de marcadores. Asimismo, la cinética obtenida mediante el estudio seriado de IL6 y sIL-2R le otorga valor en el pronóstico y en la evaluación...

Enfermedad de Chagas experimental: I estudio de diferentes condiciones de inmunización sobre el curso de la infección / Experimental Chagas disease: I study of different immunization conditions over the infection course

In previous works it has been demonstrated that Balb/c albino mice immunized with Trypanosoma rangeli developed cellular and humoral immune response to Tripanosoma cruzi. Moreover, the immunized animals were protected against lethal infection by virulent T. cruzi trypomastigotes. In fact, immunized mice had significantly lower parasitemias and longer survival than controls. To go further in this experimental model, the aim of the present work was to analyze the effect of the number of antigenic stimuli and the conservation of the antigen on the effectiveness of protective effect. For that purpose, three different immunization schedules injecting T. rangeli epimastigotes fixed with glutaraldehide and emulsified with Saponin (SAP) as adjuvant were assayed. Different lots of mice which received only phosphate buffer saline or SAP were used as controls. In another set of experiments the conservation of the antigen during 90 days at 4ºC was studied. In all the experiments mice were infected with 100 trypomastigotes of T. cruzi, Tulahuén strain. The parasitemias were analyzed on 13th, 16th and 21th post infection days, and the survival until the 60th day. The results revealed that one dose of antigen was inadequate to give an effective protection. On the other hand, mice immunized with 2 and 3 ddose showed a significant decrease of parasitemia with regard to controls (p<0.001- p<0.0001) and the survival were markedly increased. Likewise, the antigen kept during 90th days at 4ºC showed similar protective efficacy than fresh antigen. Both of these experimental groups showed significant differences with respect to control animals in parasitemia (p<0.05 - p>0.01) and survival (p>0.01). In conclusion, the results of this work showed that in the experimental conditions assayed, the immunization with T. rangeli trigger and adequate immune response when mice received at least two antigenic stimuli. Likewise, it is interesting to point out the stability of the antigenic preparation during at least 90th days

Enfermedad de Chagas crónica: efectos del tratamiento en los niveles de anticuerpos hacia antígenos crudos y semipurificados del trypanosoma cruzi / Chagas chronic disease: effects of treatment in antibodies levels to crude and partially purified trypanosoma cruzi antigens

The specific therapy in chagas' disease is useful in acute and neonatal infection and in children under three years old. The results of antiparasitic treatment during chronic infection are still controversial. It will be interesting to analyze the serological behavior in patients treated during chronic infection, to advance in the search of evolutive markers and markers of therapeutic efficacy. In the present work we have measured the antibody response by conventional serology and the response to partially purified T. cruzi antigens in chagasic patients who received nifurtimox or benznidazol 2 to 20 years before. The results showed that, by indirect immunofluorescence in 20 percent of treated patients the antibody levels were below the established cut off (1:32). By indirect hemagglutination 55 percent of treated patients showed this serological behavior. In this group a high number of discordant results was observed. By immunoenzimatic assay it was possible to detect a significative decrease of serologic reactivity to a partially purified acidic antigen (F IV) and to exoantigen of T. cruzi. It will be interesting to perform longitudinal surveys employing these antigens, to go further in the knowledge of possible immunological evolutive markers in Chagasïdisease

Enfermedad de Chagas: reactividad serologica hacia antigenos crudos y semipurificados del T. Cruzi en distintos grupos clinicos de pacientes / Chagas disease: serologic response to crude and semipurified T. cruzi antigens in different clinical groups of patients

La respuesta inmune de los infectados chagásicos difere cualitativa y cuantivamente segün el estadio de la enfermedad. La medición con fines clínicos de estas diferencias depende de la posibilidad de contar con preparaciones antigénicas adecuadas. Con el objeto de avanzar en este sentido, se estudió el comportamiento de distintos antígenos (Ag) del T. cruzi en pacientes chagásicos agudos y crónicos perttencientes a los grupos O (GO) y 1 (G1). Se midió el nivel de anticuerpos (Ac) totales por sorología convencional (hemaglutinación e inmunofluorescencia (IF)), y por enzimoinmunoanálisis, los títulos de Ac contra el extracto crudo de formas epimastigotes del parásito (F 105) y contra una fracción semipurificada por cromatoenfocado analítico (F IV). En los pacientes agudos se observó mediante la reccion de ELISA con F 105, un 60 por ciento de resultados positivos, mientras que la serología convencional reveló un alto porcentaje de sueros con ambas reacciones negativas (61 por ciento) y 35 por ciento de discordantes. Luego del tratamiento antiparasitario en los pacientes agudos, hubo una disminución en los títulos de Ac hacia cualquiera de los Ag estudiados. En los pacientes crónicos los resultados mostraron una tendencia de los individuos de G1 a presentar mayores títulos de Ac tanto por IF frente al parásito total, como por enzimoinmunoálisis (ELISA) frente a la facción F IV, con respecto a los del GO. Esta tendencia fue más evidente cuando se combinaron los resultados de ambas técnicas. En efecto, el 22 por ciento de los sueros del G1 presentaron títulos por IF = 1: 128, e índices por ELISA = 3,5, mientras que sólo el 2 por ciento de los sueros del GO presentó esta característica. Niveles más bajos no permitieron discriminar poblaciones. La conjugación de los métodos tradicionales con la utilización de técnicas que empleen antígenos purificados o sintéticos puede resultar en una mejora en la calidad y en la utilidad del inmunodiagnóstico.

Isoxazolynaphthoquinone effects on the growth of Trypanosoma cruzi

Frente a la necessidad de hallar neuveos fármacos contra el agente etiológico de la enfermedad de Chagas y en base a las propiedades biológicas y terapéuticas de naftoquinonas e isoxazoles, se resolvió estudiar el efecto de tres isoxazolilnaftoquinonas (Fig. 1) sobre el desarrollo del Trypanosoma cruzi in-vitro e in-vivo. Para evaluar la acción de las drogas sobre los epimastigotes se realizaron curvas de crecimiento con distintas concentraciones de 2-hidroxi-N-(3,4-dimetil-5-isoxazolil)-1,4-naftoquinina-4-imina (I), N-(3,4-dimetil-5-isoxazolil)-4-amino-1,2 naftoquinona (II), 2-acetil-N-(3,4-dimetil-5-isoxazolil)-1,4-naftoquinona-imina (III) y Nifurtimox como droga de referencia. Los estudios sobre la forma de tripomastigote se realizaron sobre ratones BALB/c de dos meses de edad, evaluado parasistemia en el día 13 post-infección. Los resultados obtenidos con epimastigotes mostraron que todas las drogas indujeron alteraciones consistentes es disminución de movilidad, vacuolización, pérdida de viabilidad y/o lisis de los parásitos (Fig. 2 y 3). En los tratamientos sobre tripomastigotes los resultados más concluyentes se presentaron con I que produjo una reducción de la parasitemia respecto a los controles no tratados (Fig 4)