Darren M. Reynolds

Rapid and direct determination of tryptophan in water using synchronous fluorescence spectroscopy

Water samples from lakes situated in and around the area of Tsukuba Science City, Ibaraki Prefecture, Japan, were analysed using synchronous fluorescence spectroscopy and HPLC. The spectra revealed a major emission peak (lambda emm) at 340 nm corresponding to an excitation wavelength (lambda exc) position of 280 nm and a Stokes shift, delta lambda = 60 nm. The fluorescence intensities at these wavelengths were normalised with respect to the water Raman peak, where the lambda exc was 397 nm and the lambda emm was 453 nm, and corrections were introduced to take into account absorption/re-absorption effects of the fluorescing signals caused by the sample matrix. Normalised fluorescence values correlate well with levels of tryptophan present in the samples determined by HPLC analysis (R2 = 0.99). The relationship of the observed tryptophan-like fluorescence in the water samples to the presence of free tryptophan is discussed.

Multivariate analysis of bacterial volatile compound profiles for discrimination between selected species and strains in vitro

Volatile compounds (VCs) are produced by all microorganisms as part of their normal metabolism. The aim of this study was to determine whether bacterial VC profiles could be used to discriminate between selected bacterial species and strains in vitro. Selected Ion Flow Tube Mass Spectrometry (SIFT-MS) was used to quantify the concentration of 23 microbial VCs within the head-space of various bacterial monocultures, during both the logarithmic and stationary growth phases. In comparison with existing techniques, SIFT-MS enables quantitative, high throughput, real-time head-space analysis to be performed, without need for sample preparation. The results show that most VCs were produced by >1 bacterial species or strain, and some were produced by all strains tested. Multivariate analysis using similarity matrices, cluster analysis and multidimensional scaling (MDS) was used to determine whether there was a characteristic VC profile at either the species or strain level. Significant discrimination of all bacterial species and strains was achieved by analysing the VC profiles, and the relative similarity of VC profiles could be differentiated in 2 or 3 dimensional space. This study has shown that there are significant differences in the volatile profiles obtained from various bacterial monocultures grown in vitro, and that the analysis techniques herein employed have the potential to differentiate samples at the strain level.

Aquatic Organic Matter Fluorescence

This is the first comprehensive text on the theory and practice of aquatic organic matter fluorescence analysis, written by the experts who pioneered the research area. This book covers the topic in the broadest possible terms, providing a common reference for making measurements that are comparable across disciplines, and allowing consistent interpretation of data and results. The book includes the fundamental physics and chemistry of organic matter fluorescence, as well as the effects of environmental factors. All aspects of sample handling, data processing, and the operation of both field and laboratory instrumentation are included, providing the practical advice required for successful fluorescence analyses. Advanced methods for data interpretation and modeling, including parallel factor analysis, are also discussed. The book will interest those establishing field, laboratory, or industrial applications of fluorescence, including advanced students and researchers in environmental chemistry, marine science, environmental geosciences, environmental engineering, soil science, and physical geography.

Electrochemically activated solutions: evidence for antimicrobial efficacy and applications in healthcare environments

Due to the limitations associated with the use of existing biocidal agents, there is a need to explore new methods of disinfection to help maintain effective bioburden control, especially within the healthcare environment. The transformation of low mineral salt solutions into an activated metastable state, by electrochemical unipolar action, produces a solution containing a variety of oxidants, including hypochlorous acid, free chlorine and free radicals, known to possess antimicrobial properties. Electrochemically activated solutions (ECAS) have been shown to have broad-spectrum antimicrobial activity, and have the potential to be widely adopted within the healthcare environment due to low-cost raw material requirements and ease of production (either remotely or in situ). Numerous studies have found ECAS to be highly efficacious, as both a novel environmental decontaminant and a topical treatment agent (with low accompanying toxicity), but they are still not in widespread use, particularly within the healthcare environment. This review provides an overview of the scientific evidence for the mode of action, antimicrobial spectrum and potential healthcare-related applications of ECAS, providing an insight into these novel yet seldom utilised biocides.

CoreGenes3.5: a webserver for the determination of core genes from sets of viral and small bacterial genomes

BackgroundCoreGenes3.5 is a webserver that determines sets of core genes from viral and small bacterial genomes as an automated batch process. Previous versions of CoreGenes have been used to classify bacteriophage genomes and mine data from pathogen genomes.FindingsCoreGenes3.5 accepts as input GenBank accession numbers of genomes and performs iterative BLASTP analyses to output a set of core genes. After completion of the program run, the results can be either displayed in a new window for one pair of reference and query genomes or emailed to the user for multiple pairs of small genomes in tabular format.ConclusionsWith the number of genomes sequenced increasing daily and interest in determining phylogenetic relationships, CoreGenes3.5 provides a user-friendly web interface for wet-bench biologists to process multiple small genomes for core gene determinations. CoreGenes3.5 is available at http://binf.gmu.edu:8080/CoreGenes3.5.

Characterisation of dissolved organic matter fluorescence properties by PARAFAC analysis and thermal quenching

The fluorescence intensity of dissolved organic matter (DOM) in aqueous samples is known to be highly influenced by temperature. Although several studies have demonstrated the effect of thermal quenching on the fluorescence of DOM, no research has been undertaken to assess the effects of temperature by combining fluorescence excitation - emission matrices (EEM) and parallel factor analysis (PARAFAC) modelling. This study further extends previous research on thermal quenching by evaluating the impact of temperature on the fluorescence of DOM from a wide range of environmental samples, in the range 20 °C - 0 °C. Fluorescence intensity increased linearly with respect to temperature decrease at all temperatures down to 0 °C. Results showed that temperature affected the PARAFAC components associated with humic-like and tryptophan-like components of DOM differently, depending on the water type. The terrestrial humic-like components, C1 and C2 presented the highest thermal quenching in rural water samples and the lowest in urban water samples, while C3, the tryptophan-like component, and C4, a reprocessed humic-like component, showed opposite results. These results were attributed to the availability and abundance of the components or to the degree of exposure to the heat source. The variable thermal quenching of the humic-like components also indicated that although the PARAFAC model generated the same components across sites, the DOM composition of each component differed between them. This study has shown that thermal quenching can provide additional information on the characteristics and composition of DOM and highlighted the importance of correcting fluorescence data collected in situ.

Evaluation of the efficacy of electrochemically activated solutions against nosocomial pathogens and bacterial endospores.

Aims:  Electrochemically activated solutions (ECAS) are generated from halide salt solutions via specially designed electrolytic cells. The active solutions are known to possess high biocidal activity against a wide range of target microbial species, however, literature revealing the kill‐kinetics of these solutions is limited. The aim of the study was to identify the kill‐rate and extent of population kill for a range of target species (including endospores) using ECAS generated at the anode (anolyte).

The Effect of Long-Term Storage on the Physiochemical and Bactericidal Properties of Electrochemically Activated Solutions

Electrochemically activated solutions (ECAS) are generated by electrolysis of NaCl solutions, and demonstrate broad spectrum antimicrobial activity and high environmental compatibility. The biocidal efficacy of ECAS at the point of production is widely reported in the literature, as are its credentials as a “green biocide.” Acidic ECAS are considered most effective as biocides at the point of production and ill suited for extended storage. Acidic ECAS samples were stored at 4 °C and 20 °C in glass and polystyrene containers for 398 days, and tested for free chlorine, pH, ORP and bactericidal activity throughout. ORP and free chlorine (mg/L) in stored ECAS declined over time, declining at the fastest rate when stored at 20 °C in polystyrene and at the slowest rate when stored at 4 °C in glass. Bactericidal efficacy was also affected by storage and ECAS failed to produce a 5 log10 reduction on five occasions when stored at 20 °C. pH remained stable throughout the storage period. This study represents the longest storage evaluation of the physiochemical parameters and bactericidal efficacy of acidic ECAS within the published literature and reveals that acidic ECAS retain useful bactericidal activity for in excess of 12 months, widening potential applications.

Comparative Antimicrobial Activities of Aerosolized Sodium Hypochlorite, Chlorine Dioxide, and Electrochemically Activated Solutions Evaluated Using a Novel Standardized Assay

ABSTRACT The main aim of this study was to develop a standardized experimental assay to enable differential antimicrobial comparisons of test biocidal aerosols. This study represents the first chlorine-matched comparative assessment of the antimicrobial activities of aerosolized sodium hypochlorite, chlorine dioxide, and electrochemically activated solution (ECAS) to determine their relative abilities to decontaminate various surface-associated health care-relevant microbial challenges. Standard microbiological challenges were developed by surface-associating typed Pseudomonas aeruginosa, Staphylococcus aureus, Bacillus subtilis spores, or a clinical methicillin-resistant S. aureus (MRSA) strain on stainless steel, polypropylene, or fabric. All test coupons were subjected to 20-min biocidal aerosols of chlorine-matched (100 ppm) sodium hypochlorite, chlorine dioxide, or ECAS within a standard aerosolization chamber using a commercial humidifier under defined conditions. Biocidal treatment type and material surface had a significant effect on the number of microorganisms recovered from various material surfaces following treatment exposure. Under the conditions of the assay, the order of antimicrobial efficacy of biocidal aerosol treatment was as follows: ECAS > chlorine dioxide > sodium hypochlorite. For all biocides, greater antimicrobial reductions were seen when treating stainless steel and fabric than when treating plastic-associated microorganisms. The experimental fogging system and assay protocol designed within this study were shown capable of differentiating the comparative efficacies of multiple chlorine-matched biocidal aerosols against a spectrum of target organisms on a range of test surface materials and would be appropriate for testing other biocidal aerosol treatments or material surfaces.

Application of Bacterial Bioluminescence To Assess the Efficacy of Fast-Acting Biocides

ABSTRACT Traditional microbiological techniques are used to provide reliable data on the rate and extent of kill for a range of biocides. However, such techniques provide very limited data regarding the initial rate of kill of fast-acting biocides over very short time domains. This study describes the application of a recombinant strain of Escherichia coli expressing the Photorhabdus luminescens lux operon as a whole-cell biosensor. Light emission is linked directly to bacterial metabolism; therefore, by monitoring light output, the impact of fast-acting biocides can be assessed. Electrochemically activated solutions (ECASs), bleach, Virkon, and ethanol were assessed at three concentrations (1%, 10%, 80%) in the presence of organic soiling. Over a 2-s time course, 80% ECAS produced the greatest reduction in light output in the absence of organic load but was strongly inhibited by its presence. Eighty percent ethanol outperformed all tested biocides in the presence of organic soil. Bleach and Virkon produced similar reductions in bioluminescence at matched concentrations within the time course of the assay. It was also demonstrated that the assay can be used to rapidly assess the impact of organic soiling. The use of bioluminescent bacteria as whole-cell bioreporters allows assessment of the relative efficacies of fast-acting biocides within milliseconds of application. The assay can be used to investigate activity over short or extended time domains to confirm complete metabolic inhibition of the bioreporter. Moreover, the assay may enable further elucidation of their mechanism of action by allowing the investigation of activity over time domains precluded by traditional microbiology.