The explosive growth in the ability to sequence DNA due to next-generation sequencing (NGS) technologies has brought an unprecedented ability to characterize an individual's exome inexpensively. This ability provides clinicians with additional tools to evaluate the likely genetic factors underlying heritable diseases. With this added capacity comes a need to identify relationships between the genetic variations observed in a patient and the disease with which the patient presents. This dissertation focuses on computational techniques to inform molecular diagnostics from NGS data. The techniques focus on three distinct domains in the characterization of disease-associated variants from exome sequencing. First, strategies for producing complete and non-artifactual candidate variant lists are discussed. The process of converting patient DNA to a list of variants from the reference genome is very complex, and numerous modes of error may be introduced during the process. For this, a Random Forest classifier was built to capture biases in a laboratory variant calling pipeline, and a C4.5 decision tree was built to enable discovery of thresholds for false positive reduction. Additionally, a strategy for augmenting exome capture experiments through evaluation of RNA-sequencing is discussed. Second, a novel positive and unlabeled learning for prioritization (PULP) strategy is proposed to identify candidate variants most likely to be associated with a patient's disease. Using a number of publicly available data sources, PULP ranks genes according to how alike they are to previously discovered disease genes. This strategy is evaluated on a number of candidate lists from the literature, and demonstrated to significantly enrich ordered candidate variants lists for likely disease-associated variants. Finally, the Training for Recognition and Integration of Phenotypes in Ocular Disease (TRIPOD) web utility is introduced as a means of simultaneously training and learning from clinicians about heritable ocular diseases. This tool currently contains a number of case studies documenting a wide range of diseases, and challenges trainees to virtually diagnose patients based on presented image data. Annotations by trainee and expert alike are used to construct rich phenotypic profiles for patients with known disease genotypes. The strategies presented in this dissertation are specifically applicable to heritable retinal dystrophies, and have resulted in a number of improvements to the accurate molecular diagnosis of patient diseases. However, these works also provide a generalizable framework for disease-associated variant identification in any heritable, genetically heterogeneous disease, and represent the ongoing challenge of accurate diagnosis in the information age.

The pulp and paper industry in New Zealand annually produces over one hundred thousand dry tonnes of solid waste due to the treatment of pulp and paper mill effluents, the majority of which is currently landfilled. The New Zealand Waste Management Strategy (2002) has set a target for the diversion of commercial organic wastes from landfill to beneficial use to exceed 95% by 2010 . Effluents produced by softwood pulp mills, such as in New Zealand, contain high concentrations of naturally derived resin extractive compounds that are toxic to fish and other aquatic organisms. Improvements in waste water treatment technology has enabled the industry to meet rigorous discharge standards, however, this has resulted in an increase in the production of pulp mill effluent solids which require disposal. This has become an issue for the pulp and paper industry, especially as current landfill space is limited. Land application has been used for many years as a means of disposal of pulp mill wastes. While most studies investigating land application of pulp mill effluent solids have concluded that the risk posed to the environment is low, few have investigated the potential toxicity of these wastes to soil organisms, and these studies did not directly address the effects of resin extractive compounds. Resin extractives have been shown to be recalcitrant and to accumulate in anaerobic sediments. It is not known to what extent resin extractives are bioavailable or degradable in land applied Pulp mill effluent solids (PMES), or their potential to bioaccumulate in soil organisms. This PhD thesis research sought to extend the knowledge on the environmental fate and effects of pulp mill effluent wastes. It focused on terrestrial systems, which have not been well studied in this respect. Four chemically distinct softwood pulp mill effluent solids, a primary treatment solid and three secondary treatment biosolids, were used to investigate their effects on soil organisms and soil functions. An interdisciplinary approach was adopted, which incorporated three main areas of study, as follows: 1. A comprehensive resin extractives analysis of the pulp mill effluent solids undertaken so that effects on soil organisms and soil functions could be related to the resin extractives chemistry of the individual pulp mill effluent solids tested. 2. A battery of bioassays used to investigate the toxicity of the selected pulp mill effluent solids. 3. A field trial set-up to investigate how pulp mill effluent solids affected soil functions and also to investigate the decomposition of pulp mill effluent solids and of resin extractives in these solids. Three hypotheses tested were: I. Pulp mill effluent solids are toxic to soil organisms. II. Resin extractives in pulp mill effluent solids are recalcitrant in the terrestrial environment. III. Pulp mill effluent solids will cause significant measurable negative impacts on soil functional capacity. A wide range of resin extractives compounds were identified in pulp mill effluent solids, and concentrations of individual compounds varied widely between the different pulp mill effluent solids tested. During the two years after field application of the pulp mill effluent solids, resin extractives declined, however, decay rates of individual compounds were variable. The decay rate of compounds was influenced by the type of pulp mill effluent solids containing these compounds and not by the initial concentration of compounds. All compounds, including resin acids, degraded rapidly in the applied primary solid and in one of the applied biosolids, with average half-lives calculated ranging from three to twelve months. In the other biosolids, resin acids were recalcitrant with average half-lives calculated to be nearly ten years. Laboratory bioassays conducted on oats, earthworms and enchytraeid worms indicated that pulp mill effluent solids had low toxicity to these organisms. An aquatic bioassay organism, however, was acutely affected by aqueous extracts from pulp mill effluent solids. Earthworms were shown to bioaccumulate some resin extractive compounds to a limited extent, indicating that resin extractives were bioavailable in pulp mill effluent solids. Enchytraeid reproduction was reduced by exposure to some pulp mill effluent solids but this was not correlated to resin extractives concentration of the solids tested. Field applied pulp mill effluent solids significantly enhanced the fluxes of CO2 from the soil surface. When mass losses of pulp mill effluent solids carbon were taken into account, it was shown that these solids had little effect on soil respiration. Needle litterbags were placed above and below pulp mill effluent solids and in the litter horizon of a control treatment for 12 months. Needle litter decomposition was not significantly different between treatments, however, changes in enzyme activities were detected in litter beneath pulp mill effluent solids compared to control needle litter that had not been exposed to pulp mill effluent solids. The decomposition rate of field applied pulp mill effluent solids was slow, with half-lives extrapolated to be between five and twenty five years. The decomposition rate appeared to be influenced by the pools of carbon available for decomposition, with primary solids decaying significantly faster than biosolids due to a higher cellulose component. The conclusions of the PhD thesis research are, therefore, that generally, pulp mill effluent solids used in the study were demonstrated to be relatively benign and appear to pose a low risk to the terrestrial ecosystem when applied to soil. However, a cautious approach is still recommended to land application, based on the extensive evidence of disruption to aquatic ecosystems, and because pulp mill effluent solids will take many years to decompose and resin acids are recalcitrant in some pulp mill effluent solids. Further research is recommended to elucidate mechanisms of action by resin extractives in soil organisms and the ultimate fate of these compounds in the soil compartment.

<p>The conventional viscose manufacturing process is a mature process that needs to be improved with respect to its environmental impact and its production cost structure. Therefore a research study has been done with the aim to improve the reactivity of the dissolving pulp used, in order to reduce the chemical demand in the viscose process and thus reduce the cost and indirectly the environmental impact.</p><p>The work described in this thesis has shown that it is possible to enhance the pulp reactivity and to use less carbon disulphide in the production of viscose, while maintaining a good quality viscose dope, by two entirely different pretreatment methods, one chemical and one enzymatic.</p><p>The chemical method used pressurized oxygen after the mercerisation step, which increased the reactivity of the alkali cellulose. The viscose dopes produced from the pressurized oxygen treated alkali cellulose had lower filter clogging values, Kw, compared to conventionally produced viscoses. The temperature and the oxygen treatment time of the alkali cellulose were however crucial for the viscose quality.</p><p>The best performing enzyme of several tested was a cellulase of the mono component endoglucanase preparation Carezyme®. This enzymatic treatment was optimized with respect to viscose dope preparation. The study showed that the enzyme treatment could be carried out under industrially interesting conditions with respect to temperature, enzyme dose and reaction time. A re-circulation study of the enzyme showed that it was possible to re-use the spent press water from the enzymatic treatment step several times, and thus lower the production cost. Some of the viscose process stages were modified to properly fit the enzymatically treated dissolving pulp and a comparison between viscose made from enzyme-treated pulp and viscose made from conventional pulp, showed that the enzyme-treated samples had a lower filter clogging value, Kw. This indirectly indicates that the enzyme pretreatment could reduce the carbon disulphide charge in the viscose manufacturing process. An initial study of how the Carezyme® influenced different cellulosic sources was also performed.</p>

Combustion properties of kraft black liquor solids were studied using a laminar entrained flow reactor. The tendency of black liquors to release fume (compounds containing Na⁺, K⁺, Cl⁻, SO₄²⁻, SO₃²⁻, S₂0₃²⁻ and C0₃²⁻) during combustion were observed at 1000°C. Black liquor solids with a size fraction of 63 to 100 μm were burned in a mixture of 4% 0₂ and 96% N₂ at a residence time of 0.67 seconds. Combustion properties of one black liquor (liquor #3) were studied by varying the solids feed rate from 0.47 to 1.08 g/min (liquor feed rate study). Combustion properties of five North American and Finnish black liquors were studied at a target solids feed rate of 0.73 g/min (liquor to liquor study). Black liquor fuming was observed to be a decreasing function of solids feed rate and an increasing function of excess oxygen. The appearance of char residues varied from black and porous at high solids feed rates to white and dense at low solids feed rates. Combustion may have been enhanced at low solids feed rates by liquor swelling due to a combination of heat and mass transfer effects and limited at high solids feed rates by inter-particle and bulk gas mass transfer limitations. For the liquor to liquor study, black liquors were observed to release fume differently. Chars produced during this study varied in appearance, indicating that the black liquors had unique combustion properties. Variations in temperature and mass transfer effects resulting from liquor swelling properties were likely responsible for the variability in liquor fuming behavior. The liquors that contained the most NaCl and had the highest anionic equivalents as C0₃²⁻ (or other chemical species) produced the most fume. Sodium vaporization varied from 25.2% to 33.7%: Liquors #2 and #5 vaporized the most sodium and also had the lowest concentrations of measured anions in their char residues. Potassium and chloride enrichment factors for the five liquors were slightly lower than those of common industrial boilers. Liquor #3 had a concentration of Cl⁻ that was (roughly) an order of magnitude higher than the other liquors studied; however, it also had the lowest chloride enrichment factor. / Graduation date: 2002

Graduation date: 1996

This project explored the feasibility of using fungi in a constructed wetland for the treatment of pulp mill effluent. The effluent is high in dissolved lignins (some of which are chlorinated), which have proven very difficult to degrade biologically. Mindful of work done with the (terrestrial) white rot fungi, especially Phanerochaete chtysosporium, the question is asked, Is there a fungus which can tolerate submerged conditions while degrading a significant amount of dissolved lignins? Two fungal species with lignin-degrading capability were isolated from submerged films in a log pond. These fungi have been evaluated for decolorization potential under different environmental conditions. Results of laboratory experiments show that one of these fungi, identified as Acremonium sp., was capable of 44% decolorization of pulp mill effluent under sterile, submerged, room temperature conditions. The fungal decolorization was evaluated both in floating cultures and as a film inoculated on wood chips. In addition, bench-scale examination of the potential of this fungus to decolorize pulp mill effluent in non-sterile conditions was completed. / Graduation date: 1994

The conventional viscose manufacturing process is a mature process that needs to be improved with respect to its environmental impact and its production cost structure. Therefore a research study has been done with the aim to improve the reactivity of the dissolving pulp used, in order to reduce the chemical demand in the viscose process and thus reduce the cost and indirectly the environmental impact. The work described in this thesis has shown that it is possible to enhance the pulp reactivity and to use less carbon disulphide in the production of viscose, while maintaining a good quality viscose dope, by two entirely different pretreatment methods, one chemical and one enzymatic. The chemical method used pressurized oxygen after the mercerisation step, which increased the reactivity of the alkali cellulose. The viscose dopes produced from the pressurized oxygen treated alkali cellulose had lower filter clogging values, Kw, compared to conventionally produced viscoses. The temperature and the oxygen treatment time of the alkali cellulose were however crucial for the viscose quality. The best performing enzyme of several tested was a cellulase of the mono component endoglucanase preparation Carezyme®. This enzymatic treatment was optimized with respect to viscose dope preparation. The study showed that the enzyme treatment could be carried out under industrially interesting conditions with respect to temperature, enzyme dose and reaction time. A re-circulation study of the enzyme showed that it was possible to re-use the spent press water from the enzymatic treatment step several times, and thus lower the production cost. Some of the viscose process stages were modified to properly fit the enzymatically treated dissolving pulp and a comparison between viscose made from enzyme-treated pulp and viscose made from conventional pulp, showed that the enzyme-treated samples had a lower filter clogging value, Kw. This indirectly indicates that the enzyme pretreatment could reduce the carbon disulphide charge in the viscose manufacturing process. An initial study of how the Carezyme® influenced different cellulosic sources was also performed.

Recent studies have shown that dehydroabietic acid (DHAA), a resin acid present in pulp and paper mills, may have anti-estrogenic effects in fish. A chronic-exposure toxicity experiment using immature rainbow trout (Oncorhynchus mykiss) was conducted in order to assess the endocrine disrupting and liver metabolic effects of the wood extractives DHAA and β- sitosterol (BS) regularly present in pulp and paper mills and the model estrogen 17β-estradiol (E2). It was found that exposure to 5 ppm of E2 significantly increased hepatosomatic index (HSI), vitellogenin (VTG) and plasma sorbitol dehydrogenase (SDH). This effect was reduced by mixing E2 with DHAA, indicating that DHAA does not cause its anti-estrogenic effects indirectly due to liver damage. Exposure to 5 ppm of DHAA caused a significant increase in liver citrate synthase (CS), and liver ethoxyresorufin-O-deethylase (EROD) activity after 7 days, however, the fish recovered by 28 days. This study also determined the effect of 14 different pulp and paper mill effluent extracts on liver enzyme metabolism through alterations in the activity of liver lactate dehydrogenase activity (LDH) and CS. This activity varied greatly between mills but most showed an induction of CS after 28 days exposure through i.p. injection. The results of the study indicate that DHAA may alter energy metabolism as well as cause anti-estrogenic effects in female juvenile rainbow trout. / UOIT

Graduation date: 1994

Magnesium oxide-based additives have been used in kraft recovery boilers to minimize deposit accumulation on superheater and generating bank tubes. Due to inconclusive results observed in boilers and claims made for additive effectiveness backed by little scientific evidence, only a few mills still use the additives for fouling control. A systematic study was performed to examine the effect of MgO-based additives on deposit accumulation and removal under controlled laboratory conditions, and to evaluate the additive effect in a kraft recovery boiler. The results show that while the additive coating has no effect on deposition, it could ease deposit removal if a coating greater than 5 µm can be applied on the tube surface effectively. Separate injection of additive and black liquor had insignificant effect on deposition and removal. Results of the field study show that the amount of additive collected on the tube is too small to have an impact.