2,084 results for Otago University Research Archive, Doctoral

  • The Polar Palette - The role of flower colour in polar regions

    Little, Lorna (2014)

    Doctoral thesis
    University of Otago

    Flower colour throughout the world is generally linked to various functional purposes, pollinator attraction in particular. Current knowledge regarding the functional significance of flower colour is, however, mainly based on studies from boreal, temperate, and tropical regions. This knowledge is not readily applicable to polar regions. Similar to lower latitudes, flower colour is often observed in polar regions, as well as flower colour polymorphisms. Pollinator attraction are considered to be the main purpose of flower colour, and the lack of pollinators in polar regions, where many plant species produce coloured flowers, causes the significance of colour to be unclear. Further, flower colour pigments are often quite energy expensive to produce, and in polar regions, nutrients, light and resources are limited. Hence, the question; what is the role of flower colour in reproductive success in polar regions? This thesis addresses this question of ‘What is the role of flower colour in polar regions?’ through several studies related to various aspects of floral reproduction in polar regions. This is a broad topic, and little background knowledge exists. To deal with this, a broad approach was also used, covering different geographical scales, from bipolar (Chapter Three), arctic (Chapter Two) and regional (Svalbard; Chapter Six) scales, to the population scale (Chapter Four and Five). Information from literature on hundreds of species (Chapter Two) was compiled, field experiments and thermal imaging were completed on specific plant populations (Chapter Three - Five), and, at the genetic level, amplified fragment length polymorphism (AFLPs) were conducted within a species (Papaver dahlianum; Chapter Six). Except for one study (Chapter Four), which included results from Campbell Island (Subantarctic), most information is gathered from the Arctic, in particular from the high-arctic archipelago, Svalbard. The thesis is thus somewhat biased towards the Arctic. There were several initial hypotheses around why we observe flower colour polymorphism in polar regions. One possibility was that the few pollinators present may be more specific or more efficient than formerly believed, and sufficient to drive and sustain flower colour polymorphisms in polar regions (addressed in Chapters Two, Three, Five and Six). Alternatively, these colour morphs could be remnants from ancestor populations living under a different pollinator regime, and without any current function (historical patterns are partly addressed in Chapter Two and Six). However, as the polar climate is thermodynamically unfavourable and energy budgets are tight, flower colour pigments are often too costly to produce without being linked to a functional purpose. Hence, another possibility was that flower colour is linked to functions aside from signalling to attract pollinators (addressed in Chapter Four and Five). The diversity and distribution of blue-purple flowers in the Arctic were correlated with the diversity and distribution of specialist pollinators (bumblebees) with this colour preference. The highest number of both blue-purple flowers and bumblebee diversity were registered in non-glaciated arctic regions, where populations may have persisted since the arctic biome originated (Chapter Two). These results indicate that although arctic bumblebees are few and rarely active, their presence and activity is sufficient to impact colonization efficiency in the Arctic, and in certain areas may drive further selection and diversification of flower colours. The most common flower colours throughout the Arctic were shown to be white, blue-purple and yellow (Chapter Two). White and yellow are reported to attract more general, less efficient pollinators such as Diptera. In Svalbard, no known specialist pollinators exist. To investigate possible colour preference among the putative pollinators in Svalbard, insect traps with different colours were distributed at four different sites. Eight different families of Diptera were caught. Mycetophilidae were the most common, and showed a preference for the colour red, whereas the Muscidae and Syrphidae were more common in yellow and white coloured traps. Diptera species are already known to act as pollinators of white and yellow flowers, and as such are likely performing some pollination in Svalbard. However Mycetophilidae are rarely noted for pollination, and the extent to which these Diptera are involved in outcrossing pollination of flowering species on Svalbard remains unknown. The effects of flower colour and floral heating were tested in Adventdalen, Svalbard, using Papaver dahlianum which has white and yellow flower colour morphs (Chapter Four). Floral heating effects were also briefly investigated for Pleurophyllum spp, Stilbocarpa polaris, Bulbinella rossii and Anisotome latifolia on subantarctic Campbell Island. Flower temperature was measured and compared with environmental conditions, including ambient air temperature. In P. dahlianum, yellow flowers were warmer in some situations, whereas white flowers were warmer in others. Floral shape was also of some significance. Dark purple flowers on Campbell Island were found to be significantly warmer than ambient temperatures under high levels of solar radiation. Seed production was proposed to vary between colour morphs of P. dahlianum in Svalbard due to differences in insect visitation and heating based on colour (Chapter Five). Yellow flowers were expected to produce more seeds due to being more attractive and warmer than white flowers. Seed production was measured using different pollination treatments (forced selfing, hand pollination, forced outcrossing) at four different sites within the two P. dahlianum colour morphs. P. dahlianum was capable of setting seed autogamously. The largest seeds were collected from Adventdalen, but there were no significant differences of seed size between flower colours. There were no significant differences between white and yellow flowered plants in terms of relative reproductive effort. Thus, colour is assumed to have little effect on reproductive output when few seasons are considered. Assortative mating, where mating could be structured around flower colour was investigated in Chapter Six. Genetic samples were collected from nineteen populations of P. dahlianum around Svalbard. Genetic fingerprinting via AFLPS was used to assess whether colour was associated with structuring of the population (Chapter Six). There was no genetic pattern based around colour, at regional, local or the population scale. There did appear to be several widespread genetic lineages present in Svalbard, which may represent genetic groups with reduced reproductive compatibility or cryptic species. The existence of flower colour in polar regions was concluded to be partly due to the recent colonisation processes of the areas, whereas the role held by flower colour in polar regions was variable, and strongly dependent on the season of investigation. Due to seasonal conditions differing greatly between years, flower colour polymorphisms in Svalbard are likely maintained by different conditions favouring different morphs in different seasons.

    View record details
  • “I’m not a pill-taker”: Medication and meaning for older people

    Ritchie, Lorraine Helen (2014)

    Doctoral thesis
    University of Otago

    Background: The philosophy of ageing in place has seen a greater emphasis on community care for older people. In New Zealand, access to assistance at home from health services begins with a comprehensive geriatric assessment, which includes medication management. This thesis examines specifically older participants’ self-management of medications and what medication-taking means to them. Aims 1. To explore how older people make sense of their medication-taking and medication self-management. 2. To explore the value of a narrative approach/methodology in eliciting older people’s attitudes and practices in medication-taking. 3. To critically view how the meaning older people make of their medication self- management might influence policy and practice. Methodology and Methods The overarching methodological approach to this thesis is a narrative one. Twenty participants underwent two separate sets of interviews: a needs assessment interview guided by a standardised geriatric assessment tool, and a semi-structured interview about their practices and beliefs on medication-taking. In total, forty interviews with older participants who were living in their own homes in the community were analysed using narrative analysis and presenting data in the form of themes and case studies. Results Four key findings emerged from the analysis. These were the themes of: 1. Living with illness – the impact on lives and medication-taking First and foremost, participants lived and coped with illness as part of their everyday living at home. Medication-taking was one of multiple consequences of the daily reality of living with a chronic illness. Any sense that participants made of their medication was in the first instance viewed through the lens of a recent illness, hospitalization, chronic pain or uncertainty of the future trajectory of the illness. 2. Trust and the older patient-doctor relationship The majority of participants expressed a strong sense of trust in their doctor as the main prescriber of their medication. The doctor’s word and advice was law to most participants, who imbued their doctor with an authority and competence without question. Further, many also valued a ‘social’ relationship with their doctor where a personable interaction was able to exist largely due to longevity of relationship, but also a willingness to see the doctor as a person, not just a health professional in an objective transaction. 3. Everyday routines and strategies These showed resourcefulness, coping and adaptation. Participants demonstrated a variety of unique systems and strategies related to medication self-management, from sophisticated routines where medication was checked, administered and stored correctly through to haphazard and inconsistent patterns of self-administration. Strategies depended on their beliefs but also on life circumstances and the meaning and importance placed on medication. Participants coped variously with management of their medication in the same way they coped with other aspects of daily life at home such as preparing meals, showering and socialising. Managing medication was not seen by the participants in isolation from the lived daily context of everyday managing routines and strategies. Self-management of medication required the ability to navigate through layers of complex context. For some it was more important to comply with medication taking; for others it was not a priority. 4. Advice – formal and informal Participants had valued relationships with others beyond doctors in relation to medication-taking. This was primarily with other formal health professionals such as pharmacists, but also on an ‘informal’ level, with family and friends. Some advice and recommendations came from surprising sources such as a bank teller or a women’s magazine. The gathering of information which informed medication-taking showed that older people’s lived context was far wider than a doctor-patient relationship. Views and attitudes of others all contributed to the sense older participants made of their medications. Conclusion/implications for practice Health services and professionals need to respect strategies and coping methods which older people have developed to self-manage medications and other aspects of their daily living. By listening to older people’s narratives of how they manage, health professionals can work with older people on their own terms starting with the voice of the older person as expert self-manager. Health professional education is necessary to raise awareness that meaning for older people is produced through narrative and that this is a key component of assessment and service delivery.

    View record details
  • The Soviet Legacy and Post‐Communist Civil Society in the Caucasus

    Aliyev, Huseyn (2014)

    Doctoral thesis
    University of Otago

    This study argues that the weakness of civil society in the post-Soviet Caucasus is not only a result of post-communist political and economic problems but is also due to the effects of historical legacies which continue influencing both formal and informal civil societies of the Caucasus’s countries, weakening their ability to facilitate democratization. Two decades after the break up of the USSR, democratization continues to present a challenge to all non-Baltic former Soviet states. The failure of most post-Soviet governments to overcome autocratic patrimonial habits of governance and to embark on democratic institution-building has been a characteristic of the former Soviet Union for the past two decades. Among many other malaises of post-communism, the inherent weakness of civil society has been observed in virtually all post-Soviet regimes. Unlike civil sectors of post-communist Central Europe or even the successor states of the former Yugoslavia, civil societies of ex-Soviet countries remain underdeveloped, ineffective and weak. In contrast, the entrenchment of authoritarian regimes, failures of institutional reforms, in conjunction with the continued reliance of ruling elites on informal structures rather than formal institutions is on the rise in most countries of the post-Soviet region. All of the above is most notable in the former Soviet region of Caucasus. Throughout the entire post-communist period, the political and civil actors across the Caucasus have shown themselves incapable of shedding the old forms of governance, which led to further growth of authoritarianism and weakening of independent civil society. So why does the Caucasus’s civil society fail to facilitate democratic state-building and institution-building processes, invigorating civil mobilization and serving as a balance between the state and society? This thesis examines the relationship between the weakness of civil society and the legacy of Soviet public and private spheres in the post-Soviet Caucasus. Starting from the assumption that the analysis of ‘civic traditions’ of formal and informal civil association inherited from the Soviet period can provide explanations as to why the present-day civil sector is weak, this study seeks to reveal the significance of the former regime’s legacy for contemporary civic institutions. Using qualitative methods of inquiry, this thesis conducts an in-depth examination of both Soviet and post-communist formal and informal civic association, offering fresh insights into our understanding of Soviet civic legacy and of how and why ‘civic traditions’ continue. The findings of this thesis emphasize, among others, that the antecedent regime’s institutional norms and individual attitudes can have long-lasting effects not only in particular countries but also trans-nationally.

    View record details
  • The One-China controversy, 1996-2002 : the impact of Taiwan's democratisation on the cross-strait policies of Taipei, Beijing and Washington

    Lin, Chin-sheng (2005)

    Doctoral thesis
    University of Otago

    xii, 535 p. ; 30 cm. Includes bibliographical references. University of Otago department: Political Studies. "February 2005."

    View record details
  • Enzymology and Structure of the Thiol Dioxygenase from Pseudomonas aeruginosa

    Fellner, Matthias (2015)

    Doctoral thesis
    University of Otago

    Thiol dioxygenases catalyse the oxidation of a thiol substrate with dioxygen to produce a sulfinic acid. The mammalian thiol dioxygenase cysteine dioxygenase (CDO), is specific to its substrate L-cysteine, and has been investigated for more than 40 years. However, during the last decade additional thiol dioxygenases have been discovered and all showed a high specificity for their substrate. In-depth kinetic and structural characterisation of an additional thiol dioxygenase was undertaken to support a comparative analysis of the structural basis of thiol dioxygenase activity and substrate specificity. This thesis presents the first extensive characterisation of a putative bacterial CDO homologue from the eubacterium Pseudomonas aeruginosa. For determination of kinetic parameters a new activity assay was developed. This new assay utilises a 96 well plate format and is therefore much more time efficient compared to available techniques. The depletion of any thiol substrate by a thiol dioxygenase can be studied quickly and easily without compromising data quality, however with the drawback that product formation has to be investigated with another method in parallel. For the thiol dioxygenase P. aeruginosa kinetic parameters (kcat and Km) for two different substrates (3-mercaptopropionic acid and L-cysteine) were gathered across a broad pH range, providing a fuller picture than for any thiol dioxygenase other than mammalian CDO. The results will contribute to predicting future putative thiol dioxygenase function as key differences in the sequence of putative CDOs have been compared to experimental findings. The findings show that thiol dioxygenases can have a broad substrate profile and a short list of the protein residues responsible for different substrate specificity of thiol dioxygenases is proposed. Kinetic studies were performed in tandem with X-ray crystallographic characterisation of the thiol dioxygenase structure; this combination is a first for a putative bacterial CDO homologue. The second sphere protein residues appear to be rotated in reference to the active site iron when compared with mammalian CDO. The structure also offers a new insight into an intensively researched feature of CDO, a conserved cysteine-tyrosine crosslink. The thiol dioxygenase from P. aeruginosa lacks this post-translational modification but the active site environment appears to be conserved. This was further proven by structural characterisation of single point mutation (G95C), resulting in the first artificially introduced crosslink into a naturally uncrosslinked thiol dioxygenase. However this mutation dramatically decreased enzymatic activity while having minimal affect on the structure. In addition, crystal structures of single point mutations (C93G, Y157F) abolishing the crosslink in mammalian CDO were also solved. These structures are the first crystal structures of mammalian CDO lacking the crosslink. Comparison of all crystal structures shows that the crosslink does not serve a general stabilisation purpose. The work shows that the protein possesses a dual substrate profile making it a true thiol dioxygenase, it will therefore be referred to P. aeruginosa thiol dioxygenase – pTDO.

    View record details
  • Hippocampal Place Cells Dynamically Encode Value of Available Goals During Spatial Navigation

    Cheyne, Kirsten Ruth (2014)

    Doctoral thesis
    University of Otago

    The hippocampus plays a role in spatial navigation, and place cells (cells that fire selectively for an animal’s location in an environment) are thought to provide an anatomical basis for a cognitive map (a map-like representation of space in the brain). Recently, however, place cells have been found to encode more complex aspects of navigation than simply place; including information about the location or presence of upcoming goals. The present experiments aimed to investigate the involvement of the hippocampus in representing the value of a goal (taking into account the size of the reward and the cost in obtaining it), and its contribution to navigation. Place cells were recorded while rats chose between two goals of differing value, in a spatial decision-making task conducted in a figure-of-eight maze. The value of a goal to an animal was inferred based on the proportion of trials in which that goal was chosen. It was hypothesised that cells would fire differentially in the maze depending on the available rewards, and that cell activity would be modulated by the value of upcoming rewards. It was also hypothesised that place cells representing goal value would dynamically encode changes in value when reward sizes were altered, and that this may influence the animal’s future behaviour. Additionally, it was predicted that a subset of cells would fire “out-of-field”, prior to the animal reaching the reward location those cells represented, possibly indicating the mental projection of the animal to that location. The results showed that when animals are presented with rewards of two different values, more place cells are involved in the representation of the reward of higher value to the animal, as measured by the number of cells with fields in that region, despite there being no difference in firing rate. In the maze’s central stem, many cells showed differential firing based on upcoming goal choice. Additionally, a number of cells showed out-of-field firing, firing differentially in a secondary field with regard to the animal’s next choice, and the goal that cell represented. When reward size in each goal changed every twenty trials of the 100-trial session, animals’ behaviour changed quickly to accommodate new reward values, and mean firing rate in the central stem mirrored the pattern of behaviour. The firing rate in the first pass through the stem following a change of reward size, but before a change in behaviour, was strongly correlated with the subsequent distribution of behaviour, whereas the firing rate for the last pass of each block was not. These results support previous research suggesting that the hippocampus is briefly engaged during route planning, but is not needed during the execution of a determined route. They also show for the first time that place cells dynamically encode goal value during spatial navigation. It is proposed that during spatial decision-making, value information, possibly originating from the prefrontal cortex, is transferred to the hippocampus, where it is incorporated into the spatial map of the environment, enabling the animal to most efficiently direct its behaviour.

    View record details
  • An evaluation of the clinical utility of the modified FABER’s test: Can it discriminate for low back pain of sacroiliac joint origin?

    Adhia, Divya Bharatkumar (2014)

    Doctoral thesis
    University of Otago

    Background: Innominate kinematic anomalies resulting in non-specific low back pain (LBP) of sacroiliac joints (SIJ) origin have been well recognized in the literature. While the complex anatomical orientation and position of the SIJs renders evaluation of innominate kinematic anomalies difficult, recent techniques of palpation-digitization of pelvic landmarks (in the modified FABER’s test positions) using electromagnetic tracking device have been able to accurately and non-invasively quantify subtle innominate kinematics (in healthy individuals). The aim of this thesis is to determine if the modified FABER’s test positions and the electromagnetic palpation-digitization technique are capable of discriminating for subjective (pain provocation) and objective (innominate kinematics) clinical parameters respectively, that would allow differentiation and identification of individuals clinically diagnosed with LBP of SIJ origin (SIJ-positive) when compared to individuals clinically diagnosed with LBP of Non-SIJ origin (SIJ-negative). Methods and results: This thesis comprises of three studies. The first two studies were conducted to evaluate the psychometric properties (validity and reliability) of the palpation-digitization technique, and the third study was conducted to determine the use of the palpation-digitization technique for identifying and differentiating between two clinical groups. Study 1 systematically reviewed the literature, in order to determine the level of evidence for the validity and reliability of the electromagnetic palpation-digitization technique for measurement of joint kinematics. The results of this (PRISMA guided) systematic review demonstrated an overall strong level of evidence for validity [n (3 high quality studies)] and intra-rater reliability [n (5 high & 2 low quality studies)] of palpation-digitization technique for measurement of joint kinematics. Evidence for inter-rater reliability of electromagnetic palpation-digitization technique was limited [n (1 study)] and was deemed essential for use of this procedure in the clinical utility. Study 2 evaluated the inter-tester reliability of the palpation-digitization technique for measurement of innominate kinematics. Four musculoskeletal testers, using electromagnetic tracking device, palpated and digitized the pelvic landmarks in fourteen healthy participants. The innominate vector length was calculated from the 3D co-ordinates of the palpated and digitized landmarks, in two modified FABER’s test positions. The results of this reliability study demonstrated very high inter-tester reliability [Intraclass correlation coefficient (≥ 0.97), and Standard error of measurement (≤ 1%)] of the palpation-digitization technique for non-invasive innominate kinematics measurement. This results thus demonstrate that the palpation-digitization technique satisfies the psychometric properties essential for its further research and clinical use. The next step was to explore the use of the palpation-digitization technique and the modified FABER’s test positions for determining innominate kinematics in a clinical population. Study 3 evaluated the subjective (pain reproduction) and objective (innominate movement pattern, range of motion and trend of rotation) clinical parameters of modified FABER’s test positions and the palpation-digitization technique respectively, in two clinical groups of chronic non-specific LBP individuals [SIJ-positive: n (45), and SIJ-negative: n (77)], to determine if these clinical parameters would allow differentiation and identification of SIJ-positive LBP individuals. The results of this kinematic study demonstrated that both the subjective and objective clinical parameters were capable of differentiating and identifying SIJ-positive LBP individuals. The modified FABER’s test positions reproduced familiar pain in SIJ-positive LBP individuals compared with SIJ-negative LBP individuals [p (0.001)], and had moderate levels of sensitivity (67% to 78%) and specificity (63% to 66%) for identifying SIJ-positive LBP individuals. The innominate kinematics of movement pattern and trends of rotation were significantly different [p (< 0.017)] in SIJ-positive LBP individual compared with SIJ-negative LBP individuals, but the innominate ranges of motion did not demonstrate a significant between-group difference [p (> 0.300)]. Conclusion: These between-group differences in pain reproduction and innominate kinematics in the modified FABER’s test positions, demonstrate significant associations between innominate kinematic anomalies and the SIJ pain. However, it is unknown if the innominate kinematic patterns observed in SIJ-positive LBP individuals are a cause or an effect of SIJ pain. Nevertheless, the discriminative results for palpation-digitization examination of non-invasive innominate kinematic measurement in a clinical LBP population is an important step toward a greater understanding of normative, aberrant and pathological SIJ kinematics. This thesis thus has set the groundwork for further exploration of clinical measurement, clinical relevance, and clinical management of these potentially important movement observations.

    View record details
  • S. aureus alanine racemase: "A target for structure-based drug design"

    Scaletti, Emma Rose (2014)

    Doctoral thesis
    University of Otago

    Staphylococcus aureus is an opportunistic Gram-positive bacterium which causes a wide variety of diseases ranging from minor skin infections to potentially fatal conditions such as pneumonia, meningitis and septicemia. The pathogen is a leading cause of nosocomial acquired infection, a problem exacerbated by the existence of methicillin and glycopeptide antibiotic resistant strains which are becoming exceedingly difficult to treat. Alanine racemase (Alr) is a pyridoxal-5’-phosphate dependent enzyme which catalyzes reversible racemization between enantiomers of alanine. As D-alanine is an essential component of the bacterial cell wall peptidoglycan, inhibition of Alr is lethal to prokaryotes. Additionally, while ubiquitous amongst bacteria, this enzyme is absent in humans and most eukaryotes, making it an excellent antibiotic drug target. There are two alanine racemase isozymes in S. aureus; Alr, which is constitutively expressed, and a catabolic alanine racemase (DadX), which is inducible by L-alanine. Current Alr inhibitors such as D-cycloserine are reminiscent of the alanine substrate and inhibit the enzyme by binding covalently to the enzymes’ PLP cofactor. They therefore non-specifically target other enzymes that utilize this cofactor and are associated with severe toxicity during treatment. This emphasizes the need for the identification of new non substrate-like Alr inhibitors. This research focused on the use of S. aureus Alr (AlrSas) as a template for structure-based drug design studies. This first involved solving the native structure, and kinetically characterizing the enzyme. Following structure solution, enzyme inhibition and X-ray crystallographic studies of AlrSas with nine compounds identified through high-throughput screening (HTS) (Anthony et al., 2011; Ciustea et al., 2012; Lee et al., 2013) were performed. In addition, fragment-based screening studies using differential scanning fluorimetry (DSF) were performed to identify new non substrate-like Alr inhibitors. To our knowledge this is the first reported use of this drug discovery method being applied to an alanine racemase. AlrSas from the highly antibiotic resistant Mu50 strain was over-expressed in E. coli, following which it was purified using anion-exchange, hydrophobic interaction and size-exclusion chromatography. Following crystallization, the native structure was solved to 2.15 Å resolution. Comparison of AlrSas with various alanine racemases demonstrated a conserved overall fold with the enzyme sharing most similarity with those from other Gram-positive bacteria. Structural examination indicated that the active site binding pocket, dimer interface and active site entryway of the enzyme were potential targets for structure-aided inhibitor design. AlrSas was kinetically characterized, and a comparison with selected alanine racemases indicated three orders of magnitude difference in their kinetic constants. In addition, the second alanine racemase isozyme from S. aureus (DadXSas) was studied. DadXSas from the MRSA252 strain was over-expressed in E. coli, following which it was purified to homogeneity by IMAC and size-exclusion chromatography. DadXSas was crystallized, however these crystals were severely twinned and diffracted poorly, and thus a structure did not result from this research. Kinetic characterization of DadXSas was performed and compared to other alanine racemases in the literature. DadXSas was one of the slowest enzymes, and had the weakest affinity for both enantiomers of alanine out of all the published alanine racemases. Interestingly, the enzyme differed markedly to AlrSas, having a reaction rate two orders of magnitude lower. Analysis of the nine HTS molecules against AlrSas indicated that only six compounds inhibited the enzyme to an acceptable level (BAS, JFD, S14, TCRS-151, 1944, and TO5-13). These compounds inhibited the enzyme in the low micromolar range. Further IC50 determinations to test for the possibility of non-specific inhibition indicated that of the six compounds JFD and TO5-13 were the most likely to be inhibiting AlrSas by promiscuous mechanisms. X-ray crystallography studies did not yield the structure of AlrSas in complex with any of the six HTS compounds. Preliminary screening of AlrSas via DSF against the MayBridge fragment library identified 28 fragments which either significantly increased or decreased the Tm of the enzyme. Subsequent IC50 analysis against AlrSas indicated that three of these compounds inhibited the enzyme in the low millimolar range (CC42223, GK00199, and CC42201) and thus were significantly weaker inhibitors than the HTS compounds analyzed. Promiscuous inhibition tests indicated that the fragments were not eliciting their inhibitory effects by general protein aggregation. X-ray crystallography studies identified promising electron density consistent with the presence of these fragments, which were subsequently modeled into the structures. CC42201 and CC42223 were both located in the active site, where they made similar interactions. In contrast, the fragment GK00199 did not bind in the active site of AlrSas, but was instead located in a small cavity on the surface of the protein. The location of this fragment could represent a possible allosteric site on the enzyme. Overall, this study demonstrated fragment-based drug discovery to be a viable strategy for identifying non substrate-like inhibitors of AlrSas.

    View record details
  • Malnutrition in children

    Moeeni, Vesal (2014)

    Doctoral thesis
    University of Otago

    The appropriate assessment of nutrition and ongoing review of growth are important aspects of monitoring children. Overall, hospitalised children have higher rates of under-nutrition: this altered nutritional status may impact adversely upon morbidity during hospitalisation and length of hospital stay. The overall hypothesis of this project was that the optimal use of Nutritional Risk Screening (NRS) tools in children will lead to earlier detection of children at risk of malnutrition, both in developed and developing countries. Recently, three NRS tools (STRONGkids, PYMS and STAMP) have been developed in Europe for the evaluation of hospitalised children but have not been evaluated elsewhere. Therefore, these three tools were applied to 119 and 162 paediatric hospitalised patients (in Iran and NZ) respectively. The STRONGkids tool was shown to be the most reliable and feasible tool in both settings. This tool was able to recognise 83% to 100% of under-nourished patients in its at risk groups. In both studies, the nutritional state of the children (hospitalised and the comparison groups) was assessed. 9.9% of NZ hospitalised children were under-nourished, contrasting to just 3.7% of a healthy community-based group (p=0.04). An even higher rate of under-nutrition was seen in a group of children admitted to an Iranian hospital, whilst under-nutrition was again seen infrequently in the community comparison group (25.2% versus 3%; p<0.005). The utility of the STRONGkids tool was then further evaluated, with assessment of its use by paediatric nursing staff. This tool had been designed and validated to be applied by a physician. The tool was evaluated by paired nursing and paediatrician assessments, showing substantial agreement (k=0.65). This agreement was further enhanced (k=0.86) following simplification of the tool and readjustment of the cut-off points for risk stratification. Thus, the STRONGkids tool was equally reliable and feasible when applied by a physician or paediatric nurses. The final component of this work was to evaluate an NRS tool in children with Cystic Fibrosis (CF), along with delineation of their nutritional state. CF is associated with increased risk of developing malnutrition, and nutritional status adversely affects long-term outcomes. The use of tools to simply and easily identify children at greater risk of malnutrition could ensure earlier and more focused intervention, thereby contributing to superior disease outcomes. In this study, one previously developed NRS tool for CF patients (McDonald) was compared and contrasted with the criteria published by the Australasian Clinical Practice Guidelines for Nutrition in CF for sixty nine children with CF in Iran and NZ. These results were validated by comparison to each patient’s full evaluation by their clinical team. The McDonald tool and the Australasian guidelines both were able to recognise patients at risk of malnutrition. This data illustrated the benefits and utility of the McDonald tool, and especially indicates its potential role in developing countries lacking the resources of a complete assessment team. In conclusion, NRS tools have the potential to significantly improve the early recognition of under-nutrition in hospitalised children and children with chronic disease.

    View record details
  • Paleoecology and sedimentology of the volcanically active Late Eocene continental shelf, Northeast Otago, New Zealand

    Hicks, Simone Barbara (2014)

    Doctoral thesis
    University of Otago

    In Late Cretaceous times, regional subsidence resulted in deposition of a transgressive marine continental shelf sequence across the east of the South Island, which reportedly reached its peak in the Late Oligocene, followed by a Neogene regression. The Waiareka Volcanic-Ototara Limestone sequences of this study formed in the later phases of the Late Cretaceous-Mid Cenozoic transgression when Zealandia was substantially submerged. Previous studies interpreted the volcanics as formed by short-lived localized marine Surtseyan volcanoes which erupted in a continental shelf setting probably tens of km from land, remote from a source of terrigenous components. This thesis reports on the stratigraphy, sedimentology, and especially the foraminiferal micropaleontology for the basaltic Waiareka Volcanics and the immediately overlying Ototara Limestone at 3 localities in North Otago, New Zealand. These strata have been reported widely in studies on regional geology, stratigraphy, paleontology, volcanology and igneous petrology, but few details have been published on the physical stratigraphy and biostratigraphy of limestone and associated sedimentary rocks above the Waiareka-Deborah volcanics. This study uses physical sedimentology and foraminiferal micropaleontology to deduce details of the Late Eocene marine environment associated with the Waiareka-Deborah volcanics. Composite lithostratigraphic sections of 18-20 m meters were made for the limestone-dominated strata above the Waiareka volcanics at the 3 study localities of Reidston, Maheno and Bridge Point, North Otago. Samples from those sections indicated lower to middle Runangan Stage, Late Eocene (later Priabonian) for all 3 localities, bounded by the first occurrence of Bolivina pontis at the base of Runangan, about 36.4 Ma and the last occurrence of Globigerinatheka index at 34.6 Ma (end of Runangan). The absence of G. brevis suggests that the uppermost Runangan was not represented. Thus, sequences span from the base of the Runangan stage to at least the end of the Subbotina linaperta zone, uncertainly a little older than 34.6 Ma. Paleodepths were equivalent to inner to mid shelf, as deduced from micropaleontological assessments using benthic and planktic foraminifera, taking also physical sedimentology in account. Warm to subtropical paleotemperatures were deduced by the presence of warm stenothermal fossil foraminifera including species in large benthic genera such as Asterocyclina, Asterigerina, Wadella, and Cribrorotalia, as well as by planktic species such as Globigerinatheka index, and Hantkenina alabamensis.

    View record details
  • Designing magnetically interesting supramolecular architectures: towards Single-Chain Magnets

    Dhers, Sebastien (2014)

    Doctoral thesis
    University of Otago

    Single-Molecule Magnets (SMMs) are single molecules that can behave as magnets. Similarly, Single-Chain Magnet (SCM) are 1D polymers that can exhibit slow relaxation of the magnetisation. Over the last decade, interest in SCMs has increased rapidly, with the main driving force being the possibility of increasing the blocking temperature above that realised by SMMs, in order to meet this requirement for future application as memory devices. In this project, a range of magnetically interesting complexes previously studied in the Brooker research group have been prepared and combined with appropriate linkers for the construction of 1D-chains. This field, which represents a new research direction for this group, is explored herein and the results obtained using various linkers are presented. Chapter 1 presents a review of the field of SCM to date and introduces the most important concepts of magnetism. This is followed by an overview of the methods of crystallisation used to obtain SCMs, an introduction to Schiff base ligands and macrocycles and finishes with the aims of the study. In Chapter 2 the synthesis and magnetic characterisation of the 3d-4f tetrametallic macrocyclic SMM building blocks [CuII3TbIII(L1Et)(NO3)3(MeOH)]∙MeOH ([CuII3TbIII(L1Et)]), [CuII3TbIII(L1Pr)(NO3)2(MeOH)3](NO3) ([CuII3TbIII(L1Pr)]) and [CuII3TbIII(L1Bu)(NO3)2(DMF)(H2O)](NO3) ([CuII3TbIII(L1Pr)]), originally obtained by Dr. Humphrey Feltham during his PhD thesis, is presented, as they are the main building blocks used in this project. The assembly of the macrocyclic building block, [CuII3TbIII(L1Pr)], with K3[MIII(CN)6] linkers, where M = Fe, Cr or Co, results in a range of monomer, dimer and 1D chain architectures, all of which have been structurally and magnetically characterised, and are described in Chapter 3. In Chapter 4 the assembly of the macrocyclic building block, [CuII3TbIII(L1Pr)], with a (TBA)3[WV(CN)8] linker, which results in two different 1D chains depending on the crystallisation conditions, is presented. In these two chapters, a total of nine complexes using [CuII3TbIII(L1Pr)] were structurally characterised. Although no SCM behaviour was seen, metamagnet and SMM behaviour was observed. Following these results with N-donor linkers, a different approach was taken to deliberately link the 3d-4f macrocyclic building block via the Tb(III) ion by using an O-donor linker. In Chapter 5, the assembly of the macrocyclic SMM building block, [CuII3TbIII(L1Et)], with a Na2[tpa] linker (tpa2- terephthalate ion), which results in a structurally and magnetically characterised 1D chain architecture, is described. This is a proof of principle that use of an oxygen donor linker can successfully target linking of the macrocyclic Ising centres via the Tb(III) ions rather than via the Cu(II) ions, to give a 1D chain, and shows enhanced control in the design of the supramolecular architecture by using 3d-4f macrocyclic building blocks. Another strategy envisaged to enhanced the magnetic communication along these 1D-chain systems was to use a different macrocyclic building block, [NiII3LnIII(L1Pr)]. In Chapter 6, work on the [NiII3LnIII(L1Pr)] family, reproducing the synthesis of eleven macrocyclic complexes [NiII3LnIII(L1Pr)] (LnIII = CeIII, PrIII, NdIII, SmIII, EuIII, GdIII, HoIII, ErIII, TmIII, YbIII or LuIII) for magnetic characterisations, and attempts at synthesising 1D chains using the attempts [NiII3TbIII(L1Pr)] building block with a range of linkers, are detailed. Chapter 7 describes the magnetic characterisation of a different macrocyclic building block, [MnIII(L2)(NCS)2], and the results obtained when trying to synthesise 1D chains from it. The 1D architecture obtained with a terephthalate linker has been structurally and magnetically characterised. It exhibits slow relaxation of the magnetisation, associated with the SMM behaviour of the [MnIII(L2)(NCS)2] building block, and this has been quantified. In Chapter 8, work done with a third building block, [CoII2(L3)(H2O)(MeCN)5](BF4)4 ([CoII2(L3)]), is presented. Attempts at synthesising 1D chains from this noncyclic non-SMM building block are presented, as well as the [MII3(L3)3]6+ (MII = Fe2+ and Zn2+) triangles obtained when this bis-terdentate Schiff base ligand L3 was reacted with Fe(II) and Zn(II) tetrafluoroborate salts. Finally, conclusions and suggestions for future work are presented in Chapter 9. Overall in this thesis, thirty inorganic compounds have been made. Eighteen of them had been made before but were remade in order to use them as building blocks, for magnetic characterisation or for full characterisation. Twelve are original compounds amongst which seven are 1D chains. Amongst these 1D chains, no SCM behaviour was observed, but four of them show SMM behaviour associated with the building block employed, and for two of them the SMM behaviour has been quantified.

    View record details
  • Characterisation of HCO3- secretion in the small intestine of the brushtail possum, Trichosurus vulpecula

    Gill, Michael William (2014)

    Doctoral thesis
    University of Otago

    The Australian common brushtail possum, (Trichosurus vulpecula), is the most significant mammalian pest in New Zealand. The damage done by wild possums has a major impact on New Zealand flora and fauna through destruction of habitats, competitive encroachment on native ecosystems, and as a vector for bovine tuberculosis. The intestinal physiology of the possum, a metatherian mammal, differs markedly to that of eutherian (placental) mammals. This will potentially affect the efficacy of oral toxins designed for use in the control of the possum, but in addition, could potentially provide targets for the development of a possum-specific toxin. In eutherian mammals intestinal fluid secretion is driven by electrogenic Cl- secretion, with secondary HCO3- secretion a vital component of intestinal function. However, electrogenic Cl- secretion does not occur in the ileum of the brushtail possum, instead electrogenic HCO3- secretion appears to provide the driving force for fluid secretion. The Ussing short-circuit (ISC) technique revealed the duodenum and jejunum both lack the electrogenic response to the cAMP dependent secretagogues prostaglandin E2 (PGE2) (∆ISC =2±5 µA.cm-2, and ∆ISC =14±5 uA.cm-2, respectively) seen in the possum ileum (94±7 µA.cm-2). Similar responses were also seen with direct activation of adenylyl cyclase by the agonist forskolin. This is due to an absence of NaK2Cl cotransporter (NKCC1) in the small intestine, with low CFTR expression in the duodenum and jejunum compared with ileum. High levels of NaHCO3 cotransporter (NBC) expression in the ileum and duodenum indicate the potetnial for HCO3- secretion in these tissues, with the combination of CFTR and pNBC expression in the ileum accounting for electrogenic HCO3- secretion which only occurs in the distal region of the small intestine. In addition, high levels of transcript for the Cl-/HCO3- exchangers SLC26A3 (DRA) and -A6 (PAT-1) also occur in the duodenum and ileum. In the ileum immunohistochemistry localised NBC to the basolateral membrane of the villous and crypt regions and DRA and PAT-1 to the apical membrane of the villous and crypts, consistent with a proposed role for electroneutral Cl-/HCO3- exchange. However, In the duodenum NBC was predominantly restricted to the villi tip, while both PAT-1 and DRA were largely identified in intracellular vesicle-like structures, which suggests the possibility of membrane insertion and removal as a mechanism of transporter regulation in this tissue. The pH stat technique measurement of serosal-to-mucosal HCO3- flux (JHCO3sm) in the ileum and comparison to the change in current resulting from PGE2 addition, demonstrated that mucosal [HCO3-] was found to have a marked effect on the ΔISC, the electrogenic pathway being greatly enhanced under physiological conditions of high mucosal [HCO3-], and reduced in zero mucosal HCO3-. The results of this study show the possum intestinal physiology is markedly different to that of other eutherians. Unlike eutherian intestine, Cl- secretion does not occur in the possum and HCO3- is the dominant secretory anion, resulting in a high luminal [HCO3-] and pH compared to mammalian intestine.

    View record details
  • Acculturation, Stress and Coping with Cross-Cultural Transition amongst International Rugby Players

    Tanaka, Shogo (2014)

    Doctoral thesis
    University of Otago

    As a result of globalisation, the cross-cultural movement of people, product, and process has dramatically increased over the past few decades. For example, in 2010 there were 325 New Zealand rugby players plying their trade overseas, including 72 players based in Japan (Edwards, 2011). Research has demonstrated that people can experience a number of difficulties during a process of cross-cultural transition that interfere with overall psychological well-being (e.g., Ward & Kennedy, 2001). Along with these cross-cultural difficulties, athletes relocating to a different culture face and cope with unique stressors associated with their athletic lives in an unfamiliar environment (e.g., Campbell & Sonn, 2009). Since cross-cultural transition can have a significant impact on one’s psychological well-being (Berry, 1997), understanding international athletes’ experiences and the factors associated with their adaptation is becoming more important. However, within the field of sport psychology, only limited attention has been devoted to the cross-cultural adaptation experiences of athletes. The purpose of this research project was to investigate the acculturative stressors faced by international rugby players living and playing in a different country and to identify coping strategies employed to deal with these stressors and facilitate positive adaptation. Utilizing Berry’s (1992; 1997) acculturation model, this project also aimed to further develop our understanding of athletes’ acculturation experiences and promote further research and theoretical development in this area of sport psychology. In order to accomplish these goals, three studies were conducted to examine rugby players’ acculturation experiences. The first study involved in-depth qualitative interviews with 10 elite professional New Zealand rugby players competing in the Japan Top League; study two focused on 10 amateur Japanese players competing in New Zealand. In study three, a longitudinal mixed methodology, involving focus group interviews and questionnaires, was employed to examine the acculturation experiences of 15 Japanese adolescent players participating in a 22 week residential rugby programme in New Zealand. Using grounded theory as a methodology, qualitative data sets were analysed inductively and findings were discussed and reported in relation to Berry’s (1997) acculturation framework. Results from the three studies indicated that these three groups of rugby players experienced a number of similar acculturative stressors despite their different backgrounds. Not surprisingly, communication difficulties were identified as the foremost challenge. Also a number of stressors originating from rugby settings were reported in all three studies as the participants spent considerable time and effort on their rugby activities. In addition, these rugby players encountered a variety of general adaptation challenges outside of rugby settings. While many of the stressors appeared to be common across the three groups of players, each group also experienced unique and distinctive challenges in their own acculturation environment. Results also revealed that in order to cope with identified stressors, these rugby players employed a variety of coping strategies including social support, problem-focused/active coping, emotion-focused coping, and avoidance coping. The findings produced a wealth of information regarding athletes’ acculturation experiences, particularly acculturative stressors and coping strategies.

    View record details
  • Murine Norovirus Manipulation of the Immune Response

    Waugh, Emily (2014)

    Doctoral thesis
    University of Otago

    Human norovirus is a significant human pathogen that causes morbidity and mortality worldwide. Noroviruses are a member of the Caliciviridae family of positive sense, single-stranded RNA viruses. The inability to study human norovirus in cell culture has meant that advancement in prophylactic and therapeutic treatment has been slow. The discovery of the closely related murine norovirus has allowed investigation into the pathogenesis of norovirus infection as well as the host response to infection to be studied in detail. While research has focused on the essential innate immune response and to a lesser extent the adaptive immune response, little is known about the chemokine response initiated during norovirus infection. This PhD thesis has investigated the chemokine response to murine norovirus during infection of macrophages and has identified virus-specific up-regulation of selected chemokines. This virus-induced chemokine response biases the immune system towards a Th1-type response to viral infection. This response may explain the limited efficacy of antibody responses generated during norovirus infection, resulting in poor protection against infection during an outbreak. Moreover, research identified that mRNA and protein levels of selected chemokines do not show matched up-regulation during MNV infection, suggesting viral interference with host protein production, stability or secretion. Other caliciviruses, including human norovirus, are known to interfere with host protein secretion during infection. Therefore, protein secretion was investigated during MNV infection using a secretion reporter cell line (RAW-Blue cells) in addition to examination of intracellular build up of proteins. It was determined that MNV does not affect cellular secretion during viral infection. Observations from experiments using RAW-Blue cells discovered that MNV infection was unable to induce production of the innate immune response induced reporter construct. This construct is inducible by NFκB and AP-1 therefore, MNV interactions with intracellular signaling pathways, specifically NFκB activation, were examined. The NFκB regulator protein IκBα was studied and experiments demonstrated a lack of IκBα phosphorylation and degradation during MNV infection. This revealed that MNV infection leads to impaired activation of the NFκB pathway. Moreover, investigation into the mechanism of impairment discovered that the viral protein VF1 is in part responsible, as infection of RAW-Blue cells with a mutant virus lacking VF1 restored signaling through the NFκB pathway. NFκB is a global transcription factor involved in the regulation of over 100 immune genes, thus impaired NFκB activation will likely result in delayed induction of the antiviral response. The lack of NFκB activation may partly be responsible for chemokine profile observed during MNV infection, as chemokines that are predominately inducible by interferon showed much greater up-regulation than chemokines primarily induced by NFκB. Overall, this thesis describes the interaction of murine norovirus with the host cellular response and characterizes previously unknown mechanisms of norovirus immune evasion.

    View record details
  • Macrophage Migration Inhibitory Factor and Inflammatory Bowel Disease

    Falvey, James David (2014)

    Doctoral thesis
    University of Otago

    Macrophage migration inhibitory factor (MIF) is a pleiotropic pro-inflammatory cytokine and intracellular signalling molecule that is implicated in the pathogenesis of inflammatory bowel disease. Intestinal MIF concentrations are elevated in patients with IBD, and inhibition of MIF ameliorates disease activity in animal models. MIF has diverse effects. It is a pivotal mediator of innate immunity and released by immune and non-immune cells in response to both endogenous and exogenous factors. In turn, MIF stimulates the release of pro-inflammatory cytokines and mediates immune cell migration and activation through both direct and indirect mechanisms. In addition to traditional pro-inflammatory cytokine activities, MIF counter regulates the immunosuppressive effects of glucocorticoids through both local and pituitary release, and is a potent anti-apoptotic factor able to prolong the survival of tissue, inflammatory and tumour cells. In the GI tract MIF is constitutively expressed within the apical compartment of the epithelium and is predominantly secreted in an apical direction. Provisional evidence implicates MIF in several key patho-biological processes in IBD, in particular microbial sensing through both a moderating effect on M-cell function and as an effector molecule of TLR signalling; dendritic cell function; and also in the control of intestinal hypoxia and redox sensitivity and signalling. Several small molecule inhibitors of MIF have been identified although none have reached clinical trials. MIF has recently been identified as the main cellular target of isothiocyanates, a group of plant-based chemicals that are present in cruciferous vegetables that are found in the human diet. In a proof of concept investigation, consumption of a large meal of watercress (a potent source of phenethyl isothiocyanate) was found to reduce human plasma MIF concentrations. The anti-inflammatory effects of isothiocyanates as a novel therapy in IBD was investigated in a murine model of colitis. The dextran sulphate sodium (DSS) model of colitis was established, and a novel method of clinical disease scoring was developed and prospectively validated. Animals were dosed with isothiocyanates and the MIF inhibitory capacity was investigated. Animals were examined for evidence of toxicity and the efficacy of ITC in preventing DSS colitis was investigated. No significant beneficial treatment effect was observed. Indeed, significant gastric toxicity occurred following oral administration. Gastric toxicity from ITC has not been reported previously and the mode of action of this effect is unknown. In further experiments, the efficacy of rectally administered ITC was investigated. Although a pilot investigation showed a marked effect, in subsequent experiments, no statistically significant effect was observed. No toxicicity was observed in the colon following rectal administration. The contribution of two functional promoter polymorphisms of MIF to IBD susceptibility and phenotype was investigated. The investigation, the largest of its kind, aimed to define the contribution of MIF variants to IBD risk in the Canterbury population, and to resolve, through meta-analysis, discordance in results from previous investigations. Within the Canterbury data set, no evidence was found for an association between either variant, whether considered individually or together, with respect to IBD risk, phenotype, disease behaviour or clinical course. A weak trend toward protection against ileal CD and MIF-173C was observed. With respect to rs755622; meta-analysis found no significant association between MIF-173C and risk of UC, CD or overall IBD. These data do not undermine the importance of MIF to the pathobiology of IBD, but emphasise previous observations that intestinal MIF concentrations are largely determined at the post translational level. In an investigation of MIF as a biomarker of disease in IBD, the relationship between plasma MIF and endoscopic disease severity was investigated. The investigation found no correlation between plasma MIF and gold standard disease assessment, indicating that MIF has no value as a biomarker of disease. The research group is continuing to investigate the biological properties of MIF in human disease, with particular emphasis on endogenous MIF control and the effect of MIF inhibition in vitro and in vivo. These investigations are timely given that international phase one trials are currently underway of a neutralizing anti-MIF antibody in human disease. Greater understanding of the contribution of MIF to IBD is urgently needed in order for patients with IBD to benefit from these advances

    View record details
  • An Investigation of Magnesium Alloys as Orthopaedic Biomaterials

    Walker, Jemimah (2013)

    Doctoral thesis
    University of Otago

    Magnesium (Mg) and its alloys were initially investigated as resorbable orthopaedic biomaterials more than a century ago. However, their use was abandoned due to rapid corrosion resulting in mechanical failure and excess hydrogen production. Whilst recent advances in manufacturing processes have led to a renewed interest in the field of Mg based biomaterials, the corrosion behaviour remains the feature that is the most difficult to control and characterise. In particular, the development of these materials is hindered by an inability to use in vitro techniques to accurately predict the behaviour of Mg alloys in an in vivo environment. Furthermore, the biocompatibility of the materials is often underemphasised in favour of the assessment of corrosion behaviour. The purpose of this thesis was therefore two-fold. Firstly, the aim was to develop and optimise a series of both in vitro and in vivo methodologies that would allow the prediction of corrosion rate, and the assessment of the biocompatibility of Mg materials. Secondly, these methods would be used to identify Mg alloys that could be investigated for use as orthopaedic biomaterials. The initial portion of this study focussed on the development of an in vitro immersion method that could be used to cautiously predict the in vivo corrosion rates of Mg alloys. The in vitro protocol involved the use of three different solutions with varying degrees of physiological relevance, the results from which were compared to the corrosion occurring in an in vivo subcutaneous environment. The solution providing the most comparable corrosion rates was identified and subsequently utilised to select four slowly corroding Mg alloys for further investigation. Preliminary analysis of the biocompatibility of these four alloys was carried out using an in vitro cell culture technique. The results of this investigation indicated that the use of a closed cell culture environment is likely to reduce cellular viability due to the increased pH and osmolality associated with rapid Mg corrosion. Accordingly, it was concluded that whilst cell culture techniques can provide results indicative of the degree of biocompatibility, they must be used in conjunction with the results of in vivo investigations for a comprehensive understanding of the true biocompatibility of Mg based materials. Consequently, an in vivo soft tissue study was undertaken involving the implantation of the four Mg alloys in both intermuscular and intramuscular locations in Lewis rats. This investigation indicated minimal hydrogen production associated with the Mg alloys, and biocompatibility equivalent to that seen with the implantation of a biomedical Ti alloy (Ti-6Al-4V). The results from both the in vitro and in vivo studies identified Mg-1Zn and Mg-0.4Ca as the alloys exhibiting optimal characteristics for further investigation as orthopaedic implant materials. An intraosseous study was carried out involving the implantation of these alloys into the shaft and epiphyseal region of the ovine tibia. The results indicated both alloys were biocompatible and corrosion resistant, indicating their potential as promising resorbable orthopaedic biomaterials.

    View record details
  • Sleep disordered breathing in young children: Natural history and relations to academic performance

    Luo, Rebekah Pei Ru (2015)

    Doctoral thesis
    University of Otago

    Objective: The aims of this current study were to: (i) examine the natural history of snoring and other sleep disordered breathing (SDB) related symptoms from age 3 to 7 years; (ii) examine the concurrent relationship between SDB and academic performance of 6-year old children and explore potential pathways linking SDB to academic performance; (iii) investigate whether parent-reported habitual snoring at age 3 predicts later academic performance and behavioural outcomes at age 7. Methods: Parent/s of children (n = 839) who returned a completed Age 3 Community Survey (conducted in 2008 by Dr. Amelia Gill) were re-contacted via mail four years later when children were aged 6-8 years, and asked to complete a Follow-up Survey. This follow-up survey included questions regarding their child’s sleep, academic performance and behavioural adjustment. A subgroup of 170 children from the survey sample were involved in a longitudinal study examining links between SDB, early learning and behavioural adjustment in 3- and 4-year olds. As part of the current research, these children were re-assessed at age 6 (n = 163, mean age 6.3 years) (this study was also known as the Academic Assessment Phase). SDB was assessed through parent-reported history of symptoms, and physical examination of features related to SDB. Children’s neurocognitive functioning and academic performance was assessed using researcher-administered standardised tests, and parent- and teacher-ratings. Results: At follow-up (age 7), habitual snoring was prevalent in 9.2% of the sample, similar to the 11.3% reported at age 3. However, habitual snoring changed over time; some children (36.2%) remained habitual snorers over the follow-up period, while some others were no longer snoring habitually. In addition, a small group of children (5.3%) started habitual snoring since the Age 3 Community Survey. Additionally, history of habitual snoring at age 3 predicted small but statistically significant unique variance in parent ratings of literacy outcomes and working memory functioning at age 7, even after controlling for important child socio-demographic and health correlates. Findings from the Academic Assessment Phase at age 6 revealed statistically significant correlations between SDB severity and several cognitive functioning measures (r = -.21 to -.26, p < .01), ratings of executive functioning difficulties (r = .25 to .32, p < .01), literacy and numeracy outcomes (r = -.21 to -.25, p < .01), and rating of academic performance (r = -.23, p < .01). Using a structural equation modeling (SEM) approach, indirect paths between SDB and academic performance were found, in which SDB was found to be directly related to domains of executive functioning, verbal comprehension and communication, and nonverbal reasoning; both verbal and nonverbal composites were predictive of academic performance. General pattern of findings remained even when child’s gender and Deprivation Index was taken into account, respectively. Conclusion: The current findings highlight the dynamic nature of sleep-disordered breathing in children, where related-symptoms can develop, remain present or resolved at different stages of childhood. Findings from this research also suggest that pathways from SDB to poorer academic performance were indirect, and mediated through children’s verbal and nonverbal abilities. Additionally, history of habitual snoring in preschool years were predictive of children’s later literacy achievement and working memory functioning.

    View record details
  • Quantitative ultrastructural differences in the cytoplasm of prepubertal lamb and adult ewe oocytes

    Reader, Karen Lee (2014)

    Doctoral thesis
    University of Otago

    The ability to breed from juvenile sheep and cattle can increase their lifetime productivity and the rate of genetic gain, and thus increase the profitability of the agricultural industry. Unfortunately, oocytes from prepubertal animals have limited potential to undergo normal embryogenesis and produce viable offspring, and little is known about the underlying causes of this reduced competence. The correct quantity, activity and cytoplasmic distribution of organelles are essential for oocyte maturation, fertilisation and subsequent embryo development. However, very few studies have attempted to quantify differences in the cytoplasm of prepubertal and adult oocytes to understand the underlying causes of the reduced quality. Therefore, the aims of this project were firstly; to determine if there were differences in the volumes, morphology or distribution of organelles between oocytes from prepubertal (lamb) and adult ewes both before and after in vitro maturation; and secondly, to determine if treatment of lamb oocytes in vitro with factors that may alter the volume or distribution of these organelles can improve embryo development. Embryos were produced from oocytes from slaughterhouse derived ovaries using standard sheep in vitro embryo production methods to determine blastocyst rate. Oocytes were fixed and processed for electron microscopy and stereology techniques were used to measure the distribution, density and volumes of vesicles, mitochondria and lipid droplets in 12 oocytes per age and time point. The volume of vesicles before maturation was greater in adult ewe compared to lamb oocytes (P = 0.018), and decreased during maturation in both age groups. The volume of mitochondria in adult ewe oocytes increased during maturation (P = 0.005). This was due to an increase in mitochondrial number and was greater than the mitochondrial volume (P = 0.026) and number (P = 0.003) of the mature lamb oocytes, which did not change. There were also differences in the morphology of the mitochondria and the distribution of mitochondria and lipid droplets between adult and lamb oocytes. Studies have attributed improved oocyte competence to an increased number of mitochondria and altered cytoplasmic mitochondrial distribution. However, estimates of mitochondrial number have mainly been based on mitochondrial DNA (mtDNA) copy number performed by qPCR methods while the actual number of copies of mtDNA per mitochondria is not known. Therefore, the current study compared the mtDNA copy number, measured by qPCR, with mitochondrial number estimated by stereology in oocytes from the same cohorts of adult ewes and lambs. Unlike the mitochondrial number estimated by stereology, the mean number of copies of mtDNA did not differ between the two ages either before or after in vitro maturation, but decreased in the lamb oocytes during maturation. These results indicate that mtDNA copy number does not represent actual mitochondrial organelle number. L-carnitine has been shown to increase blastocyst rate and alter mitochondrial distribution when cultured with oocytes during in vitro maturation. It has also been shown to alter expression of mitochondrial biogenesis genes in other cell types. In the current study lamb oocytes were treated with acetyl-L-carnitine during in vitro maturation and this doubled the blastocyst rate compared to untreated oocytes (P = 0.028). Stereology results showed vesicle volume and size, and lipid droplet distribution appeared to be altered. There was no effect of L-carnitine on mitochondrial volume, number or distribution, or mtDNA copy number. In conclusion the results of this research suggest a lower volume of storage vesicles prior to maturation and altered mitochondrial replication, distribution and morphology may underlie the reduced developmental competence of prepubertal lamb oocytes. Treatment with L-carnitine during in vitro maturation can also be used to improve in vitro embryo production methods for oocytes from juvenile lambs.

    View record details
  • Angiogenic Factors in Oral Cancer

    Allsobrook, Olive Florence Louise (2014)

    Doctoral thesis
    University of Otago

    Angiogenesis is a key factor in normal development and in the development and progression of most malignancies, including oral cancer. It is also an integral component of some inflammatory hyperplastic lesions including those that occur in the oral regions such as pyogenic granulomas (PG). Physiological angiogenesis is controlled mostly by vascular endothelial growth factor A (VEGF)/VEGF receptor 2 (VEGFR2) signalling, which promotes endothelial cell migration and proliferation, and blood vessel growth and by vasohibin 1 (VASH-1), an intrinsic inhibitor of angiogenesis. VEGFR2 is also known as kinase domain receptor (KDR) and this term will be used in this thesis. During pathological processes, the loss of angiogenic regulation can contribute to inflammatory hyperplasias and tumour progression. Aim: To investigate angiogenic regulation in hyperplastic vascular oral lesions and oral squamous cell carcinoma (OSCC). Methods: Archival formalin-fixed paraffin embedded (FFPE) tissue from ten gingival PGs, ten cases of OSCC and five normal oral mucosal (NOM) specimens were analysed for angiogenic markers using standard immunohistochemistry techniques with the chromogen 3, 3’-diaminobenzidine (DAB). Anti-VEGF, anti-KDR and anti-VASH-1 antibodies identified angiogenic and anti-angiogenic activity in the tissues. Mean vessel density (MVD) was assessed using the endothelial cell marker anti-CD146 antibody. Blood vessels were counted using the anti-CD34 antibody as a marker of positive endothelial cells. Immuno-positivity was evaluated by light microscopy for qualitative analysis and five randomly chosen fields were photographed for semi-quantitative and quantitative analysis. VEGF was analysed semi-quantitatively at 200x magnification and the percentage of positive cells analysed using chi-squared statistics. VASH-1+, KDR+ and CD34+ were counted at 400x magnification while CD146+ and VEGF+ cells were counted at 200x magnification. These groups were analysed using a log transformation and oneway analysis of varience (ANOVA). Statistical significance for quantitative analyses was at the 5% level. Qualitative analysis of overall staining patterns was also undertaken. Gene expression analysis was performed on granulation tissue and cell lines from three OSCC and three normal epithelial cell lines. A quality control assay was run using a granulation tissue sample as a control for specificity and validity of the assays. Gene expression was analysed using quantitative real-time polymerase chain reaction (qRT-PCR), 7500 Fast System SDS software v.1.3.1, and SA Biosciences RT2 Profiler PCR Array Data Analysis v3.5 with a 5 % level of significance, significant fold change was >2. Results: All tissues were positive for endothelial cell and angiogenic and anti-angiogenic markers. Endothelial cells were positively identified in all tissues. VEGF was found in the stroma and the overlying epithelium, especially in the OSCC group. VASH-1 was positive in blood vessels in all tissues but also in some malignant epithelial cells. The MVD was significantly greater in PGs than in normal oral mucosa and in OSCC (p<0.001) and the number of CD34+ cells was also significant (p=0.01). In respect to expression in blood vessels, qualitative analysis of the tissues showed a greater epithelial intensity of staining for VEGF in OSCC. More VASH-1+endothelial cells were present in PGs than in the other tissues with a trend towards significance (p= 0.05). There was no significant difference in VEGF staining observed in OSSC by comparison with PG or NOM. The expression of staining for KDR showed no significant difference between the tissue types. Gene expression mRNA analysis of cell lines showed a statistically significant increase in expression of VEGF in OSCC cell lines compared with normal epithelial cells (p=0.01). The levels of VASH-1 and KDR could not be determined from the analysis, because of low levels of expression. Discussion and Conclusion: The presence of VEGF, KDR and VASH-1 in OSCC tissue demonstrates the presence of angiogenic signalling. OSCC showed greater epithelial VEGF expression in IHC and OSCC cell lines but VASH-1 expression showed a tendency towards fewer stromal cells staining positively compared with PG tissues, indicating that angiogenesis self-regulating mechanisms may be altered in OSCC. PG tissues had a greater MVD and showed expression of VASH-1 in many endothelial cells. From this, it appears that angiogenic factors may be expressed differently in hyperplastic lesions compared with OSCC. Expression of these angiogenic factors was seen in mRNA analysis of vascular granulation tissue and in OSCC and epithelial cell lines, with greater relative expression of VEGF in OSCC cells compared with normal epithelium. Further research may show the implications of this for treatment of OSCC and diseases related to angiogenesis, with potential for regulation with angiogenesis inhibitors, and development of prognostic biological markers.

    View record details
  • Psychological Aspects of Inflammatory Bowel Disease

    McCombie, Andrew (2014)

    Doctoral thesis
    University of Otago

    Inflammatory bowel disease (IBD), comprising Crohn’s disease, ulcerative colitis, and IBD-unspecified, can often have severe and debilitating symptoms. These symptoms can often lead to significant depression, anxiety, and impaired health-related quality of life (HRQOL). Not only can the symptoms themselves lead to psychological distress (PD) but certain individual characteristics, such as personality and coping strategies, may determine how one responds to the disease and their subsequent psychological outcomes. This thesis explores the interrelationships between IBD and psychological factors using both observational and interventional techniques. Chapter 1 provides a general overview of IBD in terms of its symptoms, worldwide epidemiology, causes, and treatments. Chapter 2 discusses how IBD can lead to PD in many of its patients and that improvements in coping strategies may improve outcomes in IBD patients. Psychological interventions may be a treatment for at least some IBD patients. Chapter 3 systematically reviews psychotherapy for IBD. However, given that psychotherapy is expensive and difficult to access, computerised psychological interventions may be a more cost-effective and accessible treatment to many patients. No studies have been published in computerised psychological interventions in IBD patients and so Chapter 4 systematically reviews computerised cognitive behavioural therapy (CCBT) as an intervention for physical illnesses in general, the first time that this has been reviewed. Chapter 5 comprises an original observational longitudinal study of newly diagnosed IBD patients during the first six months post diagnosis. This study showed that coping changed, HRQOL improved, and neuroticism had strong associations with anxiety, depression, and HRQOL. As to whether the coping strategies are a cause or a consequence of fluctuations in HRQOL needs to be tested in an interventional study. Chapter 6 contains a study comparing patient preferences for a computerised or face-to-face psychological intervention. An appetite for a computerised psychological intervention was demonstrated and so was tested in Chapter 7 in the form of a randomised controlled trial of CCBT for IBD patients. Few participants complied with the CCBT regiment but those who did trended towards having better improvements in HRQOL post-intervention than controls. However, these effects were not maintained at six months post-intervention. Therefore, the overall result of this study is negative. One of the conclusions from Chapters 2 and 5 was that an IBD specific coping questionnaire should be developed. Chapter 8 contains such a questionnaire, namely the IBD Cope. The “good” and “bad” coping subscales of the IBD Cope were shown to match the Brief COPE adaptive and maladaptive coping subscales despite the IBD Cope having fewer questions than the Brief COPE. The most important findings from this thesis in Chapter 9 are that (1) coping is better measured in IBD patients using the IBD Cope than the Brief COPE; and (2) CCBT can improve HRQOL in the short term for IBD patients but adherence needs to be improved and long term benefits need to be maximised through “booster” sessions.

    View record details