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Abstract Introduction: A comprehensive cytometry assessment in the critical ill patient shows modifications in cell lines that estimate severity and mortality in sepsis. The objective of this study is to determine the utility of different cytometric parameters and indices as predictors of mortality in septic patients. Materials and Methods: Retrospective cohort study of adults with sepsis (SEPSIS Criteria 3) hospitalized in an Intensive Unit Care (Quito, Ecuador). Patients with neoplasms or immunodeficiency states were excluded. Different cytometric parameters have been assessed and logistic regression models were used to stablish the predictive range of mortality for each parameter and areas under the curve (AUC) for sensitivity analysis. Results: Over 159 patients, the mortality was 25%. In non-survivors, the median of the APACHE II was 25.20 points, and the median of the SOFA was 11.18, 10.44, 10.15 points at the time of admission, 48, and 72 hours respectively. About the sensitivity analysis for mortality, the cut-off point of EDW was 14.5% (AUC 0.708), and it presented an adjusted OR of 5.25 (95%CI: 1.64-16.76, p: 0.005). The cut-off point of MPV was 8.45 fL (AUC 0.666), and it had an adjusted OR of 5.28 (95%CI: 1.72-16.21, p 0.004). Conclusions: EDW and MPV are independent predictors of mortality, and they must be used with scales or biomarkers to optimize the management and therapy of patients with sepsis. They would be an alternative in centers where only blood cytometry is available as an analytical test.
Resumen Introducción: Una evaluación completa de citometría en el paciente enfermo crítico muestra modificaciones en las líneas celulares que estiman la gravedad y la mortalidad en la sepsis. El objetivo de este estudio es determinar la utilidad de diferentes parámetros e índices citométricos como predictores de la mortalidad en pacientes sépticos. Materiales y métodos: Estudio retrospectivo de cohortes de adultos con sepsis (Criterio 3 de la SEPSIS) hospitalizados en una Unidad de Cuidados Intensivos (Quito,Ecuador). Se excluyeron los pacientes con neoplasias o estados de inmunodeficiencia. Se evaluaron diferentes parámetros citométricos y se utilizaron modelos de regresión logística para establecer el rango predictivo de la mortalidad para cada parámetro y las áreas bajo la curva (AUC) para el análisis de sensibilidad. Resultados: En más de 159 pacientes, la mortalidad fue del 25%. En los no supervivientes, la mediana del APACHE II fue de 25,20 puntos, y la mediana del SOFA fue de 11,18, 10,44 y 10,15 puntos en el momento del ingreso, 48 y 72 horas respectivamente. En cuanto al análisis de sensibilidad para la mortalidad, el punto de corte del EDW fue 14,5% (AUC 0,708), y presentó un OR ajustado de 5,25 (IC 95%: 1,64-16,76, p: 0,005). El punto de corte de MPV fue de 8,45 fL (AUC 0,666), y presentó un OR ajustado de 5,28 (95%CI: 1,72-16,21, p 0,004). Conclusiones. EDW y MPV son predictores independientes de mortalidad, y deben ser utilizados con escalas o biomarcadores para optimizar el manejo y la terapia de los pacientes con sepsis. Serían una alternativa en los centros donde sólo se dispone de citometría de sangre como prueba analítica.
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Humanos , Adulto , Sepsis , Biomarcadores , Estudios de Cohortes , Mortalidad , Sobrevivientes , Citometría de Flujo , Unidades de Cuidados IntensivosRESUMEN
Introducción: La citometría de flujo permite la cuantificación de las subpoblaciones de linfocitos con una elevada sensibilidad, especificidad y objetividad. Estas ventajas solo se logran con un proceso laborioso de diseño individualizado y controlado para cada experimento. Objetivo: Diseñar un protocolo de un solo tubo policromático de citometría flujo para inmunofenotipo linfocitario periférico. Métodos: Se realizó un estudio experimental in vitro con muestras de sangre periférica obtenidas de tres voluntarios sanos, en el Centro Nacional de Genética Médica, en marzo de 2019. El tubo se compuso de seis marcadores de linaje para identificar linfocitos B, T, natural killer y natural killer T. Se desarrolló un protocolo de lisis de hematíes sin lavado. Se emplearon anticuerpos monoclonales conjugados con fluorocromos. El punto óptimo de concentración correspondió al mayor índice de tinción y conservación de los porcentajes de positividad de cada población. Se realizó la construcción progresiva del tubo y se propuso una estrategia lógica de secuencia de ventanas para el análisis de datos. Resultados: Los marcadores seleccionados permitieron realizar correctamente el inmunofenotipo linfocitario periférico. En los cinco puntos de titulación se observaron buenas discriminaciones entre las señales positivas y negativas, excepto para el anti-CD56 que presentó una tendencia decreciente del índice de tinción. El volumen total de conjugados requeridos para la determinación de los 6 antígenos fue de 3,75 μL por tubo. Conclusiones: Se obtuvo un tubo policromático que permite el inmunofenotipo periférico de forma rápida y precisa por seis antígenos linfocitarios simultáneamente, con el empleo de pequeños volúmenes de conjugado y sangre(AU)
Introduction: Flow cytometry allows quantification of lymphocyte subpopulations with high sensitivity, specificity and objectivity. These advantages are only achieved through the hardworking process of individualized and controlled design for each experiment. Objective: To design a flow cytometry protocol of a single polychromatic tube for peripheral lymphocyte immunophenotype. Methods: An experimental in vitro study was carried out, in March 2019, with peripheral blood samples obtained from three healthy volunteers, at the National Center for Medical Genetics. The tube was made up of six lineage markers for identifying natural B and T lymphocytes, natural killers and natural killer T cells. A protocol was developed for red blood cell lysis without washing. Fluorochrome-conjugated monoclonal antibodies were used. The optimal point of concentration corresponded to the highest staining index and preservation of the positivity percentages of each population. Progressive tube construction was performed and a logical window sequence strategy was proposed for data analysis. Results: The chosen markers allowed to carry out correct peripheral lymphocyte immunophenotype. Good discriminations between positive and negative signals were observed at the five titration points, except for anti-CD56, which presented a decreasing trend in the staining index. The total volume of conjugates required for determination of the six antigens was 3.75 μL per tube. Conclusions: A polychromatic tube was obtained that allows to carry out peripheral immunophenotype quickly and precisely by six lymphocyte antigens simultaneously, with the use of small volumes of conjugate and blood(AU)
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Humanos , Optimización de Procesos , Citometría de Flujo/métodos , Genética Médica , Industria de la ConstrucciónRESUMEN
RESUMEN La enfermedad del COVID-19 presenta actualmente una gama de síntomas que van desde la presencia de síntomas clásicos de compromiso de las vías respiratorias altas y bajas hasta alteraciones sensitivas como anosmia o trastornos del gusto. Las trombosis valvulares cardíacas y la endocarditis subsecuente son presentaciones extremadamente raras y poco descritas de la infección por COVID-19. Presentamos el caso de un paciente positivo para virus SARS-Cov2, quien desarrolló una bacteriemia por Staphylococcus hominis y que, ante episodios febriles persistentes y posterior evaluación de diagnósticos diferenciales, se identificó el diagnóstico de endocarditis infecciosa en válvula aórtica nativa sana -mediante ecocardiografía-, la cual fue adquirida en el hospital.
ABSTRACT Covid-19 disease shows many different manifestations, from common symptoms affecting both the upper and lower respiratory tract, and also sensorial alterations such as anosmia or taste perversion. Heart valve thrombosis and subsequent endocarditis are extremely rare manifestations and they have been scarcely described within the context of Covid-19. We present a case of a patient that was positive for SARS-Cov-2, who developed Staphylococcus hominis bacteremia. Afterwards, he developed persistent fever, and after ruling out some differential diagnoses, a diagnosis of infective endocarditis affecting a native healthy aortic valve was made, using cardiac ultrasonography. This occurrence was considered to be hospital-acquired.
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ABSTRACT Introduction: the objective of this study was to use real-time qPCR to identify and quantify the Streptococcus mutans species in samples of saliva and dental biofilm. Methods: 27 children were randomly chosen with the following criteria: 8 years of age, low socio-economic levels, residing in the northern metropolitan area of Santiago de Chile; they were asked to attend an appointment while fasting with no teeth brushing for at least 12 hours, in order to collect non-stimulated saliva and a pool of supragingival dental biofilm of all the mesio-vestibular sides of anterior and posterior teeth. The amount of S. mutans in the samples was quantified by qPCR using primers that amplify a fragment of the gtfB gene of S. mutans. Results: the amplification showed 98% efficiency with a fluorescence of 3.36 cycles. The melting curve presented a single maximum at the same temperature for all samples. Conclusion: the methodology allows the specific identification and quantification of gene gtfB of S. mutans in saliva and dental biofilm in a quick and reliable manner, contributing to the identification of individual cariogenic risk.
RESUMEN. Introducción: el objetivo del presente estudio consistió en implementar la técnica de qPCR en tiempo real para identificar y cuantificar la especie Streptococcus mutans en muestras de saliva y biopelícula dentaria. Métodos: se seleccionaron al azar 27 niños de 8 años de edad, de nivel socio-económico bajo del área norte de la región metropolitana de Santiago de Chile, que se citaron en ayunas y sin cepillado durante al menos 12 horas, para colectar saliva no estimulada y un pool de biopelícula dentaria supragingival de todas las caras mesio-vestibulares de dientes anteriores y posteriores. Se cuantificó la cantidad de S. mutans en las muestras mediante qPCR empleando partidores que amplifican un fragmento del gen gtfB de S. mutans. Resultados: la amplificación presentó 98% de eficiencia con delta de fluorescencia de 3,36 ciclos. La curva de fusión (melting) presentó un solo máximo a una misma temperatura para todas las muestras. Conclusión: la metodología permite la identificación y cuantificación específica del gen gtfB de S. mutans en muestras de saliva y biopelícula dentaria, de forma rápida y exacta, aportando a la determinación del riesgo cariogénico individual.
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Placa Dental , Saliva , Streptococcus , NiñoRESUMEN
OBJECTIVES: This study investigated the relationship between urease and arginine deiminase system (ADS) activities and dental caries through a cross-sectional study. MATERIAL AND METHODS: Urease and ADS activities were measured in saliva and plaque samples from 10 caries-free subjects and 13 caries-active. Urease activity was obtained from the ammonia produced by incubation of plaque and saliva samples in urea. ADS activity was obtained from the ammonia generated by the arginine-HCl and Tris-maleate buffer. Specific activity was defined as micromoles of ammonia per minute per milligram of protein. Shapiro-Wilk statistical test was used to analyze the distribution of the data, and Mann-Whitney test was used to determine the significance of the data. RESULTS: The specific urease activity in saliva and plaque was significantly higher in individuals with low DMFT scores. ADS activity in saliva (6.050 vs 1.350, p=0.0154) and plaque (8.830 vs 1.210, p=0.025) was also higher in individuals with low DMFT scores. CONCLUSIONS: Caries-free subjects had a higher ammonia generation activity by urease and arginine deiminase system for both saliva and plaque samples than low caries-active subjects. High levels of alkali production in oral environment were related to caries-free subjects.
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Caries Dental/enzimología , Placa Dental/enzimología , Hidrolasas/análisis , Saliva/enzimología , Ureasa/análisis , Álcalis/metabolismo , Amoníaco/análisis , Estudios de Casos y Controles , Estudios Transversales , Índice CPO , Placa Dental/química , Femenino , Humanos , Masculino , Valores de Referencia , Factores de Riesgo , Saliva/química , Estadísticas no ParamétricasRESUMEN
Objectives: This study investigated the relationship between urease and arginine deiminase system (ADS) activities and dental caries through a cross-sectional study. Material and Methods: Urease and ADS activities were measured in saliva and plaque samples from 10 caries-free subjects and 13 caries-active. Urease activity was obtained from the ammonia produced by incubation of plaque and saliva samples in urea. ADS activity was obtained from the ammonia generated by the arginine-HCl and Tris-maleate buffer. Specific activity was defined as micromoles of ammonia per minute per milligram of protein. Shapiro-Wilk statistical test was used to analyze the distribution of the data, and Mann-Whitney test was used to determine the significance of the data. Results: The specific urease activity in saliva and plaque was significantly higher in individuals with low DMFT scores. ADS activity in saliva (6.050 vs 1.350, p=0.0154) and plaque (8.830 vs 1.210, p=0.025) was also higher in individuals with low DMFT scores. Conclusions: Caries-free subjects had a higher ammonia generation activity by urease and arginine deiminase system for both saliva and plaque samples than low caries-active subjects. High levels of alkali production in oral environment were related to caries-free subjects. .
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Humanos , Masculino , Femenino , Caries Dental/enzimología , Placa Dental/enzimología , Hidrolasas/análisis , Saliva/enzimología , Ureasa/análisis , Álcalis/metabolismo , Amoníaco/análisis , Estudios de Casos y Controles , Estudios Transversales , Índice CPO , Placa Dental/química , Valores de Referencia , Factores de Riesgo , Saliva/química , Estadísticas no ParamétricasRESUMEN
There is increasing evidence that male or female genital tract represent a distinct replication compartment for human immunodeficiency virus type 1 (HIV-1) and that such compartments may serve as a virus reservoir. Forty-four paired plasma and vaginal samples from HIV-infected females undergoing HAART were collected to examine the viral responses to antiretroviral therapy and to assess the possible role of the vaginal tract as a reservoir for drug-resistant variants. Twenty-one females had detectable viral RNA both in plasma and vaginal fluid, whereas 14 females had detectable virus only in plasma. Twelve paired samples were used to analyze HIV-1 pol sequences for the presence of drug resistance-associated mutations. Nine of the twelve paired samples exhibited discordant drug resistance mutation patterns. The other three females showed identical drug resistance-associated mutations. However, further examination of protease and RT showed numerous non-drug-associated mutations that corresponded to predefined CTL epitopes. These non-drug-associated mutations were different between plasma and vaginal viruses, suggesting that evolution of HIV-1 was independent in these two compartments.