Characterisation of indoor air organic pollutants in Brisbane

The occurrence and levels of airborne polycyclic aromatic hydrocarbons and volatile organic compounds in selected non-industrial environments in Brisbane have been investigated as part of an integrated indoor air quality assessment program. The most abundant and most frequently encountered compounds include, nonanal, decanal, texanol, phenol, 2-ethyl-1-hexanol, ethanal, naphthalene, 2,6-tert-butyl-4-methyl-phenol (BHT), salicylaldehyde, toluene, hexanal, benzaldehyde, styrene, ethyl benzene, o-, m- and pxylenes, benzene, n-butanol, 1,2-propandiol, and n-butylacetate. Many of the 64 compounds usually included in the European Collaborative Action method of TVOC analysis were below detection limits in the samples analysed. In order to extract maximum amount of information from the data collected, multivariate data projection methods have been employed. The implications of the information extracted on source identification and exposure control are discussed.

New particle formation and growth at a remote, sub-tropical coastal location

A month-long intensive measurement campaign was conducted in March/April 2007 at Agnes Water, a remote coastal site just south of the Great Barrier Reef on the east coast of Australia. Particle and ion size distributions were continuously measured during the campaign. Coastal nucleation events were observed in clean, marine air masses coming from the south-east on 65% of the days. The events usually began at ~10:00 local time and lasted for 1-4 hrs. They were characterised by the appearance of a nucleation mode with a peak diameter of ~10 nm. The freshly nucleated particles grew within 1-4 hrs up to sizes of 20-50 nm. The events occurred when solar intensity was high (~1000 W m-2) and RH was low (~60%). Interestingly, the events were not related to tide height. The volatile and hygroscopic properties of freshly nucleated particles (17-22.5 nm), simultaneously measured with a volatility-hygroscopicity-tandem differential mobility analyser (VH-TDMA), were used to infer chemical composition. The majority of the volume of these particles was attributed to internally mixed sulphate and organic components. After ruling out coagulation as a source of significant particle growth, we conclude that the condensation of sulphate and/or organic vapours was most likely responsible for driving particle growth during the nucleation events. We cannot make any direct conclusions regarding the chemical species that participated in the initial particle nucleation. However, we suggest that nucleation may have resulted from the photo-oxidation products of unknown sulphur or organic vapours emitted from the waters of Hervey Bay, or from the formation of DMS-derived sulphate clusters over the open ocean that were activated to observable particles by condensable vapours emitted from the nutrient rich waters around Fraser Island or Hervey Bay. Furthermore, a unique and particularly strong nucleation event was observed during northerly wind. The event began early one morning (08:00) and lasted almost the entire day resulting in the production of a large number of ~80 nm particles (average modal concentration during the event was 3200 cm-3). The Great Barrier Reef was the most likely source of precursor vapours responsible for this event.

Long-term trends in fine particle number concentrations in the urban atmosphere of Brisbane : the relevance of traffic emissions and new particle formation

The measurement of submicrometre (< 1.0 m) and ultrafine particles (diameter < 0.1 m) number concentration have attracted attention since the last decade because the potential health impacts associated with exposure to these particles can be more significant than those due to exposure to larger particles. At present, ultrafine particles are not regularly monitored and they are yet to be incorporated into air quality monitoring programs. As a result, very few studies have analysed their long-term and spatial variations in ultrafine particle concentration, and none have been in Australia. To address this gap in scientific knowledge, the aim of this research was to investigate the long-term trends and seasonal variations in particle number concentrations in Brisbane, Australia. Data collected over a five-year period were analysed using weighted regression models. Monthly mean concentrations in the morning (6:00-10:00) and the afternoon (16:00-19:00) were plotted against time in months, using the monthly variance as the weights. During the five-year period, submicrometre and ultrafine particle concentrations increased in the morning by 105.7% and 81.5% respectively whereas in the afternoon there was no significant trend. The morning concentrations were associated with fresh traffic emissions and the afternoon concentrations with the background. The statistical tests applied to the seasonal models, on the other hand, indicated that there was no seasonal component. The spatial variation in size distribution in a large urban area was investigated using particle number size distribution data collected at nine different locations during different campaigns. The size distributions were represented by the modal structures and cumulative size distributions. Particle number peaked at around 30 nm, except at an isolated site dominated by diesel trucks, where the particle number peaked at around 60 nm. It was found that ultrafine particles contributed to 82%-90% of the total particle number. At the sites dominated by petrol vehicles, nanoparticles (< 50 nm) contributed 60%-70% of the total particle number, and at the site dominated by diesel trucks they contributed 50%. Although the sampling campaigns took place during different seasons and were of varying duration these variations did not have an effect on the particle size distributions. The results suggested that the distributions were rather affected by differences in traffic composition and distance to the road. To investigate the occurrence of nucleation events, that is, secondary particle formation from gaseous precursors, particle size distribution data collected over a 13 month period during 5 different campaigns were analysed. The study area was a complex urban environment influenced by anthropogenic and natural sources. The study introduced a new application of time series differencing for the identification of nucleation events. To evaluate the conditions favourable to nucleation, the meteorological conditions and gaseous concentrations prior to and during nucleation events were recorded. Gaseous concentrations did not exhibit a clear pattern of change in concentration. It was also found that nucleation was associated with sea breeze and long-range transport. The implications of this finding are that whilst vehicles are the most important source of ultrafine particles, sea breeze and aged gaseous emissions play a more important role in secondary particle formation in the study area.

Neurolinguistic contributions to understanding the bilingual mental lexicon

Introduction Many bilinguals will have had the experience of unintentionally reading something in a language other than the intended one (e.g. MUG to mean mosquito in Dutch rather than a receptacle for a hot drink, as one of the possible intended English meanings), of finding themselves blocked on a word for which many alternatives suggest themselves (but, somewhat annoyingly, not in the right language), of their accent changing when stressed or tired and, occasionally, of starting to speak in a language that is not understood by those around them. These instances where lexical access appears compromised and control over language behavior is reduced hint at the intricate structure of the bilingual lexical architecture and the complexity of the processes by which knowledge is accessed and retrieved. While bilinguals might tend to blame word finding and other language problems on their bilinguality, these difficulties per se are not unique to the bilingual population. However, what is unique, and yet far more common than is appreciated by monolinguals, is the cognitive architecture that subserves bilingual language processing. With bilingualism (and multilingualism) the rule rather than the exception (Grosjean, 1982), this architecture may well be the default structure of the language processing system. As such, it is critical that we understand more fully not only how the processing of more than one language is subserved by the brain, but also how this understanding furthers our knowledge of the cognitive architecture that encapsulates the bilingual mental lexicon. The neurolinguistic approach to bilingualism focuses on determining the manner in which the two (or more) languages are stored in the brain and how they are differentially (or similarly) processed. The underlying assumption is that the acquisition of more than one language requires at the very least a change to or expansion of the existing lexicon, if not the formation of language-specific components, and this is likely to manifest in some way at the physiological level. There are many sources of information, ranging from data on bilingual aphasic patients (Paradis, 1977, 1985, 1997) to lateralization (Vaid, 1983; see Hull & Vaid, 2006, for a review), recordings of event-related potentials (ERPs) (e.g. Ardal et al., 1990; Phillips et al., 2006), and positron emission tomography (PET) and functional magnetic resonance imaging (fMRI) studies of neurologically intact bilinguals (see Indefrey, 2006; Vaid & Hull, 2002, for reviews). Following the consideration of methodological issues and interpretative limitations that characterize these approaches, the chapter focuses on how the application of these approaches has furthered our understanding of (1) selectivity of bilingual lexical access, (2) distinctions between word types in the bilingual lexicon and (3) control processes that enable language selection.

The mechanism of breath aerosol formation

Background: Aerosol production during normal breathing is often attributed to turbulence in the respiratory tract. That mechanism is not consistent with a high degree of asymmetry between aerosol production during inhalation and exhalation. The objective was to investigate production symmetry during breathing. Methods: The aerosol size distribution in exhaled breath was examined for different breathing patterns including normal breathing, varied breath holding periods and contrasting inhalation and exhalation rates. The aerosol droplet size distribution measured in the exhaled breath was examined in real time using an aerodynamic particle sizer. Results and Conclusions: The dependence of the particle concentration decay rate on diameter during breath holding was consistent with gravitational settling in the alveolar spaces. Also, deep exhalation resulted in a 4 to 6 fold increase in concentration and rapid inhalation produced a further 2 to 3 fold increase in concentration. In contrast rapid exhalation had little effect on the measured concentration. A positive correlation of the breath aerosol concentration with subject age was observed. The results were consistent with the breath aerosol being produced through fluid film rupture in the respiratory bronchioles in the early stages of inhalation and the resulting aerosol being drawn into the alveoli and held before exhalation. The observed asymmetry of production in the breathing cycle with very little aerosol being produced during exhalation, is inconsistent with the widely assumed turbulence induced aerosolization mechanism.

Expression of Potato virus Y cytoplasmic inclusion protein in tobacco results in disorganization of parenchyma cells, distortion of epidermal cells, and induces mitochondrial and chloroplast abnormalities, formation of membrane whorls and atypical lipid accumulation

Infection of plant cells by potyviruses induces the formation of cytoplasmic inclusions ranging in size from 200 to 1000 nm. To determine if the ability to form these ordered, insoluble structures is intrinsic to the potyviral cytoplasmic inclusion protein, we have expressed the cytoplasmic inclusion protein from Potato virus Y in tobacco under the control of the chrysanthemum ribulose-1,5-bisphosphate carboxylase small subunit promoter, a highly active, green tissue promoter. No cytoplasmic inclusions were observed in the leaves of transgenic tobacco using transmission electron microscopy, despite being able to clearly visualize these inclusions in Potato virus Y infected tobacco leaves under the same conditions. However, we did observe a wide range of tissue and sub-cellular abnormalities associated with the expression of the Potato virus Y cytoplasmic inclusion protein. These changes included the disruption of normal cell morphology and organization in leaves, mitochondrial and chloroplast internal reorganization, and the formation of atypical lipid accumulations. Despite these significant structural changes, however, transgenic tobacco plants were viable and the results are discussed in the context of potyviral cytoplasmic inclusion protein function.

Biochemical characterization of Aprataxin, the protein deficient in Ataxia with Oculomotor Apraxia type 1

Neurodegenerative disorders are heterogenous in nature and include a range of ataxias with oculomotor apraxia, which are characterised by a wide variety of neurological and ophthalmological features. This family includes recessive and dominant disorders. A subfamily of autosomal recessive cerebellar ataxias are characterised by defects in the cellular response to DNA damage. These include the well characterised disorders Ataxia-Telangiectasia (A-T) and Ataxia-Telangiectasia Like Disorder (A-TLD) as well as the recently identified diseases Spinocerebellar ataxia with axonal neuropathy Type 1 (SCAN1), Ataxia with Oculomotor Apraxia Type 2 (AOA2), as well as the subject of this thesis, Ataxia with Oculomotor Apraxia Type 1 (AOA1). AOA1 is caused by mutations in the APTX gene, which is located at chromosomal locus 9p13. This gene codes for the 342 amino acid protein Aprataxin. Mutations in APTX cause destabilization of Aprataxin, thus AOA1 is a result of Aprataxin deficiency. Aprataxin has three functional domains, an N-terminal Forkhead Associated (FHA) phosphoprotein interaction domain, a central Histidine Triad (HIT) nucleotide hydrolase domain and a C-terminal C2H2 zinc finger. Aprataxins FHA domain has homology to FHA domain of the DNA repair protein 5’ polynucleotide kinase 3’ phosphatase (PNKP). PNKP interacts with a range of DNA repair proteins via its FHA domain and plays a critical role in processing damaged DNA termini. The presence of this domain with a nucleotide hydrolase domain and a DNA binding motif implicated that Aprataxin may be involved in DNA repair and that AOA1 may be caused by a DNA repair deficit. This was substantiated by the interaction of Aprataxin with proteins involved in the repair of both single and double strand DNA breaks (XRay Cross-Complementing 1, XRCC4 and Poly-ADP Ribose Polymerase-1) and the hypersensitivity of AOA1 patient cell lines to single and double strand break inducing agents. At the commencement of this study little was known about the in vitro and in vivo properties of Aprataxin. Initially this study focused on generation of recombinant Aprataxin proteins to facilitate examination of the in vitro properties of Aprataxin. Using recombinant Aprataxin proteins I found that Aprataxin binds to double stranded DNA. Consistent with a role for Aprataxin as a DNA repair enzyme, this binding is not sequence specific. I also report that the HIT domain of Aprataxin hydrolyses adenosine derivatives and interestingly found that this activity is competitively inhibited by DNA. This provided initial evidence that DNA binds to the HIT domain of Aprataxin. The interaction of DNA with the nucleotide hydrolase domain of Aprataxin provided initial evidence that Aprataxin may be a DNA-processing factor. Following these studies, Aprataxin was found to hydrolyse 5’adenylated DNA, which can be generated by unscheduled ligation at DNA breaks with non-standard termini. I found that cell extracts from AOA1 patients do not have DNA-adenylate hydrolase activity indicating that Aprataxin is the only DNA-adenylate hydrolase in mammalian cells. I further characterised this activity by examining the contribution of the zinc finger and FHA domains to DNA-adenylate hydrolysis by the HIT domain. I found that deletion of the zinc finger ablated the activity of the HIT domain against adenylated DNA, indicating that the zinc finger may be required for the formation of a stable enzyme-substrate complex. Deletion of the FHA domain stimulated DNA-adenylate hydrolysis, which indicated that the activity of the HIT domain may be regulated by the FHA domain. Given that the FHA domain is involved in protein-protein interactions I propose that the activity of Aprataxins HIT domain may be regulated by proteins which interact with its FHA domain. We examined this possibility by measuring the DNA-adenylate hydrolase activity of extracts from cells deficient for the Aprataxin-interacting DNA repair proteins XRCC1 and PARP-1. XRCC1 deficiency did not affect Aprataxin activity but I found that Aprataxin is destabilized in the absence of PARP-1, resulting in a deficiency of DNA-adenylate hydrolase activity in PARP-1 knockout cells. This implies a critical role for PARP-1 in the stabilization of Aprataxin. Conversely I found that PARP-1 is destabilized in the absence of Aprataxin. PARP-1 is a central player in a number of DNA repair mechanisms and this implies that not only do AOA1 cells lack Aprataxin, they may also have defects in PARP-1 dependant cellular functions. Based on this I identified a defect in a PARP-1 dependant DNA repair mechanism in AOA1 cells. Additionally, I identified elevated levels of oxidized DNA in AOA1 cells, which is indicative of a defect in Base Excision Repair (BER). I attribute this to the reduced level of the BER protein Apurinic Endonuclease 1 (APE1) I identified in Aprataxin deficient cells. This study has identified and characterised multiple DNA repair defects in AOA1 cells, indicating that Aprataxin deficiency has far-reaching cellular consequences. Consistent with the literature, I show that Aprataxin is a nuclear protein with nucleoplasmic and nucleolar distribution. Previous studies have shown that Aprataxin interacts with the nucleolar rRNA processing factor nucleolin and that AOA1 cells appear to have a mild defect in rRNA synthesis. Given the nucleolar localization of Aprataxin I examined the protein-protein interactions of Aprataxin and found that Aprataxin interacts with a number of rRNA transcription and processing factors. Based on this and the nucleolar localization of Aprataxin I proposed that Aprataxin may have an alternative role in the nucleolus. I therefore examined the transcriptional activity of Aprataxin deficient cells using nucleotide analogue incorporation. I found that AOA1 cells do not display a defect in basal levels of RNA synthesis, however they display defective transcriptional responses to DNA damage. In summary, this thesis demonstrates that Aprataxin is a DNA repair enzyme responsible for the repair of adenylated DNA termini and that it is required for stabilization of at least two other DNA repair proteins. Thus not only do AOA1 cells have no Aprataxin protein or activity, they have additional deficiencies in PolyADP Ribose Polymerase-1 and Apurinic Endonuclease 1 dependant DNA repair mechanisms. I additionally demonstrate DNA-damage inducible transcriptional defects in AOA1 cells, indicating that Aprataxin deficiency confers a broad range of cellular defects and highlighting the complexity of the cellular response to DNA damage and the multiple defects which result from Aprataxin deficiency. My detailed characterization of the cellular consequences of Aprataxin deficiency provides an important contribution to our understanding of interlinking DNA repair processes.

Initiation of embryogenic cell suspensions of taro (Colocasia esculenta var. esculenta) and plant regeneration

Embryogenic callus was initiated by culturing in vitro taro corm slices on agar-solidified half-strength MS medium containing 2.0 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D) for 20 days followed by transfer to 1.0 mg/L thidiazuron (TDZ). Callus was subsequently proliferated on solid medium containing 1.0 mg/L TDZ, 0.5 mg/L 2,4- D and 800 mg/L glutamine before transfer to liquid medium containing the same components but with reduced glutamine (100 mg/L). After 3 months in liquid culture on an orbital shaker, cytoplasmically dense cell aggregates began to form. Somatic embryogenesis was induced by plating suspension cells onto solid media containing reduced levels of hormones (0.1 mg/L TDZ, 0.05 mg/L 2,4-D), high concentrations of sucrose (40–50 g/L) and biotin (1.0 mg/L). Embryo maturation and germination was then induced on media containing 0.05 mg/L benzyladenine (BA) and 0.1 mg/L indole-3-acetic acid (IAA). Histological studies of the developing embryos revealed the presence of typical shoot and root poles suggesting that these structures were true somatic embryos. The rate of somatic embryos formation was 500–3,000 per mL settledcell volume while approximately 60% of the embryos regenerated into plants.