Análisis molecular de sistemas génicos implicados en la homeostasis de níquel y eficiencia simbiótica en Rhizobium leguminosarum

Los suelos ultramáficos, que poseen elevadas concentraciones de níquel, cobalto y cromo de manera natural, son fuente de bacterias resistentes a altas concentraciones de metales. Se realizó la caracterización físico-química de seis suelos ultramáficos del suroeste europeo, seleccionándose un suelo de la región de Gorro, Italia, como el más adecuado para aislar bacterias endosimbióticas resistentes a metales. A partir de plantas-trampa de guisante y lenteja inoculados con suspensiones de ese suelo, se obtuvieron 58 aislados de Rhizobium leguminosarum bv. viciae (Rlv) que fueron clasificados en 13 grupos según análisis de PCR-RAPDs. Se determinó la resistencia a cationes metálicos [Ni(II), Co(II), Cu(II), Zn(II)] de una cepa representante de cada grupo, así como la secuencia de los genomas de las cepas que mostraron altos niveles (UPM1137 y UPM1280) y bajos niveles (UPM1131 y UPM1136) de tolerancia a metales. Para identificar mecanismos de resistencia a metales se realizó una mutagénesis al azar en dicha cepa mediante la inserción de un minitransposón. El análisis de 4313 transconjugantes permitió identificar 14 mutantes que mostraron una mayor sensibilidad a Ni(II) que la cepa silvestre. Se determinó el punto de inserción del minitransposón en todos ellos y se analizaron en más detalle dos de los mutantes (D2250 y D4239). En uno de los mutantes (D2250), el gen afectado codifica para una proteína que presenta un 44% de identidad con dmeF (divalent efflux protein) de Cupriavidus metallidurans. Cadena arriba de dmeF se identificó un gen que codifica una proteína con un 39% de identidad con el regulador RcnR de Escherichia coli. Se decidió nombrar a este sistema dmeRF, y se generó un mutante en ambos genes en la cepa Rlv SPF25 (Rlv D15). A partir de experimentos de análisis fenotípico y de regulación se pudo demostrar que el sistema dmeRF tiene un papel relevante en la resistencia a Ni(II) y sobre todo a Co(II) en células en vida libre y en simbiosis con plantas de guisante. Ambos genes forman un operón cuya expresión se induce en respuesta a la presencia de Ni(II) y Co(II). Este sistema se encuentra conservado en distintas especies del género Rhizobium como un mecanismo general de resistencia a níquel y cobalto. Otro de los mutantes identificados (D4239), tiene interrumpido un gen que codifica para un regulador transcripcional de la familia AraC. Aunque inicialmente fue identificado por su sensibilidad a níquel, experimentos posteriores demostraron que su elevada sensibilidad a metales era debida a su sensibilidad al medio TY, y más concretamente a la triptona presente en el medio. En otros medios de cultivo el mutante no está afectado específicamente en su tolerancia a metales. Este mutante presenta un fenotipo simbiótico inusual, siendo inefectivo en guisantes y efectivo en lentejas. Análisis de complementación y de mutagénesis dirigida sugieren que el fenotipo de la mutación podría depender de otros factores distintos del gen portador de la inserción del minitransposón. ABSTRACT Ultramafic soils, having naturally high concentrations of nickel, cobalt and chrome, are potential sources of highly metal-resistant bacteria. A physico-chemical characterization of six ultramafic soils from the European southwest was made. A soil from Gorro, Italy, was chosen as the most appropriated for the isolation of heavy-metal-resistant endosymbiotic bacteria. From pea and lentil trap plants inoculated with soil suspensions, 58 isolates of Rhizobium leguminosarum bv. viciae (Rlv) were obtained and classified into 13 groups based on PCR-RAPDs analysis. The resistance to metallic cations [Ni(II), Co(II), Cu(II), Zn(II)] was analyzed in a representative strain of each group. From the results obtained in the resistance assays, the Rlv UPM1137 strain was selected to identify metal resistance mechanism. A random mutagenesis was made in UPM1137 by using minitransposon insertion. Analysis of 4313 transconjugants allowed to identify 14 mutants with higher sensitivity to Ni(II) than the wild type strain. The insertion point of the minitransposon was determined in all of them, and two mutants (D2250 and D4239) were studied in more detail. In one of the mutants (D2250), the affected gene encodes a protein with 44% identity in compared with DmeF (divalent efflux protein) from Cupriavidus metallidurans. Upstream R. leguminosarum dmeF, a gene encoding a protein with 39% identity with RcnR regulator from E. coli was identified. This protein was named DmeR. A mutant with both genes in the dmeRF deleted was generated and characterized in Rlv SPF25 (Rlv D15). From phenotypic and regulation analysis it was concluded that the dmeRF system is relevant for Ni(II) and specially Co(II) tolerance in both free living and symbiotic forms of the bacteria. This system is conserved in different Rhizobium species like a general mechanism for nickel and cobalt resistance. Other of the identified mutants (D4239) contains the transposon insert on a gene that encodes for an AraC-like transcriptional regulator. Although initially this mutant was identified for its nickel sensitivity, futher experiments demonstrated that its high metal sensitivity is due to its sensitivity to the TY medium, specifically for the tryptone. In other media the mutant is not affected specifically in their tolerance to metals. This mutant showed an unusual symbiotic phenotype, being ineffective in pea and effective in lentil. Complementation analysis and directed mutagenesis suggest that the mutation phenotype could depend of other factors different from the insertion minitransposon gene.

Implementación de sistema domótico con servidor Raspberry

Este proyecto consiste en el diseño e implementación un sistema domótico que puede ser instalado en una vivienda para controlar distintas variables ambientales y conseguir así la máxima comodidad de los habitantes de manera automática o manual según los gustos y necesidades de los usuarios. La característica principal de este sistema, es que cuenta con un funcionamiento distribuido donde entran en juego un servidor, encargado de tomar las decisiones generales para el comportamiento de la casa, y una serie de controladores esclavo cuya función es mantener constantes las variables ambientales con los valores fijados por el servidor. Así se consigue mantener la vivienda en una situación de bienestar constante para cualquier persona que se encuentre dentro. El sistema ha sido pensado de manera que se intenta reducir al máximo el cableado para facilitar su instalación por lo que la comunicación entre los distintos dispositivos se hace de manera inalámbrica por medio de un protocolo descrito en la norma IEEE 802.15.4 llamado ZigBee. Para ello se ha utilizado un módulo de comunicación wireless llamado Xbee, el cual permite la comunicación entre dos dispositivos. Para el control de dicho sistema distribuido se cuenta con una aplicación web, que mediante una interfaz gráfica permite al usuario controlar los distintos dispositivos dentro de la vivienda consiguiendo así controlar las variables ambientales a gusto del usuario. Dicha interfaz gráfica no depende de un software específico, sino que sólo es necesario un cliente http como podría ser Internet Explorer, Mozilla Firefox, Google Chrome, etc. Para integrar dicho sistema se ha usado un mini ordenador de bajo coste llamado RaspBerryPi, en el que se encuentra alojado un servidor Apache con el fin de gestionar y automatizar las variables ambientales. El control de los dipositivos encargados de modificar y estabilizar las variables ambientales se realiza mediante unos controladores genéricos implementados mediante mcontroladores 80C51F410, pertenecientes a la familia 80C51, y una serie de componentes y circuitería que permiten el correcto funcionamiento de éstos. Existen dos tipos de controladores distintos, los cuales son: Controlador Sensor: Encargados de las tomas de valores ambientales como puede ser la luz y la temperatura. Controladores Actuadores: Encargados de actuar sobre los dispositivos que modifican y estabilizan las variables ambientales como pueden ser la calefacción, tiras de leds de iluminación, persianas, alarmas, etc. El conjunto de la RaspBerryPi y los diferentes controladores forman el prototipo diseñado para este proyecto fin de carrera, el cual puede ser ampliado sencillamente para abarcar una amplia gama de posibilidades y funcionalidades dentro de la comodidad de una vivienda. ABSTRACT. The project described in this report consisted designing and implementing a home automation system that could be installed in a house in order to control environmental variables and thus get the maximum comfort of the inhabitant automatically or manually according to their tastes and needs. The main feature of this system consists in a distributed system, formed by a server which is responsible for making the main decisions of the actions performed inside the house. In addition, there are a series of slave controlers whose function consists in keeping the environmental variables within the values established by the server. Thus gets to keep the home in a situation of constant wellbeing to anyone who is inside. The system has been designed in order to reduce the amount of wire needed for the inter-connection of the devices, by means of wireless communication. The devices chosen for the solution are Xbee modules, which use the Zigbee protocol in order to comunicate one between each other. The Zigbee protocol is fully described in the IEEE 802.15.4 standard. A web application has been used to control the distributed system. This application allows users to control various devices inside the house and subsequently the different environmental variables. This implementation allows obtaining the maximum comfort by means of a very simple graphical interface. In addition, the Graphical User Interface (GUI) does not depend on any specific software. This means that it would only be necessary a http client (such as Internet Explorer, Mozilla Firefox, Google Chrome, etc.) for handling the application. The system has been integrated using a low-cost mini computer called RaspBerryPi.This computer has an Apache server allocated which allows to manage and to automatize the different environmental variables. Furthermore, for changing and stabilizing those variables, some generic controllers have been developed, based on mcontrollers 80C51F410. There have been developed mainly two different types of controllers: Sensor Controllers, responsible for measuring the different environmental values, such as light and temperature; and Actuator Controllers, which purpose is to modify and stabilize those environmental variables by actuating on the heating, the led lamps, the blinders, the alarm, etc. The combination of the RaspBerryPi and the different controllers conform the prototype designed during this project. Additionally, this solution could be easily expanded in order to intake further functionalities adapted to new needs that could arise in the future.

Modes of occurrence and geochemistry at DSDP Holes 69-505A and 69-505B

The compositions of chrome spinels of Costa Rica Rift basalts from Deep Sea Drilling Project Site 505 vary depending on their occurrences as (1) inclusions in olivine crystals, (2) inclusions in Plagioclase crystals, and (3) isolated crystals in variolitic or glassy samples. The variations are a consequence of (1) changes of melt compositions as crystallization proceeds, and (2) contrasting behavior of olivine and Plagioclase in competition with spinels for Al and Mg. Some spinels have skeletal rims compositionally less magnesian than mineral cores; however, the cores do not appear to be xenocrysts, unlike some texturally similar spinels in Mid-Atlantic Ridge basalts.

Geochemistry at DSDP Sites 69-505, 69-504 and 70-504

Deep basement penetration during Legs 69 and 70 at Hole 504B in the Panama Basin allowed the recovery of a 561.5-meter sequence of basaltic pillows, thin flows, and breccias interspersed with thick massive flows. The lavas, which are aphyric to moderately plagioclase-olivine-clinopyroxene phyric, are petrologically indistinguishable from typical mid-ocean-ridge basalts (MORB). Some units are distinctive in that they carry accessory chrome-spinel microphenocrysts or emerald green clinopyroxene phenocrysts. Major and trace element analyses were carried out on 67 samples using X-ray fluorescence techniques. The basalts resemble normal MORB in terms of major elements. However, the trace element analyses show that most of the basalts are characterized by very strong depletion in the more incompatible elements compared with, for instance, normal (N type) MORB from the Atlantic at 22°N. Interdigitated with these units are one or two units that have distinctly higher incompatible element concentrations similar to those in basalts of the transitional (T) type from the Reykjanes Ridge (63°N in the Mid-Atlantic Ridge). All the basalts appear to have undergone some high-level crystal fractionation, although this has not proceeded to the extent of yielding ferrobasalts as it has at the adjacent Galapagos Spreading Center or along the East Pacific Rise. The magnetic anomalies are of lower amplitude than in the latter two regions, which suggests that the absence of ferrobasalts may be a general feature of the ocean crust generated at the Costa Rica Rift. The presence of two distinct magma types, one strongly depleted and the other moderately enriched in incompatible elements, suggests that magma chambers at the spreading center are discontinuous rather than continuous and that there is some chemical heterogeneity in the underlying mantle source. Observed variations in incompatible element ratios of basalts from the more depleted group could, however, reflect mixing between these two magma types. In general it would appear that the mantle feeding the Costa Rica Rift is significantly more depleted in incompatible trace elements than that feeding the Mid-Atlantic Ridge.

The manufacture of leather /

Disbound Original Held in Oak Street Library Facility.

Report on elevated temperature properties of chromium steels (12-17 per cent) Data compiled by and issued under the auspices of the Data and Publications Panel of the ASTM-ASME Joint Committee on Effect of Temperature on the Properties of Metals.

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