Characterisation of soils containing naturally occurring radioactive material (NORM) using in situ and rapid laboratory techniques. Case study: South Terras, Cornwall

In situ analysis has become increasingly important for contaminated land investigation and remediation. At present, portable techniques are used mainly as scanning tools to assess the spread and magnitude of the contamination, and are an adjunct to conventional laboratory analyses. A site in Cornwall, containing naturally occurring radioactive material (NORM), provided an opportunity for Reading University PhD student Anna Kutner to compare analytical data collected in situ with data generated by laboratory-based methods. The preliminary results in this paper extend the author‟s poster presentation at last September‟s GeoSpec2010 conference held in Lancaster.

Rapid protein domain assignment from amino acid sequence using predicted secondary structure

The elucidation of the domain content of a given protein sequence in the absence of determined structure or significant sequence homology to known domains is an important problem in structural biology. Here we address how successfully the delineation of continuous domains can be accomplished in the absence of sequence homology using simple baseline methods, an existing prediction algorithm (Domain Guess by Size), and a newly developed method (DomSSEA). The study was undertaken with a view to measuring the usefulness of these prediction methods in terms of their application to fully automatic domain assignment. Thus, the sensitivity of each domain assignment method was measured by calculating the number of correctly assigned top scoring predictions. We have implemented a new continuous domain identification method using the alignment of predicted secondary structures of target sequences against observed secondary structures of chains with known domain boundaries as assigned by Class Architecture Topology Homology (CATH). Taking top predictions only, the success rate of the method in correctly assigning domain number to the representative chain set is 73.3%. The top prediction for domain number and location of domain boundaries was correct for 24% of the multidomain set (±20 residues). These results have been put into context in relation to the results obtained from the other prediction methods assessed

Assimilation of RAPID array observations into an ocean model

Experiments assimilating the RAPID dataset of deep temperature and salinity profiles at 26.5°N on the western and eastern Atlantic boundaries into a 1° global NEMO ocean model have been performed. The meridional overturning circulation (MOC) is then assessed against the transports calculated directly from observations. The best initialization found for this short period was obtained by assimilating the EN3 upper-ocean hydrography database prior to 2004, after which different methods of assimilating 5-day average RAPID profiles at the western boundary were tested. The model MOC is strengthened by ∼ 2 Sv giving closer agreement with the RAPID array transports, when the western boundary profiles are assimilated only below 900 m (the approximate depth of the Florida Straits, which are not well resolved) and when the T,S observations are spread meridionally from 10 to 35°N along the deep western boundary. The use of boundary-focused covariances has the largest impact on the assimilation results, otherwise using more conventional Gaussian covariances has a very local impact on the MOC at 26°N with strong adverse impacts on the MOC stream function at higher and lower latitudes. Even using boundary-focused covariances only enables the MOC to be strengthened for ∼ 2 years, after which the increased transport of warm waters leads to a negative feedback on water formation in the subpolar gyre which then reduces the MOC. This negative feedback can be mitigated if EN3 hydrography data continue to be assimilated along with the RAPID array boundary data. Copyright © 2012 Royal Meteorological Society and Crown in the right of Canada.

A modified rapid enzymatic microtiter plate assay for the quantification of intracellular γ-aminobutyric acid and succinate semialdehyde in bacterial cells

The GABase assay is widely used to rapidly and accurately quantify levels of extracellular γ-aminobutyric acid (GABA). Here we demonstrate a modification of this assay that enables quantification of intracellular GABA in bacterial cells. Cells are lysed by boiling and ethanolamine-O-sulphate, a GABA transaminase inhibitor is used to distinguish between GABA and succinate semialdehyde.

Use of a LightCycler gyrA mutation assay for rapid identification of mutations conferring decreased susceptibility to ciprofloxacin in multiresistant Salmonella enterica serotype typhimurium DT104 isolates

A LightCycler-based PCR-hybridization gyrA mutation assay (GAMA) was developed to rapidly detect gyrA point mutations in multiresistant (MR) Salmonella enterica serotype Typhimurium DT104 with decreased susceptibility to ciprofloxacin (MIC, 0.25 to 1.0 mg/liter). Ninety-two isolates (49 human, 43 animal) were tested with three individual oligonucleotide probes directed against an Asp-87-to-Asn (GAC --> AAC) mutation, an Asp-87-to-Gly (GAC --> GGC) mutation, and a Ser-83-to-Phe (TCC --> TTC) mutation. Strains homologous to the probes could be distinguished from strains that had different mutations by their probe-target melting temperatures. Thirty-seven human and 30 animal isolates had an Asp-87-to-Asn substitution, 6 human and 6 animal isolates had a Ser-83-to-Phe substitution, and 5 human and 2 animal isolates had an Asp-87-to-Gly substitution. The remaining six strains all had mismatches with the three probes and therefore different gyrA mutations. The sequencing of gyrA from these six isolates showed that one human strain and two animal strains had an Asp-87-to-Tyr (GAC --> TAC) substitution and two animal strains had a Ser-83-to-Tyr (TCC --> TAC) substitution. One animal strain had no gyrA mutation, suggesting that this isolate had a different mechanism of resistance. Fifty-eight of the strains tested were indistinguishable by several different typing methods including antibiograms, pulsed-field gel gel electrophoresis, and plasmid profiling, although they could be further subdivided according to gyrA mutation. This study confirmed that MR DT104 with decreased susceptibility to ciprofloxacin from humans and food animals in England and Wales may have arisen independently against a background of clonal spread of MR DT104.

Infra-red laser ablative micromachining of parylene-C on SiO2substrates for rapid prototyping, high yield, human neuronal cell patterning

Cell patterning commonly employs photolithographic methods for the micro fabrication of structures on silicon chips. These require expensive photo-mask development and complex photolithographic processing. Laser based patterning of cells has been studied in vitro and laser ablation of polymers is an active area of research promising high aspect ratios. This paper disseminates how 800 nm femtosecond infrared (IR) laser radiation can be successfully used to perform laser ablative micromachining of parylene-C on SiO2 substrates for the patterning of human hNT astrocytes (derived from the human teratocarcinoma cell line (hNT)) whilst 248 nm nanosecond ultra-violet laser radiation produces photo-oxidization of the parylene-C and destroys cell patterning. In this work, we report the laser ablation methods used and the ablation characteristics of parylene-C for IR pulse fluences. Results follow that support the validity of using IR laser ablative micromachining for patterning human hNT astrocytes cells. We disseminate the variation in yield of patterned hNT astrocytes on parylene-C with laser pulse spacing, pulse number, pulse fluence and parylene-C strip width. The findings demonstrate how laser ablative micromachining of parylene-C on SiO2 substrates can offer an accessible alternative for rapid prototyping, high yield cell patterning with broad application to multi-electrode arrays, cellular micro-arrays and microfluidics.

Rapid assessment of historic, current and future habitat quality for biodiversity around UK Natura 2000 sites

Changes in landscape composition and structure may impact the conservation and management of protected areas. Species that depend on specific habitats are at risk of extinction when these habitats are degraded or lost. Designing robust methods to evaluate landscape composition will assist decision- and policy-making in emerging landscapes. This paper describes a rapid assessment methodology aimed at evaluating landcover quality for birds, plants, butterflies and bees around seven UK Natura 2000 sites. An expert panel assigned quality values to standard Coordination of Information on the Environment (CORINE) landcover classes for each taxonomic group. Quality was assessed based on historical (1950, 1990), current (2000) and future (2030) land-cover data, the last projected using three alternative scenarios: a growth applied strategy (GRAS), a business-as-might-beusual (BAMBU) scenario, and sustainable European development goal (SEDG) scenario. A quantitative quality index weighted the area of each land-cover parcel with a taxa-specific quality measure. Land parcels with high quality for all taxonomic groups were evaluated for temporal changes in area, size and adjacency. For all sites and taxonomic groups, the rate of deterioration of land-cover quality was greater between 1950 and 1990 than current rates or as modelled using the alternative future scenarios (2000– 2030). Model predictions indicated land-cover quality stabilized over time under the GRAS scenario, and was close to stable for the BAMBU scenario. The SEDG scenario suggested an ongoing loss of quality, though this was lower than the historical rate of c. 1% loss per decade. None of the future scenarios showed accelerated fragmentation, but rather increases in the area, adjacency and diversity of high quality land parcels in the landscape.

Analysis of the courtship behavior of the Navel Orangeworm, Amyelois transitella (Walker) (Lepidoptera: Pyralidae), with a commentary on methods for the analysis of sequences of behavioral transitions

The courtship behavior of the navel orangeworm, Amyelois transitella, was examined in a wind tunnel. Sixty nine courtship sequences were analyzed and successful sequences divided into two categories: rapid courtship sequences, which involved few breaks in contact, short or no periods of male/female chasing and lasted <10 s between initial contact and mating; and prolonged courtship sequences, which involved many breaks in contact, extended periods of male/female chasing and lasted >10 s. Fifty six (81%) courtships were successful (50.7% rapid courtship and 30.4% prolonged courtship); the remaining 13 (18.8%) sequences were failed courtships. Of failed courtships, 9 (13.0%) were due to males losing contact with females during courtship chases and 4 (5.8%) due to females flying away immediately after male contact. Of all courtship sequences involving a break in contact during a chase, 38.5% resulted in an unsuccessful mating attempt. These findings contrast with previous studies of the courtship behavior of the navel orangeworm, potentially indicating that the type of bioassay used to study courtship may have a large effect on the behavioral sequences displayed. We evaluate several diagnostic techniques for the analysis of sequences of behavioral transitions.

Rapid, automated detection of stem canker symptoms in woody perennials using artificial neural network analysis

Background Pseudomonas syringae can cause stem necrosis and canker in a wide range of woody species including cherry, plum, peach, horse chestnut and ash. The detection and quantification of lesion progression over time in woody tissues is a key trait for breeders to select upon for resistance. Results In this study a general, rapid and reliable approach to lesion quantification using image recognition and an artificial neural network model was developed. This was applied to screen both the virulence of a range of P. syringae pathovars and the resistance of a set of cherry and plum accessions to bacterial canker. The method developed was more objective than scoring by eye and allowed the detection of putatively resistant plant material for further study. Conclusions Automated image analysis will facilitate rapid screening of material for resistance to bacterial and other phytopathogens, allowing more efficient selection and quantification of resistance responses.

Comparison of agar gel immunodiffusion test, rapid slide agglutination test, microbiological culture and PCR for the diagnosis of canine brucellosis

The performance of the rapid slide agglutination test, with and without 2-mercaptoethanol (RSAT and 2ME-RSAT) and agar gel immunodiffusion test (AGID) was evaluated for the diagnosis of brucellosis in naturally infected dogs. The microbiological culture, PCR and clinical parameters were used as reference. A total of 167 dogs were clinically examined and tested by blood culture, culture of semen/vaginal swab and PCR in blood and semen/vaginal swab. According to the results observed the 167 dogs were divided into three groups: Brucella canis infected dogs (Group 1). B. canis non-infected dogs (Group 2) and dogs with suspected brucellosis (Group 3). The dogs were then tested by RSAT, 2ME-RSAT and AGID. Groups 1 and 2 were used to calculate the diagnostic sensitivity and specificity of the serological tests and the results observed in Group 3 were also discussed. The diagnostic sensitivity of RSAT, 2ME-RSAT and AGID was respectively 70.58%, 31.76%, and 52.94%. The diagnostic specificity of RSAT, 2ME-RSAT and AGID was respectively 83.34%, 100%, and 100%. In dogs with suspected brucellosis 15% were RSAT positive, none was 2ME-RSAT positive and 5% were AGID positive. Although the serological tests are the most commonly used methods for brucellosis diagnosis, a significant proportion of false-negative results were observed highlighting the importance of the direct methods of diagnosis, like blood culture and PCR to improve the diagnosis of canine brucellosis. (c) 2008 Elsevier Ltd. All rights reserved.

False-positive results of a rapid K39-based strip test and Chagas disease

Background: The definitive diagnosis of visceral. leishmaniasis (VL) requires invasive procedures with demonstration of amastigotes in tissue or promastigotes in culture. Unfortunately, these approaches require laboratory materials not available in poor countries where the disease is endemic. The correct diagnosis of VL is important, and made more difficult by the fact that several common tropical diseases such as malaria, disseminated tuberculosis, and enteric fever share the same clinical presentation. Serological tests have been developed to replace parasitological diagnosis in the field. A commercially available K39-based strip test for VL has been developed for this purpose. The endemic area of leishmaniasis in Brazil overlaps the endemic area of Chagas disease, a disease that can cause false-positive serological test results. The aim of this study was to evaluate the incidence of false-positive exams using a rapid test for VL in patients with Chagas disease. Methods: A rapid test based on the recombinant K39 antigen of Leishmania was used in: (1) 30 patients with confirmed Chagas disease, (2) 30 patients with a serological diagnosis of Chagas disease by ELISA, indirect immunofluorescence, indirect hemagglutination, and chemiluminescence, (3) 30 healthy patients from a non-endemic area as the control group, (4) 30 patients with confirmed VL, and (5) 20 patients with proved cutaneous leishmaniasis. Results: The sensitivity and specificity of the rapid strip test were 100% when compared with healthy volunteers and those with confirmed Chagas disease. One false-positive result occurred in the group with Chagas disease diagnosed by serological tests (specificity of 96%). Conclusion: The rapid test based on recombinant K39 is a useful diagnostic assay, and a false-positive result rarely occurs in patients with a serological diagnosis of Chagas disease. (C) 2008 International Society for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Rapid and direct determination of glyphosate and aminomethylphosphonic acid in water using anion-exchange chromatography with coulometric detection

A simple, rapid, and low-cost coulometric method for direct detection of glyphosate and aminomethylphosphonic acid (AMPA) in water samples using anion-exchange chromatography and coulometric detection with copper electrode is presented. Under optimized conditions, the limits of detection (LODs) (S/N = 3) were 0.038 mu g ml(-1) for glyphosate and 0.24 mu g ml(-1) for AMPA, without any preconcentration method. The calibration curves were linear and presented an excellent correlation coefficient. The method was successfully applied to the determination of glyphosate and AMPA in water samples without any kind of extraction, clean-up, or preconcentration step. No interferent was found in the water, like this, the recovery was, practically, 100%. (c) 2008 Elsevier B.V. All rights reserved.

A rapid and reliable bonding process for microchip electrophoresis fabricated in glass substrates

In this report, we describe a rapid and reliable process to bond channels fabricated in glass substrates. Glass channels were fabricated by photolithography and wet chemical etching. The resulting channels were bonded against another glass plate containing a 50-mu m thick PDMS layer. This same PDMS layer was also used to provide the electrical insulation of planar electrodes to carry out capacitively coupled contactless conductivity detection. The analytical performance of the proposed device was shown by using both LIF and capacitively coupled contactless conductivity detection systems. Efficiency around 47 000 plates/m was achieved with good chip-to-chip repeatability and satisfactory long-term stability of EOF. The RSD for the EOF measured in three different devices was ca. 7%. For a chip-to-chip comparison, the RSD values for migration time, electrophoretic current and peak area were below 10%. With the proposed approach, a single chip can be fabricated in less than 30 min including patterning, etching and sealing steps. This fabrication process is faster and easier than the thermal bonding process. Besides, the proposed method does not require high temperatures and provides excellent day-to-day and device-to-device repeatability.

Drug resistance in Mycobacterium tuberculosis clinical isolates from Brazil: Phenotypic and genotypic methods

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Rapid detection of resistance to pyrazinamide in Mycobacterium tuberculosis using the resazurin microtitre assay

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)