Genome analysis and characterisation of the exopolysaccharide produced by Bifidobacterium longum subsp. longum 35624™

The Bifibobacterium longum subsp. longum 35624™ strain (formerly named Bifidobacterium longum subsp. infantis) is a well described probiotic with clinical efficacy in Irritable Bowel Syndrome clinical trials and induces immunoregulatory effects in mice and in humans. This paper presents (a) the genome sequence of the organism allowing the assignment to its correct subspeciation longum; (b) a comparative genome assessment with other B. longum strains and (c) the molecular structure of the 35624 exopolysaccharide (EPS624). Comparative genome analysis of the 35624 strain with other B. longum strains determined that the sub-speciation of the strain is longum and revealed the presence of a 35624-specific gene cluster, predicted to encode the biosynthetic machinery for EPS624. Following isolation and acid treatment of the EPS, its chemical structure was determined using gas and liquid chromatography for sugar constituent and linkage analysis, electrospray and matrix assisted laser desorption ionization mass spectrometry for sequencing and NMR. The EPS consists of a branched hexasaccharide repeating unit containing two galactose and two glucose moieties, galacturonic acid and the unusual sugar 6-deoxy-L-talose. These data demonstrate that the B. longum 35624 strain has specific genetic features, one of which leads to the generation of a characteristic exopolysaccharide.

Mapping the gene for autosomal dominant restless legs syndrome in an Irish family

Restless Legs Syndrome (RLS) is a common neurological disorder affecting nearly 15% of the general population. Ironically, RLS can be described as the most common condition one has never heard of. It is usually characterised by uncomfortable, unpleasant sensations in the lower limbs inducing an uncontrollable desire to move the legs. RLS exhibits a circadian pattern with symptoms present predominantly in the evening or at night, thus leading to sleep disruption and daytime somnolence. RLS is generally classified into primary (idiopathic) and secondary (symptomatic) forms. Primary RLS includes sporadic and familial cases of which the age of onset is usually less than 45 years and progresses slowly with a female to male ratio of 2:1. Secondary forms often occur as a complication of another health condition, such as iron deficiency or thyroid dysfunction. The age of onset is usually over 45 years, with an equal male to female ratio and more rapid progression. Ekbom described the familial component of the disorder in 1945 and since then many studies have been published on the familial forms of the disorder. Molecular genetic studies have so far identified ten loci (5q, 12q, 14p, 9p, 20p, 16p, 19p, 4q, 17p). No specific gene within these loci has been identified thus far. Association mapping has highlighted a further five areas of interest. RLS6 has been found to be associated with SNPs in the BTBD9 gene. Four other variants were found within intronic and intergenic regions of MEIS1, MAP2K5/LBXCOR1, PTPRD and NOS1. The pathophysiology of RLS is complex and remains to be fully elucidated. Conditions associated with secondary RLS, such as pregnancy or end-stage renal disease, are characterised by iron deficiency, which suggests that disturbed iron homeostasis plays a role. Dopaminergic dysfunction in subcortical systems also appears to play a central role. An ongoing study within the Department of Pathology (University College Cork) is investigating the genetic characteristics of RLS in Irish families. A three generation RLS pedigree RLS3002 consisting of 11 affected and 7 unaffected living family members was recruited. The family had been examined for four of the known loci (5q, 12q, 14p and 9p) (Abdulrahim 2008). The aim of this study was to continue examining this Irish RLS pedigree for possible linkage to the previously described loci and associated regions. Using informative microsatellite markers linkage was excluded to the loci on 5q, 12q, 14p, 9p, 20p, 16p, 19p, 4q, 17p and also within the regions reported to be associated with RLS. This suggested the presence of a new unidentified locus. A genome-wide scan was performed using two microsatellite marker screening sets (Research Genetics Inc. Mapping set and the Applied Biosystems Linkage mapping set version 2.5). Linkage analysis was conducted under an autosomal dominant model with a penetrance of 95% and an allele frequency of 0.01. A maximum LOD score of 3.59 at θ=0.00 for marker D19S878 indicated significant linkage on chromosome 19p. Haplotype analysis defined a genetic region of 6.57 cM on chromosome 19p13.3, corresponding to 2.5 Mb. There are approximately 100 genes annotated within the critical region. Sequencing of two candidate genes, KLF16 and GAMT, selected on the assumed pathophysiology of RLS, did not identify any sequence variant. This study provides evidence of a novel RLS locus in an Irish pedigree, thus supporting the picture of RLS as a genetically heterogeneous trait.

Soft sediment ecology of Lough Hyne Marine Reserve - spatial and temporal patterns

Distribution of soft sediment benthic fauna and the environmental factors affecting them were studied, to investigate changes across spatial and temporal scales. Investigations took place at Lough Hyne Marine Reserve using a range of methods. Data on the sedimentation rates of organic and inorganic matter were collected at monthly intervals for one year at a number of sites around the Lough, by use of vertical midwater-column sediment traps. Sedimentation of these two fractions were not coupled; inorganic matter sedimentation depended on hydrodynamic and weather factors, while the organic matter sedimentation was more complex, being dependent on biological and chemical processes in the water column. The effects of regular hypoxic episodes on benthic fauna due to a natural seasonal thermocline were studied in the deep Western Trough, using camera-equipped remotely-operated vehicle to follow transects, on a three-monthly basis over one year. In late summer, the area below the thermocline of the Western Trough was devoid of visible fauna. Decapod crustaceans were the first taxon to make use of ameliorating oxygen conditions in autumn, by darting below the thermocline depth, most likely to scavenge. This was indicated by tracks that they left on the surface of the Trough floor. Some species, most noticeably Fries’ goby Lesueurigobius friesii, migrated below the thermocline depth when conditions were normoxic and established semi-permanent burrows. Their population encompassed all size classes, indicating that this habitat was not limited to juveniles of this territorial species. Recolonisation by macrofauna and burrowing megafauna was studied during normoxic conditions, from November 2009 to May 2010. Macrofauna displayed a typical post-disturbance pattern of recolonisation with one species, the polychaete Scalibregma inflatum, occurring at high abundance levels in March 2010. In May, this population had become significantly reduced and a more diverse community was established. The abundance of burrowing infauna comprising decapods crabs and Fries’ gobies, was estimated by identifying and counting their distinctive burrow structures. While above the summer thermocline depth, burrow abundance increased in a linear fashion, below the thermocline depth a slight reduction of burrow abundance occurred in May, when oxygen conditions deteriorated again. The majority of the burrows occurring in May were made by Fries’ gobies, which are thought to encounter low oxygen concentrations in their burrows. Reduction in burrow abundance of burrowing shrimps Calocaris macandreae and Callianassa subterranea (based on descriptions of burrow structures from the literature), from March to May, might be related to their reduced activity in hypoxia, leading to loss of structural burrow maintenance. Spatial and temporal changes to macrofaunal assemblage structures were studied seasonally for one year across 5 sites in the Lough and subject to multivariate statistical analysis. Assemblage structures were significantly correlated with organic matter levels in the sediment, the amounts of organic matter settling out of the water column one month before macrofaunal sampling took place as well as current speed and temperature. This study was the first to investigate patterns and processes in the Lough soft sediment ecology across all 3 basins on a temporal and spatial scale. An investigation into the oceanographic aspects of the development, behaviour and break-down of the summer thermocline of Lough Hyne was performed in collaboration with researchers from other Irish institutions.

Development of a triple stage heat transformer for the recycling of low temperature heat energy

Absorption heat transformers are thermodynamic systems which are capable of recycling industrial waste heat energy by increasing its temperature. Triple stage heat transformers (TAHTs) can increase the temperature of this waste heat by up to approximately 145˚C. The principle factors influencing the thermodynamic performance of a TAHT and general points of operating optima were identified using a multivariate statistical analysis, prior to using heat exchange network modelling techniques to dissect the design of the TAHT and systematically reassemble it in order to minimise internal exergy destruction within the unit. This enabled first and second law efficiency improvements of up to 18.8% and 31.5% respectively to be achieved compared to conventional TAHT designs. The economic feasibility of such a thermodynamically optimised cycle was investigated by applying it to an oil refinery in Ireland, demonstrating that in general the capital cost of a TAHT makes it difficult to achieve acceptable rates of return. Decreasing the TAHT's capital cost may be achieved by redesigning its individual pieces of equipment and reducing their size. The potential benefits of using a bubble column absorber were therefore investigated in this thesis. An experimental bubble column was constructed and used to track the collapse of steam bubbles being absorbed into a hotter lithium bromide salt solution. Extremely high mass transfer coefficients of approximately 0.0012m/s were observed, showing significant improvements over previously investigated absorbers. Two separate models were developed, namely a combined heat and mass transfer model describing the rate of collapse of the bubbles, and a stochastic model describing the hydrodynamic motion of the collapsing vapour bubbles taking into consideration random fluctuations observed in the experimental data. Both models showed good agreement with the collected data, and demonstrated that the difference between the solution's temperature and its boiling temperature is the primary factor influencing the absorber's performance.

Factors relating to motivation to change behaviour in individuals who are overweight: application of the health belief model

Background: Obesity is the most important health challenge faced at a global level and represents a rapidly growing problem to the health of populations. Given the escalating global health problem of obesity and its co-morbidities, the need to re-appraise its management is more compelling than ever. The normalisation of obesity within our society and the acceptance of higher body weights have led to individuals being unaware of the reality of their weight status and gravity of this situation. Recognition of the problem is a key component of obesity management and it remains especially crucial to address this issue. A large amount of research has been undertaken on obesity however, limited research has been undertaken using the Health Belief Model. Aim: The aim of the research was to determine factors relating to motivation to change behaviour in individuals who perceive themselves to be overweight and investigate whether the constructs of the Health Belief Model help to explain motivation to change behaviour. Method: The research design was quantitative, correlational and cross-sectional. The design was guided by the Health Belief Model. Data Collection: Data were collected online using a multi-section and multi-item questionnaire, developed from a review of the theoretical and empirical research. Descriptive and inferential statistical analyses were employed to describe relationships between variables. Sample: A sample of 202 men and women who perceived themselves to be overweight participated in the research. Results: Following multivariate regression analysis, perceived barriers to weight loss and perceived benefits of weight loss were significant predictors of motivation to change behaviour. The perceived barriers to weight loss which were significant were psychological barriers to weight loss (p =<0.019) and environmental barriers to physical activity (p=<0.032).The greatest predictor of motivation to change behaviour was the perceived benefits of weight loss (p<0.001). Perceived susceptibility to obesity and perceived severity of obesity did not emerge as significant predictors in this model. Total variance explained by the model was 33.5%. Conclusion: Perceived barriers to weight loss and perceived benefits of weight loss are important determinants of motivation to change behaviour. The current study demonstrated the limited applicability of the Health Belief Model constructs to motivation to change behaviour, as not all core dimensions proved significant predictors of the dependant variable.

Risk indicators associated with root caries in independently living older adults

Objective: To determine the risk indicators associated with root caries experience in a cohort of independently living older adults in Ireland. Methods: The data reported in the present study were obtained from a prospective longitudinal study conducted in a cohort of independently living older adults (n = 334). Each subject underwent an oral examination, performed by a single calibrated examiner, to determine the root caries index and other clinical variables. Questionnaires were used to collect data on oral hygiene habits, diet, smoking and alcohol habits and education level. A regression analysis with the outcome variable of root caries experience (no/yes) was conducted. Results: A total of 334 older dentate adults with a mean age of 69.1 years were examined. 53.3% had at least one filled or decayed root surface. The median root caries index was 3.13 (IQR 0.00, 13.92). The results from the multivariate regression analysis indicated that individuals with poor plaque control (OR 9.59, 95% CI 3.84–24.00), xerostomia (OR 18.49, 95% CI 2.00–172.80), two or more teeth with coronal decay (OR 4.50, 95% CI 2.02–10.02) and 37 or more exposed root surfaces (OR 5.48, 95% CI 2.49–12.01) were more likely to have been affected by root caries. Conclusions: The prevalence of root caries was high in this cohort. This study suggests a correlation between root caries and the variables poor plaque control, xerostomia, coronal decay (≥2 teeth affected) and exposed root surfaces (≥37). The significance of these risk indicators and the resulting prediction model should be further evaluated in a prospective study of root caries incidence. Clinical significance Identification of risk indicators for root caries in independently living older adults would facilitate dental practitioners to identify those who would benefit most from interventions aimed at prevention.

On classical and other methods of discriminant analysis and estimation of log-odds

For two multinormal populations with equal covariance matrices the likelihood ratio discriminant function, an alternative allocation rule to the sample linear discriminant function when n1 ≠ n2 ,is studied analytically. With the assumption of a known covariance matrix its distribution is derived and the expectation of its actual and apparent error rates evaluated and compared with those of the sample linear discriminant function. This comparison indicates that the likelihood ratio allocation rule is robust to unequal sample sizes. The quadratic discriminant function is studied, its distribution reviewed and evaluation of its probabilities of misclassification discussed. For known covariance matrices the distribution of the sample quadratic discriminant function is derived. When the known covariance matrices are proportional exact expressions for the expectation of its actual and apparent error rates are obtained and evaluated. The effectiveness of the sample linear discriminant function for this case is also considered. Estimation of true log-odds for two multinormal populations with equal or unequal covariance matrices is studied. The estimative, Bayesian predictive and a kernel method are compared by evaluating their biases and mean square errors. Some algebraic expressions for these quantities are derived. With equal covariance matrices the predictive method is preferable. Where it derives this superiority is investigated by considering its performance for various levels of fixed true log-odds. It is also shown that the predictive method is sensitive to n1 ≠ n2. For unequal but proportional covariance matrices the unbiased estimative method is preferred. Product Normal kernel density estimates are used to give a kernel estimator of true log-odds. The effect of correlation in the variables with product kernels is considered. With equal covariance matrices the kernel and parametric estimators are compared by simulation. For moderately correlated variables and large dimension sizes the product kernel method is a good estimator of true log-odds.

Survival analysis of adult tuberculosis disease

Background: We conducted a survival analysis of all the confirmed cases of Adult Tuberculosis (TB) patients treated in Cork-City, Ireland. The aim of this study was to estimate Survival time (ST), including median time of survival and to assess the association and impact of covariates (TB risk factors) to event status and ST. The outcome of the survival analysis is reported in this paper. Methods: We used a retrospective cohort study research design to review data of 647 bacteriologically confirmed TB patients from the medical record of two teaching hospitals. Mean age 49 years (Range 18–112). We collected information on potential risk factors of all confirmed cases of TB treated between 2008–2012. For the survival analysis, the outcome of interest was ‘treatment failure’ or ‘death’ (whichever came first). A univariate descriptive statistics analysis was conducted using a non- parametric procedure, Kaplan -Meier (KM) method to estimate overall survival (OS), while the Cox proportional hazard model was used for the multivariate analysis to determine possible association of predictor variables and to obtain adjusted hazard ratio. P value was set at <0.05, log likelihood ratio test at >0.10. Data were analysed using SPSS version 15.0. Results: There was no significant difference in the survival curves of male and female patients. (Log rank statistic = 0.194, df = 1, p = 0.66) and among different age group (Log rank statistic = 1.337, df = 3, p = 0.72). The mean overall survival (OS) was 209 days (95%CI: 92–346) while the median was 51 days (95% CI: 35.7–66). The mean ST for women was 385 days (95%CI: 76.6–694) and for men was 69 days (95%CI: 48.8–88.5). Multivariate Cox regression showed that patient who had history of drug misuse had 2.2 times hazard than those who do not have drug misuse. Smokers and alcohol drinkers had hazard of 1.8 while patients born in country of high endemicity (BICHE) had hazard of 6.3 and HIV co-infection hazard was 1.2. Conclusion: There was no significant difference in survival curves of male and female and among age group. Women had a higher ST compared to men. But men had a higher hazard rate compared to women. Anti-TNF, immunosuppressive medication and diabetes were found to be associated with longer ST, while alcohol, smoking, RICHE, BICHE was associated with shorter ST.