Heather Faye Chapman

Comparison of five in vitro bioassays to measure estrogenic activity in environmental waters

Bioassays are well established in the pharmaceutical industry and single compound analysis, but there is still uncertainty about their usefulness in environmental monitoring. We compared the responses of five bioassays designed to measure estrogenic activity (the yeast estrogen screen, ER-CALUX, MELN, T47D-KBluc, and E-SCREEN assays) and chemical analysis on extracts from four different water sources (groundwater, raw sewage, treated sewage, and river water). All five bioassays displayed similar trends and there was good agreement with analytical chemistry results. The data from the ER-CALUX and E-SCREEN bioassays were robust and predictable, and well-correlated with predictions from chemical analysis. The T47D-KBluc appeared likewise promising, but with a more limited sample size it was less compelling. The YES assay was less sensitive than the other assays by an order of magnitude, which resulted in a larger number of nondetects. The MELN assay was less predictable, although the possibility that this was due to laboratory-specific difficulties cannot be discounted. With standardized bioassay data analysis and consistency of operating protocols, bioanalytical tools are a promising advance in the development of a tiered approach to environmental water quality monitoring.

Characterisation of atmospheric deposition as a source of contaminants in urban rainwater tanks

To characterise atmospheric input of chemical contaminants to urban rainwater tanks, bulk deposition (wet+dry deposition) was collected at sixteen sites in Brisbane, Queensland, Australia on a monthly basis during April 2007-March 2008 (N=175). Water from rainwater tanks (22 sites, 26 tanks) was also sampled concurrently. The deposition/tank water was analysed for metals, soluble anions and selected samples were additionally analysed for PAHs, pesticides, phenols, organic & inorganic carbon. Flux (mg/m(2)/d) of total solids mass was found to correlate with average daily rainfall (R(2)=0.49) indicating the dominance of the wet deposition contribution to total solids mass. On average 97% of the total mass of analysed components was accounted for by Cl(-) (25.0%), Na (22.6%), organic carbon (20.5%), NO(3)(-) (10.5%), SO(4)(2-) (9.8%), inorganic carbon (5.7%), PO(4)(3-) (1.6%) and NO(2)(-) (1.5%). For other minor elements the average flux from highest to lowest was in the order of Fe>Al>Zn>Mn>Sr>Pb>Ba>Cu>Se. There was a significant effect of location on flux of K, Sb, Sn, Li, Mn, Fe, Cu, Zn, Ba, Pb and SO(4)(2-) but not other metals or anions. Overall the water quality resulting from the deposition (wet+dry) was good but 10.3%, 1.7% and 17.7% of samples had concentrations of Pb, Cd and Fe respectively greater than the Australian Drinking Water Guidelines (ADWG). This generally occurred in the drier months. In comparison 14.2% and 6.1% of tank samples had total Pb and Zn concentrations exceeding the guidelines. The cumulative mean concentration of lead in deposition was on average only 1/4 of that in tank water over the year at a site with high concentrations of Pb in tank water. This is an indication that deposition from the atmosphere is not the major contributor to high lead concentrations in urban rainwater tanks in a city with reasonable air quality, though it is still a significant portion.

Habitat characteristics and eggshell distribution of the salt marsh mosquito, Aedes vigilax, in marshes in subtropical Eastern Australia.

Abstract Research at 10 locations in coastal subtropical Queensland, Australia, has shown that salt marshes contained heterogeneous distributions of eggshells of the pest and vector mosquito Aedes vigilax (Skuse) (Diptera:Culicidae). The eggshell distribution was related to specific vegetation assemblages, with a mix of the grass, Sporobolus virginicus (L.) Kunth (Poales: Poaceae), and the beaded glasswort, Sarcocornia quinqueflora (Bunge ex (Ung.-Stern) A.J. Scott (Caryophyllales: Chenopodiaceae), as significantly higher in eggshells than any other vegetation. There were also high numbers in the mix of S. virginicus with the arrowgrass, Triglochin striata Ruiz & Pavón (Alismatales: Juncaginaceae). Both mixed types are found in relatively wetter areas, despite very few eggshells being found generally in the low marsh. Most sites contained S. virginicus and eggshell locations were variable for this species alone. This was probably related to its life form variability in response to salinity and location on the marsh. Location on the marsh was important for eggshell distribution with most eggshells around the edges of pools and depressions, followed by, but to a significantly lesser extent, the marsh surface. Eggshells were fewest in the low marsh. Partition analysis resulted in a tree that simplified and summarised the factors important for eggshell distribution confirming the individual analyses. The potential effects of climate, sea level and other change are also briefly discussed in the context of likely changes to land cover and relative location on the marsh. For example, increased sea level may lead to low marsh conditions extending into higher marsh area with implications for oviposition and numbers of eggshells.

Development and evaluation of a species diagnostic polymerase chain reaction-restriction fragment-length polymorphism procedure for cryptic members of the Culex sitiens (Diptera : Culicidae) subgroup in Australia and the southwest Pacific

Abstract Members of the Culex sitiens subgroup are important vectors of arboviruses, including Japanese encephalitis virus, Murray Valley encephalitis virus and Ross River virus. Of the eight described species, Cx. annulirostris Skuse, Cx. sitiens Wiedemann, and Cx. palpalis Taylor appear to be the most abundant and widespread throughout northern Australia and Papua New Guinea (PNG). Recent investigations using allozymes have shown this subgroup to contain cryptic species that possess overlapping adult morphology. We report the development of a polymerase chain reaction–restriction fragment-length polymorphism (PCR-RFLP) procedure that reliably separates these three species. This procedure utilizes the sequence variation in the ribosomal DNA ITS1 and demonstrates species-specific PCR-RFLP profiles from both colony and field collected material. Assessment of the consistency of this procedure was undertaken on mosquitoes sampled from a wide geographic area including Australia, PNG, and the Solomon Islands. Overlapping adult morphology was observed for Cx. annulirostris and Cx. palpalis in both northern Queensland and PNG and for all three species at one site in northwest Queensland.

Population Structure and Dispersal of the Freshwater Mosquitoes Culex annulirostris and Culex palpalis (Diptera: Culicidae) in Papua New Guinea and Northern Australia

Abstract In 1995 and 1998, outbreaks of Japanese encephalitis (JE) virus occurred for the first time in Australia. Virus isolation from pools of mosquitoes indicated Culex annulirostris Skuse was the most likely vector. It was hypothesized that wind-blown mosquitoes introduced JE from Papua New Guinea to the Torres Strait and Cape York, northern Australia. This study used levels of genetic differentiation, as indirect evidence of dispersal of mosquitoes between the two continents. The results have demonstrated that in the region incorporating Western Province in PNG, the Torres Strait and Cape York Peninsula, Cx. annulirostris is represented by a panmictic population, indicating frequent widespread dispersal throughout the region of investigation. The closely related Cx. Palpalis Taylor was also present at some locations but the population structure of this species is uncertain. This supports the hypothesis that wind-blown Cx. annulirostris and possibly Cx. Palpalis could be a possible mechanism for introduction of JE virus into Australia.

Quantification of vitellogenin mRNA induction in mosquitofish (Gambusia affinis) by reverse transcription real-time polymerase chain reaction (RT-PCR)

Abstract A method to quantify induction of vitellogenin (Vtg) mRNA in adult male mosquitofish was developed. Male mosquitofish were exposed to 0, 1, 20 and 250 ng l−1 17β-oestradiol (E2) for 4 and 8 days in static exposures, and liver Vtg mRNA and 18S rRNA expression were quantified in duplex RT-PCR. Liver 18S rRNA expression was very consistent among individuals, and there was a highly significant increase in Vtg mRNA expression after exposure of mosquitofish for just 4 days at 250 ng l−1 E2. Lower doses did not induce Vtg mRNA expression even at 4 or 8 days. This method could be used as a rapid test to detect exposure of mosquitofish to oestrogenic chemicals. Further work is needed to determine if increased Vtg mRNA levels in male mosquitofish induce Vtg synthesis, and to determine the usefulness of the method in field sampling.

Analysis of polymeric quaternary ammonium salts as found in cosmetics by metachromatic polyelectrolyte titration

Metachromatic polyelectrolyte titration was investigated as a means of analyzing polymeric quaternary ammonium salts (polyquaterniums) of relatively low charge density used as ingredients in cosmetics. They are of possible toxicological concern, but little is known of their environmental fate and behavior. With o-toluidine blue as the indicator, determination of the concentration of a range of polyquaterniums of commercial importance and cosmetic origin was possible. Using a visual endpoint, normalities as low as 3 × 10−5 N could be determined, while with a spectrophotometric endpoint, the technique was effective to approximately 3 × 10−6 N. The concentrations (g L−1) to which these normalities correspond depend on the charge density of the polymer. Polyquaterniums are frequently used in association with anionic surfactants in cosmetic formulations and the presence of an anionic surfactant even in excess did not affect results although the color change was less stable. Titration in the presence of prepared humic acid was also possible.

Balancing the budget of environmental estrogen exposure: the contribution of recycled water

Estrogenic endocrine disrupting compounds (e-EDCs) are present in treated sewage and there is concern about estrogenicity of potable recycled water. However e-EDCs are also present in other environmental media and intake from water needs to be considered in relation to these other sources. The concentrations of 13 e-EDCs in foodstuffs and drinking water are reviewed, their predicted concentrations in recycled water are estimated, and the daily estrogenic intake as 17beta-estradiol equivalent (EEq) based on both in vitro and in vivo potencies is calculated as 1.39 and 9.65 microg EEq/d, respectively. Dietary intake accounts for more than 99.8% of that total, and more than 84.2% is due to phytosterols. Drinking 2 L of recycled water per day is expected to add 0.001 to 0.016 microg EEq/d based on in vitro and in vivo potencies, respectively. Exposure to e-EDCs in recycled water is therefore likely to be insignificant compared to current dietary intakes.

CONSTRUCTED WETLANDS FOR SEWAGE EFFLUENT TREATMENT AND MOSQUITO LARVAE AT TWO SITES IN SUBTROPICAL AUSTRALIA

ABSTRACT This study of 2 wetlands in subtropical Australia, constructed to treat sewage effluent, examined the relationships between dips positive for mosquito larvae and water quality, operational status of the system, vegetation, and nontarget macroinvertebrates. One site is inland and the other is close to the coast. Larvae of disease vector mosquitoes were present at various times in the wetlands, especially in summer and autumn. The proportion of early instars (1st and 2nd) was greater than that of later ones (3rd and 4th). Dissolved oxygen was negatively, and temperature was positively, associated with the proportion of dips containing larvae. For the coastal site we noted that larvae were more common during draw-down of water for maintenance and also as the system started to come online. Vegetation associated with larvae included dense Typha orientalis and algae. Where there were several types of plants, such as at the coastal site, plant density and water depth were not significantly related to larval presence. Where there were several types of macroinvertebrates there were fewer dips positive for larvae. To provide water treatment capacity and minimal mosquito production we concluded that design should include a variety of plant types, discouraging low dissolved oxygen (for example, by aeration) and ongoing maintenance should be carried out in winter or spring, when mosquitoes are fewer than in summer.