Stepwise engineering of a Pichia pastoris D-amino acid oxidase whole cell catalyst

Trigonopsis variabilis D-amino acid oxidase (TvDAO) is a well characterized enzyme used for cephalosporin C conversion on industrial scale. However, the demands on the enzyme with respect to activity, operational stability and costs also vary with the field of application. Processes that use the soluble enzyme suffer from fast inactivation of TvDAO while immobilized oxidase preparations raise issues related to expensive carriers and catalyst efficiency. Therefore, oxidase preparations that are more robust and active than those currently available would enable a much broader range of economically viable applications of this enzyme in fine chemical syntheses. A multi-step engineering approach was chosen here to develop a robust and highly active Pichia pastoris TvDAO whole-cell biocatalyst. As compared to the native T. variabilis host, a more than seven-fold enhancement of the intracellular level of oxidase activity was achieved in P. pastoris through expression optimization by codon redesign as well as efficient subcellular targeting of the enzyme to peroxisomes. Multi copy integration further doubled expression and the specific activity of the whole cell catalyst. From a multicopy production strain, about 1.3 x 103 U/g wet cell weight (wcw) were derived by standard induction conditions feeding pure methanol. A fed-batch cultivation protocol using a mixture of methanol and glycerol in the induction phase attenuated the apparent toxicity of the recombinant oxidase to yield final biomass concentrations in the bioreactor of >or= 200 g/L compared to only 117 g/L using the standard methanol feed. Permeabilization of P. pastoris using 10% isopropanol yielded a whole-cell enzyme preparation that showed 49% of the total available intracellular oxidase activity and was notably stabilized (by three times compared to a widely used TvDAO expressing Escherichia coli strain) under conditions of D-methionine conversion using vigorous aeration. Stepwise optimization using a multi-level engineering approach has delivered a new P. pastoris whole cell TvDAO biocatalyst showing substantially enhanced specific activity and stability under operational conditions as compared to previously reported preparations of the enzyme. The production of the oxidase through fed-batch bioreactor culture and subsequent cell permeabilization is high-yielding and efficient. Therefore this P. pastoris catalyst has been evaluated for industrial purposes.

Manoeuvring control of underactuated surface vessels using manifold regulation for Port-Hamiltonian systems

This paper considers the manoeuvring of underactuated surface vessels. The control objective is to steer the vessel to reach a manifold which encloses a waypoint. A transformation of configuration variables and a potential field are used in a Port-Hamiltonian framework to design an energy-based controller. With the proposed controller, the geometric task associated with the manoeuvring problem depends on the desired potential energy (closed-loop) and the dynamic task depends on the total energy and damping. Therefore, guidance and motion control are addressed jointly, leading to model-energy-based trajectory generation.

Eldercare demands, mental health, and work performance : the moderating role of satisfaction with eldercare tasks

Due to demographic changes, a growing number of employees provide in-home care to an elderly family member. Previous research suggested a negative relationship between employees' eldercare demands and their work performance. However, the empirical nature of this relationship and its boundary conditions and mediating mechanisms have been neglected. The goal of this multisource study was to examine a mediated-moderation model of eldercare demands, mental health, and work performance. Drawing on the theory of conservation of resources (Hobfoll, 1989), it was expected that employees' satisfaction with eldercare tasks would buffer the negative relationship between eldercare demands and work performance, and that mental health would mediate this moderating effect. Data were collected from 165 employees providing in-home eldercare, as well as from one colleague and one family member of each employee. Results of mediated-moderation analyses supported the hypothesized model. The findings suggest that interventions that aim to increase employees' satisfaction with eldercare tasks may help protect employees from the negative effects of high eldercare demands on mental health and, subsequently, on work performance.

Rabobank : a process mining case study BPI Challenge 2014 report

The increased interest in the area of process improvement persuaded Rabobank Group ICT in examining its own Change-process in order to improve its competitiveness. The group is looking for answers about the effectiveness of changes applied as part of this process, with particular interest toward the presence of predictive patterns and their parameters. We conducted an analysis of the log using well established process mining techniques (i.e. Fuzzy Miner). The results of the analysis conducted on the log of the process show that a visible impact is missing.

The Thoughts Questionnaire (TQ) for family caregivers of people with dementia

Objectives: To develop a new measure of dysfunctional thoughts for family caregivers of people living with dementia. These thoughts can contribute to negative outcomes, but they may be modifiable. Method: A stepwise process was used to develop the Thoughts Questionnaire, commencing with item generation, concept mapping, and pilot testing in a sample of professional and nonprofessional caregivers of people with dementia (n = 18). Next, an independent sample of 35 family caregivers of people with dementia (30 female; M age = 64.30, standard deviation = 10.65) completed: (a) the Thoughts Questionnaire; (b) an existing measure of dysfunctional thoughts, the Dementia Thoughts Caregivers Questionnaire; and (c) separate validated measures of depressive symptoms, caregiver stress, and coping, respectively. Results: The level of agreement with dysfunctional thought statements from the Dementia Thoughts Caregivers Questionnaire and Thoughts Questionnaire was low. However, a small number of Thoughts Questionnaire statements were strongly endorsed by over 85% of the sample. Both dysfunctional thought measures had adequate reliability, but total scores were not significantly intercorrelated (r = .287, p = .095). Only the Thoughts Questionnaire was significantly, positively correlated with most caregiver stress measures. Thoughts Questionnaire items required a much lower reading level than the Dementia Thoughts Caregivers Questionnaire items. Discussion: This study provides preliminary data on a tool for assessing the negative role-related thoughts that family caregivers of people with dementia may experience. Given that these thoughts are implicated in depression but they may be modified, the capacity to identify dysfunctional thoughts may prove useful in caregiver support programs.

Refining user and item profiles based on multidimensional data for top-n item recommendation

In recommender systems based on multidimensional data, additional metadata provides algorithms with more information for better understanding the interaction between users and items. However, most of the profiling approaches in neighbourhood-based recommendation approaches for multidimensional data merely split or project the dimensional data and lack the consideration of latent interaction between the dimensions of the data. In this paper, we propose a novel user/item profiling approach for Collaborative Filtering (CF) item recommendation on multidimensional data. We further present incremental profiling method for updating the profiles. For item recommendation, we seek to delve into different types of relations in data to understand the interaction between users and items more fully, and propose three multidimensional CF recommendation approaches for top-N item recommendations based on the proposed user/item profiles. The proposed multidimensional CF approaches are capable of incorporating not only localized relations of user-user and/or item-item neighbourhoods but also latent interaction between all dimensions of the data. Experimental results show significant improvements in terms of recommendation accuracy.

A robot application to marine vessel inspection

Seagoing vessels have to undergo regular inspections, which are currently performed manually by ship surveyors. The main cost factor in a ship inspection is to provide access to the different areas of the ship, since the surveyor has to be close to the inspected parts, usually within arm's reach, either to perform a visual analysis or to take thickness measurements. The access to the structural elements in cargo holds, e.g., bulkheads, is normally provided by staging or by 'cherry-picking' cranes. To make ship inspections safer and more cost-efficient, we have introduced new inspection methods, tools, and systems, which have been evaluated in field trials, particularly focusing on cargo holds. More precisely, two magnetic climbing robots and a micro-aerial vehicle, which are able to assist the surveyor during the inspection, are introduced. Since localization of inspection data is mandatory for the surveyor, we also introduce an external localization system that has been verified in field trials, using a climbing inspection robot. Furthermore, the inspection data collected by the robotic systems are organized and handled by a spatial content management system that enables us to compare the inspection data of one survey with those from another, as well as to document the ship inspection when the robot team is used. Image-based defect detection is addressed by proposing an integrated solution for detecting corrosion and cracks. The systems' performance is reported, as well as conclusions on their usability, all in accordance with the output of field trials performed onboard two different vessels under real inspection conditions.

Spectrum of ankylosing spondylitis in Portugal. Development of BASDAI, BASFI, BASMI and mSASSS reference centile charts

The availability of population-specific normative data regarding disease severity measures is essential for patient assessment. The goals of the current study were to characterize the pattern of ankylosing spondylitis (AS) in Portuguese patients and to develop reference centile charts for BASDAI, BASFI, BASMI and mSASSS, the most widely used assessment tools in AS. AS cases were recruited from hospital outpatient clinics, with AS defined according to the modified New York criteria. Demographic and clinical data were recorded. All radiographs were evaluated by two independent experienced readers. Centile charts for BASDAI, BASFI, BASMI and mSASSS were constructed for both genders, using generalized linear models and regression models with duration of disease as independent variable. A total of 369 patients (62.3% male, mean ± (SD) age 45.4 ± 13.2 years, mean ± (SD) disease duration 11.4 ± 10.5 years, 70.7% B27-positive) were included. Family history of AS in a first-degree relative was reported in 17.6% of the cases. Regarding clinical disease pattern, at the time of assessment 42.3% had axial disease, 2.4% peripheral disease, 40.9% mixed disease and 7.1% isolated enthesopatic disease. Anterior uveitis (33.6%) was the most common extra-articular manifestation. The centile charts suggest that females reported greater disease activity and more functional impairment than males but had lower BASMI and mSASSS scores. Data collected through this study provided a demographic and clinical profile of patients with AS in Portugal. The development of centile charts constitutes a useful tool to assess the change of disease pattern over time and in response to therapeutic interventions.

A manager-centred perspective on work-life integration

Decades of research has shown that the uptake of workplace ‘flexibility’ provisions set out in organizational/HR policies rests heavily on the support of line managers. However, the majority of scholarship addressing the intersection of managers’ roles and work-life integration has been employee-centred. That is, the literature primarily situates managers as gatekeepers to the effective implementation of work and family policies as they affect employees or workers, examining their role in, for example, approving requests to adjust or personalise employees’ work schedules; influencing whether employees are cross-trained to undertake the work of others during absences; publicising available policies; and creating norms supporting the use of formal provisions (Ryan & Ernst Kossek, 2008). Managers’ actions are primarily seen as key, contingent phenomena affecting the adoption and diffusion of work-life initiatives in an organization; consequently impacting on the work-life outcomes of subordinate employees (Bardoel, 2003; Gregory & Milner, 2012).

New genetic loci link adipose and insulin biology to body fat distribution

Body fat distribution is a heritable trait and a well-established predictor of adverse metabolic outcomes, independent of overall adiposity. To increase our understanding of the genetic basis of body fat distribution and its molecular links to cardiometabolic traits, here we conduct genome-wide association meta-analyses of traits related to waist and hip circumferences in up to 224,459 individuals. We identify 49 loci (33 new) associated with waist-to-hip ratio adjusted for body mass index (BMI), and an additional 19 loci newly associated with related waist and hip circumference measures (P < 5 × 10−8). In total, 20 of the 49 waist-to-hip ratio adjusted for BMI loci show significant sexual dimorphism, 19 of which display a stronger effect in women. The identified loci were enriched for genes expressed in adipose tissue and for putative regulatory elements in adipocytes. Pathway analyses implicated adipogenesis, angiogenesis, transcriptional regulation and insulin resistance as processes affecting fat distribution, providing insight into potential pathophysiological mechanisms.

Genetic studies of body mass index yield new insights for obesity biology

Obesity is heritable and predisposes to many diseases. To understand the genetic basis of obesity better, here we conduct a genome-wide association study and Metabochip meta-analysis of body mass index (BMI), a measure commonly used to define obesity and assess adiposity, in up to 339,224 individuals. This analysis identifies 97 BMI-associated loci (P < 5 × 10−8), 56 of which are novel. Five loci demonstrate clear evidence of several independent association signals, and many loci have significant effects on other metabolic phenotypes. The 97 loci account for ~2.7% of BMI variation, and genome-wide estimates suggest that common variation accounts for >20% of BMI variation. Pathway analyses provide strong support for a role of the central nervous system in obesity susceptibility and implicate new genes and pathways, including those related to synaptic function, glutamate signalling, insulin secretion/action, energy metabolism, lipid biology and adipogenesis.

Eldercare demands, strain, and work engagement: The moderating role of perceived organizational support

Demographic changes give rise to an increasing number of middle-aged employees providing home-based care to an elderly family member. However, the potentially important role of employees' perceptions of organizational support for eldercare has so far not been investigated. The goal of this study was to examine a stressor–strain–outcome model (Koeske & Koeske, 1993) of eldercare strain as a mediator of the relationship between eldercare demands and caregivers' work engagement. Perceived organizational eldercare support was expected to attenuate the positive relationship between eldercare demands and eldercare strain and to buffer the negative relationship between eldercare strain and work engagement. Results of mediation and moderated mediation analyses with data collected from 147 employees providing eldercare supported the hypotheses. The findings suggest that perceived organizational eldercare support is especially beneficial for employees' work engagement when eldercare demands and strain are high.

What consumers think, do and say about antibiotic use

Objective The objectives of this study were to investigate (1) the attitudes and behaviours of Australian consumers in antibiotic use, and (2) their understanding of antibiotic resistance. Methods Semi-structured interviews were conducted with consumers in May/June 2015. Convenience sampling was used to recruit consumers between 18–54 years old. Thirty-two consumers were interviewed. Transcripts were analysed to identify themes. Lessons Learned Dominant themes for attitudes and behaviours regarding antibiotics were (a) avoidance of antibiotic use unless clinically warranted; (b) antibiotics were useful but “weakened the body”; and (c) use of complementary medicines as adjuncts to antibiotics or to strengthen the immune system. Key information needs were (a) unambiguous instructions from GPs when prescribed antibiotics, to avoid inappropriate medicine-taking behaviour; (b) rationale for antibiotic selection; and (c) treatment duration. Antibiotic resistance was conceptualised in three ways: as a property of the body (body becomes resistant to antibiotics); the medication (antibiotic no longer effective); and the bacteria (bacteria is resistant). Antibiotic resistance was perceived as an issue that would only affect the wider community in the future, although most recognised that it is a current challenge for hospitals. Personal good health and/or avoidance of antibiotics were perceived as insurance against being adversely affected by antibiotic resistance. Implications A structured survey (discrete choice experiment) will be developed from these findings to investigate how consumers trade-off on factors influencing antibiotic use. Public health campaigns promoting conservation of antibiotics can benefit from these findings.