Kathryn L. Hassell

The salinity tolerance of freshwater macroinvertebrate eggs and hatchlings in comparison to their older life-stages: a diversity of responses - The salinity tolerance of freshwater macroinvertebrate eggs and hatchlings

The tolerance to high salinity of older life stage macroinvertebrates could underestimate the effects of increasing salinity on freshwater macroinvertebrates. The salinity tolerance of the eggs and/or hatchlings of Glyptophysa gibbosa (Planorbidae), Glyptophysa aliciae, Glacidorbis sp. (Glacidorbidae), a Glossiponiidae, Piona cumberlandis (Pionidae), and Chironomus sp. (Chironomidae) were determined. The salinity tolerances of younger life-stages of species studied here and elsewhere were then compared to salinity tolerances of their mature aquatic or dominant life-stage. A diversity of responses have been observed with some species having similar salinity tolerances in all life-stages examined, whilst the eggs or hatchlings of other species had salinity tolerances ranging from 4% to 88% of their older life stages. On present knowledge, this diversity of responses presents some difficulties for simple rules of thumb to approximate sensitivity of young life-stages of freshwater macroinvertebrates inferred from their dominant stage’s tolerance.

Hypoxia impairs embryo development and survival in black bream (Acanthopagrus butcheri)

Coastal environments are threatened by the increasing frequency, extent and severity of hypoxic events. Hypoxia affects vast areas around the world and often causes fish kills, reduced abundance, altered distribution, low benthic biomass and declines in fisheries. In Australia, many fisheries are based on sparid fishes and in the southern states black bream (Acanthopagrus butcheri) is important to both the recreational and commercial sectors. This species completes its entire life cycle in estuaries and annual recruitment is highly variable and very likely influenced by environmental conditions during the spawning season. In a laboratory-based experiment, fertilised black bream eggs (embryos) were exposed to five different levels of dissolved oxygen (DO). The DO levels were maintained in small test wells using nitrogen gas in a novel chamber design. Embryo development was assessed over a 2-day period and hatched larvae were observed until Day 2 post-hatch. Significant differences (p<0.05) were observed in embryonic development and survival as a function of DO level. In severely hypoxic conditions (30% saturation) survival to 1 day was reduced and no hatching occurred. In moderately hypoxic conditions (45-55%S), both precocious and delayed hatching was observed and hatch rates were reduced, whilst the number of hatched larvae with deformities increased, resulting in reduced larval lengths. No larvae survived to Day 2 post-hatch when held in hypoxic conditions (<55%S). This study demonstrates the detrimental effect that severe hypoxia can have on the early development of black bream which could result in reduced recruitment and lowered abundance. Other species that share similar early life histories may also be at risk.

Ecological evidence links adverse biological effects to pesticide and metal contamination in an urban Australian watershed

Summary: Aquatic ecosystems near urban areas are often ecologically impaired, but causative factors are rarely identified. Effects may be revealed by considering multiple lines of evidence at different levels of biological organization. Biological impairment is evident in the urban section of the Upper Dandenong Creek Catchment (Victoria, Australia). We assessed whether episodic sewage spills or other pollutants were the cause of poor ecological condition in the stream. The evidence evaluated included chemical and invertebrate assessments, caging studies of mudsnails Potamopyrgus antipodarum, antioxidant biomarkers and endocrine disruption-related endpoints in fish (Carassius auratus and Gambusia holbrooki) and toxicological studies with chironomids (Chironomus tepperi). A combination of metals and pesticides is likely to be affecting the aquatic fauna across all biological levels, with macroinvertebrate communities, P. antipodarum and C. tepperi populations and C. auratus individuals all ecologically impaired. Adverse alterations to aquatic fauna were consistently seen in Bungalook Creek and persisted downstream of this confluence into Dandenong Creek. In addition, chemical assessments and toxicity identification evaluation (TIEs) resulted in several point sources of both metals and pesticides being identified as origins of impairment. This contrasted with an expectation that adverse effects were likely to be associated with sewer-related pollution. As a consequence, target areas and specific pollutants were identified for remediation instead of an expensive sewer upgrade. Synthesis and applications. The results demonstrate that it is important to investigate biological effects in different taxa, in both the laboratory and field, to understand which stressors are causing adverse effects on faunal assemblages. When adverse effects are seen across multiple levels of biological organization and caused by the same pollutant from an identifiable source, there is a clear

Fluctuations in natural and synthetic estrogen concentrations in a tidal estuary in south-eastern Australia

Estuaries are often the final repositories for aquatic pollutants but how estuarine hydrology influences the availability of marine- and freshwater-derived pollutants is not well understood, particularly for micro-pollutants such as endocrine disrupting chemicals. To address this knowledge gap, this study measured natural and synthetic estrogen concentrations within the Little River, a tidal estuary in close vicinity to the major discharge point of Melbournes largest waste water treatment plant (WWTP), the Western Treatment Plant. Quantitative enzyme-linked immunosorbent assays (ELISA) were used to determine concentrations of natural estrogens (ES: ∑E1 (estrone), E2 (17β-estradiol), E3 (estriol)) and the synthetic estrogen, 17α-ethinylestradiol (EE2). The highest concentrations were measured in samples taken from the WWTP effluent discharge channel (29.0 ng/L and 0.35 ng/L, respectively). Within the estuary, concentrations of ES (2.25-23.16 ng/L) varied somewhat between locations and sampling periods (p < 0.05), however patterns were not consistent. Significant spatial variation was observed on only one sampling occasion, and likewise temporal variation was only observed once. In the upstream freshwaters, ES (2.95-7.26 ng/L) concentrations were lower than in the estuary, although their presence suggests an additional source of ES to the environment, most likely of agricultural origin. The EE2 concentrations measured in both the estuarine and freshwater areas were all low (mostly below 0.20 ng/L), which created difficulties in interpretation due to problems associated with trying to measure such low concentrations with confidence. However, some patterns did emerge, with EE2 concentrations exhibiting significant temporal and tidal variation (p < 0.05), with concentrations greatest during low and flooding (incoming) tides. Physico-chemical properties explained 30% of the variation in ES concentrations, whereby concentrations increased with decreasing pH and DO and increasing salinity. Given the higher concentrations observed during flooding tides and the association of higher estrogen concentrations with increased salinity and low DO, we suggest that estrogens might accumulate in estuarine bottom waters and upon disturbance from the incoming tidal flows, may be a contributing source of estrogens into the estuary. This study contributes the first comprehensive investigation of estrogen dynamics in an Australian estuary, and provides the foundation for further research aimed at identifying which compounds are present in estuarine waterways, where they are coming from and how their concentrations vary through space and time.

The assessment of vitellogenin as a biomarker of exposure to estrogenic compounds in two Australian perciformes

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 calcarifer) and the temperate black bream (Acanthopagrus butcheri). Blood plasma from 17beta-estradiol exposed (E2) male barramundi (20 mg kg(-1)) and male black bream (2.5-5.0 mg kg(-1)) 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 bream was most cross-reactive with ND-1C8 (Mab) and PO-2 (Pab). Next, plasma was assessed for Vtg induction in E2-dosed (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-dosed 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 17beta-estradiol induces significant Vtg production in males of the two Australian Perciformes, with potential use as a biomarker for exposure to estrogenic compounds.

Contaminant (PAHs, OCs, PCBs and trace metals) concentrations are declining in axial tissue of sand flathead (Platycephalus bassensis) collected from an urbanised catchment (Port Phillip Bay, Australia)

Concentrations of PAHs, OCs, PCBs and trace metals were determined in the white muscle of sand flathead Platycephalus bassensis collected at 6 locations in Port Phillip Bay during 2015. No PAHs, OCs or PCBs were detected in the white muscle of sand flathead at any of the locations, however measurable levels of As, Cu, Hg, Se and Zn were detected at all sites. Only As and Hg exhibited regional difference in white muscle concentrations, with As present only in a non-toxic organic form and Hg measured at levels that are comparable to levels reported in reference sites in other studies. All contaminants detected in the white muscle of sand flathead collected in Port Phillip Bay in 2015 were below Australian Food Standards guideline values, and by world standards, the Port Phillip Bay sand flathead population is considered minimally contaminated. Furthermore, tissue contaminant concentrations appear to be decreasing over time.

No evidence of exposure to environmental estrogens in two feral fish species sampled from the Yarra River, Australia: A comparison with Northern Hemisphere studies

Environmental estrogens originate from a variety of sources including sewage treatment plant (STP) effluents and adverse physiological effects (endocrine disruption) have been observed in several fish species sampled downstream of STP discharges. In this study we examined common carp (Cyprinus carpio) and roach (Rutilis rutilis) for signs of exposure to environmental estrogens in the iconic Yarra River, Melbourne, Australia. The Yarra River flows through the city of Melbourne and more than 2 million people live within the catchment. Two STPs discharge water into the Yarra River within the middle reaches, and the areas immediately downstream of these discharge locations were the focus of this study. Carp and roach were chosen as test species since both have been utilised extensively for endocrine disruption research throughout Europe, North America and Asia, and data from various international studies was used for comparison with the results of the present study. Neither species showed evidence of exposure to environmental estrogens, with no elevation of plasma vitellogenin levels in males and no incidence of intersex gonads. Most physiological endpoints in both species from this study were within ranges reported in carp and roach from reference sites in other studies, however some degenerative histological changes in both male and female gonads were observed. Surface water samples showed no estrogenic activity (measured by the yeast-estrogen screen, YES), but did display strong anti-estrogenic and weak androgenic activity (measured by the yeast-androgen screen, YAS). Whilst the results show no evidence of impacts from environmental estrogens in the Yarra River, the presence of both anti-estrogenic and androgenic activity in water samples, as well as some gonadal changes in carp is concerning and indicates that our focus needs to broaden, in order to look for biological impacts in resident fauna that might be due to environmental pollutants other than environmental estrogens.

Municipal wastewater effluent licensing: A global perspective and recommendations for best practice

Advances in wastewater treatment have greatly improved the quality of municipal wastewater effluents in many parts of the world, but despite this, treated wastewaters can still pose a risk to the environment. Licensing plays a crucial role in the regulation of municipal wastewater effluents by setting standards or limits designed to protect the economic, environmental and societal values of waterbodies. Traditionally these standards have focused on physical and chemical water quality parameters within the discharge itself, however these approaches do not adequately account for emerging contaminants, potential effects of chemical mixtures, or variations in the sensitivity and resilience of receiving environments. In this review we focus on a number of industrialised countries and their approach to licensing. We consider how we can ensure licensing is effective, particularly when considering the rapid changes in our understanding of the impacts of discharges, the technical advances in our ability to detect chemicals at low concentrations and the progress in wastewater treatment technology. In order to meet the challenges required to protect the values of our waterways, licensing of effluents will need to ensure that there is no disconnect between the core values to be protected and the monitoring system designed to scrutinise performance of the WWTP. In many cases this may mean an expansion in the monitoring approaches used for both the effluent itself and the receiving waterbody.

Using a multi-biomarker approach to assess the effects of pollution on sand flathead (Platycephalus bassensis) from Port Phillip Bay, Victoria, Australia

Hepatic gene expression and liver histology were examined in sand flathead (Platycephalus bassensis) from six locations in Port Phillip Bay, Victoria, Australia. Four sets of genes including thyroid-related genes (D1, D2, TTR, TRα and TRβ), metal metabolism-related genes (MT, MTF1, TF, Ferritin and FPN1), apoptosis-related genes (Diablo/SMAC1, Diablo/SMAC2 and CYP1A) and an endoplasmic reticulum stress biomarker gene (GRP78) were examined in female flathead using qRT-PCR. TRβ and Diablo/SMAC1 gene expression was significantly up-regulated in fish from all polluted sites compared to those from a reference site. The transcripts of TRα and FPN1 were significantly higher in flathead from Corio Bay, while the hepatic mRNA of TTR and GRP78 were significantly lower in those fish. Positive correlations were observed between Diablo/SMAC1 and CYP1A, D2 and TRβ, TRα and TRβ. This study demonstrates that application of pathway-based biomarker genes and histopathology can provide comprehensive information on the impact of environmental pollutants on fish.

Smell no evil: Copper disrupts the alarm chemical response in a diadromous fish, Galaxias maculatus

Fish, at all life stages, utilize olfactory information in the decision-making processes essential to survival. Olfaction is a sensitive sensory process, and toxicants within urban aquatic environments can have destructive or depreciating effects. In the present study, the authors exposed Galaxias maculatus, a native fish commonly found in urban waterways throughout southeastern Australia, to 1 of 5 ecologically relevant copper (II) chloride concentrations (<1 μg/L, 1 μg/L, 6 μg/L, 8 μg/L, 18 μg/L) for 16 h. After exposure, the authors tested the response of individual fish to 1 of 3 stimuli: a conspecific skin extract containing a stress-inducing alarm chemical odor, a conspecific odor, and distilled water as a control. Stress responses were quantified through behavioral assays. The authors found evidence for distinct changes in behavioral response with increasing copper concentration and a marked difference in response between control fish and fish exposed to the alarm chemical odor. Copper, even at relatively low concentrations, can have a significant effect on the stress response behavior shown by G. maculatus. Environ Toxicol Chem 2016;35:2209-2214.