Functional thin films fabricated by plasma polymerization and their biological applications

Plasma polymerization technique is widely accepted as an effective and simple method for the preparation of functional thin films. By careful choice of precursors and deposition parameters, plasma polymers bearing various functional groups could be easily obtained. In this work, I explored the deposition of four kinds of plasma polymerised functional thin films, including the protein-resistant coatings, the thermosensitive coatings, as well as, the coatings bearing amine or epoxide groups. The deposited plasma polymers were characterized by various techniques, such as X-ray photoelectron spectroscopy, atom force microscopy, Fourier transform infrared spectroscopy, surface plasmon resonance spectroscopy, optical waveguide spectroscopy, and so on. As expected, high retention of various functional groups could be achieved either at low plasma input power or at low duty cycle (duty cycle = Ton/(Ton+Toff)). The deposited functional thin films were found to contain some soluble materials, which could be removed simply by extraction treatment. Besides the thermosentive plasma polymer (see Chapter 9), other plasma polymers were used for developing DNA sensors. DNA sensing in this study was achieved using surface plasmon enhanced fluorescence spectroscopy. The nonfouling thin films (i.e., ppEO2, plasma polymerization of di(ethylene glycol) monovinyl ether) were used to make a multilayer protein-resistant DNA sensor (see Chapter 5). The resulted DNA sensors show good anti-fouling properties towards either BSA or fibrinogen. This sensor was successfully employed to discriminate different DNA sequences from protein-containing sample solutions. In Chapter 6, I investigated the immobilization of DNA probes onto the plasma polymerized epoxide surfaces (i.e., ppGMA, plasma polymerization of glycidyl methacrylate). The ppGMA prepared at a low duty cycle showed good reactivity with amine-modified DNA probes in a mild basic environment. A DNA sensor based on the ppGMA was successfully used to distinguish different DNA sequences. While most DNA detection systems rely on the immobilization of DNA probes onto sensor surfaces, a new homogeneous DNA detection method was demonstrated in Chapter 8. The labeled PNA serves not only as the DNA catcher recognizing a particular target DNA, but also as a fluorescent indicator. Plasma polymerized allylamine (ppAA) films were used here to provide a positively charged surface.

Gespeicherte Ionen in der Paulfalle: Voruntersuchungen zur Lebensdauermessung des metastabilen 6D-3/2-Zustands des 226 Ra +, Speichereigenschaften von 138 Ba + und 28 N 2 + bei Variation des Puffergasdrucks

Auf Paulfallen basierende Experimente spielen eine wichtige Rolle in verschiedenen Bereichen der Physik, z.B. der Atomphysik zum Test theoretischer Modelle und der Massenspektroskopie. Die vorliegende Arbeit widmet sich beiden Themengebieten und gliedert sich entsprechend in zwei Teilbereiche: 1) Erdalkali-Ionen sind aufgrund ihrer Energieniveaus optimale Kandidaten für Laserspektroskopie-Experimente mit Ionenfallen und bestens geeignet, um mittels der spektroskopischen Daten die theoretischen Modelle zu testen. Lediglich für Ra+ fehlen bislang als einzigem Erdalkali-Ion diese Daten wie z.B. die Lebensdauern der metastabilen Niveaus. Diese wären auch von Interesse für bereits geplante Radium-Experimente zur Paritätsverletzung. Im ersten Teil dieser Arbeit wird der Aufbau eines Laser-Paulfallenexperiments zur Messung der Lebensdauer des 6D3/2 Zustands von 226Ra+ dokumentiert und es werden Testmessungen mit 138Ba+ vorgestellt. 2) Für die Verwendung der Paulfalle in der Massenspektroskopie und zur Analyse von Reaktionsprodukten ist die Kenntnis der Lage der im Speicherbereich auftretenden nichtlinearen Resonanzen wesentlich, ebenso wie deren Veränderung durch Dämpfung und Raumladung. Im zweiten Teil dieser Arbeit werden detaillierte Untersuchungen der Speicherung großer puffergasgekühlter Ionenwolken an zwei unterschiedlichen Paulfallen-Experimenten vorgestellt. Am ersten wurden 138Ba+-Ionenwolken kontinuierlich durch Laserspektroskopie bzw. über einen elektronischen Nachweis beobachtet, während das zweite N2+-Molekülionen automatisiert destruktiv nachwies. Am N2+-Experiment wurden zwei hochaufgelöste Messungen des ersten Speicherbereichs durchgeführt, die erstmals eine direkte Überprüfung der theoretisch berechneten Verläufe der Resonanzen mit experimentellen Daten erlauben. Die Nachweiseichung ermöglichte dabei zum ersten Mal die Angabe absoluter Ionenzahlen. Im Gegensatz zu vergleichbaren früheren Messungen wurden hierbei die sich überlagernden Speicherbereiche von 4 simultan gespeicherten Ionensorten beobachtet und zur Analyse der Resonanzen herangezogen. Die nichtlinearen Resonanzen wurden untersucht bei Variation von Puffergasdruck und Ionenzahl, wobei kollektive Resonanzen ohne zusätzliche externe Anregung beobachtet wurden. Die gemessenen Raumladungsverschiebungen wurden mit theoretischen Modellen verglichen. Bei Variation des Puffergasdrucks wurde mit Bariumionen die räumliche Ausdehnung der Ionenwolke gemessen und mit Stickstoffionen die Verschiebung des Punktes optimaler Speicherung bestimmt. Dabei wurde festgestellt, daß der zum Ioneneinfang optimale Puffergasdruck kleiner ist als der, bei dem die längsten Speicherdauern erzielt werden. Mit gespeicherten N2+-Ionen wurde die Position extern angeregter kollektiver und individueller Resonanzen im Frequenzspektrum bei Änderung der Parameter Ionenzahl, Puffergasdruck und Dauer der Anregung untersucht, ebenso wie die Resonanzform, die mit theoretischen Linienformen verglichen wurde. Bei Änderung der Fallenparameter wurden verstärkende Effekte zwischen nahen kollektiven Resonanzen festgestellt. Die Beobachtung, welche der im Frequenzspektrum vorher identifizierten Bewegungs-Resonanzen sich bei Variation der Fallenparameter a bzw. q überlagern, ermöglicht eine bislang nicht beschriebene einfache Methode der Bestimmung von nichtlinearen Resonanzen im Stabilitätsdiagramm.

Isolierung, Charakterisierung und Nachweis methanogener Archaea aus laufenden Praxis-NawaRo-Biogasanlagen unter besonderer Beruecksichtigung der Gattung Methanobacterium

Im Verlauf der Forschungsarbeit wurden Proben aus fünf, mit nachwachsenden Rohstoffen (NawaRo) beschickten, landwirtschaftlichen Biogasanlagen (BGA) auf die Biozönose methanogener Archaea hin molekularbiologisch untersucht. Über „amplified rDNA restriction analysis“-Screening (ARDRA) von Bibliotheken auf Basis von 16S rRNA-Genfragmenten konnte anhand zweier beispielhafter BGA das Vorkommen von Vertretern der Gattungen Methanoculleus (Mcu.), Methanobacterium (Mb.), Methanosarcina (Msc.) und Methanosaeta (Mst.) nachgewiesen werden. Mittels denaturierender Gradienten-Gelelektrophorese (DGGE) wurde das Vorkommen dieser Mikroorganismen auch in den übrigen Anlagen gezeigt. Ergänzend dazu wurde in drei Anlagen Methanospirillum hungatei nachgewiesen. Nach Ausarbeitung gattungsspezifischer Isolierungsstrategien konnten insgesamt zehn Vertreter der Gattung Methanobacterium (Isolate Mb1 bis Mb10) und jeweils ein Vertreter der Gattungen Methanoculleus (Isolat Mcu(1)), Methanosarcina (Isolat NieKK) und Methanosaeta (Isolat Mst1.3) aus den BGA-Proben isoliert werden. Durch in silico-Abgleich der partiellen 16S rRNA-Gensequenzen wurden diese als Verwandte von Mb. formicicum MFT, Mcu. bourgensis MS2T, Msc. mazei S-6T und Mst. concilii FE mit einer Sequenzidentität > 97% identifiziert. Im Laufe weiterer molekularbiologischer Untersuchungen mittels DGGE und ARDRA-Analyse konnten die Isolate den Referenzstämmen zugeordnet werden. In Bezug auf die Gattung Methanobacterium ergaben sich jedoch leichte Abweichungen. Diese bestätigten sich in vergleichenden Analysen des genomischen Fingerabdrucks in der „specifically amplified polymorphic DNA“-PCR (SAPD-PCR), welche im Rahmen dieser Arbeit erstmalig erfolgreich auf archaeelle Organismen angewandt wurde. Hier zeigten die Isolate zwei von den Fingerabdrücken der untersuchten Referenzstämme verschiedene Hauptamplifikationsmuster. Aufgrund der Vielzahl der Isolate sowie dem signifikanten Vorkommen in qPCR-Analysen und Klonbibliotheken fokussierten sich die weiteren Arbeiten zur genauen Untersuchung dieser Abweichungen auf phylogenetische Analysen der Gattung Methanobacterium und die Entwicklung von Nachweissystemen. Die Aufklärung eines Großteils der 23S rRNA-Gensequenzen der Isolate und von ausgewählten Typstämmen ermöglichte ergänzende phylogenetische Untersuchungen zu durchgeführten 16S rRNA-Analysen. Dabei wurden die Isolate jeweils in einem eigenen Cluster abseits der meisten Referenzstämme aus der Gattung Methanobacterium positioniert. Analog zur Musterbildung im Rahmen der SAPD-Analyse zeigte sich eine Differenzierung in zwei Äste und ergab in Übereinstimmung mit den in silico-Sequenzabgleichen den höchsten Verwandtschaftsgrad mit Mb. formicicum MFT. Die Eignung der SAPD-PCR zur Ableitung spezifischer Primerpaare konnte erstmals auch für methanogene Archaea gezeigt werden. Die Ableitung zweier Primerpaare mit Spezifität für die Methanobacterium-Isolate Mb1 bis Mb10 sowie für den Typstamm Mb. formicicum MFT gelang und konnte im Rahmen eines Direkt-PCR-Nachweises erfolgreich auf Reinkulturen und Fermenterproben angewandt werden. Unter Einbezug der sequenzierten 23S rRNA-Genfragmente gelang die Erstellung von Oligonukleotid-Sonden für den Einsatz in Fluoreszenz in situ-Hybridisierungsexperimenten. Im Praxistest ergab sich für diese Sonden eine Spezifität für alle getesteten Vertreter der Gattung Methanobacterium sowie für Methanosphaera stadtmanae MCB-3T und Methanobrevibacter smithii PST.rnSomit konnten im Laufe der Arbeit die dominanten methanogenen Archaea in NawaRo-BGA in mehrphasigen Experimenten nachgewiesen, quantifiziert und auf nur wenige Gattungen eingegrenzt werden. Vertreter der vier dominanten Gattungen wurden isoliert und Nachweissysteme für Arten der Gattung Methanobacterium erstellt.rn

Der g-Faktor des Protons

In dieser Arbeit wird die bisher präziseste und erste direkte Hochpräzisionsmessung des g-Faktors eines einzelnen Protons präsentiert. Die Messung beruht auf der nicht-destruktiven Bestimmung der Zyklotronfrequenz und der Larmorfrequenz eines in einer Penning-Falle gespeicherten Protons. Zur Bestimmung der Larmorfrequenz wird die Spin-Flip-Wahrscheinlichkeit als Funktion einer externen Spin-Flip-Anregung aufgenommen. Zu diesem Zweck wird der kontinuierliche Stern-Gerlach Effekt verwendet, welcher zu einer Kopplung des Spin-Moments an die axiale Bewegung des Protons führt. Ein Spin-Flip zeigt sich dabei in einem Sprung der axialen Bewegungsfrequenz. Die Schwierigkeit besteht darin, diesen Frequenzsprung auf einem Hintergrund axialer Frequenzfluktuationen zu detektieren. Um diese Herausforderung zu bewältigen, wurden neuartige Methoden und Techniken angewandt. Zum einen wurden supraleitende Nachweise mit höchster Empfindlichkeit entwickelt, welche schnelle und damit präzise Frequenzmessungen erlauben. Zum anderen wurde eine auf dem statistischen Bayes Theorem basierende Spin-Flip-Analyse-Methode angewandt. Mit diesen Verbesserungen war es möglich, einzelne Spin-Flips eines einzelnen Protons zu beobachten. Dies wiederum ermöglichte die Anwendung der sogenannten Doppelfallen-Methode, und damit die eingangs erwähnte Messung des g-Faktors mit einer Präzision von 4.3 10^-9.

On-chip non-invasive and label-free cell discrimination by impedance spectroscopy

OBJECTIVES: Many flow-cytometric cell characterization methods require costly markers and colour reagents. We present here a novel device for cell discrimination based on impedance measurement of electrical cell properties in a microfluidic chip, without the need of extensive sample preparation steps and the requirement of labelling dyes. MATERIALS AND METHODS, RESULTS: We demonstrate that in-flow single cell measurements in our microchip allow for discrimination of various cell line types, such as undifferentiated mouse fibroblasts 3T3-L1 and adipocytes on the one hand, or human monocytes and in vitro differentiated dendritic cells and macrophages on the other hand. In addition, viability and apoptosis analyses were carried out successfully for Jurkat cell models. Studies on several species, including bacteria or fungi, demonstrate not only the capability to enumerate these cells, but also show that even other microbiological life cycle phases can be visualized. CONCLUSIONS: These results underline the potential of impedance spectroscopy flow cytometry as a valuable complement to other known cytometers and cell detection systems.

Road Ecology: A Practitioner's Tale

Road Ecology is a relatively new sub-discipline of ecology that focuses on understanding the interactions between road systems and the natural environment. Wildlife crossings that allow animals to safely cross human-made barri-ers such as roads, are intended not only to reduce animal-vehicle collisions, but ideally to provide connectivity of habitat areas, combating habitat fragmentation. Wildlife mitigation strategies to improve the permeability of our infrastructure can include a combination of structures (overpasses/underpasses), at-grade crossings, fencing, animal-detection systems, and signage. One size does not fit all and solutions must be considered on a case-by-case ba-sis. Often, the feasibility of the preferred mitigation solution depends on a combination of variables including road geometrics, topography, traffic patterns, funding allocations, adjacent land use and landowner cooperation, the target wildlife species, their movement patterns, and habitat distribution. Joe and Deb will speak to the current road ecolo-gy practices in Montana and some real-world applications from the Department of Transportation.

Bases moleculares de la resistencia a insecticidas en la mosca mediterránea de la fruta Ceratitis Capitata (Wiedemann)

La mosca mediterránea de la fruta Ceratitis capitata (Wiedemann, 1824) está considerada una de las plagas clave para la fruticultura. El malatión es un insecticida organofosforado que fue empleado mayoritariamente en España para el control de C. capitata hasta 2009, año en el que dejó de utilizarse por no estar incluido en el anexo I de la Directiva Europea 91/414/ECC. El incremento del uso del malatión, debido a las graves pérdidas económicas causadas por C. capitata, provocó la aparición de poblaciones de campo resistentes. El estudio de una población resistente a malatión, recogida en Castelló en 2004, permitió la identificación de dos mecanismos de resistencia: una mutación puntual (G328A) en la acetilcolinesterasa (AChE) y un mecanismo de resistencia metabólica, probablemente mediado por carboxilesterasas. Teniendo en cuenta estos antecedentes, nos propusimos estudiar los mecanismos implicados en la resistencia a malatión en C. capitata. Además, durante el desarrollo de esta Tesis, el malatión fue sustituido por otros insecticidas como el espinosad y la lambda-cialotrina para el control de la plaga. En este nuevo contexto, es extremadamente importante analizar la susceptibilidad de poblaciones de campo frente a espinosad y estudiar la posible existencia de resistencia cruzada a estos insecticidas, así como sentar las bases para el estudio de futuros mecanismos de resistencia. En primer lugar, analizamos mediante bioensayos con dosis discriminante la susceptibilidad a malatión y espinosad en doce poblaciones de C. capitata de Andalucía, Aragón, Cataluña, Comunidad Valenciana e Islas Baleares; y nuestros resultados sugirieron la presencia de individuos resistentes a malatión en la mayoría de las poblaciones analizadas. En el caso del espinosad, observamos que la susceptibilidad a este insecticida de origen biológico fue elevada en la mayoría de las poblaciones, sin embargo, la población recogida en Xàbia (Alicante) mostró un nivel de susceptibilidad unas dos veces menor al resto de poblaciones. Mediante la selección en laboratorio, obtuvimos dos líneas resistentes a malatión, W-4Km y W-10Km, con unos niveles de resistencia con respeto a la línea susceptible C de 178 y 400 veces, respectivamente. Además, se seleccionó por primera vez en C. capitata una línea altamente resistente a espinosad (Xàbia-W-100s), que actualmente es unas 500 veces más resistente que la línea de laboratorio C. Con el objetivo de escoger la estrategia más adecuada para el manejo de la plaga, estudiamos la susceptibilidad a diferentes tipos de insecticidas en la línea resistente a malatión W- 4Km. En esta línea detectamos resistencia cruzada moderada a los organofosforados fentión, diazinón, fosmet, triclorfón y metil-clorpirifos (de 7 a 16 veces) y frente al carbamato carbaril, al piretroide lambda-cialotrina y al quimioesterilizante lufenurón (de 4 a 6 veces). Por otra parte, la resistencia cruzada frente a espinosad fue baja (1,5 veces). Es importante destacar que los niveles de resistencia estimados frente a todos los insecticidas fueron de uno o dos órdenes de magnitud inferiores al observado en la línea W-4Km frente a malatión (178 veces), hecho que podría deberse, al menos, a dos posibles hipótesis: que la mutación AChE G328A confiera mayor insensibilidad al malaoxón (forma activa del malatión) que a otros insecticidas que tienen como diana la AChE y/o, en segundo lugar, que el mecanismo de resistencia mediado por carboxilesterasas hidrolice el malatión de manera más eficiente que los otros insecticidas analizados. En el estudio de nuevos mecanismos de resistencia en C. capitata, por un lado, analizamos la diversidad de enzimas citocromo P450, asociadas con resistencia metabólica en otras especies, y por otro lado, desarrollamos un sistema para la detección de nuevas mutaciones puntuales que pudiesen aparecer en los genes que codifican la AChE (Ccace2) y la aliesterasa (Ccae7). Mediante el empleo de cebadores degenerados obtuvimos 37 genes CYP, que codifican enzimas P450, pertenecientes a cinco familias. Posteriormente, en un estudio de inducción con fenobarbital, observamos que la expresión de cuatro de los seis genes analizados era susceptible de ser inducida. Por otro lado, se puso a punto un sistema que permite amplificar y secuenciar, a partir de DNA genómico, los exones de los genes Ccace2 y Ccae7 en los que se han encontrado mutaciones relacionadas con resistencia a insecticidas en otras especies. Los resultados obtenidos facilitarán el estudio de nuevos mecanismos de resistencia mediados por estas enzimas en C. capitata. Se diseñó un método PCR-RFLP para identificar los individuos portadores de la mutación AChE G328A (alelo de resistencia Ccace2R) sin la necesidad de realizar bioensayos y que, además, permite detectar resistencia cuando ésta se encuentra a baja frecuencia. Según el análisis realizado, el alelo Ccace2R se observó en 25 de las 27 localidades españolas muestreadas en el territorio español, incluyendo las Islas Baleares y Canarias. Sin embargo, este alelo no se detectó en poblaciones procedentes de once países y de cinco continentes. El análisis de la presencia del alelo Ccace2R en las líneas resistentes a malatión durante el proceso de selección en el laboratorio mostró una rápida disminución de los homocigotos, tanto para el alelo susceptible como para el alelo de resistencia, en favor de los individuos heterocigotos. Así, después de 52 generaciones de selección, se observó que la totalidad de los individuos analizados de la línea W-10Km presentaban un genotipo heterocigoto para la mutación AChE G328A. Este desequilibrio contradice la segregación mendeliana esperada para un gen con dos alelos pero podría ser explicado por la existencia de una duplicación del gen Ccace2. La demostración de la presencia de esta duplicación se realizó mediante: i) el cruzamiento de individuos heterocigotos de la línea W-10Km con homocigotos susceptibles de la línea C, que dio lugar a una descendencia en la que el 100% de los individuos eran heterocigotos; ii) la evaluación del número de copias del gen Ccace2 por PCR cuantitativa en tiempo real (qPCR), que resultó dos veces mayor en individuos de la línea W-10Km en comparación con los de la línea C; iii) el análisis del nivel de expresión de Ccace2, que fue el doble en la línea W-10Km con respecto a la línea C, y iv) el estudio de la actividad AChE, que resultó mayor en los individuos de la línea W-10Km. Según los resultados obtenidos, una duplicación del gen Ccace2 provoca la coexistencia en un mismo cromosoma del alelo silvestre y del alelo mutado y, además, las dos copias del gen Ccace2, al estar ligadas, producen una heterocigosis permanente (Ccace2RS). De esta manera se explica que el hecho de que 100% de los individuos de la línea W-10Km mostrasen un perfil de restricción correspondiente a un individuo heterocigoto ya que, en realidad, eran homocigotos estructurales para la duplicación (genotipo CCace2RS/RS). Se ha detectado un coste biológico asociado a la duplicación que consiste en un incremento en la mortalidad acumulada de los adultos a partir del séptimo día después de la emergencia. La descripción de la duplicación Ccace2RS supone la identificación de un nuevo mecanismo de resistencia a malatión en C. capitata. Finalmente, mediante el diseño de un método de doble PCR-RFLP se determinó la presencia de la duplicación Ccace2RS en la mayoría de las poblaciones españolas. La proporción de individuos portadores de la duplicación osciló entre el 5% y el 35%, observándose los mayores valores de frecuencia en las poblaciones de C. capitata recogidas en la cuenca mediterránea. Podemos por lo tanto concluir que la resistencia a malatión asociada a la mutación AChE G328A y a la duplicación Ccace2RS está ampliamente establecida en las poblaciones españolas de C. capitata. Nuestros resultados desaconsejan la utilización del malatión (si fuera de nuevo autorizado) o de otros organofosforados para el control de esta plaga. Además, una de las líneas resistentes a malatión mostró resistencia cruzada frente a insecticidas con diferentes modos de acción y que se utilizan actualmente para el control de C. capitata, tales como lambda-cialotrina y lufenurón. La alta susceptibilidad a espinosad observada en las poblaciones españolas, así como la reducida resistencia cruzada estimada para este insecticida, sugieren que su utilización es adecuada para el control de la plaga. Sin embargo, la utilización de un sólo insecticida puede entrañar riesgos por favorecer la selección de resistencia, de hecho, mediante selección en laboratorio se obtuvo una población altamente resistente a espinosad. Por tanto, es recomendable implementar programas de control integrado y de manejo de la resistencia en C. capitata utilizando distintos sistemas de control e insecticidas con diferentes mecanismos de acción que permitan su sostenibilidad en el tiempo. Los sistemas de detección de alelos de resistencia desarrollados en este trabajo permitirán la detección precoz de resistencia en campo, facilitando la decisión sobre el sistema de control más adecuado. Además, los conocimientos generados podrán contribuir al desarrollo de nuevos sistemas de detección para otros mecanismos de resistencia. Abstract. The Mediterranean fruit fly, Ceratitis capitata (Wiedemann, 1824), is considered one of the most harmful pests in fruit crops. Until 2009, when malathion use was banned due to its not inclusion in the Annex I of Directive 91/414/EEC, the application of this organophosphate (OP) insecticide in Spain increased gradually due to the large economic losses caused by C. capitata. The increase in the frequency of treatments resulted in the development of resistant field populations. The study of a malathion-resistant population, collected in 2004 in Castelló (Comunidad Valenciana), allowed the identification of two resistance mechanisms: a single point mutation (G328A) in the target acetylcholinesterase (AChE), as well as a metabolic resistance mechanism, most likely carboxylesterase-mediated. Taking all the preceding into account, we studied the malathion resistance mechanisms in C. capitata. During the development of this PhD Thesis malathion use was banned by the European Union, being replaced by other insecticides, such as spinosad and lambda-cyhalotrin. Within this new working frame, the need to analyse the possible existence of cross-resistance to these insecticides and the susceptibility to spinosad in field populations was raised. This would define the baseline for future studies on resistance mechanisms. Firstly, through discriminant dose bioassays, we analysed malathion and spinosad susceptibility in twelve C. capitata populations from Andalucia, Aragon, Cataluña, C. Valenciana and the Baleares Islands. Our results suggest the presence of malathion-resistant individuals in most of the populations analysed. Regarding spinosad, we noticed a high susceptibility to this biologically derived insecticide in most of the populations, but in the one collected in Xabia (Alicante), which had a susceptibility level two times lower than the rest of populations. Through laboratory selection, we obtained two malathion-resistant strains, W-4Km and W-10Km, with resistance levels 178- and 400-fold, respectively, compared to the control susceptible C strain. Besides, a strain highly-resistant to spinosad (Xabia-W-100s), 500-times more resistant than control C strain, was selected. In order to decide the most appropriate management strategy for the pest, we studied the susceptibility to different insecticides in the malathion-resistant W-4Km strain. We detected a moderated cross-resistance to the OPs fenthion, diazinon, phosmet, trichlorphon and methylchlorpyrifos (7- to 16-fold), and to the carbamate carbaryl, the pyretroid lambda-cyhalotrin and the chemosterilizer lufenuron (4- to 6-fold). On the other hand, cross-resistance to spinosad was low (1.5-fold). It is important to note that resistance levels to all insecticides were one or two orders of magnitude less than that observed against malathion in W-4Km strain (178-fold), a fact that might be due to, at least, two possible causes: mutation AChE G328A may provide a higher insensitivity to malaoxon (the active form of malathion) than to other insecticides having AChE as target, and/or, secondly, the carboxylesterase-mediated resistance mechanism hydrolyzes malathion more efficiently than all other analysed insecticides. To investigate new resistance mechanisms in C. capitata we analysed the diversity of the cytochrome P450 enzymes, which have been associated to metabolic resistance in insects, and we developed a new method to detect single point mutations in acetylcholinesterase (Ccace2) and aliesterase (Ccae7) genes that could appear. Using degenerate primers we obtained 37 CYP genes, coding P450 enzymes, included in five families. Afterwards, in a phenobarbital-induction study, we observed that the expression of 4 out of the 6 analysed genes could be induced. On the other hand, a system was set up to amplify and to sequence from genomic DNA the exons of genes Ccace2 and Ccae7 where mutations related to insecticide resistance have been found in other species. The results obtained could facilitate the study of new resistance mechanisms in C. capitata mediated by these enzymes. A PCR-RFLP method was designed to detect the presence of the mutation AChE G328A (resistance allele Ccace2R), with no need to perform bioassays and allowing detecting resistance at low frequency. According to the analysis, the resistance allele was found in 25 out of 27 sampled locations in Spain, including the Balearic and the Canary Islands. However, this allele was not detected in other populations collected in 11 countries from 5 continents. The follow-up of the presence of the allele Ccace2R in the malathion-resistant strains during the selection process in the laboratory showed a quick decrease in homozygous individuals, for both the susceptible and the resistant alleles, favouring heterozygous. Thus, after 52 generations of selection, all the individuals analysed from W-10Km strain showed a heterozygous genotype for mutation AChE G328A, contradicting mendelian segregation as expected for a gene with two alleles. Afterwards, we were able to demonstrate that this was caused by the presence of a duplication of the gene coding acetylcholinesterase by: i) crossing heterozygous individuals from W-10Km strain with susceptible homozygous from C strain, originating a F1 population in which 100% of individuals were heterozygous; ii) evaluating the number of copies of gen Ccace2 by quantitative PCR in real time (qPCR), that happened to be twice higher in individuals from W-10Km VII strain when compared with C strain; iii) analysing the level of expression of Ccace2, twice in W- 10Km strain when compared to C strain; iv) studying the acetylcholinesterase activity, that was higher in individuals from W-10Km strain. According to these results, duplication of gen Ccace2 originates the coexistence of the susceptible and the resistant allele in the same chromosome. The two linked copies of the gene Ccace2 provoke the existence of permanent heterozygosis (Ccace2RS). This explains why the 100% of individuals from W-10Km strain showed an heterozygous restriction pattern since, in fact, they were structural homozygotes for the duplication (genotype Ccace2RS/RS). A biological cost has been detected associated to this duplication, consisting in a rise in accumulated adult mortality from the seventh day after emergence. The Ccace2RS duplication described in this study represents a new resistance mechanism to malathion in C. capitata. Finally, by the design of a double PCR-RFLP method, the presence of Ccace2RS duplication was confirmed in most of the Spanish populations. We observed that the proportion of individuals carrying the duplication oscillated between 5 and 35%, the frequency being higher in those C. capitata populations collected in the area of the Mediterranean basin. Therefore, we can conclude that malathion resistance associated to mutation AChE G328A and to Ccace2RS duplication are widely distributed in Spanish populations of C. capitata. Our results advice against the use of malathion (if it came to be newly authorized for use) or other OPs for the control of this pest. Besides, one of the malathion-resistant strains showed cross-resistance against insecticides with diverse action modes that are currently used for pest control, such as lambdacyhalotrin and lufenuron. High susceptibility to spinosad in the Spanish populations, as well as the reduced cross-resistance estimated for this insecticide suggests its adequacy for Medfly control. However, the use of a single insecticide is a risky strategy since it favours the selection of resistance. In fact, a population highly resistant to spinosad was obtained through laboratory selection. Therefore, it is advisable to implement integrated pest management (IPM) and resistance management programs for C. capitata control. Using insecticides with different modes of action and diverse control systems would contribute to the sustainability of the pest control. The resistance allele detection systems developed through this work will allow the early detection of resistance in the field, making possible the selection of the most appropriate method for pest control. Besides, the generated knowledge may also contribute to the development of new detection systems for other resistance mechanisms.

Contribución a la automatización de sistemas de respuesta frente a intrusiones mediante ontologías

La seguridad en redes informáticas es un área que ha sido ampliamente estudiada y objeto de una extensa investigación en los últimos años. Debido al continuo incremento en la complejidad y sofisticación de los ataques informáticos, el aumento de su velocidad de difusión, y la lentitud de reacción frente a las intrusiones existente en la actualidad, se hace patente la necesidad de mecanismos de detección y respuesta a intrusiones, que detecten y además sean capaces de bloquear el ataque, y mitiguen su impacto en la medida de lo posible. Los Sistemas de Detección de Intrusiones o IDSs son tecnologías bastante maduras cuyo objetivo es detectar cualquier comportamiento malicioso que ocurra en las redes. Estos sistemas han evolucionado rápidamente en los últimos años convirtiéndose en herramientas muy maduras basadas en diferentes paradigmas, que mejoran su capacidad de detección y le otorgan un alto nivel de fiabilidad. Por otra parte, un Sistema de Respuesta a Intrusiones (IRS) es un componente de seguridad que puede estar presente en la arquitectura de una red informática, capaz de reaccionar frente a los incidentes detectados por un Sistema de Detección de Intrusiones (IDS). Por desgracia, esta tecnología no ha evolucionado al mismo ritmo que los IDSs, y la reacción contra los ataques detectados es lenta y básica, y los sistemas presentan problemas para ejecutar respuestas de forma automática. Esta tesis doctoral trata de hacer frente al problema existente en la reacción automática frente a intrusiones, mediante el uso de ontologías, lenguajes formales de especificación de comportamiento y razonadores semánticos como base de la arquitectura del sistema de un sistema de respuesta automática frente a intrusiones o AIRS. El objetivo de la aproximación es aprovechar las ventajas de las ontologías en entornos heterogéneos, además de su capacidad para especificar comportamiento sobre los objetos que representan los elementos del dominio modelado. Esta capacidad para especificar comportamiento será de gran utilidad para que el AIRS infiera la respuesta óptima frente a una intrusión en el menor tiempo posible. Abstract Security in networks is an area that has been widely studied and has been the focus of extensive research over the past few years. The number of security events is increasing, and they are each time more sophisticated, and quickly spread, and slow reaction against intrusions, there is a need for intrusion detection and response systems to dynamically adapt so as to better detect and respond to attacks in order to mitigate them or reduce their impact. Intrusion Detection Systems (IDSs) are mature technologies whose aim is detecting malicious behavior in the networks. These systems have quickly evolved and there are now very mature tools based on different paradigms (statistic anomaly-based, signature-based and hybrids) with a high level of reliability. On the other hand, Intrusion Response System (IRS) is a security technology able to react against the intrusions detected by IDS. Unfortunately, the state of the art in IRSs is not as mature as with IDSs. The reaction against intrusions is slow and simple, and these systems have difficulty detecting intrusions in real time and triggering automated responses. This dissertation is to address the existing problem in automated reactions against intrusions using ontologies, formal behaviour languages and semantic reasoners as the basis of the architecture of an automated intrusion response systems or AIRS. The aim is to take advantage of ontologies in heterogeneous environments, in addition to its ability to specify behavior of objects representing the elements of the modeling domain. This ability to specify behavior will be useful for the AIRS in the inference process of the optimum response against an intrusion, as quickly as possible.

Desarrollo e implementación de un sistema robótico aéreo con programación tridimensional orientado al recorrido de rutas

En la actualidad se estudia en numerosos campos cómo automatizar distintas tareas ejecutadas por aeronaves con tripulación humana. Estas tareas son en todos los casos muy costosos, debido al gran consumo de combustible, gran coste de adquisición y mantenimiento de la propia aeronave, todo ello sin contar el riesgo para los mismos tripulantes. Como ejemplo de estas tareas se puede incluir la vigilancia policial y fronteriza, revisiones de tendidos de alta tensión, la alerta temprana de incendios forestales y la medición de parámetros contaminantes. El objetivo de este proyecto es el diseño y la construcción de un prototipo electrónico empotrado basado en microcontrolador con núcleo C8051 de Silicon labs, que sea capaz de gobernar una aeronave de radiocontrol de forma transparente, de manera que en un futuro se pueda sustituir el propio aeromodelo, con la modificación de algunos parámetros, para poder incorporar sistemas de video o distintos medios de detección de variables. El prototipo seguirá una ruta confeccionada y transferida como un archivo de texto con un formato determinado que contendrá los datos necesarios para poder navegar mediante GPS. El trabajo con los modelos de motorización térmica (motores de combustión interna tipo glow, en este caso) resulta peligroso debido a la gran energía que son capaces de alcanzar. A fin de mantener la máxima seguridad durante la evolución del proyecto, se ha diseñado un proceso de tres partes independientes que permitan la correcta familiarización de los distintos componentes que se emplearán. Las fases son las siguientes: 1. Test y modelado de todos los componentes mediante pequeños montajes con protoboard de inserción y programas individuales. Se realizará mediante una tarjeta multipropósito que contendrá un microcontrolador similar en características, aunque de menor complejidad, al del prototipo final. 2. Integración de todos los componentes mediante una tarjeta especialmente diseñada que servirá de interfaz entre la tarjeta multipropósito y todo el hardware necesario para el control de un vehículo terrestre de iguales características (actuadores y motorización) al aeromodelo. 3. Diseño de un sistema embebido que concentre todos los subsistemas desarrollados en las fases anteriores y que integre todos los componentes necesarios para el gobierno de una aeronave de ala fija. ABSTRACT. Nowadays, the way of automating different tasks done by manned vehicles is studied. These tasks are any case very expensive, due to large fuel consumption, costs of aircraft buying, without taking into account the risk for human crew. As an example of these tasks, we can include policing or border surveillance, maintenance of high voltage lines, early warning of forest fire and measuring of pollution parameters. The target of this project is the design and construction of an embedded electronic prototype, based on a microcontroller with C8051 core from Silicon labs, and it will be able to controlling an aircraft transparently, in order that in the future the flying model could be changed with the modification of some parameters, and video or any variables detection systems could be added. The prototype will follow a designed and transferred path as an plain text file with a given format, that will contain all the necessary data for GPS navigation. Working with heat engine models (internal combustion engine, glow type, in this case) becomes dangerous due to the large energy that can be able to acquire. In order to keep the maximum safety level during the project evolution a three independent stages process have been designed, this allows familiarizing properly with the parts that will be used. The stages are as follows: 1. Test and modeling of all of the parts by little assemblies with through-hole protoboard and stand alone programs. It will be done with a multipurpose card which contains a microcontroller of similar characteristics, although less complex, of the final prototype. 2. Integrating of all of parts through a dedicated design card that will serve as interface between multipurpose card and all the necessary hardware for controlling a ground vehicle with the same characteristics (actuators and engine) of the flying model. 3. Embedded system designing that contains all the developed subsystems in the previous stages and integrates all the necessary parts for controlling a fixed-wing aircraft.

Histograms of oriented gradients for fast on-board vehicle verification

Histograms of Oriented Gradients (HoGs) provide excellent results in object detection and verification. However, their demanding processing requirements bound their applicability in some critical real-time scenarios, such as for video-based on-board vehicle detection systems. In this work, an efficient HOG configuration for pose-based on-board vehicle verification is proposed, which alleviates both the processing requirements and required feature vector length without reducing classification performance. The impact on classification of some critical configuration and processing parameters is in depth analyzed to propose a baseline efficient descriptor. Based on the analysis of its cells contribution to classification, new view-dependent cell-configuration patterns are proposed, resulting in reduced descriptors which provide an excellent balance between performance and computational requirements, rendering higher verification rates than other works in the literature.

Análisis de ataques avanzados y sus técnicas de detección

Los ataques a redes de información son cada vez más sofisticados y exigen una constante evolución y mejora de las técnicas de detección. Para ello, en este proyecto se ha diseñado e implementado una plataforma cooperativa para la detección de intrusiones basada en red. En primer lugar, se ha realizado un estudio teórico previo del marco tecnológico relacionado con este ámbito, en el que se describe y caracteriza el software que se utiliza para realizar ataques a sistemas (malware) así como los métodos que se utilizan para llegar a transmitir ese software (vectores de ataque). En el documento también se describen los llamados APT, que son ataques dirigidos con una gran inversión económica y temporal. Estos pueden englobar todos los malware y vectores de ataque existentes. Para poder evitar estos ataques, se estudiarán los sistemas de detección y prevención de intrusiones, describiendo brevemente los algoritmos que se tienden a utilizar en la actualidad. En segundo lugar, se ha planteado y desarrollado una plataforma en red dedicada al análisis de paquetes y conexiones para detectar posibles intrusiones. Este sistema está orientado a sistemas SCADA (Supervisory Control And Data Adquisition) aunque funciona sobre cualquier red IPv4/IPv6, para ello se definirá previamente lo que es un sistema SCADA, así como sus partes principales. Para implementar el sistema se han utilizado dispositivos de bajo consumo llamados Raspberry PI, estos se ubican entre la red y el equipo final que se quiera analizar. En ellos se ejecutan 2 aplicaciones desarrolladas de tipo cliente-servidor (la Raspberry central ejecutará la aplicación servidora y las esclavas la aplicación cliente) que funcionan de forma cooperativa utilizando la tecnología distribuida de Hadoop, la cual se explica previamente. Mediante esta tecnología se consigue desarrollar un sistema completamente escalable. La aplicación servidora muestra una interfaz gráfica que permite administrar la plataforma de análisis de forma centralizada, pudiendo ver así las alarmas de cada dispositivo y calificando cada paquete según su peligrosidad. El algoritmo desarrollado en la aplicación calcula el ratio de paquetes/tiempo que entran/salen del equipo final, procesando los paquetes y analizándolos teniendo en cuenta la información de señalización, creando diferentes bases de datos que irán mejorando la robustez del sistema, reduciendo así la posibilidad de ataques externos. Para concluir, el proyecto inicial incluía el procesamiento en la nube de la aplicación principal, pudiendo administrar así varias infraestructuras concurrentemente, aunque debido al trabajo extra necesario se ha dejado preparado el sistema para poder implementar esta funcionalidad. En el caso experimental actual el procesamiento de la aplicación servidora se realiza en la Raspberry principal, creando un sistema escalable, rápido y tolerante a fallos. ABSTRACT. The attacks to networks of information are increasingly sophisticated and demand a constant evolution and improvement of the technologies of detection. For this project it is developed and implemented a cooperative platform for detect intrusions based on networking. First, there has been a previous theoretical study of technological framework related to this area, which describes the software used for attacks on systems (malware) as well as the methods used in order to transmit this software (attack vectors). In this document it is described the APT, which are attacks directed with a big economic and time inversion. These can contain all existing malware and attack vectors. To prevent these attacks, intrusion detection systems and prevention intrusion systems will be discussed, describing previously the algorithms tend to use today. Secondly, a platform for analyzing network packets has been proposed and developed to detect possible intrusions in SCADA (Supervisory Control And Data Adquisition) systems. This platform is designed for SCADA systems (Supervisory Control And Data Acquisition) but works on any IPv4 / IPv6 network. Previously, it is defined what a SCADA system is and the main parts of it. To implement it, we used low-power devices called Raspberry PI, these are located between the network and the final device to analyze it. In these Raspberry run two applications client-server developed (the central Raspberry runs the server application and the slaves the client application) that work cooperatively using Hadoop distributed technology, which is previously explained. Using this technology is achieved develop a fully scalable system. The server application displays a graphical interface to manage analytics platform centrally, thereby we can see each device alarms and qualifying each packet by dangerousness. The algorithm developed in the application calculates the ratio of packets/time entering/leaving the terminal device, processing the packets and analyzing the signaling information of each packet, reating different databases that will improve the system, thereby reducing the possibility of external attacks. In conclusion, the initial project included cloud computing of the main application, being able to manage multiple concurrent infrastructure, but due to the extra work required has been made ready the system to implement this funcionality. In the current test case the server application processing is made on the main Raspberry, creating a scalable, fast and fault-tolerant system.

Autonomous classification models in ubiquitous environments

Stream-mining approach is defined as a set of cutting-edge techniques designed to process streams of data in real time, in order to extract knowledge. In the particular case of classification, stream-mining has to adapt its behaviour to the volatile underlying data distributions, what has been called concept drift. Moreover, it is important to note that concept drift may lead to situations where predictive models become invalid and have therefore to be updated to represent the actual concepts that data poses. In this context, there is a specific type of concept drift, known as recurrent concept drift, where the concepts represented by data have already appeared in the past. In those cases the learning process could be saved or at least minimized by applying a previously trained model. This could be extremely useful in ubiquitous environments that are characterized by the existence of resource constrained devices. To deal with the aforementioned scenario, meta-models can be used in the process of enhancing the drift detection mechanisms used by data stream algorithms, by representing and predicting when the change will occur. There are some real-world situations where a concept reappears, as in the case of intrusion detection systems (IDS), where the same incidents or an adaptation of them usually reappear over time. In these environments the early prediction of drift by means of a better knowledge of past models can help to anticipate to the change, thus improving efficiency of the model regarding the training instances needed. By means of using meta-models as a recurrent drift detection mechanism, the ability to share concepts representations among different data mining processes is open. That kind of exchanges could improve the accuracy of the resultant local model as such model may benefit from patterns similar to the local concept that were observed in other scenarios, but not yet locally. This would also improve the efficiency of training instances used during the classification process, as long as the exchange of models would aid in the application of already trained recurrent models, that have been previously seen by any of the collaborative devices. Which it is to say that the scope of recurrence detection and representation is broaden. In fact the detection, representation and exchange of concept drift patterns would be extremely useful for the law enforcement activities fighting against cyber crime. Being the information exchange one of the main pillars of cooperation, national units would benefit from the experience and knowledge gained by third parties. Moreover, in the specific scope of critical infrastructures protection it is crucial to count with information exchange mechanisms, both from a strategical and technical scope. The exchange of concept drift detection schemes in cyber security environments would aid in the process of preventing, detecting and effectively responding to threads in cyber space. Furthermore, as a complement of meta-models, a mechanism to assess the similarity between classification models is also needed when dealing with recurrent concepts. In this context, when reusing a previously trained model a rough comparison between concepts is usually made, applying boolean logic. The introduction of fuzzy logic comparisons between models could lead to a better efficient reuse of previously seen concepts, by applying not just equal models, but also similar ones. This work faces the aforementioned open issues by means of: the MMPRec system, that integrates a meta-model mechanism and a fuzzy similarity function; a collaborative environment to share meta-models between different devices; a recurrent drift generator that allows to test the usefulness of recurrent drift systems, as it is the case of MMPRec. Moreover, this thesis presents an experimental validation of the proposed contributions using synthetic and real datasets.

A yellow fluorescent protein-based assay for high-throughput screening of glycine and GABA(A) receptor chloride channels

There is a significant clinical need to identify novel ligands with high selectivity and potency for GABA(A), GABA(C) and glycine receptor Cl- channels. Two recently developed, yellow fluorescent protein variants (YFP-I152L and YFP-V163S) are highly sensitive to quench by small anions and are thus suited to reporting anionic influx into cells. The aim of this study was to establish the optimal conditions for using these constructs for high-throughput screening of GABA(A), GABA(C) and glycine receptors transiently expressed in HEK293 cells. We found that a 70% fluorescence reduction was achieved by quenching YFP-I152L with a 10 s influx of I- ions, driven by an extemal I- concentration of at least 50 mM. The fluorescence quench was rapid, with a mean time constant of 3 s. These responses were similar for all anion receptor types studied. We also show the assay is sufficiently sensitive to measure agonist and antagonist concentration-responses using either imaging- or photomultiplier-based detection systems. The robustness, sensitivity and low cost of this assay render it suited for high-throughput screening of transiently expressed anionic ligand-gated channels. (c) 2005 Elsevier Ireland Ltd. All rights reserved.

“I didn’t mean to steal someone else’s words!”:a forensic linguistic approach to detecting intentional plagiarism

The concept of plagiarism is not uncommonly associated with the concept of intellectual property, both for historical and legal reasons: the approach to the ownership of ‘moral’, nonmaterial goods has evolved to the right to individual property, and consequently a need was raised to establish a legal framework to cope with the infringement of those rights. The solution to plagiarism therefore falls most often under two categories: ethical and legal. On the ethical side, education and intercultural studies have addressed plagiarism critically, not only as a means to improve academic ethics policies (PlagiarismAdvice.org, 2008), but mainly to demonstrate that if anything the concept of plagiarism is far from being universal (Howard & Robillard, 2008). Even if differently, Howard (1995) and Scollon (1994, 1995) argued, and Angèlil-Carter (2000) and Pecorari (2008) later emphasised that the concept of plagiarism cannot be studied on the grounds that one definition is clearly understandable by everyone. Scollon (1994, 1995), for example, claimed that authorship attribution is particularly a problem in non-native writing in English, and so did Pecorari (2008) in her comprehensive analysis of academic plagiarism. If among higher education students plagiarism is often a problem of literacy, with prior, conflicting social discourses that may interfere with academic discourse, as Angèlil-Carter (2000) demonstrates, we then have to aver that a distinction should be made between intentional and inadvertent plagiarism: plagiarism should be prosecuted when intentional, but if it is part of the learning process and results from the plagiarist’s unfamiliarity with the text or topic it should be considered ‘positive plagiarism’ (Howard, 1995: 796) and hence not an offense. Determining the intention behind the instances of plagiarism therefore determines the nature of the disciplinary action adopted. Unfortunately, in order to demonstrate the intention to deceive and charge students with accusations of plagiarism, teachers necessarily have to position themselves as ‘plagiarism police’, although it has been argued otherwise (Robillard, 2008). Practice demonstrates that in their daily activities teachers will find themselves being required a command of investigative skills and tools that they most often lack. We thus claim that the ‘intention to deceive’ cannot inevitably be dissociated from plagiarism as a legal issue, even if Garner (2009) asserts that generally plagiarism is immoral but not illegal, and Goldstein (2003) makes the same severance. However, these claims, and the claim that only cases of copyright infringement tend to go to court, have recently been challenged, mainly by forensic linguists, who have been actively involved in cases of plagiarism. Turell (2008), for instance, demonstrated that plagiarism is often connoted with an illegal appropriation of ideas. Previously, she (Turell, 2004) had demonstrated by comparison of four translations of Shakespeare’s Julius Caesar to Spanish that the use of linguistic evidence is able to demonstrate instances of plagiarism. This challenge is also reinforced by practice in international organisations, such as the IEEE, to whom plagiarism potentially has ‘severe ethical and legal consequences’ (IEEE, 2006: 57). What plagiarism definitions used by publishers and organisations have in common – and which the academia usually lacks – is their focus on the legal nature. We speculate that this is due to the relation they intentionally establish with copyright laws, whereas in education the focus tends to shift from the legal to the ethical aspects. However, the number of plagiarism cases taken to court is very small, and jurisprudence is still being developed on the topic. In countries within the Civil Law tradition, Turell (2008) claims, (forensic) linguists are seldom called upon as expert witnesses in cases of plagiarism, either because plagiarists are rarely taken to court or because there is little tradition of accepting linguistic evidence. In spite of the investigative and evidential potential of forensic linguistics to demonstrate the plagiarist’s intention or otherwise, this potential is restricted by the ability to identify a text as being suspect of plagiarism. In an era with such a massive textual production, ‘policing’ plagiarism thus becomes an extraordinarily difficult task without the assistance of plagiarism detection systems. Although plagiarism detection has attracted the attention of computer engineers and software developers for years, a lot of research is still needed. Given the investigative nature of academic plagiarism, plagiarism detection has of necessity to consider not only concepts of education and computational linguistics, but also forensic linguistics. Especially, if intended to counter claims of being a ‘simplistic response’ (Robillard & Howard, 2008). In this paper, we use a corpus of essays written by university students who were accused of plagiarism, to demonstrate that a forensic linguistic analysis of improper paraphrasing in suspect texts has the potential to identify and provide evidence of intention. A linguistic analysis of the corpus texts shows that the plagiarist acts on the paradigmatic axis to replace relevant lexical items with a related word from the same semantic field, i.e. a synonym, a subordinate, a superordinate, etc. In other words, relevant lexical items were replaced with related, but not identical, ones. Additionally, the analysis demonstrates that the word order is often changed intentionally to disguise the borrowing. On the other hand, the linguistic analysis of linking and explanatory verbs (i.e. referencing verbs) and prepositions shows that these have the potential to discriminate instances of ‘patchwriting’ and instances of plagiarism. This research demonstrates that the referencing verbs are borrowed from the original in an attempt to construct the new text cohesively when the plagiarism is inadvertent, and that the plagiarist has made an effort to prevent the reader from identifying the text as plagiarism, when it is intentional. In some of these cases, the referencing elements prove being able to identify direct quotations and thus ‘betray’ and denounce plagiarism. Finally, we demonstrate that a forensic linguistic analysis of these verbs is critical to allow detection software to identify them as proper paraphrasing and not – mistakenly and simplistically – as plagiarism.

Learning and Exploiting Camera Geometry for Computer Vision

This work explores the use of statistical methods in describing and estimating camera poses, as well as the information feedback loop between camera pose and object detection. Surging development in robotics and computer vision has pushed the need for algorithms that infer, understand, and utilize information about the position and orientation of the sensor platforms when observing and/or interacting with their environment.

The first contribution of this thesis is the development of a set of statistical tools for representing and estimating the uncertainty in object poses. A distribution for representing the joint uncertainty over multiple object positions and orientations is described, called the mirrored normal-Bingham distribution. This distribution generalizes both the normal distribution in Euclidean space, and the Bingham distribution on the unit hypersphere. It is shown to inherit many of the convenient properties of these special cases: it is the maximum-entropy distribution with fixed second moment, and there is a generalized Laplace approximation whose result is the mirrored normal-Bingham distribution. This distribution and approximation method are demonstrated by deriving the analytical approximation to the wrapped-normal distribution. Further, it is shown how these tools can be used to represent the uncertainty in the result of a bundle adjustment problem.

Another application of these methods is illustrated as part of a novel camera pose estimation algorithm based on object detections. The autocalibration task is formulated as a bundle adjustment problem using prior distributions over the 3D points to enforce the objects' structure and their relationship with the scene geometry. This framework is very flexible and enables the use of off-the-shelf computational tools to solve specialized autocalibration problems. Its performance is evaluated using a pedestrian detector to provide head and foot location observations, and it proves much faster and potentially more accurate than existing methods.

Finally, the information feedback loop between object detection and camera pose estimation is closed by utilizing camera pose information to improve object detection in scenarios with significant perspective warping. Methods are presented that allow the inverse perspective mapping traditionally applied to images to be applied instead to features computed from those images. For the special case of HOG-like features, which are used by many modern object detection systems, these methods are shown to provide substantial performance benefits over unadapted detectors while achieving real-time frame rates, orders of magnitude faster than comparable image warping methods.

The statistical tools and algorithms presented here are especially promising for mobile cameras, providing the ability to autocalibrate and adapt to the camera pose in real time. In addition, these methods have wide-ranging potential applications in diverse areas of computer vision, robotics, and imaging.