VictorJuiz/thesis huggingface dataset:
Quantifying the adaptive power of the mobile integron

Mobile integrons are widespread and clinically relevant drivers of antibiotic resistance. Acting as genetic platforms, they allow bacteria to capture, express and re-order mobile antibiotic resistance gene cassettes, whose expression levels depend on their distance from the promoter, located at the integration site. Cassettes can be integrated and excised by the integrase enzyme, produced by the bacteria in time of stress. It has been hypothesized that integrons allow bacteria to adapt quickly to changing antibiotic pressures by reshuffling the cassette order, and therefore their resistance levels, such that integrons provide ‘adaptation on demand’.
To test this ‘adaptation on demand’ hypothesis, I used a custom three-cassette integron system transformed in both E. coli and P. aeruginosa. I first investigated the effect of cassette position on cassette expression at the phenotypic, transcriptional and translational level and confirmed a transcriptional origin of the cassette expression gradient, which was either strengthened or reduced by additional translational effects. I then combined experimental evolution and whole genome sequencing to quantify the evolvability benefits of cassette shuffling against increasing concentrations of antibiotics. By comparing strains with and without a functional integrase and focusing on two different cassettes and antibiotics, I observed striking differences in population survival rates and evolutionary pathways depending on the cassette and on the antibiotic. On one hand, with the antibiotic gentamicin and the aadB cassette, I saw an increase in evolvability and identified extensive cassettes rearrangements as well as cassette duplications in the populations with a functional integrase. On the other hand, with piperacillin and blaVEB-1, integrase activity was linked with decreased evolvability and a much lower rearrangement frequency, but I observed unexpected integron-mediated recombinations in the plasmid conjugation machinery.
These results highlight the key impact of antibiotics and cassette characteristics on the potential integrase shuffling benefits and provide novel insights on the dynamics controlling cassettes and integrase prevalence in the environment. In my experiments, high cassette mobility was necessary to compensate for the high fitness cost of integrase expression, or the integrase was a burden for bacteria survival. Moreover, I showed integrase activity was not limited to cassette shuffling or cassette acquisition but may also promote cassette duplication and beneficial off-target recombination, with far-reaching consequences on plasmid and antibiotic resistance evolution.

Keywords: experimental evolution
integron
antibiotic resistance
Advisor: MacLean, R
Foster, K
Structural determinants of Bunyavirus entry and neutralisation

The order Bunyavirales is comprised of numerous emerging zoonotic pathogens which present a global threat to human health. Many bunyaviruses are arboviruses including members of the family Nairovirdae such as the tick-borne Crimean-Congo haemorrhagic fever virus that is responsible for deadly outbreaks in Africa, East Asia and Southern Europe. Also among them are members of the family Hantaviridae which include Andes and Sin Nombre virus in the American continent and Hantaan and Puumala virus in Asia and Europe respectively. These are a hosted predominantly by rodents and frequently spill over into humans populations leading to hantavirus pulmonary syndrome and haemorrhagic fever with renal syndrome.
Bunyaviruses are often pleomorphic and are enveloped in a membrane harbouring just two surface glycoproteins, Gn and Gc. These glycoproteins are responsible for budding of the virus from the host cell, attachment to new target cells and release of viral RNA by membrane fusion. Furthermore, these glycoproteins are the target of the antibody-mediated host immune response and could form the basis for the development of future vaccines and immunotherapeutic medical interventions. The precise arrangement of these glycoproteins, and how these processes are mediated, are only partially understood for members of Hantaviridae and Nairoviridae. The goal of this thesis is to contribute to our understanding by addressing these topics using cryogenic electron microscopy (cryo-EM).
An initial description of the nairovirus surface is presented which displays a novel lattice of glycoproteins across its surface. An improvement to our understanding of the hantavirus glycoprotein array is made which enables, in combination with available crystallographic models, a near-complete pseudo-atomic model of the hantavirus surface to be proposed. In addition, the metastable nature of this lattice is demonstrated after exposure to low pH conditions equivalent of the endocytic pathway. A hantaviral virus-like particle (VLP) system is developed to provide a safe platform for structural studies of otherwise dangerous pathogenic species. This is then used to gain insights into hantavirus receptor interactions including Andes virus and the cell surface protein protocadherin-1. The VLP system is also used to study the structural mechanism of antibody-mediated host immune responses to Puumala virus and Hantaan virus. Finally, a number of tools and optimisations to assist and improve the cryo-ET and subtomogram averaging workflows that enable these structural studies are presented.

Keywords: cryo-EM
virology
virus
hantavirus
structural biology
electron microscopy
Advisor: Huiskonen, JT
Bowden, TA
Tailoring nanomaterials for energy applications

Silicon-based materials present high theoretical capacities (4200 mAh/g), making them among the most promising materials for Li-ion battery anodes. However, the major problem with them is massive volume change (∼ 300%) that occurs during cycling, resulting in pulverisation and significant capacity drop of these anode materials. Besides, the use of additives and metallic current collectors to make the anodes reduces the overall battery energy density as these components could not contribute any capacities to the battery. In this work, free-standing silicon-coated multi-wall carbon nanotube (MWCNT) anodes were developed without any polymeric binders and carbonaceous additives being added. These materials were achieved through exploiting atmospheric pressure chemical vapour deposition as well as advanced nanomaterials fabrication and processing methods. Electrodes were prepared utilising MWCNT papers (so-called buckypapers) as robust, electrically conductive flexible (22.2 S/cm) and free-standing scaffolds since the capacity of these buckypapers is too low to use as a new active material for the battery (0.3 mAh/cm2 in the 300th cycle). Hence, these buckypapers were functionalised with silicon nanoparticles attached through a strong silicon carbide interface and coated with a protective amorphous carbon layer. The SiC interface was partially developed between Si NPs and MWCNTs as excessive SiC could reduce electrochemical capability on the material due to its low electroactivity. The coin cell battery test demonstrated a high areal capacity of up to 2.01 mAh/cm2. These free-standing Si-CNT-SiC-C materials showed good rate capability and stability performance (0.55 mAh/cm2 after 300 cycles with coulombic efficiency of 99.7%). As a result, the material interface design, which is the key of this work, enhances the connection of Si NPs to the MWCNT walls and addresses the issue of Si detachment. A post-mortem result also revealed the structural stabilisation of the electrode after the repeated cycling test. In addition, the pouch-cell battery was fabricated to confirm the use of this free-standing Si-CNT-SiC-C electrode without the influence of the battery component. This new anode architecture performs better than the buckypaper, thereby increasing the areal capacity by 83.3%, allowing better functionality for this material to be implemented in a full-cell battery system.

Keywords: silicon
MWCNTs
carbon nanotube
Li-ion battery
anode
Advisor: Grobert, N
Pursuing the universal: formalism, pragmatism and the project of modernity in contemporary critical democratic theory

This thesis critically evaluates the role of form and formalism in contemporary Frankfurt School critical democratic theory’s treatment of ethical pluralism. It examines how three contemporary theorists, Jurgen Habermas, Rainer Forst and Alessandro Ferrara, deploy methodological formalism, and a certain ideal of aesthetic or theoretical ‘form’, in support of the Kantian ‘project of modernity’. The thesis has two related aims: the first is to diagnose and critically evaluate the role of form and formalism as an instrument of mediation between ethical identity, value pluralism and the ‘project of modernity.’ In the course of identifying and assessing the intellectual resources that underpin the role of form and formalism in the relevant critical theories of democracy, this thesis argues that the strategies for normative and constitutional integration pursued by Habermas, Forst and Ferrara suffer from a hermeneutical deficit. Consequently, while they set out to develop an ethically non-impositional framework for negotiating between normative universality and ethical particularity, these three theorists also continue several of the weaknesses of Kantianism, including its dichotomous separation of normative ‘form’ from the texture of ethical life and its resident meanings. While Habermas, Forst and Ferrara all seek creatively to integrate the insights of philosophical pragmatism by developing immanently grounded ‘inter-subjective’ theories of normativity, their deployment of form as an ethically diminutive binding agent displays a limited reflexivity regarding the ideological specificity of aesthetic and methodological formalism.
The second aim, in light of these criticisms, is to suggest an alternative method by which to theorise a deliberatively developed normative commonality amidst ethical pluralism - one that draws upon the philosophical pragmatism of John Dewey, Hans Joas, James Bohman and others. I argue, in conversation with these thinkers of a pragmatist bent, and with reference to recent contributions to the sociology of religion, for an account of value generalisation as a form of ‘world-building’ deliberative action oriented towards what Hannah Arendt calls ‘a common world.’ In so doing, I question the role of the ‘comprehensive social theory’ of normative integration amidst value pluralism, and instead look to theoretically enunciate and develop deliberative practices that plausibly effect ‘practical transformations’ of existing social challenges.

Keywords: Frankfurt School
democratic theory
Habermas
pragmatism
Arendt
modernity
Advisor: McNay, L
Study of electrode kinetics

This thesis reports the use of Tafel analysis in the study of electrode kinetics from both theoretical and experimental perspectives.
During electrochemical measurements, any changes in temperature cause changes in diffusion coefficient of the species, the electrochemical rate constant and the equilibrium potential. Concequently, the improtance of temperature control in electrochemical systems is first investigated. When and how thermally induced convective flows in bulk solution influence the votlammetric behaviour are presented in Chapter 3.
Chapters 4 and 5 theoretically discuss what fraction of a voltammetric wave is appropriate to use as the Tafel region for accurate analysis under linear, quasi-steady-state and steady-state mass-transport regimes for an irreversible one-electron transfer process. The measured transfer coefficient is found to deviate significantly from its true value as a function of potential due to the mass-transport limitation at high overpotentials. If and how a simple analytical mass-transport correction using a plot of ln |1I-1Ilim| against potential can be used to improve the measurement of transfer coefficient is investigated. The methodology of measuring transfer coefficient is further employed in the electrochemical characterisation of a single microband electrode with unknown dimensions (Chapter 6). Such Tafel analysis is applied to an experimental study where the intrinsic surface quinones on carbon substrates can catalyse Fe2+/3+ redox reaction evidenced by a potential dependent transfer coefficient (Chapter 7).
Last but not least, a new simulation techinique is developed in Chapter 8 to extract the kinetic information from experimental voltammograms for electrodes under both radial and liner regimes on the basis of the prior knowledge of the physical parameters defining the system, most importantly the diffusion coefficient, analyte concentration and electrode radius.

Keywords: Surface quinone
Thermal convection
Mass transport corrected Tafel analysis
Electrode kinetics
Mass transport
Band electrode
Microelectrode
Electrocatalysis
Tafel analysis
Advisor: Compton, RG
Investigation of type 2 cytokine pathways as modulators of haematopoiesis during chronic inflammation

Chronic inflammatory diseases often promote bone marrow (BM) myelopoiesis at the expense of erythropoiesis and lymphopoiesis, facilitating production of mature myeloid cells to sustain the inflammatory response. In previous studies, type 2 cytokines, including IL-4, IL-5, and IL-33, have been suggested as possible regulators of haematopoiesis, but their role in modulation of haematopoiesis during inflammation was unknown. Spondyloarthritis (SpA) is a chronic inflammatory disease of people that has systemic sequelae, including anaemia. Although associated with type 17 immune responses, there is evidence of a concurrent type 2 response in the intestine of people with SpA. In this thesis, I sought to bring these areas of investigation together by asking if a possible type 2 immune module in murine SpA might be implicated in changes in haematopoiesis, and whether this would be important for progression of disease.
I find inflammation perturbs haematopoiesis in murine SpA, causing a marked bias towards myelopoiesis in the BM, as well as accumulation of myeloid progenitors in spleen and inflamed joints. Conversely, BM erythropoiesis is suppressed, resulting in anaemia. I find that mice also have evidence of a type 2 immune module in the intestine, with increased numbers of cells producing IL-4 and IL-5, and increased production of IL-33. Of these cytokines, IL-4 has pro-myelopoietic effects in vivo and ex vivo, but my results suggest this pathway is not important for development of clinical disease or expansion of myelopoiesis in SpA. Conversely, I find that IL-33 specifically suppresses differentiation of erythroid progenitors. This effect is implicated in development of anaemia in SpA, with IL-33 also causing resistance to the effects of erythropoietin (EPO). Interleukin-33 also promotes myelopoiesis through a combination of direct and indirect effects on progenitor stages, indicating that IL-33 has co-ordinated effects in different parts of the haematopoietic system that may be consistent with its role as an alarmin.
Collectively, my work reveals IL-4 and IL-33 as regulators of myelopoiesis, which may be important in disease settings associated with increased production of these cytokines. I further identify IL-33 as a mediator of anaemia of inflammatory disease and EPO resistance in murine SpA, suggesting that IL-33 could be a therapeutic target for this prevalent and debilitating comorbidity in people with chronic inflammation.

Keywords: Anaemia
Haematopoiesis
Spondyloarthritis
Erythropoiesis
Immunology
Inflammation
Haematology
Advisor: Griseri, T
Powrie, F
Understanding why quality circles work in primary health care: a realist approach about how, why and under what circumstances they work

Quality circles and similarly structured small groups are commonly used in primary health care in Europe to reflect on and improve standard practice over time. They are a complex social intervention in the fast-changing system of primary health care. Many controlled trials and reviews have documented their positive effects on behaviour change. As effect sizes vary, I sought to understand how and why quality circles achieved their results by using a realist approach.
I conducted a co-inquiry with stakeholders to determine their interests and preferences concerning the direction of my research, and their mental model of quality circles, which I used as a framework for my realist review. After identifying relevant papers and selecting and extracting appropriate data, I labelled and analysed configurations of context and mechanism that shaped the outcomes of quality circle work. From these, I built propositions to develop a programme theory underlying quality circles, then refined and consolidated this in sequential interviews with participants, facilitators and managers of quality circles.
The programme theory suggests that government trust in GPs’ ability to achieve quality improvement, with appropriate professional and administrative support, will motivate them to participate in quality circle work. Professional support includes training in relevant techniques, access to educational materials and personal performance data. Administrative support includes protected time, an appropriate venue and financial resources for meetings. Group dynamics are at the core of the process. Facilitators help participants to build social bonds and mutual trust so that the group becomes a safe environment where participants link insights to everyday practice to develop their professional role. They reflect on their personal experiences, add information from evidence-based or personal performance data and thus create new concepts for improving their practice. With repetition, they gain confidence in their quality improvement skills and can put their innovations into practice.

Keywords: Quality in Health Care
Quality Circle
Primary Health Care
Continuous Professional Development
Quality Improvement
Continuous Medical Education
Knowledge Translation
Peer Review Group
Advisor: Wong, G
Harris, J
Mickan, S
Investigation of the nascent human transcriptome

High-throughput sequencing of transcripts through RNA-Seq has revolutionised the way gene expression can be understood. However, RNA-Seq only measures the steady-state levels of stably-expressed transcripts rather than the act of transcription. To measure nascent transcription, a range of sequencing methods have been developed, all with a range of different benefits and disadvantages. This study aims to advance the already established transient transcriptome sequencing (TT-Seq) approach and develop two novel nascent transcriptomic methods in human cells that utilise metabolic labelling and deliver high resolution profiles, called SNU-Seq (single nucleotide resolution 4sU-Seq), and size-selected SNU-Seq. The former is particularly well-suited to measure elongation of transcription throughout the gene body whereas the latter is used for studying transcription at the very 5’ end of genes. This study therefore includes a comprehensive analysis of promoter-proximal pausing behaviour and the development of a new size-selected SNU-Seq-based pipeline for annotations of actively-transcribed transcription start sites.

Advisor: Mellor, E
Time-lapse super-resolution imaging reveals key principles for cis-limited Xist RNA localisation

Dosage compensation in XX female mammals is achieved by X chromosome inactivation (XCI). The key player in this process is the long noncoding RNA Xist (X inactive specific transcript), which associates in cis with the inactive X chromosome elect (Xi) to mediate gene silencing and heterochromatization. Xist spreads across the entire Xi, but little is known about the dynamics of this process, particularly in the context of single cells. This thesis documents the development of a new, time-resolved super-resolution imaging methodology: RNA-SPLIT (Sequential Pulse Localisation Imaging over Time) combining Bgl-stem-loop labelling, HaloTag technology and 3D structured illumination microscopy (3D-SIM) to detect individual Xist RNA molecules in single mouse embryonic stem cells (mESC) with high spatial and temporal resolution.
XY and XX mESCs expressing inducible autosomal transgenic or endogenous Bgl-stem-loop tagged Xist in tandem with BglG-Halo fusion protein were used to visualise distinct, functional Xist foci and analyse their spreading behaviour across the Xi during early XCI. RNA-SPLIT yielded important insights into Xist RNA kinetics, localisation and spreading behaviour during XCI and a feedback mechanism linking Xist RNA synthesis and degradation. RNA-SPLIT also uncovered novel coupling behaviour of Xist RNA molecules which, using perturbation experiments with CIZ-1 and SPEN, was shown to be independent of Xist localisation to the Xi, but dependent upon SPEN. In depth analysis demonstrated SPEN to have separable, SPOC domain independent functions in Xist RNA localisation, stability and coupling behaviour.

Keywords: SPEN
3D-SIM
X chromosome inactivation
Xist
Super-resolution microscopy
Ciz-1
Advisor: Coker, H
Schermelleh, L
Brockdorff, N
Optimal transport and 1-Lipschitz maps

The main topic of this work is concerned with optimal transport approach to the localisation technique. The localisation technique allows to reduce a high-dimensional problem to a collection of one-dimensional problems. Significance of this approach is illustrated by its applications to Poincaré inequality, log-Sobolev inequality, isoperimetric inequality in the context of metric measure spaces satisfying the curvature-dimension condition. We investigate multi-dimensional generalisation of the technique and related conjectures of Klartag. The settlement of these conjectures would constitute a step towards a proof of the waist inequality in the setting of metric measure spaces satisfying the curvature-dimension condition. This, in turn, would help in answering the Bourgain’s hyperplane conjecture and the isoperimetric conjecture of Kannan, Lovász and Simonovits.
We provide a partial affirmative answer to the conjecture that, whenever Euclidean space equipped with a measure satisfies the curvature-dimension condition, then any vector-valued 1-Lipschitz map induces a partition of the domain such that the leaves equipped with the conditional measures satisfy the curvature-dimension condition with the same parameters as the initial space did. Our approach is based on the ideas of Sudakov and their further instances in works of Caffarelli, Feldman, McCann and of Klartag.
We provide a counterexample to another conjecture of Klartag that the conditional measures, with respect to a partition induced by a certain 1-Lipschitz map, satisfy so-called mass balance condition. We develop a theory of optimal transport of vector measures and employ it to provide a sufficient condition for the mass balance condition to hold true.
One way of proving the mass balance condition in one-dimensional setting is via approximation of Lipshitz functions. This leads us to a study of continuity properties of extensions of vector-valued Lipschitz maps. We provide a strengthening of the classical Kirszbraun theorem, which shows a discrepancy between one-dimensional and multi-dimensional cases. We identify a sharp rate of continuity of extensions of 1-Lipschitz maps.
Recently, several variants of the optimal transport problem have been studied. Martingale optimal transport is a one of the greatest importance. Interestingly, also in this case a localisation result has been established. Therefore, with help of a novel version of Strassen’s theorem, we investigate the duality theorems in this problem. We provide a reformulation of the optimal transport in two-marginal, in multi-marginal and in martingale setting. We reprove the Kantorovich duality formulae. As an application we characterise the uniformly smooth and the uniformly convex functions.
We develop a divergence formulation of optimal transport of vector measures and use it to provide a novel proof of the representation formula for polar cone to monotone maps. We find several generalisations of this result. A tool that we use is the matrix Hölder’s inequality, for which we characterise the equality cases.

Advisor: Qian, Z