Tuberculosis (TB) is an important disease worldwide. Currently, one-third of the world’s population is infected with TB, and it is a leading cause of death among people living with HIV. Immediate but also accurate diagnosis is required for disease control, yet available diagnostics cannot do both simultaneously. Therefore, designing a technique that can diagnose the disease correctly in the shortest possible time is in great demand in order to stop its spread. Diffraction-based sensing is a novel technique for measuring of biomolecular interaction that has potential for disease diagnosis. In this study, diffraction-based sensing successfully demonstrated its usefulness for diagnostics of TB using recombinant TB antigen, or by detection of interferon-γ that is produced from white blood cells when the immune system activates. The feasibility of the technology was also evaluated in terms of providing real time observation, reducing diagnostic duration, and increasing sensitivity of detection.

This dissertation combines aspects of lasers, nonlinear optics and interferometry to measure the linear optical properties involved in phase matched second harmonic generation, (SHG). A new measuring technique has been developed to rapidly analyze the phase matching performance of potential SHGs. The data taken is in the form of interferograms produced by the self referencing nonlinear Fizeau interferometer (NLF), and correctly predicts when phase matched SHG will occur in the sample wedge. Data extracted from the interferograms produced by the NLF, allows us to predict both phase matching temperatures for noncritically phase matchable crystals and crystal orientation for angle tuned crystals. Phase matching measurements can be made for both Type I and Type II configurations. Phase mismatch measurements were made at the fundamental wavelength of 1.32 (mu)m, for: calcite, lithium niobate, and gadolinium molybdate (GMO). Similar measurements were made at 1.06 (mu)m. for calcite. Phase matched SHG was demonstrated in calcite, lithium niobate and KTP, while phase matching by temperature tuning is ruled out for GMO.

Master of Science / Department of Diagnostic Medicine/Pathobiology / Sanjeev K. Narayanan / Antibiotics are substances produced by bacteria or fungi that are inhibitory to other bacteria and fungi. Antimicrobial compounds include substances that are naturally produced, chemically modified or completely synthetic (chemically designed or synthesized). The chemical modification of naturally produced antibiotic generally results in increase stability, solubility, increased spectrum of activity, or efficacy. Antimicrobial compounds are used in animals to treat and control infectious diseases, and also for growth promotion. Bacteria may gain resistance to antibacterial agents via a variety of mechanisms. There is growing evidence that antimicrobial resistance has significant public health consequences. Rationale use of antimicrobial drugs using appropriate medication at the proper dosage and for duration is one of the important means to reduce selective pressure that helps reduce life of resistant organism. It is also vital to reduce the spread of multi drug resistant organisms in the environment especially in health care facilities. Bacteria evolve rapidly not only by mutation, but also by horizontal gene transfer through the transformation, transduction, and conjugation. Conjugation involves a close contact between two bacteria and transfer of the plasmid that carry many genetic elements. The pathogenic bacteria have the ability to sense as well as respond to the stress in the recipient. The epinephrine and norepinephrine play a key role in stress situations in animals. A previous study showed that norepinephrine (NE), a catecholamine at physiological concentrations promoted the conjugation efficiencies of a conjugative plasmid from a clinical strain of Salmonella typhimurium to an E. coli recipient in vitro. The objective of this study was to determine the effect of norepinephrine on conjugation of two E. coli strains. Both filter mating and liquid mating assays were used. The results revealed that there was no significance difference between the presence and the absence of norepinephrine on conjugative transfer of RP4 plasmid between E. coli strains (FS1290 and C600N) either in filter mating or liquid mating. Further studies are needed to determine whether higher concentration of (more than 20 mM) has any effects on conjugation in E. coli.

As vacinas conjugadas contra Streptococcus pneumoniae mostram-se eficazes na redução da doença pneumocócica, no entanto depois de sua introdução tem se verificado a substituição de sorotipos. Para evitar estes problemas o emprego de proteínas da própria bactéria como carreadoras tem sido usado. Neste trabalho a PspA foi conjugada ao sorotipo 6B de S. pneumoniae por dois métodos de conjugação. Inicialmente a PspA clado 1 e 3 foram purificadas, recuperando as proteínas com mais de 90% de pureza. A conjugação intermediada pelo agente ativador cloreto de 4-(4,6-dimetoxi-1,3,5-triazin-2-il-)-4-metilmorfolino (DMT-MM) permitiu rendimentos de Ps entre 20 e 30%, no entanto este tipo de conjugado não foi imunogênico resultando na não indução de anticorpos anti-PspA. Por outro lado a conjugação por aminação redutiva possibilitou obter rendimentos de Ps entre 50 e 60% e os conjugados induziram elevados títulos de anticorpos anti-PspA em camundongos Balb/c, que foram capazes de promover a fagocitose da bactéria; no entanto, a resposta de anticorpos anti-Ps 6B foi baixa. / Conjugate vaccines against Streptococcus pneumonaie have had an important public health benefit; nevertheless after its introduction it has been observed a serotype replacement. To solve this problems conjugate vaccines using pneumococcal surface proteins as carriers has been studied. In this work, Pneumococal surface protein A (PspA) was employed as a carrier protein, conjugated to serotype 6B. Initially PspA clade 1 and PspA clade 3 were purified; both proteins were recovered with more than 90% purity. The conjugation mediated by the activating agent 4- (4,6-dimethoxy-1,3,5-triazin-2-yl -) - 4-methylmorpholine chloride(DMT-MM) allowed Ps yields between 20 and 30%, however this conjugation chemistry had a negative impact on the immunogenicity of the protein. On the other hand conjugation by reductive amination led to Ps yields between 50 and 60% and induced high anti-PspA antibodies titers in Balb /c mice that were able to promote phagocytosis of the bacterium; however, polysaccharide 6B induced low antibody titers.

The unifying theme of this thesis is the exploitation of the reactivity of aryliodonium salts as electrophile transfer reagents. In the first part of the thesis, diaryliodonium salts are employed as arylation reagents for the enantioselective copper-catalysed arylative semipinacol rearrangement (SPR) of various tertiary allylic alcohols. This cascade reaction is a rare example of asymmetrically activating SPR using carbon electrophiles. Different substrate classes - including dihydropyran, indene and dihydronaphthalene moieties - are converted to enantioenriched beta-aryl spirocyclic ketones in excellent yields and enantioselectivities, and often as a single diastereomer. These are in turn useful functional handles for transformations into other moieties, including further rearrangements via Baeyer-Villiger oxidation. In the second part of this thesis, a two-step process for the macrocyclisation of native peptides via a non-natural linkage is developed. This study exploits previous work conducted in the group on the use of aryliodonium salts as methionine-selective diazoacetate transfer reagents. The functionalised methionine is in turn used for an intramolecular rhodium-catalysed insertion into tryptophan. Eventual translation onto solid-phase enables facile access into various macrocyclic peptides.

Philosophiae Doctor - PhD / The field of nanotechnology continues to offer solutions for biotechnologists whose target is to improve the quality of life by finding new therapies to combat diseases. Gold nanoparticles (AuNPs) have been showing great potentials in many biomedical applications. The antibacterial activity of the AuNPs presents a therapeutic option for conditions caused by bacterial infections such as chronic wounds. Also, these versatile particles can offer solutions in the treatments of infectious diseases and can also be exploited as “smart” vehicles to carry drugs, such as antibiotics, for improved efficiency. Moreover, the anti-inflammatory activity of AuNPs makes them useful in the management of prolonged inflammation caused by bacterial infections. The synthesis of AuNPs can be achieved by variety of physical and chemical methods that have been successfully applied in labs and industry. Nonetheless, the drawbacks of these “conventional” methods in terms of high cost, adverse health side effects and incompatibility with the ecosystem cannot be overlooked. Thus, new safer and more cost-effective protocols have been reported for the synthesis of AuNPs. Plants have provided alternate synthesis methods in which the reducing capabilities of the phytochemicals, found in the aqueous plant extracts, can be used to chemically synthesize AuNPs from gold precursors. The biosynthesis and characterization of AuNPs from the phytochemicals of several South African plants is investigated in this study. The study also reports the optimization of the AuNPs biosynthesis by varying reaction conditions such as temperature and plant extracts’ concentrations. Furthermore, the study highlights the wound healing activity of the AuNPs synthesized from selected plants by investigating their antibacterial activity on bacterial strains known to cause chronic wounds. The ability of these AuNPs to carry ampicillin in order to enhance the antibacterial activity is also described herein. The cytotoxicity of the biosynthesized AuNPs was evaluated on human normal fibroblasts cells (KMST-6). Additionally, the immunomodulatory effect of the biosynthesized AuNPs on the cytokines production from macrophages and Natural Killer (NK) cells was examined. The study was successful to produce biocompatible and safe AuNPs synthesized from the tested aqueous plant extracts. The resulted AuNPs showed different physicochemical properties by varying the reaction conditions. The AuNPs exhibited antibacterial activity against several Gram-positive and Gram-negative bacteria. Also, ampicillin was successfully loaded on the biosynthesized AuNPs, which led to the formation of more antibacterial active conjugated AuNPs compared to the free AuNPs. The green synthesized AuNPs were also found to have anti-inflammatory responses as shown by the reduction of pro-inflammatory cytokines from immune cells. In vitro assays showed that the biogenic AuNPs were not toxic to KMST-6 cells. Overall, the data suggest that plant extracts produce biologically safe AuNPs with antibacterial and anti-inflammatory activities that can be exploited in the treatment of chronic wounds and in the management of chronic inflammation.

Single-molecular electronics and organic material electronics are expanding research fields that ultimately aim for a vast variety of different applications, ranging from organic light-emitting diodes, to novel ways to improve the performance and decrease the size of electronics components. To achieve these goals, research has to be focused both on the development of functional molecules, but also on device fabrication. The work of this thesis is focused on the development of synthetic routes towards novel molecules for potential organic electronics applications, together with an investigation of their optical and electronic properties. The first part of the thesis describes the synthesis of butadiyne-substituted and diacetylenic phosphaalkenes. Theoretical, spectroscopic and electrochemical techniques have been used to understand key steps during their synthesis, and to gain further information on the conjugative properties of their π-systems. A mechanism is proposed for the formation of the butadiyne-substituted and diacetylenic phosphaalkenes and it is shown that the phosphorus heteroatom is an intrinsic part of the π-conjugated system. The incorporation of the phosphorus heteroatom leads to decreased HOMO-LUMO gaps compared to all-carbon based reference compounds. In the second part of the thesis, acetylenic phosphaalkenes are utilized for the preparation of phosphaalkene-substituted phospholes. A first step towards the exploration of the difference in reactivity of the σ2, λ3 phosphaalkene-P and the σ3, λ3 phosphole-P is presented as the oxidation of the compounds by sulfur proceeds selectively at the σ3, λ3–P. Spectroscopic and electrochemical investigations show that the phosphaalkene is an integral part of the compounds’ π-systems, and induces a HOMO-LUMO gap decrease compared to reference compounds that lack the P=C substituent. The third part of this thesis presents an exploratory study concerning the suitability of metathesis reactions for the assembly of alkene-bridged phosphaalkenes.

No description available.

<p>Metamaterials have gained extensive attention in recent years due to their ability to exhibit material properties otherwise difficult or impossible to obtain using natural materials. Nonlinear and active metamaterials in particular exhibit great promise for exploring new effects and applications, from tunability to mixing. However, nonlinear and active metamaterials have been explored significantly less than linear metamaterials to this point and much work has focused on the fundamental physics of nonlinear metamaterials. Our aim is to further extend the knowledge of practical nonlinear metamaterials and to demonstrate how they can be transformed to real-world applications through the use of embedded devices. In this dissertation, we demonstrate a variety of ways that devices can be embedded within metamaterial unit cells to provide nonlinear and active effects. </p><p>Chapter 1 introduces the basic theory of metamaterials, background of existing work, and the current limitations of nonlinear and active metamaterial design. In Chapter 2, we present the design, simulation, fabrication, and verification of an RF limiter metamaterial. We show how a metamaterial can be designed using RF engineering principles to act as an effective limiter in a new topology, relying on nonlinear devices embedded within a metamaterial. Chapter 3 shows our design and demonstration of a power harvesting metamaterial. We design a nonlinear metamaterial towards a potential application, discussing how the selection of an appropriate embedded device provides our desired functionality. In Chapter 4 we show how nonlinear and active metamaterials can be used to realize phase conjugation, including demonstration of negative refraction and imaging through the use of these metamaterials. We also discuss design approaches to moving these metamaterials towards real-world applications. Chapter 5 discusses our work concerning metamaterials based on transistors. First we show that appropriate design of a transistor circuit allows us to tune the quality factor and resonant frequency of a metamaterial. We use this metamaterial for time-varying mixing, as well, demonstrating a mixing metamaterial that remains linear. We then illustrate how using transistors as nonlinear devices provides much greater design freedom for use with metamaterials. We show that the nonlinearity of a metamaterial can be dramatically enhanced through the use of transistors and even dynamically tuned, applying these nonlinear metamaterials to applications including phase conjugation and acoustoelectromagnetic modulation. In Chapter 6 we summarize the achievements of the presented research and directions for future work that build on the work described in this thesis.</p> / Dissertation

Catheter-associated urinary tract infection (CAUTI) is one of the most common nosocomial infections and is caused by a range of different uropathogens, particularly by uropathogenic Escherichia coli (UPEC). Amongst the different virulence factors, biofilm formation and bacterial aggregation, often mediated by cell surface structures such as fimbriae, are common traits among uropathogens that cause CAUTI. In this study, a collection of UPEC isolates were screened for virulence genes and phenotypes associated with urinary tract infections such as biofilm formation and mannose-sensitive haemagglutination. Two strains, E. coli MS2027 (which formed a strong biofilm) and E. coli M184 (which aggregated strongly) were analysed in detail to determine the molecular mechanisms associated with these phenotypes. Transposon mutagenesis of E. coli MS2027 identified type 3 fimbriae as the factor responsible for its strong biofilm growth. Further screening revealed the presence of type 3 fimbriae in uropathogenic Citrobacter freundii, Citrobacter koseri, Klebsiella oxytoca, Klebsiella pneumoniae and other E. coli. Phylogenetic analysis of the type 3 fimbrial (mrkABCD) genes from these strains revealed they clustered into five distinct clades (A-E) ranging from one to twenty-three members. The majority of the sequences grouped in clade A, which was represented by the mrk gene cluster from the genome sequenced K. pneumoniae strain MGH78578. We demonstrated that type 3 fimbriae are functionally expressed by different Gram negative nosocomial pathogens and present evidence to suggest that they contribute significantly to catheter colonisation. The type 3 fimbrial genes from E. coli MS2027 were found to be located on a conjugative plasmid. Sequencing and annotation revealed that this 42,644 bp plasmid, named pMAS2027, contains 58 putative genes. Bioinformatic analysis identified pMAS2027 as an incompatibility X (IncX1) plasmid. Plasmid pMAS2027 contained genes encoding two important virulence factors, type 3 fimbriae and a type IV secretion (T4S) system. The biofilm ability was solely based on the expression of type 3 fimbriae and not the T4S system. The T4S system, however, accounted for the conjugative ability of pMAS2027. Differential tagging with fluorescent reporter genes demonstrated conjugative transfer of pMAS2027 between cells during biofilm growth. Finaly, transposon mutagenesis of E. coli M184 revealed a number of putative genes potentially responsible for bacterial aggregation. Of these, genes involved in the synthesis of the enterobacterial common antigen (ECA) were shown to be associated with an aggregation phenotype.