Genetic analysis for carrier diagnosis in hemophilia A and B in the Mexican population: 25 years of experience.

Our 25 years of experience in carrier diagnosis of hemophilia A (HA) and B (HB) in Mexican population comprises linkage analysis of intragenic F8/F9 neutral variants along with, in severe HA (SHA), detection of F8 int22h and int1h inversions. In symptomatic carriers (SCs) we explored Lyonization to explain their symtomatology. From a DNA-Bank of 3,000 samples, intragenic restriction fragment length (RFLPs) and short tandem repeats (STRs) of F8/F9 genes were assessed by PCR-PAGE and GeneScan. In SHA patients, F8 inversions were detected by inverse shifting-PCR/diagnostic and complementary tests. In SCs, we evaluated hemorrhagic symptoms, clotting FVIII/FIX and X-chromosome inactivation (XCI) patterns were assessed by HUMARA assay and the search of XIST promoter pathogenic variants. Informativeness of linkage analysis for HA carrier diagnosis with RFLP's/STR's increased to 74% and reached 80% with five RFLPs for HB. Combined Inv22/Inv1 diagnosed 113 possible carriers, three de novo Inv22-1, and confirmed 45 mothers as obligate or sporadic carriers. Among 21 SCs, four showed extreme skewed XCI pattern (~80:20) but had normal karyotype and no C43G pathogenic variant in XIST promoter. Clotting FVIII/FIX correlated with the active X in leukocytes. Our data integrate the largest comprehensive research worldwide on the molecular diagnosis of HA and HB carriers in terms of the number of studied and diagnosed cases, in addition to the genetic analysis in SCs. Intragenic RFLPs and STRs of F8/F9 genes along with F8 int22h/int1h inversions in SHA emerge as optimal variants for molecular diagnosis in Mexican population. In counseling SCs, inheritance of skewed X-inactivation should be considered.

The Chemical Transformation of the Cellular Toxin INT (2-(4-Iodophenyl)-3-(4-Nitrophenyl)-5-(Phenyl) Tetrazolium Chloride) as an Indicator of Prior Respiratory Activity in Aquatic Bacteria.

In the ocean, the prokaryote respiration rates dominate the oxidation of organics, but the measurements may be biased due to pre-incubation size filtration and long incubation times. To overcome these difficulties, proxies for microbial respiration rates have been proposed, such as the in vitro and in vivo estimation of electron transport system rates (ETS) based on the reduction of tetrazolium salts. INT (2-(4-Iodophenyl)-3-(4-Nitrophenyl)-5-(Phenyl) Tetrazolium Chloride) is the most commonly applied tetrazolium salt, although it is toxic on time scales of less than 1 h for prokaryotes. This toxicity invalidates the interpretation of the rate of in vivo INT reduction to formazan as a proxy for oxygen consumption rates. We found that with aquatic bacteria, the amount of reduced INT (F; µmol/L formazan) showed excellent relation with the respiration rates prior to INT addition (R; O2 µmol/L/hr), using samples of natural marine microbial communities and cultures of bacteria (V. harveyi) in batch and continuous cultures. We are here relating a physiological rate with the reductive potential of the poisoned cell with units of concentration. The respiration rate in cultures is well related to the cellular potential of microbial cells to reduce INT, despite the state of intoxication.

Intubación nasotraqueal con ‘videolaringoscopio artesanal’ en paciente con vía aérea dificultosa prevista

RESUMEN El objetivo de este trabajo fue analizar un caso de vía aérea dificultosa prevista que requirió intubación nasotraqueal (INT) para cirugía maxilofacial, y se resolvió con un laringoscopio convencional modificado mediante el agregado de una mini cámara. Paciente de sexo femenino de 68 años, coordinada para osteosíntesis de maxilar inferior y órbita con INT. Antecedentes: neoplasma de mama operado. Fumadora intensa. Traumatismo encéfalocraneano 3 semanas antes, con fractura de peñasco y sin lesiones parenquimatosas. Parálisis facial periférica secundaria al trauma. Del examen se destaca: desviación de rasgos a izquierda, parálisis de Bell, apertura bucal disminuida (2,5 cm), Mallampati IV, flexo-extensión de cuello limitada, distancias conservadas. Se preparó la narina derecha con solución de adrenalina aplicada con torundas. Premedicación con Fentanil 2 mg/kg, preoxigenación al 100%, inducción con propofol. Se comprobó una buena permeabilidad de la narina derecha digitalmente, por lo que se pasó una sonda endotraqueal (SET) 7.0 hasta atravesar los cornetes; laringoscopía con pala curva número 3 preparada con una cámara de 7 mm (7 mm USB Endoscope, Welsky Technologies Limited) pegada en el extremo distal, conectada a una notebook en la cual se visualizaron las estructuras. No se administró relajante muscular hasta lograr la intubación. Al visualizar las cuerdas vocales en la pantalla, se progresó la SET en la vía aérea sin necesidad de utilizar pinza de Magill. Conclusión: el laringoscopio modificado con el agregado de una cámara y bajo visión en una pantalla fue efectivo para realizar la INT y resolver esta situación de vía aérea dificultosa prevista.

SUMMARY The objective of this work was to analyze a case of expected difficult airway that required nasotracheal intubation (INT) for maxillofacial surgery, which was resolved with a traditional laryngoscope modified by the addition of a mini-camera. Female patient, 68 years of age, scheduled for osteosynthesis of jaw and orbit with INT. History: operation of breast cancer. Heavy smoker. Traumatic brain injury 3 weeks before, with petrous bone fracture and no parenchymal injuries. Peripheral facial paralysis secondary to trauma. Examination shows: features deviation to the left, Bell's palsy, reduced mouth opening (2.5 cm), Mallampati IV, limited flexion and extension of the neck, distances are preserved. Right nostril was prepared with adrenaline solution applied with swabs. Premedication with Fentanyl 2 g/kg, preoxygenation at 100%, induction with propofol. Good permeability of right nostril was confirmed by digital examination, allowing to insert a 7.0 endotracheal tube (ETT) until passing through the turbinate bones; laryngoscopy with #3 curved blade equipped with a 7 mm camera (7 mm USB Endoscope, Welsky Technologies Limited) at the distal tip, connected to a notebook wherein structures were visualized. No muscle relaxant was administered until intubation was completed. ETT was introduced after visualizing the vocal cords on the screen; Magill forceps were not used. Conclusion: the laryngoscope modified by the addition of a camera for visualization on a monitor screen was effective to carry out INT and manage this expected difficult airway situation.

RESUMO O objetivo deste trabalho foi analisar um caso de previsão de via aérea difícil que requereu intubacão nasotraqueal (INT) para cirurgia maxilofacial, que foi solucionado com um laringoscópio convencional modificado mediante o acoplamento de uma câmera. Paciente do sexo feminino de 68 anos, marcada para realizar osteossintese do maxilar inferior e orbita com INT. Antecedentes. Operada de neoplasia de mama. Tabagista intensa. Traumatismo encefalocrãniano há 3 semanas, com fratura de rochedo e sem lesões parenquimatosas. Paralisia facial periférica secundaria ao trauma. Do exame destaca-se : desvio de simetria facial a esquerda, paralisia de Bell, abertura bucal diminuída (2.5 cm) Mallampatti IV, flexo-extensão do pescoço limitada, distancias conservadas. Preparou-se a narina direita com solução de adrenalina aplicada em torundas. Premedicação com Fentanil 2 µg /kg, pré-oxigenacão a 100%, indução com Propofol. Comprovou-se boa permeabilidade da narina direita digitalmente, pela qual se introduziu uma sonda endotraqueal (SET) 7.0 até atravessar os cornetos , laringoscopia com lamina curva numero 3 preparada com uma câmera de 7 mm (7 mm USB Endoscope , Welsky Technologies Limited) acoplada ao extremo distal, conectada a um notebook no qual se visualizaram as estruturas. Não se administrou relaxante muscular ate que houve-se sucesso na intubação. Ao visualizar as cordas vocais na tela, se progrediu a SET na via aérea sem necessidade de utilização de pinça de Magill. Conclusão: o laringoscópio modificado com uma câmera acoplada e visualização numa tela foi efetivo para realizar a INT e resolver essa situação de previsão de via aérea difícil.

Aplicación de slles de tetrazolio de nueva geneación (xtt) para la estimación de la densidad de microorganismos degradadores de hidrocarburos empleando la técnica del número más poobable / Application of the New Generation Tetrazolium Salt (XTT) for the Enumeration of Hydrocarbon Degrading Microorganisms Using the Most Probable Number Method

El presente estudio evaluó el desempeño de dos sales de tetrazolio, una tradicional: INT y una de nueva generación: XTT, para estimar la densidad de microorganismos degradadores de hidrocarburos (HCs) en suelos empleando la técnica del Número Más Probable (NMP). Se analizaron 96 muestras de suelo provenientes de la Ecorregión Cafetera de Colombia. Los microorganismos fueron recuperados en agar mínimo de sales en atmósfera saturada de HCs y la capacidad degradadora fue confirmada por repiques sucesivos utilizando diesel como fuente de carbono. No se observaron diferencias significativas en los recuentos de microorganismos degradadores obtenidos con las dos sales (t de Student, p < 0,05), pero el XTT permitió mejor visualización de los pozos positivos dada la solubilidad del producto reducido, mientras que el INT produjo precipitación, debido al formazán insoluble generado, dificultando su lectura. Se obtuvo un mayor porcentaje de aislamientos empleando XTT (67%), lo cual podría indicar que el tipo de sal es determinante en la viabilidad de estas bacterias. Adicionalmente, se evaluó el límite de detección celular, las condiciones óptimas de concentración de XTT y el tiempo de incubación necesario para la detección de actividad degradadora utilizando la cepa Acinetobacter sp. El aumento en la concentración de XTT de 0,5 mM a 2 mM y el tiempo de incubación tuvieron un efecto inhibitorio y favorable respectivamente, en la recuperación de células viables, adicionalmente, límite de detección de la técnica fue de 10² UFC/ml.

The objective of this study was to evaluate the performance of two tetrazolium indicators: a traditional one: INT and a new generation one: XTT, for the estimation of hydrocarbon (HC) degrading microorganism s density using the Most Probable Number Technique (MPN). Ninety six composite soil samples were taken and analyzed from Ecorregión Cafetera Colombiana. Degrading microorganisms were recovered in minimum salt medium with saturated HC atmosphere. Degrading HC capacity of the microorganisms was confirmed by successive subcultures in the same medium using diesel as only carbon source. Counts obtained with the two salts were not significantly different (Student t test, p < 0,05) but XTT allowed an easier visualization of positive wells due to product solubility of the reduce product. A greater percentage of isolates was obtained using XTT (67%), which suggests that salt type is relevant for recovering of these microorganisms. Additionally, cell detection limit, optimal conditions of XTT concentration and incubation times for detection of activity were evaluated. This evaluation was performed by means of microplate format for hydrocarbon degrading microorganisms using Acinetobacter sp. An inhibitory effect was observed in the recovering of cultivable cells when XTT concentrations increased from 0,5 mM to 2 mM. Incubation time favored this recovering. Detection limit of this technique was established at 10² UFC/ml. Production of the XTT-formazan was positively related with initial cell concentration and negatively with incubation time.