Of Bioavailable Metal Contamination Across South Australia Using Translocated Mussels Mytilus Galloprovincalis

Transactions of the Royal Society of South Australia (2011), 135(1): 39–54

AN ASSESSMENT OF THE CURRENT STATUS OF BIOAVAILABLE METAL CONTAMINATION ACROSS SOUTH AUSTRALIA USING TRANSLOCATED MUSSELS MYTILUS GALLOPROVINCALIS

S. GAYLARD1, S. THOMAS1, AND M. NELSON1

South Australian Environment Protection Authority, GPO Box 2607 Adelaide, South Australia 5001 1 – corresponding author, GPO Box 2607 Adelaide, South Australia 5001 Fax: +61881244673

Abstract

This study presents results from the first regional assessment of metal levels in translocated blue mussels (Mytilus galloprovincialis Lamarck, 1819) in the nearshore coastal environment across South Australia. A total of 422 replicate samples of translocated mussels were deployed throughout 11 regions across the South Australian near shore marine environment. Mussels were recovered after approximately 70 days and analysed for total metal load in the flesh of the mussel. Results showed vast differences in metal concentrations between different regions and also compared to the baseline metal concentration. Many of these differences can be attributed to the level of industrial activity in the regions which may have implications for environmental regulation in these regions.

KEY WORDS: Mytilus galloprovincialis, lead, cadmium, mercury, biomonitor, industrialisation.

Introduction

Spatial and temporal monitoring of levels and trends of metals and other contaminants in the coastal environment using bivalve mussels as bio-monitors is well established since Goldberg (1975) proposed the concept. Metals accumulated in the mussel tissues represent a time integrated response to bioavailable metals, where the concentration may be several orders of magnitude higher than what would be found in seawater [Phillips and Rainbow, 1993; Rainbow, 1995]. Using this time integrated approach can reduce the frequency and cost of analysis and can also produce results which are less likely to be affected by analytical limits of reporting or by frequent changes in dissolved concentrations which may vary on each tidal cycle or season [Rainbow, 1995]. Importantly using biological indicators of the available fraction of metals in the marine environment is of direct ecotoxicological relevance [Phillips and Rainbow, 1993; Boening, 1999] and is preferable to other methods where the sampling and/or chemical analysis method artificially define the bioavailable fraction. State- or country-wide monitoring programs using field-collected or translocated mussels have been underway in many locations since the early 1980s either as Government funded “state of the environment” style programs e.g.: Mussel Watch in the National Status & Trends program US NOAA: [Farrington et al., 1983; O’Conner, 1996], or the French Reseau National d’Observation de la Qualité du Mulieu Marin [Beliaeff et al, 1998] or Biointegrator network (RINBIO) [Andral et al., 2004]. Alternatively specific studies have been conducted to investigate known or suspected contamination issues via research bodies [Ward et al., 1986; Jamil et al., 1999; Chou et al., 2003].

The uptake and hence bioavailability of metals in the marine environment by a biomonitor is highly dependant on a number of geochemical and biological factors. The geochemical factors that can affect bioavailability include the concentration of organic carbon, water hardness, salinity, temperature, pH, dissolved oxygen and hydrologic features of the particular environment [Boening, 1999]. These parameters may vary between regions which may contribute to spatial variability. The biological factors that can influence the uptake of bioavailable metals are significant between bivalve species [Beliaeff et al., 1998]. This issue can be somewhat overcome by using a standardised or cosmopolitan species such as Mytilus spp. [Rainbow, 1995]. Even within the same species variability can occur through different ages, sex and reproductive state [Boening, 1999]. Translocating mussels from an area such as a commercial lease where the age and reproductive state are known and homogeneous can help to alleviate some of the variability seen within the same species [Fabris et al., 1994].

39 S. GAYLARD, S. THOMAS, & M. NELSON

South Australia’s nearshore coastline is varied both hydrodynamically and geomorphologically. There are two large inverse estuaries which reduce flushing with the open ocean [Petrusevics, 1993; Nunes Vaz et al., 1990], numerous large embayments and open southern ocean. The city of Adelaide, with its 1.1 million people [ABS, 2007], lies on the eastern shores of Gulf St Vincent (Figure 1) is the major population centre in South Australia and there are numerous smaller towns, the majority of which are located on the coast. There are a number of potential sources of metals into the nearshore coastal environment in South Australia. The most significant of these is a lead/zinc smelter at Port Pirie located in the north of Spencer Gulf and also the city of Adelaide located on the eastern side of Gulf St Vincent (Fig. 1). Significant metal accumulation and biological effects within the coastal waters surrounding Port Pirie have been well documented over the last 30 years [Ross et al., 2002; Edwards et al., 2001; Ward & Hutchings, 1996; Ward et al., 1986; Ward & Young, 1981]. Other potential sources of metals across South Australia include wastewater treatment plants, urban and agricultural runoff, heavy industry and atmospheric deposition. Like most locations throughout Australia coastal towns are experiencing a large amount of development in close proximity to the coast. To date there has been little documented assessment of metal levels or quantification of risk in all regions of South Australia, with the exception of Port Pirie, other than a small number of desktop risk assessments or specific pollution event driven investigations [Gaylard, 2009; Bryars et al, 2006; Mortimer, 2004; Westphalen et al., 2004].

The aim of this study is to assess regional water quality rather than investigating specific sources of metal pollution. Mussels have been used in this survey as biomonitors to assess the metal status of the marine environment, not to determine whether the concentrations of metals recorded in the environment are adversely affecting the Mytilus galloprovincialis that have been exposed. This monitoring is designed to provide information to quantify the risks from heavy metals on a regional scale in South Australian coastal waters as this may affect the level of regulation exerted on each of the sources in the regions.

Methods

A large proportion of South Australia’s nearshore coastal environment was sampled throughout this study. In order to make regional assessments of metal levels the state was arbitrarily divided into 11 regions based on potential risk factors, hydrodynamics and logistical constraints.

A total of 422 replicates were sampled across 11 regions throughout South Australia’s nearshore coastal waters (Fig. 1). At each replicate location within each region, nylon bags containing 10 mussels were placed in the water using a clean “Besser block” weight on a polypropylene rope to maintain its position. Each replicate analysis was taken from the 10 organism pooled which has been shown to provide an excellent estimate of the population mean generally with a lower variance than the population variance [Gordon et al, 1980]. The bags were suspended approximately 2 m below mean low water mark using a 100 mm sub-surface float and marked by GPS. The number of replicates within each region varies according to the perceived level of contamination which was assessed prior to deployment. Studies have shown that disturbed areas can be more variable than undisturbed and as such sample size is often needed to be higher in order to find differences (whether in means or variances) between regions [Fraterrigo and Rusak, 2008; Fairweather, 1991]. In some areas replicate numbers are lower than anticipated due to a number of replicates being un- recoverable or lost. All regions, with the exception of the South East indicated significant differences for a number of metals (see later) indicating that there was sufficient power to detect a difference at the 0.05 level. There were a number of replicates that were not recovered from the South East which greatly reduced the sample size in this region. This is likely to have adversely influenced the power to detect a difference between this location and others. The higher wave energy typical of this region is likely to be a contributing factor to the loss of replicates in this region.

Mussel sampling

Mussels (Mytilus galloprovincialis, Lamarck 1819, length µ=85.3 mm σ=5.8 mm) were obtained from a commercial mussel aquaculture lease located approximately 10 km from the township of Port Lincoln on South Australia’s Eyre Peninsula (Figure 1). On receipt of each 20 kg batch of mussels a sub-sample of

40 STATUS OF METAL CONTAMINATION USING TRANSLOCATED MUSSELS MYTILUS GALLOPROVINCALIS

1 11 .. WHYALLA

1'i;--- ~ PORTPIRIE Study Area \j

EE COFFIN BAY D

o-"% , 0/o PORT LINCOLN

> .; / Yorke Peninsula

GAWLER ~ A~ ELAIDE

Port River

Adelaide North ---.. ~ j 1\ t: '"' - ADELAIDE Adelaide 0 50 100 150 200 250 Km Central

# , X

Adelaide X So"th ------. , KINGSCOTE 1%

McLAREN VALE # X # Built Up Area I

Figure 1. Map of 87 mussel locations within 11 regions across South Australia.

41 S. GAYLARD, S. THOMAS, & M. NELSON mussels (n=20) were sent to the laboratory for metal analysis to establish a baseline. All mussels were deployed between February-March 2008 and February-March 2009 in order to maintain similar season between years. All mussels were retrieved as close to 70 days after deployment as possible. Depuration to clear the digestive tract of sediment and detritus was not undertaken in this survey as it was considered to lead to inaccuracies in metal results due to possible metal elimination depending on the specific depuration time, which is different for each metal species and physical properties of the depuration water, particularly food, temperature and salinity [Wang et al., 1995; McKinney & Rogers, 1992]. The potential for subsequent uptake of trace concentrations of metals in the “clean” waters was also a factor, particularly in the regions where very low concentrations of metals were expected.

Analytical procedure

Mussel shells were removed and the flesh freeze-dried for 24 h. The freeze-dried samples were thoroughly homogenised and 0.30 g sub-samples were weighed for microwave digestion (CEM MDS 2100, Matthews, USA) with 4 mL of high purity nitric acid (69%, GFS Chemicals). The digested samples were diluted with Milli-Q deionised water (>18 MWcm-1, Millipore Element System) to 40 mL. The metal levels were analysed by inductively coupled plasma mass spectrometry (ICP-MS) (Agilent 7500a, Japan) after appropriate addition of internal standards. The method used in this laboratory is based on methods described elsewhere (Sakao and Uchida, 1999; Cubadda et al., 2001). The standard reference materials of Bovine Liver AGAL-4 (Australian Government Analytical Laboratories, NSW), Oyster Tissue SRM 1566b (NIST, Gaithersburg, USA) and the laboratory in-house reference material of seafood tissue (QC 180 fish mix) were used for quality control and assurance, and these reference materials were treated similarly to the samples throughout the study. All QC recoveries and certified standards were in the range of 88.6 – 113.6% of the reference values (Table 1).

The limits of reporting (LOR) of the method were 0.05 µg.g-1 for arsenic, chromium, copper, lead, manganese and zinc, and 0.01 µg.g-1 for cadmium and mercury. Sample preparation and trace element analysis was undertaken at the Queensland Health Forensic and Scientific Services which is NATA accredited.

Statistical methods

Because of the large amount of data within this investigation (422 samples across 11 regions), analysis has been performed on a regional scale to evaluate broad differences in metal levels across South Australia. This is consistent with other reporting undertaken by the South Australian Environment Protection Authority and State of the Environment reporting. In regions where some replicates indicated anomalous results, the data were extracted and analysed at a finer resolution to investigate whether patterns within that region exist.

Results have been presented using box plots indicating mean, median, inter-quartile range, maximum and minimum. A Shapiro-Wilks test for normality was undertaken and in all cases, at least one region was not normally distributed after transformation, hence non-parametric methods have been used to look for differences between locations. Differences in metal concentrations between regions were assessed using the Kruskal Wallace multiple comparison test, applying the Bonferroni correction for the significance level. Principal components analyses (PCA) with Spearman correlations have been undertaken to investigate associations within metal species and location. All statistical analysis has been using XLSTAT (Addinsoft, 2008) or Minitab 14.

Results and Discussion

Spatial patterns in metal concentrations

The results from the PCA are shown as a 2-dimensional correlation biplot (Figure 2). The variabilities for PCA axis 1 and PCA axis 2 were 43 % and 24 % respectively accounting for 67.4 % of the total variance. The third and fourth PCA axis accounted for only minor proportions of the total variability (11 and 10% respectively) and therefore will not be discussed further. Spearman correlation coefficients indicated that copper,

42 STATUS OF METAL CONTAMINATION USING TRANSLOCATED MUSSELS MYTILUS GALLOPROVINCALIS manganese and lead are all strongly positively correlated, similarly PCA indicates that there are distinct similarities in bioavailable metals at Port Pirie, Port River and Whyalla (Figure 2), the Port Pirie region being the most important factor on the F1 axis. Cadmium, mercury, arsenic and chromium are all strongly correlated with each other, but not to the other metals.

PCA Biplot (axes F1 and F2: 67.40 %)

3 Zn Port Pirie • 2 South_East ♦

0~ Baseline (") • Kangaroo_lsland ~ •Port_River '

♦ -3 Coffin_Bay

-4 Ade~ide_ stuth -5 -4 -3 -2 -1 0 2 3 4 5 6 7 8 F1 (42.96 %)

Figure 2. PCA biplot of regional metal concentrations (µg g-1) in Mytilus galloprovincialis and metal speciation.

Port Pirie, Whyalla and Port River regions

As outlined above, the PCA indicated correlations between bioavailable metals at Port Pirie, Port River and Whyalla regions which were different to the other regions sampled with the PCA indicating important associations between lead, copper and manganese. Port Pirie, Port River and Whyalla regions had significantly elevated concentrations of many bioavailable metals compared not only to the baseline mussels but also other regions. Table 1 and Figures 3-5 show that Port Pirie had significantly elevated arsenic (p < 0.0001), copper (p < 0.0001), manganese (p < 0.0001), cadmium (p < 0.0001), mercury (p < 0.0001) and zinc concentrations (p < 0.0001). By far the most noteworthy of these were the lead concentrations, which across the Port Pirie region the median lead content in the translocated mussels was 22.0 µg.g-1 with a maximum of 310, µg.g-1 (n = 55) which is almost 19,000% and significantly (p < 0.0001) higher than the baseline lead concentration (Figure 3). It should be noted that this result was not unexpected give that Port Pirie is a known ‘hot spot’ for metal contamination due to the presence of a lead zinc smelter since the late 1800s; significant contamination in sediment and biota has been documented since the mid 1970s [Corbin and Wade, 2004; Ross et al., 2002; Ward and Hutchings, 1996; Ward et al., 1986]. However, the magnitude of the lead concentration was surprising given the installation of wastewater treatment systems within the plant over the last decade. It should be noted that while this observation is very high indeed, care needs to be applied when comparing values from representative sites as compared to sites known as “hot spots”. All samples within the Port Pirie region (n = 55 over a 180 km2 area) exceeded the 3.20 µg.g-1 criterion proposed by Cantillo (1998) as a level indicative of contamination. At this stage, it is unclear whether the bioavailable lead has simply arisen from historical contamination or may have a more

43 S. GAYLARD, S. THOMAS, & M. NELSON recent component in its causation. A more detailed investigation is being undertaken and could be the subject of additional publications. In addition to very high lead levels, the Port River, Port Pirie and Whyalla were also significantly elevated (p < 0.0001) in manganese compared to the Baseline concentrations of 15.3, 12.1 and 5.94 µg.g-1 respectively. This represents up to a 600% increase over the manganese baseline. Similarly, copper results were also strongly associated with Port River, Whyalla and Port Pirie with median concentrations of 6.88, 5.88 and 5.32 µg.g-1 respectively. These results are up to 57% higher and significantly greater than the baseline copper content in the M. galloprovincialis (p < 0.0001). Interestingly, both Port Pirie and Whyalla had relatively low bioavailable mercury concentrations. Port Pirie had the highest regional median bioavailable cadmium concentration across South Australian coastal waters of 2.51 µg.g-1 (Table 1) which represented a 110% and significant increase from the baseline state (p < 0.0001). Also of interest was the Port River region which, although it is a heavily industrialised region with well documented metal pollution [EPA, 1997a&b; Edwards et al., 2001; Wade, 2002], showed the lowest cadmium concentrations throughout South Australia. This result was a 27% decrease from the baseline cadmium content over the 70-day deployment period.

Table 1 and Figure 4 show an apparent distinction between industrialised, urbanised and rural/semi rural regions in bioavailable manganese concentrations in translocated M. galloprovincialis. The highest concentrations were observed in the more industrialised areas of Port River and Port Pirie with 15.26 and 12.13 µg.g-1 respectively and to a lesser extent Whyalla (5.95 µg.g-1). This represents up to 600% increase from the baseline level of manganese (Table 1). Similarly, copper concentrations were generally elevated in the more industrialised areas with Port River, Whyalla and Port Pirie regions having the highest median results (6.88, 5.88 and 5.32 µg.g-1 respectively); the baseline state of copper concentrations was 4.39 µg.g-1 so that copper concentrations in the industrialised areas were up to 57% higher than the baseline (Table 1). When the median values were ranked from all metals in all regions, the locations that had the highest ranks in the whole data set were Port Pirie, Port River and Whyalla. Again this reinforces the results shown here that these three regions were ranked higher in median metal concentrations compared to all other areas in the study (Table 2).

The most significant metal source in these regions is the large lead/zinc smelter at Port Pirie which has a long and well documented history of metal contamination throughout the Port Pirie region. The Whyalla region is dominated by the steel manufacturing facility at Whyalla and again there is a long history of metal discharges into the marine environment from this facility. Similarly the Port River has many industrial discharges, urban stormwater and one large sewage treatment plant discharge (a second was diverted in 2006) which all contribute to the metal load in the region. Unfortunately it is difficult to differentiate between current metal pollution and historical contamination within sediment being resuspended, particularly in an environment frequently disturbed by dredging and shipping [Kalnejais et al, 2010]. The current load of metal pollution being discharged to the marine environment by large facilities in all three regions has significantly declined over the last 10 years [NPI, 2010] but these results show that the current level of metal pollution in addition to the significant extent of historical contamination in the marine environment can be detected in bivalve mussels and could be having long term consequences on ecosystems in these regions.

Adelaide regions

In general, the three Adelaide regions (Adelaide North, Central and South) were elevated in a number of bioavailable metals when compared to the less urbanised regions but were broadly lower in most metals than the industrialised locations of Port Pirie, Whyalla and Port River (Figures 3-5). The most noteworthy results from the Adelaide regions were the significantly greater bioavailable manganese concentrations in all three regions compared to the Baseline concentration (p < 0.0001); of these the manganese at Adelaide North was slightly higher than the Adelaide Central and Adelaide North regions (Table 1 & Figure 4). Similarly bioavailable lead concentrations were significantly higher in Adelaide North, Adelaide Central and Adelaide South regions (p<0.0001, p < 0.0001 and p <0.01 respectively) compared to the Baseline concentration. The cadmium concentrations in the Adelaide Central region were significantly higher than the baseline (p < 0.0001) and were also higher than the other two Adelaide regions. This represents a 65% increase from the baseline cadmium body burden.

44 Figure 3. STATUS OFMETAL CONTAMINATION USING TRANSLOCATED MUSSELS Total Mercury (ug.g-1) Total manganese (ug.g-1) Lead (µg.g Total Copper (ug.g-1) 0 0 0 0 0 0 0 0 ~ C) C) C) C) C) ~ ~ ~ I\) .I>, 0) 00 0 ~ ~ ~ ~ ~ 0 I\) .I>, 0) 00 0 I\) .I>, 0 0 0 0 0 0 0 I\) .I>, 0) 00 0 I\) .I>, 0) 00 -1 dry weight)intranslocatedmussels andleadintranslocatedmusselsexcludingPortPirie. Baseline ~ ~ O• Coffin Bay f- ~ Hlli Kangaroo Island f- ~ • ~ Adelaide Central ~ -~· m • Adelaide North ~ • • +- ~ •

45 ~ Adelaide South ~ ~- t ~- Port Lincoln ~ w ID Port Pirie ~ -~ • 0------, • • • ~. Port River ~ -~. 0-, • • ~ • MYTILUS GALLOPROVINCALIS South East ~ [] I ~ Whyalla f- HIH 0 Kn Yorke Peninsula ~ ~ • ~ S. GAYLARD, S. THOMAS, & M. NELSON

Also noteworthy is that Adelaide South copper concentrations were significantly lower than many other regions including the Adelaide North and Adelaide Central regions. Ranks of median metal concentrations indicated that the Adelaide North was consistently higher in metal concentrations, followed by Adelaide Central and Adelaide South (Table 2), these regions were lower than the three industrialised regions (Port Pirie, Port River and Whyalla) but higher than all remaining regions.

The northern and central regions and to a lesser extent the southern region are exposed to numerous industrial and stormwater discharges typical of waters bordering on a large city. The cumulative effect of these discharges have been shown to be significantly influencing water quality [Gaylard, 2004], seagrass condition [Westphalen et al., 2004] and macroalgal reef condition [Turner et al., 2007] in the northern and central regions and to a lesser extent the southern region. Of the three regions adjacent the City of Adelaide, Adelaide Central and Adelaide North both were frequently higher in bioavailable metal concentrations than Adelaide South which is likely to reflect the fewer discharges in this region. As outlined above, ranked median metal concentrations showed a north to south gradient along Adelaide coast line which may be influenced by local hydrodynamic flushing patterns along a south-north gradient which in turn may affect exposure time of discharged metals along this section of coastline.

Other regions

The remaining regions of Coffin Bay, Kangaroo Island, Port Lincoln, Yorke Peninsula and South East were all generally low in bioavailable metal concentrations in translocated M. galloprovincialis, which is generally what would be expected given that these regions are typically free from large industrial discharges and only subject to relatively small stormwater or agricultural runoff loads. Ranking the median metal levels indicated that all regions were higher than the baseline but were lower than both the three Adelaide regions and the more industrialised regions of Port Pirie, Port River and Whyalla.

Mercury

Table 1 indicates that mercury concentrations ranged from 0.04 µg.g-1 at Port Lincoln to 0.07 µg.g-1 in the Port River region. Kruskal Wallace tests indicated many statistically significant differences between the regions, of which the most noteworthy was the elevated mercury concentrations in the regions bordering the City of Adelaide, particularly the Port River and Adelaide North. The mercury concentrations at the Port River and Adelaide North were both 0.07 µg.g-1 which represented a 75% increase from the baseline mercury concentration. It is interesting to note that the two major industrial regions of Port Pirie and

Table 1. Mean metal concentrations in translocated Mytilus galloprovincialis (µg.g-1 dry weight)

n As Cr Cd Cu Mn Pb Hg Zn

Baseline 23 14.8 0.71 1.20 4.39 2.18 0.17 0.04 143 Coffin Bay 12 17.6 0.84 1.53 3.91 3.85 0.15 0.05 114 Kangaroo Island 26 16.1 0.73 1.32 4.80 3.43 0.26 0.05 138 Adelaide North 85 16.1 0.88 1.97 4.85 4.81 0.73 0.07 128 Adelaide Central 66 15.3 0.79 1.41 4.71 5.11 0.80 0.07 139 Adelaide South 69 18.3 0.84 1.53 3.99 4.42 0.50 0.07 120 Port Lincoln 12 18.6 0.72 1.05 5.09 2.89 0.34 0.04 122 Port Pirie 58 16.3 0.73 2.51 5.32 12.1 22.0 0.06 172 Port River 31 18.0 0.82 0.87 6.88 15.26 1.00 0.07 139 South East 6 14.4 0.50 1.20 4.30 3.18 0.21 0.05 148 Whyalla 10 17.0 0.73 1.40 5.88 5.94 0.91 0.04 178 Yorke Peninsula 36 16.7 0.54 1.18 4.27 2.85 0.46 0.06 141

46 STATUS OF METAL CONTAMINATION USING TRANSLOCATED MUSSELS MYTILUS GALLOPROVINCALIS

Table 2. Rank of median metal levels in Mytilus galloprovincialis (ug.g-1). 1= highest ranked relative metal concentrations and 12 = lowest ranked relative metal concentrations

Region As Cr Cd Cu Mn Pb Hg Zn Total Rank

Port Pirie 6 5 12 10 11 12 7 11 74 1 Port River 10 9 1 12 12 11 12 6 73 2 Whyalla 8 7 7 11 10 10 3 12 68 3 Adelaide North 4 12 11 8 8 8 9 4 64 4 Adelaide Central 3 8 8 6 9 9 11 7 61 5 Adelaide South 11 11 10 2 7 7 10 2 60 6 Coffin Bay 9 10 9 1 6 1 6 1 43 7 Kangaroo Island 5 6 6 7 5 4 5 5 43 7 Port Lincoln 12 4 2 9 3 5 1 3 39 9 Yorke Peninsula 7 2 3 3 2 6 8 8 39 9 South East 1 1 4 4 4 3 4 10 31 11 Baseline 2 3 5 5 1 2 2 9 29 12

Whyalla both had relatively low bioavailable mercury concentrations in translocated M. galloprovincialis with 0.04 and 0.06 µg.g-1, which indicates a probably different composition and nature of potential sources between urban and industrial regions. This result could also could be influenced by local geology which has been highlighted as a potential mercury source by a number of authors [Butterfield and Gaylard, 2005; Corbin & Wade, 2004; Gustin 2003; Olsen, 1983; Working Group on Mercury in Fish, 1980; Williams et al., 1976].

Zinc

Across all 11 regions, bioavailable zinc results were largely similar, suggesting that M. galloprovincialis may be able to regulate zinc concentrations, as can a number of other species of bivalve including Mytilus edulis [Ross et al, 2003; Riget et al., 1997] and Perna viridis [Chan 1988]. As for many other metals, Whyalla and Port Pirie were observed to have the highest concentrations which is likely showing an excess of zinc in the environment over which the Mytilus are able to excrete. These results represented a change from the baseline of only 25%. As stated above, Port Pirie is the location of a large lead and zinc smelter and Whyalla is the home to a large steelworks, both of which are likely to be key drivers in zinc concentrations above what is able to be regulated by mussels in these regions.

Arsenic

Table 1 shows that median bioavailable arsenic concentrations in translocated M. galloprovincialis ranged from 14.4 µg.g-1 at the South East region and 18.6 µg.g-1 in Port Lincoln (Figure 5); these represent a -3% and a 26% difference from the baseline level of metal content. The maximum result was 32.7 µg.g-1 in the Adelaide North region, representing a 121% increase in arsenic body burden. There were many statistically significant differences across regions but these did not reflect known anthropogenic inputs, so it is likely that South Australia has natural geological sources of arsenic that can be accumulated by M. galloprovincialis. Many studies have shown a high degree of accumulation of arsenic in seafood, particularly shellfish, and in these circumstances the arsenic is present as organoarsenic compounds, of which the water soluble arsenobetaine and arsenocholine are the dominant forms [WHO, 1988]. Toxicological research has shown that while arsenobetaine and arsenocholine are absorbed on consumption, they are rapidly and completely excreted unchanged [Vahter et al., 1983, Marafante et al., 1984]. In fact, there are no reports to date of toxicity in man or animals from the consumption of organoarsenicals in seafood [WHO, 1988].

47 Figure 4.

Boxplots formetalconcentrations (µg.g Total Cadmium (ug.g-1) Total Arsenic (ug.g-1) Total Chromium (ug.g-1)

~ ~ ~ ~ N N c.> c.> 0 N .... 0) ex, 0 N 0 N c.> .... (11 0) (11 0 (11 0 (11 0 (11 t-1 I I I I I - I I I I I I

Baseline f- 0 - ~- (I]-, Coffin Bay f- ~ - ~ ID

Kangaroo Island f- - S. @• ~ . ~ . GAYLARD, S. THOMAS, &M.NELSON

-1 Adelaide Central >- - dry weight)intranslocatedmussels throughoutSouth Australia. ~· ~ • Adelaide North - ~· ~

48 ~- ~- Adelaide South ~· - ~ • - ~- Port Lincoln [H ~ ·~ Port Pirie f- ~ • - @• ·~· Port River ~ ~ - @ • •

- ·~ South East ~ ~ [I] ~ Whyalla ~ HI}l - ~ ~ Yorke Peninsula~ @ - @. ·~· - Figure 5. STATUS OFMETAL CONTAMINATION USING TRANSLOCATED MUSSELS Total Mercury (ug.g-1)

Boxplots formetalconcentrations (µg.g Total manganese (ug.g-1) Total Copper (ug.g-1) 0 0 0 0 0 0 0 0 ~ 0 0 0 0 0 ~ ~ ~ N .I>, O> ex, 0 ~ ~ ~ ~ 0 N .I>, O> ex, 0 N .I>, 0 0 0 0 0 0 0 N .I>, O> ex, 0 N .I>, O> ex,

Baseline f- ~ O•

Coffin Bay f-- ~ HIB Kangaroo Island f-- ~ • ~ -1 Adelaide Central f- dry weight)intranslocatedmussels throughoutSouth Australia •Kil--1• m • Adelaide North f.- • • +- ~ • 49 ~ Adelaide South f.- ~- t ,ITH• Port Lincoln f- ~ ID Port Pirie f- •HTI---i • 0------, • • • ~. f- Port River -~ • m-, . . ~ • MYTILUS GALLOPROVINCALIS South East f- DJ I ~

Whyalla f-- HIH 0 KTu Yorke Peninsula f- -Hil---1• ID S. GAYLARD, S. THOMAS, & M. NELSON

Comparisons to Food Standards Mussel metal concentrations were compared to the maximum residue limits in the Australian & New Zealand Standards Code [FSANZ, 2010]. There were a number of locations which exceeded the maximum residue limits as published by FSANZ (2009). The most significant of these were the excesses of lead throughout the entire Port Pirie region. This result was not unexpected, as already discussed. As a response to this contamination, the South Australian Department of Health requested Primary Industries and Resources Department of South Australia (PIRSA) to declare a shellfish collection prohibition zone around the Port Pirie region due to the elevated lead levels [SAGG, 1996]. Cadmium along the Adelaide metropolitan coastline (Adelaide North, Adelaide Central and Adelaide South regions) also frequently exceeded the FSANZ maximum residue limit (MRL) Guideline of 2.00 mg/kg. As mussels translocated in the three Adelaide regions are exposed to cadmium inputs via the three coastal wastewater treatment plants (~55kg/annum [NPI, 2010]), it is likely that stormwater would make up a similar proportion entering the marine environment. The sites which showed exceedances in cadmium followed no discernible spatial pattern: in some instances samples collected from further offshore were higher than adjacent inshore sites, and a number of locations significantly removed from terrestrial sources also exceeded the MRL, e.g. Kangaroo Island, Yorke Peninsula and Coffin Bay. These results suggest that terrestrial sources of cadmium may not be the cause of the elevated results. Therefore the likely source is naturally occurring cadmium, possibly from geological sources across Southern Australia. There are a number of authors who have quoted natural background levels of cadmium in marine water across Australia and the Untied States [ANZECC, 2000] and the ability for cadmium to accumulate in filter feeding organisms can lead to elevated concentrations. This comparison with FSANZ MRL’s was undertaken as a point of reference rather than as a program designed to test food safety, which is undertaken by a separate program designed specifically for this purpose [see SASQAP, 2009]. There are a number of methods used in this study that are not consistent with this protocol and as such may not be representative of the products as they are sold for food (e.g.: transplantation of mussels). Queries regarding the food safety of metal concentrations in mussels should be directed to the SASQAP program.

South Australia in a global perspective Metal concentrations in translocated mussels were compared to other published data from around the world. This has been carried out with caution because not all programs translocate mussels to study uptake, rather many programs harvest natural (wild) populations. This different methodology may result in slight differences in uptake rates if translocated mussels have not reached equilibrium with the environment or have been exposed for differing amounts of time, which may alter the actual concentration. Notwithstanding these considerations, in most cases metal levels in Mytilus galloprovincialis throughout South Australia were on a par with other locations around the world when monitoring methods were comparable. Cantillo (1998) describes the median metal concentration in a review of three major mussel contamination programs (World Mussel Watch, French Reseau National d’Observation de la Qualité du Mulieu Marin RNO and the US NOAA National Status & Trends program). With the exception of arsenic, the South Australian mussel data collected in this study as a consolidated program were found to have generally lower medians than these three programs. Arsenic was found to be significantly higher than the WMW, RNO & NS&T programs (total -1 As Median this program = 17.0; Median WMW = 7.1; Median NS&T = 9.6 µg.g ); possible natural geological sources are discussed above. A number of individual regions within this program had higher levels than those reported by Cantillo (1998) and also other authors. The most significant of these were concentrations of lead at Port Pirie, which were some of the highest mean lead concentrations (mean = 99.9 ug.g-1) found anywhere in literature, only one study reporting higher lead levels (mean = 480 µg.g-1) using similar methods (Riget et al, 1997). It is worthy to note that this site was located in close proximity to a lead/zinc mine in Greenland. The vast majority of lead levels published in the literature were in the range of 0 – 5.0 µg.g-1 [Deudero et al., 2009; Schintu et al., 2008; Acker et al., 2005; Andral, 2004; Cantillo, 1998].

50 STATUS OF METAL CONTAMINATION USING TRANSLOCATED MUSSELS MYTILUS GALLOPROVINCALIS

Conclusions

The PCA and the ranking of median metal levels indicated a clear progression from relatively low concentrations to high. Comparison to possible sources in each region has allowed a hierarchy to be developed with a clear separation between bioavailable metal loads in translocated M. galloprovincialis from industrialised regions (Port Pirie, Port River and Whyalla), to urbanised regions (Adelaide North, Adelaide Central and Adelaide South), and then to remaining rural/semi-rural regions throughout South Australia. Generally, bioavailable metal concentrations were significantly higher in the above mentioned industrialised regions, and the rank order was industrialised > urbanised > rural regions > baseline metal state. This suggests that activities currently occurring in the industrialised regions such as the lead/zinc smelter and steel manufacturing facilities are significantly contributing to, or have contributed historically to, metal loads which may still be affecting the near-shore coastal environment. Bioavailable lead and manganese concentrations in translocated mussels from Port Pirie were high across the whole region and while it is currently unclear whether these metals are coming from current practices at the lead/zinc smelter or are due to the significant historical contamination in the region, it is possible that the metal concentrations observed are damaging some sections of the ecology in the region. This issue is currently being been investigated further and could be the subject of additional publications. The conclusion to be drawn from this, should it be confirmed, would be that the industrialised regions may need further environmental regulation or remediation if improvement in bioavailable metal concentrations is desirable.

A comparison to the FSANZ Food Standards Code showed a number of exceedances compared to the maximum residue limits (MRL) particularly for lead and cadmium. All samples taken within the Port Pirie region exceeded the MRL for lead. This result was not unexpected given the long history of metal contamination in this region. Indeed, the Primary Industries and Resources and the Department of Health have implemented a ban on collection of shellfish throughout the Port Pirie region. There were a number of exceedances for cadmium particularly along the Adelaide metropolitan coast and it is likely that the source is natural background cadmium concentrations. Caution should be applied in interpreting comparison with Food standards because of differences in methodologies within this program compared to sampling undertaken specifically for this purpose.

This survey demonstrates that translocated mussels in the near-shore coastal waters of South Australia take up a range of metals but in most cases the concentrations are not outside of what would be considered to be normal given the adjacent terrestrial inputs when compared to a number of studies undertaken throughout the world. This study again shows the utility of translocating standardised mussels as a method to measure bioavailable metal levels in nearshore coastal waters. This study is the first regional assessment of metal levels to use translocated mussels in South Australia. The results observed here set a baseline which can be used as a marker for whether current regulation is adequate also as a comparison of the effectiveness of future regulation of discharges containing metals throughout regions of South Australia. Future work will need to investigate specific locations and assess sediment metal concentrations (dilute acid soluble), sediment toxicity and sediment ecology (the sediment quality triad) in order to assess fully the quantity and risk posed by metals in receiving environments.

Acknowledgements

The authors would like to acknowledge Glenn Boucher from Coffin Bay Fishing Charters, Allan Slater from Fish 'N Trips in Whyalla and Lawrie Commercial Diving Services for aiding in the collection of mussels. Additionally thanks to Ujang Tinggi and Pieter Scheelings from the Queensland Health Scientific Services for undertaking the analysis on mussel tissues. The authors would also like to thank Clive Jenkins from the EPA, David Cunliffe and Michelle Wittholz from the South Australian Department of Health, Vic Neverauskas and Clinton Wilkinson for reviewing the document and providing valuable feedback.

51 S. GAYLARD, S. THOMAS, & M. NELSON

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