Estrategia de PCR múltiple para la caracterización molecular simultánea de Staphylococcus aureus y enterotoxinas estafilocócicas en aislamientos de brotes de origen alimentario

Introduction: Staphylococcal food poisoning is the most frequent type of food poisoning around the world. Staphylococcus aureus enterotoxins cause significant loss of water in the intestinal lumen, followed by vomiting and diarrhea. Objective: To report a fast, reliable and inexpensive strategy based on multiplex PCR for the simultaneous identification of S. aureus and detection of five classical S. aureus enterotoxin genes (sea, seb, sec, sed, see) in Staphylococcus spp. strains isolated from food poisoning outbreaks. Materials and methods: We analyzed isolates from 12 food poisoning outbreaks occurred in Santa Fe province (Argentina). Isolation and phenotypic characterization were carried out by standard procedures. Genotypic analysis was performed by multiplex PCR, using primers for nuc, sea-see and 16S rRNA genes simultaneously. Results: Of all the strains tested, 58% were found to carry toxigenic genes. Sea and seb toxins were found at the same percentage (29%) while sec, sed and see genes were found in a lower and identical proportion (14%). We did not find more than one different type of S. aureus enterotoxin in the isolates analyzed. Conclusions: The multiplex PCR strategy designed in this work has enabled us to identify strains of S. aureus and detect -at the same time- their enterotoxigenic ability. At present, our efforts are devoted to the detection of genes encoding enterotoxins other than the classical ones, in order to know their impact on staphylococcal food poisoning, as well as to investigate their relevance to our country’s public health. http://dx.doi.org/10.7705/biomedica.v33i1.816  Introducción. La intoxicación alimentaria estafilocócica es la más frecuente de las intoxicaciones alimentarias. La acción de las enterotoxinas de Staphylococcus aureus en la luz intestinal provoca una importante pérdida de agua que origina vómitos y diarrea. Objetivo. Presentar una estrategia rápida, fiable y de bajo costo, basada en una reacción en cadena de la polimerasa (PCR) múltiple para, simultáneamente, identificar S. aureus y detectar genes de las cinco enterotoxinas clásicas de S. aureus (sea, seb, sec, sed, see) en cepas de Staphylococcus spp. aisladas de alimentos. Materiales y métodos. Se trabajó con aislamientos recuperados de 12 brotes de intoxicación alimentaria estafilocócica ocurridos en la provincia de Santa Fe, Argentina. El aislamiento y la caracterización fenotípica se llevaron a cabo mediante procedimientos estándar. La evaluación genotípica se hizo por una PCR múltiple, utilizando simultáneamente cebadores para los genes nuc, sea-see y 16S rRNA. Resultados. En las cepas analizadas se detectó 58 % de portadoras de genes toxigénicos. Las toxinas sea y seb se encontraron en igual porcentaje (29 %), mientras que las sec, sed y see se encontraron en menor e idéntica proporción (14 %). No encontramos más de un tipo diferente de enterotoxinas de S. aureus en los aislamientos analizados. Conclusiones. La estrategia de PCR múltiple diseñada en este trabajo permitió identificar cepas de S. aureus y confirmar, al mismo tiempo, su enterotoxigenicidad. En estos momentos, nuestros esfuerzos están dirigidos a detectar los genes que codifican enterotoxinas distintas de las clásicas, a fin de conocer su incidencia en las intoxicaciones alimentarias estafilocócicas e investigar su relevancia en la salud pública de nuestro país.  doi: http://dx.doi.org/10.7705/biomedica.v33i1.816 

Adverse events during reoperative cardiac surgery: Frequency, characterization, and rescue

ObjectivesTo (1) determine frequency of occurrence and risk factors for intraoperative adverse events (IAE) during reoperative cardiac surgery, (2) characterize them with respect to structure injured, timing, and use of preventive strategies, and (3) identify the impact on outcome in terms of successful and unsuccessful rescue and cost.MethodsOperative notes of 1847 patients undergoing reoperative cardiac surgery were reviewed to identify and characterize documented intraoperative adverse events. Logistic regression modeling was used to identify risk factors for intraoperative adverse events and outcomes. Expected versus observed poor outcomes (stroke, myocardial infarction, death) was used to measure rescue.ResultsAmong 127 patients, 145 (7%) intraoperative adverse events occurred. These included injuries to bypass grafts (n = 47), heart (n = 38), and great vessels (n = 28) and ischemia without graft injury (n = 22). Most occurred on opening (n = 34, 23%) and during prebypass dissection (n = 57, 39%). Risk incremented as reoperations increased. Seventy-seven patients experienced 1 or more lapses in preventive strategies. Patients with intraoperative adverse events had a greater number of poor outcomes (n = 24 [19%] vs n = 107 [6.2%]; P < .0001) and incurred higher direct technical intraoperative and postoperative costs (ratio 1.3). Twelve patients with intraoperative adverse events were predicted to have poor outcomes versus 24 who did (P < .0001), indicating 12 “failures to rescue.”ConclusionsAdverse events still occur regularly during cardiac reoperation, are related to complexity of the procedure, and occur particularly during dissection and often when preventive strategies have not been used. Compensatory rescue measures are not always successful. Adverse events lead to poor patient outcome and higher cost

La firma electrónica: la necesidad de una compatiblización legislativa a nivel internacional

Introducci&oacute;n. Los servicios de la Sociedad de la Informaci&oacute;n. La firma electr&oacute;nica en el comercio internacional: su importancia. La armonizaci&oacute;n com t&eacute;nica legislativa en torno de la firma digital. La t&eacute;cnica de las Leyes Modelo como reguladoras del comercio inyernacional. La Ley Modelo de la CNUDMI para las firmas electr&oacute;nicas (2001). Cuestiones en torno a la t&eacute;cnica legislativa en materia de firmas electr&oacute;nicas a nivel nacional. Regulaciones en materia de firma electr&oacute;nica, con especial referencia al reconocimiento de la firma digital

Descripción de un brote de intoxicación alimentaria estafilocócica ocurrido en Las Rosas, Provincia de Santa Fe, Argentina

En febrero de 2008 se denunció en la localidad santafecina de Las Rosas un presunto brote de una enfermedad transmitida por alimentos. En los procedimientos oficiales no fue posible determinar la cantidad de personas afectadas luego de consumir canelones de verdura adquiridos en un local comercial. Se auditó el establecimiento elaborador y se tomaron muestras del citado producto, muestras ambientales e hisopados de los manipuladores de alimentos. Se entrevistó a los afectados y se recuperaron restos del alimento ingerido. Mediante análisis microbiológicos de rutina se determinó la presencia de S. aureus subespecie aureus coagulasa positivo en muestras del producto consumido, del producto crudo y de los manipuladores del alimento. Los microorganismos indicadores no mostraron niveles significativos y no se aisló otro patógeno de transmisión alimentaria. Se investigó la presencia de genes productores de enterotoxinas estafilocócicas, con resultado positivo para la enterotoxina B en las cepas aisladas de un manipulador y en el alimento vinculado con el brote. Al analizar los aislamientos por SmaI-PFGE se demostró un 100% de similitud entre ellos. La notificación oportuna, las acciones sanitarias coordinadas y la disponibilidad de las herramientas de laboratorio apropiadas permitieron identificar los factores de protección y de riesgo, y cortar la cadena de transmisión de la enfermedad.On February 2008, a suspected foodborne outbreak was reported in Las Rosas (Santa Fe Province, Argentina). The formal procedures indicated that an undetermined number of individuals had experienced food poisoning following consumption of vegetable cannelloni bought at a local shop. The manufacturer establishment was audited. Samples from the suspected food, as well as environmental samples and swabs from food handlers were obtained and involved subjects were interviewed. Remnants of ingested food were also obtained. Routine microbiological procedures of the foodborne outbreak revealed the presence of coagulase positive S. aureus subspecies aureus in samples from ingested and raw food, and from manipulators. Indicator microorganisms did not show significant levels and no other foodborne pathogen was isolated. Presence of staphylococcal enterotoxin-producing genes was subsequently investigated, and a positive result for enterotoxin B was shown in S. aureus strains isolated from a food handler as well as from food linked to the outbreak Moreover, these isolates showed 100% similarity by SmaI-PFGE. Timely notification together with coordinated sanitary measures and the availability of appropriate laboratory tools allowed to interrupt the chain of disease transmission by identifying risk and protective factors

Lrg1p Is a Rho1 GTPase-Activating Protein Required for Efficient Cell Fusion in Yeast

To identify additional cell fusion genes in Saccharomyces cerevisiae, we performed a high-copy suppressor screen of fus2Δ. Higher dosage of three genes, BEM1, LRG1, and FUS1, partially suppressed the fus2Δ cell fusion defect. BEM1 and FUS1 were high-copy suppressors of many cell-fusion-defective mutations, whereas LRG1 suppressed only fus2Δ and rvs161Δ. Lrg1p contains a Rho-GAP homologous region. Complete deletion of LRG1, as well as deletion of the Rho-GAP coding region, caused decreased rates of cell fusion and diploid formation comparable to that of fus2Δ. Furthermore, lrg1Δ caused a more severe mating defect in combination with other cell fusion mutations. Consistent with an involvement in cell fusion, Lrg1p localized to the tip of the mating projection. Lrg1p-GAP domain strongly and specifically stimulated the GTPase activity of Rho1p, a regulator of β(1-3)-glucan synthase in vitro. β(1-3)-glucan deposition was increased in lrg1Δ strains and mislocalized to the tip of the mating projection in fus2Δ strains. High-copy LRG1 suppressed the mislocalization of β(1-3) glucan in fus2Δ strains. We conclude that Lrg1p is a Rho1p-GAP involved in cell fusion and speculate that it acts to locally inhibit cell wall synthesis to aid in the close apposition of the plasma membranes of mating cells