Dennis McNevin

Biofiltration as an odour abatement strategy.

The chemical, physical and biological processes occurring in biofiltration are reviewed. A survey of operating biofilter performances is also presented and includes some novel comparative methods. It is concluded that biofiltration is a simple and cost-effective technology for odour removal and that an understanding of the many interactions occuring within the biofilter is essential for the optimal performance of the biofilter.

Alternative analysis of BOD removal in subsurface flow constructed wetlands employing Monod kinetics

A new, mechanistic approach for design and analysis of subsurface flow (SSF) constructed wetlands is presented. The model is based on the assumption that the biological processes in wetlands, like other biological systems, exhibit Monod kinetics. A Monod approach fits well with observed wetland performance. It predicts first-order behaviour at low concentrations, that is, pollutant removal rates which increase with increasing pollutant concentration; and zero-order or saturated behaviour at high pollutant concentrations, that is, a maximum pollutant removal rate. A kinetic analysis of subsurface flow constructed wetlands exhibiting Monod kinetics reveals that loading rate, as well as the zero-order degradation rate constant, are essential parameters for efficient wetlands design for the removal of organic carbon. In particular, Monod kinetics enables the identification of an absolute maximum removal rate which is necessary to prevent undersizing in design. This is significant because it represents a theoretical upper bound on loading rate for wetlands design. The analysis is applied to wetlands data collected in North America by the US EPA in order to extract design criteria for BOD removal. It reveals that maximum loadings for SSF wetlands are at least 80 kg ha-1 d-1 for BOD. In addition, a new dimensionless performance efficiency parameter, omega, is presented as a more effective means of comparing wetland performance.

Differences in Carbon Isotope Discrimination of Three Variants of D-Ribulose-1,5-bisphosphate Carboxylase/Oxygenase Reflect Differences in Their Catalytic Mechanisms

The carboxylation kinetic (stable carbon) isotope effect was measured for purified d-ribulose-1,5-bisphosphate carboxylases/oxygenases (Rubiscos) with aqueous CO2 as substrate by monitoring Rayleigh fractionation using membrane inlet mass spectrometry. This resulted in discriminations (Δ) of 27.4 ± 0.9‰ for wild-type tobacco Rubisco, 22.2 ± 2.1‰ for Rhodospirillum rubrum Rubisco, and 11.2 ± 1.6‰ for a large subunit mutant of tobacco Rubisco in which Leu335 is mutated to valine (L335V). These Δ values are consistent with the photosynthetic discrimination determined for wild-type tobacco and transplastomic tobacco lines that exclusively produce R. rubrum or L335V Rubisco. The Δ values are indicative of the potential evolutionary variability of Δ values for a range of Rubiscos from different species: Form I Rubisco from higher plants; prokaryotic Rubiscos, including Form II; and the L335V mutant. We explore the implications of these Δ values for the Rubisco catalytic mechanism and suggest that Rubiscos that are associated with a lower Δ value have a less product-like carboxylation transition state and/or allow a decarboxylation step that evolution has excluded in higher plants.

Measurement of (carbon) kinetic isotope effect by Rayleigh fractionation using membrane inlet mass spectrometry for CO2-consuming reactions

Methods for determining carbon isotope discrimination, Δ, or kinetic isotope effects, α, for CO2-consuming enzymes have traditionally been cumbersome and time-consuming, requiring careful isolation of substrates and products and conversion of these to CO2 for measurement of isotope ratio by mass spectrometry (MS). An equation originally derived by Rayleigh in 1896 has been used more recently to good effect as it only requires measurement of substrate concentrations and isotope ratios. For carboxylation reactions such as those catalysed by d-ribulose-1,5-bisphosphate carboxylase / oxygenase (RuBisCO, EC 4.1.1.39) and PEP carboxylase (PEPC, EC 4.1.1.31), this has still required sampling of reactions at various states of completion and conversion of all inorganic carbon to CO2, as well as determining the amount of substrate consumed. We introduce a new method of membrane inlet MS which can be used to continuously monitor individual CO2 isotope concentrations, rather than isotope ratio. This enables the use of a simplified, new formula for calculating kinetic isotope effects, based on the assumptions underlying the original Rayleigh fractionation equation and given by.

Inter-relationship between adsorption and pH in peat biofilters in the context of a cation-exchange mechanism

A mathematical model of biofiltration McNevin and Barford (1998) has been augmented to include speciation, acid/base equilibria and pH dependence of adsorptive equilibria. It accurately predicts qualitative aspects of dynamic transients observed in an experimental perfusion column and supported a mechanism of adsorption by cation exchange with acidic functional groups on the surface of peat. It mirrored the buffering capacity of peat when solutions of high and low pH flow over the peat surface. This is a direct result of cation exchange where adsorption of cations increases with pH. This buffering capacity makes peat an attractive medium for engineered biological systems which must often operate within narrow pH bands to optimise biological activity.

A quantitative assessment of a reliable screening technique for the STR analysis of telogen hair roots

Human telogen hairs are commonly recovered as trace evidence but currently have limited use for forensic DNA analysis. Recent studies have revealed that telogen roots may be shed with adhering material that may contain cells, thus providing a potential source of nuclear DNA. A simple histological stain can be used to screen telogen roots for the presence of nuclei, thus increasing the chance of selecting roots that may yield nuclear DNA. Using this method to visualise nuclei, we surveyed 998 hairs from 136 individuals, quantified the number of nuclei, extracted DNA and evaluated corresponding DNA yield and STR profiling success. Of the hairs screened, 35% of telogen roots contained nuclei and in total 6% of all roots screened had more than 100 nuclei. The number of nuclei associated with telogen roots was independent of the presence or absence of visibly adhering material, highlighting the importance of using histological staining rather than simple microscopic examination. DNA yield and STR profiling were significantly and positively correlated with nuclei number. The methods presented here can be incorporated into routine trace and DNA analysis providing an efficient and cost effective method to screen telogen hairs, and predict STR profiling success prior to destructive DNA analysis. The results of this study indicate telogen hairs may provide a reliable source of nuclear DNA for use in routine casework.

Human tissue preservation for disaster victim identification (DVI) in tropical climates

Disaster victim identification (DVI) poses unique challenges for forensic personnel. Typical scenarios may involve many bodies or body parts to identify in remote locations with limited access to laboratory facilities and in extreme temperatures. Transportation of tissue samples to a forensic laboratory for DNA profiling can take weeks without refrigeration. As well as protecting DNA for subsequent analysis, tissue preservation methods ideally should be safe, readily available and easy to transport to the scene at relatively low cost. We examined eight tissue preservatives (salt, DMSO, ethanol, ethanol with EDTA, TENT buffer, RNAlater(®), DNA Genotek Tissue Stabilising Kit and DNAgard(®)) and compared the quantity and quality of DNA recovered from human tissue and preservative solution stored at 35°C. Salt, DMSO, ethanol solutions, DNA Genotek and DNAgard(®) produced full Identifiler(®) genotypes up to one month from DNA extracts. In addition, DMSO, DNA Genotek and DNAgard(®) produced full profiles from aliquots of the liquid preservative.

Towards an integrated performance model for subsurface flow constructed wetlands

Abstract Detailed investigations have been conducted on a set of four pilot scale subsurface flow (SSF) constructed wetlands in order to characterise heat transfer, mass dispersion and biological performance mechanisms. These studies have followed the beds from post construction through unplanted hydraulic base line studies to the current status of mature stands of Phragmites australis. Experimental observations indicate that in unplanted beds, daily thermal fluctuations are depth dependent and range from 1 to 9 degrees Celsius. These fluctuations result in daily thermal inversions, and enhanced mixing and oxygen transport. For planted beds, thermal fluctuations are depth independent, and have a constant amplitude of 2 degrees Celsius. Planted beds may be thermally stratified. Lithium tracer studies corroborate these results for the planted bed. In addition, performance studies indicate that organic pollutant removal is probably limited to organic suspended solids removal, with subsequent biological breakdown. Current first‐order plug flow models can not account for these operational issues. A combined model is necessary to account for lateral dispersion, temperature gradients and settling of suspended solids to accurately reflect real biological removal mechanisms.

Forensically relevant SNaPshot® assays for human DNA SNP analysis: a review

Short tandem repeats are the gold standard for human identification but are not informative for forensic DNA phenotyping (FDP). Single-nucleotide polymorphisms (SNPs) as genetic markers can be applied to both identification and FDP. The concept of DNA intelligence emerged with the potential for SNPs to infer biogeographical ancestry (BGA) and externally visible characteristics (EVCs), which together enable the FDP process. For more than a decade, the SNaPshot® technique has been utilised to analyse identity and FDP-associated SNPs in forensic DNA analysis. SNaPshot is a single-base extension (SBE) assay with capillary electrophoresis as its detection system. This multiplexing technique offers the advantage of easy integration into operational forensic laboratories without the requirement for any additional equipment. Further, the SNP panels from SNaPshot® assays can be incorporated into customised panels for massively parallel sequencing (MPS). Many SNaPshot® assays are available for identity, BGA and EVC profiling with examples including the well-known SNPforID 52-plex identity assay, the SNPforID 34-plex BGA assay and the HIrisPlex EVC assay. This review lists the major forensically relevant SNaPshot® assays for human DNA SNP analysis and can be used as a guide for selecting the appropriate assay for specific identity and FDP applications.

Recovery and identification of bacterial DNA from illicit drugs

Bacterial infections, including Bacillus anthracis (anthrax), are a common risk associated with illicit drug use, particularly among injecting drug users. There is, therefore, an urgent need to survey illicit drugs used for injection for the presence of bacteria and provide valuable information to health and forensic authorities. The objectives of this study were to develop a method for the extraction of bacterial DNA from illicit drugs and conduct a metagenomic survey of heroin and methamphetamine seized in the Australian Capital Territory during 2002-2011 for the presence of pathogens. Trends or patterns in drug contamination and their health implications for injecting drug users were also investigated. Methods based on the ChargeSwitch(®)gDNA mini kit (Invitrogen), QIAamp DNA extraction mini kit (QIAGEN) with and without bead-beating, and an organic phenol/chloroform extraction with ethanol precipitation were assessed for the recovery efficiency of both free and cellular bacterial DNA. Bacteria were identified using polymerase chain reaction and electrospray ionization-mass spectrometry (PCR/ESI-MS). An isopropanol pre-wash to remove traces of the drug and diluents, followed by a modified ChargeSwitch(®) method, was found to efficiently lyse cells and extract free and cellular DNA from Gram-positive and Gram-negative bacteria in heroin and methamphetamine which could then be identified by PCR/ESI-MS. Analysis of 12 heroin samples revealed the presence of DNA from species of Comamonas, Weissella, Bacillus, Streptococcus and Arthrobacter. No organisms were detected in the nine methamphetamine samples analysed. This study develops a method to extract and identify Gram-positive and Gram-negative bacteria from illicit drugs and demonstrates the presence of a range of bacterial pathogens in seized drug samples. These results will prove valuable for future work investigating trends or patterns in drug contamination and their health implications for injecting drug users as well as enabling forensic links between seizures to be examined.