Context: The human digestive system contains trillions of microbes collectively called the gut microbiota. Important discoveries have been made through better understanding the role the gut microbiota has in maintaining physical and mental health. Imbalances in the oral microbial community, and poor dental health, have been associated with impaired cardiovascular health, but the role of the oral microbiota as a potential modulator of human health is not well understood. One mechanism that may link the oral microbiota to good health is the nitrate-nitrite-nitric oxide (NO) reduction pathway. This, for its first step, crucially relies on the reduction of nitrate, which has been ingested through the diet, to nitrite by bacteria residing in the mouth. Among many other physiological roles, NO regulates vascular endothelial function, and therefore blood pressure (BP). In the UK, at least 25% of adults and more than 50% of people over 60 years old have elevated BP and this is considered a significant cause of premature morbidity and mortality. Epidemiological studies indicate that a greater consumption of high nitrate foodstuffs (specifically, green leafy vegetables and some fruits, such as in the 'Mediterranean' or '5 a day' diets) may protect against adverse cardiovascular events, and it is known that dietary nitrate supplementation significantly reduces BP in both young and older adults. Differences between individuals in the ability to reduce nitrate to nitrite and in the effects on BP indicate that the benefits derived from a high nitrate diet may be contingent on individuals having the 'right' oral microbiota. The ability to produce NO in the body through the classical pathway involving nitric oxide synthase (NOS) enzymes is impaired in older age, leading to a gradual increase in BP. We propose that the nitrate-nitrite-NO pathway has significant potential to compensate for dysfunctional NOS in older age. Little is known, however, about how the symbiotic relationship between the human host and the oral microbiota might change with ageing and how the microbiota could be harnessed to maintain cardiovascular health during ageing. Aims and Objectives: We aim to investigate whether the prevalence of specific nitrate-reducing bacteria residing on the surface of the tongue influences the cardiovascular benefits that may be derived from the consumption of a high nitrate diet and whether there are differences between young and older adults. This builds on our preliminary data which indicate differences in the oral microbiome between young and older adults as well as correlations between the prevalence of specific oral bacteria in an individual and their ability to reduce a standardised dietary nitrate bolus to nitrite and the observed reduction in BP. This suggests that augmentation of this nitrate-nitrite-NO pathway through manipulation of diet and/or the oral microbiome might compensate for a reduced ability to generate NO endogenously with older age. Objectives: 1) Identify the bacterial species responsible for the reduction of inorganic nitrate to nitrite in the human oral cavity. 2) Compare the oral microbiomes and nitrate reduction ability of young and older adults. 3) Establish the in vivo effects of dietary nitrate supplementation and antibacterial mouthwash on the oral microbiome, NO bioavailability/bioactivity and indices of cardiovascular health (blood pressure, arterial stiffness, flow-mediated dilation). 4) Assess the effects of nitrate, antibacterial mouthwash and the sweetening agent, xylitol, on the quantities and relative abundances of oral bacteria in an in vitro biofilm model. Potential Applications and Benefits: The results of this work, which will be undertaken in collaboration with industrial sponsor DuPont Nutrition & Health, may ultimately lead to the development of prebiotic and/or probiotic interventions that help to maintain NO homeostasis and cardiovascular health across the human lifespan.
The production of the chemical solvents acetone-butanol (AB) by the bacterium Clostridium acetobutylicum was one of the first large-scale industrial processes to be developed, and in the first part of the last century ranked second in importance only to ethanol (alcohol) production. Since its development, however, its fortunes have shown considerable fluctuations. From the peak of activity in between the first and second world wars, there has been a steep decline as new technologies made it more economic to produce these chemicals from fossil fuels. In recent years, with current concerns over global warming and severe rises in the costs of crude oil, there has been resurgence in interest cumulating in the announcement by BP/Dupont to begin biobutanol (that is butanol produced by a biological process) production in 2007. Butanol is used primarily as an industrial solvent, but it is also a replacement for petrol as a fuel. Currently, the use of ethanol as a petrol additive is widespread in the developed world. The development of alternatives to petroleum as fuels is essential if we are to reduce our reliance on finite crude oil resources. However, butanol has many properties that make it far superior to ethanol. It has a higher energy content than ethanol, and its low vapour pressure and its tolerance to water contamination in petrol blends facilitate its use in existing petrol supply and distribution channels. Moreover, butanol can be blended into petrol at higher concentrations than existing biofuels, without the need to make expensive modifications to car engines. It also gives better fuel economy than petrol-ethanol blends. A key stage in the re-establishment of the AB process will be the generation of stable strains engineered to maximise butanol production. This can be achieved by making mutations in genes that lead to the production of products other than butanol. However, despite many decades of intense research, progress has been hampered by our inability to make the necessary mutations. Due to these limitations, a complete and thorough mutational analysis of all the key steps in the fermentation process responsible for butanol production has never been undertaken and the design of butanol hyperproducing strains by means of genetic manipulation has been impossible. In recent months, University of Nottingham scientists have developed a highly effective gene tool, the ClosTron. In proof of principle studies, over a dozen genes have now been inactivated in three different clostridial species, including 7 in C. acetobutylicum. Typically, the number of mutants obtained per experiment number in the 100s, and from start to finish are generated within 8-14 days. To date the system has been 100% effective, and opens up the possibility of revolutionizing metabolic engineering in Clostridium, through both gene inactivation and gene addition. We are, therefore, uniquely able to now undertake large scale targeted mutagenesis in C. acetobutylicum with very high efficiency. This will enable us to manipulate the AB fermentation pathway in a way that will either increase or abolish the generation of unwanted products. Constructing such a series of mutants is not only an essential prerequisite for the development of effective industrial strains, but will also allow us to identify the signals that control solvent formation. This should allow industry to more effectively control the production of butanol. Upon completion of ours studies we will, therefore, have generated a prototype production strain in which the yields of butanol have been maximised. Our analysis will have also have identified those signals which control solvent production. The results obtained will provide valuable information and strains useful to the fledgling biobutanol industry.
With worldwide production of light crude oil reserves expected to last ~50 years, there is a need to exploit alternative fuel resources e.g. oil sands. Vast oil-sand resources are already being exploited, resulting in large-scale pollution. They contain complex mixtures of aliphatic and aromatic acids known as 'naphthenic acids' (NAs) that are highly toxic to humans and the environment. During refining, over 1 billion m3 of wastewaters are generated containing high NA concentrations (40-120 mg/L). These toxic wastewaters are stored in large ponds for many years (often decades) before their toxicity is reduced to acceptable levels. NAs can also block or corrode pipes and oil-processing equipment causing further pollution and billion-dollar losses to the industry. High NA concentrations found in oil also reduce the saleable value of petroleum products. Thus, removing NA contamination is of great importance to the global economy, environment and human health. Microbial treatment of NAs has clear cost-environmental advantages. However, the transformation of organic compounds is complex and influenced by a combination of microbial activities/ interactions, biogeochemical factors and the physical-chemical properties of the compound. Our aims and objectives will be to identify the main organisms responsible for NA biodegradation, investigate their interactions, obtain and optimize NA-degrading pure cultures and mixed communities, and validate the rapidity of degradation/ detoxification of NA-contaminated wastewaters. We will follow the degradation process, metabolite accumulation, toxicity, biosurfactant production and microbial community composition. We will design gene probes based on molecular analysis of the main microbes found in the environment, and our new isolates. However, almost nothing is known about the metabolic pathways of NA-degrading microbes (and thus we lack suitable gene probes). The University of Essex (UoE), is at the forefront of research into pollution microbiology, and has significantly advanced of our understanding of NA biodegradation and already begun to elucidate NA catabolic pathways and we will build on our existing knowledge in order to develop suitable gene probes. This study has two potential applications and benefits. A: It will provide a better understanding of the microbes and specific conditions required for the rapid removal of these recalcitrant, toxic compounds from the environment. B: It will provide a better understanding of novel microbial interactions and degradation pathways involved. This study will also have several beneficial outcomes, it will: 1) Provide a cost-effective rapid bioremediation strategy for ecosystems with severe NA contamination 2) Develop cleaner more saleable fuels 3) Identify novel microbes and catabolic pathways with potential applications in cleaner biotechnological processes 4) It will allow gene probes to be developed to determine the degradative potential of other NA-contaminated sites elsewhere 5) Exploit novel fuel resources 6) It may allow possible new discoveries to be made e.g. reveal novel biosurfactants for biotechnological exploitation e.g. biodegradation & microbial enhanced oil recovery, anti-corrosion, oil up-grade etc.
Fluidised bed dryers in the pharmaceutical industry are currently operated by trial-and-error because of the lack of means for online measurement of granule moisture. This proposed project is to follow a current EPSRC project Product quality control of fluidised bed dryers with tomographic imaging and online process modelling (GR/S14726/01). While we have made good progress on fundamental understanding of the complicated fluidisation processes and online measurement of granule moisture and also our research results have attracted considerable interest from industry, it is necessary to bridge the gap between the current funding and future commercial funding. The aim of this proposed Follow-on project is to meet this need by scaling up from a lab scale dryer to semi-industrial scale dryers, re-engineering electrical capacitance tomography (ECT) sensors and demonstrating the principle of online measurement of granule moisture and feedback control of semi-industrial scale fluidised bed dryers in an industrial lab environment. The success of this project will advance the technology to a stronger position to attract commercial interest and further commercial seed funding, helping the UK fluidised bed dryer manufacturing industry and the UK pharmaceutical industry to increase their competitiveness in the world. As a key collaborator, GEA Process Engineering (NPS) Ltd will provide strong support to the project by providing semi-industrial scale facilities, re-engineering sensors and giving technical and market advice. Sherwood Scientific Ltd, Sensatech Research Ltd and DuPont will also support the project as industrial partners and have expressed their interest in commercialisation. The University of Manchester Intellectual Property (UMIP) Ltd has filed a UK Patent (GB0717080.6) to protect the outputs of the current project, has managed the commercial aspect of the current EPSRC project, and has included this proposed project in their active project portfolio. It is anticipated that by demonstrating this patented technology on semi-industrial scale fluidised bed dryers, a number of other applications would open up in other industries, e.g. the food, marine, power and process industries.
The polymer industry is intimately involved in every aspect of our lives, producing commodity plastics, modern electronics, biomedical materials, and much more. However, the vast majority of these polymers are derived from petroleum resources, creating both economic uncertainty and environmental risk. Biodegradable polyesters like poly(lactic acid) have provided one of the most promising solutions to this challenge, building plastics from renewable resources through a catalysed polymerisation reaction. Despite the hype and expectation, these renewable polymers account for less than 5% of all commercial polymers. Why this limitation? Replacing all commodity plastics is difficult because of the limited range of properties accessible with PLA and its copolymers. One strategy to overcome this final roadblock is to develop polyesters with different functional groups - however current synthetic methods provide low yields of monomer feedstocks or derive from toxic reagents. This proposal builds on an important recent discovery of a new synthetic strategy to target these structurally divergent plastics. New monomers, built entirely from renewable resources, can be ring-opened to afford plastics with a broad range of functional groups. The products have the potential to transform traditional polymer markets, potentially serving as biodegradable mimics of polystyrene, as new biodegradable feedstocks for health applications, and as commodity plastics with a significantly broader range of thermal properties. Importantly, this project will also address the first stages of monomer and polymer scale up, moving the discovery from laboratory towards an industrial scale to facilitate commercialisation and materials testing of the new plastics.