Diabetes mellitus por mutación en el gen de glucokinasa: caso clínico / Diabetes mellitus caused by a mutation of glucokinase gene: report of an affected family

Maturity-Onset Diabetes of the Young (MODY) refers to a heterogeneous group of monogenic diabetes. Unlike other types of MODY characterized by genetic defects in transcription factors, MODY 2 is triggered by metabolic alterations caused by mutations of glucokinase (GCK), the first enzyme of the glycolytic pathway. We report a three-generation Chilean family with multiple cases affected with this disease. The index case is a patient who presented severe neonatal hyperglycemia (831 mg/dl, without ketosis) requiring continuous infusion of insulin, which was suspended after 48 hours with normalization of blood glucose. Subsequently, continuous glucose monitoring at 4 months of age revealed 47% of tissue glucose levels above 140 mg/dl, with fasting glucose levels between 120 and 166 mg/dl. The genetic analysis revealed a previously reported mutation in heterozygous state of the GCK gene (c.148C>T; p.His50Tyr). This mutation was also identified in more than one affected relative in the last two generations, with a transmission pattern suggestive of dominant inheritance. GCK gene sequencing led to a correct molecular diagnosis of MODY 2 while bioinformatic analysis indicated the possible molecular causes of the enzyme dysfunction. The knowledge of the molecular diagnosis allowed an adequate medical treatment for this disease.

[Diabetes mellitus caused by a mutation of glucokinase gene. Report of an affected family]. / Diabetes mellitus por mutación en el gen de glucokinasa. Caso clínico.

Maturity-Onset Diabetes of the Young (MODY) refers to a heterogeneous group of monogenic diabetes. Unlike other types of MODY characterized by genetic defects in transcription factors, MODY 2 is triggered by metabolic alterations caused by mutations of glucokinase (GCK), the first enzyme of the glycolytic pathway. We report a three-generation Chilean family with multiple cases affected with this disease. The index case is a patient who presented severe neonatal hyperglycemia (831 mg/dl, without ketosis) requiring continuous infusion of insulin, which was suspended after 48 hours with normalization of blood glucose. Subsequently, continuous glucose monitoring at 4 months of age revealed 47% of tissue glucose levels above 140 mg/dl, with fasting glucose levels between 120 and 166 mg/dl. The genetic analysis revealed a previously reported mutation in heterozygous state of the GCK gene (c.148C>T; p.His50Tyr). This mutation was also identified in more than one affected relative in the last two generations, with a transmission pattern suggestive of dominant inheritance. GCK gene sequencing led to a correct molecular diagnosis of MODY 2 while bioinformatic analysis indicated the possible molecular causes of the enzyme dysfunction. The knowledge of the molecular diagnosis allowed an adequate medical treatment for this disease.

Diagnóstico de la infección por Treponema pallidum en pacientes con sífilis temprana y neurosífilis mediante reacción de la polimerasa en cadena / Laboratory diagnosis of Treponema pallidum infection in patients with early syphilis and neurosyphilis through a PCR-based test

Syphilis is a sexually transmitted disease caused by Treponema pallidum. The diagnosis is based mainly in clinical presentation and non-specific assays. PCR-based diagnosis has been suggested as an attractive alternative method. The aim of this study was the validation of a PCR-based test for the diagnosis of early syphilis (ES) and neurosyphilis (NS). Clinical samples of mucocutaneous lesions and cerebrospinal fluid (CSF) specimens from patients previously diagnosed for ES and NS respectively using an enlarged gold standard, were tested by PCR. The reaction was done using primers targeting the tpN47gene. Twenty out of 21 mucocutaneous samples from patients diagnosed with ES were positive by PCR, with a clinical sensitivity of 95 percent. Four out of 8 CSF samples from patients previously diagnosed with NS were positive by PCR, with a clinical sensitivity of 50 percent. The clinical specificity for both ES and NS was 100 percent. The PCR sensitivity and specificity for mucocutaneous samples allowed us to implement this assay in our laboratory for routine diagnosis. Although the sensitivity of the PCR in CSF was low, it may be useful to support clinical diagnosis.

La sífilis es una enfermedad de transmisión sexual producida por Treponema pallidum, cuyo diagnóstico se realiza presuntivamente basándose en aspectos clínicos y análisis de especificidad limitada. La reacción de la polimerasa en cadena (RPC) ha sido planteada como una alternativa diagnóstica de mayor sensibilidad y especificidad. El objetivo de este trabajo fue validar una RPC para el diagnóstico de sífilis temprana (ST) y neurosífilis (NS). Se utilizaron muestras de lesiones muco-cutáneas y de LCR de pacientes con sospecha de cursar ST y NS respectivamente, previamente diagnosticados, utilizando un estándar de oro ampliado. La RPC fue realizada con partidores dirigidos al gen tpN47. De las 21 muestras de pacientes con ST, la RPC resultó positiva en 20, lo que resulta en una sensibilidad clínica de 95 por ciento. De las 8 muestras de pacientes con NS, la RPC resultó positiva en 4, obteniéndose una sensibilidad clínica de 50 por ciento. La especificidad clínica para ST y NS fue de 100 por ciento. La excelente sensibilidad y especificidad de la RPC para muestras muco-cutáneas permitió la exitosa implementación de este análisis en nuestro laboratorio para el diagnóstico de rutina. Si bien la sensibilidad de la RPC en LCR es baja, es muy útil para apoyar el diagnóstico clínico.

Conceptos básicos sobre el estudio de paternidad / Basic concepts about paternity testing

Nowadays, the analysis of genetic markers is a very important and validated tool for the identification of individuals, and for paternity testing. To do so, highly variable regions of the human genome are analyzed, making it possible to obtain the genetic profle of an individual, and to distinguish between different individuals. The methodology used is basically the same all over the world, consisting in the analysis of 13 to 15 markers. To assign biological paternity the child must have inherited the characteristics from the alleged father in each of the genetic markers analyzed. This analysis achieves a certainty higher than with any other test, which is expressed as the probability of paternity. This probability has to be at least 99.9 percent, but greater probabilities are usually obtained, especially if the mother is included in the analysis. If the characteristics of two or more genetic markers from the alleged father are absent in the child, biological paternity is excluded.

El análisis de marcadores genéticos se ha convertido en una herramienta muy importante y ampliamente reconocida para la identificación de individuos y para el estudio de paternidad. Para esto se estudian distintas regiones del genoma humano que son altamente variables en la población y que permiten obtener el perfil genético y distinguir entre distintos individuos. La metodología que se utiliza es básicamente la misma en todo el mundo y consiste en el análisis de entre 13 a 15 marcadores. La paternidad biológica se asigna cuando el hijo/a presenta las características que debe heredar del presunto padre en cada uno de los marcadores genéticos estudiados. A través de este análisis es posible asignar paternidad con un grado de certeza más alto que con cualquier otro sistema, el que se expresa como probabilidad de paternidad. Esta probabilidad debe alcanzar al menos 99,9 por ciento. Sin embargo, es posible obtener probabilidades mucho más altas, sobre todo si se incluye a la madre en el estudio. Si las características genéticas del supuesto padre están ausentes en el hijo/a en al menos dos marcadores, se excluye la paternidad biológica.

[Genetic tests: definition, methods, validity and clinical utility]. / Tests genéticos: Definición, métodos, validación y utilidad clínica.

The knowledge of the human genome has led to an explosion of available genetic tests for clinical use. The methodologies used in these tests vary widely, allowing the study from chromosomes to the analysis of a single nucleotide. Prior to its use in the clinical setting, these tests should have an evaluation that includes analytical and clinical validation and determination of the clinical utility, as any other tests, including requirements for quality assurance. Recently, the CDC (Centers for Disease Control and Prevention, USA) published a guideline for Good Laboratory Practices for Molecular Genetic Testing for Heritable Diseases and Conditions, covering the pre-analytical, analytical and post-analytical phases of the tests. The document covers the importance of proper selection of tests, the availability of information on the performance of the techniques used, the quality control practices, the training of personnel involved and the report of results, to allow the adequate interpretation, including sensitivity and specificity. Considering that recent advances in genetics have changed and will continue to affect clinical practice, genetic tests must meet quality and safety requirements to enable optimal use of them.

Tests genéticos: Definición, métodos, validación y utilidad clínica / Genetic tests: Definition, methods, validity and clinical utility

The knowledge of the human genome has led to an explosion of available genetic tests for clinical use. The methodologies used in these tests vary widely, allowing the study from chromosomes to the analysis of a single nucleotide. Prior to its use in the clinical setting, these tests should have an evaluation that includes analytical and clinical validation and determination of the clinical utility, as any other tests, including requirements for quality assurance. Recently, the CDC (Centers for Disease Control and Prevention, USA) published a guideline for Good Laboratory Practices for Molecular Genetic Testing for Heritable Diseases and Conditions, covering the pre-analytical, analytical and post-analytical phases of the tests. The document covers the importance of proper selection of tests, the availability of information on the performance of the techniques used, the quality control practices, the training of personnel involved and the report of results, to allow the adequate interpretation, including sensitivity and specificity. Considering that recent advances in genetics have changed and will continue to affect clinical practice, genetic tests must meet quality and safety requirements to enable optimal use of them.

El conocimiento del genoma humano ha dado lugar a un aumento explosivo de los test genéticos disponibles para uso clínico. Las metodologías utilizadas en este tipo de tests son muy variadas, permitiendo desde el estudio de los cromosomas hasta el análisis de una base nucleotídica. Previo a su utilización en el ámbito clínico, estos tests deben tener una evaluación que incluya su validación analítica y clínica y determinación de la utilidad clínica, además de cumplir, como cualquier otro examen, con requisitos para el aseguramiento de la calidad. Recientemente, el CDC (Centersfor Disease Control and Prevention, EE. UU) hapublicado recomendaciones para las buenas prácticas de laboratorio de tests moleculares que se utilizan para el diagnóstico de enfermedades genéticas, que abarcan la fase pre- analítica, analítica y post-analítica. Dentro de éstas destacan: la importancia de la selección adecuada de los tests, la disponibilidad de la información sobre el desempeño de las técnicas utilizadas, las prácticas de control de calidad, la capacitación del personal involucrado y la elaboración de un informe de resultados que permita al clínico interpretarlos adecuadamente, incluyendo sensibilidad y especificidad. Tomando en cuenta que los recientes avances en genética han modificado y seguirán modificando la práctica clínica, los test genéticos deben cumplir con las exigencias de calidady seguridad que permitan su uso óptimo.

[Laboratory alert value reporting by the clinical laboratory at an academic medical network]. / Aviso de valores de alerta por parte del laboratorio clínico en una red de salud universitaria.

BACKGROUND: An alert value is a result suggesting that the patient is at imminent danger unless appropriate remedial actions begin promptly. Report of alert values (AV) by the clinical laboratories has taken special relevance in recent years due to its contribution to patient's care. AIM: To report results of AV informed during 2007 within the Health Network of the Pontificia Universidad Católica de Chile. MATERIAL AND METHODS: Analysis of AV recorded in a centralized database of the laboratories of the health network, between January and December, 2007. RESULTS: Total number of AV was 5.366, which represented 0.3% of total examinations and corresponded mainly to the clinical chemistry area. Potassium levels generated the higher number of AV detected, followed by positive blood cultures. Eighty two percent of AV corresponded to hospitalized patients. The greater number of AV was reported to intermediate and intensive care services. Thirty two percent of AV was informed to the physician or professional in charge of the patient within 5 minutes of obtaining the results and 79% within 30 minutes. CONCLUSIONS: To obtain a real impact on patient management, it is fundamental to shorten the lapse between the obtainment of tests results and the warning, supported on appropriate computerized systems, and to spread the procedure to all personnel involved in patient's care.

Análisis retrospectivo del rendimiento de Amplicor-PCR® para la detección de Mycobacterium tuberculosis en muestras respiratorias y no respiratorias con baciloscopia negativa: A retrospective analysis / Assessment of the Amplicor PCR® for the detection of Mycobacterium tuberculosis in smear negative respiratory and non respiratory specimens

Background: Commercial polymerase chain reaction (PCR) kits are widely accepted for analysis of smear positive respiratory specimens, but the sensitivity is variable for smear negative ones. Objective: To assess the PCR method usefulness in smear negative respiratory and non respiratory specimens. Methods: We compared the PCR results (AMPLICOR MTB test™, Roche) of 235 specimens subjected to culture in Loewenstein-Jensen agar (as the gold standard). Results: 181 (76 percent) were respiratory and 54 (24 percent) extra-respiratory specimens. The sensitivity was 88 percent) and 50 percent>, respectively, specificity and PPV was 100 percent> in both cases. NPV was 99.4 percent> in respiratory specimens and 96.1 percent in non-respiratory specimens. Conclusions: The good performance of this PCR in smear negative respiratory specimens allows the clinician to take decisions based on the result of this exam. In extra-respiratory specimens the contribution is important only when the PCR result is positive.

[Universal or broad-range polymerase chain reaction (PCR): a contribution to the detection and identification of bacteria and fungi in clinical practice]. / "PCR universal o de amplio espectro": un aporte a la detección e identificación de bacterias y hongos en la práctica clínica.

The use of techniques for the detection of nucleic acids such as the polymerase chain reaction (PCR) has had a major impact on microbiological analysis, playing an important role in the clinical laboratory. Most of the techniques currently used are designed for specific detection of a particular microorganism. However, infectious agents can also be identified even if genus or species are unknown, using universal primers to amplify bacterial or fungal DNA and then identify the species by sequence (universal or wide spectrum PCR). This methodology is applied in cultures that are difficult to identify using phenotypic techniques, and more recently it is also being used directly in clinical samples, where the detection and identification of the infectious agent by traditional techniques is difficult or not possible.

Aviso de valores de alerta por parte del laboratorio clínico en una red de salud universitaria / Laboratory alert value reporting by the clinical laboratory at an academic medical network

Background: An alert value is a result suggesting that the patient is at imminent danger unless appropriate remedial actions begin promptly. Report of alert values (AV) by the clinical laboratories has taken special relevance in recent years due to its contribution to patient's care. Aim: To report results of AV informed during 2007 within the Health Network of the Pontificia Universidad Católica de Chile. Material and methods: Analysis of AV recorded in a centralized database of the laboratories of the health network, between January and December, 2007. Results: Total number of AV was 5.366, which represented 0.3 percent of total examinations and corresponded mainly to the clinical chemistry area. Potassium levels generated the higher number of AV detected, followed by positive blood cultures. Eighty two percent of AV corresponded to hospitalized patients. The greater number of AV was reported to intermediate and intensive care services. Thirty two percent of AV was informed to the physician or professional in charge of the patient within 5 minutes of obtaining the results and 79 percent within 30 minutes. Conclusions: To obtain a real impact on patient management, it is fundamental to shorten the ¡apse between the obtainment of tests results and the warning, supported on appropriate computerized systems, and to spread the procedure to all personnel involved in patient's care (RevMéd Chile 2009; 137: 1137-44).

PCR universal o de amplio espectro: Un aporte a la detección e identificación de bacterias y hongos en la práctica clínica / Universal or broad-range polymerase chain reaction (PCR): A contribution to the detection and identification of bacteria and fungi in clinical practice

The use of techniques for the detection of nucleic acids such as the polymerase chain reaction (PCR) has had a major impact on microbiological analysis, playing an important role in the clinical laboratory. Most of the techniques currently used are designed for specific detection of a particular microorganism. However, infectious agents can also be identified even if genus or species are unknown, using universal primers to amplify bacterial or fungal DNA and then identify the species by sequency (universal or wide spectrum PCR). This methodology is applied in cultures that are difficult to identify usingphenotypic techniques, and more recently it is also being used directly in clinical samples, where the detection and identification of the infectious agent by traditional techniques is difficult or not possible.

El uso de técnicas para la detección de ácidos nucleicos como la reacción en cadena de la polimerasa (PCR) ha tenido un gran impacto en el diagnóstico microbiológico, ocupando un lugar importante en el laboratorio clínico. La mayoría de ¡as técnicas en uso han sido diseñadas para la detección específica de un microorganismo. Sin embargo, también es posible identificar el agente etiológico aunque se desconozca la especie o el género, utilizando partidores universales para amplificar el ADN de bacterias y hongos, y luego secuenciar para identificar la especie (PCR universal o de amplio espectro). Esta metodología se aplica en cultivos difíciles de clasificar por técnicas fenotípicas, pero también se ha comenzado a utilizar directamente en muestras clínicas, en las que la detección e identificación del agente infeccioso por técnicas tradicionales resulta difícil o no es posible.

[The contribution of critical laboratory values to patients' safety and care]. / Valores de alerta en el laboratorio clínico y su aporte al cuidado y seguridad del paciente.

Critical values are those laboratory values that are so abnormal that may threaten the life of a patient unless immediate corrective or therapeutic actions are undertaken. Among laboratory procedures, this definition has been incorporated to standards that watch over patients' safety. Health institutions should incorporate this practice and monitor its effectiveness.

Presencia de macroprolactina en sueros con normo e hiperprolactinemia: comparación de dos métodos de detección / Comparison of two techniques to detect the presence of macroprolactin in serum

Background: Macroprolactin is biologically inactive but may be detected by immnoassays. This leads to errors in diagnosis and inadequate treatment of patients with hyperprolactinemia. Aim:To assess two techniques to detect the presence of macroprolactin. Material and Methods: Prolactin was measured by immunoassay in 57 serum samples (from 4 males and 53 females aged33 +/- 13 years), before and after precipitation with polyethyleneglycol (PEG) and separation by ultrafiltration. A significant level of macroprolactin was considered to be present when prolactin detected in the supernatant after PEG precipitation or in the ultrafiltrate was less than 40 percent of the initial concentration of prolactin. Results: Prolactin levels fluctuated from 5 to 411 ng/ml. The percentages of recuperation were independent of the initial prolactin concentration. In 12 and 14 percent of samples, using polyethyleneglycol and ultrafiltration respectively, there was a prolactin recuperation of less than 40 percent. Eight and 11 percent of samples with a prolactin concentration of more than 30 ng/ml, had a recuperation of less than 40 percent using polyethyleneglycol and ultrafiltration respectively. Conclusions: Approximately 10 percent of samples with a prolactin concentration over 30ng/ml have recuperation values suggestive of the presence of macroprolactin. There is a good concordance between precipitation using polyethyleneglycol or ultrafiltration.

Valores de alerta en el laboratorio clínico y su aporte al cuidado y seguridad del paciente / The contribution of critical laboratory values to patients' safety and care

Critical values are those laboratory values that are so abnormal that may threaten the life of a patient unless immediate corrective or therapeutic actions are undertaken. Among laboratory procedures, this definition has been incorporated to standards that watch over patients' safety. Health institutions should incorpórate this practice and monitor its effectiveness.

Micetoma por Actinomadura madurae en el pie: reporte de un caso en Chile / Actinomadura madurae mycetoma of the foot: report of one case

Mycetoma is a chronic infection that affects skin, subcutaneous tissue and bone. Its etiology can be mycotic or bacterial. It affects mainly the lower extremities ofmiddie age men livingin tropical climates. We repon a 44 year-old male ¡ivingin a template zone, consulting for swelling and pain in the left foot, lasting for 10 years. Physical examination showed a swollen left foot with hyperpigmented skin and a few crustedpapules. Radiology showed an extensive bone involvement of the midfoot with several oval and radiolucid images. Magnetic resonance showed son and bone tissue involvement, with múltiple oval and low intensity images in TI and T2. The biopsy was compatible with an unspecific chronic osteomyelitis. A bacterial identification by polymerase chain reaction and sequencing in the biopsy determined the presence of an Actinomadura madurae. Treatment with cotrimoxazol was started).