A step to the gigantic genome of the desert locust: chromosome sizes and repeated DNAs

The desert locust (Schistocerca gregaria) has been used as material for numerous cytogenetic studies. Its genome size is estimated to be 8.55 Gb of DNA comprised in 11 autosomes and the X chromosome. Its X0/XX sex chromosome determinism therefore results in females having 24 chromosomes whereas males have 23. Surprisingly, little is known about the DNA content of this locust's huge chromosomes. Here, we use the Feulgen Image Analysis Densitometry and C-banding techniques to respectively estimate the DNA quantity and heterochromatin content of each chromosome. We also identify three satellite DNAs using both restriction endonucleases and next-generation sequencing. We then use fluorescent in situ hybridization to determine the chromosomal location of these satellite DNAs as well as that of six tandem repeat DNA gene families. The combination of the results obtained in this work allows distinguishing between the different chromosomes not only by size, but also by the kind of repetitive DNAs that they contain. The recent publication of the draft genome of the migratory locust (Locusta migratoria), the largest animal genome hitherto sequenced, invites for sequencing even larger genomes. S. gregaria is a pest that causes high economic losses. It is thus among the primary candidates for genome sequencing. But this species genome is about 50 % larger than that of L. migratoria, and although next-generation sequencing currently allows sequencing large genomes, sequencing it would mean a greater challenge. The chromosome sizes and markers provided here should not only help planning the sequencing project and guide the assembly but would also facilitate assigning assembled linkage groups to actual chromosomes.

Phosphorus fractions in soils of the mangrove, restinga and Atlantic forest ecosystems from Cardoso Island, Brazil

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Theses, Projects, and Africana Materials Management in Nigerian Academic Libraries: The Case of Hezekiah Oluwasanmi Library

This is a survey of the procurement, organization and use of unpublished projects, theses, and Africana mateials in Hezekiah Oluwasanmi Library, Obafemi Awolowo University, Ile-Ife, Nigeria. The Africana section conserves knowledge, preserves cultural heritage, provides information, and supports education and research. This paper the location, mode of processing, circulation, and terms of availability of these materials. Recommendations are made on how to manage Africana materials in academic libraries where they constitute a vital component of collections.

Comparative Genomics of Early-Diverging Brucella Strains Reveals a Novel Lipopolysaccharide Biosynthesis Pathway

Brucella species are Gram-negative bacteria that infect mammals. Recently, two unusual strains (Brucella inopinata BO1(T) and B. inopinata-like BO2) have been isolated from human patients, and their similarity to some atypical brucellae isolated from Australian native rodent species was noted. Here we present a phylogenomic analysis of the draft genome sequences of BO1(T) and BO2 and of the Australian rodent strains 83-13 and NF2653 that shows that they form two groups well separated from the other sequenced Brucella spp. Several important differences were noted. Both BO1(T) and BO2 did not agglutinate significantly when live or inactivated cells were exposed to monospecific A and Mantisera against O-side chain sugars composed of N-formyl-perosamine. While BO1(T) maintained the genes required to synthesize a typical Brucella O-antigen, BO2 lacked many of these genes but still produced a smooth LPS (lipopolysaccharide). Most missing genes were found in the wbk region involved in O-antigen synthesis in classic smooth Brucella spp. In their place, BO2 carries four genes that other bacteria use for making a rhamnose-based O-antigen. Electrophoretic, immunoblot, and chemical analyses showed that BO2 carries an antigenically different O-antigen made of repeating hexose-rich oligosaccharide units that made the LPS water-soluble, which contrasts with the homopolymeric O-antigen of other smooth brucellae that have a phenol-soluble LPS. The results demonstrate the existence of a group of early-diverging brucellae with traits that depart significantly from those of the Brucella species described thus far. IMPORTANCE This report examines differences between genomes from four new Brucella strains and those from the classic Brucella spp. Our results show that the four new strains are outliers with respect to the previously known Brucella strains and yet are part of the genus, forming two new clades. The analysis revealed important information about the evolution and survival mechanisms of Brucella species, helping reshape our knowledge of this important zoonotic pathogen. One discovery of special importance is that one of the strains, BO2, produces an O-antigen distinct from any that has been seen in any other Brucella isolates to date.

Architectures and design patterns for functional design of logic control and diagnostics in industrial automation

Recently in most of the industrial automation process an ever increasing degree of automation has been observed. This increasing is motivated by the higher requirement of systems with great performance in terms of quality of products/services generated, productivity, efficiency and low costs in the design, realization and maintenance. This trend in the growth of complex automation systems is rapidly spreading over automated manufacturing systems (AMS), where the integration of the mechanical and electronic technology, typical of the Mechatronics, is merging with other technologies such as Informatics and the communication networks. An AMS is a very complex system that can be thought constituted by a set of flexible working stations, one or more transportation systems. To understand how this machine are important in our society let considerate that every day most of us use bottles of water or soda, buy product in box like food or cigarets and so on. Another important consideration from its complexity derive from the fact that the the consortium of machine producers has estimated around 350 types of manufacturing machine. A large number of manufacturing machine industry are presented in Italy and notably packaging machine industry,in particular a great concentration of this kind of industry is located in Bologna area; for this reason the Bologna area is called “packaging valley”. Usually, the various parts of the AMS interact among them in a concurrent and asynchronous way, and coordinate the parts of the machine to obtain a desiderated overall behaviour is an hard task. Often, this is the case in large scale systems, organized in a modular and distributed manner. Even if the success of a modern AMS from a functional and behavioural point of view is still to attribute to the design choices operated in the definition of the mechanical structure and electrical electronic architecture, the system that governs the control of the plant is becoming crucial, because of the large number of duties associated to it. Apart from the activity inherent to the automation of themachine cycles, the supervisory system is called to perform other main functions such as: emulating the behaviour of traditional mechanical members thus allowing a drastic constructive simplification of the machine and a crucial functional flexibility; dynamically adapting the control strategies according to the different productive needs and to the different operational scenarios; obtaining a high quality of the final product through the verification of the correctness of the processing; addressing the operator devoted to themachine to promptly and carefully take the actions devoted to establish or restore the optimal operating conditions; managing in real time information on diagnostics, as a support of the maintenance operations of the machine. The kind of facilities that designers can directly find on themarket, in terms of software component libraries provides in fact an adequate support as regard the implementation of either top-level or bottom-level functionalities, typically pertaining to the domains of user-friendly HMIs, closed-loop regulation and motion control, fieldbus-based interconnection of remote smart devices. What is still lacking is a reference framework comprising a comprehensive set of highly reusable logic control components that, focussing on the cross-cutting functionalities characterizing the automation domain, may help the designers in the process of modelling and structuring their applications according to the specific needs. Historically, the design and verification process for complex automated industrial systems is performed in empirical way, without a clear distinction between functional and technological-implementation concepts and without a systematic method to organically deal with the complete system. Traditionally, in the field of analog and digital control design and verification through formal and simulation tools have been adopted since a long time ago, at least for multivariable and/or nonlinear controllers for complex time-driven dynamics as in the fields of vehicles, aircrafts, robots, electric drives and complex power electronics equipments. Moving to the field of logic control, typical for industrial manufacturing automation, the design and verification process is approached in a completely different way, usually very “unstructured”. No clear distinction between functions and implementations, between functional architectures and technological architectures and platforms is considered. Probably this difference is due to the different “dynamical framework”of logic control with respect to analog/digital control. As a matter of facts, in logic control discrete-events dynamics replace time-driven dynamics; hence most of the formal and mathematical tools of analog/digital control cannot be directly migrated to logic control to enlighten the distinction between functions and implementations. In addition, in the common view of application technicians, logic control design is strictly connected to the adopted implementation technology (relays in the past, software nowadays), leading again to a deep confusion among functional view and technological view. In Industrial automation software engineering, concepts as modularity, encapsulation, composability and reusability are strongly emphasized and profitably realized in the so-calledobject-oriented methodologies. Industrial automation is receiving lately this approach, as testified by some IEC standards IEC 611313, IEC 61499 which have been considered in commercial products only recently. On the other hand, in the scientific and technical literature many contributions have been already proposed to establish a suitable modelling framework for industrial automation. During last years it was possible to note a considerable growth in the exploitation of innovative concepts and technologies from ICT world in industrial automation systems. For what concerns the logic control design, Model Based Design (MBD) is being imported in industrial automation from software engineering field. Another key-point in industrial automated systems is the growth of requirements in terms of availability, reliability and safety for technological systems. In other words, the control system should not only deal with the nominal behaviour, but should also deal with other important duties, such as diagnosis and faults isolations, recovery and safety management. Indeed, together with high performance, in complex systems fault occurrences increase. This is a consequence of the fact that, as it typically occurs in reliable mechatronic systems, in complex systems such as AMS, together with reliable mechanical elements, an increasing number of electronic devices are also present, that are more vulnerable by their own nature. The diagnosis problem and the faults isolation in a generic dynamical system consists in the design of an elaboration unit that, appropriately processing the inputs and outputs of the dynamical system, is also capable of detecting incipient faults on the plant devices, reconfiguring the control system so as to guarantee satisfactory performance. The designer should be able to formally verify the product, certifying that, in its final implementation, it will perform itsrequired function guarantying the desired level of reliability and safety; the next step is that of preventing faults and eventually reconfiguring the control system so that faults are tolerated. On this topic an important improvement to formal verification of logic control, fault diagnosis and fault tolerant control results derive from Discrete Event Systems theory. The aimof this work is to define a design pattern and a control architecture to help the designer of control logic in industrial automated systems. The work starts with a brief discussion on main characteristics and description of industrial automated systems on Chapter 1. In Chapter 2 a survey on the state of the software engineering paradigm applied to industrial automation is discussed. Chapter 3 presentes a architecture for industrial automated systems based on the new concept of Generalized Actuator showing its benefits, while in Chapter 4 this architecture is refined using a novel entity, the Generalized Device in order to have a better reusability and modularity of the control logic. In Chapter 5 a new approach will be present based on Discrete Event Systems for the problemof software formal verification and an active fault tolerant control architecture using online diagnostic. Finally conclusive remarks and some ideas on new directions to explore are given. In Appendix A are briefly reported some concepts and results about Discrete Event Systems which should help the reader in understanding some crucial points in chapter 5; while in Appendix B an overview on the experimental testbed of the Laboratory of Automation of University of Bologna, is reported to validated the approach presented in chapter 3, chapter 4 and chapter 5. In Appendix C some components model used in chapter 5 for formal verification are reported.

Modeling the spatial dynamics of economic models

The advances that have been characterizing spatial econometrics in recent years are mostly theoretical and have not found an extensive empirical application yet. In this work we aim at supplying a review of the main tools of spatial econometrics and to show an empirical application for one of the most recently introduced estimators. Despite the numerous alternatives that the econometric theory provides for the treatment of spatial (and spatiotemporal) data, empirical analyses are still limited by the lack of availability of the correspondent routines in statistical and econometric software. Spatiotemporal modeling represents one of the most recent developments in spatial econometric theory and the finite sample properties of the estimators that have been proposed are currently being tested in the literature. We provide a comparison between some estimators (a quasi-maximum likelihood, QML, estimator and some GMM-type estimators) for a fixed effects dynamic panel data model under certain conditions, by means of a Monte Carlo simulation analysis. We focus on different settings, which are characterized either by fully stable or quasi-unit root series. We also investigate the extent of the bias that is caused by a non-spatial estimation of a model when the data are characterized by different degrees of spatial dependence. Finally, we provide an empirical application of a QML estimator for a time-space dynamic model which includes a temporal, a spatial and a spatiotemporal lag of the dependent variable. This is done by choosing a relevant and prolific field of analysis, in which spatial econometrics has only found limited space so far, in order to explore the value-added of considering the spatial dimension of the data. In particular, we study the determinants of cropland value in Midwestern U.S.A. in the years 1971-2009, by taking the present value model (PVM) as the theoretical framework of analysis.

Modelling, simulation and optimization of maintenance considerations on condition based maintenance

Globalization has increased the pressure on organizations and companies to operate in the most efficient and economic way. This tendency promotes that companies concentrate more and more on their core businesses, outsource less profitable departments and services to reduce costs. By contrast to earlier times, companies are highly specialized and have a low real net output ratio. For being able to provide the consumers with the right products, those companies have to collaborate with other suppliers and form large supply chains. An effect of large supply chains is the deficiency of high stocks and stockholding costs. This fact has lead to the rapid spread of Just-in-Time logistic concepts aimed minimizing stock by simultaneous high availability of products. Those concurring goals, minimizing stock by simultaneous high product availability, claim for high availability of the production systems in the way that an incoming order can immediately processed. Besides of design aspects and the quality of the production system, maintenance has a strong impact on production system availability. In the last decades, there has been many attempts to create maintenance models for availability optimization. Most of them concentrated on the availability aspect only without incorporating further aspects as logistics and profitability of the overall system. However, production system operator’s main intention is to optimize the profitability of the production system and not the availability of the production system. Thus, classic models, limited to represent and optimize maintenance strategies under the light of availability, fail. A novel approach, incorporating all financial impacting processes of and around a production system, is needed. The proposed model is subdivided into three parts, maintenance module, production module and connection module. This subdivision provides easy maintainability and simple extendability. Within those modules, all aspect of production process are modeled. Main part of the work lies in the extended maintenance and failure module that offers a representation of different maintenance strategies but also incorporates the effect of over-maintaining and failed maintenance (maintenance induced failures). Order release and seizing of the production system are modeled in the production part. Due to computational power limitation, it was not possible to run the simulation and the optimization with the fully developed production model. Thus, the production model was reduced to a black-box without higher degree of details.

The Contegra bovine jugular vein graft versus the Shelhigh pulmonic porcine graft for reconstruction of the right ventricular outflow tract: a comparative study

Reconstruction of the right ventricular outflow tract plays a major role in congenital cardiac surgery. With the advent of the Contegra bovine jugular vein graft and the Shelhigh pulmonic xenograft, hopes were high that the lack of availability of homografts would be overcome. The present study evaluated both grafts and investigated the influence of known risk factors for premature graft failure.

Can Andean medicine coexist with biomedical healthcare? A comparison of two rural communities in Peru and Bolivia

Background It is commonly assumed that indigenous medical systems remain strong in developing countries because biomedicine is physically inaccessible or financially not affordable. This paper compares the health-seeking behavior of households from rural Andean communities at a Peruvian and a Bolivian study site. The main research question was whether the increased presence of biomedicine led to a displacement of Andean indigenous medical practices or to coexistence of the two healing traditions. Methodology Open-ended interviews and free listing exercises were conducted between June 2006 and December 2008 with 18 households at each study site. Qualitative identification of households’ therapeutic strategies and use of remedies was carried out by means of content analysis of interview transcriptions and inductive interference. Furthermore, a quantitative assessment of the incidence of culture-bound illnesses in local ethnobiological inventories was performed. Results Our findings indicate that the health-seeking behavior of the Andean households in this study is independent of the degree of availability of biomedical facilities in terms of quality of services provided, physical accessibility, and financial affordability, except for specific practices such as childbirth. Preference for natural remedies over pharmaceuticals coexists with biomedical healthcare that is both accessible and affordable. Furthermore, our results show that greater access to biomedicine does not lead to less prevalence of Andean indigenous medical knowledge, as represented by the levels of knowledge about culture-bound illnesses. Conclusions The take-home lesson for health policy-makers from this study is that the main obstacle to use of biomedicine in resource-poor rural areas might not be infrastructural or economic alone. Rather, it may lie in lack of sufficient recognition by biomedical practitioners of the value and importance of indigenous medical systems. We propose that the implementation of health care in indigenous communities be designed as a process of joint development of complementary knowledge and practices from indigenous and biomedical health traditions.

Annotated record of the detailed examination of Mn deposits from the Albatross 1884 stations

he exploration of the Gulf Stream region was continued in 1884, under nearly the same conditions as in 1883, by the steamer Albatross, Lieut. Z. L. Tanner, commander. During the four trips, between July 20 and Sept. 13, sixty nine dredgings (at stations 2170 to 2238) were made. The results were highly satisfactory, both in the way of physical observatidns and zoological discoveries. In some localities, in 1000 to 1600 fathoms, the bottom was found covered with 0or largely composed of hard, very irregular, flattened, crust-like concretions of clay and iron-oxide, with more or less manganese?oxide in the crevices and worm-burrows with which they are filled. Sometimes a barrel-full, or more, of such masses were brought up, varying in size from a few ounces up to 20 pounds or more in weight alld from one inch to six inches in thickness.

Mejora del comportamiento térmico de los morteros de cal aditivados y su empleo en la rehabilitación de inmuebles = Improvement of thermal behaviour of additived lime mortars and their use in rehabilitation of buildings

La envolvente de la edificación es la responsable de equilibrar el intercambio energético entre el interior y el exterior, por lo tanto cualquier actuación encaminada a la reducción del consumo energético ha de establecer, como uno de sus objetivos prioritarios, la mejora del comportamiento de la misma. Las edificaciones anteriores a 1940 constituyen la mayor parte de las existentes en áreas rurales y centros urbanos. En ellas, la repercusión de la fachada sobre las transmitancias globales pone de manifiesto la necesidad de intervención. Sin embargo, su elevada inercia térmica y los importantes saltos térmicos característicos de gran parte de España plantean la importancia de que aquélla se efectúe por el exterior. A tal respecto, la falta de disponibilidad de espesor suficiente para implantar sistemas tipo SATE deriva en que, frecuentemente, la única solución viable sea la de aislar por el interior perdiendo con ello la capacidad de acumulación térmica del muro y con el asociado riesgo de condensaciones. La amplia tradición en el empleo de revestimientos, especialmente en base de cal, permiten que éstos sean utilizados no sólo como elemento estético o de protección de la obra de fábrica antigua sino también para la mejora del comportamiento térmico del soporte, si se aprovecha el mecanismo de transmisión térmica por radiación. Éste es el objetivo de la presente Tesis Doctoral en la que se estudia la modificación de las propiedades radiantes de los morteros de revestimiento para la mejora de la eficiencia energética de las construcciones históricas, principalmente las constituidas por muros monolíticos, aunque podría ser de aplicación a otro tipo de construcciones compuestas por diversas capas. Como punto de partida, se estudió y revisó la documentación disponible sobre las investigaciones de las tres áreas científico-tecnológicas que convergen en la Tesis Doctoral: rehabilitación, material y comportamiento térmico, a partir de lo cual se comprobó la inexistencia de estudios similares al objeto de la presente Tesis Doctoral. Complementariamente, se analizaron los revestimientos en lo concerniente a los materiales que los constituyen, la composición de las mezclas y características de cada una de las capas así como al enfoque que, desde el punto de vista térmico, se estimaba más adecuado para la obtención de los objetivos propuestos. Basándonos en dichos análisis, se preseleccionaron ochenta materiales que fueron ensayados en términos de reflectancia y emisividad para elegir cuatro con los que se continuó la investigación. Éstos, junto con la cal elegida para la investigación y el árido marmóreo característico de la última capa de revestimiento, fueron caracterizados térmicamente, de forma pormenorizada, así como química y físicamente. Los fundamentos teóricos y los estudios preliminares desarrollados con distintos materiales, en estado fresco y endurecido, fueron empleados en la dosificación de componentes de las mezclas, en dos proporciones distintas, para el estudio del efecto del agregado. Éstas se ensayaron en estado fresco, para comprobar su adecuación de puesta en obra y prever su VI adherencia al soporte, así como en estado endurecido a 28 y 90 días de curado, para conocer las propiedades que permitieran prever su compatibilidad con aquél y estimar el alcance de la reducción de transferencias térmicas lograda. Además, se estudiaron las características generales de las mezclas que sirvieron para establecer correlaciones entre distintas propiedades y entender los resultados mecánicos, físicos (comportamiento frente al agua) y energéticos. Del estudio conjunto de las distintas propiedades analizadas se propusieron dos mezclas, una blanca y otra coloreada, cuyas características permiten garantizar la compatibilidad con la obra de fábrica antigua, desde el punto de vista físico y mecánico, y preservar la autenticidad de los revestimientos, en cuanto a la técnica de aplicación tradicional en sistemas multicapa. El comportamiento térmico de las mismas, sobre una obra de fábrica de 40 cm de espesor, se estimó, en estado estacionario y pseudo-transitorio, demostrándose reducciones del flujo térmico entre 16-48%, en condiciones de verano, y entre el 6-11%, en invierno, en función de la coloración y de la rugosidad de la superficie, en comparación con el empleo de la mezcla tradicional. Por lo que, se constata la viabilidad de los materiales compuestos propuestos y su adecuación al objetivo de la investigación. VII ABSTRACT The envelope is responsible for balancing the energy exchange between the inside and outside in buildings. For this reason, any action aimed at reducing energy consumption must establish, as one of its key priorities, its improvement. In rural areas and urban centers, most of the constructions were built before 1940. In them, the impact of the façade on the global transmittance highlights the need for intervention. However, its high thermal inertia and fluctuation of temperatures in the majority of Spain bring up that it should be placed outside the insulation. In this regard, the lack of availability of enough thickness to implement systems such as ETICS results in that often the only viable solution is to isolate the interior, losing thereby the wall’s heat storage capacity with the associated risk of condensation. The tradition in the use of renders, especially lime-based, allows us to use them not only as an aesthetic element or to protect the ancient masonry, but also for improved thermal performance of the support by taking advantage of the heat transfer mechanism by radiation. This is the aim of this Doctoral Thesis in which the modification of the radiative properties of lime mortars for renders to improve the energy efficiency of historic buildings, mainly composed of monolithic walls, is studied, although it could be applied to other structures composed of several layers. As a starting point, the available literature in the three scientific-technological areas that converge at the Doctoral Thesis: rehabilitation, material and thermal behaviour, was reviewed, and confirmed the absence of researches similar to this Doctoral Thesis. Additionally, the renders were studied with regard to the materials that constitute them, the composition of the mixtures and the characteristics of each layer, as well as to the approach which, from a thermal point of view, was deemed the most suitable for achieving the objectives sets. Based on thre aforementioned analysis, eighty materials tested in terms of reflectance and emissivity were pre-selected, to choose four with which the research was continued. Common marble sand, used in the last layer of the renders, together with the appointed materials and hydrated lime were characterized thermally, in detail, as well as chemically and physically. The theoretical bases and preliminary studies with different materials, in fresh and hardened state, were used in the dosage of the composition of the mixtures. In order to study their effect they were used in two different proportions, that is, ten mixtures in total. These were tested in their fresh state to evaluate their setting-up suitability and foresee their adhesion to the support, as well as in their hardened state, at 28 and 90 days of curing, to establish the properties which enabled us to anticipate their compatibility with the old masonry walls and estimate the scope of the reduction of heat transfers achieved. In addition, the general characteristics of the mixtures used to establish correlations and to understand the mechanical, physical and energy results were studied. Two mixtures, one white and one colored, were proposed as the result of the different properties analysed, whose characteristics allow the guarantee of mechanical and physical compatibility VIII with the old masonry and preserve the authenticity of the renders. The thermal behavior of both, applied on a masonry wall 40 cm thick, was estimated at a steady and pseudo-transient state, with heat flow reductions between 16-48% during summertime and 6-11% during wintertime, depending on the color and surface roughness, compared to the use of the traditional mixture. So, the viability of the proposed composite materials and their fitness to the aim of the research are established.

Native bradyrhizobial symbionts of Lupinus mariae-josephae, a unique endemism thriving in alkaline soils in Eastern Spain

Lupinus mariae-josephae (Lmj) es una especie de lupino endémica de una pequeña y específica área de Comunidad Valenciana (Este de España), donde prospera en suelos alcalinoscalcáreos, un hábitat singular para los altramuces, que crecen preferentemente en suelos ácidos o neutros. Esto hace de Lmj una especie de lupino única. Cuando se inició este trabajo, la extensión conocida de este endemismo abarcaba unos 700 kilómetros cuadrados, confinados en la provincia de Valencia. En esta área, Lmj prospera en pequeñas poblaciones aisladas que contienen un número reducido de plantas por lo que se la consideró una especie en peligro de extinción. Todos los esfuerzos, utilizando estrategias clásicas dirigidas a ampliar el área de crecimiento de Lmj y garantizar su conservación, han tenido un éxito limitado. El trabajo que se presenta está dirigido a mejorar el conocimiento de la ecología de Lmj, en particular la interacción simbiótica que establece con bacterias del suelo denominadas rizobios y se centra en la caracterización fenotípica, filogenética y genómica de esos rizobios. También se investiga la posible contribución de la simbiosis en mejorar la conservación de Lmj. Para este fin, se han estudiado diferentes aspectos que se describen a continuación. El primero objetivo se centró en aislar y estudiar de la diversidad genética de las bacterias endosimbióticas de Lmj. . Se realizó un análisis filogenético de genes esenciales que mostró que las cepas de Lmj pertenecen al género Bradyrhizobium y que presentan una gran diversidad con características fenotípicas y simbióticas diferentes de cepas de Bradyrhizobium que nodulan otras especies de lupinos nativos de España (cepas ISLU). Las cepas estudiadas se dividieron en dos grupos (Clado I y Clado II). El Clado I, incluye a las cepas Lmj, definiendo un nuevo linaje, filogenéticamente relacionado con otras especies de Bradyrhizobium, como B. jicamae y B. elkanii. El Clado II contiene cepas ISLU relacionadas con cepas de B. canariense y B. japonicum que establecen simbiosis con lupinos de suelos ácidos. Otro análisis filogenético basado en genes simbióticos, distribuyó las cepas de Lmj en sólo dos grupos diferentes. La singularidad y gran diversidad de estas cepas en una pequeña área geográfica, hacen de este, un atractivo sistema para el estudio de la evolución y adaptación de las bacterias simbióticas a su respectiva planta huésped. Adicionalmente, se estudio la presencia de bacterias capaces de nodular Lmj en suelos básicos de Chiapas, México. Sorprendentemente, estos suelos contienen bacterias capaces establecer interacciones simbióticas eficientes con Lmj en ensayos de invernadero. A continuación se investigó la taxonomía de los endosimbiontes de Lmj analizando la secuencia de cuatro genes esenciales (16S rRNA, recA, glnII y atpD) y el promedio de identidad de nucleótidos de genomas completos de algunas cepas representativas de la diversidad (ANIm). Se identificaron nuevas especies de Bradyrhizobium dentro del Clado I y se definió una de ellas: 'Bradyrhizobium valentinum' sp. nov (cepa tipo LmjM3T = CECT 8364T, LMG 2761T). También se abordó cómo conservar Lmj en su hábitat natural mediante inoculación con alguna de las cepas aisladas. Se demostró la ausencia de bacterias capaces de nodular Lmj en suelos rojos alcalinos o ‘‘terra rossa’’ de la Península Ibérica y Baleares. Dos cepas, altamente eficientes en cuanto a la fijación de nitrógeno, LmjC y LmjM3T, fueron seleccionadas para ser empleadas como inoculantes. Dos experimentos de campo llevados a cabo en años consecutivos en áreas con características edafoclimáticas similares a las que presentan las poblaciones de Lmj, lograron la reproducción exitosa de la planta. Se concluyó que un ciclo reproductivo exitoso de Lmj es absolutamente dependiente de la inoculación con sus simbiontes naturales y que la simbiosis debe ser considerada un factor esencial en estrategias de conservación de leguminosas en peligro. La obtención de varias secuencias genómicas de cepas aisladas de Lmj y de otras cepas de Bradyrhizobium reveló una alta similitud entre los genomas de las cepas del Clado I, y permitió la identificación de cinco posibles nuevas especies. Además, se estudiaron tres agrupaciones de genes relacionados con la simbiosis (nod, nif y fix) definiendo un nuevo linaje para las cepas de Lmj, diferente del symbiovar “genistearum” de B. canariense y B. japonicum. La baja diversidad encontrada en el análisis filogenético de los genes simbióticos contrasta con la gran diversidad asociada a genes esenciales. La presencia de plásmidos en cepas del género Bradyrhizobium ha sido descrita en muy pocas ocasiones, sin embargo el análisis de la secuencia genómica de la cepa ISLU101, aislada de Lupinus angustifolius, reveló la presencia de un origen de replicación extracromosómico homólogo al operón repABC, presente en el plásmido de Bradyrhizobium sp BTAi1. Gracias a esta secuencia se identificaron genes homólogos en 19 de 72 cepas ISLU. Filogenéticamente, las secuencias de repABC se agruparon en un grupo monofilético con las de pBTAi1 y separadas de los rizobios de crecimiento rápido. Finalmente, se identificaron sistemas de secreción de proteínas de tipo III (T3SS) en nueve genomas de cepas de Lmj. Los T3SS pueden inyectar proteínas efectoras al interior de células vegetales. Su presencia en rizobios se ha relacionado con la gama de hospedador que pueden nodular y puede tener un efecto beneficioso, neutro o perjudicial en la simbiosis. Los T3SS de las cepas de Lmj codifican para una proteína efectora similar a NopE, un efector dependiente de T3SS descrito en B. diazoefficiens USDA 110T. La proteína NopE de la cepa LmjC se ha caracterizado bioquímicamente. ABSTRACT Lupinus mariae-josephae (Lmj) is a lupine species endemic of a unique small area in Valencia region (Eastern Spain) where the lupine plants thrive in alkaline-limed soils, which preferentially grow in acid or neutral soils. This is the type of soils native lupines of Spain. When this work was initiated, the extension of the endemic area of Lmj was of about 700 squared kilometers confined to the Valencia province. In this area, Lmj thrives in small, isolated patches containing a reduced number of plants, and points to an endemism that can easily became endangered or extinct. Consequently, the Valencia Community authorities gave a ‘‘microreserve” status for conservation of the species. All efforts, using classical strategies directed to extend the area of Lmj growth and ensure its conservation have been so far unsuccessful. The work presented here is directed to improve our knowledge of Lmj ecology and it is centered in the characterization of the rhizobial symbiosis by phenotypic, phylogenetic and genomic analysis as well as in investigate the potential contribution of the symbiosis to improve its conservation. To this end, five different topics have been studied, and results are briefly described here. Extensive details can be followed en the attached, published articles. The first topic deals with the indigenous rhizobial symbionts of the Lmj endemism, and its genetic diversity was investigated. The Lmj root symbionts belong to the Bradyrhizobium genus, and phylogenetic analysis based on core genes identified a large diversity of Bradyrhizobium strains with phenotypic and symbiotic characteristics different from rhizobia nodulating other Lupinus spp. native of Spain. The strains were split in two clades. Clade II contained strains close to classical B. canariense and B. japonicum lineages that establish symbioses with lupines in acid soils of the Mediterranean area. Clade I included Lmj strains that define a new lineage, close to other Bradyrhizobium species as B. jicamae and B. elkanii. The phylogenetic analysis based on symbiotic genes identified only two distinct clusters. The singularity and large diversity of these strains in such a small geographical area makes this an attractive system for studying the evolution and adaptation of the rhizobial symbiont to the plant host. Additionally, the presence of bacteria able to nodulate Lmj in basic soils from Chiapas, Mexico was investigated. Surprisingly, these soils contain bacteria able to effectively nodulate and fix nitrogen with Lmj plants in greenhouse assays. In the second topic, the taxonomic status of the endosymbiotic bacteria of Lmj from Valencia endemism and Chiapas was investigated. Results from phylogenetic analysis of core genes and Average Nucleotide Identity (ANIm) using draft genomic sequences identified new Bradyrhizobium species within strains of Clade I of Lmj endosymbiotic bacteria. Only one of these potentially new species has been defined, meanwhile the others are under process of characterization. The name ‘Bradyrhizobium valentinum’ sp. nov. was proposed for the defined species (type strain LmjM3T= CECT 8364T, LMG 2761T). The third topic was directed to conservation of endangered Lmj in its natural habitat. The relevant conclusion of this experimentation is that the symbiosis should be considered as a relevant factor in the conservation strategies for endangered legumes. First, we showed absence of bacteria able to nodulate Lmj in all the inspected ‘‘terra rossa’’ or alkaline red soils of the Iberian Peninsula and Balearic Islands. Then, two efficient nitrogen fixing strains with Lmj plants, LmjC and LmjM3T, were selected as inoculum for seed coating. Two planting experiments were carried out in consecutive years under natural conditions in areas with edapho-climatic characteristics identical to those sustaining natural Lmj populations, and successful reproduction of the plant was achieved. The relevant conclusion from these assays was that the successful reproductive cycle was absolutely dependent on seedling inoculation with effective bradyrhizobia The forth topic deep into the analysis of the genomic of Lmj representative strains. To this end, draft genomic sequences of selected Lmj strains and type strains of Bradyrhizobium spp. were assembled. The comparison analysis of the draft genomic sequences of Lmj strains and related Bradyrhizobium species grouped in Clade I, revealed a high genomic homology among them, and allowed the definition of five potentially new species of Lmj nodulating bacteria. Also, based on the available draft genomic sequences, only three clusters of nod, fix and nif genes from Lmj strains were identified and showed to define a new symbiotic lineage, distant from that of B. canariense and B. japonicum bv. genistearum. The low diversity exhibited by the phylogenetic analysis of symbiotic genes contrast with the large diversity of strains as regards the housekeeping genes analyzed. Besides, the genomic analysis of a Lupinus angustifolius strain ISLU101, revealed the presence of an extrachromosomal replication origin homologous to repABC cluster from plasmid present in Bradyrhizobium spp BTAi1. This repABC cluster gene sequence allowed the identification of extrachromosomic replication origin in 19 out of 72 Bradyrhizobium strains from Lupinus spp., a highly significant result since the absence of plasmids in the Bradyrhizobium genus was traditionally assumed. The repABC gene sequences of these strains grouped them in a unique monophyletic group, related to B. sp. BTAi1 plasmid, but differentiated from the repABC gene cluster of plasmids in fast growing rhizobium strains. The last topic was focused on characterization of type III secreted effectors present in Lmj endosymbiotic bacteria. Type III secretion systems (T3SS) are specialized protein export machineries which can deliver effector proteins into plant cells. The presence of T3SS in rhizobia has frequently been related to the symbiotic nodulation host-range and may have a beneficial or detrimental effect on the symbiosis with legumes. In this context, the presence of T3SS in genomes of nine Lmj strains was investigated, and it was shown the presence of clusters encoding NopE type III-secreted protein similar to the NopE1 and NopE2 of B. diazoefficiens USDA 110T. The putative NopE protein of LmjC strain is at present being characterized regarding its structure and function.

Use of cDNA subtraction and RNA interference screens in combination reveals genes required for germ-line development in Caenorhabditis elegans

Caenorhabditis elegans is an ideal organism for the study of the molecular basis of fundamental biological processes such as germ-line development, especially because of availability of the whole genome sequence and applicability of the RNA interference (RNAi) technique. To identify genes involved in germ-line development, we produced subtracted cDNA pools either enriched for or deprived of the cDNAs from germ-line tissues. We then performed differential hybridization on the high-density cDNA grid, on which about 7,600 nonoverlapping expressed sequence tag (EST) clones were spotted, to identify a set of genes specifically expressed in the germ line. One hundred and sixty-eight clones were then tested with the RNAi technique. Of these, 15 clones showed sterility with a variety of defects in germ-line development. Seven of them led to the production of unfertilized eggs, because of defects in spermatogenesis (4 clones), or defects in the oocytes (3 clones). The other 8 clones led to failure of oogenesis. These failures were caused by germ-line proliferation defect (Glp phenotype), meiotic arrest, and defects in sperm–oocyte switch (Mog phenotype) among others. These results demonstrate the efficacy of the screening strategy using the EST library combined with the RNAi technique in C. elegans.

Restrictions on Aliens in the United Kingdom, 1914-1919 : a Study of the Origins and Political Background of the Aliens Restrictions Acts of 1914 and 1919.

At first glance the Aliens Restriction Act of 1914, which was introduced and passed on the first day of World War One, seems a hasty and ill-prepared piece of legislation. Actually, when examined in the light of Arthur Marwick's thesis that war is a forcing house for pre-existent social and governmental ideas, it becomes clear that the act was not after all the product of hastily formed notions. In point of fact it followed the precedent of detailed draft clauses produced in 1911 by a sub-committee of the Committee of Imperial Defence established to consider the treatment of aliens in the event of war. Indeed the draft clauses and the restrictions embodied in the 1914 act were strikingly similar to restrictions on aliens legislated in 1793. Hostility to aliens had been growing from 1905 to 1914 and this hostility blossomed into xeno-phobia on the outbreak of war, a crucial precondition for the specifically anti-enemy fears of the time. In 1919 the Aliens Restriction (Amendment) Bill was introduced into parliament to extend temporarily the provisions of the 1914 act thus permitting the Home Secretary to plan permanent, detailed legislation. Two minority groups of MPs with extreme views on the treatment of aliens were prominent in the debates on this bill. The extreme Liberal group which advocated leniency in the treatment of aliens had little effect on the final form of the bill, but the extreme Conservative group, which demanded severe restrictions on aliens, succeeded in persuading the government to include detailed restrictions. Despite its allegedly temporary nature, the Aliens Restriction (Amendment) Act of 1919 was renewed annually until 1971.