Towards the prediction of mutations in genomic sequences

Bio-systems are inherently complex information processing systems. Furthermore, physiological complexities of biological systems limit the formation of a hypothesis in terms of behavior and the ability to test hypothesis. More importantly the identification and classification of mutation in patients are centric topics in today's cancer research. Next generation sequencing (NGS) technologies can provide genome-wide coverage at a single nucleotide resolution and at reasonable speed and cost. The unprecedented molecular characterization provided by NGS offers the potential for an individualized approach to treatment. These advances in cancer genomics have enabled scientists to interrogate cancer-specific genomic variants and compare them with the normal variants in the same patient. Analysis of this data provides a catalog of somatic variants, present in tumor genome but not in the normal tissue DNA. In this dissertation, we present a new computational framework to the problem of predicting the number of mutations on a chromosome for a certain patient, which is a fundamental problem in clinical and research fields. We begin this dissertation with the development of a framework system that is capable of utilizing published data from a longitudinal study of patients with acute myeloid leukemia (AML), who's DNA from both normal as well as malignant tissues was subjected to NGS analysis at various points in time. By processing the sequencing data at the time of cancer diagnosis using the components of our framework, we tested it by predicting the genomic regions to be mutated at the time of relapse and, later, by comparing our results with the actual regions that showed mutations (discovered at relapse time). We demonstrate that this coupling of the algorithm pipeline can drastically improve the predictive abilities of searching a reliable molecular signature. Arguably, the most important result of our research is its superior performance to other methods like Radial Basis Function Network, Sequential Minimal Optimization, and Gaussian Process. In the final part of this dissertation, we present a detailed significance, stability and statistical analysis of our model. A performance comparison of the results are presented. This work clearly lays a good foundation for future research for other types of cancer.^

New Organogermanium Substrates for Palladium-Catalyzed Cross-Coupling Reactions. Application of Organogermanes towards the Synthesis of Carbon-5 Modified Uridine Analogues

The diverse biological properties exhibited by uridine analogues modified at carbon-5 of the uracil base have attracted special interest to the development of efficient methodologies for their synthesis. This study aimed to evaluate the possible application of vinyl tris(trimethylsilyl)germanes in the synthesis of conjugated 5-modified uridine analogues via Pd-catalyzed cross-coupling reactions. The stereoselective synthesis of 5-[(2-tris(trimethylsilyl)germyl)ethenyl]uridine derivatives was achieved by the radical-mediated hydrogermylation of the protected 5-alkynyluridine precursors with tris(trimethylsilyl)germane [(TMS)3GeH]. The hydrogermylation with Ph3GeH afforded in addition to the expected 5-vinylgermane, novel 5-(2-triphenylgermyl)acetyl derivatives. Also, the treatment with Me3GeH provided access to 5-vinylgermane uridine analogues with potential biological applications. Since the Pd-catalyzed cross-coupling of organogermanes has received much less attention than the couplings involving organostannanes and organosilanes, we were prompted to develop novel organogermane precursors suitable for transfer of aryl and/or alkenyl groups. The allyl(phenyl)germanes were found to transfer allyl groups to aryl iodides in the presence of sodium hydroxide or tetrabutylammonium fluoride (TBAF) via a Heck arylation mechanism. On the other hand, the treatment of allyl(phenyl)germanes with tetracyanoethylene (TCNE) effectively cleaved the Ge-C(allyl) bonds and promoted the transfer of the phenyl groups upon fluoride activation in toluene. It was discovered that the trichlorophenyl,- dichlorodiphenyl,- and chlorotriphenylgermanes undergo Pd-catalyzed cross-couplings with aryl bromides and iodides in the presence of TBAF in toluene with addition of the measured amount of water. One chloride ligand on the Ge center allows efficient activation by fluoride to promote transfer of one, two or three phenyl groups from the organogermane precursors. The methodology shows that organogermanes can render a coupling efficiency comparable to the more established stannane and silane counterparts. Our coupling methodology (TBAF/moist toluene) was also found to promote the transfer of multiple phenyl groups from analogous chloro(phenyl)silanes and stannanes.

A study of methods used to analyze total oil and polycyclic aromatic hydrocarbons in produced water: steps towards the validation of molecularly imprinted polymers for use in marine environmetns

Produced water is a by-product of offshore oil and gas production, and is released in large volumes when platforms are actively processing crude oil. Some pollutants are not typically removed by conventional oil/water separation methods and are discharged with produced water. Oil and grease can be found dispersed in produced water in the form of tiny droplets, and polycyclic aromatic hydrocarbons (PAHs) are commonly found dissolved in produced water. Both can have acute and chronic toxic effects in marine environments even at low exposure levels. The analysis of the dissolved and dispersed phases are a priority, but effort is required to meet the necessary detection limits. There are several methods for the analysis of produced water for dispersed oil and dissolved PAHs, all of which have advantages and disadvantages. In this work, EPA Method 1664 and APHA Method 5520 C for the determination of oil and grease will be examined and compared. For the detection of PAHs, EPA Method 525 and PAH MIPs will be compared, and results evaluated. APHA Method 5520 C Partition-Infrared Method is a liquid-liquid extraction procedure with IR determination of oil and grease. For analysis on spiked samples of artificial seawater, extraction efficiency ranged from 85 – 97%. Linearity was achieved in the range of 5 – 500 mg/L. This is a single-wavelength method and is unsuitable for quantification of aromatics and other compounds that lack sp³-hybridized carbon atoms. EPA Method 1664 is the liquid-liquid extraction of oil and grease from water samples followed by gravimetric determination. When distilled water spiked with reference oil was extracted by this procedure, extraction efficiency ranged from 28.4 – 86.2%, and %RSD ranged from 7.68 – 38.0%. EPA Method 525 uses solid phase extraction with analysis by GC-MS, and was performed on distilled water and water from St. John’s Harbour, all spiked with naphthalene, fluorene, phenanthrene, and pyrene. The limits of detection in harbour water were 0.144, 3.82, 0.119, and 0.153 g/L respectively. Linearity was obtained in the range of 0.5-10 g/L, and %RSD ranged from 0.36% (fluorene) to 46% (pyrene). Molecularly imprinted polymers (MIPs) are sorbent materials made selective by polymerizing functional monomers and crosslinkers in the presence of a template molecule, usually the analytes of interest or related compounds. They can adsorb and concentrate PAHs from aqueous environments and are combined with methods of analysis including GC-MS, LC-UV-Vis, and desorption electrospray ionization (DESI)- MS. This work examines MIP-based methods as well as those methods previously mentioned which are currently used by the oil and gas industry and government environmental agencies. MIPs are shown to give results consistent with other methods, and are a low-cost alternative improving ease, throughput, and sensitivity. PAH MIPs were used to determine naphthalene spiked into ASTM artificial seawater, as well as produced water from an offshore oil and gas operation. Linearity was achieved in the range studied (0.5 – 5 mg/L) for both matrices, with R² = 0.936 for seawater and R² = 0.819 for produced water. The %RSD for seawater ranged from 6.58 – 50.5% and for produced water, from 8.19 – 79.6%.

Towards the design of monolithic decoupled XYZ compliant parallel mechanisms for multi-function applications

This paper deals with the monolithic decoupled XYZ compliant parallel mechanisms (CPMs) for multi-function applications, which can be fabricated monolithically without assembly and has the capability of kinetostatic decoupling. At first, the conceptual design of monolithic decoupled XYZ CPMs is presented using identical spatial compliant multi-beam modules based on a decoupled 3-PPPR parallel kinematic mechanism. Three types of applications: motion/positioning stages, force/acceleration sensors and energy harvesting devices are described in principle. The kinetostatic and dynamic modelling is then conducted to capture the displacements of any stage under loads acting at any stage and the natural frequency with the comparisons with FEA results. Finally, performance characteristics analysis for motion stage applications is detailed investigated to show how the change of the geometrical parameter can affect the performance characteristics, which provides initial optimal estimations. Results show that the smaller thickness of beams and larger dimension of cubic stages can improve the performance characteristics excluding natural frequency under allowable conditions. In order to improve the natural frequency characteristic, a stiffness-enhanced monolithic decoupled configuration that is achieved through employing more beams in the spatial modules or reducing the mass of each cubic stage mass can be adopted. In addition, an isotropic variation with different motion range along each axis and same payload in each leg is proposed. The redundant design for monolithic fabrication is introduced in this paper, which can overcome the drawback of monolithic fabrication that the failed compliant beam is difficult to replace, and extend the CPM’s life.

Attitudes towards the Marine Environment and Implications for Marine Resource Management in Seward, Alaska

This research, conducted in 2006-2008, examines the ways in which various groups involved with the marine resources of Seward, Alaska construct attitudes towards the environment. Participant observation and semi-structured interviews are used to assess how commercial halibut fishers, tour boat operators, local residents and government officials understand the marine environment based on their previous experiences. This study also explores how ideologies relate to the current practices of each group. Two theories orient the analyses: The first, cultural modeling provided a theoretical and methodological framework for pursuing a more comprehensive analysis of resource management. The second, Theory of Reasoned Action (Ajzen and Fishbein 1980), guided the analysis of the ways in which each participant’s ideology towards the marine environment relates to their practice. Aside from contributing to a better understanding of a coastal community’s ideologies and practices, this dissertation sought to better understand the role of ecological ideologies and behaviors in fisheries management. The research illustrates certain domains where ideologies and practices concerning Pacific halibut and the marine environment differ among commercial fishers, government, and management officials, tour boat operators and residents of Seward, AK. These differences offer insights into how future collaborative efforts between government officials, managers and local marine resource users might better incorporate local ideology into management, and provide ecological information to local marine resource users in culturally appropriate ways.

Temporal variation of temperature and related proxies from the penultimate glacial towards the Eemian in the Black Sea

The last glacial-interglacial transition or Termination I (T I) is well documented in the Black Sea, whereas little is known about climate and environmental dynamics during the penultimate Termination (T II). Here we present a multi-proxy study based on a sediment core from the SE Black Sea covering the penultimate glacial and almost the entire Eemian interglacial (133.5 ±0.7-122.5 ±1.7 ka BP). Proxies comprise ice-rafted debris (IRD), O and Sr isotopes as well as Sr/Ca, Mg/Ca, and U/Ca ratios of benthic ostracods, organic and inorganic sediment geochemistry, as well as TEX86 and UK'37derived water temperatures. The ending penultimate glacial (MIS 6, 133.5 to 129.9 ±0.7 ka BP) is characterised by mean annual lake surface temperatures of about 9°C as estimated from the TEX86 palaeothermometer. This period is impacted by two Black Sea melt water pulses (BSWP-II-1 and 2) as indicated by very low Sr/Ca ostracods but high sedimentary K/Al values. Anomalously high radiogenic 87Sr/86Sr ostracod values (max. 0.70945) during BSWP-II-2 suggest a potential Himalayan source communicated via the Caspian Sea. The T II warming started at 129.9 ±0.7 ka BP, witnessed by abrupt disappearance of IRD, increasing d18O ostracod values, and a first TEX86 derived temperature rise of about 2.5°C. A second, abrupt warming step to ca. 15.5°C as the prelude of the Eemian warm period is documented at 128.3 ka BP. The Mediterranean-Black Sea reconnection most likely occurred at 128.1 ±0.7 ka BP as demonstrated by increasing Sr/Ca ostracods and U/Ca ostracods values. The disappearance of ostracods and TOC contents >2% document the onset of Eemian sapropel formation at 127.6 ka BP. During sapropel formation, TEX86 temperatures dropped and stabilised at around 9°C, while UK'37 temperatures remain on average 17°C. This difference is possibly caused by a habitat shift of Thaumarchaeota communities from surface towards nutrient-rich deeper and colder waters located above the gradually establishing halo-and redoxcline.

Karst hazard analysis towards the development of predictive karst models for southern Ontario

An investigation into karst hazard in southern Ontario has been undertaken with the intention of leading to the development of predictive karst models for this region. The reason these are not currently feasible is a lack of sufficient karst data, though this is not entirely due to the lack of karst features. Geophysical data was collected at Lake on the Mountain, Ontario as part of this karst investigation. This data was collected in order to validate the long-standing hypothesis that Lake on the Mountain was formed from a sinkhole collapse. Sub-bottom acoustic profiling data was collected in order to image the lake bottom sediments and bedrock. Vertical bedrock features interpreted as solutionally enlarged fractures were taken as evidence for karst processes on the lake bottom. Additionally, the bedrock topography shows a narrower and more elongated basin than was previously identified, and this also lies parallel to a mapped fault system in the area. This suggests that Lake on the Mountain was formed over a fault zone which also supports the sinkhole hypothesis as it would provide groundwater pathways for karst dissolution to occur. Previous sediment cores suggest that Lake on the Mountain would have formed at some point during the Wisconsinan glaciation with glacial meltwater and glacial loading as potential contributing factors to sinkhole development. A probabilistic karst model for the state of Kentucky, USA, has been generated using the Weights of Evidence method. This model is presented as an example of the predictive capabilities of these kind of data-driven modelling techniques and to show how such models could be applied to karst in Ontario. The model was able to classify 70% of the validation dataset correctly while minimizing false positive identifications. This is moderately successful and could stand to be improved. Finally, suggestions to improving the current karst model of southern Ontario are suggested with the goal of increasing investigation into karst in Ontario and streamlining the reporting system for sinkholes, caves, and other karst features so as to improve the current Ontario karst database.

The European Union Foreign Policy in the Caucasus and Central Asia: The Limitations of the Opening towards the East

This article intends to study the evolution of the European Union foreign policy in the Southern Caucasus and Central Area throughout the Post-Cold War era. The aim is to analyze Brussels’ fundamental interests and limitations in the area, the strategies it has implemented in the last few years, and the extent to which the EU has been able to undermine the regional hegemons’ traditional supremacy. As will be highlighted, the Community’s chronic weaknesses, the local determination to preserve sovereignty and an increasing international geopolitical competition undermine any European aspiration to become a pre-eminent actor at the heart of the Eurasian continent in the near future.

Stimulating Neoblast-Like Cell Proliferation in Juvenile Fasciola hepatica Supports Growth and Progression towards the Adult Phenotype In Vitro

Fascioliasis (or fasciolosis) is a socioeconomically important parasitic disease caused by liver flukes of the genus Fasciola. Flukicide resistance has exposed the need for new drugs and/or a vaccine for liver fluke control. A rapidly improving 'molecular toolbox' for liver fluke encompasses quality genomic/transcriptomic datasets and an RNA interference platform that facilitates functional genomics approaches to drug/vaccine target validation. The exploitation of these resources is undermined by the absence of effective culture/maintenance systems that would support in vitro studies on juvenile fluke development/biology. Here we report markedly improved in vitro maintenance methods for Fasciola hepatica that achieved 65% survival of juvenile fluke after 6 months in standard cell culture medium supplemented with 50% chicken serum. We discovered that this long-term maintenance was dependent upon fluke growth, which was supported by increased proliferation of cells resembling the "neoblast" stem cells described in other flatworms. Growth led to dramatic morphological changes in juveniles, including the development of the digestive tract, reproductive organs and the tegument, towards more adult-like forms. The inhibition of DNA synthesis prevented neoblast-like cell proliferation and inhibited growth/development. Supporting our assertion that we have triggered the development of juveniles towards adult-like fluke, mass spectrometric analyses showed that growing fluke have an excretory/secretory protein profile that is distinct from that of newly-excysted juveniles and more closely resembles that of ex vivo immature and adult fluke. Further, in vitro maintained fluke displayed a transition in their movement from the probing behaviour associated with migrating stage worms to a slower wave-like motility seen in adults. Our ability to stimulate neoblast-like cell proliferation and growth in F. hepatica underpins the first simple platform for their long-term in vitro study, complementing the recent expansion in liver fluke resources and facilitating in vitro target validation studies of the developmental biology of liver fluke.

Investigations on selected chalcogenide glasses towards the realization of photonic devices

Among various optical sensing schemes, infrared spectroscopy is a powerful tool for detecting and determining the composition of complex organic samples since vibrational finger prints of all biomolecules and organic species are located in this window. This spectroscopic technique is simple, reliable, fast, non-destructive, cost-effective while having low sensitivity. Use of metallic nanoparticles in association with a good IR transparent sensing substrate, is one of the promising solutions to enhance the sensitivity. Chalcogenide glasses are promising substrate material because of their extended optical transmission window starting from the visible to the far infrared range up to 20 μm, high refractive index usually between 2 and 3 and high optical nonlinearity, which make them good candidates as IR sensors and optical ultrafast nonlinear devices. These glasses are favorable sensor materials for the infrared spectral range because of their high IR transparency to allow for low optical loss at wavelengths corresponding to the characteristic optical absorption bands of organic molecules, high refractive index for tight confinement of optical energy within the resonator structure, processibility into thin film form, chemical compatibility for adhesion of silver nano particles and thin films and resistance to the chemical environment to be sensed. Molecules adsorbed to silver island structures shows enhanced IR absorption spectra and the extent of enhancement is determined by many factors such as the size, density and morphology of silver structures, optical and dielectric properties of the substrate material etc.