Isoprene oxidation by the gram-negative model bacterium variovorax sp. WS11

Plant-produced isoprene (2-methyl-1,3-butadiene) represents a significant portion of global volatile organic compound production, equaled only by methane. A metabolic pathway for the degradation of isoprene was first described for the Gram-positive bacterium Rhodococcus sp. AD45, and an alternative model organism has yet to be characterised. Here, we report the characterisation of a novel Gram-negative isoprene-degrading bacterium, Variovorax sp. WS11. Isoprene metabolism in this bacterium involves a plasmid-encoded iso metabolic gene cluster which differs from that found in Rhodococcus sp. AD45 in terms of organisation and regulation. Expression of iso metabolic genes is significantly upregulated by both isoprene and epoxyisoprene. The enzyme responsible for the initial oxidation of isoprene, isoprene monooxygenase, oxidises a wide range of alkene substrates in a manner which is strongly influenced by the presence of alkyl side-chains and differs from other well-characterised soluble diiron monooxygenases according to its response to alkyne inhibitors. This study presents Variovorax sp. WS11 as both a comparative and contrasting model organism for the study of isoprene metabolism in bacteria, aiding our understanding of the conservation of this biochemical pathway across diverse ecological niches

The Role of Harvey-ras in Mouse Skin Tumorigenesis

This thesis outlines two alternative approaches to investigating the role of H-ras in mouse skin tumour development. The first, the in vitro approach utilises a number of unique cell lines representative of a late stage of tumorigenesis, the conversion from a well differentiated squamous carcinoma to a undifferentiated spindle form. Through manipulation of the gene dosage ratio of mutant:normal H-ras in these cell lines, by plasmid transfection and virus infection, it was hoped to provide some clues as to the function of H-ras in this late stage transition event. The second approach involves the technique of gene targeting. The aim of this approach was to create a mouse hemizygous or completely deficient in H-ras and to examine the effects in the context of both chemical carcinogenesis studies and normal mouse development. Initial observations of the squamous/spindle transition event showed that the cell lines representative of both phenotypes were characteristically very different. The squamous cell line B9 produced a well differentiated tumour when injected into nude mice whereas the spindle cell lines A5 and D3 gave rise to aggressive, disorganised tumours with a much reduced latency. Genetic analysis of the cell lines showed that the most dramatic difference between the squamous and spindle derivatives was in the copy number and expression levels of H-ras. Both spindle cell lines A5 and D3 showed an approximate 5-10 fold increase in mutant H-ras, best explained by a genetic amplification mechanism. Experimentally overexpressing mutant H-ras in the squamous cells, and vice versa the levels of normal H-ras in the spindle variants, was unable to mimic the conversion event. However it was found that several A5 and D3 transfectant clones showed a dramatic reduction in tumorigenicity. This correlated not with the levels of the introduced normal H-ras but the absolute levels of endogenous mutant H-ras. Spontaneous loss of mutant copies of H-ras resulted in clones that morphologically resembled spindle cells yet produced tumours with a vastly increased latency period. It seems therefore that in this system mutant H-ras is the critical determinant of tumorigenicity, and relates perhaps to the growth of the tumour cell in vivo, possibly providing the driving force for the conversion. However, alteration of the H-ras gene dosage ratio was not in itself sufficient for the transition of a squamous to a spindle cell carcinoma, implying that another locus as yet unknown must play a role in controlling the epithelial phenotype. Finally, the gene targeting approach, provides a means to examining the earlier stages of mouse skin tumorigenesis, in particular the proposed role of normal H-ras in suppressing the development of papillomas from an initiated cell. For instance, if the consistent duplication of the chromosome carrying the mutant H-ras gene, seen in the development of papillomas, is related to overcoming a suppressive effect of normal H-ras, then carcinogen treatment of a mouse deficient in H-ras may result in a mouse more susceptible to tumorigenesis. Experiments with the gene targeting technique are on going and as yet no targeted ES cell clone has been isolated, but hope remains that this approach will provide new insights into the functions of H-ras both in early tumorigenesis and normal development

Precipitation Hardenable High Temperature Shape Memory Alloy

A composition of the invention is a high temperature shape memory alloy having high work output, and is made from (Ni+Pt+Y),Ti(100-x) wherein x is present in a total amount of 49-55 atomic % Pt is present in a total amount of 10-30 atomic %, Y is one or more of Au, Pd. and Cu and is present in a total amount of 0 to 10 atomic %. The alloy has a matrix phase wherein the total concentration of Ni, Pt, and the one or more of Pd. Au, and Cu is greater than 50 atomic %

Sphingopyxis sp. Strain OPL5, an Isoprene-Degrading Bacterium from the Sphingomonadaceae Family Isolated from Oil Palm Leaves

The volatile secondary metabolite, isoprene, is released by trees to the atmosphere in enormous quantities, where it has important effects on air quality and climate. Oil palm trees, one of the highest isoprene emitters, are increasingly dominating agroforestry over large areas of Asia, with associated uncertainties over their effects on climate. Microbes capable of using isoprene as a source of carbon for growth have been identified in soils and in the tree phyllosphere, and most are members of the Actinobacteria. Here, we used DNA stable isotope probing to identify the isoprene-degrading bacteria associated with oil palm leaves and inhabiting the surrounding soil. Among the most abundant isoprene degraders of the leaf-associated community were members of the Sphingomonadales, although no representatives of this order were previously known to degrade isoprene. Informed by these data, we obtained representatives of the most abundant isoprene degraders in enrichments, including Sphingopyxis strain OPL5 (Sphingomonadales), able to grow on isoprene as the sole source of carbon and energy. Sequencing of the genome of strain OPL5, as well as a novel Gordonia strain, confirmed their pathways of isoprene degradation and broadened our knowledge of the genetic and taxonomic diversity of this important bacterial trait

Inhibition of ammonia monooxygenase from ammonia oxidising archaea by linear and aromatic alkynes

Ammonia monooxygenase (AMO) is a key nitrogen-transforming enzyme belonging to the same copper-dependent membrane monooxygenase family (CuMMO) as the particulate methane monooxygenase (pMMO). The AMO from ammonia-oxidizing archaea (AOA) is very divergent from both the AMO of ammonia-oxidizing bacteria (AOB) and the pMMO from methanotrophs, and little is known about the structure or substrate range of the archaeal AMO. This study compares inhibition by C 2 to C 8 linear 1-alkynes of AMO from two phylogenetically distinct strains of AOA, " Candidatus Nitrosocosmicus franklandus" C13 and " Candidatus Nitrosotalea sinensis" Nd2, with AMO from Nitrosomonas europaea and pMMO from Methylococcus capsulatus (Bath). An increased sensitivity of the archaeal AMO to short-chain-length alkynes (≤C 5) appeared to be conserved across AOA lineages. Similarities in C 2 to C 8 alkyne inhibition profiles between AMO from AOA and pMMO from M. capsulatus suggested that the archaeal AMO has a narrower substrate range than N. europaea AMO. Inhibition of AMO from " Ca Nitrosocosmicus franklandus" and N. europaea by the aromatic alkyne phenylacetylene was also investigated. Kinetic data revealed that the mechanisms by which phenylacetylene inhibits " Ca Nitrosocosmicus franklandus" and N. europaea are different, indicating differences in the AMO active site between AOA and AOB. Phenylacetylene was found to be a specific and irreversible inhibitor of AMO from " Ca Nitrosocosmicus franklandus," and it does not compete with NH 3 for binding at the active site. IMPORTANCE Archaeal and bacterial ammonia oxidizers (AOA and AOB, respectively) initiate nitrification by oxidizing ammonia to hydroxylamine, a reaction catalyzed by ammonia monooxygenase (AMO). AMO enzyme is difficult to purify in its active form, and its structure and biochemistry remain largely unexplored. The bacterial AMO and the closely related particulate methane monooxygenase (pMMO) have a broad range of hydrocarbon cooxidation substrates. This study provides insights into the AMO of previously unstudied archaeal genera, by comparing the response of the archaeal AMO, a bacterial AMO, and pMMO to inhibition by linear 1-alkynes and the aromatic alkyne, phenylacetylene. Reduced sensitivity to inhibition by larger alkynes suggests that the archaeal AMO has a narrower hydrocarbon substrate range than the bacterial AMO, as previously reported for other genera of AOA. Phenylacetylene inhibited the archaeal and bacterial AMOs at different thresholds and by different mechanisms of inhibition, highlighting structural differences between the two forms of monooxygenase

Water consumption, not expectancies about water consumption, affects cognitive performance in adults

Research has shown that water supplementation positively affects cognitive performance in children and adults. The present study considered whether this could be a result of expectancies that individuals have about the effects of water on cognition. Forty seven participants were recruited and told the study was examining the effects of repeated testing on cognitive performance. They were assigned either to a condition in which positive expectancies about the effects of drinking water were induced, or a control condition in which no expectancies were induced. Within these groups, approximately half were given a drink of water, while the remainder were not. Performance on a thirst scale, letter cancellation, digit span forwards and backwards and a simple reaction time task was assessed at baseline (before the drink) and 20 minutes and 40 minutes after water consumption. Effects of water, but not expectancy, were found on subjective thirst ratings and letter cancellation task performance, but not on digit span or reaction time. This suggests that water consumption effects on letter cancellation are due to the physiological effects of water, rather than expectancies about the effects of drinking water

Randomized trial of an education and support intervention to preventreadmission of patients with heart failure

AbstractObjectivesWe determined the effect of a targeted education and support intervention on the rate of readmission or death and hospital costs in patients with heart failure (HF).BackgroundDisease management programs for patients with HF including medical components may reduce readmissions by 40% or more, but the value of an intervention focused on education and support is not known.MethodsWe conducted a prospective, randomized trial of a formal education and support intervention on one-year readmission or mortality and costs of care for patients hospitalized with HF.ResultsAmong the 88 patients (44 intervention and 44 control) in the study, 25 patients (56.8%) in the intervention group and 36 patients (81.8%) in the control group had at least one readmission or died during one-year follow-up (relative risk = 0.69, 95% confidence interval [CI]: 0.52, 0.92; p = 0.01). The intervention was associated with a 39% decrease in the total number of readmissions (intervention group: 49 readmissions; control group: 80 readmissions, p = 0.06). After adjusting for clinical and demographic characteristics, the intervention group had a significantly lower risk of readmission compared with the control group (hazard ratio = 0.56, 95% CI: 0.32, 0.96; p = 0.03) and hospital readmission costs of $7,515 less per patient.ConclusionsA formal education and support intervention substantially reduced adverse clinical outcomes and costs for patients with HF

Poplar phyllosphere harbors disparate isoprene-degrading bacteria

The climate-active gas isoprene (2-methyl-1,3-butadiene) is released to the atmosphere in huge quantities, almost equaling that of methane, yet we know little about the biological cycling of isoprene in the environment. Although bacteria capable of growth on isoprene as the sole source of carbon and energy have previously been isolated from soils and sediments, no microbiological studies have targeted the major source of isoprene and examined the phyllosphere of isoprene-emitting trees for the presence of degraders of this abundant carbon source. Here, we identified isoprene-degrading bacteria in poplar tree-derived microcosms by DNA stable isotope probing. The genomes of isoprene-degrading taxa were reconstructed, putative isoprene metabolic genes were identified, and isoprene-related gene transcription was analyzed by shotgun metagenomics and metatranscriptomics. Gram-positive bacteria of the genus Rhodococcus proved to be the dominant isoprene degraders, as previously found in soil. However, a wider diversity of isoprene utilizers was also revealed, notably Variovorax, a genus not previously associated with this trait. This finding was confirmed by expression of the isoprene monooxygenase from Variovorax in a heterologous host. A Variovorax strain that could grow on isoprene as the sole carbon and energy source was isolated. Analysis of its genome confirmed that it contained isoprene metabolic genes with an identical layout and high similarity to those identified by DNA-stable isotope probing and metagenomics. This study provides evidence of a wide diversity of isoprene-degrading bacteria in the isoprene-emitting tree phyllosphere and greatly enhances our understanding of the biodegradation of this important metabolite and climate-active gas

Use of measurement theory for operationalization and quantification of psychological constructs in systems dynamics modelling

The analytical tools available to social scientists have traditionally been adapted from tools originally designed for analysis of natural science phenomena. This article discusses the applicability of systems dynamics - a qualitative based modelling approach, as a possible analysis and simulation tool that bridges the gap between social and natural sciences. After a brief overview of the systems dynamics modelling methodology, the advantages as well as limiting factors of systems dynamics to the potential applications in the field of social sciences and human interactions are discussed. The issues arise with regards to operationalization and quantification of latent constructs at the simulation building stage of the systems dynamics methodology and measurement theory is proposed as a ready and waiting solution to the problem of dynamic model calibration, with a view of improving simulation model reliability and validity and encouraging the development of standardised, modular system dynamics models that can be used in social science research

A systematic review of school-based sexual health interventions to prevent STI/HIV in sub-Saharan Africa

Background The HIV/AIDS epidemic remains of global significance and there is a need to target (a) the adolescent age-groups in which most new infections occur; and (b) sub-Saharan Africa where the greatest burden of the epidemic lies. A focused systematic review of school-based sexual health interventions in sub-Saharan Africa to prevent HIV/AIDS and Sexually Transmitted Infections (STI) in this age group was therefore conducted. Methods Searches were conducted in Medline, Embase, Cinahl and PsychINFO according to agreed a priori criteria for studies published between 1986 and 2006. Further searches were conducted in UNAIDS and WHO (World Health Organization) websites, and 'Google'. Relevant journals were hand-searched and references cited in identified articles were followed up. Data extraction and quality assessment was carried out on studies selected for full text appraisal, and results were analysed and presented in narrative format. Results Some 1,020 possible titles and abstracts were found, 23 full text articles were critically appraised, and 12 articles (10 studies) reviewed, reflecting the paucity of published studies conducted relative to the magnitude of the HIV epidemic in sub-Saharan Africa. Knowledge and attitude-related outcomes were the most associated with statistically significant change. Behavioural intentions were more difficult to change and actual behaviour change was least likely to occur. Behaviour change in favour of abstinence and condom use appeared to be greatly influenced by pre-intervention sexual history. Conclusion There is a great need in sub-Saharan Africa for well-evaluated and effective school-based sexual health interventions