Cost-effective reduction in the uncertainty surrounding global indicators of biodiversity change is a central goal of conservation. In this thesis, I identify and resolve the effects of IUCN Data Deficient species on the estimation of global patterns and levels of extinction risk. I show that gaps in our knowledge of species' conservation status are primarily driven by spatial patterns of ecological research (Chapter 2). Large numbers of species are extremely poorly known, highlighting the importance of basic taxonomic and natural history information in conservation assessments. Using sensitivity analyses (Chapter 3), I show that Data Deficient species contribute to considerable uncertainty in patterns of extinction risk in freshwater invertebrates, limiting our understanding of the factors influencing extinction risks and our capacity to design reliable conservation schemes. To determine the likely conservation status of Data Deficient species, I develop seven machine learning models based on species' life-history traits, niche and threat exposure (Chapter 4). I find that machine learning models accurately predict species conservation status and geographical patterns of threatened species richness. I predict 64% of Data Deficient mammals to be at risk of extinction, increasing the estimated proportion of threatened mammals from 22% to 27% globally. Finally, I use sampling theory to compare the cost-effectiveness of predictive models and IUCN Red List assessments in mammals, amphibians, reptiles and crayfish (Chapter 5). Double sampling with predictive models reduces the cost of determining the proportion of Data Deficient species at risk of extinction by up to 69%, and can be used to reduce the impact of uncertainty in the Red List and Red List Index. My thesis demonstrates how predictive models and decision theory can strengthen indicators of biodiversity change to monitor progress towards international biodiversity targets.

The behaviour of a fishing fleet is a critical, but all too often overlooked, uncertainty in the implementation of fisheries management. Unexpected responses by fishers to management controls, such as effort restrictions or spatial closures, can result in unintended and potentially undesirable outcomes. Whilst this uncertainty can be reduced by anticipating the behavioural response of a fishing fleet, it is first necessary to understand the characteristics and drivers of fleet behaviour. The aim of this thesis was to address gaps in knowledge of the behaviour of offshore tuna fleets, using the Indian Ocean tropical tuna purse seine fishery as a case study example. I used statistical modelling to examine the factors that influence the spatial behaviour of the purse seine fleet at broad spatiotemporal scales. This analysis revealed consistency in the use of seasonal fishing grounds by the fleet, as well as a forcing influence of biophysical ocean conditions on the allocation of effort. These findings, which suggested strong inertia in fleet spatial behaviour, have important implications for predicting the response of the fleet to certain natural events or management measures (e.g. spatial closures). To better understand the impact of spatial closures on purse seine fleet dynamics, I used the statistical model of fleet behaviour to isolate the policy effect of two recent closures on fleet behaviour. By comparing the observed behaviour of the fleet against a model-generated counterfactual scenario I revealed, in the case of one of the closures, a policy effect that was inconsistent between years, and that the absence of fishing effort in the closed area was explained primarily by biophysical ocean conditions. These findings demonstrate the importance of using a counterfactual approach to evaluate spatial closures in open ocean systems where fleet behaviour is influenced by highly variable biophysical conditions. Fish aggregating devices (FADs) have become a dominant fishing practice in tuna purse seine fisheries worldwide, and I examined the influence of the use of FADs on purse seine fleet dynamics in the Indian Ocean. I reviewed historical catch trends and spatiotemporal patterns of fleet behaviour and linked this to the use of FADs. I also reviewed the existing management of FAD-fishing and speculated at the influence of possible future management measures on the behaviour of the fleet. Finally, I used a scenario planning approach to think about how the main drivers of purse seine fleet behaviour might change in the future, and how this might affect fleet dynamics. This analysis served to highlight aspects of purse seine fleet behaviour that should be a priority consideration of tuna fishery managers and policy makers. This thesis showed fleet behaviour to be a dynamic aspect of tuna fisheries management, and stressed the importance of anticipating the response of fleets to management measures in order to avoid unintentional outcomes. The understanding of purse seine fleet behaviour developed throughout this thesis provides a good basis for building the anticipation of fleet behaviour into existing management tools and processes.

NaV1.8 is a voltage gated sodium channel mainly expressed on the membrane of thin diameter c-fibre neurons involved in the transmission of pain signals. In these neurons NaV1.8 is essential for the propagation of action potentials. NaV1.8 is located in lipid rafts along the axons of sensory neurons and disruption of these lipid rafts leads to NaV1.8 dependant conduction failure. Using computational modelling, I show that the clustering of NaV1.8 channels in lipid rafts along the axon of thin diameter neurons is energetically advantageous and requires fewer channels to conduct action potentials. During an action potential NaV1.8 currents across the membrane in these thin axons are large enough to dramatically change the sodium ion concentration gradient and thereby void the assumptions upon which the cable equation is based. Using scanning electron microscopy NaV1.8 is seen to be clustered, as are lipid raft marker proteins, on neurites at scales below 200nm. FRET signals show that the lipid raft marker protein Flotillin is densely packed on the membrane however disruption of rafts does not reduce the FRET signal from dense protein packing. Using mass spectrometry I investigated the population of proteins found in the lipid rafts of sensory neurons. I found that the membrane pump NaK-ATPase, which restores the ion concentrations across the membrane, is also contained in lipid rafts. NaK-ATPase may help to offset concentration changes due to NaV1.8 currents enabling the repeated firing of c-fibres, which is associated with spontaneous pain in chronic pain disorders.

Isoprene is the most important biogenic organic volatile compound emitted by terrestrial vegetation into the atmosphere, in term of amount and effects on atmospheric chemistry. Primary environmental drivers of isoprene production are photosynthetic photon flux density (PPFD), leaf temperature (T) and internal CO2 concentration (Ci). Robust process-based modelling approaches are needed to assess how future changes in these environmental drivers may affect isoprene emissions and consequently atmospheric chemistry, air quality and (indirectly) the radiative forcing of climate. I present an original, conceptually simple model for isoprene emission by plants based on the hypothesis that the electron flux available for isoprene biosynthesis depends on the balance between the supply of reducing power generated by the light reactions of photosynthesis and the demand for reducing power in carbon fixation and photorespiration. I explain the physiological reasoning that led me to propose this. Using various leaf-scale measurements of carbon assimilation and isoprene emission, including a laboratory study I conducted on black poplar, I show that the model can reproduce well the variations of isoprene emission with PPFD, temperature, and Ci. The model also reproduces the tendency for the fraction of carbon re-emitted as isoprene to increase with increasing PPFD, and for the quantum efficiency of isoprene emission to decrease with increasing CO2 concentration. The model is shown to systematically outperform models that are in common use today. I also analysed the PPFD and temperature responses of carbon assimilation and isoprene emission as measured above the forest canopy. The model was upscaled and shown to reproduce key responses shown in two long-term flux monitoring datasets from temperate mixed forests. I discuss future research needs and the potential for this model to be further scaled up for global analyses.

Predatory mites are important components in biological control of arthropod pests in protected crops. The whitefly and thrips predator Amblyseius swirskii (Acari: Phytoseiidae) is an efficient biocontrol agent used widely for pest control in protected vegetable and ornamental crops. Amblyseius swirskii can be mass-reared on at least three species of astigmatid mites but little is known about their predator-prey interactions and population dynamics. These factitious prey allow for large-scale efficient rearing systems and novel crop inoculation methods. The use of breeding sachets offer a predatory mite delivery method with prolonged and sustained crop inoculation. This study endeavoured to assess the suitability of the factitious prey Suidasia medanensis (Acari: Suidasidae) for mass-rearing and field deployment of A. swirskii by studying the life table parameters of the predator on a diet of the said prey. The underlying predator-prey interactions were examined through a series of laboratory experiments focusing on the response of A. swirskii to prey density, preference of prey life stage, capture success ratio and the defence volatiles of adult S. medanensis against predators. Furthermore, in order to understand the behaviour and performance of a breeding sachet the internal population dynamics were studied in relation to release rates from the sachet. These studies were extended to examine the effect of different simulated crop conditions on predator release, focusing on temperature and relative humidity at constant and alternating controlled conditions. In addition to different crop conditions, A. swirskii may be exposed to various other crop protection products in the field as part of an IPM programme. The compatibility of A. swirskii with one such product, the fungal entomopathogen Beauveria bassiana (Balsamo), for concomitant use in the field was investigated through pathogenicity studies and sublethal effects under controlled conditions. Suidasia medanensis was found to be of good nutritional value to A. swirskii resulting in population growth rates similar to target pests, as reported in literature. Specific predator-prey interactions were identified, such as Type II functional response, preference to egg stages of the prey and the defence volatile of S. medanensis, the significance of which are discussed in depth in the thesis. Underlying dispersal strategies and the association between breeding sachet productivity and predator output was established. Furthermore, climatic conditions were found to have significant effects on sachet performance with clear indications of what constitutes favourable and unfavourable conditions. Amblyseius swirskii was found to be a physiological host to B. bassiana. Due to low-to-moderate mortality rates under ideal laboratory conditions, little effect on juvenile mites and no effect on offspring of treated mites these two biocontrol agents were concluded to have good potential for concomitant use, but with further trials required.

As multiple myeloma (MM) progresses, Natural Killer (NK) cell responses decline against malignant plasma cells. The immunomodulatory drug lenalidomide is widely used for the treatment of MM but its influence on NK cell biology is unclear. Here, a combination of functional assays and microscopy techniques were used to investigate how lenalidomide affects NK cell activation and effector function. First, lenalidomide lowered the threshold for NK cell activation, causing a 66% decrease in the EC50 for activation through CD16, and a 38% decrease in the EC50 for NKG2D-mediated activation, allowing NK cells to respond to lower doses of ligand. In addition, lenalidomide augmented NK cell responses, causing a 2-fold increase in the proportion of primary NK cells producing IFN-γ, and a 20-fold increase in the amount of IFN-γ produced per cell. Importantly, lenalidomide did not trigger IFN-γ production in unstimulated NK cells. Thus, lenalidomide enhances the NK cell arm of the immune response, without activating NK cells inappropriately. Of particular clinical importance, lenalidomide also allowed NK cells to be activated by lower doses of rituximab, an anti-CD20 mAb widely used to treat B cell malignancies. This supports the combined use of lenalidomide and rituximab in a clinical setting. Second, super resolution STED microscopy revealed that lenalidomide increased the periodicity of cortical actin at immune synapses, resulting in an increase in the area of the actin mesh predicted to be penetrable to vesicles containing IFN-γ. Finally, lenalidomide augmented IFN-γ production and enhanced cortical actin rearrangements in NK cells from MM patients. Interestingly, NK cells from relapsing MM patients showed defective F-actin remodelling compared to NK cells from MM patients in remission. This could be rescued with lenalidomide treatment. This establishes that nanometre-scale rearrangements in cortical actin, a recently discovered step in immune synapse assembly, are a potential new target for therapeutic compounds.

The anciently asexual bdelloid rotifers are a ubiquitous and ecologically important group coined 'evolutionary scandals' owing to their diversification and persistence over evolutionary time despite the pressures of obligate parthenogenesis. Understanding the biodiversity of rotifers, in the context of reproductive mode, will aid in the understanding of how differences in sex and ecology drive biodiversity patterns. Given that resolved taxonomy is a prerequisite to investigating larger scale macroevolutionary patterns, and that rotifer taxonomy is confounded by morphostasis, cryptic diversity, and a dearth of expert taxonomists, this thesis first deals with the use of DNA taxonomy as a tool to alleviate the taxonomic crisis so prevalent in the group. Results from multiple meta-analyses exploring the effects of choice in gene, phylogenetic reconstruction methods, and species delimitation metrics, provide better guidelines for DNA taxonomy. The analyses suggest that coalescent-based DNA taxonomy using the Generalised Mixed Yule Coalescent model in conjunction with the cytochrome oxidase 1 subunit c gene analysed in a Bayesian inference phylogenetic framework using BEAST, provides realistic and tangible species clusters (evolutionarily significant units; ESU) in both asexual and sexual organisms. Using these ESUs, the efficacy of DNA barcoding for identifying rotifer species is examined. These analyses suggest that the sexual monogonont rotifers are more readily identifiable than the asexual bdelloid rotifers. Explicit comparison of genetic discreteness of asexual and sexual rotifers indicates that sexual rotifers are separated by larger genetic and phylogenetic gaps than asexual rotifers and thus are more discrete. Combined, these analyses indicate that sex, specifically reproductive isolation, is the predominant factor in explaining why species form discrete clusters rather than a continuum of forms. Finally, a dated multilocus molecular phylogeny of Bdelloidea is reconstructed. The phylogeny suggests that bdelloid rotifers have persisted for at least 40 million years, but that bdelloid higher taxa are typically polyphyletic.

The dimorphic fungus Candida albicans, a harmless commensal in approximately 60% of healthy adults, is also an opportunistic pathogen in humans. Immuno-compromised individuals are mainly at risk, for example those under immunosuppressive therapies, HIV/AIDS patients, the elderly and early neonates. Diseases caused include invasive candidiasis, which has a 40-60% mortality rate. The majority of invasive candidiasis is caused by C. albicans forming biofilms on medical devices (e.g. catheters, heart valves) from which yeast cells disperse and enter the blood stream. The adherence properties of C. albicans are crucial for colonisation and biofilm formation. An important adhesive factor in the early stages of colonisation is Als1 (Agglutinin-like Sequence protein 1). Another crucial virulence factor is hyphal-expressed Als3, which contributes to adhesion and invasion of host cells. These proteins belong to the Als family of glycoproteins (Als1-7 and Als9), all of which share similar domain organisation: an N-terminal adhesion region comprised of two immunoglobulin-like domains, plus central and C-terminal regions, which are highly glycosylated. Previously, the N-terminus of Als9 had been shown to bind the flexible C-termini of host cell proteins as ligands. Due to their relevance in C. albicans adhesion, the N-terminal domains of Als1 and Als3 are ideal templates for ligand binding studies and the development of antifungal compounds. Shortened constructs Als1 and Als3 with improved solubility and structural properties (relative to the full length adhesins) were employed in this project. Here I describe the X-ray crystallographic structure of sNT-Als1 with a peptide ligand, the backbone resonance assignments by Nuclear Magnetic Resonance (NMR) of the ligand-free protein and ligand-binding experiments using Differential Scanning Fluorimetry (DSF). Comparison of the structure and ligand binding behavior of sNT-Als1 with other Als proteins will guide the design of therapeutic molecules that block the adhesive properties of these adhesins, and thus the formation of disease-causing biofilms by C. albicans.

Geobacillus thermoglucosidasius has been identified as an organism capable of producing bioethanol from lignocellulosic biomass based on its ability to ferment both hexose and pentose sugars. Engineering of the wild-type strain DL33 (wt) has produced a single knock out strain DL44 (Δldh) and a double knock out strain DL66 (ΔldhΔpfl↑pdh), both of which have increased capacity for bioethanol production. The nutritional requirements of the strains under anaerobic conditions are yet to be fully understood. In this study, a systems approach to understanding the metabolism of the wild-type and engineered strains has been taken in order to further understand the changes in metabolism resulting from the mutations introduced. For the first time 13C-metabolic flux analysis has been applied to the comparative study of the wild-type and engineered strains using global isotopomer balancing. This has revealed flux through the anaplerotic reactions has reversed from being in the direction of pyruvate/phosphoenolpyruvate in the wild-type, to being in the direction of oxaloacetate/malate in the engineered strains. Alterations in TCA cycle flux between the strains were also seen. Furthermore alanine was found to be produced as a fermentation product in each strain. Analysis of the genome sequence has revealed an unusual oxidative branch of the pentose phosphate pathway, missing 6-phosphogluconolactonase but with genes encoding the rest of the pathway still present, suggesting that flux through this pathway may still proceed, dependent on the themolability of glucono-1,5-lactone-6-phosphate. It has been found that RNA extracted from G. thermoglucosidasius is prone to rapid degradation which may affect the outcome of analysis of the transcriptome by RNA-seq. Nonetheless, it has been possible to apply RNA-seq to the wild-type organism grown aerobically and use this to identify transcripts for the major pathways of central carbon metabolism and the most highly expressed transcripts of the culture.

Mutations in the human BRCA2 gene are a leading cause of susceptibility to breast, ovarian and prostate cancers. Its protein equivalent BRCA2, is the key mediator in repair of double-stranded DNA breaks and interstrand crosslinks - the most dangerous forms of DNA damage, via RAD51 effected homologous recombination. Thereby, preferably the sister chromatid, or homologous chromosome is utilised to drive the repair. Described here is the first structural and related mechanistic characterisation of full-length (384 kDa) BRCA2, and its complex with RAD51 and DNA. Electron microscopy reconstruction reveals that BRCA2 exists as a dimer in head-to-tail conformation, which binds two oppositely directed sets of RAD51 molecules. Single-stranded DNA (derived from exonucleolytic processing of double-strand breaks) binds BRCA2 along its long axis, such that one set of RAD51 monomers always binds thereon regardless of its binding polarity. These then self-assemble into RAD51-ssDNA nucleoprotein filaments that catalyse the main reaction, having been nucleated and primed for growth by BRCA2. In complementary work, it is shown that BRCA2 increases the frequency of RAD51-ssDNA filament formation, but does not impact filament length itself. It is also shown that BRCA2 initiates monodirectional (3'→5') extension of the RAD51 filament on single-stranded DNA from nucleation sites (thence at the 3' end). Furthermore, BRCA2 initiates such nucleations at multiple sites along individual ssDNA molecules, suggesting that these extend and merge into a single nucleoprotein filament, which would be most capable of catalysing the subsequent homologous pairing and strand exchange reactions. Together, the structures and biochemical data define the molecular mechanism via which BRCA2 orchestrates the formation of RAD51 nucleoprotein filaments, which constitute the central reaction intermediate in the repair of DNA double-strand breaks.