The impacts of global change on ecosystems from climate change and invasive species are likely to be complex. Rising atmospheric CO2 concentrations, the associated climate forcing and greater frequency of extreme weather are serious challenges to natural ecosystems. In tandem with climate change, globalisation has led to the spread of invasive alien species around the globe that threaten to interrupt food web dynamics. Advancing understanding of the effects of global change on trophic interactions therefore requires study of interspecific and multi-trophic interactions. The aim of this thesis was to examine how host-plant heterogeneity, native–invasive species interactions and climate change effects (elevated atmospheric CO2 (eCO2) or drought) influence trophic interactions. An experimental approach was used which centred on a study system comprising the European raspberry (Rubus idaeus), the herbivorous large raspberry aphid (Amphorophora idaei) and coccinellid beetle predators (native species: Adalia bipunctata, Coccinella septempunctata; invasive alien species Harmonia axyridis). Under eCO2, R. idaeus resistance to A. idaei was unchanged for two cultivars (Glen Clova, Glen Ample) partially susceptible to A. idaei, but significantly reduced for another (Octavia) with complete resistance in ambient climatic conditions. The inclusion of a coccinellid predator, however, mitigated the reduction in the resistance of Octavia by reducing aphid abundance. Behavioural responses to predation by A. idaei were also impaired under eCO2 after feeding on Glen Ample. The role of natural enemies in controlling herbivore abundance in future climates is therefore crucial. Native coccinellid species are currently declining in much of Europe, attributed to the occurrence of the invasive species, H. axyridis. Despite the declines in native coccinellid species, it was found that behavioural modification to feeding by both native and invasive coccinellid species can, theoretically, result in coexistence. Plant resistance in a future climate is likely to be modified significantly. Reduced resistance to aphid herbivory demonstrated here mirrors previous studies, highlighting the future importance of natural enemies to control aphid abundance. Changes to the abundance and behaviour of aphid prey and intraguild predators will modify the effectiveness of native and invasive natural enemies. Further mechanistic research is required to understand multi-trophic interactions in dynamic environments.

Kinetochores are multi-protein machines that control chromosome movements by regulating the dynamics of attached microtubules. In human cells chromosome movements are orchestrated by the leading kinetochore tracking a shrinking microtubule whilst its sister tracks a growing microtubule. Directional switches occur when (both) kinetochore-attached microtubules fl ip between these two states, adaptive and coordinated switching then giving rise to the oscillations observed during metaphase. However the mechanisms (and rules) controlling directional switching are poorly understood. This work demonstrates that by tracking kinetochores with sub-pixel resolution in HeLa cells and fitting stochastic mathematical models that a sensor on the leading sister triggers switching when the tension across the centromeric spring connecting the sisters builds up sufficiently rapidly. Further it is shown that the trailing sisters polymerisation state is stabilised by high spring tension. These mechanisms pre-empt trail-first switching that would otherwise impose abnormal pulling forces between sister chromatids. As a consequence sister-switching is biased towards lead-first switching, switching of the trailing sister rapidly following as the spring tension falls, this removing the force dependent stabilisation of the trailing sisters K-fibre (kinetochore bound microtubules). This model explains how switching events are initiated and resolved, the centromeric spring tension providing a means for inter-sister communication and cross regulation that results in coordinated oscillations within a context of low spring tension. This study demonstrates that high throughput analysis and modelling pipelines can provide novel mechanistic insight into mechanochemical systems.

G protein-coupled receptors (GPCRs) are a superfamily of membrane proteins that bind to a diverse array of stimuli and are involved in a large number of physiological functions. The family A GPCRs are the largest and most comprehensively studied. The family B GPCRs are a small but important group of receptors (~15 members) that bind to peptide ligands and are involved in physiological processes that include vasodilation, stress, digestion and glucose homeostasis. The CGRP receptor is a unique member of this family as it is a heterodimer consisting of a GPCR subunit (calcitonin receptor-like receptor, CLR) and a single transmembrane accessory protein (receptor activity modifying protein, RAMP1). The extracellular loop two (ECL2) domain is involved in ligand binding and activation in a number of studied GPCRs. This makes it vital both with respect to receptor function and in the design of therapeutics. The main focus of this thesis is to study the structure and function of the ECL2 domain in the CGRP receptor. This was initially done through individual alanine substitutions of each ECL2 residue and measuring the effect of this on a number of receptor processes. Residues that were identified as important for receptor function through this investigation were selected for an extensive set of mutagenesis to identify the precise molecular interactions that were involved at each position. These experiments have shown that ECL2 is the most important domain of the CGRP receptor for ligand-based activation. The N-terminal half of ECL2 contains residues predicted to have structural function and the C-terminal half is predicted to be involved in direct ligand binding. These results have been used in collaboration to refine a computer model of receptor structure and ligand binding to predict specific ligand docking sites that can be used to design therapeutics for migraine, heart attack and hypertension. The final part of thesis produced preliminary data to support proof of concept for two techniques that can be used in the study of CGRP receptor function.

In last decade, there has been a tremendous progress in scanning probe microscopies, some of which have achieved atomic resolution. However, there still exist some problems which have to be solved before the instrument can be used as a metrological measurement tool. The object of the project introduced in this thesis was to develop a scanning force microscope of metrological capability with the aim of making significant improvement in scanning force microscopy from the viewpoint of instrumentation. A capacitance based force probe has been studied theoretically and experimentally with the main concern being its dynamic properties, characterized by squeeze air film damping, which are believed to have direct effects on the fidelity of measurement. The optimization of design is investigated so as to achieve the results of both high displacement sensitivity and force sensitivity. An x-y scanning stage has been designed and built, which consists of a two axis linear flexure system of motion amplifying mode machined from a single aluminium alloy block. The stage is driven by two piezo actuators with two capacitance sensors monitoring the actual position of the platform to form a closed loop control system. The design strategy is introduced and the performances and characteristics of two commonly used types of flexure translation mechanisms, leaf spring and notch hinge spring system, are analyzed. The finite element analysis method is employed in the analysis and design of translation mechanism. Finally, a metrological scanning force microscope has been constructed, combining a constant force probe system, an x-y scanning stage and a 3D coarse positioning mechanism into a metrological system. The performance of the instrument system has been systematically evaluated and its measuring capability investigated on the. specimens of various properties and features. The results from this first prototype of the instrument demonstrated a subnanometer resolution with comparable stability and repeatability in all three axes.

Methanotrophic bacteria convert methane to methanol using a methane monooxygenase enzyme (MMO). Two types of MMO exist: a membrane bound enzyme (PMMO) and a cytoplasmic enzyme (sMMO). A system for the site-directed mutagenesis of residues in the active site of sMMO has recently been developed that uses a sMMO-minus strain of Methylosinus trichosporium OB3b as the expression host (Smith et al., 2002. Appl. Environ. Microbiol. 68:5265-5273); this strain, designated Mutant F, was created by disrupting the mmoX gene by marker-exchange mutagenesis. In this study a Ms. trichosporium OB3b strain was created in which all the sMMO structural genes were deleted or disrupted. This mutant was designated Ms. trichosporium SMDM. The recombinant expression of sMMO was performed in Ms. trichosporium SMDM using the same sMMO expression plasmid used for Ms. trichosporium Mutant F. The effect of sMMO expression in the absence of the enigmatic mmoD gene was investigated. Preliminary results indicate that mmoD is required for active expression of sMMO. The sMMO genes from Methylocella silvestris BL2T were sequenced and conjugated into Ms. trichosporium SMDM on a broad host range plasmid. No expression of Methylocella silvestris BL2 T sMMO was detected in Ms. trichosporium SMDM. A new system for the mutagenesis of the Ms. trichosporium OB3b sMMO a-subunit was created. Chimaeric sMMO mutants were created by introducing gene sequence from the alkene monooxygenase enzyme of Rhodococcus corallin us into the mmoX. The chimaeric sMMO enzymes appeared to be unstable in Ms. trichosporium Mutant F. An attempt was made to improve the stability of sMMO mutants in Ms. trichosporium Mutant F by disrupting the gene encoding the Lon protease. The Ms. trichosporium OB3b Ion gene was cloned and sequenced and attempts were made to disrupt the Ms. trichosporium OB3b Ion by marker-exchange mutagenesis. A mutant was not obtained, suggesting that Lon may be essential for vegetative growth of Ms. trichosporium OB3b. The diversity of sMMO in several environmental samples was investigated using PCR. The objective was to isolate novel mmoX sequences from uncultivated methanotrophs that could be used to design sMMO mutagenesis experiments. New PCR primers targeting the mmoX were developed. The primers were used to generate libraries from a blanket bog peat (UK) and from cave water (Romania). A group of sequences that did not cluster with the mmoX of any cultivated methanotroph was obtained from the cave water. The use of a PCR independent approach to clone methanotroph genes from environmental samEles was also investigated. This was performed by developing a method to clone 1 C-DNA from a stable isotope probing experiment with 13CH4 into a BAC vector. A library of 2300 clones was generated. Greater than 95 % of plasmids analysed contained inserts, which ranged in size from approximately 10 - 30 kb. The library was screened for mxaF, mmoX and pmoA by colony hybridization. A clone (15 kb) containing pmoA was completely sequenced. Other genes encoding proteins with (potential) roles in methylotrophy were contained on the clone, includingpmoC, pmoB, folP, folK, mptG and moxF.

No description available.

The variety of life forms within a given species, ecosystem, biome or planet is known as biodiversity. Biodiversity can also be referred as species diversity and species richness. Understanding the drivers of biodiversity requires an understanding of intertwined biotic and abiotic factors, including climate patterns over the earth, primary productivity processes, e.g. photosynthetic pathways which change with climate and latitude; latitude, geology, soil science, ecology and behavioural science. Diversity of living organisms is not evenly distributed; instead it differs significantly across the globe as well as within regions. The aim of my study is to try to understand the diversity patterns of aquatic plants, using both information derived from previous studies and by collecting new data across the globe, allowing me to examine the underlying mechanisms driving biodiversity at regional and local scales. Both geographical location and local environmental factors were found to contribute to variation in macrophyte assemblage and alpha diversity (i.e. number of species in a locality), with important roles being played by local biotic interactions and abiotic environmental factors. Overall aquatic plants, or macrophytes, play a significant role in the ecology of large numbers of freshwater ecosystems worldwide. For the purpose of my study only calcareous steams, located in both temperate and tropical/subtropical regions were included. Such streams are common in catchments throughout the world because approximately one fifth of the earth’s surface is underlain by carbonate-containing rock. Overall my findings in Chapter 3 provide evidence that there is a high variation in macrophyte assemblages of calcareous rivers across the different countries included in my study, broadly agreeing with information from the literature. I found two large groups based on species assemblages across the different countries included, i.e. a subtropical/tropical and a temperate group. As demonstrated in different parts of Chapter 4, it is possible to identify different 4 diversity responses of macrophyte functional groups to environmental conditions, at local scale, in hardwater rivers. Width and flow were found to be significantly affecting the distribution patterns of diversity of free-floating and floating-leaved rooted species, whereas diversity of marginal species was significantly related to alkalinity and width, and floating-leaved rooted diversity was significantly related to alkalinity. Last but not least submerged species were related to shading. Chapter 5 shows that variation in richness and community structure for hardwater river macrophytes can be partly explained by environmental variation relative to spatial processes in the British Isles (temperate scenario) and in Zambia (tropical scenario). Among the environmental variables, climatic ones explained a great part of species richness and composition distribution for the British Isles. Conversely in Zambia spatial processes made the greatest contribution to variation in hardwater river macrophyte species richness and community structure. Moreover Chapter 6 illustrates how macrophyte species richness, measured as alpha-diversity in calcareous rivers, was at best only very weakly attributed to latitudinal gradient. This is most likely due to the effect of other physical, chemical and biotic variables overriding broader-scale influences on species richness, at more local scales.

Biomineralisation is the process by which living things produce hard mineral tissues with unique physical properties. The study of this process can help us produce biomimetic materials, reproducing such properties, with the study of nucleation and crystallisation of the materials being particularly important. I have used molecular simulation techniques to help gain a greater understanding of these processes, focussing particularly on identifying the conformations and solid phases available to nanoparticles of two biomineral compounds. The bones and teeth of mammals are made largely of calcium phosphates. I have used metadynamics to study nanoparticles of tricalcium phosphate (TCP) and have identified high and lower order configurations. To facilitate this work I reviewed the extant empirical potentials for calcium phosphate systems, selecting the most appropriate for TCP. Calcium carbonate, found in examples throughout the animal kingdom, has three crystalline polymorphs relevant to biomineralisation: calcite, aragonite and vaterite. While nanoparticles of calcite have been extensively studied the other polymorphs have been neglected to date. In this work I present a technique for predicting crystalline morphologies for all three polymorphs across a range of sizes, and compare the energetic ordering. In water the energetic ordering of the nanoparticles is heavily dependent on nanoparticle size. Furthermore, I present work calculating the surface enthalpies of a variety of calcium carbonate surfaces, many of which are negative. It appears that entropic penalty of ordered water is key to understanding the stability of nanocrystals. Also presented is an application of the nudged elastic band method to study transitions between nanoparticle crystal conformations. Between all three crystal polymorphs the nanoparticles passed through an amorphous region of phase space. These results have also been used to evaluate order parameters for use in metadynamics simulations.

This thesis examines the colony size structure and taxonomic composition of coral communities from eight regions of the Indian Ocean approximately 10 years after thermal stress-induced mass mortality events. Coral community composition and population structure differed widely within and between regions, reflecting the different climatic and anthropogenic impacts experienced by each over the past decade. Coral communities in most areas started from a similarly depleted condition but after 1998 their recovery trajectories varied significantly, reflecting different surviving adult communities and continuing, different local stressors; some have remained highly depleted, while others have shown marked recovery. Profound differences between coral communities at intra and inter-regional spatial scales are identified and related to diversity and taxonomic composition, colony abundance, surface area, size frequency distributions, and population demographic parameters within taxa. These are analysed through multivariate techniques and univariate graphical representations to illustrate the significantly different size frequency distributions, taxonomic composition, taxonomic richness and dominance patterns at different spatial scales. A novel technique is assessed for surveying juvenile coral communities, using ultraviolet light, which causes new, growing tissue to fluoresce. This method significantly increases detected juveniles, with important consequences to size frequency patterns and to some previously published views on juvenile densities. The surveying methodologies used are far more revealing than most commonly-used conventional benthic assessments such as intercept surveys, cover values and diversity, which rarely capture discriminatory information on overall composition of coral communities, let alone the structure of populations within them. These colony size-based studies of individual genera are extremely sensitive for interpreting spatial and temporal variations in reefs and greatly enhance understanding of coral reef condition and complexity. The spatial differences demonstrate the applicability of the methods for advising reef management, specifically in identifying areas where ecological resilience is impeded by recruitment failure. Long-term consequences of changes in coral communities may include reduced ecological functional redundancy, reduced structural complexity, reduced carbonate accretion and reef growth, and impaired recovery potential.

There has been growing concern by policy and other decision makers that timber extraction by local communities is the main threat to achieving sustainable management of mangrove systems in Unguja Island, Zanzibar, Tanzania. However, this concern, and responses to the perceived threat to date, do not appear to be informed by a clear understanding of the complexity and capacity of mangrove Social-Ecological Systems (SES) at different scales to adapt to this and other disturbances. The aim of this study was to assess the resilience of mangroves to the increased demand for provisioning ecosystem services and other drivers with a view to identifying options for sustainable mangrove management on Unguja Island. This study was guided by broad resilience concepts and specific approaches, particularly the components-relationship-innovation-continuity framework developed by Cumming et al. (2005). Data relating to both social and ecological components of the mangrove system was collected. A total of 185 plots were surveyed within mangrove forests from three case study sites of Pete- Jozani, Charawe and Michamvi Shehia (lowest administrative unit) on Unguja Island in which mangrove tree species, diameter and height of trees, the numbers of seedlings and stumps were collected to assess the ecological condition of the forests. Key informant interviews (with government officials and village stakeholders), semi-structured household interviews, village meetings and focus group discussions (with beekeepers, mangrove harvesters, village elders and village conservation organisations) were used to collect social-economic data from the three case study sites. The results showed that between the 1920s and 1970s at each case study site local communities reported that they were able to obtain diverse ecosystem services while the key variables that defined the identities of the mangrove SES were maintained. The mangrove SES from each case study site was found to have changed over the past three decades in temporal and spatial scales and currently reside at different phases of change. The current mangrove ecological systems of Pete, Charawe and Kinani (part of Michamvi) were found to have been degraded compared to the past. This was evidenced by the quality and quantity of trees present, with a relatively high density of small-sized mature trees with correspondingly small basal areas and volumes, together with significant numbers of tree stumps in the ecosystems. The areas covered by mangrove vegetation in the study sites were also found to have declined. The decline in quality and quantity of trees was found to correspond with a reduction in desirable ecosystem services as reported by communities. The levels of dependence on mangrove wood provisioning ecosystem services and management approaches have changed across the case study sites. Excessive rates of harvesting of mangrove wood were identified as the key direct driver on mangrove ecological systems, which was fuelled by several underlying drivers including poverty, population change, limited livelihood activities, inappropriate management regimes, and markets for trading mangrove wood ecosystem services. Vijichuni mangrove (another part of Michamvi) was found to be an exceptional case whereby the quality and quantity of mangrove ecological variables had improved. Availability of reliable alternative income sources by the majority of villagers and effective management institutions had contributed to these changes. The drivers identified were used to develop three alternative future scenarios to explore whether projected changes will result in the mangrove SES maintaining their identities in the future. The findings suggest that the Non-inclusive State Control scenario strictly conserves the mangroves, but does not provide alternative livelihood opportunities to improve the well-being of local communities and so is not desirable. Coastal Boom scenario, characterised by unregulated economic growth, particularly in the tourism sector and community forest management with limited benefits for local communities, results in complete degradation of mangrove and reduced wellbeing of local people. However, the Techno-green scenario which includes green growth, access to low-cost cooking energy and co-managed mangrove forests with benefits for local communities, provides decision makers and other stakeholders with an alternative pathway towards more resilient mangrove SES in Unguja.