Metabolism of host-derived carbohydrates by Bifidobacterium breve UCC2003

Bifidobacteria are Gram positive, anaerobic, typically Y-shaped bacteria which are naturally found in the digestive tract of certain mammals, birds and insects. Bifidobacterium breve strains are numerically prevalent among the gut microbiota of many healthy breast-fed infants. The prototypical B. breve strain UCC2003 has previously been shown to utilise numerous carbohydrates of plant origin. Various aspects of host-derived carbohydrate metabolism occurring in this bacterium will be described in this thesis. Chapter II describes B. breve UCC2003 utilisation of sialic acid, a nine-carbon monosaccharide, which is found in human milk oligosaccharides (HMOs) and the mucin glycoprotein. B. breve UCC2003 was also shown to cross-feed on sialic acid released from 3’ sialyllactose, a prominent HMO, by the extracellular sialidase activity of Bifidobacterium bifidum PRL2010. Chapter III reports on the transcriptional regulation of sialic acid metabolism in B. breve UCC2003 by a transcriptional repressor encoded by the nanR gene. NanR belongs to the GntR-family of transcriptional regulators and represents the first bifidobacterial member of this family to be characterised. Chapter IV investigates B. breve UCC2003 utilisation of mucin. B. breve UCC2003 was shown to be incapable of degrading mucin; however when grown in co-culture with B. bifidum PRL2010 it exhibits enhanced growth and survival properties. A number of methods were used to investigate and identify the mucin components supporting this enhanced growth/viability phenotype. Chapter V describes the characterisation of two sulfatase-encoding gene clusters from B. breve UCC2003. The transcriptional regulation of both sulfatase-encoding gene clusters was also investigated. The work presented in this thesis represents new information on the metabolism of host-derived carbohydrates in bifidobacteria, thus increasing our understanding of how these gut commensals are able to colonise and persist in the gastrointestinal tract.

Screening the human exome: a comparison of whole genome and whole transcriptome sequencing.

BACKGROUND: There is considerable interest in the development of methods to efficiently identify all coding variants present in large sample sets of humans. There are three approaches possible: whole-genome sequencing, whole-exome sequencing using exon capture methods, and RNA-Seq. While whole-genome sequencing is the most complete, it remains sufficiently expensive that cost effective alternatives are important. RESULTS: Here we provide a systematic exploration of how well RNA-Seq can identify human coding variants by comparing variants identified through high coverage whole-genome sequencing to those identified by high coverage RNA-Seq in the same individual. This comparison allowed us to directly evaluate the sensitivity and specificity of RNA-Seq in identifying coding variants, and to evaluate how key parameters such as the degree of coverage and the expression levels of genes interact to influence performance. We find that although only 40% of exonic variants identified by whole genome sequencing were captured using RNA-Seq; this number rose to 81% when concentrating on genes known to be well-expressed in the source tissue. We also find that a high false positive rate can be problematic when working with RNA-Seq data, especially at higher levels of coverage. CONCLUSIONS: We conclude that as long as a tissue relevant to the trait under study is available and suitable quality control screens are implemented, RNA-Seq is a fast and inexpensive alternative approach for finding coding variants in genes with sufficiently high expression levels.

Modelling of the time-dependent flow behaviour of lead-free solder pastes used for flip-chip assembly applications

The market for solder paste materials in the electronic manufacturing and assembly sector is very large and consists of material and equipment suppliers and end users. These materials are used to bond electronic components (such as flip-chip, CSP and BGA) to printed circuit boards (PCB's) across a range of dimensions where the solder interconnects can be in the order of 0.05mm to 5mm in size. The non-Newtonian flow properties exhibited by solder pastes during its manufacture and printing/deposition phases have been of practical concern to surface mount engineers and researchers for many years. The printing of paste materials through very small-sized stencil apertures is known to lead to increased stencil clogging and incomplete transfer of paste to the substrate pads. At these very narrow aperture sizes the paste rheology and particle-wall interactions become crucial for consistent paste withdrawal. These non-Newtonian effects must be understood so that the new paste formulations can be optimised for consistent printing. The focus of the study reported in this paper is the characterisation of the rheological properties of solder pastes and flux mediums, and the evaluation of the effect of these properties on the pastes' printing performance at the flip-chip assembly application level. Solder pastes are known to exhibit a thixotropic behaviour, which is recognised by the decrease in apparent viscosity of paste material with time when subjected to a constant shear rate. The proper characterisation of this time-dependent theological behaviour of solder pastes is crucial for establishing the relationships between the pastes' structure and flow behaviour; and for correlating the physical parameters with paste printing performance. In this paper, we present a number of methods which have been developed for characterising the time-dependent and non-Newtonian rheological behaviour of solder pastes and flux mediums as a function of shear rates. We also present results of the study of the rheology of the solder pastes and flux mediums using the structural kinetic modelling approach, which postulates that the network structure of solder pastes breaks down irreversibly under shear, leading to time and shear dependent changes in the flow properties. Our results show that for the solder pastes used in the study, the rate and extent of thixotropy was generally found to increase with increasing shear rate. The technique demonstrated in this study has wide utility for R&D personnel involved in new paste formulation, for implementing quality control procedures used in solder paste manufacture and packaging; and for qualifying new flip-chip assembly lines

The Impact of Offshore Wind Farms on Marine Ecosystems: A Review Taking an Ecosystem Services Perspective

The rapid increase in renewable energy generation from wind has increased concerns about the impacts that wind arrays have on the marine environment and what these impacts mean for society. One method for identifying the impacts of offshore wind farms (OWFs) on human welfare is through the assessment and valuation of ecosystem services. Using an ecosystem services approach, this paper reviews the impacts of OWFs on the ecosystem services delivered by marine environments. During the construction phase, supporting services such as reduced energy capture and nutrient cycling are changed due to the introduction of hard substrate and the reduction in soft sediment habitat at turbine bases. This may lead to changes in all other ecosystem services, both negative and positive. Quantifying these changes, however, is a challenge partly due to data limitations and a lack of clear understanding of the impacts of OWFs on the marine ecosystems. Scientific effort needs to quantitatively explore the impacts of OWFs on ecosystem functionality and the gathering of data that enables the assessment of changes to ecosystem services. Data needed to better quantify and value the impacts of OWFs on ecosystem services are suggested. The development of methods which integrate socioeconomic valuation of ecosystem services into the evaluation of renewable energy devices compliments efforts in assessing the environmental impacts and should enable a holistic assessment of the impact of renewable energy production and greenhouse gas mitigation technologies on the U. K. carbon footprint.

A methodology for the assessment of local-scale changes in marine environmental benefits and its application

Local-scale planning decisions are required by the existing Environmental Impact Assessment process to take account of the implications of a development on a range of environmental and social factors, and could therefore be supported by an ecosystem services approach. However, empirical assessments at a local scale within the marine environment have focused on only a single or limited set of services. This paper tests the applicability of the ecosystem services approach to environmental impact appraisal by considering how the identification and quantification of a comprehensive suite of benefits provided at a local scale might proceed in practice. A methodology for conducting an Environmental Benefits Assessment (EBA) is proposed, the underlying framework for which follows the recent literature by placing the emphasis on ecosystem benefits, as opposed to services. The EBA methodology also proposes metrics that can be quantified at local scale, and is tested using a case study of a hypothetical tidal barrage development in the Taw Torridge estuary in North Devon, UK. By suggesting some practical steps for assessing environmental benefits, this study aims to stimulate discussion and so advance the development of methods for implementing ecosystem service approaches at a local scale.

Integrating methods for ecosystem service assessment and valuation: Mixed methods or mixed messages?

A mixed-method approach was used to assess and value the ecosystem services derived from the Dogger Bank, an extensive shallow sandbank in the southern North Sea. Three parallel studies were undertaken that 1) identified and quantified, where possible, how indicators for ecosystem service provision may change according to two future scenarios, 2) assessed members of the public's willingness-to-pay for improvements to a small number of ecosystem services as a consequence of a hypothetical management plan, and 3) facilitated a process of deliberation that allowed members of the public to explore the uses of the Dogger Bank and the conflicts and dilemmas involved in its management. Each of these studies was designed to answer different and specific research questions and therefore contributes different insights about the ecosystem services delivered by the Dogger Bank. This paper explores what can be gained by bringing these findings together post hoc and the extent to which the different methods are complementary. Findings suggest that mixed-method research brings more understanding than can be gained from the individual approaches alone. Nevertheless, the choice of methods used and how these methods are implemented strongly affects the results obtained.

A method to assess the sensitivity of sedimentary communities to fishing activities.

1.Methods of sensitivity assessment to identify species and habitats in need of management or protection have been available since the 1970s. 2.The approach to sensitivity assessment adopted by the Marine Life Information Network (MarLIN) assumes that the sensitivity of a community or biotope is dependent on the species within it. However, the application of this approach to sedimentary communities, especially offshore, is complex because of a lack of knowledge of the structural or functional role of many sedimentary species. 3.This paper describes a method to assess the overall sensitivity of sedimentary communities, based on the intolerance and recoverability of component species to physical disturbance. A range of methods were applied to identify the best combinations of abundant, dominant or high biomass species for the assessment of sensitivity in the sedimentary communities examined. 4.Results showed that reporting the most frequent species' sensitivity assessment, irrespective of the four methods used to select species, consistently underestimated the total sensitivity of the community. In contrast, reporting the most sensitive assessment from those species selected resulted in a range of biotope sensitivities from very low to very high, that was better able to discriminate between the sensitivities of the communities examined. 5.The assumptions behind the methodology, its limitations and potential application are discussed.

Grading gems: Appraising the quality of research about social work and social care.

The impetus towards basing practice and policy decisions more explicitly on sound research requires tools to facilitate the systematic appraisal of the quality of research encompassing a diverse range of methods and designs. Five exemplar tools were developed and assessed in terms of their usefulness in selecting studies for inclusion in a systematic review. The widely used ‘hierarchy of evidence’ was adapted and used to appraise internal validity. Four tools were then developed to appraise the external validity dimensions of generalizability (two scales) and methods of data collection (two scales). Methods of combining the scores generated by each tool were explored. Qualitative and quantitative studies were appraised, not separated into two spheres but by using complementary tools developed to appraise different aspects of rigour. There was a high level of agreement between researchers in applying the tools to twenty-two studies on decision making by professionals about the longterm care of older people. The scales for internal validity and generalizability discriminated between the qualities of studies appropriately. The two tools to appraise data collection gave diverse results. Excluding studies that scored in the lowest category on any scale appeared to be the scoring system that was most justifiable. This approach is presented to stimulate debate about the practical application of the evidence-based initiative to social work and social care. This study may assist in developing clearer definitions and common language about appraising rigour that should further the process of selecting robust research for synthesis to inform practice and policy decisions.

Novel approaches to the analysis of the Maillard reaction of proteins

The Maillard reaction comprises a complex network of reactions which has proven to be of great importance in both food science and medicine. The majority of methods developed for studying the Maillard reaction in food have focused on model systems containing amino acids and monosaccharides. In this study, a number of electrophoretic techniques, including two-dimensional gel electrophoresis and capillary electrophoresis, are presented. These have been developed specifically for the analysis of the Maillard reaction of food proteins, and are giving important insights into this complex process.

Developing a conceptual model of lead performance measurement and benchmarking: A multiple case analysis

Purpose: Environmental turbulence including rapid changes in technology and markets has resulted in the need for new approaches to performance measurement and benchmarking. There is a need for studies that attempt to measure and benchmark upstream, leading or developmental aspects of organizations. Therefore, the aim of this paper is twofold. The first is to conduct an in-depth case analysis of lead performance measurement and benchmarking leading to the further development of a conceptual model derived from the extant literature and initial survey data. The second is to outline future research agendas that could further develop the framework and the subject area.

Design/methodology/approach: A multiple case analysis involving repeated in-depth interviews with managers in organisational areas of upstream influence in the case organisations.

Findings: It was found that the effect of external drivers for lead performance measurement and benchmarking was mediated by organisational context factors such as level of progression in business improvement methods. Moreover, the legitimation of the business improvement methods used for this purpose, although typical, had been extended beyond their original purpose with the development of bespoke sets of lead measures.

Practical implications: Examples of methods and lead measures are given that can be used by organizations in developing a programme of lead performance measurement and benchmarking.

Originality/value: There is a paucity of in-depth studies relating to the theory and practice of lead performance measurement and benchmarking in organisations.

A preliminary investigation into the use of testate amoebae for the discrimination of forensic soil samples

We present the results of an initial investigation into the efficacy of using testate amoebae for the discrimination of soils from wet ground and puddles, as little attention has been given to these organisms in forensic science. The preservation of testate amoebae in these sediments is generally good, although test concentrations are low. Statistical analysis suggests that restate amoebae assemblages are somewhat spatially distinct and have potential to be used for soil discrimination. A case study is presented where mineralogical (X-ray diffraction) and restate amoebae analyses are used in conjunction to clarify the scene of crime in a 'cold case' murder enquiry. Testate amoebae were recovered from dried sediment residues on clothing 10 years after the murder. Despite these promising results, further experimental work is crucial to examine the spatial and temporal variation of amoebae assemblages in water films, wet ground and puddles before they can be added to the armoury of methods available to the forensic biologist.

‘Stakeholders’ views of legal and advice services for people admitted to psychiatric hospital’

This paper describes a study that used a mixed method approach to elicit the views of a range of stakeholders about experiences of compulsory admission to psychiatric hospital, and the use of the Mental Health Review Tribunal (MHRT). The paper begins with an introduction to the background of the study, one that took place in Northern Ireland, a region in the UK with its own mental health legislation and policy. A review of literature is then presented. This highlights some of the disadvantages that service users and carers face when dealing with professionals during and following compulsory admission to hospital. This section concludes with an overview of literature on the MHRT in the UK. A range of methods was used to gather data from the following stakeholders: five service user and carer focus group interviews (n = 44); interviews with four lawyers experienced in Tribunal work; an interview with a legal member of the Tribunal; a survey of solicitors who identified themselves as equipped to carry out Tribunal work; interviews with three managers of organisations that provided patient advocacy services; letters to hospital managers requesting information provided to patients and carers. The findings reveal a number of themes associated with these experiences of compulsory admission to hospital and subsequent use of the Tribunal. Service users and carers generally found it difficult to access relevant information about rights, information provided by hospital managers was uneven and lawyers were often not familiar with processes associated with compulsory admission. There was a range of views about the Tribunal. Most respondents felt that the Tribunal was necessary and mostly satisfactory in the way it carried out its functions, but stakeholders raised a number of issues. Carers in particular felt that they should be more involved in decision-making processes, whereas lawyers tended to be focused on more technical, legal issues. Problems of regrading prior to the Tribunal and in examining medical evidence were highlighted by lawyers. There was an appeal for better information and advice by service users and carers, and recognition of the need for better training and education for lawyers. The paper concludes with a brief discussion about current mental health law in the UK, arguing that, in this context, professionals should more proactively use information and advice that can enable service users and carers to defend their rights. Keywords: compulsory mental health; law; legal and advice services

Remote, subsurface detection of the algal toxin domoic acid onboard the Environmental Sample Processor: Assay development and field trials

The ability to detect harmful algal bloom (HAB) species and their toxins in real- or near real-time is a critical need for researchers studying HAB/toxin dynamics, as well as for coastal resource managers charged with monitoring bloom populations in order to mitigate their wide ranging impacts. The Environmental Sample Processor (ESP), a robotic electromechanical/fluidic system, was developed for the autonomous, subsurface application of molecular diagnostic tests and has successfully detected several HAB species using DNA probe arrays during field deployments. Since toxin production and thus the potential for public health and ecosystem effects varies considerably in natural phytoplankton populations, the concurrent detection of HAB species and their toxins onboard the ESP is essential. We describe herein the development of methods for extracting the algal toxin domoic acid (DA) from Pseudonitzschia cells (extraction efficiency >90%) and testing of samples using a competitive ELISA onboard the ESP. The assay detection limit is in the low ng/mL range (in extract), which corresponds to low ng/L levels of DA in seawater for a 0.5 L sample volume acquired by the ESP. We also report the first in situ detection of both a HAB organism (i.e., Pseudo-nitzschia) and its toxin, domoic acid, via the sequential (within 2-3 h) conduct of species- and toxin-specific assays during ESP deployments in Monterey Bay, CA, USA. Efforts are now underway to further refine the assay and conduct additional calibration exercises with the aim of obtaining more reliable, accurate estimates of bloom toxicity and thus their potential impacts. Published by Elsevier B.V.