Phage library screening for the rapid identification and in vivo testing of candidate genes for a DNA vaccine against Mycoplasma mycoides subsp. mycoides small colony biotype

A new strategy for rapidly selecting and testing genetic vaccines has been developed, in which a whole genome library is cloned into a bacteriophage λ ZAP Express vector which contains both prokaryotic (Plac) and eukaryotic (PCMV) promoters upstream of the insertion site. The phage library is plated on Escherichia coli cells, immunoblotted, and probed with hyperimmune and/or convalescent-phase antiserum to rapidly identify vaccine candidates. These are then plaque purified and grown as liquid lysates, and whole bacteriophage particles are then used directly to immunize the host, following which PCMV-driven expression of the candidate vaccine gene occurs. In the example given here, a semirandom genome library of the bovine pathogen Mycoplasma mycoides subsp. mycoides small colony (SC) biotype was cloned into λ ZAP Express, and two strongly immunodominant clones, λ-A8 and λ-B1, were identified and subsequently tested for vaccine potential against M. mycoides subsp. mycoides SC biotype-induced mycoplasmemia. Sequencing and immunoblotting indicated that clone λ-A8 expressed an isopropyl-β-d-thiogalactopyranoside (IPTG)-inducible M. mycoides subsp. mycoides SC biotype protein with a 28-kDa apparent molecular mass, identified as a previously uncharacterized putative lipoprotein (MSC_0397). Clone λ-B1 contained several full-length genes from the M. mycoides subsp. mycoides SC biotype pyruvate dehydrogenase region, and two IPTG-independent polypeptides, of 29 kDa and 57 kDa, were identified on immunoblots. Following vaccination, significant anti-M. mycoides subsp. mycoides SC biotype responses were observed in mice vaccinated with clones λ-A8 and λ-B1. A significant stimulation index was observed following incubation of splenocytes from mice vaccinated with clone λ-A8 with whole live M. mycoides subsp. mycoides SC biotype cells, indicating cellular proliferation. After challenge, mice vaccinated with clone λ-A8 also exhibited a reduced level of mycoplasmemia compared to controls, suggesting that the MSC_0397 lipoprotein has a protective effect in the mouse model when delivered as a bacteriophage DNA vaccine. Bacteriophage-mediated immunoscreening using an appropriate vector system offers a rapid and simple technique for the identification and immediate testing of putative candidate vaccines from a variety of pathogens.

Combining passive sampling and toxicity testing for evaluation of mixtures of polar organic chemicals in sewage treatment plant effluent

Effluent from sewage treatment plants has been associated with a range of pollutant effects. Depending on the influent composition and treatment processes the effluent may contain a myriad of different chemicals which makes monitoring very complex. In this study we aimed to monitor relatively polar organic pollutant mixtures using a combination of passive sampling techniques and a set of biochemistry based assays covering acute bacterial toxicity (Microtox™), phytotoxicity (Max-I-PAM assay) and genotoxicity (umuC assay). The study showed that all of the assays were able to detect effects in the samples and allowed a comparison of the two plants as well as a comparison between the two sampling periods. Distinct improvements in water quality were observed in one of the plants as result of an upgrade to a UV disinfection system, which improved from 24× sample enrichment required to induce a 50% response in the Microtox™ assay to 84×, from 30× sample enrichment to induce a 50% reduction in photosynthetic yield to 125×, and the genotoxicity observed in the first sampling period was eliminated. Thus we propose that biochemical assay techniques in combination with time integrated passive sampling can substantially contribute to the monitoring of polar organic toxicants in STP effluents.

Cytotoxicity testing of silver-containing burn treatments using primary and immortal skin cells

A novel burn wound hydrogel dressing has been previously developed which is composed of 2-acrylamido-2-methylpropane sulfonic acid sodium salt with silver nanoparticles (silver AMPS). This study compared the cytotoxicity of this dressing to the commercially available silver products; Acticoat™, PolyMem Silver® and Flamazine™ cream. Human keratinocytes (HaCaT and primary HEK) and normal human fibroblasts (NHF) were exposed to dressings incubated on Nunc™ polycarbonate inserts for 24, 48 and 72h. Four different cytotoxicity assays were performed including; Trypan Blue cell count, MTT, Celltiter-Blue™ and Toluidine Blue surface area assays. The results were expressed as relative cell viability compared to an untreated control. The cytotoxic effects of Acticoat™ and Flamazine™ cream were dependent on exposure time and cell type. After 24h exposure, Acticoat™ and Flamazine™ cream were toxic to all tested cell lines. Surprisingly, HaCaTs treated with Acticoat™ and Flamazine™ had an improved ability to survive at 48 and 72h while HEKs and NHFs had no improvement in survival with any treatment. The novel silver hydrogel and PolyMem Silver® showed low cytotoxicity to all tested cell lines at every time interval and these results support the possibility of using the novel silver hydrogel as a burn wound dressing. Researchers who rely on HaCaT cells as an accurate keratinocyte model should be aware that they can respond differently to primary skin cells.

A frontier functions approach to optimal scales of sustainable production

This paper translates the concepts of sustainable production to three dimensions of economic, environmental and ecological sustainability to analyze optimal production scales by solving optimizing problems. Economic optimization seeks input-output combinations to maximize profits. Environmental optimization searches for input-output combinations that minimize the polluting effects of materials balance on the surrounding environment. Ecological optimization looks for input-output combinations that minimize the cumulative destruction of the entire ecosystem. Using an aggregate space, the framework illustrates that these optimal scales are often not identical because markets fail to account for all negative externalities. Profit-maximizing firms normally operate at the scales which are larger than optimal scales from the viewpoints of environmental and ecological sustainability; hence policy interventions are favoured. The framework offers a useful tool for efficiency studies and policy implication analysis. The paper provides an empirical investigation using a data set of rice farms in South Korea.

Optimal HMM processing of differentially phase modulated communication systems

This paper investigates demodulation of differentially phase modulated signals DPMS using optimal HMM filters. The optimal HMM filter presented in the paper is computationally of order N3 per time instant, where N is the number of message symbols. Previously, optimal HMM filters have been of computational order N4 per time instant. Also, suboptimal HMM filters have be proposed of computation order N2 per time instant. The approach presented in this paper uses two coupled HMM filters and exploits knowledge of ...

Optimal HMM filtering and decision feedback equalisation for differential encoded transmission systems

In this paper conditional hidden Markov model (HMM) filters and conditional Kalman filters (KF) are coupled together to improve demodulation of differential encoded signals in noisy fading channels. We present an indicator matrix representation for differential encoded signals and the optimal HMM filter for demodulation. The filter requires O(N3) calculations per time iteration, where N is the number of message symbols. Decision feedback equalisation is investigated via coupling the optimal HMM filter for estimating the message, conditioned on estimates of the channel parameters, and a KF for estimating the channel states, conditioned on soft information message estimates. The particular differential encoding scheme examined in this paper is differential phase shift keying. However, the techniques developed can be extended to other forms of differential modulation. The channel model we use allows for multiplicative channel distortions and additive white Gaussian noise. Simulation studies are also presented.

Testing the effectiveness of a self-efficacy based exercise intervention for adults with venous leg ulcers : protocol of a randomised controlled trial

Background: Exercise and adequate self-management capacity may be important strategies in the management of venous leg ulcers. However, it remains unclear if exercise improves the healing rates of venous leg ulcers and if a self-management exercise program based on self-efficacy theory is well adhered to. Method/Design: This is a randomised controlled in adults with venous leg ulcers to determine the effectiveness of a self-efficacy based exercise intervention. Participants with venous leg ulcers are recruited from 3 clinical sites in Australia. After collection of baseline data, participants are randomised to either an intervention group or control group. The control group receive usual care, as recommended by evidence based guidelines. The intervention group receive an individualised program of calf muscle exercises and walking. The twelve week exercise program integrates multiple elements, including up to six telephone delivered behavioural coaching and goal setting sessions, supported by written materials, a pedometer and two follow-up booster calls if required. Participants are encouraged to seek social support among their friends, self-monitor their weekly steps and lower limb exercises. The control group are supported by a generic information sheet that the intervention group also receive encouraging lower limb exercises, a pedometer for self-management and phone calls at the same time points as the intervention group. The primary outcome is the healing rates of venous leg ulcers which are assessed at fortnightly clinic appointments. Secondary outcomes, assessed at baseline and 12 weeks: functional ability (range of ankle motion and Tinetti gait and balance score), quality of life and self-management scores. Discussion: This study seeks to address a significant gap in current wound management practice by providing evidence for the effectiveness of a home-based exercise program for adults with venous leg ulcers. Theory-driven, evidence-based strategies that can improve an individual’s exercise self-efficacy and self-management capacity could have a significant impact in improving the management of people with venous leg ulcers. Information gained from this study will provide much needed information on management of this chronic disease to promote health and independence in this population. Trial registration: Australian New Zealand Clinical Trials Registry ACTRN12612000475842 Trial status: Current follow up

Environmental, personal, and genetic determinants of response to vitamin D supplementation in older adults

CONTEXT AND OBJECTIVE: Suboptimal vitamin D status can be corrected by vitamin D supplementation, but individual responses to supplementation vary. We aimed to examine genetic and nongenetic determinants of change in serum 25-hydroxyvitamin D (25(OH)D) after supplementation. DESIGN AND PARTICIPANTS: We used data from a pilot randomized controlled trial in which 644 adults aged 60 to 84 years were randomly assigned to monthly doses of placebo, 30 000 IU, or 60 000 IU vitamin D3 for 12 months. Baseline characteristics were obtained from a self-administered questionnaire. Eighty-eight single-nucleotide polymorphisms (SNPs) in 41 candidate genes were genotyped using Sequenom MassArray technology. Serum 25(OH)D levels before and after the intervention were measured using the Diasorin Liaison platform immunoassay. We used linear regression models to examine associations between genetic and nongenetic factors and change in serum 25(OH)D levels. RESULTS: Supplement dose and baseline 25(OH)D level explained 24% of the variability in response to supplementation. Body mass index, self-reported health status, and ambient UV radiation made a small additional contribution. SNPs in CYP2R1, IRF4, MC1R, CYP27B1, VDR, TYRP1, MCM6, and HERC2 were associated with change in 25(OH)D level, although only CYP2R1 was significant after adjustment for multiple testing. Models including SNPs explained a similar proportion of variability in response to supplementation as models that included personal and environmental factors. CONCLUSION: Stepwise regression analyses suggest that genetic variability may be associated with response to supplementation, perhaps suggesting that some people might need higher doses to reach optimal 25(OH)D levels or that there is variability in the physiologically normal level of 25(OH)D.

Development of realistic design fire time-temperature curves for the testing of cold-formed steel wall system

Fire resistance rating of light gauge steel frame (LSF) wall systems is obtained from fire tests based on the standard fire time-temperature curve. However, fire severity has increased in modern buildings due to higher fuel loads as a result of modern furniture and light weight constructions that make use of thermoplastics materials, synthetic foams and fabrics. Some of these materials are high in calorific values and increase both the spread of fire growth and heat release rate, thus increasing the fire severity beyond that of the standard fire curve. Further, the standard fire curve does not include a decay phase that is present in natural fires. Despite the increasing usage of LSF walls, their behaviour in real building fires is not fully understood. This paper presents the details of a research study aimed at developing realistic design fire curves for use in the fire tests of LSF walls. It includes a review of the characteristics of building fires, previously developed fire time-temperature curves, computer models and available parametric equations. The paper highlights that real building fire time-temperature curves depend on the fuel load representing the combustible building contents, ventilation openings and thermal properties of wall lining materials, and provides suitable values of many required parameters including fuel loads in residential buildings. Finally, realistic design fire time-temperature curves simulating the fire conditions in modern residential buildings are proposed for the testing of LSF walls.

Differential effects of refractive blur on day and night-time driving performance

Purpose To investigate the effect of different levels of refractive blur on real-world driving performance measured under day and nighttime conditions. Methods Participants included 12 visually normal, young adults (mean age = 25.8 ± 5.2 years) who drove an instrumented research vehicle around a 4 km closed road circuit with three different levels of binocular spherical refractive blur (+0.50 diopter sphere [DS], +1.00 DS, +2.00 DS) compared with a baseline condition. The subjects wore optimal spherocylinder correction and the additional blur lenses were mounted in modified full-field goggles; the order of testing of the blur conditions was randomized. Driving performance was assessed in two different sessions under day and nighttime conditions and included measures of road signs recognized, hazard detection and avoidance, gap detection, lane-keeping, sign recognition distance, speed, and time to complete the course. Results Refractive blur and time of day had significant effects on driving performance (P < 0.05), where increasing blur and nighttime driving reduced performance on all driving tasks except gap judgment and lane keeping. There was also a significant interaction between blur and time of day (P < 0.05), such that the effects of blur were exacerbated under nighttime driving conditions; performance differences were evident even for +0.50 DS blur relative to baseline for some measures. Conclusions The effects of blur were greatest under nighttime conditions, even for levels of binocular refractive blur as low as +0.50 DS. These results emphasize the importance of accurate and up-to-date refractive correction of even low levels of refractive error when driving at night.

A multiple case study of NAPLAN numeracy testing of Year 9 students in three Queensland secondary schools

This thesis reports on a multiple case study of the actions of three Queensland secondary schools in the context of Year 9 NAPLAN numeracy testing, focusing on their administrative practices, curriculum, pedagogy and assessment. It was established that schools have found it both challenging and costly to operate in an environment of educational reform generally, and NAPLAN testing in particular. The lack of a common understanding of numeracy and the substantial demands of implementing the Australian Curriculum have impacted on schools' ability to prepare students appropriately for NAPLAN numeracy tests. It was concluded that there is scope for schools to improve their approaches to NAPLAN numeracy testing in a way that maximises learning as well as test outcomes.

Confirmatory factor analysis and invariance testing of the Young Carer of Parents Inventory (YCOPI)

Objective Research into youth caregiving in families where a parent experiences a significant medical condition has been hampered by a lack of contextually sensitive measures of the nature and breadth of young caregiving experiences. This study examined the factor structure and measurement invariance of such a measure called the Young Carer of Parents Inventory (YCOPI; Pakenham et al., 2006) using confirmatory factor analysis across 3 groups of youth. The YCOPI has 2 parts: YCOPI-A with 5 factors assessing caregiving experiences that are applicable to all caregiving contexts; YCOPI-B with 4 factors that tap dimensions related to youth caregiving in the context of parent illness. Design Two samples (ages 9–20 years) were recruited: a community sample of 2,429 youth from which 2 groups were derived (“healthy” family [HF], n = 1760; parental illness [PI], n = 446), and a sample of 130 youth of a parent with multiple sclerosis). Results With some modification, the YCOPI-A demonstrated a replicable factor structure across 3 groups, and exhibited only partial measurement invariance across the HF and PI groups. The impact of assuming full measurement invariance on latent mean differences appeared small, supporting use of the measure in research and applied settings when estimated using latent factors and controlling for measurement invariance. PI youth reported significantly higher scores than did HF youth on all YCOPI-A subscales. The YCOPI-B requires some modifications, and further development work is recommended. Conclusion The factor structure that emerged and the addition of new items constitutes the YCOPI-Revised. Findings support the use of the YCOPI-Revised in research and applied settings.