The detection of microcontaminants in aquatic environments raises concerns about their potential to exert ecotoxicological effects and impact human health. In contrast to freshwater habitats, little information is available on environmental concentrations in urban estuarine and marine environments. This study investigated an extensive range of organic and inorganic microcontaminants in the Darwin Harbour catchment, a tropical estuary in northern Australia undergoing rapid urbanisation and industrial development. We sampled wastewater effluent and surface water from seven sites in Darwin Harbour for pharmaceuticals and personal care products, alkylphenols, hormones, pesticides, herbicides and metals. In vitro bioassays were used to estimate the (anti)estrogenic and (anti)androgenic activities of samples. Seventy-nine of 229 organic microcontaminants analysed were detected at concentrations ranging from 0.01 to 20 μg/L, with acesulfame, paracetamol, cholesterol, caffeine, DEET and iopromide detected at the highest concentrations in wastewater effluent (20 μg/L, 17 μg/L, 11 μg/L, 11 μg/L, 10 μg/L and 7.6 μg/L, respectively). Levels of estrogenic activity ranged from estradiol equivalency quotients (EEQs) of <0.10 to 6.29±0.16 ng/L while levels of androgenic activity ranged from dihydrotestosterone equivalency quotients (DHTEQs) of <3.50 to 138.23±3.71 ng/L. Environmental concentrations of organic microcontaminants were comparable to ranges reported from aquatic environments worldwide with sewage effluent discharges representing the dominant source of entry into Darwin Harbour. The measured concentration range of DEET was higher than ranges reported in previous studies.

A joint collaboration was undertaken to complement the existing Water Quality Monitoring Program in Darwin Harbour. The aim was to assess in situ environmental contaminants and their bioavailability at various sites around Darwin Harbour. Time integrated samplers known as semi-permeable membrane devices (SPMDs) for dissolved polycyclic aromatic hydrocarbons and diffusive gradients in thin films (DGTs) for labile dissolved metal species were deployed in four locations (Sadgroves Creek, Shell Island, Hudson Creek and West Arm) for a period of 10 days. In addition, at Sadgroves Creek and West Arm, biological monitoring was conducted using caged and wild caught barramundi (Lates calcifer). Several biomarkers, already established in barramundi, were evaluated and they included bile metabolites, cytochrome P450s and 7-ethoxyresorufin-O-deethylase (EROD) glutathione S-transferase (GSTs) and cholinesterase activity (ChE). Two other biomarkers were evaluated: vitellogenin in blood to assess potential impact from sewerage into Darwin Harbour, and sorbitol dehydrogenase (SDH), a potential biomarker of liver damage. Results highlight PAH concentrations at the West Arm site were below detection whilst PAH concentration at Sadgroves Creek, Hudson Creek and Shell Island were all similar. For labile dissolved metals, Aluminum, Manganese and Nickel were higher at impact sites as compared to the reference site. Mean concentrations of EROD activity, cytochrome P450, SDH activity, GST, ChE and vitellogenin were not significantly difference (p > 0.05) in barramundi from Sadgroves Creek and West Arm and they were comparable to basal levels measured in hatchery-reared barramundi (T = 0) used as a non-exposed reference. These results provided a baseline survey of PAHs and labile dissolved metals at several sites in Darwin Harbour, which compliments the data being generated from the Darwin Harbour Water Quality Monitoring Program. As development of Darwin Harbour progresses, impacts to the harbour can continue to be monitored and assessed in order to protect the environmental values for Darwin Harbour.

Mud crabs, Scylla serrata, were sampled from four estuaries (the Normanby, Herbert, Burdekin and Fitzroy Rivers) along the coast of northern Queensland, Australia, representing a pollution gradient from low to high contamination based upon previous chemical monitoring. Four biomarkers; glutathione-S-transferase (GST) activity, cholinesterase (ChE) inhibition and the urinary metabolite concentrations of naphthalene (NPH) and benzo-a-pyrene (BaP) were evaluated in S. serrata hepatopancreas, haemolymph and urine. Site-specific evidence of elevated GST activity and BaP metabolite concentrations and significant ChE inhibition was detected. Biomarker responses from this field study provide evidence of contaminant exposure of S. serrata from river estuaries along the coast of northern Queensland and indicate that further investigation is warranted. Based on the current results, and with further work on characterising the dose-response and seasonal variation in this species, mud crabs have great potential as indicator species for water quality and ecosystem monitoring programs across tropical coastal regions of Australia.

Vitellogenin (Vtg) is a yolk protein precursor that has been identified as a sensitive biomarker for exposure to estrogenic compounds. We evaluated specific monoclonal and polyclonal antibodies for reactivity with plasma Vtg from two Australian Perciformes, the tropical barramundi (Lates calcanifer) and the temperate black brearn (Acanthopagrus butcheri). Blood plasma from 17p-estradiol exposed (EA male barramundi (20 mg kg-1) and male black bream (2.5-5.0 mg kg-') were sent to Biosense Laboratories (Norway) for cross-reactivity testing using their extensive anti-Vtg antibody selection. Indirect ELISA results determined barramundi plasma displayed the highest binding affinities to ND-3G2 (monoclonal-Mab) and PO-1 (polyclonal-Pab). Black brearn was most cross-reactive with ND-1C8 (Mab) and PC2 (Pab). Next, plasma was assessed for Vtg induction in E2-closed (5 mg kg-1), hatchery-reared barramundi and black bream, versus a non-injected control group. Vtg production was assessed by Western blot and indirect ELISA using ND-3G2 and ND-1C8 Mabs, respectively. A prominent band was identified in the range of 100-200 kDa for all female black bream and for all E2-treated barramundi and black bream, males, which was confirmed as Vtg by Western blot. Indirect ELISA results for barramundi demonstrated highly significant differences in E2-closed fish as compared to control fish (Student t, P < 0.001). E2 male black bream were significantly different than control males (Student t, P < 0.001) and control and E2 females displayed highly significant differences (Student t, P < 0.001). These results indicate that exposure to 17p-estradiol induces significant Vtg production in males of the two Australian Perciformes, with potential use as a biomarker for exposure to estrogenic compounds. Crown Copyright (C) 2008 Published by Elsevier Ltd. All rights reserved.

Normal operation of oil well platforms results in the discharge of produced formation water (PFW). The expression of CYP1A, CYP2M1- and 2K1-like proteins was examined for use as possible biomarkers of PFW exposure. A pilot study on the Northwest Shelf of Australia had indicated that PFW contamination possibly contributes to induction of CYP1A-like proteins in Gold-Spotted Trevally (Carangoides fulvoguttatus). The pilot study samples were re-examined for CYP1A, and, in addition, CYP2K1/2M1-like proteins. In a subsequent caged fish study in the same location a second species, Stripey seaperch (Lutjanus carponotatus), caught at a clean site, were distributed to three caging sites in a PFW gradient from the Harriet A production platform: A (near-field), B (far-field) and C (a non-impacted reference site). Fish were sampled at time (T) T = 0, T = 3 and T = 10 days. Significant increases of CYP1A, one CYP2K1- and two CYP2M1-like proteins were noted at Site A at T = 10 d. For another CYP2K1-like protein, a significant increase was observed at Site A only at T = 3 d. These results support a previous study indicating that CYP1A protein is sensitive to PFW exposure. Importantly, statistically significant environmental induction of both CYP2M1- and CYP2K1-like proteins in tropical fish due to PFW exposure had not previously been described and induction of enzymes in the CYP2 family suggest new biomarkers for PFW. In addition, the novel response of one CYP2K-like protein requires further verification, but offers promise for improved monitoring of sub-lethal responses in marine organisms.

Normal operation of oil well platforms results in the discharge of “produced formation water” (PFW). The expression of CYP1A, CYP2M1- and 2K1-like proteins was examined for use as possible biomarkers of PFW exposure. A pilot study at the Harriet A production platform, on the Northwest Shelf of Australia, had indicated that PFW contamination possibly contributes to induction of CYP1A- and 2K1/2M1-like proteins in Gold-Spotted Trevally (Carangoides fulvoguttatus). In a subsequent caged fish study in the same location, Stripey seaperch (Lutjanus carponotatus) were caught at a clean site, then distributed to three caging sites: A (near-field), B (far-field) and C (a non-impacted reference site). Fish were sampled at time T=0, 3 and 10 days. Significant increases of CYP1A, one CYP2K1- and two CYP2M1-like proteins were noted at Site A at T=10. For another CYP2K1-like protein, a significant increase was observed at Site A only at T=3. Prevailing winds changed between days 6 and 8 of sampling, moving the surface water plume directly west, possibly impacting in situ PFW exposure. The results indicate that tropical fish CYP1A protein is sensitive to PFW exposure. Importantly, statistically significant environmental induction of both CYP2M1- and CYP2K1-like proteins in tropical fish due to PFW exposure had not previously been described and CYP2 family induction may represent possible new biomarkers (other than CYP1A) of PFW exposure. In addition, the novel fraction-specific response of CYP2K-like proteins requires further verification but offers promise for improved monitoring of sub-lethal responses in marine organisms. (Supported by Australian Institute of Marine Science, Apache Energy Ltd. and the Environmental Toxicology Research Program at The University of Mississippi).

The Australian Institute of Marine Science (AIMS) conducted a pilot study around the Harriet A oil production platform on the Northwest Shelf of Australia. We evaluated hepatic ethoxyresorufin-O-deethylase (EROD) activity, fluorescent aromatic compounds (FACs) in bile and immunodetection of CYP1A-like proteins in two Australian tropical fish species, Gold-Spotted Trevally (Carangoides fulvoguttatus) and Bar-Cheeked Coral Trout (Plectropomus maculatus) to assess exposure to petroleum hydrocarbons associated with produced formation water (PFW). Additionally, the incidence of hydrocarbon-degrading bacteria isolated from the liver and bile of all fish captured was examined. Low EROD activity was found in both species, with EROD activity in C. fulvoguttatus showing significant site differences. FACs and CYP1A protein levels in C. fulvoguttatus showed a clear trend in hydrocarbon exposure consistent with hydrocarbon chemistry data: Harriet A>Harriet C>reference site. P. maculatus showed elevated levels of FACs at Harriet A as compared to the reference site and demonstrated detectable levels of CYP1A-like proteins at these two sites. Hydrocarbon-degrading bacteria were found in the liver and bile of both species, yet there was no correlation by sites. Our results demonstrate that C. fulvoguttatus and P. maculatus have potential as indicator species for assessing the effects from exposure to petroleum hydrocarbons. Both FACs and CYP1A are providing warning signs that there is potential for biological effects on fish populations exposed to PFW around the Harriet A production platform.

The dose‐response relationship for hepatic 7‐ethoxyresorufin‐ O ‐deethylase (EROD) induction in barramundi ( Lates calcarifer ) was examined under controlled laboratory conditions for 15 d using farm‐reared barramundi. These results were compared with EROD activity measured in barramundi collected from two rivers catchments (impacted and nonimpacted) in northern Queensland, Australia. Barramundi were dosed by intraperitoneal injection with a known cytochrome P4501A (CYP1A) inducer, β‐naphthoflavone (β‐NF), at 5, 10, and 50 mg kg −1 using two controls: A vehicle control (corn‐oil injected) and an experimental control (no injection). The EROD induction occurred within 4 h in the 5, 10, and 50 mg β‐NF kg −1 exposures, reaching mean maximum activities of 88.6 (±51.9), 85.5 (±91.7), and 149.1 (±106.4) pmol min −1 mg protein −1 , respectively. Mean EROD activities remained low in the corn‐oil controls (2.1 ± 1.8 pmol min −1 mg protein −1 ) and experimental controls (5.3 ± 4.4 pmol min −1 mg protein −1 ) throughout the study. Barramundi demonstrated a rapid response curve, which was dose dependent (50 > 10 > 5 mg β‐NF kg −1 ) and decreased progressively over time from induction. Measurement of total cytochrome P450 content (nmol mg protein −1 ) was not dose dependent. The EROD activities from field‐collected barramundi from the Johnstone River (impacted) and Olive River (nonimpacted) suggest exposure to low‐level contaminants in the Johnstone River fish only. With more controlled laboratory and field studies, barramundi have the potential to become a major indicator species in assessing exposure to environmental contaminants in coastal areas throughout northern Queensland, Australia.