A range of diseases commonly affect cattle in the UK and worldwide including bovine mastitis, brucellosis, bluetongue, bovine viral diarrhoea, foot and mouth disease, Johne's disease and bovine TB. These have major implications for animal health, food security and the economy. Vaccination is widely considered the most effective control strategy, but in most cases vaccines still need to be developed or improved, necessitating screening studies, and all licenced vaccines need to be regularly batch-tested. Current in vitro models fail to capture the complex features of the adaptive immune response required to assess antigen-specific vaccine immunogenicity in a satisfactory way, and rodents are a poor model due to key differences in their immune systems compared with cattle. Thus in vivo cattle experiments are commonly used, which are ethically, logistically and economically costly. We propose to develop bovine organoids based on secondary lymphoid organs as a new tool for screening bovine vaccines to replace the use of cattle (and rodents) in early vaccine development and vaccine batch-testing. Organoids are self-organised 3D tissue that mimic the key functional, structural and biological complexity of an organ. Bovine organoids have been developed for several other anatomical locations in cattle including the mammary glands and intestine, indicating feasibility. Immune organoids have recently been reported in humans, with demonstrated ability to measure primary and memory responses to antigens, adjuvants and vaccines. Our preliminary data shows that it is possible to grow organoids from cryopreserved bovine lymph node material, with the expected reaggregation of cells and good survival over an extended culture period. We aim to optimise, validate and implement the bovine immune organoid model using bovine TB as an exemplar for proof-of-concept in collaboration with the Animal and Plant Health Agency. Project Partner Lisa Wagar from the University of Irvine, California, pioneered the human immune organoid model and will provide hands-on training and expertise. For the optimisation phase, we will obtain secondary lymphoid tissue from naïve and BCG vaccinated cattle, and use this to identify the most appropriate tissue type and assay conditions including composition and concentrations of the components as well as the kinetic of response over time. We will also assess reproducibility at multiple levels. We will then validate the model by demonstrating that it can mirror immunogenicity read-outs from in vivo studies. The approach to this will be three-fold: i) comparing immune responses induced by the Ad85A bovine TB vaccine in vivo in previously-conducted studies in the literature with those induced in our organoid model; ii) comparing immune responses induced by a panel of vaccine adjuvants in a previously-conducted study in the literature with those induced in our organoid model; and iii) directly comparing immune responses induced by four novel bovine TB vaccine candidates using an in vivo cattle study and our immune organoid model in parallel. To demonstrate that the model is fit for purpose, we will then implement it to screen an expanded panel of bovine TB vaccine candidates. Finally, we will establish a 3Rs legacy locally and globally by actively disseminating methods, protocols and results; by supporting training and tech-transfer to relevant end-user laboratories; and by generating a small-scale bovine lymphoid tissue bank. The advantages of bovine immune organoids include the clear 3Rs benefits in replacing the use of cattle and rodents in early bovine vaccine screening and batch-testing, the scientific benefits in offering a high-throughput and tractable model to expedite bovine vaccine development, cost-effectiveness, transferability including to resource-limited settings, and adaptability for use across a range of bovine diseases as well as other ungulate species of importance in vaccine development.

Injuries caused by trauma, infections and inflammation to the surface of the eye can cause scarring that 'clouds' the transparent window of the eye called the cornea, interfering with vision and is sight-threatening. 'Corneal Blindness' affects millions of people and the World Health Organisation have made curing the problem a priority area programme to prevent world-wide blindness. The current treatment for damaged eyes caused by infection is to treat with antibiotic agents followed by strategies to promote healing. We are developing a synthetic, optically-transparent, anti-scarring dressing (biomembrane) suitable for the management of patients world-wide at risk of corneal scarring following injury, by promoting cells and molecules in tissues to heal without scarring improving patient visual outcomes. To date we have: (1), engaged closely with clinical colleagues to define and refine the characteristics and technical specifications of the fluid biomembrane dressing to ensure suitability for clinical use (2), investigated processes for its manufacture (3), performed limited testing of a dressing prototype (4), prepared a technical portfolio of the performance data generated thus far. We now seek further funding to allow progression of the project so that we can: (1), refine and improve the characteristics and performance of the dressing (2), scale-up its manufacture for use in humans (3), obtain regulatory approvals for testing in humans and finally (4), undertake a small clinical trial to check for safety and see how well the anti-scarring dressing works on patients with infected corneas. At this same time we will be (5), developing a commercialisation plan so that the new dressing becomes widely available for use in the clinic to reduce ocular scar formation that can cause blindness.

Humans are modifying the diversity of life on Earth, changing the number and composition of species in ecosystems. It is essential to understand how these changes will affect how natural systems function. This project evaluates a promising method for predicting the impacts of biodiversity change by linking species' physical characteristics - their shapes and forms - to the key roles that they play in communities and ecosystems. These links between traits and emergent roles are poorly described, especially in coastal marine environments. This project involves screening the traits of seaweed species along gradients in environmental conditions on rocky shorelines - tide height, exposure to heavy surf, and latitude - along the U.S. West Coast and (via a partnership with the Natural Environment Research Council in the United Kingdom) the coast of Europe. This project includes experiments and outreach in regions with substantial exposure to the public, and the investigators will work with community, university, and museum outreach personnel to communicate this research to broader audiences. The project includes mechanisms for curriculum development and outreach and trains undergraduate and graduate students and postdoctoral researchers in marine science. The project also supports a visual arts exhibit for public display that illustrates the loss of seaweed species. The investigators are using a suite of innovative approaches to understand the links between functional traits and emergent functional attributes of seaweeds, providing an unprecedented dataset linking seaweed morphology, physiology, and ecology: (i) characterizing the functional structure of seaweed communities and how that structure changes across environmental gradients, (ii) evaluating how that suite of easily measured traits is related to physiological and ecological processes, (iii) measuring species and traits at multiple sites along U.S. and European coastlines spanning gradients in latitude, wave exposure, and tidal elevation, and (iv) evaluating changes in functional redundancy along these gradients to quantify vulnerability of community interactions and ecosystem functioning to species loss These methods and approaches facilitate exploration of relationships between algal form and function and provide insights into the functional consequences of changes in seaweed diversity, with the potential to transform understanding of marine biodiversity across levels of biological organization. These advances in linking seaweeds' functional traits, functional groups, and roles in community- and ecosystem-level processes enhance prediction of the impacts of changing biodiversity - especially along environmental gradients and species ranges - and serve as a template for a mechanistic understanding of the roles of species in communities and ecosystems.

Abstracts are not currently available in GtR for all funded research. This is normally because the abstract was not required at the time of proposal submission, but may be because it included sensitive information such as personal details.

Hydrogen bonds are the key to understanding aqueous chemistry and nearly all of biochemistry. Of course, hydrogen bonds determine the structure of pure water and give it many of its bizarre properties such as its density maximum at 4 degrees C and its expansion on freezing. Hydrogen bonds between amino-acid residues and cooperative effects determine the structure of peptides and proteins. Finally yet importantly, the surfaces of proteins and enzyme binding pockets are often solvated by water and there are strong indications that the protein changes the structure of the surrounding water while the water changes the dynamics of the protein. Protein-bound water appears to have many properties like that of crystalline or perhaps glassy water and often even shows up in X-ray crystal structures. It is therefore vital to study the hydrogen-bond structure and dynamics of supercooled water and especially glassy water, which can be made through nano-confinement. It is also essential to study the structure and dynamics of peptide model systems in order to be able to isolate the effects of hydrogen-bond dynamics and cooperativity.Hydrogen-bond dynamics takes on many forms. Equilibrium dynamics ranges from hydrogen-bond bend and stretch modes with periods of ~0.2-0.5 ps to diffusive translational and rotational relaxation as slow as 8 ps. Non-equilibrium dynamics (such as after the excitation of an OH-stretch mode) involve the breaking of hydrogen bonds on a ~1-2 ps timescale. These processes are difficult to study with one technique. Infrared and dielectric techniques suffer from limited frequency ranges. Raman scattering techniques suffer from weak signals and turn out not to be sensitive to rotational motion in water.Here we propose the development of the little-used ultrafast spectroscopy technique of terahertz-field-induced second-harmonic generation (TFISH). It is known to yield large signals in water and covers the entire range from rotational diffusion to hydrogen-bond bends and stretches. Moreover, it lends itself to be expanded into a multi-dimensional spectroscopy (2D-TFISH) that can be used to measure non-equilibrium dynamics. This would give us the unique opportunity to study the effects of non-equilibrium relaxation dynamics (such as the relaxation of OH-stretch and bend modes) on hydrogen bonds directly. The techniques will be used on supercooled water (bulk or confined in silica nanopores) to determine the presence of a liquid-liquid phase transition at ~220K and for the first time the effects of the phase transition on the structure and dynamics of water. They will also be used on peptide model systems such as N-methylacetamide and analogues to study cooperativity and non-equilibrium relaxation dynamics.