Michelle A Templeman
email: michelle.templeman@jcu.edu.au

Michael J Kingsford

Trace element accumulation in Cassiopea sp. (Scyphozoa) from urban marine environments in Australia

Keywords: jellyfish, Cassiopea, trace metals, bio-uptake, Australia, biomonitor

Jellyfish are robust, short-lived animals, tolerant to a wide range of environmental conditions and pollutants. The benthic jellyfish, Cassiopea sp. was collected from five locations along the north eastern coast of Australia and analysed for trace elements to determine if this species has potential as a marine biomonitor. Both the oral arm and bell tissues readily accumulated aluminium, arsenic, barium, cadmium, chromium, copper, iron, manganese and zinc above ambient seawater levels. In contrast, lithium appeared to be actively regulated within the tissues while calcium, magnesium and strontium reflected the ambient environment. The multi-element signatures showed spatial variation, reflecting the geographical separations between locations, with locations closer together showing more similar elemental patterns. The combination of bioaccumulative capacity, life history traits and 10 biophysical aspects indicate that this species has high potential as a biomonitor in coastal marine systems.

Introduction

Many trace metals have important cellular and tissue functions within organisms. However, excessive concentrations of some elements can have a detrimental effect on the environment and its inhabitants [START_REF] Sunda | Processes regulating cellular accumulation and physiological effects: Phytoplankton as model systems[END_REF]. Heavy metals and organic compounds that have no known biological function can also be taken up and concentrated in biological tissue [START_REF] Wright | The effect of salinity on cadmium uptake by the tissues of the shore crab, Carcinus maenus[END_REF].

Although a large body of research exists on the levels, fluxes and cycling of chemicals in marine environments (eg [START_REF] Furness | Heavy metals in the marine environment[END_REF][START_REF] Sadiq | Toxic metal chemistry in marine environments[END_REF], concentrations of contaminants in water do not necessarily describe their ecological significance to biota.

The use of organisms as biomonitors for trace metals and environmental contaminants is a wellestablished practice in both terrestrial and aquatic environments. The extent to which any particular organism can serve as a biomonitor depends on a number of factors including the system under investigation, life history traits of the organism, the specific contaminants being monitored and the extent of uptake and retention of those contaminants (Rainbow & Phillips 1993).

Gelatinous zooplankton (including jellyfish) can comprise a significant proportion of the total biomass of pelagic marine assemblages [START_REF] Shenker | Carbon content of the neritic scyphomedusa Chrysaora fuscescens[END_REF][START_REF] Romeo | Importance of gelatinous plankton organisms in storage and transfer of trace metals in the northwestern Mediterranean[END_REF]. Jellyfishes can tolerate a wide range of environmental and water quality conditions [START_REF] Lucas | Reproduction and life history strategies of the common jellyfish, Aurelia aurita, in relation to its ambient environment[END_REF][START_REF] Purcell | Jellyfish blooms: ecological and societal importance. Report No. MASGC-W-00-001, Mississippi-Alabama Sea[END_REF]. They are short-lived generally, capable of rapid population explosions, high growth rates and plasticity of resource allocation. As a consequence, they can have detrimental economic, health and aesthetic effects in coastal systems [START_REF] Graham | Ecological and economic implications of a tropical jellyfish invader in the Gulf of Mexico[END_REF][START_REF] Uye | Population biomass, feeding, respiration and growth rates, and carbon budget of the scyphomedusa Aurelia aurita in the inland Sea of Japan[END_REF], although they are valuable in some regions as a fisheries resource [START_REF] Kingsford | Management of jellyfish fisheries, with special reference to the order Rhizostomeae[END_REF][START_REF] Pitt | Temporal and spatial variation in recruitment and growth of medusae of the jellyfish Catostylus mosaicus (Scyphozoa: Rhizostomeae)[END_REF].

Although data is limited on the capacity of jellyfish to take up metals, the available information suggests they are capable of accumulating metals and nutrients above ambient seawater concentrations and potentially transferring these further up the foodchain. [START_REF] Heymans | A carbon flow model and network analysis of the northern Benguela Upwelling system, Namibia[END_REF][START_REF] Kingsford | Management of jellyfish fisheries, with special reference to the order Rhizostomeae[END_REF][START_REF] Fukuda | Potential dietary effects on the fatty acid composition of the common jellyfish Aurelia aurita[END_REF][START_REF] Fowler | Applied radiotracer techniques for studying pollutant bioaccumulation in selected marine organisms (jellyfish, crabs and sea stars)[END_REF][START_REF] Hay | Marine ecology: Gelatinous bells may ring change in marine ecosystems[END_REF]. [START_REF] Caurant | Bioaccumulation of Cadmium, Copper and Zinc in some Tissues of Three Species of Marine Turtles Stranded Along the French Atlantic Coasts[END_REF] linked elevated cadmium levels in tissues of certain turtle species with their jellyfish prey. [START_REF] Romeo | Importance of gelatinous plankton organisms in storage and transfer of trace metals in the northwestern Mediterranean[END_REF] also identified that gelatinous zooplankton may have a role in bioconcentration of metals up the food chain.

Although previously not considered as biomonitors, scyphozoan jellyfish possess many features considered important in biomonitors. Rainbow & Phillips (1983) identified a number of features that were essential for an organism to be considered a suitable biomonitor. These include the capacity to accumulate metals, sedentary behaviour, ease of identification, presence at locations of interest, and tolerance to physico-chemical changes within their environment (Rainbow & Phillips 1993).

The benthic scyphozoan jellyfish, Cassiopea spp. is euryhaline, tolerant of physio-chemical changes to its environment, is readily identifiable, can be present at high densities, has a wide distribution in tropical coastal environments and due to its benthic nature is relatively sedentary, and as such meets the biomonitoring criteria set out above. Cassiopea spp. are typically found in sheltered locations (eg man-made lagoons and lakes, inshore seagrass meadows, mangroves, protected reefs, etc) and exhibit atypical behaviour of resting upside down on the substrate giving rise to their common name -'Upside Down Jellyfish'. They possess an endosymbiotic relationship with Symbodium spp. and thus are capable of photosynthesis, which is associated with demonstrated efficiencies in uptake of inorganic nutrients and elements from the surrounding water column [START_REF] Estes | Localization and Quantification of Carbonic Anhydrase Activity in the Symbiotic Scyphozoan Cassiopea xamachana[END_REF][START_REF] Fowler | Applied radiotracer techniques for studying pollutant bioaccumulation in selected marine organisms (jellyfish, crabs and sea stars)[END_REF].

The objective of this study was to compare trace element concentrations in bell and oral arm tissues of Cassiopea sp. collected from urban waterbodies across eastern and northern Australia. Both essential trace elements and priority pollutants were measured and compared to ambient seawater concentrations from the collection sites to determine if Cassiopea sp. were capable of accumulating trace elements. Trace element signatures in jellyfish and seawater were also compared among locations to determine the potential for jellyfish to provide time-integrated measures of local conditions.

Materials & Methods

Cassiopea sp. and water samples were collected from five locations adjacent to urban areas: Myora Drain A & B on the Gold Coast; Lake Magellan A & B on the Sunshine Coast in Queensland; and Lake Alexander in Darwin, Northern Territory, Australia. Locations A & B for Myora Drain & Lake Magellan were separated from each other by approximately 1 -1.5 km. Five jellyfish and two water samples were collected at each location (Table 1).

INSERT TABLE 1

Specimen Handling

Following collection, jellyfish were rinsed with local water to remove any visible sediment or other material adhering to the bell or oral arms. The oral arms were separated from the bell by hand and placed in either clean, acid washed plastic resealable bags or plastic vials. Tissue samples were frozen as soon as practicable and kept at -18 0 C until they were digested.

Replicate water samples (50 -100 ml) were collected and filtered through a 0.45µ m syringe filter and stored in acid washed vials. Water samples were acidified with Suprapur grade nitric acid (HNO 3 ) to 2% and stored at 4 0 C until analysed.

Tissue Processing & Analysis

Approximately 3 -5 g tissue was accurately weighed and digested in five millilitres of concentrated (69%) Suprapur grade HNO 3 . Samples were digested for approximately 2 hours, allowed to evaporate to approximately five millilitres. Three to five millilitres of AR grade hydrogen peroxide was added to remove any residual organic carbon. Following digestion, the tissue was made up to 25 millilitres with Milli-Q water. All equipment and containers were pre-cleaned and soaked in 10% HNO 3 , triple rinsed in deionised water and air dried in a Class 100 laminar flow unit before use.

Samples were analysed using Varian 820-MS ICP-MS and Varian Liberty Series II AES. ICP-MS was used to determine aluminium (0.5) ** , arsenic (1), barium (0.1), copper (0.1), cadmium (0.05), chromium (1), lithium (0.1), manganese (2), lead (0.05), strontium (0.5), silver (0.1g) and zinc (2), while ICP-AES was used to measure calcium (10), magnesium (10) and iron (10). Elements were selected based on either their importance as essential biological elements or their consideration as anthropogenic or priority pollutants. Due to issues with signal suppression, it was necessary to dilute seawater samples 1:10 (seawater:diluent) prior to analysis.

Subsets of samples were spiked with known concentrations of all elements for quality control purposes and to determine recoveries (90-117%). Indium, gallium and yttrium were used as internal standards to correct for potential instrument drift and matrix effects. Analytical data was checked to ensure signal strength exceeded three standard deviations for all analyses. Digestion blanks were included to ensure integrity of the digestion process. Digestion blanks had low levels of contamination and tissue data was corrected for blank results before statistical analysis.

Statistical Procedures

Prior to statistical analysis of tissue data, seawater results that were below detection were recalculated to half the reported detection limit (Section 2.2). Elements below detection in seawater samples were included at half the reported detection limit in statistical analyses.

Analytical data was tested for homogeneity of variance (Bartlett's Test). All tissue data was log 10 transformed to meet assumptions of normality. Univariate data was analysed using Statistica Version 8.1. Univariate data among locations were analysed using a one-way ANOVA. Comparisons between tissue type (bell, oral arm) were undertaken using a pair-wise two-tailed t-test. Correlations between tissue elements and tissue -water relationships were also performed using Statistica Version 8.1.

Bioconcentration plots were prepared by calculating the bioconcentration factor for each metal using ratio of tissue metal concentration in tissue to the seawater metal concentration at each location [START_REF] Sadiq | Toxic metal chemistry in marine environments[END_REF]. Principal Component Analyses (PCA) were performed using SYSTAT Version 10 (Crane Software) after log 10 transformation to describe spatial variation in multi-element signatures.

Results

Element Concentrations Between & Within Tissues

Trace element concentrations varied among locations and between tissues for most elements (Table 2). Barium and zinc concentrations were significantly higher in oral arm tissues than bell tissue at all locations. Mean concentrations of manganese, arsenic, aluminium and cadmium were also ** detection limits for elements (ug/L) higher in oral arm tissue than bell tissue at all locations but the differences were not always significant (Table 2). Concentrations of copper, iron and strontium were higher in oral arm tissue at some locations and higher in bell tissue at others, although higher bell concentrations of elements were not significant. No significant differences were found between tissues for chromium, calcium, magnesium and lithium. There was no significant relationship between animal size and element concentration, thus size did not confound the spatial comparisons.

There were significant inter-element relationships in both bell and oral arm tissue, although these patterns were not always consistent between the tissues (Table 3). There was a very significant positive correlation between Ca-Mg (r = 0.96 bell; r = 0.95 oral arm) in both tissues. Ca-Cd (r = 0.73 bell; r = 0.52 arm) and As-Mn (r = 0.67 bell; r = 0.70 arm) were also positively correlated in both tissues. Other significant positive correlations in both tissues included As-Ba, Ba-Mn, Cr-Zn, Cd-Mg and Mn-Zn (Table 3). In contrast, Cr-Li was negatively correlated in both tissues (r = -0.54 bell; r = -0.52 arm).

For many other elements, the relationships were different for element pairs in the two tissues, with many being significantly correlated in one tissue but not the other (Table 3). Aluminium had significant correlations with a number of elements in the bell tissue but was only significantly correlated with zinc in oral arm tissue. Strontium also correlated positively with a number of other elements in the oral arm tissue but was only negatively correlated with iron in the bell tissue (Table 3).

Zinc was also correlated with a number of elements in both tissues but the relationships were not consistent between tissues (Table 3). Barium had a number of negative correlations with other elements although again, these patterns varied between the tissues.

INSERT TABLE 2 INSERT TABLE 3

Comparisons of Tissue and Ambient Seawater Concentration

Most elements were detected in higher concentrations in the jellyfish tissues by at least one order of magnitude compared to ambient seawater (Figure 1). Lithium was the only element found in Cassiopea sp. at concentrations below that of seawater. This was consistent across all locations and indicated that the animals were potentially regulating uptake and/or excretion of this element. The relative patterns of element accumulation were similar among locations although the degree of accumulation for some elements (particularly Al, Cd, Mn and Fe) was higher at Lake Alexander than the other locations (Figure 1). Tissue concentrations of calcium, magnesium and strontium were similar to ambient seawater concentrations at all locations. With the exception of magnesium in oral arm tissue and water (p < 0.05, n=5, r=0.94) there were no significant correlations between tissue concentrations of any given metal with the corresponding water concentrations, indicating a lack of regulation in elemental uptake.

Tissue levels of lead and silver were below detection limit in most animals and all waters.

Measurable levels of silver were only found in eight animals at tissue concentrations <2 ug/kg (wet weight) while lead was found in only seven animals with concentrations less than 10 ug/kg (wet weight).

INSERT FIGURE 1

Spatial Variation in Tissue Elements

There were significant differences in tissue concentrations among locations and between tissues for most elements (Tables 4 &5). Lithium and chromium were the only elements that were not significantly different among locations for either tissue type. Copper exhibited significant differences among locations for bell tissue but no significant difference among locations for oral arm tissue, while strontium showed the opposite effect (Tables 4 &5). Variation in barium concentrations was observed at large spatial scales in oral arm tissue but not at local scales (ie between locations A & B

for Myora Drain and Lake Magellan). This relationship was not as robust in the bell tissue (Table 5).

Bell tissue iron concentrations were significantly different among locations; however, post-hoc tests could not discriminate between them (Table 5).

Samples from Myora Drain A & B had similar elemental concentrations with no significant differences in bell tissue for any element (Table 5). Iron and arsenic were the only elements significantly different in oral arm tissue between the two locations. The similarity in concentration patterns between locations in close proximity to each other reflects a system with a short water residence time and regular tidal flushing.

In contrast, the patterns of element concentration for Lake Magellan A & B varied by element and tissue type. Tissues were significantly different in calcium and magnesium. Arsenic, cadmium, iron and strontium concentrations were significantly different in oral arm tissue, while bell tissue was significantly different for copper between the two locations (Tables 2 &5). The variation in calcium and magnesium concentrations may reflect changes in salinity within the lake between collection dates. Differences in Ca and Mg were also measured in the water samples at these locations.

Differences in the concentrations of other elements may be due to the long water residence time in the system (Table 1).

INSERT TABLE 4

Tissue results from Lake Alexander demonstrated no consistent relationship with any other location (Table 5). Significant differences could be identified between Lake Alexander and individual locations in both Myora Drain and Lake Magellan but there were no consistent patterns among the elements. Post-hoc tests separated Lake Alexander from at least one other location for all elements except barium and copper (in bell tissue), and manganese and strontium (in oral arm tissue). Overall, variability was least between locations in closest proximity (eg Myora Drain A & B), with greater variation among locations with the greatest geographical separation. The rate of flushing and length of water residence time is also likely to have influenced variation in tissue concentrations among locations.

INSERT TABLE 5

Patterns in Elemental Fingerprints

Differences in elemental fingerprints were found among locations in both bell and oral arm tissue.

Multivariate analysis (PCA) of elements in bell tissue indicated variation among locations was greater for those locations with a wider geographical separation (Figure 2A). Some variation among replicate samples within locations was found in all locations. Total variance in the matrix was 70.4% explained by Factor 1 (41.7%) and Factor 2 (28.7% were also very similar although there was greater spread within the Lake Magellan A samples. Lake

Alexander was separate to all other locations (Figure 2A).

Multi-element patterns in oral arm tissue were not as discrete as bell tissue (Figure 2B). Overall the total variation in the matrix for oral arms was 65.7% with 35.1% explained by Factor 1 and 30.6%

explained by Factor 2. Drain and Lake Magellan locations remained reasonably distinct from each other. Unlike the bell tissue, the oral arm elemental signature from Lake Alexander reflected an intermediate pattern between Myora Drain and Lake Magellan (Figure 2B). This suggests that different processes were influencing uptake and concentration of elements in oral arm compared with bell tissue.

INSERT FIGURE 2 4 Discussion

Trace Element Uptake and Accumulation

Studies have demonstrated that jellyfishes and other gelatinous plankton are capable of absorbing trace elements from the environment in measurable concentrations [START_REF] Romeo | Importance of gelatinous plankton organisms in storage and transfer of trace metals in the northwestern Mediterranean[END_REF][START_REF] Ki | Occurrence of organo-arsenicals in jellyfishes and their mucus[END_REF][START_REF] Fowler | Applied radiotracer techniques for studying pollutant bioaccumulation in selected marine organisms (jellyfish, crabs and sea stars)[END_REF]. However, data on bioaccumulation in scyphozoan and other jellyfishes is limited (Table 6).

Investigations into the extent of uptake and / or accumulation of elements from the surrounding water by jellyfish are even more limited. [START_REF] Cimino | Trace elements in tentacles from the jellyfish Pelagia noctiluca[END_REF] found tissue concentrations between 100 times (Al, As, Cd, Co, Hg, Mn, Pb, Sn & Zn) and 10 000 times (Fe) greater than typical seawater concentrations in tentacles of Pelagia noctiluca. The current study showed that Cassiopea sp. accumulated iron and aluminium at concentrations greater than 1000 times ambient seawater concentrations. Other metals including copper, zinc, cadmium, manganese and arsenic were accumulated greater than 10 fold above ambient seawater concentrations (Figure 1), while lithium seemed to be actively regulated in the tissues.

Tissue copper concentrations in Cassiopea sp. were comparable to the literature for other jellyfish (Table 6), although accumulation in this study was lower than levels reported for other marine organisms (eg Rainbow & Phillips 1993). This may reflect the high water content in jellyfish and the different metabolic requirements of other phyla (eg molluscs, crustaceans).

Zinc is a key component of the metalloenzyme carbonic anhydrase. Carbonic anhydrase is particularly abundant in organisms with a symbiotic association with photosynthetic algae [START_REF] Furla | Involvement of H+-ATPase and carbonic anhydrase in inorganic carbon uptake for endosymbiotic photosynthesis[END_REF] and the presence of zooxanthellae may therefore be an important contributor to tissue concentrations of zinc within symbiotic jellyfish. Concentrations of zinc measured in tissues from this study were higher than reported literature values for non-symbiotic jellyfish species for both oral arm and bell tissues (Table 6). This is consistent with the observation that zooxanthellae potentially influence zinc concentrations in the jellyfish. Zinc concentrations were below detection in waters at all locations indicating that Cassiopea sp. must either be efficient at obtaining zinc from food and water, or recycling zinc.

INSERT TABLE 6

The uptake and accumulation of non-essential elements in this study was variable. Chromium was present in low concentrations in the water but present at higher levels in the tissues, while arsenic and cadmium, although below detection in the seawater, were found in measurable concentrations in all animals (Table 2). This suggests that tissue concentrations of these metals may reflect a timeintegrated measure of metal concentrations in the water (either from pulse or press events).

Cadmium generally correlates with phosphate in seawater [START_REF] Lares | Temporal variability of bioavailable Cd, Hg, Zn, Mn and Al in an upwelling regime[END_REF] and uptake of this element may be linked with nutrient levels at the sampling locations. Phosphate was not measured in this study but there were no obvious signs of eutrophication (eg colour, algal blooms etc) at any location during sampling that would suggest pulses or presses of nutrients including phosphates.

Tissue concentrations of arsenic vary among scyphozoan jellyfish and differences were found between tissues in this study and others (Table 6). Investigations of arsenic in Mastigias papua [START_REF] Ki | Occurrence of organo-arsenicals in jellyfishes and their mucus[END_REF] found that levels in oral arm tissue containing zooxanthellae were higher than bell tissue but lower than levels in gastric filaments and gonads. Hanaoka et al (2001) also found similar differences between umbrella (bell) tissues and other tissues in other asymbiotic jellyfish species. The arsenic results for Cassiopea sp. (this study) are within the range of literature data, with bell concentrations generally higher than other studies. In contrast, arsenic concentrations in oral arm tissue from this study tended to be lower than for other studies that focussed on zooxanthellae containing tissues (Table 6).

Lithium was the only element present at lower concentrations in tissue samples than seawater samples. [START_REF] Kszos | Toxicity of lithium to three freshwater organisms and the antagonistic effect of sodium[END_REF] showed lithium was toxic to freshwater organisms in the absence of sodium. However, in marine environments, sodium levels are sufficiently elevated that lithium is not generally a problem [START_REF] Sadiq | Toxic metal chemistry in marine environments[END_REF]. It is interesting that lithium appears to be actively regulated within the body in this study, although the mechanism/s and requirements for regulation are not known. It is possible that this result reflects a lag between lithium uptake and change in water concentration but it is unlikely that pulses of lithium occurred at all five locations immediately prior to sampling.

The correlations in metal accumulation in the tissues provided some significant relationships.

Positive correlations such as calcium and magnesium in tissue were not unexpected given the osmoconforming nature of the animal [START_REF] Arai | A functional biology of Scyphozoa[END_REF]. Other relationships however, were more difficult to explain. It is recognised that metal transport sites, whilst having a high affinity for metals, may not have a high selectivity and can bind with non-essential metals of similar geometry [START_REF] Worms | Bioavailability of trace metals to aquatic microorganisms: importance of chemical, biological and physical processes on biouptake[END_REF].

Given the variability in element correlation between the tissues, it is feasible that elemental absorption and sequestration may occur in at different locations within the jellyfish. The ability of the zooxanthellae to migrate between the tissues within amoebocytes [START_REF] Estes | Localization and Quantification of Carbonic Anhydrase Activity in the Symbiotic Scyphozoan Cassiopea xamachana[END_REF] may allow ready migration of elements between the tissues as well. In addition, it is possible that there may be deliberate absorption of some elements as a detoxification mechanism [START_REF] Wang | Influence of metal exposure history on trace metal uptake and accumulation by marine invertebrates[END_REF].

Food as a source of metal uptake may be important in explaining some of these patterns, and has been identified by many authors as a potentially significant source (eg Depledge & Rainbow 1990).

Importantly, the presence of these correlations indicates that uptake and sequestration of elements may not be independent from each other.

Relationships between tissue and water for the different elements for the most part were not significant. However, a grab water sample may not reflect the time integrated exposure of the jellyfish to elements. It is likely that for many of the elements, tissue concentrations were the result of long term uptake and/or sequestration of elements while the water measurements reflected the current water quality. Again, the contribution of diet in tissue element concentrations could obscure any relationships between tissue and water concentrations. There was a positive correlation between tissue magnesium and water magnesium in the oral arm (p < 0.05, n = 5, r = 0.94) and is likely to be due to the osmoconforming nature of jellyfish [START_REF] Arai | A functional biology of Scyphozoa[END_REF].

Although data on the long term accumulation of trace metals in Cassiopea sp. are limited, Fowler et al ( 2004) used radiotracers on asymbiotic Aurelia aurita and symbiotic Cassiopea andromeda to assess efficiencies of uptake and half-life retention of Zn, Co, Ag and Cd. C. andromeda was found to be very efficient at both accumulating and retaining radiotracers from seawater [START_REF] Fowler | Applied radiotracer techniques for studying pollutant bioaccumulation in selected marine organisms (jellyfish, crabs and sea stars)[END_REF].

This capability occurred under both light and dark conditions. Results for A. aurita were lower than for C. andromeda but still measurable with shorter retention times for A. aurita, indicating that zooxanthellae may be important in trace element retention [START_REF] Fowler | Applied radiotracer techniques for studying pollutant bioaccumulation in selected marine organisms (jellyfish, crabs and sea stars)[END_REF]. The reported biological half life of zinc in Cassiopea sp. [START_REF] Fowler | Applied radiotracer techniques for studying pollutant bioaccumulation in selected marine organisms (jellyfish, crabs and sea stars)[END_REF] was comparable to the 66 days reported for the sea snail Rapana venosa [START_REF] Onat | A laboratory study of Zn and 134 Cs depuration by the sea snail (Rapana venosa)[END_REF] and longer than the biological half lives of 16.7-30.1 and 25.1 days reported for Crassostrea gigas and Mytilus smarangdium respectively [START_REF] Han | Depuration of copper and zinc by green oysters and blue mussels of Taiwan[END_REF]. [START_REF] Fowler | Applied radiotracer techniques for studying pollutant bioaccumulation in selected marine organisms (jellyfish, crabs and sea stars)[END_REF] is one of the few studies to date that quantifies the efficiencies of uptake, and demonstrates that scyphozoan jellyfish have the potential to accumulate and retain metals over time with similar efficiencies to other invertebrate species.

Differences found in elemental signatures between oral arms and bell tissues in this study could be due to several factors. Different tissue functions and densities of zooxanthellae will certainly contribute to the variation [START_REF] Estes | Localization and Quantification of Carbonic Anhydrase Activity in the Symbiotic Scyphozoan Cassiopea xamachana[END_REF]. The variation in uptake and half-life efficiencies in A.

aurita and C. andromeda [START_REF] Fowler | Applied radiotracer techniques for studying pollutant bioaccumulation in selected marine organisms (jellyfish, crabs and sea stars)[END_REF] suggest that zooxanthellae may explain differences in accumulation and sequestration of trace elements. Jellyfish may also actively transport elements within their bodies leading to differences between uptake and storage tissues. Elemental concentrations in prey and undigested food could also contribute to variation in tissue elemental loads, as could other differences in uptake mechanisms (eg direct absorption from the environment).

Spatial Variation in Elemental Signatures

Multivariate analyses have been used widely in ecological studies and more recently it has also been used to try and characterise processes and trace anthropogenic pollutants to their sources (eg Del [START_REF] Valls | Determining contamination sources in marine sediments using multivariate analysis[END_REF][START_REF] Zitko | Characterization of PCBs by principal component analysis (PCA of PCB)[END_REF]. The locations in this study reflected a variety of urban marine environments with different catchment characteristics, adjacent population densities, and water turnover rates. Spatial variation was found among locations, separated by kilometres to hundreds of kilometres and was consistent for both tissue types. Accumulation patterns in bell tissue were more discrete than oral arm tissue (Figure 2). Notwithstanding this, it was still possible to separate locations based on the oral arm signature. Variation was, as predicted, smallest between locations separated by hundreds of metres (eg Myora Drain A & B; Lake Magellan A & B). In addition, the patterns of accumulation tended to reflect differences in the extent of the adjacent urban areas. The sedentary nature of Cassiopea sp. is likely to be fundamental to this capability, as individual animals do not readily move large distances within their environment.

Biomonitoring Potential of Cassiopea sp.

Rainbow & Phillips (1993) identified a number of key criteria that organisms need to have in order to be considered an effective biomonitor. Cassiopea sp. has a wide distribution in tropical environments, in particular coastal marine environments. It is largely sedentary due to its atypical behaviour of resting upside down on the substrate and which also makes its identification relatively simple. Due to its presence in coastal marine systems, Cassiopea sp. is considered euryhaline and thus tolerant physico-chemical changes in its environment. These attributes meet many of the criteria for biomonitors (Rainbow & Phillips 1993). A primary attribute of biomonitors is the ability to accumulate elements of concern. This study and the few other studies to date (Table 6) indicate that Cassiopea sp. (among others) is capable of accumulating elements above ambient seawater concentration (Figure 1). The extent of this capability requires further exploration to determine limitations to uptake and also retention times within tissues.

350

Conclusions

This study has demonstrated that Cassiopea sp. is capable of accumulating elements above ambient seawater concentrations. It is also possible to discriminate different geographical populations based on chemical signatures in their tissues. Whilst their persistence in polluted systems has been documented, historically jellyfish have not been considered useful biomonitors. We suggest it would be useful to consider them as a future tool in the biomonitoring toolbox, as they meet many of the identified criteria.

Acknowledgements

Thank you to Dr Yi Hu for reviewing & providing constructive suggestions on the digestion 360 method and for undertaking the ICP-MS analyses. Thank you to Darwin City Council for providing Cassiopea sp. samples. Thank you also to Caloundra City council for providing background information on Lake Magellan. Table 1: Sampling locations and site descriptions. For Lake Alexander collection (A) = Jellyfish, (W) = Water

FIGURE CAPTIONS

  Factor 1 was described by positive loadings for Zn, Cu, Cr and Mn (0.854, 0.833, 0.728 & 0.709, respectively) and a negative loading for Li (-0.562). Factor 2 loadings were positive for Mg & Ca (0.928 & 0.916 respectively) and negative for Fe and Ba (-0.789 & -0.681, respectively). Individual samples within locations displayed greater variability in signatures and there was greater overlap among locations. Although exhibiting greater individual variations, the Myora

Figure 1 :Figure 2 :

 12 Figure 1: Elemental bioconcentration in Cassiopea sp. tissues as a function of ambient seawater concentration measured at each location. Equation to calculate bioconcentration = (tissue concentration/water concentration). A= Bell tissue; B= Oral arm tissue. Bioconcentration >1 is considered accumulation; bioconcentration = 1 reflects no accumulation; bioconcentration <1 reflects regulation / excretion

  ). Factor 1 was characterised by positive loadings for Mn,

	Al, As, Zn and Ba (0.872, 0.829, 0.797, 0.726 and 0.707, respectively) and negative loadings for Cu (-
	0.617). Factor 2 was characterised by positive loadings for Ca & Mg (0.987 & 0.986, respectively)
	and negative loadings for Ba and Zn (-0.417 & -0.373, respectively). The Myora Drain A & B

locations had the greatest similarity in elemental fingerprint. Patterns between Lake Magellan A & B

Table 2 :

 2 Trace elements (mean ± 1SE) in Cassiopea sp. tissues (wet weight) with 2-tailed pair-wise t-test between tissues for each location. ns p>0.05; * p<0.05; ** p<0.01; ***p<0.005. df=4. All concentration expressed as ug/kg except Ca, Mg and Fe (mg/kg) wet weight

	Element	Tissue	GC	p	EC	p	SC	p	JB	p	LA	p
	Al	Bell	358+81	ns	251+73	***	1727+680	ns	481+117	ns	6975+2577	ns
		Oral Arm	1323+484		628+96		5418+2874		1142+284		6681+2128
	As	Bell	45.9+6.6	*	84.1+9.0	*	94.0+21.1	ns	170+15	**	143+25	ns
		Oral Arm	69.2+10.6		153+25		135+15		378+38		156+25	ns
	Ba	Bell	14.4+0.7	*	12.9+0.6	**	27.0+5.7	***	22.8+1.7	*	20.2+2.1	*
		Oral Arm	19.4+1.7		21.3+1.7		53.3+7.3		47.6+4.8		30.8+4.4
	Ca	Bell	372+29	ns	427+18	ns	258+11	ns	342+7	ns	466+19	ns
		Oral Arm	429+36		446+20		272+17		412+28		430+20
	Cd	Bell	7.58+0.97	ns	7.11+0.69	ns	3.31+1.41	ns	4.86+0.56	***	15.1+2.1	*
		Oral Arm	8.29+1.48		12.4+2.4		5.42+1.18		13.1+1.1		40.6+8.0
	Cr	Bell	108+30	ns	110+47	ns	141+84	ns	121+52	***	88.8+39.5	ns
		Oral Arm	104+35		117+51		141+74		162+54		116+50
	Cu	Bell	124+9	ns	79.4+13.1	ns	36.8+7.6	**	79.8+10.9	***	64.2+16.8	*
		Oral Arm	119+14		134+29		142± +26		196+16		139+27
	Fe	Bell	1.79+0.65	*	4.38+2.21	*	8.73+2.69	***	1.52+0.36	***	19.2+7.8	ns
		Oral Arm	3.46+0.46		12.9+2.7		38.8+3.6		13.4+1.8		13.0+3.7
	Li	Bell	120+32	ns	94.1+39.3	ns	52.6+17.9	ns	71.1+30.2	ns	154+33	ns
		Oral Arm	137+55		124+41		52.7+19.3		70.6+31.9		127+31
	Mg	Bell	1050+79	ns	1221+69	ns	744+28	ns	979+13	ns	1297+55	ns
		Oral Arm	1251+115		1257+29		734+44		1034+75		1251+55
	Mn	Bell	123+15	**	248+25	***	402+159	**	338+48	***	590+200	ns
		Oral Arm	354+42		687+80		644+166		1077+105		669+246
	Sr	Bell	7340+699	ns	8022+707	**	5812+862	***	7228+518	*	8358+647	ns
		Oral Arm	9184+560		10343+799		6970+828		10666+874		7634+599
	Zn	Bell	1245+178	***	1040+121	ns	2633+441	*	2959+421	***	2085+275	*
		Oral Arm	1897+170		2431+575		4835+836		7559+828		4999+833

Table 3 :

 3 Correlations among trace elements in bell and oral arm tissue of Cassiopea sp. n=25 for both tissues

	Bell Tissue
		Al	As	Ba	Ca	Cd	Cr	Cu	Fe	Li	Mg	Mn	Sr
	As	0.475 *
	Ba	0.488 *	0.534 **
	Ca	0.101	0.127	-0.396
	Cd	0.294	0.134	-0.077	0.729 ***
	Cr	-0.050	0.109	0.131	0.233	0.248
	Cu	-0.543 **	-0.324	-0.494 *	0.440 *	0.300	0.362
	Fe	0.671 ***	0.271	0.161	-0.090	0.051	-0.199	-0.580 ***
	Li	0.324	0.061	0.023	0.260	0.231	-0.542 **	-0.096	0.317
	Mg	0.102	0.143	-0.411 *	0.970 ***	0.723 ***	0.191	0.430 *	-0.094	0.275
	Mn	0.589 ***	0.674 ***	0.543 **	0.189	0.252	0.433 *	-0.452 *	0.346	-0.208	0.120
	Sr	-0.336	-0.139	0.179	-0.187	-0.293	0.044	0.033	-0.454 *	-0.361	-0.254	0.063
	Zn	0.002	0.131	-0.010	0.717 ***	0.690 ***	0.606 ***	0.318	-0.305	-0.116	0.663 ***	0.464 *	0.095
	Oral Arm Tissue
		Al	As	Ba	Ca	Cd	Cr	Cu	Fe	Li	Mg	Mn	Sr
	As	-0.100
	Ba	0.322	0.526 **
	Ca	-0.132	0.168	-0.406 *
	Cd	0.324	0.202	-0.134	0.525 **
	Cr	0.267	0.058	-0.056	0.278	0.270
	Cu	0.124	0.326	0.216	0.100	0.317	0.811 ***
	Fe	0.279	0.313	0.592 ***	-0.596 ***	-0.220	-0.071	0.084
	Li	-0.175	-0.024	-0.212	0.222	0.081	-0.552 ***	-0.533 **	-0.176
	Mg	-0.090	0.005	-0.539 **	0.948 ***	0.494 *	0.108	-0.123	-0.629 ***	0.370
	Mn	0.139	0.702 ***	0.477 *	0.202	0.079	0.238	0.292	0.273	-0.266	0.071
	Sr	-0.200	0.249	-0.101	0.714 ***	0.157	0.539 **	0.423 *	0.333	-0.192	0.566 **	0.497 *
	Zn	0.454 *	0.593 ***	0.682 ***	-0.128	0.340	0.455 *	0.695 ***	0.431 *	-0.464 *	-0.323	0.542 **	0.140
		* significant at p < 0.05, ** significant at p <0.01, ** * significant at p < 0.005

Table 4 :

 4 One-way ANOVA trace element concentrations in water, bell and oral arm tissue. Data log10 transformed. n.d. = not determined (results below reported detection limit). ns p>0.05; * p<0.05; ** p<0.01; ***p<0.005. df (Water) = 4,9; df (Bell & Oral Arms) = 4,20.

	Element Water	Bell	Oral Arm
		MS	Residual
	Ag	n.d	n.d
	Al	0.426	0.013
	As	n.d	n.d
	Ba	0.040	0.000
	Ca	0.011	0.000
	Cd	n.d	n.d
	Cr	0.003	0.006
	Cu	0.035	0.007
	Fe	0.300	0.000
	Li	0.019	0.001
	Mg	0.007	0.000
	Mn	0.145	0.000
	Pb	n.d	n.d
	Sr	0.010	0.000
	Zn	n.d	n.d

Table 5 :

 5 .05) from one-way ANOVA for trace element concentrations among locations. Locations sharing the same letters are not significantly different from each other.

	Element	Tissue
	Al
	As
	Ba
	Ca
	Cd
	Cr
	Cu
	Fe
	Li
	Mg
	Mn
	Sr
	Zn

Table 6 :

 6 Trace element concentration in tissues of scyphozoan jellyfish from the literature. Results are mean for tissue type. # indicates concentration as ug/g dry weight, all others ug/g wet weight. Results below detection <d.l. Results from current study (both wet and dry) are the means from all locations.

	Species	Location	Tissue Type	Ag	Al	As	Ba	Ca	Cd	Cr	Cu	Fe	Li	Mg	Mn	Pb	Sr	Zn	Source
		Collected
	Pelagia noctiluca	Straits of Messina, Sicily, Italy	Tentacles		0.71	1.23		119	0.055		0.19	33		11.7	1.15	0.33		3.01	Cimino et al
	Pelagia noctiluca	Bay of Villefranche-sur-Mer, Cote	Whole animal						0.4 #		2.0 #					0.6#		46 #	Romeo et al
		d'Azur, France
	Auralia sp.	Coastal waters	Whole animal					18800 #	0.04 #		16.1 #	178.8 #		75700 #	0.04 #			19.9 #	Siddiqui
		Karachi, Pakistan																	et al
	Unidentified	Pertuis charentais,	Unknown						0.27 #										Caurant
		coastal waters,																	et al
		France
	Chrysaora	Coastal waters	Umbrella			0.074													Hanaoka
	melanaster	Yoshimi, Japan	Other			0.428													et al
	Pelagia	Coastal waters	Whole animal			0.197													Hanaoka
	noctiluca	Yoshimi, Japan																	et al
	Mastigias	Coastal waters	Umbrella			0.039													Hanaoka
	papua	Yoshimi, Japan	Oral Arm			0.073													et al
			Gastral Filament
			/Gonad			0.578
			Zooxanthellae
			Tissue			0.308
	Stromolophus	Coastal waters	Umbrella			0.183													Hanaoka
	nomurai	Yoshimi, Japan	Oral Arm			0.111													et al
			Tentacle			0.664
	Cassiopea sp.	Gold & Sunshine Coasts, Queensland;	Bell	<d.l.	1.959 39.5 #	0.107 2.16 #	0.019 0.383 #	373 7520 #	0.008 0.161 #	0.114 2.30 #	0.077 1.41 #	7.12 144 #	0.098 1.98 #	1058 21300 #	0.340 6.85 #	<d.l.	7.352 148 #	1.993 40.2 #	Current Study
		& Darwin, Northern
		Territory, Australia	Oral Arm	<d.l	3.038 54.3 #	0.178 3.18 #	0.034 0.607 #	398 7110 #	0.016 0.286 #	0.128 2.29 #	0.146 2.61 #	16.33 292 #	0.102 1.82 #	1106 19750 #	0.686 12.3 #	<d.l.	8.959 160 #	4.344 77.6 #	Current Study

Bell Oral Arm Bell Oral Arm Bell Oral Arm Bell Oral Arm Bell Oral Arm Bell Oral Arm Bell Oral Arm Bell Oral Arm Bell Oral Arm Bell Oral Arm Bell Oral Arm Bell Oral Arm Bell Oral Arm