Bathycranium: synonymised with Syntormon, distinction between Parasyntormon and Syntormon discussed and S. bicolorellus and S. luteicornis (Diptera: Dolichopodidae) redescribed

It is demonstrated that there is no valid basis on which to sustain the monotypic genus Bathycranium Strobl and concluded that Bathycranium should be recognised as a junior synonym of Syntormon Loew (new status). The species Syntormon bicolorellus Zetterstedt (new combination) falls into a natural grouping of Syntormon species with downcurved facial hairs in females. This species and S. luteicornis Parent are redescribed. Distinctions between Syntormonand Parasyntormon are discussed

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

<p>Abstract</p> <p>Background</p> <p><it>Trigonopsis variabilis </it><smcaps>D</smcaps>-amino acid oxidase (<it>Tv</it>DAO) 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 <it>Tv</it>DAO 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 <it>Pichia pastoris Tv</it>DAO whole-cell biocatalyst.</p> <p>Results</p> <p>As compared to the native <it>T. variabilis </it>host, a more than seven-fold enhancement of the intracellular level of oxidase activity was achieved in <it>P. pastoris </it>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 × 10³U/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 ≥ 200 g/L compared to only 117 g/L using the standard methanol feed. Permeabilization of <it>P. pastoris </it>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 <it>Tv</it>DAO expressing <it>Escherichia coli </it>strain) under conditions of <smcaps>D</smcaps>-methionine conversion using vigorous aeration.</p> <p>Conclusions</p> <p>Stepwise optimization using a multi-level engineering approach has delivered a new <it>P. pastoris </it>whole cell <it>Tv</it>DAO 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 <it>P. pastoris </it>catalyst has been evaluated for industrial purposes.</p

Antimicrobial adhesive films by plasma-enabled polymerisation of m-cresol

This work reveals a versatile new method to produce films with antimicrobial properties that can also bond materials together with robust tensile adhesive strength. Specifically, we demonstrate the formation of coatings by using a dielectric barrier discharge (DBD) plasma to convert a liquid small-molecule precursor, m-cresol, to a solid film via plasma-assisted on-surface polymerisation. The films are quite appealing from a sustainability perspective: they are produced using a low-energy process and from a molecule produced in abundance as a by-product of coal tar processing. This process consumes only 1.5 Wh of electricity to create a 1 cm2 film, which is much lower than other methods commonly used for film deposition, such as chemical vapour deposition (CVD). Plasma treatments were performed in plain air without the need for any carrier or precursor gas, with a variety of exposure durations. By varying the plasma parameters, it is possible to modify both the adhesive property of the film, which is at a maximum at a 1 min plasma exposure, and the antimicrobial property of the film against Escherichia coli, which is at a maximum at a 30 s exposure

Effect of plasmid design and type of integration event on recombinant protein expression in Pichia pastoris

(syn. ) is one of the most common eukaryotic expression systems for heterologous protein production. Expression cassettes are typically integrated in the genome to obtain stable expression strains. In contrast to , where short overhangs are sufficient to target highly specific integration, long overhangs are more efficient in and ectopic integration of foreign DNA can occur. Here, we aimed to elucidate the influence of ectopic integration by high-throughput screening of >700 transformants and whole-genome sequencing of 27 transformants. Different vector designs and linearization approaches were used to mimic the most common integration events targeted in Fluorescence of an enhanced green fluorescent protein (eGFP) reporter protein was highly uniform among transformants when the expression cassettes were correctly integrated in the targeted locus. Surprisingly, most nonspecifically integrated transformants showed highly uniform expression that was comparable to specific integration, suggesting that nonspecific integration does not necessarily influence expression. However, a few clones

Synthesis of 2,6-disubstituted dihydropyrans via an efficient BiBr3-initiated three component, one-pot cascade

The rapid synthesis of cis-2,6-disubstituted dihydropyrans is achieved in a three-component, one-pot cascade reaction. BiBr3-mediated addition of ketene silyl acetals or silyl enol ethers to beta,gamma-unsaturated cis-4-trimethylsilyl-3-butenal provides a Mukaiyama aldol adduct containing a vinylsilane moiety tethered to a silyl ether. Addition of a second aldehyde initiates a domino sequence involving intermolecular addition followed by an intramolecular silyl-modified Sakurai (ISMS) reaction. Isolated yields of this one-pot reaction vary from 44 to 80% and all compounds were isolated as the cis-diastereomers (10 examples). (C) 2009 Elsevier Ltd. All rights reserved

Self-Organizing Maps For Knowledge Discovery From Corporate Databases To Develop Risk Based Prioritization For Stagnation

Stagnation or low turnover of water within water distribution systems may result in water quality issues, even for relatively short durations of stagnation / low turnover if other factors such as deteriorated aging pipe infrastructure are present. As leakage management strategies, including the creation of smaller pressure management zones, are implemented increasingly more dead ends are being created within networks and hence potentially there is an increasing risk to water quality due to stagnation / low turnover. This paper presents results of applying data driven tools to the large corporate databases maintained by UK water companies. These databases include multiple information sources such as asset data, regulatory water quality sampling, customer complaints etc. A range of techniques exist for exploring the interrelationships between various types of variables, with a number of studies successfully using Artificial Neural Networks (ANNs) to probe complex data sets. Self Organising Maps (SOMs), are a class of unsupervised ANN that perform dimensionality reduction of the feature space to yield topologically ordered maps, have been used successfully for similar problems to that posed here. Notably for this application, SOM are trained without classes attached in an unsupervised fashion. Training combines competitive learning (learning the position of a data cloud) and co-operative learning (self-organising of neighbourhoods). Specifically, in this application SOMs performed multidimensional data analysis of a case study area (covering a town for an eight year period). The visual output of the SOM analysis provides a rapid and intuitive means of examining covariance between variables and exploring hypotheses for increased understanding. For example, water age (time from system entry, from hydraulic modelling) in combination with high pipe specific residence time and old cast iron pipe were found to be strong explanatory variables. This derived understanding could ultimately be captured in a tool providing risk based prioritisation scores

Recommendations for improving integration in national end-to-end flood forecasting systems: an overview of the FFIR (flooding from intense rainfall) programme

Recent surface-water and flash floods have caused millions of pounds worth of damage in the UK. These events form rapidly and are difficult to predict due to their short-lived and localised nature. The interdisciplinary Flooding From Intense Rainfall (FFIR) programme investigated the feasibility of enhancing the integration of an end-to-end forecasting system for flash and surface-water floods to help increase the lead time for warnings for these events. Here we propose developments to the integration of an operational end-to-end forecasting system based on the findings of the FFIR programme. The suggested developments include methods to improve radar-derived rainfall rates and understanding of the uncertainty in the position of intense rainfall in weather forecasts; the addition of hydraulic modelling components; and novel education techniques to help lead to effective dissemination of flood warnings. We make recommendations for future advances such as research into the propagation of uncertainty throughout the forecast chain. We further propose the creation of closer bonds to the end users to allow for an improved, integrated, end-to-end forecasting system that is easily accessible for users and end users alike, and will ultimately help mitigate the impacts of flooding from intense rainfall by informed and timely action

Cell-free pipeline for discovery of thermotolerant xylanases and endo-1,4-β-glucanases

The rapid expansion in the number of sequenced genomes and metagenomes provides an exceptional resource for mining of the enzymes with biotechnologically relevant properties. However, the majority of protein production and analysis methods are not sufficiently cost-efficient and scalable to experimentally verify the results of computational genomic mining. Here, we present a pipeline based on Leishmania tarentolae cell-free system that was used to characterize 30 putative thermostable endo-1,4-beta-glucanases and xylanases identified in public genomic databases. In order to analyse the recombinant proteins without purification, novel high-throughput assays for glucanase and xylanase activities were developed. The assays rely on solubilisation of labelled particulate substrates performed in multiwell plates. Using this approach both acidophilic and thermophilic enzymes were identified. The developed approach enables rapid discovery of new biotechnologically useful enzymes

Characterization of pre-transplant psychosocial burden in an integrated national islet transplant programme

The psychological burden experienced by people with diabetes prior to islet transplantation is recognized but has not been studied comprehensively, especially in relation to glycemia. Therefore, we conducted a rigorous pre-operative psychosocial profile of UK islet transplant recipients, and compared groups with higher/lower HbA1 c to test the null hypothesis that pre-transplant hypoglycemia awareness and psychosocial burden would not be related to baseline HbA1 c in this high-risk cohort. Pre-transplant, recipients (n = 44) completed validated hypoglycemia awareness questionnaires and generic/diabetes-specific measures of psychological traits and states. Scores were compared in groups, dichotomized by HbA1 c (≤8% versus >8%). Participants were aged (mean±SD) 53 ± 10 years; 64% were women; with HbA1 c 8.3 ± 1.7%. Median rate of severe hypoglycemia over the preceding 12 months was 13 events/person-year and 90% had impaired awareness of hypoglycemia (Gold/Clarke score ≥4). Participants had elevated fear of hypoglycemia (HFS-II Worry), impaired diabetes-specific quality of life (DQoL) and low generic health status (SF-36; EQ-5D). One quarter reported scores indicating likely anxiety/depression (HAD). Dispositional optimism (LOT-R) and generalized self-efficacy (GSE) were within published ‘norms.’ Despite negative perceptions of diabetes (including low personal control), participants were confident that islet transplantation would help (BIPQ). Hypoglycemia awareness and psychosocial profile were comparable in lower (n = 24) and higher (n = 20) HbA1 c groups. Islet transplant candidates report sub-optimal generic psychological states (anxiety/depressive symptoms), health status and diabetes-specific psychological states (fear of hypoglycemia, diabetes-specific quality of life). While their generic psychological traits (optimism, self-efficacy) are comparable with the general population, they are highly optimistic about forthcoming transplant. HbA1 c is not a proxy measure of psychosocial burden, which requires the use of validated questionnaires to systematically identify those who may benefit most from psychological assessment and support