Plants as sessile organisms are susceptible to many environmental stresses such as drought, and salinity. They have therefore evolved mechanisms to acclimate and tolerate environmental stresses. Knowledge of the molecular aspects of abiotic stress gleaned from extensive studies in Arabidopsis has provided much information on the complex processes underlying plant response to abiotic stresses. Nevertheless, there is a need for integration of the knowledge gained and a systematic molecular genetic dissection of the complex responses to abiotic stress. In this study in Arabidopsis, comparative expression profiling analysis of progressive (pDr) and moderate (mDr) drought treatments revealed common drought responses, as well as treatment specific signatures responses to drought stress. Under prolonged moderate drought plants develop different mechanisms for acclimation: induction of cell wall loosening at early stage, and a change in hormonal balance (ABA: JA) at late stage of moderate drought. Taking a reverse genetics approach, a MYB transcription factor (MYB109) has been identified as a regulator of growth under drought and salt stress. Global expression profiling showed possible mechanisms of how MYB109 modulates growth under drought conditions: as a regulator of RNA processing and splicing and as a negative regulator of jasmonic acid biosynthesis and signaling. A forward genetics screen for drought and salt tolerance of transposon activation tag (ATag) lines led to the discovery of novel genes, which shed light on unexplored areas of abiotic stress biology. Utilizing this strategy, a potential role for cell wall modification and MATE transporters in response to drought and salt stress has been discovered, which needs further analysis to integrate this information on the role of these biological processes in plant stress biology. / Ph. D.

Key Words: Thrombosis, Vascular Graft, Microfluidics, Wall Shear Stress / Vascular grafts are essential for the management of cardiovascular disease. However, the lifesaving potential of these devices is undermined by thrombosis arising from material and flow interactions on the blood contacting surface. To combat this issue, the use of antithrombogenic coatings has emerged as a promising strategy for modulating blood and graft interaction in vivo. Although an important determinant of graft performance, hemodynamics are frequently overlooked in the in vitro testing of coatings and their translatability remains poorly understood. We address this limitation with a microscale platform that incorporates vascular prosthesis and coatings with tuneable flow and surface conditions in vitro. As a proof of concept, we use the platform to test the thrombogenic performance of a novel class of lubricant infused (LIS) and antibody lubricant infused (anti-CD34 LIS) coated ePTFE vascular grafts in the presence of arterial wall shear stress, with and without the presence of endothelial cells. Our findings suggest lubricant infused coated ePTFE vascular grafts are thromboresistant under flow and may have potential for in vivo arterial grafting applications. It is moreover apparent that the microscale properties of the device could be advantageous for the testing and translation of novel antithrombogenic coatings or blood contacting prosthesis in general. / Thesis / Master of Applied Science (MASc)

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The biosynthetic enzymes involved in wall teichoic acid biogenesis in Grampositive bacteria have been the subject of renewed investigation in recent years with the benefit of modem tools of biochemistry and genetics. Nevertheless, there have been only limited investigations into the enzymes that glycosylate wall teichoic acid. Decades-old experiments in the model Gram-positive bacterium, Bacillus subtilis 168, using phage resistant mutants implicated tagE (also called gtaA and rodD) as the gene coding for the wall teichoic acid glycosyltransferase. This study and others have provided only indirect evidence to support a role for TagE in wall teichoic acid glycosylation. In this work, we showed that deletion of tagE results in the loss of a-glucose at the C-2 position of glycerol in the poly(glycerol phosphate) polymer backbone. We also report the first kinetic characterization of pure, recombinant wall teichoic acid glycosyltransferase using clean synthetic substrates. We investigated the substrate specificity ofTagE using a wide variety of acceptor substrates and showed that this enzyme has a strong kinetic preference for the transfer of glucose from UDP-glucose to glycerol phosphate in polymeric form. Further, we showed that the enzyme recognizes its polymeric (and repetitive) substrate with a sequential kinetic mechanism. This work provides direct evidence that TagE is the wall teichoic acid glycosyltransferase in B. subtilis 168 and provides a strong basis for further studies on the mechanism of wall teichoic acid glycosylation, a largely uncharted aspect of wall teichoic acid biogenesis. / Thesis / Doctor of Philosophy (PhD)

<p> The increase in microbes resistant to a wide array of antibiotics has led to the need for the development of novel antimicrobials. However in order to develop new antimicrobials, novel pathways need to be targeted. Teichoic acid is an anionic polymer covalently attached to the cell wall of Gram-positive bacteria. Recent research has demonstrated that teichoic acid genes are indispensable to the viability of Bacillus subtilis. This makes teichoic acid biosynthetic proteins ideal candidates for the development of a new antimicrobial. Of the teichoic acid glycerol phosphate (tag) genes involved in the biosynthesis of teichoic acid in B. subtilis 168, a conditional deletion mutant of tagD, whose protein product encodes the proposed glycerol-3-phosphatecytidylyltransferase, has been previously constructed and was shown to have a lethal phenotype upon depletion of TagD. This was used in a microarray analysis to find genes that were transcriptionally up-regulated upon the depletion of TagD in B. subtilis 168. Ten candidate genes were selected from those up-regulated and used in the design of a novel, real-time, cell-based luminescent reporter system that responds to lesions in wall biosynthesis. Characterization of these reporter systems in tag gene deletion backgrounds and an examination of their response to antibiotics of various mechanism of action led to the identification of our candidate reporter system P ywac, a robust reporter of both lesions in teichoic acid and peptidoglycan synthesis. In a proof-of-principle screen, the use of Pywac as a reporter of lesions in the cell wall was validated. This reporter system is unique in that it combines conventional genetics with a high throughput capacity. It will not only be amenable for screening small molecules to find inhibitors that impinge on teichoic acid biosynthesis, but it can also be used to probe genetic interactions in B. subtilis. </p> / Thesis / Master of Science (MSc)

<p> Atherosclerosis generally occurs near the branching in the arteries where there tends to be flow irregularities. A build up of fatty deposits (plaque) occurs in the blood vessel in such regions making it to lose its elasticity. Such hardening of the arteries and the narrowing of the lumen can cause severe atheromas and even high blood pressure and blockage of the vessels. It is observed in North America that nearly 47% of the deaths are caused due to cardiovascular diseases and hence determination of such regions becomes very critical and can be very beneficial if done at an earlier stage. In this thesis, we present: an approach to model the pulsating flow of blood through such an atherosclerosis affected region of the artery using finite element method and further discuss the statistical model used to implement the optimization techniques to estimate the region of maximum rigidity. Here within we present a numerical and non-invasive approach to predict such regions. The computational modeling is carried out under two categories: a. The mathematical model and b. The statistical model. </p> <p> The mathematical model which is the forward model, comprises of the artery and the cardiac muscle as hyperelastic material modeled with the neo-hookean model and the three dimensional Navier-Stokes equations solve for the blood flowing through it. We perform fluid dynamic analysis for the blood flowing through the vessel to compute the velocity at different time instances and mechanical analysis to compute the deformation of the artery which is a function of the elasticity of the vessel. The two models are interconnected to each other by boundary conditions as the normal component of the surface force provides the coupling between the two models. The shear modulus represents a measure of the elasticity of the vessel. We use linear spatial basis functions to model the shear modulus which spatially varies along the geometry of the vessel thus we have a region of atherosclerosis and the geometry shows the stenosis. The change in the shear modulus affects the velocity of blood through the vessel. </p> <p> In the statistical model, we propose an inverse computational model for estimating the elasticity profile of the arterial wall where we implement the inverse modeling approach to estimate the maximum shear modulus which helps us to predict the region of atherosclerosis. The velocity and the deformation obtained for a particular shear modulus from our COMSOL forward model provide the realistic simulated measurements that are made noisy by introduction of white Gaussian noise with different SNR and we try to estimate the shear modulus that minimizes the error-function. We use COMSOL with MATLAB for simultaneous iterative computations of velocity and deformation measurements by running the optimization code. We estimate these unknown parameters using optimization algorithm that minimizes the cost function of our model. For our estimation we use the least squares estimator and we derive the maximum likelihood estimator. The unconstrained optimization is carried out with Neider Mead Simplex Method and the Trust Region Method which uses only the function evaluations to find the minimum: making it a very robust algorithm and very efficient for problems that are nonlinear or have a number of discontinuities. Our preliminary results demonstrate significant change in velocity of the blood and occurrence of vortices in the region of less elasticity and the tendency of the artery to deform minimum in the hardened less elastic region. Our estimation results show that the parameters are identifiable. The mean square error of the estimate as a function of SNR shows accuracy of the estimation. </p> / Thesis / Master of Applied Science (MASc)

Turbulent flow over a rough surface produces sound that radiates outside the near wall region. This noise source is often at a lower level than the noise created by edges and bluff body flows, but for applications with large surface area to perimeter ratios at low Mach number, this noise source can have considerable levels. In the first part of this dissertation, a detailed study is made of the ability of the Glegg & Devenport (2009) scattering theory to predict roughness noise. To this end, comparisons are made with measurements from cuboidal and hemispherical roughness with roughness Reynolds numbers, hu_Ï /ν, ranging from 24 to 197 and roughness height to boundary layer thickness ratios of 5 to 18. Their theory is shown to work very accurately to predict the noise from surfaces with large roughness Reynolds numbers, but for cases with highly inhomogeneous wall pressure fields, differences grow between estimation and measurement. For these surfaces, the absolute levels were underpredicted but the spectral shape of the measurement was correctly determined indicating that the relationship of the radiated noise with the wavenumber wall pressure spectrum and roughness geometry appears to remain relatively unchanged. In the second part of this dissertation, delay and sum beamforming and least-squares analyses were used to examine roughness noise recorded by a 36-sensor linear microphone array. These methods were employed to estimate the variation of source strengths through short fetches of large hemispherical and cuboidal element roughness. The analyses show that the lead rows of the fetches produced the greatest streamwise and spanwise noise radiation. The least-squares analysis confirmed the presence of streamwise and spanwise aligned dipoles emanating from each roughness element as suggested by the LES of Yang & Wang (2011). The least-squares calculated source strengths show that the streamwise aligned dipole is always stronger than that of the spanwise dipole, but the relative magnitude of the difference varies with frequency. / Ph. D.