ABSTRACT This work examined the role of micro-organisms in the biogeochemical cycling of selenium within the benthic ecosystem of Lake Macquarie, a coastal lake in New South Wales with a history of anthropogenic heavy metal contamination. Certain micro-organisms possess the ability to oxidise or reduce selenium (Fleming and Alexander, 1973; Doran and Alexander, 1977), and microbial volatilisation of selenium from contaminated sediments and soils utilising naturally-occurring microflora has been shown in overseas research (Thompson-Eagle and Frankenberger, 1992) to be a potentially effective remediation strategy. In examining the impact of micro-organisms upon the oxidation state of selenium in Lake Macquarie sediments, this work also investigated and characterised selenium (and heavy metal) concentrations, speciation and geochemical phase associations (an indicator of potential bioavailability) in the sediments. Seven distinct bacterial species indigenous to Lake Macquarie were identified in this work with the ability to reduce selenium as selenite to elemental selenium, and selenium as selenate to organic forms of selenium, including volatile methylated selenium compounds. Metabolic parameters calculated for these organisms compared favourably with those reported in the literature by other researchers. Mixed populations of sediment micro-organisms were also isolated and studied in this work for their selenite and selenate reduction abilities. Total reduction of added selenite at levels up to 100 mg/L was recorded for a number of the organisms studied in this work. A maximum specific uptake rate for selenite of 3040 mgSe(IV).(gcells)-1.(h)-1 for one isolate (Shewanella putrefaciens) was determined, exceeding rates reported in the literature by other authors. Use of the indigenous micro-organisms from Lake Macquarie for the bioremediation of selenium containing waste streams was also examined in this work and selenium reduction in an immobilised cell reactor was demonstrated with such organisms. Concentrations, speciation, sediment core profiles and geochemical phase associations for selenium were determined for sediment samples collected at a variety of sites throughout Lake Macquarie and from Wyee Creek, a selenium-impacted fluvial input to the lake. The maximum concentration of selenium obtained in this work for the lake proper was 4.04 mg/kg, considerably lower than values reported over a decade ago (Batley, 1987) but consistent with reported reductions of selenium input into the lake from the lead-zinc smelter. Selective extraction methodology (Tessier et al. and BCR methods) studied geochemical phase association of selenium in Lake Macquarie sediments and found up to 44% of selenium was in bioavailable forms. Of interest and environmental concern was levels of selenium found in sediments of Wyee Creek, which previously received overflows from the ash dam associated with the Vales Point Power Station. Sediment selenium levels of up to 300 mg/kg were determined for this creek. These were an order of magnitude or more greater than those recorded for the lake itself and are of concern as to the potential impact on benthic organisms and those animals, including humans, who consume them. While this work can only provide a 'snapshot' of conditions within Lake Macquarie at the time of the sampling events recorded herein, it does make several important contributions to the understanding of selenium biogeochemistry in Lake Macquarie. These include: presentation of the hypothesis that selenium levels in surficial sediments being deposited in the north of the lake have decreased in recent years as a result of selenium reduction measures undertaken by the lead-zinc smelter; determination that up to 44% of selenium in surficial sediments from the lake is associated with sediment phases in which selenium has the potential to become remobilized and hence possibly bioavailable; and documentation of selenium concentrations in Wyee Creek, identifying the area as having selenium concentrations an order of magnitude or more greater than the lake itself. Concerning the role played by microorganisms in the biogeochemical cycling of selenium in Lake Macquarie, this work has: identified individual isolated and mixed cultures of bacteria that can reduce selenium as selenite to lower oxidation states; identified individual isolated and mixed cultures of bacteria that can reduce selenium as selenate to lower oxidation states; identified volatile methylated selenium compounds in the headspace gases of microorganisms reducing selenate; determined Minimum Inhibitory Concentrations for selenate and selenite for organisms isolated from Lake Macquarie; identified casein hydrolysate as a preferred carbon source for selenium reducing microorganisms from Lake Macquarie; and demonstrated that bioremediation of selenium contaminated waste streams using indigenous organisms from Lake Macquarie is feasible on the laboratory scale. Further research areas suggested by this work include: additional investigations of elevated selenium levels in Wyee Creek sediments; determination of the role of microbes in in-situ selenium reduction; and optimisation of selenium biotreatment/bioremediation of selenium-containing waste streams and sediments. In summary, this work, in rejecting the null hypothesis that the oxidation states of selenium in sediments from Lake Macquarie, NSW, are independent of microbial activity and accepting the alternate hypothesis that these oxidation states are not independent of microbial activity, contributes to the understanding of the role of microorganisms in the biogeochemical cycling of selenium, having applicability to both the specific ecosystem of Lake Macquarie, NSW, and also to selenium cycling in the environment in general. In addition, this work has identified selenium contamination in Wyee Creek, one of the fluvial inputs to Lake Macquarie, which was previously been undocumented in the literature and which may pose significant potential risk to humans and the ecosystem due to sediment selenium levels one or more orders of magnitude higher than those recorded in the lake itself. Finally, this work has also identified a number of microorganisms indigenous to Lake Macquarie with the ability to reduce selenium from toxic, mobile forms to less toxic, immobile or volatile forms, and these organisms have been shown to have the potential for use in treatment of selenium contaminated waste streams and also in the bioremediation of selenium-contaminated sediments.

ABSTRACT This work examined the role of micro-organisms in the biogeochemical cycling of selenium within the benthic ecosystem of Lake Macquarie, a coastal lake in New South Wales with a history of anthropogenic heavy metal contamination. Certain micro-organisms possess the ability to oxidise or reduce selenium (Fleming and Alexander, 1973; Doran and Alexander, 1977), and microbial volatilisation of selenium from contaminated sediments and soils utilising naturally-occurring microflora has been shown in overseas research (Thompson-Eagle and Frankenberger, 1992) to be a potentially effective remediation strategy. In examining the impact of micro-organisms upon the oxidation state of selenium in Lake Macquarie sediments, this work also investigated and characterised selenium (and heavy metal) concentrations, speciation and geochemical phase associations (an indicator of potential bioavailability) in the sediments. Seven distinct bacterial species indigenous to Lake Macquarie were identified in this work with the ability to reduce selenium as selenite to elemental selenium, and selenium as selenate to organic forms of selenium, including volatile methylated selenium compounds. Metabolic parameters calculated for these organisms compared favourably with those reported in the literature by other researchers. Mixed populations of sediment micro-organisms were also isolated and studied in this work for their selenite and selenate reduction abilities. Total reduction of added selenite at levels up to 100 mg/L was recorded for a number of the organisms studied in this work. A maximum specific uptake rate for selenite of 3040 mgSe(IV).(gcells)-1.(h)-1 for one isolate (Shewanella putrefaciens) was determined, exceeding rates reported in the literature by other authors. Use of the indigenous micro-organisms from Lake Macquarie for the bioremediation of selenium containing waste streams was also examined in this work and selenium reduction in an immobilised cell reactor was demonstrated with such organisms. Concentrations, speciation, sediment core profiles and geochemical phase associations for selenium were determined for sediment samples collected at a variety of sites throughout Lake Macquarie and from Wyee Creek, a selenium-impacted fluvial input to the lake. The maximum concentration of selenium obtained in this work for the lake proper was 4.04 mg/kg, considerably lower than values reported over a decade ago (Batley, 1987) but consistent with reported reductions of selenium input into the lake from the lead-zinc smelter. Selective extraction methodology (Tessier et al. and BCR methods) studied geochemical phase association of selenium in Lake Macquarie sediments and found up to 44% of selenium was in bioavailable forms. Of interest and environmental concern was levels of selenium found in sediments of Wyee Creek, which previously received overflows from the ash dam associated with the Vales Point Power Station. Sediment selenium levels of up to 300 mg/kg were determined for this creek. These were an order of magnitude or more greater than those recorded for the lake itself and are of concern as to the potential impact on benthic organisms and those animals, including humans, who consume them. While this work can only provide a 'snapshot' of conditions within Lake Macquarie at the time of the sampling events recorded herein, it does make several important contributions to the understanding of selenium biogeochemistry in Lake Macquarie. These include: presentation of the hypothesis that selenium levels in surficial sediments being deposited in the north of the lake have decreased in recent years as a result of selenium reduction measures undertaken by the lead-zinc smelter; determination that up to 44% of selenium in surficial sediments from the lake is associated with sediment phases in which selenium has the potential to become remobilized and hence possibly bioavailable; and documentation of selenium concentrations in Wyee Creek, identifying the area as having selenium concentrations an order of magnitude or more greater than the lake itself. Concerning the role played by microorganisms in the biogeochemical cycling of selenium in Lake Macquarie, this work has: identified individual isolated and mixed cultures of bacteria that can reduce selenium as selenite to lower oxidation states; identified individual isolated and mixed cultures of bacteria that can reduce selenium as selenate to lower oxidation states; identified volatile methylated selenium compounds in the headspace gases of microorganisms reducing selenate; determined Minimum Inhibitory Concentrations for selenate and selenite for organisms isolated from Lake Macquarie; identified casein hydrolysate as a preferred carbon source for selenium reducing microorganisms from Lake Macquarie; and demonstrated that bioremediation of selenium contaminated waste streams using indigenous organisms from Lake Macquarie is feasible on the laboratory scale. Further research areas suggested by this work include: additional investigations of elevated selenium levels in Wyee Creek sediments; determination of the role of microbes in in-situ selenium reduction; and optimisation of selenium biotreatment/bioremediation of selenium-containing waste streams and sediments. In summary, this work, in rejecting the null hypothesis that the oxidation states of selenium in sediments from Lake Macquarie, NSW, are independent of microbial activity and accepting the alternate hypothesis that these oxidation states are not independent of microbial activity, contributes to the understanding of the role of microorganisms in the biogeochemical cycling of selenium, having applicability to both the specific ecosystem of Lake Macquarie, NSW, and also to selenium cycling in the environment in general. In addition, this work has identified selenium contamination in Wyee Creek, one of the fluvial inputs to Lake Macquarie, which was previously been undocumented in the literature and which may pose significant potential risk to humans and the ecosystem due to sediment selenium levels one or more orders of magnitude higher than those recorded in the lake itself. Finally, this work has also identified a number of microorganisms indigenous to Lake Macquarie with the ability to reduce selenium from toxic, mobile forms to less toxic, immobile or volatile forms, and these organisms have been shown to have the potential for use in treatment of selenium contaminated waste streams and also in the bioremediation of selenium-contaminated sediments.

In response to concern that excessive macroalgal growth and accumulation was occurring in some inshore areas of Lake Macquarie, the distribution, abundance and seasonality of macroalgae was investigated in relation to nutrient input and power station cooling water. Macroalgal dry weight biomass was measured at ten sites on a monthly basis for two years, with an analysis of community structure conducted using Multi-Dimensional Scaling. The macroalgal community in Lake Macquarie was typical of those found in other New South Wales coastal lagoons, exhibiting considerable spatial and temporal variation. A close similarity was observed in macroalgal community structure at sites affected by urban nutrient input, these being characterised by a high biomass often attributable to only one or two species. These nuisance macroalgae were mostly green algae, which were almost entirely absent from other sites. Biomass at sites affected by urban nutrient input was generally within the range documented for eutrophic estuaries elsewhere. Analysis of macroalgal community structure showed no evidence of large-scale changes macroalgal communities attributable to the effects of power station cooling water except within 500m of the outfall. At sites affected by a 1-2??C temperature increase, community structure and the magnitude of the biomass were similar to sites deemed as being relatively free of human impact. A reduction in species diversity occurred only within the immediate discharge zone, where water temperatures were approximately 6??C above ambient temperatures. Excessive growth of nuisance macroalgal species was not observed at any of the sites influenced by power station cooling water. There were no distinct patterns in seasonality of macroalgal growth in this study, though the greatest biomass appeared to occur in spring. The irregular temporal variation in macroalgal growth suggests that the most significant factors affecting growth occur on a time scale of weeks to months. It is therefore likely nutrient input to the nearshore through surface runoff is an important influence on the distribution and abundance of macroalgae in Lake Macquarie. This emphasises the need to reduce nitrogen and phosphorus input from urban sources in Lake management.

This study examines the occurrence, distribution and storage of selenium in seagrass communities of Lake Macquarie, which is a heavily industrialised area of NSW, where notably high trace metal concentrations have been reported previously. Initially a suite of organisms was collected from a seagrass bed (Zostera capricornia) in the southeastern section of the lake to investigate the bioaccumulation and biomagnification of selenium. All organisms contained selenium, with sediment-dwelling organisms containing the highest Se concentrations. No consistent pattern of significant Se-metal correlations with Cu, Zn and Cd was found. Biomagnification of Se was evident, as concentrations increased from sediments and water to flora (algae and seagrass) to bivalves to Crustacea. However, this trend was not continued to the higher trophic groups of invertebrate predators and teleost fish. The bivalve Anadara trapezia was further studied. Intrinsic and extrinsic factors affecting the accumulation of Se were examined. Gender and mass were assessed in a single day study, followed by a temporal investigation of the effect of gender, reproductive cycle, temperature and salinity on Se accumulation. Gender had no effect on the accumulation of Se in A. trapezia. Se was present in all tissues (adductor, blood, foot, gills, intestine and mantle) and not immobilised or stored in a specific tissue, indicating that it plays a structural role in the tissues of A. trapezia. Se burden increased relative to size (shell length and dry mass) suggesting that Se is metabolically controlled within the organism. Se concentrations were found to fluctuate temporally because of: food availability in response to water temperatures; the reproductive cycle; and associated metabolic activities responding to temperature changes and food availability. Subcellular selenium associations in A. trapezia were examined to assist in the understanding of the fate of Se in marine tissues. Most of the Se was associated with proteins, suggesting that Se has a metabolic role in this marine organism. Proteins are intrinsically associated with the lipid bilayer of the cell membranes. A number of proteins (94, 85, 43, 36.5, 30, 23.4, 17.4 and 15 kDa) were separated by SDS PAGE from ethanol fractions. Determination of the Se concentration within individual proteins was not possible because the ratio of Se to protein was too low for further analysis. These findings indicated that Se plays a metabolic role in the tissues of the marine organism A. trapezia. The biochemical regulatory mechanism responsible for maintaining Se concentrations within the tissues is currently unknown.

In response to concern that excessive macroalgal growth and accumulation was occurring in some inshore areas of Lake Macquarie, the distribution, abundance and seasonality of macroalgae was investigated in relation to nutrient input and power station cooling water. Macroalgal dry weight biomass was measured at ten sites on a monthly basis for two years, with an analysis of community structure conducted using Multi-Dimensional Scaling. The macroalgal community in Lake Macquarie was typical of those found in other New South Wales coastal lagoons, exhibiting considerable spatial and temporal variation. A close similarity was observed in macroalgal community structure at sites affected by urban nutrient input, these being characterised by a high biomass often attributable to only one or two species. These nuisance macroalgae were mostly green algae, which were almost entirely absent from other sites. Biomass at sites affected by urban nutrient input was generally within the range documented for eutrophic estuaries elsewhere. Analysis of macroalgal community structure showed no evidence of large-scale changes macroalgal communities attributable to the effects of power station cooling water except within 500m of the outfall. At sites affected by a 1-2??C temperature increase, community structure and the magnitude of the biomass were similar to sites deemed as being relatively free of human impact. A reduction in species diversity occurred only within the immediate discharge zone, where water temperatures were approximately 6??C above ambient temperatures. Excessive growth of nuisance macroalgal species was not observed at any of the sites influenced by power station cooling water. There were no distinct patterns in seasonality of macroalgal growth in this study, though the greatest biomass appeared to occur in spring. The irregular temporal variation in macroalgal growth suggests that the most significant factors affecting growth occur on a time scale of weeks to months. It is therefore likely nutrient input to the nearshore through surface runoff is an important influence on the distribution and abundance of macroalgae in Lake Macquarie. This emphasises the need to reduce nitrogen and phosphorus input from urban sources in Lake management.

Lake Macquarie is the largest estuarine lake in New South Wales and is located on the eastern seaboard of Australia, approximately 85 km north of Sydney.The study of heavy metal concentrations in surficial sediments, sediment cores, seagrasses and fish conducted by the New South Wales Department of Mines in 1974 revealed that significant heavy metal contamination of Lake Macquarie has occurred. The metalloid selenium was found in elevated concentrations in 1987. Selenium has a complex chemistry and is both an important nutrient and in high concentrations toxic. This review describes briefly the Lake Macquarie environment and suggests some approaches of modelling the remobilisation and cycling of selenium in sediment. The biogeochemical cycling of selenium in an estuarine environment is exceedingly complex. Factors include Eh, pH, ligand complexing ability, solubility of selenium containing minerals, sediment/soil characteristics, microbially-mediated reactions and physical reworking by biota (bioturbation). The goal of the model will be to be able to estimate the transport rates from a calculated value of Se concentration in sediment pore water resulting in a Se flux value for the interface between sediment and overlaying water. / www.ima.kth.se

THESIS ORGANISATION This thesis is organised into nine chapters that include seven international and national publications (six accepted and one submitted for publication). The initial overview chapter outlines the justification and direction for this thesis. With the exception of chapter 8 (accepted for publication on the 1st May 2005); all chapters are exact duplicates of published articles in international and national refereed journals (chapters 2 to 7). The initial chapters (2 and 3) presents research findings using a marine fish species, mullet (Mugil cephalus), to measure trace metal bioavailability in Lake Macquarie, NSW Australia. While subsequent chapters (4 to 8) are presenting research under taken to improve the understanding of arsenic cycling in marine and estuarine environments. The final chapter (chapter 9) is a synopsis of the major findings presented in this thesis. Due to the publication nature of this thesis, an unavoidable degree of replication exists within chapters (publications).

Three approaches were employed to examine the effects of elevated sediment trace metal concentrations on estuarine/marine macrobenthic invertebrate assemblages. The initial study examined macroinvertebrate communities along a known polymetallic gradient, Lake Macquarie, NSW (gradient study). The second study experimentally tested if sediments sourced from different locations within Lake Macquarie differentially influenced the recolonisation of benthic invertebrates. The third study investigated the different recolonisation patterns of benthic invertebrates into sediments spiked with increasing concentrations of sediment-bound cadmium. In the Lake Macquarie gradient study, four locations (Cockle Bay, Warner's Bay, Kooroora Bay and Nord's Wharf) were sampled in winter 2000 and summer 2003 using a hierarchical design (location > site > plot). On both sampling occasions, the sediments showed strong gradients in lead, cadmium and zinc concentrations emanating from the Cockle Bay industrialised region in the lake's north, with concentrations being significantly lower in the most southern and less urbanised location (Nord's Wharf). In general, concentrations of lead, cadmium and zinc in the sediments increased among locations in the following order: Nord's Wharf > Kooroora Bay > Warner's Bay > Cockle Bay. AVSJSEM analyses indicated that in some sites in Cockle Bay, and to a lesser extent Warner's Bay, SEM concentrations exceeded their molar equivalence of AVS, indicating the potential for trace metals to be labile within the porewaters. Granulometry also changed along the gradient, with a higher proportion of silt/clay occurring in the locations with high metal concentrations. Conversely, the percentage of total organic carbon was higher in the less contaminated locations. In winter 2000, changes in benthic communities along the gradient supported the a priori hypotheses, with diversity and richness being greater in locations with lower concentrations of metals. Polychaetes were most numerous in Cockle Bay and Warner's Bay, whilst bivalves and gastropods were more abundant in Nord's Wharf and Kooroora Bay. Crustaceans were more numerous in Nord's Wharf; with all other locations having similar, lower, abundances. Ordination maps of the assemblages provided relatively clear separation of the assemblages among locations, with nonparametric multivariate analysis of variance (NPMANOVA) and subsequent pair-wise comparisons finding significant differences among the assemblages from all locations. SIMPER analyses found the highest level of dissimilarity was between the Nord's Wharf and Cockle Bay assemblages - primarily attributable to differences in the relative contributions of isopods; tellenid bivalves; and the polychaete families Spionidae, Opheliidae and Nephytidae. Weighted Spearman rank correlations (BIOENV) identified cadmium (Pw =0.74) as the strongest environmental (single or combination) variable to correlate with biotic assemblages. Benthic patterns along the gradient were less defined in summer 2003 due to a dramatic reduction in the abundance and diversity of fauna in Nord's Wharf. This decline was possibly attributable to a sustained reduction in salinity caused by a prolonged rainfall event. With the exception of Nord's Wharf, trends in the community indices and abundances of key taxa among the other locations were similar to those reported in winter 2000. Multivariate analyses discriminated the benthic assemblages from the four locations, with the findings from the NPMANOVA pair-wise comparisons indicating that the assemblages from all four locations were significantly different. SIMPER analyses showed the highest level of dissimilarity was between Nord's Wharf and Warner's Bay, with these differences being primarily attributable to their relative abundances of amphipods and polychaetes from the families Spionidae, Cirratulidae, Opheliidae and Capitellidae. BIOENV found that the combination of the sedimentary concentrations of cadmium and iron provided the best correlation (Pw =0.73) with biotic patterns, with similar correlations occumng with the addition of lead and its covariate, zinc (Pw =0.72). The combined findings from the gradient study established a strong correlation between trace metal concentrations within the sediments and suite of univariate and multivariate measurements. The low abundance and diversity of fauna in Nord's Wharf in the summer of 2003 highlighted the dynamic changes which can occur in the distributions of macrobenthic invertebrates. Although the study indicated that there was a strong relationship between trace metal concentrations and benthic community structure, the study was correlative, and requires subsequent experimental testing to confirm the causality of the observed relationships. The second component of the research was a translocation experiment using benthic recolonisation as an end-point. The experiment was performed to identify if the sediments, and not location, were influencing the composition of benthic assemblages in Lake Macquarie. Sediments were collected from three locations (Cockle Bay, Warner's Bay and Nord's Wharf), defaunated, and transplanted in three new locations along the south-east edge of the lake. At each location, 10 containers of each treatment were randomly placed in the sediment and allowed to recolonise for 22 weeks. Upon retrieval, the benthic communities were sampled and enumerated in conjunction with a variety of chemical and sedimentary measurements. Ten replicate invertebrate samples were also collected in the sediments adjacent to the experiment (ambient samples) at the completion of the experiment. Due to human interference, the containers from only two locations were analysed. Upon retrieval, pH and redox profiles of the sediments were similar to those expected in natural sediments. In general, concentrations of metals were low in the porewaters; however, iron precipitation on the porewater collection devices may have artificially increased the diffusion of metals, increasing concentrations near the sediment-water interface. Concentrations of SEM exceeded their AVS equivalence in some samples taken from the Cockle Bay and Warner's Bay treatments. Two-way ANOVAs found significant interactions between location and sediment treatments in diversity, evenness and the number of polychaetes, as well as significant differences in the number of capitellids and crustaceans among locations. Post-hoc comparisons of means found the Nord's Wharf sediment contained a higher mean number of individuals than the other treatments, including the ambient samples. nMDS ordination plots for both locations provided poor graphical discrimination of the assemblages among treatments; however, NPMANOVA detected significant location and treatment interactions. In both locations, pair-wise comparisons indicated that the assemblages within the Nord's Wharf treatments were significantly different to the Cockle Bay, Warner's Bay and ambient assemblages. No significant differences were detected between the Cockle Bay and Warner's Bay assemblages at either location. SIMPER analyses found the highest level of dissimilarity occurred between the ambient assemblages in Location 2 and the Nord's Wharf treatment, primarily due to the relative difference in the abundances of Capitellidae, Spionidae, Oweniidae, Nereididae and isopods among the assemblages. The findings from the translocation experiment suggest that the sediments are influencing the recolonisation of benthos. However, because differences were not detected between the Cockle Bay and Warner's Bay treatments, the approach used in the study shows potential as an in situ technique which could be used to assess the potential ecological risks of sediments fiom specific locations. Excluding cost and time considerations, the technique's primary disadvantage is the lack of a true control. As a result, the technique can only identify if the sediments are modifying benthic recolonisation, and not causality. The final component of the research experimentally tested if elevated concentrations of sediment-bound cadmium affected benthic invertebrate recolonisation. Sediments from the south coast of New South Wales (Durras Lake) were defaunated, and spiked with cadmium under anaerobic conditions to obtain three targeted cadmium concentrations: control (<O.1 ug/g), Low-Cd (15 Cd ug/g) and High-Cd (150 Cd ug/g). The physio-chemical properties of the waters and porewater concentrations of cadmium were monitored over a 28-day equilibration period, with declines in pH mediated with the addition of NaOH(aq). At the end of the equilibration period, porewater concentrations of cadmium were low in the Low-Cd and High-Cd treatments (maximum <l.5 ug/L in High-Cd), and below the detection limit in the control. Cadmium was not detected in the control sediments, with concentrations in the Cd-Low and Cd-High sediments exceeding their targeted concentrations, with final mean concentrations of 17 ug/g and 183 ug/g, respectively. The experimental design was similar to that employed in the translocation experiment, with 10 containers from each treatment transplanted into the sediments at three locations within Lake Macquarie. After 20 weeks, the containers were collected, along with benthic invertebrate samples from the ambient sediments. Data was not used from Location C due to extensive sediment deposition on the transplanted treatments. Significant declines occurred in the concentrations of cadmium in both the Low-Cd and High-Cd sediments, with the greatest loss occumng in the surficial sediments. The loss of cadmium was probably due to the differential loss of the fine fraction through physical means (hydrodynamic) rather than fluxing, as it assumed that the cadmium was primarily sediment-bound and relatively insoluble under anoxic conditions. Mean porewater concentrations of cadmium were below the detection limit in the control treatments; < 1 ug/L in the Low-Cd treatment, and generally < 2ug/L in the High-Cd, with the exception of some samples in Location B (maximum 5.6 ug/L) Concentrations of ammonia were low in the porewaters from the surficial sediments, with concentrations being significantly higher, and potentially toxic, in the anoxic porewaters (7 cm depth). In comparison to the previous recolonisation experiment, the number of individuals which recolonised the cadmium-spiked treatments was low, and significantly lower than the mean number of individuals sampled in the ambient sediments. No significant differences were detected among the treatments or locations (and their interactions) in diversity (H'), richness (d) or evenness (J). The number of polychaetes and molluscs significantly differed among the treatments, with post-hoc analyses indicating these differences were not among the cadmium-spike treatments, but were due to a greater mean abundance of these taxa in the ambient sediments. A significant interaction between treatment and location was detected in the mean abundance of crustaceans, with the ambient sediments having significantly lower mean abundances in both Location A and B. Ordination plots of the experiments in Location A and B provided poor graphical discrimination among the spiked treatments, although the ambient assemblages appear to be separated from the cadmium-spiked assemblages. NPMANOVA detected a significant interaction between treatments and locations, as well as among treatments. In both Location A and B, pair-wise analyses found the assemblages in the ambient sediments to be significantly different to the assemblages in all three cadmium treatments, with no differences being detected among the latter. SIMPER analyses found the highest levels of dissimilarity occurred between the spike-treatments and the ambient sediments, with these differences being primarily due to the relatively higher abundance of decapods in the spiked treatments, and capitellids in the ambient sediments. The cadmium-spiking component of the experiment clearly illustrated that artificially increasing the trace metal concentrations of metals in estuarine sediments is a complex process which needs to be performed in a methodological manner in order to obtain homogenous treatments with low porewater concentrations, and minimal artefacts. Furthermore, the results confirmed that the equilibration time for sediments can be extensive (several weeks), even in the case of organically rich sediments. The timing of the experiment (commenced late summer, February, 2003) appears to the major factor for the relatively low recolonisation rates, with the experiment missing the main larval recolonisation period between spring and early summer. Even in the highest treatment, elevated concentrations of cadmium did not appear to affect benthic recolonisation. This finding is supported by other experimental studies which suggest that concentrations of a single isolated metal must considerably exceed current guideline values (or contain high porewater concentrations) in order to elicit a biological effect. Nevertheless, as trace metals generally co-occur with other contaminants - with the response of multiple contaminants being possibly additive or synergistic - a conservative guideline value may be suitable in the interim as a precautionary measure. The findings of this thesis suggest that elevated concentrations of trace metal mixtures in estuarine sediments can affect the structure and composition of benthic communities; however, identifying causality is difficult. Although there has been an increase in the use of manipulative field experiments as a means of reducing the confounding influence of covariables found in field studies, this approach also has limitations, e.g. spatial and temporal scale issues, container effects, cost and biogeochemical changes to the sediments. Measuring stress at a community level is a fundamental component of estuarine risk assessment programs; and in isolation this approach can produce subjective and confounded findings. In order to accurately assess the risks associated with trace metal contaminated sediments, an integrated approach (e.g. weight of evidence) is required, one which uses multiple lines of evidence sourced from various chemical, environmental biological measurements.