Influencia de un panel molecular de reacción de polimerasa en cadena múltiple en el diagnóstico o descarte de neumonía asociada a ventilación mecánica en una cohorte chilena de pacientes críticos con COVID-19

Resumen Introducción: La neumonía asociada a ventilación mecánica (NAVM) es frecuente en pacientes críticos con COVID-19. Su diagnóstico precoz es fundamental para su pronóstico. Objetivo: Demostrar la utilidad del panel FilmArray Pneumo (PFA-P) en la confirmación o descarte de NAVM en estos pacientes. Métodos: Estudio retrospectivo de 71 pacientes críticos con COVID-19 con sospecha de NAVM en quienes se realizó cultivos y PFA-P para confirmación diagnóstica. Se describen las características clínicas, microbiología y mortalidad. Se define la validez y seguridad de PFA-P. Resultados: El uso de FAP-P y cultivos descartó NAVM en 29 pacientes (40,8%). En 41 pacientes se confirmó NAVM y la mortalidad a 30 días fue 48,8%. Se estudiaron 48 muestras, los cultivos fueron positivos en 30 (62,5%) y se detectaron 33 bacterias, PFA-P detectó 32 de estas 33 bacterias; 37 bacterias fueron detectadas exclusivamente por PFA-P. Las bacterias prevalentes fueron Klebsiella pneumoniae (31,4%) Pseudomonas aeruginosa (21,4%) y Acinetobacter calcoaceticusbaumannii (14,2%). La sensibilidad, especificidad, valor predictor positivo y valor predictor negativo de PFA-P con respecto a cultivos fue 96,9%, 92,5%, 46,4% y 99,8, respectivamente. Un paciente tuvo NAVM por Burkholderia cepacia bacteria no detectada por FAP-P. Conclusiones: La FAP-P es una técnica molecular eficaz para descartar y diagnosticar la NAVM permitiendo una suspensión rápida de los antimicrobianos o un tratamiento dirigido temprano.

Abstract Background: Ventilator-associated pneumonia (VAP) is frequent in critical COVID-19 patients. Its early diagnosis is essential for its prognosis. Aim: To demonstrate the usefulness of the FilmArray Pneumo panel (FAP-P) in confirming or ruling out VAP in these patients. Methods: Retrospective study of 71 critical COVID-19 patients with suspected VAP in whom cultures and FAP-P were performed for diagnostic confirmation. Clinical characteristics, microbiology and mortality are described. The validity and safety of FAP-P is defined. Results: The use of FAP-P and cultures ruled out VAP in 29 patients (40.8%). In 41 patients, VAP was confirmed and the 30-day mortality was 48.8%. Forty-eight samples were studied, the cultures were positive in 30 (62.5%) and 33 bacteria were detected, FAP-P detected 32 of these 33 bacteria; 37 bacteria were exclusively detected by PFA-P. The most prevalent bacteria were Klebsiella pneumoniae (31.4%), Pseudomonas aeruginosa (21.4%) and Acinetobacter calcoaceticus-baumannii (14.2%). The sensitivity, specificity, positive predictive value and negative predictive value of FAP-P with respect to cultures were 96.9%, 92.5%, 46.4% and 99.8%, respectively. One patient had VAP due to Burkholderia cepacia bacteria not detected by FAP-P. Conclusions: FAP-P is an effective molecular technique to rule out and diagnose VAP, allowing rapid suspension of antibiotics or early targeted treatment.

Comportamiento del consumo de oxígeno durante una sesión de rehabilitación cardíaca / Behavior of Oxygen Consumption During a Cardiac Rehabilitation Session

RESUMEN Objetivo: estudiar el comportamiento del consumo de oxígeno en pacientes con miocardiopatía dilatada durante una sesión de rehabilitación cardíaca. Material y métodos: diseño observacional, transversal, analítico relacional. Muestra: 10 pacientes masculinos con miocardiopatía dilatada, con deterioro de moderado a grave de la función ventricular. Se evaluó a los pacientes en laboratorio y en una sesión de rehabilitación mediante un analizador de gases portátil Medgraphics® VO 2000. La sesión de rehabilitación consistió en ejercicios en bicicleta fija de 10 minutos, step, escalera coordinativa, fuerza con mancuerna para bíceps y hombros, cuádriceps en camilla y dorsales en máquina. Resultados: Edad (años) 57,4 ± 14,6. Peso (kg) 91,4 ± 22,2. Talla (cm) 168,1 ± 6,2. Laboratorio: VO2max relativo (ml/kg/min) 21,8 ± 7,3. Tasa de intercambio respiratorio, RER (VCO2/VO2)1,05 ± 0,09. Volumen ventilado (L/min) 65,7 ± 18,5. Frecuencia cardíaca (lat./min) en VO2max 127,8 ± 23,8. Sesión: Duración (min) 37,5 ± 10. VO2pico (ml/kg/min) 14,6 ± 3 (69,9 ± 16,7% del VO2max). Coeficiente de correlación entre VO2max y tiempo con VO2 < 50% del VO2max (min) 0,662 (p = 0,037) y entre VO2pico en rehabilitación y tiempo en RER entre 0,85-1(min) 0,787 (p = 0,007). Los pacientes con mejor aptitud ejercitaron en zona de baja intensidad. Al aumentar el esfuerzo, aumentaron los minutos en intensidad moderada. Conclusión: Se constató en este estudio que los pacientes alcanzaron un VO2pico en las sesiones inferiores a sus máximos obtenidos en laboratorio. Si bien cualquier dosis de entrenamiento en estos pacientes es más beneficiosa que la inactividad física, el diseño y la planificación de las sesiones de RHC, valorando las intensidades de trabajo intrasesión, podrían generar mayor impacto en la mortalidad, las reinternaciones y en la calidad de vida.

ABSTRACT Objective: The aim of this study was to analyze the response of oxygen consumption in patients with dilated cardiomyopathy during a cardiac rehabilitation session. Methods: This was an observational, cross-sectional, relational analytical study. Ten male patients with dilated cardiomyopathy and moderate to severe ventricular dysfunction were included in the study. Patients were evaluated in the laboratory and during a rehabilitation session using a Medgraphics VO 2000 portable gas analyzer. The rehabilitation session consisted in 10 minutes of stationary bike exercises, step, coordinaton stairs, and muscle strength using dumbbells for biceps and shoulder, a quadriceps stretcher and a dorsal muscle machine. Results: Mean age was 57.4 ± 14.6 years, weight 91.4 ± 22.2 kg and height 168.1 ± 6.2 cm. In the laboratory, VO2max was 21.8 ± 7.3 ml/kg/min, respiratory exchange rate (RER) (VCO2/VO2) 1.05 ± 0.09, ventilated volume 65.7 ± 18.5 L/min and heart rate in VO2max 127.8 ± 23.8 beats/min. Rehabilitation session duration was 37.5 ± 10 min with peakVO2 14.6 ± 3 ml kg/min (69.9 ± 16.7 % VO2max). The correlation coefficient between VO2max and time with VO2 <50% VO2max(min) was 0.662 (p = 0.037) and between peakVO2 in rehabilitation and time in RER between 0.85-1 (min) was 0.787 (p = 0.007). Patients with better fitness exercised in the low-intensity zone. As exercise increased, the minutes in moderate intensity also increased. Conclusion: The study showed that patients reached peakVO2 in sessions below the maximum values obtained in the laboratory. Even though any dose of training in these patients was more beneficial than physical inactivity, cardiac rehabilitation session design and planning, taking into account intrasession exercise intensities, could generate greater impact on mortality, rehospitalizations and quality of life.

Tumor sólido pseudopapilar de páncreas: reporte de caso / Pseudopapilar solid tumor of pancreas: case Report

Resumen Introducción: El tumor sólido pseudopapilar del páncreas es una rara entidad que representa menos del 1% de las neoplasias pancreáticas. Suele presentarse en mujeres jóvenes y solo da síntomas de carácter compresivo una vez que alcanza un gran tamaño. Dado su comportamiento biológico incierto el tratamiento es la cirugía. Caso Clínico: Presentamos el caso de una mujer de 23 años con historia de 1 año de evolución de dolor epigástrico y baja de peso. El estudio imagenológico demostró una masa heterogénea sólida-quística dependiente de la cabeza del páncreas de aspecto neoplásico. Se realizó una biopsia incisional laparoscópica cuyo resultado fue de un tumor maligno indiferenciado, por lo que se optó por la resección quirúrgica. Se realizó una pancreatoduodenectomía abierta sin incidentes con un postoperatorio favorable. Los análisis histopatológicos e inmunohistoquímico fueron compatibles con un tumor sólido pseudopapilar de páncreas.

Introduction: The pseudopapillary solid tumor of the pancreas is a rare entity that represents less than 1% of pancreatic neoplasms. It usually occurs in young women and only gives symptoms of a compressive nature once it has reached a large size. Given its uncertain biological behavior, the treatment is surgery. Case Report: We present the case of a 23-year-old woman with a 1-year history of epigastric pain evolution and weight loss. The imaging study demonstrated a solid-cystic heterogeneous mass dependent on the head of the pancreas of neoplasic appearance. A laparoscopic incisional biopsy was performed, the result of which was an undifferentiated malignant tumor, which was why the surgical resection was chosen. An open pancreatoduodenectomy was performed without incident with a favorable post operative. Histopathological and immunohistochemical analyzes were compatible with a solid pseudopapillary tumor of the pancreas.

Aportes de la genética al estudio y manejo clínico de las miocardiopatías / Genetic contributions to the study and clinical management of cardiomyopathies

Resumen: Las miocardiopatías son patologías muy heterogéneas asociadas a muerte súbita en jóvenes, cuyo diagnóstico y pronóstico son difíciles de establecer. El estudio genético puede ser una herramienta importante para el abordaje de estos pacientes y sus familias. Son generalmente patologías monogénicas, con penetrancia incompleta y expresividad clínica variable. Múltiples causas genéticas subyacen a la misma patología y un mismo gen puede estar asociado con fenotipos diferentes. La tecnología disponible en la actualidad permite analizar todos los genes importantes para cada fenotipo, con costos y tiempos de entrega de los resultados muy razonables. La rentabilidad del estudio genético depende de cada patología y de la probabilidad pretest de cada paciente particular. La interpretación de un estudio genético es una tarea compleja y un aspecto limitante para una correcta implementación clínica. Depende de múltiples variables y deberá ser realizado por equipos multidisciplinares con experiencia clínica en las patologías y los genes asociados. Un estudio genético positivo aportará mucha información diagnóstica y pronóstica para el paciente y su familia. Esto permitirá hacer recomendaciones en el estilo de vida, seleccionar tratamientos específicos para determinadas patologías, y decidir con más precisión el momento oportuno para implementar técnicas preventivas invasivas. Está demostrado que el screening genético en cascada luego de un resultado positivo es una estrategia costo-efectiva, que permite grandes ahorros en seguimientos clínicos innecesarios para focalizar los recursos en individuos genéticamente predispuestos.

Summary: Cardiomyopathies are heterogeneous diseases associated with sudden death in the young. The diagnosis and associated prognosis is sometimes difficult to establish. The genetic study could be an important tool for the clinical work-up of patients and families with these diseases. Cardiomyopathies are usually monogenic diseases, with incomplete penetrance and variable clinical expressivity. Several genes are associated with the same phenotype, and a particular gene could be related with different diseases. All the genes related with a particular phenotype could be study with the available sequencing technology at a reasonable price and turnaround time for the results. The yield of genetic tests depends on the type of cardiomyopathy and is specifically driven by the clinical pre-test probability of each case. The interpretation of genetic studies is complex and the main challenge for the correct clinical application of the results. Interpretation depends on several variables and should be performed by multidisciplinary teams with clinical and genetic expertise on cardiomyopathies. A positive genetic study could contribute with important diagnostic and prognostic information for the patient and the family. This information could be useful for life-style modifications, specific treatment selection and, in some cases, to decide the correct moment for primary prevention device's implantation. Cascade family screening after a positive genetic diagnosis is a cost-effective strategy for health-care systems.

Tiroiditis de Riedel, una entidad fácilmente confundible con carcinoma anaplásico tiroideo / Riedel's thyroiditis, an entity easily confused with anaplastic thyroid carcinoma

INTRODUCTION: Riedel´s thyroiditis is a thyroid inflammation with invasive fibrosis. CASE REPORT: Female 84 years old, studied by dysphagia to liquids and dysphonia, with cervical mass poorly demarcated and immobile. Computed Tomography (CT) (CT) showed tumor that diverted the trachea and esophagus compressed. Fine Needle Aspiration (FNA) obtained cells with atypia. Thyroidectomy was performed, appreciating a stony tumor which included the right recurrent laryngeal nerve. Given the high suspicion of thyroid malignancy tracheostomy it was performed. The pathology report as fibrosing Riedel thyroiditis. DISCUSSION: This is the less common type of chronic thyroiditis. Etiology is unknown. The normal thyroid parenchyma is replaced by fibrous connective tissue. Clinically usually a hard mass, fast growing, why is confused with thyroid carcinoma and Hashimoto's thyroiditis. It´ss more common in women and is usually bilateral. The CT or MRI (Magnetic Resonance Imaging) help assess the extent. Puncturing the mass could give us the diagnosis but is often difficult. Treatment to relieve symptoms or rule out malignancy is surgery. The prognosis is usually good. CONCLUSIONS: Riedel´s thyroiditis is an entity with histopathologic diagnosis essential

Angiosarcoma primario de mama y carcinoma papilar de tiroides sincrónico: presentación de un caso / Primary angiosarcoma of breast and synchronous thyroid papillary carcinoma: a case report

Antecedentes: El angiosarcoma primario de mama constituye un tumor infrecuente que se desarrolla de forma predominante en mujeres en la tercera y cuarta décadas de la vida. Los nódulos tiroideos incidentales son aquellos que se objetivan en pruebas de imagen realizadas por otra causa y se observan en menos de un 10 por ciento con la tomografía por emisión de positrones (PET). Objetivo: Presentar el caso excepcional de una paciente con angiosarcoma primario de mama y un carcinoma papilar de tiroides sincrónicos. Caso clínico: Mujer de 34 años con angiosarcoma primario en la mama derecha sometida a mastectomía simple. En el estudio de extensión se halló un incidentaloma tiroideo derecho compatible con carcinoma papilar que requirió tiroidectomía total y linfadenectomía del compartimento central cervical en un segundo tiempo. Comentarlo: Los angiosarcomas primarios suponen menos del 0,05 por ciento de todos los tumores primarios malignos de la mama. El diagnóstico definitivo de estos tumores viene definido por el estudio anatomopatológico, que establece tres grados: alto, bajo e intermedio, los cuales se relacionan de forma directa con la supervivencia. La prevalencia de los incidentalomas malignos tiroideos objetivados por PET varía del 14-30,9 por ciento, según diferentes estudios. La planificación del tratamiento quirúrgico de las neoplasias incidentales tiroideas en pacientes con otro tumor primario debe analizarse de forma individualizada, según la supervivencia esperada del tumor primario.

Background: The primary angiosarcoma of breast is a rare tumor that develops predominantly in women in the third and fourth decades of life. Incidental thyroid nodules are observed in imaging tests performed for other reasons and it's observed in less than 10 percent with the positron emission tomography (PET). Aims: To report the exceptional case of a patient with primary angiosarcoma of the breast and synchronous thyroid papillary carcinoma. Clinical case: Female patient 34 year old with primary angiosarcoma in the right breast, submitted to a simple mastectomy. In the extension study we found an incidental thyroid papillary carcinoma that required a total thyroidectomy and central cervical lymphadenectomy in a second time. Comments: The primary angiosarcoma are less than 0.05 percent of all primary malignant tumors of the breast. The definitive diagnosis is determined by the pathology, which defines three grades: high, low and intermediate, which relate directly to survival. Prevalence of malignant thyroid incidentalomas observed by PET varies from 14 to 30.9 percent, according different studies. Planning the surgical treatment of incidental thyroid malignancies in patients with other primary tumor should be analyzed individually, according the expected survival of the primary tumor.

Síndrome de Sweet localizado sobre linfedema: descripción de un caso y revisión de la literatura / Sweet´s syndrome located on lymphoedema: a case report and review of literature

Comunicamos el caso de una mujer de 63 años de edad, que desarrolló lesiones compatibles con un síndrome de Sweet, localizadas en el miembro sometido a linfadenectomía e irradiación, debido a una neoplasia maligna neural. Efectuamos comentarios del caso y los comparamos con la bibliografía publicada.

A 63 years-old woman treated wit lymphadenectomy and radiotherapy due a malignant neural neoplasia, developed lesions of a Sweet's syndrome, localized in the area treated. A review and comparative study of the previous papers are made.

Haga su diagnóstico / Do your diagnostic

Presentamos un caso de esta entidad en una paciente caucásica, con lesiones maculosas purpúricas en zona tibial posterior de pierna derecha, sin afectación sistémica, de curso crónico y persistente en la actualidad, tras dieciocho meses de seguimiento con mejoría parcial, después de recibir tratamiento con esteroides e inmunosupresores tópicos. Los anticonceptivos orales y la hipercolesterolemia leve, podrían ser datos asociados en nuestro caso a dicha entidad, aunque se necesitarían más estudios para confirmar esta hipótesis.

We present a case of this entity in a Caucasian, patient with purpuric macular lesion area with her right posterior tibial, without affecting systemic, chronic and persistent course now, after eighteen months of follow-up with partial improvement after treatment with topical steroids and immunosuppressors. Oral contraceptives and mild hypercholesterolemia may be associated data in our case to the entity, althought further studies are needed to confirm this hypothesis.

Leucemia Cutis de presentación atípica / An atypical presentation of Leukemia Cutis

El trabajo aporta un nuevo caso de leucemia cutis, raro en frecuencia, que presenta un comienzo clínico muy poco habitual, a tener en cuenta. Fueron necesarias varias biopsias para llegar al diagnóstico de la paciente. La supervivencia de la paciente está por encima de la media en estos casos.

The work presents a new case of leukemia cutis, rare in frequency, which has a very unusual clinical onset to take into account. It took several biopsies for the diagnosis of the patient. The survival of the patient is above average in these cases.

Metabolomics: A Novel Approach to Early and Noninvasive Prostate Cancer Detection

Prostate cancer (PCa) is the most commonly diagnosed visceral cancer in men and is responsible for the second highest cancer-related male mortality rate in Western countries, with increasing rates being reported in Korea, Japan, and China. Considering the low sensitivity of prostate-specific antigen (PSA) testing, it is widely agreed that reliable, age-independent markers of the presence, nature, and progression of PCa are required to facilitate diagnosis and timely treatment. Metabolomics or metabonomics has recently emerged as a novel method of PCa detection owing to its ability to monitor changes in the metabolic signature, within biofluids or tissue, that reflect changes in phenotype and function. This review outlines the physiology of prostate tissue and prostatic fluid in health and in malignancy in relation to metabolomics as well as the principles underlying the methods of metabolomic quantification. Promising metabolites, metabolic profiles, and their correlation with the presence and stage of PCa are summarized. Application of metabolomics to biofluids and in vivo quantification as well as the direction of current research in supplementing and improving current methods of detection are discussed. The current debate in the urology literature on sarcosine as a potential biomarker for PCa is reviewed and discussed. Metabolomics promises to be a valuable tool in the early detection of PCa that may enable earlier treatment and improved clinical outcomes.

Desvascularización preoperatoria de tumor del glomus mediante colocación de stent cubierto en la carótida externa / Preoperative Devascularization of a Glomus Tumor with a Covered Stent in the External Carotid Artery

Los tumores del glomus carotídeo se caracterizan por su vascularización importante y su manejo preoperatorio puede incluir la embolización percutánea previa a la resección quirúrgica. Esta técnica disminuye la hemorragia y el tamaño del tumor y hace menos riesgosa la disección, con reducción de la morbimortalidad. Una técnica alternativa es la interrupción de la irrigación del tumor mediante la colocación de un stent cubierto en la carótida externa, que es la vía principal de irrigación. Esta técnica es útil en especial en tumores grandes y evita el riesgo de embolia intracraneal cuando se emplean coils para realizar la embolización. En esta presentación se describe el caso de una paciente de 31 años tratada de esta manera; se le colocó el stent y 24 horas después se realizó la resección del tumor.

Glomus tumors are hypervascularized neoplasms which may require preoperative percutaneous embolization. This technique reduces the incidence of bleeding, the tumor size, the risk of resection-related complications, and morbidity and mortality. The interruption of the tumor blood supply placing a covered stent in the external carotid artery, the main tumor-supplying vessel, is an alternative option. This technique is especially useful in large tumors and prevents the risk of intracranial embolism when coils are used during embolization. We describe the case of a 31 year-old female patient who underwent stent placement 48 hours before tumor resection.

Effect of magnesium oxide (MgO) addition in diets for lactating holstein cows / Efecto de la adición de óxido de magnesio (MgO) en dietas para vacas holstein en lactación

It was evaluated the effect of magnesium oxide (MgO) addition in diets for Holstein cows on milk production and quality. Nineteen Holstein cows in second third of lactation were assigned to two treatments (T), with 9 and 10 animals. Both treatments were similar for days in lactation and milk yield. Cows were assigned at random to two treatments (T) groups: T1 received 0.2% and T2 received 0.4% of MgO. The MgO (alkalinizing agent) was mixed with a commercial concentrate and offered in feed bunk with total diet, which consisted of 9 kg/d of concentrate plus alfalfa hay fed ad libitum. Total diet was offered three times a day (0600; 1200 and 1700 h). The experimental period was of 30 d, with other 30 for adaptation. Milk production (kg/d) and milk quality (fat, protein, and total solids) were evaluated. Data were analyzed using the statistical package SAS through of a randomized block design. Animals on T2 (0.40% MgO) produced 2.85 kg/d more milk (P<0.50) than those on T1 (0.20% MgO). The fat, protein and total solids content in milk were higher (P<0.05) in T1 than in T2. It is concluded that the addition of 0.4% MgO in diets for lactating dairy cows resulted in milk production increased, although there was a small reduction in milk quality, compared with 0.2% MgO supplementation.

Se evaluó el efecto de adición de óxido de magnesio (MgO) en dietas para vacas Holstein sobre la producción y calidad de la leche. Diecinueve vacas Holstein en el segundo tercio de lactancia fueron asignadas a dos tratamientos (T), con 9 y 10 animales. Ambos tratamientos fueron similares para días de lactancia y producción de leche. Las vacas fueron asignadas al azar a dos grupos o tratamientos (T): T1 recibió 0,2%, y T2 recibió 0,4% de MgO. El MgO (agente alcalinizante) se mezcló con un concentrado comercial y se ofreció en el comedero junto con la dieta total, la cual consistió de 9 kg/d del concentrado mas heno de alfalfa ad libitum. La dieta total se ofreció 3 veces al día (06000, 1200 and 1700 h). El periodo experimental fue de 30 d, con otros 30 d de adaptación. Se evaluó la producción de leche (kg/d) y la calidad de la leche (grasa, proteína, y sólidos totales). Los datos se analizaron por medio del paquete estadístico SAS en un diseño de bloques al azar. Los animales en T2 (0,4% MgO) produjeron más leche (2,5 kg/d; P<0,05) que aquellos en T1 (0,20% MgO). El contenido de grasa, proteína, y sólidos totales en leche fue más alto (P<0,05) en T1 que en T2. Se concluye que la adición de 0,40% de MgO en dietas para vacas lactantes incrementa la producción de leche, sin embargo puede haber una pequeña reducción en la calidad de la leche, comparado con la suplementación de 0,20% de MgO.

Patterns of woody plant species diversity in lebanon as affected by climatic and soil properties

Lebanese biodiversity is threatened by tourist and urban development, political instability, over-collection of medicinal and aromatic plants, lack of compliance to the regulations prohibiting over-exploitation from the wild, over-grazing and forest fires. A large number of the native species have unexplored economic potential for either medicinal or ornamental use. One way to preserve these species is by propagation and reintroduction into appropriate habitats. However, this requires an understanding of the species biology and environment. The relationship of nine species to the soil and climatic conditions in eight sites along an altitudinal gradient was studied. Individual species were counted and identified within transects at each site. Climatic data were collected and soil samples were taken and analyzed for soil texture, soil pH, EC, CaCOs, organic matter content and the following nutrients: Ca, Mn, Na, Fe, P, K, Cu, Mg, and Zn. Each ecosystem had a unique environment that could be described using the first two factors [70.3% of variation] in a Factor Analysis of the six most important variables. Some species' densities were affected by soil conditions [the first factor] while climatic conditions [the second factor] explained the densities of other species. Recommendations are made for the in-situ and ex-situ preservations of the nine species and their ecosystems

T-DNA transfer from Agrobacterium tumefaciens to the ectomycorrhizal fungus Pisolithus microcarpus / Transferencia de T-DNA de Agrobacterium tumefaciens al hongo ectomicorrícico Pisolithus microcarpus

The model ectomycorrhizal fungus Pisolithus microcarpus isolate 441 was transformed by using Agrobacterium tumefaciens LBA1100 and AGL-1. The selection marker was the Shble gene of Streptoallotecius hidustanus, conferring resistance to phleomycin, under the control of the gpd gene promoter and terminator of Schizophyllum commune. Transformation resulted in phleomycin resistant clones which were confirmed by PCR to contain the resistance cassette. A. tumefaciens-mediated gene transfer would allow the development of RNA interference technology in P. microcarpus.

El hongo ectomicorrícico modelo Pisolithus microcarpus aislamiento 441 fue transformado utilizando Agrobacterium tumefaciens LBA 1100 y AGL-1. El marcador de selección fue el gen Shble de Streptoallotecius hidustanus, el cual confiere resistencia a fleomicina, bajo el control del promotor y terminador del gen gpd de Schizophyllum commune. La transformación resultó en clones resistentes a fleomicina comprobándose por PCR la presencia del transgen. La transferencia génica mediada por Agrobacterium podría permitir el desarrollo de la tecnología de interferencia por ARN en P. microcarpus.

The Shiga toxin 2 B subunit inhibits net fluid absorption in human colon and elicits fluid accumulation in rat colon loops

Shiga toxin (Stx)-producing Escherichia coli (STEC) colonizes the large intestine causing a spectrum of disorders, including watery diarrhea, bloody diarrhea (hemorrhagic colitis), and hemolytic-uremic syndrome. It is estimated that hemolytic-uremic syndrome is the most common cause of acute renal failure in infants in Argentina. Stx is a multimeric toxin composed of one A subunit and five B subunits. In this study we demonstrate that the Stx2 B subunit inhibits the water absorption (Jw) across the human and rat colonic mucosa without altering the electrical parameters measured as transepithelial potential difference and short circuit current. The time-course Jw inhibition by 400 ng/ml purified Stx2 B subunit was similar to that obtained using 12 ng/ml Stx2 holotoxin suggesting that both, A and B subunits of Stx2 contributed to inhibit the Jw. Moreover, non-hemorrhagic fluid accumulation was observed in rat colon loops after 16 h of treatment with 3 and 30 ng/ml Stx2 B subunit. These changes indicate that Stx2 B subunit induces fluid accumulation independently of A subunit activity by altering the usual balance of intestinal absorption and secretion toward net secretion. In conclusion, our results suggest that the Stx2 B subunit, which is non-toxic for Vero cells, may contribute to the watery diarrhea observed in STEC infection. Further studies will be necessary to determine whether the toxicity of Stx2 B subunit may have pathogenic consequences when it is used as a component in an acellular STEC vaccine or as a vector in cancer vaccines.

Peptide and protein modulation of local Ca2+ release events in permeabilized skeletal muscle fibers

Local discrete elevations in myoplasmic Ca2+ (Ca2+ sparks) arise from the opening of a small group of RyRs. Summation of a large number of Ca2+ sparks gives rise to the whole cell Ca2+ transient necessary for muscle contraction. Unlike sarcoplasmic reticulum vesicle preparations and isolated single channels in artificial membranes, the study of Ca2+ sparks provides a means to understand the regulation of a small group of RyRs in the environment of a functionally intact triad and in the presence of endogenous regulatory proteins. To gain insight into the mechanisms that regulate the gating of RyRs we have utilized laser scanning confocal microscopy to measure Ca2+ sparks in permeabilized frog skeletal muscle fibers. This review summarizes our recent studies using both exogenous (ImperatoxinA and domain peptides) and endogenous (calmodulin) modulators of RyR to gain insight into the number of RyR Ca2+ release channels underlying a Ca2+ spark, how domain-domain interactions within RyR regulate the functional state of the channel as well as gating mechanisms of RyR in living muscle fibers.