Bacteria can quickly adapt to changing environmental conditions by activating alternative sigma factors. It has been shown previously that single cell approaches can reveal hidden dynamics in sigma factor activation. Here, we investigate the single cell response dynamics of the B. subtilis extracytoplasmic function sigma factors, which are an important part of the cell envelope stress response, under their specific stresses. To do this we use transcriptional reporters of sigma factors, quantitative single cell snapshots, time-lapse microscopy, and microfluidics. By developing an improved microfluidics setup for single cell time-lapse microscopy, as well as improved single cell analysis code, we are able to observe new sigma factor dynamics. First, we observe heterogeneous entry into a higher $\sigma^{V}$ activity state in response to lysozyme, which displays a memory, as the heterogeneity is lost on removal and reapplication of the stress. Next, we observe a pulse amplitude and duration modulated sigma factor response of $\sigma^{M}$ to bacitracin. Finally, for $\sigma^{M}$ under ethanol and acidic stress, and for $\sigma^{Y}$ under ethanol stress, we observe a noisy increase in activity to a new steady state level, where the degree of variability between cells depends on the stress condition. This thesis also discusses efforts on building a single cell microfluidic device based on the ”mother machine” design, for the rod-shaped cyanobacterium, S. elongatus, which forces the cells to grow in a straight line. Growing this organism in a traditional mother machine device has, so far, proved challenging. To adapt the mother machine for cyanobacteria we modify the channel geometry using electron beam lithography, and improve the loading protocol. The research presented here reveals the range of regulatory dynamics possible for ECF sigma factors in B. subtilis, and provides improved microfluidics and analysis code that will enable easier quantification of bacterial gene circuits at the single cell level in the future.

Peanut allergy is a public health concern which affects a significant proportion of the population. Accidental exposure to peanut can cause severe and fatal reactions in peanut allergic individuals and currently their only safeguard is to practise careful avoidance. Identification and protection of at-risk members of the allergic population is critical in managing this life-threatening condition. This thesis produces key data to enable this. A prospective study was performed on 60 peanut allergic participants to determine thresholds of reactivity to peanut using oral challenges with incrementally increasing amounts of peanut protein. Following a double-blind placebo-controlled peanut challenge, participants received three further peanut challenges, two with co-factors: sleep deprivation and exercise, and one without. Severity was measured using a numerical scale derived from symptoms and serum tryptase was measured at each challenge. A total of 187 challenges were performed. Findings were that the median amount of peanut protein which induces a reaction in 10% of the population (ED10) was 12.3mg (95% CI 7.3,20.4) equivalently this suggests that 90% of the allergic population will not react to doses below this level. Both sleep deprivation and exercise have a significant effect on lowering reaction threshold (ED10), by 5 times and 2.5 times respectively. Separately there is a reduction in threshold with successive challenges. Co-factors also significantly increased symptom severity during challenge reactions. In particular sleep deprivation significantly increased the severity of gastrointestinal symptoms suggesting that a stressful stimulus may affect intestinal permeability. Evidence was provided for the importance of asthma as a risk factor which increased the severity of respiratory symptoms during reaction. Using a novel visual analogue scale for measuring the participant’s perception of severity, a poor correlation was observed between the participant’s perception of the reaction and the overall numerical severity score, suggesting that participants misperceive severe symptoms. This thesis provides the first data showing that symptom patterns in repeated challenges show a high degree of homogeneity within individuals, but importantly that this symptom homogeneity is also observed across individuals. Lastly the utility of serum tryptase in identifying food allergic reactions has been disputed previously. This thesis provides evidence of its value and identifies a rise cut-off of 30% as being diagnostic of a food allergic reaction, but cautions that acute levels must be compared with baseline as this rise may occur within the normal range.

In order to understand gene regulatory networks (GRNs) in mammalian cells, it is pivotal to assess the interaction between proteins and DNA. In particular, the specific DNA binding activity of transcription factors (TFs) determines the expression of target genes and in general the overall connectivity of the GRN. However, the genomic location of TF binding cannot be predicted just from the DNA sequence, and functional assays are required to detect this interaction. The investigation of the binding of TF to DNA is usually accomplished by chromatin immunoprecipitation followed by next-generation sequencing (ChIP-seq). While in the last 10 years this method enabled a better understanding of how transcription is regulated in living cells, it does have some drawbacks. In particular, the need for very highly specific antibodies and the large amount of starting material limit the ability of ChIP-seq to address biological questions when dealing with samples of small quantity. A technique called DNA Adenine Methyltransferase Identification (DamID) was developed in Drosophila as an alternative method for the detection of protein- DNA interactions and it is based on the fusion of a protein of interest (POI) with the DNA adenine methyltransferase (Dam). This fusion causes DNA methylation of adenines surrounding the sites where POI binds and the subsequent identification of the methylation sites allows mapping of the binding event without antibodies and using less cells as starting material. While this technology was successful in detecting the interaction between nuclear lamina and DNA in mammalian cells, to date little reports are present in the literature about TF DamID. This is mainly due to the different nature of TF binding compared to Lamin (punctuated instead of broad) and to the elevated intrinsic activity of Dam that makes the detection of real signal above the noise challenging. I here demonstrate a step-by-step optimization of the DamID technology coupled to next-generation sequencing (DamID-seq) that I used to map the binding of the mouse embryonic stem cell master regulator Oct4 in as few as 1,000 cells. This new technology paves the way for exciting new experiments where the number of cells is scarce such as in vitro cell state change or in vivo processes.

Introduction Non-union (NU) is a fracture that will not unite. With over one million fractures per annum in the UK long bone non-union has serious social and economical implications. There is little epidemiological data available specifically looking at this NU patient cohort. Studies that are bone specific quote rates of non-union as a proportion of their study group but there is no data quantifying the incidence of NU in the population or per fracture. Studies have highlighted risk factors associated with atrophic non-union including age, diabetes, non-steroidals, and cigarette smoking. There is scientific interest regarding how best to classify non-unions and the role of biological agents in treating them. Aims • To quantify the incidence of non-union in a large population and calculate the risk of non-union per fracture according to age, sex and anatomical distribution. • To assess the causes contributing to non-union and outcomes of treatment in a non-union cohort and validate a new non-union scoring system. • To test the treatment potential of a novel molecule (monobutyrin) and a growth factor in a small animal model of non-union. Method • Using the ICD-10 data from the Scottish population as collected by NHS Scotland the incidence of non-union and fractures were calculated. • A cohort of 100 non-union patients were studied for risk factors associated with their non-union, treatment outcome and to assess a new NU classification system. • A rat model of tibial non-union was used to assess the potential of monobutyrin and BMP-2 in treating non-union in an animal model. Results • Fracture non-union is very rarely found in children (1 in 500 fractures) and occurs in up to 1 in 50 adult fractures. Non-union of a fracture has a significantly higher risk in young adults than the elderly by about 3 fold. Osteoporosis may not be a risk for non-union. The tibia and clavicle are the sites with the greatest potential for fracture non-union. • Non-union is multifactorial in two out of three patients. Biomechanical stability, patient host factors and infection must all be considered in every patient. Occult or unexpected recurrent infection is present in up to 10% of patients. When all factors are considered in treatment the outcome is 95% successful with 88% requiring 2 or less procedures to heal the non-union and only a minority requiring adjuvant graft or biological agents. The proposed new classification system is complex and did not clearly identify those patients who would require adjuvant treatment (eg bone grafting or BMP) or those likely to have unsuccessful non-union treatment. • Monobutyrin and BMP-2 when tested on the small animal non-union model did not improve the success rate of union. Conclusions Non-union affects approximately 1000 people per year in Scotland, this figure is not as high as 5-10% of all fractures. It is associated with fractures in young adults and of the clavicle and tibia, treatment can have a very high success rate without the need for adjuvant biological polytherapy when all contributing factors are considered and managed appropriately. A new non-union classification needs to incorporate the multifactorial aspects of non-union without being too complex to use in everyday clinical situations.

nÃo hà / As mortes desencadeadas em crianÃas menores de um ano de vida sÃo influenciadas por fatores biolÃgicos, culturais, demogrÃficos, socioeconÃmicos e assistenciais. Este trabalho teve como objetivo avaliar a evitabilidade dos Ãbitos infantis ocorridos na Secretaria Executiva Regional VI em Fortaleza-CE, em 2008 e 2009. Trata-se de um estudo do tipo ecolÃgico, descritivo e quantitativo, tendo como desfecho a ocorrÃncia do Ãbito infantil. Foram estudados 192 Ãbitos, de uma populaÃÃo de 240. As variÃveis foram agrupadas de acordo com as caracterÃsticas sociodemogrÃficas e maternas. Analisaram-se as mÃdias, medianas e IC95% de variÃveis quantitativas. Para avaliaÃÃo da evitabilidade dos Ãbitos, foram usados os critÃrios propostos pela Lista Brasileira de Mortes EvitÃveis no Sistema Ãnico de SaÃde, apÃs anÃlise pelo Comità Regional de PrevenÃÃo do Ãbito Infantil e Fetal. Em 2008,54% e 2009,52% dos Ãbitos em menores de um ano foram considerados evitÃveis pelo ComitÃ. Comprovou-se que 30% dos Ãbitos de menores de um ano ocorridos em 2008 sÃo reduzÃveis por aÃÃes adequadas de diagnÃstico e tratamento; 26% por adequada atenÃÃo à mulher na gestaÃÃo; 22% por aÃÃes adequadas de promoÃÃo à saÃde; 12% reduzÃveis por adequada atenÃÃo ao recÃm-nascido; 8% por adequada atenÃÃo à mulher no parto, e 2% reduzÃveis por aÃÃes de imunoprevenÃÃo. Dos Ãbitos evitÃveis em 2009, 44% foram classificados como reduzÃveis por adequada atenÃÃo à mulher na gestaÃÃo; 33% por aÃÃes adequadas de diagnÃstico e tratamento; 10% por adequada atenÃÃo ao recÃm-nascido; 7% por aÃÃes adequadas de promoÃÃo à saÃde; e 6% reduzÃveis por adequada atenÃÃo à mulher no parto. Os resultados encontrados podem ser utilizados para o direcionamento de intervenÃÃes efetivas que visem a diminuir a mortalidade infantil, tanto no Ãmbito da Secretaria Executiva Regional VI, como nas demais secretarias do municÃpio de Fortaleza. / Deaths triggered in children less than one year of life are influenced by biological, cultural, demographic, socio-economic and assistance factors. This study describes the epidemiology profile of infant deaths that happened in the 6th Regional Executive Office in Fortaleza-CE, Brazil, in 2008 and 2009 and assesses its avoidability according to the results of the Regional Committee for the Prevention of Infant and Fetal Death (CRPOIF). This is an ecological, descriptive and quantitative study, with the outcome of the occurrence of infant death. 192 deaths were studied in a population of 240. Variables were grouped according to maternal socio-demographic characteristics. For data processing we used the Epi-Info software version 3.5.1, with analysis of means, medians of some variables and adopted the Confidence Interval of 95%. To assess the avoidability of deaths, we used the criteria proposed by the Brazilian List of Preventable Deaths in the National Health System after being reviewed by the committee. In 2008 (54%) and 2009 (52%) of deaths in children under one year of life were considered preventable by the CRPOIF based on this list, and following the classification adopted to analyze the deaths. It was concluded that 30% of deaths of children under one year old in 2008 are avoidable by appropriate actions of diagnosis and treatment, 26% by appropriate care to women in pregnancy, 22% by appropriate actions for health promotion, 12% are avoidable by appropriate care to newborns, 8% by appropriate care to women during childbirth, and 2% reduced through actions of vaccine prevention. From the avoidable deaths in 2009, 44% were reduced through appropriate care to women in pregnancy, 33% by appropriate actions for diagnosis and treatment, 10% by appropriate care to newborns, 7% by appropriate actions to health promotion and 6% reduced through appropriate care to women in childbirth. These results can be used to address effective interventions that aim to reduce infant mortality, both within the 6th Regional Executive Office, as in the other offices of this city.

In this thesis, I explore a number of epistemological issues concerning the relations between knowledge, belief and practical matters. In particular, I defend a view, which I call credal pragmatism. This view is compatible with moderate invariantism, a view that takes knowledge to depend exclusively on truth-relevant factors and to require an invariant epistemic standard of knowledge that can be quite easily met. The thesis includes a negative and a positive part. In the negative part (Ch. 1-4) I do two things: i) I critically examine some moderate invariantist accounts of the intuitive influence of practical factors on knowledge ascriptions, and ii) I provide a criticism of the idea that knowledge is the norm of practical reasoning. In Chapter 1, I provide a general overview of the issues that constitute the background for the views and arguments defended in my thesis. In particular, I provide a thorough discussion of two aspects of the relation between knowledge and practical matters: one is constituted by the practical factors' effects on knowledge ascriptions; the other is the intuitive normative role of knowledge in the regulation and assessments of action and practical reasoning. In Chapter 2, I consider and criticize Timothy Williamson's account according to which an alleged failure to acknowledge the distinction between knowing and knowing that one knows generates the intuition that knowledge ascriptions are sensitive to practical factors. In Chapter 3, I argue against the idea that practical reasoning is governed by a knowledge norm. The argument generalizes to other candidate epistemic norms of practical reasoning. In Chapter 4, I criticise a number of accounts which explain effects of practical factors on knowledge ascriptions in terms of the influence of practical factors on belief. These include the accounts of Brain Weatherson, Dorit Ganson, Kent Bach and Jennifer Nagel. In the positive part of the thesis (Ch. 5-6), I develop and argue for credal pragmatism, an original account of the nature and interaction of different doxastic attitudes and the role of practical factors in their rational regulation. On this view, given a certain fixed amount of evidence, the degree of credence of an adaptively rational agent varies in different circumstances depending on practical factors, while the threshold on the degree of credence necessary for outright belief remains fixed across contexts. This account distinguishes between two kinds of outright belief: occurrent belief, which depends on the actual degree of credence, and dispositional belief, which depends on the degree of credence in normal circumstances. In Chapter 5, I present the view and I show how credal pragmatism can explain the practical factors' effects on knowledge ascriptions. In Chapter 6, I develop a fallibilist account of several features about knowledge ascriptions including i) why in folk epistemological practices knowledge is often taken to be a necessary and sufficient epistemic condition for relying on a proposition in practical reasoning; ii) concessive knowledge attributions and related data; and iii) the infallibilist intuition that knowledge excludes error possibilities.

Scientific interest in the clinical implications, public health importance, and risk factors of anxiety disorders has grown substantially in the past two decades. Despite this, the evidence base on anxiety is insufficient to inform health care planning and policy-making. Further research on the outcomes and risk factors associated with anxiety disorders, and ways of mitigating these risks is needed. One of the aims of this thesis was to provide an overview of the existing literature on the prevalence of anxiety in adults living in countries across the globe, and to describe the prevalence in the context of various health states and life stages. Because generalised anxiety disorder is one of the most common psychiatric conditions in the population, the remainder of the thesis focused on this disorder and aimed to explore its links with health service use and mortality. Risk for this condition was also explored and area deprivation was studied as a possible determinant. Since depression is commonly studied alongside anxiety, the relationship between the residential environment and major depressive disorder was also assessed. Finally, to provide insight into the mitigation of risks of generalised anxiety disorder, a study of coping mechanisms was undertaken. Primary study findings from this thesis are based on the European Prospective Investigation of Cancer in Norfolk, a large, population study of British people over the age of 40. Results from the systematic review showed that anxiety is common in population sub-groups around the world, with women, younger people, and those suffering from chronic physical conditions, such as cancer and cardiovascular disease being particularly affected. Results from the primary studies of the thesis showed that generalised anxiety disorder is associated with increased risk for deaths, though it is not significantly associated with non-psychiatric hospital admissions. Results from the risk factor analyses showed that living in a deprived area is associated with generalised anxiety disorder in women and major depressive disorder in men. The risk mitigation analysis indicated that sense of coherence is an important coping mechanism that can protect against generalised anxiety disorder among women living in disadvantaged circumstances. My work has shown novel associations and attempted to provide a more complete picture of one of the most common psychiatric conditions in the population by focusing on several angles: health outcomes, risk factors, and ways of mitigating risks.

Employees have begun to be seen as the most important resource for many companies and, according to researchers, the focus on human capital is expected to be even greater in the future. The purpose of the study was to investigate which organizational psychological factors that had the strongest effect and connection with success. The factors studied were leadership, skills development, freedom, pride and corporate culture. All 79 participants, from five different companies, answered the survey with questions regarding these five factors. Success was studied from four perspectives: financial profitability, staff turnover, employees' experience of focusing on human capital in the company and their experience of success in comparison with other companies in the same industry. Stepwise multiple regression was used as a statistical analysis in the study and the result showed that the factor of corporate culture was the one that consistently had the greatest effect on all of the dependent variables. On a shared second place was the variables competence development and freedom. All correlations were significant except for success based on economic profitability where the effects were too low to become significant. / Medarbetare har börjat ses som den viktigaste resursen på många företag och enligt forskare förväntas fokus på humankapitalet bli än större framöver. Syftet med studien var att undersöka vilka organisationspsykologiska faktorer som hade starkast effekt och samband med framgång. Faktorerna som studerats var ledarskap, kompetensutveckling, frihet, stolthet samt företagskultur. Samtliga 79 deltagare, från fem olika företag, besvarade enkäten med frågor rörande dessa fem faktorerna. Framgång studerades utifrån fyra perspektiv: ekonomisk lönsamhet, personalomsättning, medarbetarnas upplevelse av fokus på humankapitalet i företaget samt deras upplevelse av framgång i jämförelse med andra företag inom samma bransch. Stegvis multipel regression användes som statistik analys i studien och resultatet visade på att faktorn företagskultur var den som genomgående hade störst effekt på samtliga av de beroende variablerna. På en delad andra plats kom variablerna kompetensutveckling och frihet. Alla korrelationer blev signifikanta med undantag för framgång baserat på ekonomisk lönsamhet där effekterna var för låga för att ge utslag.

abstract: Previous research has shown that perceptual illusions can enhance golf putting performance, and the effect has been explained as being due to enhanced expectancies. The present study was designed to further understand this effect by measuring putting in 3 additional variations to the Ebbinghaus illusion and by measuring putting kinematics. Nineteen ASU students with minimal golf experience putted to the following illusion conditions: a target, a target surrounded by small circles, a target surrounded by large circles, a target surrounded by both large and small circles, no target surrounded by small circles and no target surrounded by large circles. Neither perceived target size nor putting error was significantly affected by the illusion conditions. Time to peak speed was found to be significantly greater for the two conditions with no target, and lowest for the condition with the target by itself. Suggestions for future research include having split groups with and without perceived performance feedback as well as general performance feedback. The size conditions utilized within this study should continue to be explored as more consistent data could be collected within groups. / Dissertation/Thesis / Masters Thesis Human Systems Engineering 2019

Domestic violence perpetrated by men against women persists as a major human rights issue in the United States and around the world with some estimates showing that one in four women will be victimized in their lifetimes. Victims face many obstacles when they try to leave their abusive partners. A major barrier faced by victims is secondary victimization. Secondary victimization is the process in which informal (e.g., family, friends, etc.) and formal (e.g., police officer, judges, etc.) supporters re-victimize the victim by questioning the validity of the abuse, excusing the perpetrator, and blaming the victim. Because most victims seek help from informal supporters first, predictive personality factors of supportive attitudes towards DV victims from informal supporters was the main focus of this study and a measure, the Support for the Victim scale, was developed for the current study based on a review of the literature and a consultation with a licensed psychologist. Using a sample of college students from Brigham Young University, an online survey was conducted to examine which factors are predictive of supportive attitudes towards DV victims.