Structural basis for divergent and convergent evolution of catalytic machineries in plant aromatic amino acid decarboxylase proteins.

Radiation of the plant pyridoxal 5'-phosphate (PLP)-dependent aromatic l-amino acid decarboxylase (AAAD) family has yielded an array of paralogous enzymes exhibiting divergent substrate preferences and catalytic mechanisms. Plant AAADs catalyze either the decarboxylation or decarboxylation-dependent oxidative deamination of aromatic l-amino acids to produce aromatic monoamines or aromatic acetaldehydes, respectively. These compounds serve as key precursors for the biosynthesis of several important classes of plant natural products, including indole alkaloids, benzylisoquinoline alkaloids, hydroxycinnamic acid amides, phenylacetaldehyde-derived floral volatiles, and tyrosol derivatives. Here, we present the crystal structures of four functionally distinct plant AAAD paralogs. Through structural and functional analyses, we identify variable structural features of the substrate-binding pocket that underlie the divergent evolution of substrate selectivity toward indole, phenyl, or hydroxyphenyl amino acids in plant AAADs. Moreover, we describe two mechanistic classes of independently arising mutations in AAAD paralogs leading to the convergent evolution of the derived aldehyde synthase activity. Applying knowledge learned from this study, we successfully engineered a shortened benzylisoquinoline alkaloid pathway to produce (S)-norcoclaurine in yeast. This work highlights the pliability of the AAAD fold that allows change of substrate selectivity and access to alternative catalytic mechanisms with only a few mutations.

Making spatial-temporal marine ecosystem modelling better - A perspective.

Marine Ecosystem Models (MEMs) provide a deeper understanding of marine ecosystem dynamics. The United Nations Decade of Ocean Science for Sustainable Development has highlighted the need to deploy these complex mechanistic spatial-temporal models to engage policy makers and society into dialogues towards sustainably managed oceans. From our shared perspective, MEMs remain underutilized because they still lack formal validation, calibration, and uncertainty quantifications that undermines their credibility and uptake in policy arenas. We explore why these shortcomings exist and how to enable the global modelling community to increase MEMs' usefulness. We identify a clear gap between proposed solutions to assess model skills, uncertainty, and confidence and their actual systematic deployment. We attribute this gap to an underlying factor that the ecosystem modelling literature largely ignores: technical issues. We conclude by proposing a conceptual solution that is cost-effective, scalable and simple, because complex spatial-temporal marine ecosystem modelling is already complicated enough.

Production of nonnatural straight-chain amino acid 6-aminocaproate via an artificial iterative carbon-chain-extension cycle.

Bioplastics produced from microbial source are promising green alternatives to traditional petrochemical-derived plastics. Nonnatural straight-chain amino acids, especially 5-aminovalerate, 6-aminocaproate and 7-aminoheptanoate are potential monomers for the synthesis of polymeric bioplastics as their primary amine and carboxylic acid are ideal functional groups for polymerization. Previous pathways for 5-aminovalerate and 6-aminocaproate biosynthesis in microorganisms are derived from L-lysine catabolism and the citric acid cycle, respectively. Here, we show the construction of an artificial iterative carbon-chain-extension cycle in Escherichia coli for simultaneous production of a series of nonnatural amino acids with varying chain length. Overexpression of L-lysine α-oxidase in E. coli yields 2-keto-6-aminocaproate (2K6AC) as a non-native substrate for the artificial iterative carbon-chain-extension cycle. The chain-extended α-ketoacid products are decarboxylated and oxidized by an α-ketoacid decarboxylase and an aldehyde dehydrogenase, respectively, to yield their corresponding nonnatural straight-chain amino acids. The engineered system demonstrated simultaneous in vitro production of 99.16 mg/L of 5-aminovalerate, 46.96 mg/L of 6-aminocaproate and 4.78 mg/L of 7-aminoheptanoate after 8 h of enzyme catalysis starting from 2K6AC as the substrate. Furthermore, simultaneous production of 2.15 g/L of 5-aminovalerate, 24.12 mg/L of 6-aminocaproate and 4.74 mg/L of 7-aminoheptanoate was achieved in engineered E. coli. This work illustrates a promising metabolic-engineering strategy to access other medium-chain organic acids with -NH2, -SCH3, -SOCH3, -SH, -COOH, -COH, or -OH functional groups through carbon-chain-elongation chemistry.

Referral of patients to diabetes prevention programmes from community campaigns and general practices: mixed-method evaluation using the RE-AIM framework and Normalisation Process Theory.

BACKGROUND: Each year around 5-10% of people with non-diabetic hyperglycaemia will develop type 2 diabetes mellitus. Diabetes prevention is a national and global public health concern. Diabetes Prevention Programmes, which seek to identify at-risk individuals and support entry to health improvement initiatives, recognise that enhanced identification and referral of at-risk individuals is required within primary care and beyond, through community-focused prevention approaches. We report an evaluation of a demonstrator site for the NHS Diabetes Prevention Programme in the UK, which piloted an enhanced Primary Care referral programme (sampling from patients identified as at-risk from general practice databases) and a Community identification programme (sampling from the general population through opportunistic identification in community locations) in an effort to maximise participation in prevention services. METHODS: We used mixed-methods evaluation to assess the impact of the two referral routes on participation in the Diabetes Prevention Programmes in line with the RE-AIM Framework (Reach, Effectiveness, Adoption, Implementation and Maintenance). Individual level patient data was descriptively analysed to assess identifications and eligible referrals made in each route. Semi-structured interviews conducted with referral staff and key stakeholders were analysed using thematic analysis and informed by Normalisation Process Theory. RESULTS: The nurse facilitated primary care referral route provided 88% of all referrals to the telephone DPP, compared to the community referral route which provided 5%, and the proportion joining the programme was higher among primary care referrals (45%) than community referrals (22%), and retention rates were higher (73% compared to 50%). The nurse-facilitated route integrated more easily into existing clinical processes. The community programme was impeded by a lack of collaborative inter-agency working which obscured the intended focus on high-risk populations despite conversion rates (numbers identified at risk who entered prevention programmes) being highest in areas of high deprivation. CONCLUSIONS: The study demonstrates the interaction of components, with effective Adoption and Implementation necessary to support Reach. The NPT analysis demonstrated the importance of consensus around not only the need for such programmes but agreement on how they can be delivered. Future programmes should support inter-agency communication and collaboration, and focus identification efforts on areas of high-risk.

Getting messier with TIDieR: embracing context and complexity in intervention reporting.

BACKGROUND: The Template for Intervention Description and Replication (TIDieR) checklist and guide was developed by an international team of experts to promote full and accurate description of trial interventions. It is now widely used in health research. The aim of this paper is to describe the experience of using TIDieR outside of trials, in a range of applied health research contexts, and make recommendations on its usefulness in such settings. MAIN BODY: We used the TIDieR template for intervention description in six applied health research projects. The six cases comprise a diverse sample in terms of clinical problems, population, settings, stage of intervention development and whether the intervention was led by researchers or the service deliverers. There was also variation in how the TIDieR description was produced in terms of contributors and time point in the project. Researchers involved in the six cases met in two workshops to identify issues and themes arising from their experience of using TIDieR. We identified four themes which capture the difficulties or complexities of using TIDieR in applied health research: (i) fidelity and adaptation: all aspects of an intervention can change over time; (ii) voice: the importance of clarity on whose voice the TIDieR description represents; (iii) communication beyond the immediate context: the usefulness of TIDieR for wider dissemination and sharing; (iv) the use of TIDieR as a research tool. CONCLUSION: We found TIDieR to be a useful tool for applied research outside the context of clinical trials and we suggest four revisions or additions to the original TIDieR which would enable it to better capture these complexities in applied health research: An additional item, 'voice' conveys who was involved in preparing the TIDieR template, such as researchers, service users or service deliverers. An additional item, 'stage of implementation' conveys what stage the intervention has reached, using a continuum of implementation research suggested by the World Health Organisation. A new column, 'modification' reminds authors to describe modifications to any item in the checklist. An extension of the 'how well' item encourages researchers to describe how contextual factors affected intervention delivery.

A qualitative study of professional stakeholders' perceptions about the implementation of a stepped care pain platform for people experiencing chronic widespread pain.

BACKGROUND: Chronic widespread pain (CWP) is a major public health problem. Many people experiencing CWP experience mental health problems such as anxiety or depression. Complete relief of skeletal and body pain symptoms is unlikely but with appropriate treatment the impact upon quality of life, functioning and mental health symptoms can be reduced. Cognitive behavioural therapy (CBT) is widely used for a range of health conditions and can have short and long-term improvements in patients with CWP. This research aimed to explore, from a professional stakeholder perspective, the implementation of a local Pain Platform offering a stepped care approach for interventions including telephone delivered CBT (T-CBT). METHODS: Fourteen professional stakeholders holding various roles across primary and secondary care services within the Pain Platform took part in semi-structured interviews. Their views and experiences of the implementation of the Pain Platform were explored. Interviews were recorded, transcribed verbatim and analysed according to Normalisation Process Theory (NPT). RESULTS: Professional stakeholders were positive about the Pain Platform and its potential to overcome previously identified existing access issues to psychological interventions for CWP patients. It was considered a valuable part of ensuring that patients' preferences and needs are more readily addressed. In some circumstances, however, introducing psychological interventions to patients was considered challenging and the introduction of new referral processes was raised concerns. To ensure sustainability more work is required to reduce professional isolation and ensure efficient referral procedures between primary and secondary care services are established to reduce concerns over issues related to clinical governance and potential risk to patient. CONCLUSIONS: The findings provide professional insight into the key challenges of introducing a Pain Platform incorporating psychological support across primary and secondary care services within a local service. These included development of sustainable procedures and closer working relationships. Areas requiring future development are identified.

Transcriptomes of Plant Gametophytes Have a Higher Proportion of Rapidly Evolving and Young Genes than Sporophytes.

Reproductive traits in plants tend to evolve rapidly due to various causes that include plant-pollinator coevolution and pollen competition, but the genomic basis of reproductive trait evolution is still largely unknown. To characterize evolutionary patterns of genome wide gene expression in reproductive tissues in the gametophyte and to compare them to developmental stages of the sporophyte, we analyzed evolutionary conservation and genetic diversity of protein-coding genes using microarray-based transcriptome data from three plant species, Arabidopsis thaliana, rice (Oryza sativa), and soybean (Glycine max). In all three species a significant shift in gene expression occurs during gametogenesis in which genes of younger evolutionary age and higher genetic diversity contribute significantly more to the transcriptome than in other stages. We refer to this phenomenon as "evolutionary bulge" during plant reproductive development because it differentiates the gametophyte from the sporophyte. We show that multiple, not mutually exclusive, causes may explain the bulge pattern, most prominently reduced tissue complexity of the gametophyte, a varying extent of selection on reproductive traits during gametogenesis as well as differences between male and female tissues. This highlights the importance of plant reproduction for understanding evolutionary forces determining the relationship of genomic and phenotypic variation in plants.

Involving healthcare professionals and family carers in setting research priorities for end-of-life care.

It is important to ensure regional variances are considered when setting future end-of-life research priorities, given the differing demographics and service provision. This project sought to identify end-of-life research priorities within Greater Manchester (United Kingdom). Following an initial scoping exercise, six topics within the 10 national priorities outlined by The Palliative and end-of-life care Priority Setting Partnership were selected for exploration. A workshop involving 32 healthcare professionals and a consultation process with 26 family carers was conducted. Healthcare professionals and carers selected and discussed the topics important to them. The topics selected most frequently by both healthcare professionals and carers were 'Access to 24 hour care', 'Planning end-of-life care in advance' and 'Staff and carer education'. Healthcare professionals also developed research questions for their topics of choice which were refined to incorporate carers' views. These questions are an important starting point for future end-of-life research within Greater Manchester.

The role of boundary spanners in delivering collaborative care: a process evaluation.

BACKGROUND: On average, people with schizophrenia and psychosis die 13-30 years sooner than the general population (World Psychiatry 10 (1):52-77, 2011). Mental and physical health care is often provided by different organisations, different practitioners and in different settings which makes collaborative care difficult. Research is needed to understand and map the impact of new collaborative ways of working at the primary/secondary care interface (PloS One 7 (5); e36468). The evaluation presented in this paper was designed to explore the potential of a Community and Physical Health Co-ordinator role (CPHC) (CPHCs were previously Care Co-ordinators within the Community Mental Health Team, Community in the title CPHC refers to Community Mental Health) and Multi-Disciplinary Team (MDT) meetings across primary and community care, with the aim of improving collaboration of mental and physical health care for service users with Severe Mental Illness (SMI). METHODS: Data collection took place across five general practices (GPs) and a Community Mental Health Team (CMHT) in the Northwest of England, as part of a process evaluation. Semi-structured interviews were conducted with a purposive sample of GP staff (n= 18) and CMHT staff (n=4), a focus group with CMHT staff (n=8) and a survey completed by 13 CMHT staff, alongside cardiovascular risk data and MDT actions. Framework analysis was used to manage and interpret data. RESULTS: The results from the evaluation demonstrate that a CPHC role and MDT meetings are effective mechanisms for improving the collaboration and co-ordination of physical health care for SMI service users. The findings highlight the importance of embedding and supporting the CPHC role, with an emphasis on protected time and continuing professional roles and integrating multiple perspectives through MDT meetings. Considering the importance of physical health care for SMI service users and the complex environment, these are important findings for practitioners, researchers and policy makers in the field of primary care and mental health. CONCLUSION: There is an increasing focus on integration and collaborative working to ensure the delivery of quality care across the whole patient pathway, with a growing need for professionals to work together across service and professional boundaries. The introduction of a two pronged approach to collaboration has shown some important improvements in the management of physical health care for service users with SMI.

Facilitating large-scale implementation of evidence based health care: insider accounts from a co-operative inquiry.

BACKGROUND: Facilitators are known to be influential in the implementation of evidence-based health care (EBHC). However, little evidence exists on what it is that they do to support the implementation process. This research reports on how knowledge transfer associates (KTAs) working as part of the UK National Institute for Health Research 'Collaboration for Leadership in Applied Health Research and Care' for Greater Manchester (GM CLAHRC) facilitated the implementation of EBHC across several commissioning and provider health care agencies. METHODS: A prospective co-operative inquiry with eight KTAs was carried out comprising of 11 regular group meetings where they reflected critically on their experiences. Twenty interviews were also conducted with other members of the GM CLAHRC Implementation Team to gain their perspectives of the KTAs facilitation role and process. RESULTS: There were four phases to the facilitation of EBHC on a large scale: (1) Assisting with the decision on what EBHC to implement, in this phase, KTAs pulled together people and disparate strands of information to facilitate a decision on which EBHC should be implemented; (2) Planning of the implementation of EBHC, in which KTAs spent time gathering additional information and going between key people to plan the implementation; (3) Coordinating and implementing EBHC when KTAs recruited general practices and people for the implementation of EBHC; and (4) Evaluating the EBHC which required the KTAs to set up (new) systems to gather data for analysis. Over time, the KTAs demonstrated growing confidence and skills in aspects of facilitation: research, interpersonal communication, project management and change management skills. CONCLUSION: The findings provide prospective empirical data on the large scale implementation of EBHC in primary care and community based organisations focusing on resources and processes involved. Detailed evidence shows facilitation is context dependent and that 'one size does not fits all'. Co-operative inquiry was a useful method to enhance KTAs learning. The evidence shows that facilitators need tailored support and education, during the process of implementation to provide them with a well-rounded skill-set. Our study was not designed to demonstrate how facilitators contribute to patient health outcomes thus further prospective research is required.

Biochemical evaluation of the decarboxylation and decarboxylation-deamination activities of plant aromatic amino acid decarboxylases.

Plant aromatic amino acid decarboxylase (AAAD) enzymes are capable of catalyzing either decarboxylation or decarboxylation-deamination on various combinations of aromatic amino acid substrates. These two different activities result in the production of arylalkylamines and the formation of aromatic acetaldehydes, respectively. Variations in product formation enable individual enzymes to play different physiological functions. Despite these catalytic variations, arylalkylamine and aldehyde synthesizing AAADs are indistinguishable without protein expression and characterization. In this study, extensive biochemical characterization of plant AAADs was performed to identify residues responsible for differentiating decarboxylation AAADs from aldehyde synthase AAADs. Results demonstrated that a tyrosine residue located on a catalytic loop proximal to the active site of plant AAADs is primarily responsible for dictating typical decarboxylase activity, whereas a phenylalanine at the same position is primarily liable for aldehyde synthase activity. Mutagenesis of the active site phenylalanine to tyrosine in Arabidopsis thaliana and Petroselinum crispum aromatic acetaldehyde synthases primarily converts the enzymes activity from decarboxylation-deamination to decarboxylation. The mutation of the active site tyrosine to phenylalanine in the Catharanthus roseus and Papaver somniferum aromatic amino acid decarboxylases changes the enzymes decarboxylation activity to a primarily decarboxylation-deamination activity. Generation of these mutant enzymes enables the production of unusual AAAD enzyme products including indole-3-acetaldehyde, 4-hydroxyphenylacetaldehyde, and phenylethylamine. Our data indicates that the tyrosine and phenylalanine in the catalytic loop region could serve as a signature residue to reliably distinguish plant arylalkylamine and aldehyde synthesizing AAADs. Additionally, the resulting data enables further insights into the mechanistic roles of active site residues.

Diverse functional evolution of serine decarboxylases: identification of two novel acetaldehyde synthases that uses hydrophobic amino acids as substrates.

BACKGROUND: Type II pyridoxal 5'-phosphate decarboxylases are an important group of phylogenetically diverse enzymes involved in amino acid metabolism. Within plants, this group of enzymes is represented by aromatic amino acid decarboxylases, glutamate decarboxylases and serine decarboxylases. Additional evolutionary divergence of plant aromatic amino acid decarboxylases has resulted in further subcategories with distinct substrate specificities and enzymatic activities. Despite shared homology, no such evolutionary divergence has been characterized within glutamate decarboxylases or serine decarboxylases (SDC). RESULTS: Comparative analysis of two previously characterized serine decarboxylase-like (SDC-like) enzymes demonstrates distinct substrate specificities despite their highly conserved primary sequence. The alternate substrate preference of these homologous SDC-like proteins indicated that functional divergence might have occurred with in SDC-like proteins. In an effort to identify additional SDC-like functional divergence, two uncharacterized SDC-like enzymes were recombinantly expressed and characterized. CONCLUSIONS: An extensive biochemical analysis of two serine decarboxylases-like recombinant proteins led to an interesting discovery; both proteins catalyze the formation of acetaldehyde derivatives from select hydrophobic amino acids substrates. Specifically, Medicago truncatula [GenBank: XP_003592128] and Cicer arietinum [GenBank: XP_004496485] catalyze the decarboxylation and oxidative deamination of phenylalanine, methionine, leucine and tryptophan to generate their corresponding acetaldehydes. The promiscuous aldehyde synthase activity of these proteins yields novel products of 4-(methylthio) butanal, 3-methylbutanal (isovaleraldehyde) and indole-3-acetaldehyde from methionine, leucine and tryptophan respectively. A comparative biochemical analysis of the Medicago truncatula and Cicer arietinum enzymes against two previously characterized SDC-like enzymes further emphasizes the unusual substrate specificity and activity of these novel aldehyde synthases. Due to the strong substrate preference towards phenylalanine, it is likely that both enzymes function as phenylacetaldehyde synthesis in vivo. However, due to their significant sequence divergence and unusual substrate promiscuity these enzymes are functionally and evolutionary divergent from canonical phenylacetaldehyde synthesis enzymes. This work further elaborates on the functional complexity of plant type II PLP decarboxylases and their roles in secondary metabolite biosynthesis.

Mechanism of cysteine-dependent inactivation of aspartate/glutamate/cysteine sulfinic acid α-decarboxylases.

Animal aspartate decarboxylase (ADC), glutamate decarboxylase (GDC) and cysteine sulfinic acid decarboxylase (CSADC) catalyze the decarboxylation of aspartate, glutamate and cysteine sulfinic acid to ß-alanine, γ-aminobutyric acid and hypotaurine, respectively. Each enzymatic product has been implicated in different physiological functions. These decarboxylases use pyridoxal 5-phosphate (PLP) as cofactor and share high sequence homology. Analysis of the activity of ADC in the presence of different amino determined that beta-alanine production from aspartate was diminished in the presence of cysteine. Comparative analysis established that cysteine also inhibited GDC and CSADC in a concentration-dependent manner. Spectral comparisons of free PLP and cysteine, together with ADC and cysteine, result in comparable spectral shifts. Such spectral shifts indicate that cysteine is able to enter the active site of the enzyme, interact with the PLP-lysine internal aldimine, form a cysteine-PLP aldimine and undergo intramolecular nucleophilic cyclization through its sulfhydryl group, leading to irreversible ADC inactivation. Cysteine is the building block for protein synthesis and a precursor of cysteine sulfinic acid that is the substrate of CSADC and therefore is present in many cells, but the presence of cysteine (at comparable concentrations to their natural substrates) apparently could severely inhibit ADC, CSADC and GDC activity. This raises an essential question as to how animal species prevent these enzymes from cysteine-mediated inactivation. Disorders of cysteine metabolism have been implicated in several neurodegenerative diseases. The results of our study should promote research in terms of mechanism by which animals maintain their cysteine homeostasis and possible relationship of cysteine-mediated GDC and CSADC inhibition in neurodegenerative disease development.

Counter culture: causes, extent and solutions of systematic bias in the analysis of behavioural counts.

We often quantify the rate at which a behaviour occurs by counting the number of times it occurs within a specific, short observation period. Measuring behaviour in such a way is typically unavoidable but induces error. This error acts to systematically reduce effect sizes, including metrics of particular interest to behavioural and evolutionary ecologists such as R2 , repeatability (intra-class correlation, ICC) and heritability. Through introducing a null model, the Poisson process, for modelling the frequency of behaviour, we give a mechanistic explanation of how this problem arises and demonstrate how it makes comparisons between studies and species problematic, because the magnitude of the error depends on how frequently the behaviour has been observed as well as how biologically variable the behaviour is. Importantly, the degree of error is predictable and so can be corrected for. Using the example of parental provisioning rate in birds, we assess the applicability of our null model for modelling the frequency of behaviour. We then survey recent literature and demonstrate that the error is rarely accounted for in current analyses. We highlight the problems that arise from this and provide solutions. We further discuss the biological implications of deviations from our null model, and highlight the new avenues of research that they may provide. Adopting our recommendations into analyses of behavioural counts will improve the accuracy of estimated effect sizes and allow meaningful comparisons to be made between studies.

Biochemical evaluation of a parsley tyrosine decarboxylase results in a novel 4-hydroxyphenylacetaldehyde synthase enzyme.

Plant aromatic amino acid decarboxylases (AAADs) are effectively indistinguishable from plant aromatic acetaldehyde syntheses (AASs) through primary sequence comparison. Spectroscopic analyses of several characterized AASs and AAADs were performed to look for absorbance spectral identifiers. Although this limited survey proved inconclusive, the resulting work enabled the reevaluation of several characterized plant AAS and AAAD enzymes. Upon completion, a previously reported parsley AAAD protein was demonstrated to have AAS activity. Substrate specificity tests demonstrate that this novel AAS enzyme has a unique substrate specificity towards tyrosine (km 0.46mM) and dopa (km 1.40mM). Metabolite analysis established the abundance of tyrosine and absence of dopa in parsley extracts. Such analysis indicates that tyrosine is likely to be the sole physiological substrate. The resulting information suggests that this gene is responsible for the in vivo production of 4-hydroxyphenylacetaldehyde (4-HPAA). This is the first reported case of an AAS enzyme utilizing tyrosine as a primary substrate and the first report of a single enzyme capable of producing 4-HPAA from tyrosine.

A Comparison of Different Textured and Non-Textured Anti-Reflective Coatings for Planar Monolithic Silicon-Perovskite Tandem Solar Cells.

Multijunction solar cells offer a route to exceed the Shockley-Queisser limit for single-junction devices. In a few short years, silicon-perovskite tandems have significantly passed the efficiency of the best silicon single-junction cells. For scalable solution processing of silicon-perovskite tandem devices, with the avoidance of vacuum processing steps, a flat silicon sub-cell is normally required. This results in a flat top surface that can lead to higher optical reflection losses than conformal deposition on textured silicon bottom cells. To overcome this, textured anti-reflective coatings (ARCs) can be used on top of the finished cell, with textured polydimethylsiloxane (PDMS), a promising candidate. In this work, we vary the texture geometry and film thickness of PDMS anti-reflective foils to understand the effect of these parameters on reflectance of the foil. The best film is selected, and anti-reflective performance is compared with two common planar ARCs-lithium fluoride (LiF) and magnesium fluoride (MgF2) showing considerable reduction in reflectance for a non-textured silicon-perovskite tandem cell. The application of a PDMS film is shown to give a 3-5% increase in integrated J SC in each sub-cell of a silicon-perovskite tandem structure.

Distribution of ETBE-degrading microorganisms and functional capability in groundwater, and implications for characterising aquifer ETBE biodegradation potential.

Microbes in aquifers are present suspended in groundwater or attached to the aquifer sediment. Groundwater is often sampled at gasoline ether oxygenate (GEO)-impacted sites to assess the potential biodegradation of organic constituents. However, the distribution of GEO-degrading microorganisms between the groundwater and aquifer sediment must be understood to interpret this potential. In this study, the distribution of ethyl tert-butyl ether (ETBE)-degrading organisms and ETBE biodegradation potential was investigated in laboratory microcosm studies and mixed groundwater-aquifer sediment samples obtained from pumped monitoring wells at ETBE-impacted sites. ETBE biodegradation potential (as determined by quantification of the ethB gene) was detected predominantly in the attached microbial communities and was below detection limit in the groundwater communities. The copy number of ethB genes varied with borehole purge volume at the field sites. Members of the Comamonadaceae and Gammaproteobacteria families were identified as responders for ETBE biodegradation. However, the detection of the ethB gene is a more appropriate function-based indicator of ETBE biodegradation potential than taxonomic analysis of the microbial community. The study shows that a mixed groundwater-aquifer sediment (slurry) sample collected from monitoring wells after minimal purging can be used to assess the aquifer ETBE biodegradation potential at ETBE-release sites using this function-based concept.

The Skinner Burial of Ontario, Canada, and the Question of Paget's Disease in the Americas.

PURPOSE: To examine a possible case of Paget's disease of bone (PDB) in an Indigenous pre-contact male from Canada, individual D of the Skinner site in Ontario. METHODS: Radiographs, CT scan and histological analysis. RESULTS: The histological analysis revealed the mosaic pattern that characterizes PDB. CT scans show advanced sclerosis of the cranium and a diminished diplÓ§e with osteolytic lesions. CONCLUSIONS: The pathological features that have been identified are collectively characteristic of PDB. SIGNIFICANCE: The Skinner case advances our understanding of the global history and distribution of PDB. LIMITATIONS OF STUDY: Only two New World cases have been identified and neither has been studied in sufficient detail. SUGGESTIONS FOR FUTURE RESEARCH: The older individuals in precolonial New World skeletal series should be given CT scans, which are non-intrusive, to be followed by histological and genetic analyses when indicated.

The behavioural response of European seabass (Dicentrarchus labrax) to chlorinated seawater effluents.

Seawater chlorination is widely used for coastal, marine industries for the prevention of fouling. Using a choice chamber system, we investigated the influence of chlorinated seawater at typical concentrations occurring near chlorinated cooling water discharges, on the behaviour of juvenile seabass (Dicentrachus labrax). These studies showed that there was evidence of an effect of chlorination, with models predicting 22% fewer fish present in the chlorine dosed chamber compared to the undosed chamber in all control runs (mean number of fish in the dosed chamber in all control runs was 2.23 in comparison to 1.74 when chlorine was present). These data suggest that when food is available D. labrax will enter areas that are at mean chlorination levels of 0.04 mg l-1, which are typical close to the discharge of power station cooling water but are more likely to move away once the food supply is reduced.

Imine chemistry in plant metabolism.

Imine chemistry represents an important category of chemical reactions involved in the biosynthesis of plant natural products, ranging from the newly discovered mobile defense hormone N-hydroxy-pipecolic acid to the red-to-yellow tyrosine-derived betalain pigments. Spontaneous imine formation reactions have also served as the basis for the evolution of numerous plant metabolic enzymes, such as specialized Pictet-Spenglerases that produce the backbone structures of benzylisoquinoline and monoterpene indole alkaloids and pyridoxal 5'-phosphate-dependent enzymes of diverse functions. Here, we review occurrences of imine chemistry in plant metabolism and their chemical and biochemical mechanisms. A better understanding of plant imine chemistry will ultimately facilitate synthetic biology approaches to further expand the scope of imine natural product biosynthesis for broad biotechnological applications.