Biological effects of gold mine tailings on the intertidal marine environment in Nova Scotia, Canada

MPB-07983; No of Pages 13 Marine Pollution Bulletin xxx (2016) xxx–xxx

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Biological effects of gold mine tailings on the intertidal marine environment in Nova Scotia, Canada K. Doe a,1, R. Mroz b,⁎, K.-L. Tay b,1, J. Burley b, S. Teh c, S. Chen d a

Environment and Climate Change Canada, Mount Uniacke, Nova Scotia, Canada Environment and Climate Change Canada, 45 Alderney Drive, Dartmouth, Nova Scotia B2Y 2N6, Canada University of California-Davis, Anatomy, Physiology, and Cell Biology, One Shields Avenue, 1321 Haring Hall, Davis, CA 95616-8732, USA d Canadian Food Inspection Agency, Dartmouth Laboratory, 1992 Agency Drive, PO Box 1060, Dartmouth, Nova Scotia B2Y 3Z7, Canada b c

a r t i c l e

i n f o

Article history: Received 8 December 2015 Received in revised form 6 July 2016 Accepted 22 August 2016 Available online xxxx Keywords: Biological effects Gold mine tailings Arsenic Mercury Molluscs Bioaccumulation

a b s t r a c t From 1861 to the 1940s, gold was produced from 64 mining districts in Nova Scotia, where mercury amalgamation was the dominant method for the extraction of gold from ore until the 1880s. As a result, wastes (tailings) from the milling process were contaminated by mercury and were high in naturally occurring arsenic. In 2004 and 2005, sediments, water and mollusc tissues were collected from 29 sampling stations at nine former gold mining areas along the Atlantic coastline and were analysed for arsenic and mercury. The resulting data were compared with environmental quality guidelines. Samples indicated high potential risk of adverse effects in the intertidal environments of Seal Harbour, Wine Harbour and Harrigan Cove. Arsenic in Seal Harbour was bioavailable, resulting in high concentrations of arsenic in soft-shell clam tissues. Mercury concentrations in tissues were below guidelines. This paper presents results of the sampling programs and implications of these findings. Crown Copyright © 2016 Published by Elsevier Ltd. All rights reserved.

1. Introduction Nova Scotia experienced its first gold rush in the 1860s after the discovery of gold in a quartz vein at Mooseland in 1858 (Bates, 1987). This was followed by two additional gold rushes which accounted for the development of N60 gold districts throughout Nova Scotia from the 1860s to the mid 1900s. During this time, total gold production for Nova Scotia was estimated to be 1.2 MOZ. The fine grained wastes from the milling process (tailings), were contaminated by mercury and arsenic and these contaminated tailings were generally deposited without any form of control into natural water bodies or into low lying areas next to water bodies. In Nova Scotia, mercury was used in gold mining for over 100 years, and the primary method used to extract the majority of gold during the three gold rushes was mercury amalgamation until the 1880s (Smith and Kontak, 1996), when it was supplemented by cyanidation (Bates, 1987). In general, 1 oz. of mercury was used for each ounce of gold recovered (Parsons et al., 2004). Of the total mercury used in the extraction process, 10–30% is typically lost per season (Alpers and Hunerlach, 2000) while another study found that up to 40% was lost

⁎ Corresponding author. E-mail addresses: [email protected] (K. Doe), [email protected] (R. Mroz), [email protected] (K.-L. Tay), [email protected] (J. Burley), [email protected] (S. Teh), [email protected] (S. Chen). 1 Retired.

when panning for gold (Ogola et al., 2002). The excess mercury was distributed with the tailings or lost through airborne emissions. Consequently, elevated mercury concentrations have been found in the environment associated with abandoned mine sites in Nova Scotia, although many of them have been closed for 70 years or more (Parsons et al., 2004). Eisler (2004) suggested that mercury has no beneficial biological function. Instead, its presence in living organisms causes neurotoxicity, birth defects and other adverse effects. Its ability to transform to more toxic forms (methyl mercury) and biomagnify within the food chain poses risk of adverse biological effects in ecosystems, especially to organisms in higher trophic levels (for example, fish eating mammals and birds), and in humans who consume mercury contaminated species. Arsenic is a naturally occurring element in many bedrock types. In Canada, arsenic occurs in gold ore deposit minerals such as arsenopyrite (FeAsS) (Eisler, 2004; Wang and Mulligan, 2006). Arsenopyrite is the most abundant arsenic containing mineral found and is commonly associated with gold (Francesconi and Kuehnelt, 2002). In Nova Scotia, historical mining activities resulted in the release of approximately 3 Mt of tailings containing 20,700 kg of arsenic (Wong et al., 1999). Since arsenic is found abundantly in the gold ore, when disturbed and altered, it can have serious repercussions on the environment and human health. The World Health Organization (WHO, 1992) reported that inorganic arsenic is a documented human carcinogen. The Agency for Toxic Substances and Disease Registry (2007) reports a variety of adverse effects of arsenic exposure in humans, including irritation of the stomach

http://dx.doi.org/10.1016/j.marpolbul.2016.08.056 0025-326X/Crown Copyright © 2016 Published by Elsevier Ltd. All rights reserved.

Please cite this article as: Doe, K., et al., Biological effects of gold mine tailings on the intertidal marine environment in Nova Scotia, Canada, Marine Pollution Bulletin (2016), http://dx.doi.org/10.1016/j.marpolbul.2016.08.056

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K. Doe et al. / Marine Pollution Bulletin xxx (2016) xxx–xxx

and intestines, blood vessel damage, skin changes, reduced nerve function, decreased production of red and white blood cells, skin cancer, and increased risk of cancer in the liver, bladder, and lungs. Environment Canada and Health Canada (1993) concluded that arsenic and its compounds are considered to be “toxic” to the environment and constitute a danger to human life or health. Gomez-Caminero et al. (2001) found that marine biota tend to accumulate much higher concentrations of arsenic than freshwater species. Neff (1997) suggested that the highest concentrations of arsenic are found in tissues of marine animals that feed primarily on phytoplankton or macroalgae. These species include planktonic crustaceans, bivalve molluscs, herbivorus snails, and some polychaete worms. Much of the bioaccumulated arsenic is present in a relatively nontoxic organic form, such as arsenobetaine (Cullen and Reimer, 1989; Francesconi and Kuehnelt, 2002; Phillips and Depledge, 1985). Mercury and arsenic have a long residence time in the environment although local concentrations may decline due to dispersion processes. Releases from gold mining operations pose potential risks to the ecosystems and human health either through direct exposure to tailings, or indirectly from contaminated air, water or through the consumption of arsenic or mercury contaminated organisms. Mollusc species can provide valuable insight on the availability of metal(oid)s to the food web. Dixon and Wilson (2000) suggest that mussels are a good choice for the detection of pollution because they accumulate a variety of chemical substances due to their filtration capacity, their contact with sediments and the water column, and because of their low mobility. The use of bivalves to assess the bioaccumulation of metal(oid)s has been used in many studies around the world. The National Oceanic and Atmospheric Administration's National Status and Trends Program for Marine Environmental Quality has been using mussels and oysters since 1986 to monitor spatial and temporal trends in contamination of the coastal USA (Valetee-Silver et al., 1999). The San Francisco Estuary Institute, after a five year period (1993–1997) of analyzing heavy metals in mussels, oysters, and clams, concluded that bivalves are effective tools for monitoring long-term trends and that they provide valuable information that water or sediment data alone would not supply (Hardin et al., 1999). In 2003, Natural Resources Canada initiated a study to examine the dispersion, speciation and fate of mercury and arsenic in terrestrial and shallow marine environments surrounding 14 abandoned gold mines in Nova Scotia (Parsons et al., 2012). The field studies confirmed that most of those sites contain large volumes of unconfined tailings and that gold mine tailings throughout Nova Scotia contain elevated concentrations of mercury and arsenic, which were widely dispersed in local sediments and surface waters. Based on the Natural Resources Canada findings, Environment Canada undertook studies in 2004 and 2005 to measure the magnitude and distribution of arsenic and mercury in sediments, water, and bivalve molluscs and this paper presents the results of these studies. 2. Methods and materials 2.1. 2004 study The 2004 study focused on six intertidal sampling stations in Seal Harbour, Nova Scotia, located in the immediate vicinity of the mouth of West Brook, the main source of transport for tailings that were originally deposited approximately 1 km upstream (Fig. 1). At each sampling station, location was determined on a hand-held Global Positioning System device and is presented in Table 1. Sediment and soft-shell clam (M. arenaria) samples were taken from each sampling station. All samples were collected on 9 August 2004. Intertidal sediment was sampled from a depth of 0–10 cm using disposable polystyrene spatulas. Soft-shell clams were depurated in a bucket full of water from the site for approximately 10 h, after which they were measured, weighed, and the meat was removed from the shell with stainless

steel disposable scalpels. All sediment and tissue samples were then frozen for transport to laboratories for analyses of arsenic and mercury. Clams sold for human consumption were purchased from a Moncton, NB, fish market and used as reference to compare with clams collected from Seal Harbour. 2.2. 2005 study The field program was expanded in 2005. In order to prioritize mining areas that could be having an impact on the marine environment, each of the 64 gold mining districts in Nova Scotia was assessed against the following criteria: • Sites which had a relatively high ore throughput (N10,000 t); • Sites located within 5 km of a marine receiving environment; • Sites which have the potential for tailings to enter a receiving environment and that are “classified shellfish growing areas” where shellfish might be harvested for human consumption; • Site accessibility; • Sites that have any “species at risk” that potentially might be present; • Shoreline should have intertidal sediments (i.e. favourable for clams, one of the indicator species to be collected during this study).

Nine former gold mining areas met the above criteria and were sampled in 2005, along with a reference area at New Harbour, for a total of 23 sampling stations. Their locations are shown on Fig. 1. The reference area was chosen at New Harbour, Nova Scotia, since it was from the same rock formation where the gold vein or lode deposits are predominantly located (the Meguma Supergroup slates and greywackes, Bates, 1987). No gold mining had been carried out in the New Harbour watershed area and so there was no influence from mine tailings. At Seal Harbour, because of the contamination identified in the 2004 study, a total of eight intertidal stations were chosen to examine the distribution of arsenic and mercury throughout the intertidal areas of the Harbour (Table 1), from 2.54 km west to 1.6 km east of the mouth of West Brook. Sampling occurred at low tide. At each sampling station, location was determined on a hand-held Global Positioning System device and is presented in Table 1. 100 mL of intertidal sediment was collected near the location of the clams from a depth of 0–10 cm using a new disposable polystyrene spatula. Water (250 mL) was collected adjacent to the blue mussels into an acid-washed plastic bottle. Five or more clams, and ten mussels (both N 5 cm in length), were collected. Softshelled clams, and mussels were depurated in a food grade polyethylene bucket full of water from the station for approximately 6 h. Water samples and sediment samples were kept cool. At the end of the day, clams and mussels were measured, weighed, and the measurements recorded. The tissues were removed from the shell using new stainless steel disposable scalpels for each species at each station. The meat of all soft-shelled clams were pooled for each sampling station, and the meat of all blue mussels were pooled for each sampling station, and placed in acid-washed glass jars. Water samples were preserved using a few drops of nitric acid. All samples but water were then frozen for transport to laboratories for analyses. At each sampling station, samples were taken for: total arsenic and mercury in unfiltered seawater; total arsenic and mercury in sediments; total arsenic and mercury in tissues of soft-shell clam (M. arenaria) and blue mussel (Mytilus edulis); and inorganic arsenic speciation as arsenic (III) and arsenic (V) in tissues of clams and mussels. As a quality assurance measure and to ensure that data were comparable between the laboratories, homogenized and split samples of clam tissue (29% of samples, n = 4) and sediment (30% of samples, n = 7) were sent in 2005 to both the Environment Canada laboratory in Moncton, NB and to the laboratories used in 2005 for the majority of analyses for comparative purposes. The samples were thoroughly homogenized so that a representative sample could be used for analysis by each laboratory. The tissue samples were

Please cite this article as: Doe, K., et al., Biological effects of gold mine tailings on the intertidal marine environment in Nova Scotia, Canada, Marine Pollution Bulletin (2016), http://dx.doi.org/10.1016/j.marpolbul.2016.08.056

K. Doe et al. / Marine Pollution Bulletin xxx (2016) xxx–xxx

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Gold Mine Sampling Locations – 2004 and 2005

Fig. 1. Study sites for the 2004 and 2005 sampling program, Nova Scotia, Canada.

placed in a clean blender jar and homogenized on a blender until smooth (approximately 2 to 4 min). The jar and blade were cleaned between samples. The blended sample was stored in the original sample jar or pre-cleaned jar if it needed to be transferred. For sediment samples, large pieces of rock and vegetation were removed and discarded. The sediment samples were transferred to a wide mouth Wheaton® bottle and mixed and stirred in the sample container until homogenous, using a large disposable polystyrene spatula. 2.2.1. Analysis of arsenic and mercury in sediment, tissue, and water For the 2004 study, all samples were sent to the Environment Canada laboratory in Moncton, New Brunswick. For total arsenic, sediments and tissues were digested in strong acid (US EPA, 1996) using a microwave, and measured using inductively coupled plasma-mass spectrometry (ICP-MS). Results were reported on a dry weight basis for sediment, and as wet weight for tissue samples. Mercury content of sediments and tissues was determined by wet oxidation digestion and measured using cold vapour atomic absorption spectrometry (US EPA, 1994a). Results were reported as dry weight basis for sediment, and as wet weight for tissue samples. For the 2005 study, samples were sent to three laboratories. Sediment and water samples were sent to Maxxam Analytics Inc. in Bedford, Nova Scotia for analysis of total arsenic and mercury. Total arsenic in sediment and water was analysed following US EPA Method 6020 (US EPA, 1994b), using inductively coupled plasma-mass spectrometry (ICP-MS). Total arsenic in sediment was reported on a dry weight basis. Mercury in water and sediment was determined using US EPA Methods 245.2 (US EPA, 1983a) and 245.5 (US EPA, 1983b) respectively, which reduce mercury compounds and ions to the elemental state of mercury vapour to analyze total mercury. The vapor passes through a

drying tube and through a flow cell positioned in the light path of an atomic absorption spectrophotometer. Sediment results were reported as dry weight. Blue mussel and soft-shell clam tissue samples were sent to the Canadian Food Inspection Agency (CFIA), Dartmouth Laboratory in Nova Scotia, for the analysis of mercury, total arsenic and arsenic speciation as As (III) and As (V). For mercury, tissue was digested with nitric acid using a closed vessel microwave digester. The digests were then analyzed by a Varian 600Z series AAS, with mercury analyzer using a VGA-77 hydride generator attached. Mercury in tissues was reported as wet weight. For the analysis of total arsenic, tissue was digested in an open vessel digester with a mixture of nitric acid and sulfuric acid. A small amount of peroxide was added to break up lipids. A final cleanup step using hydrogen peroxide was performed in the microwave to reduce the sulfate interference prior to analysis by mass spectrometry. The digests were analyzed through the ICP in DRC (Dynamic Reaction Cell) mode. For the analysis of As (III) and As (V), approximately 1 g of tissue, 20 mL of 75% (3:1) methanol water, 100 μL of protease at 20 mg/100 μL and 100 IU of lipase was added to a 50 mL polypropylene centrifuge tube for each sample. The tubes were capped tightly and put on a rotating shaker for 18–20 h in a 37 °C incubator. The resulting extract was allowed to settle for approximately 0.5 h and then a portion was filtered through a 0.45 μm filter. Approximately 200 μL of the extract was analyzed by ICP-MS. Samples were then compared to standards of mixed, arsenobetaine, arsenocholine, methyl-arsonate, dimethyl-arsinate, As (III), As (V) at 20, 40, 60, 80, and 100 ppb concentrations. At the time of the analysis, there were no certified reference materials available for As (III) and As (V), so spike recoveries were done on individual tissue types. The CFIA lab has finalized these methods as internal CFIA methods.

Please cite this article as: Doe, K., et al., Biological effects of gold mine tailings on the intertidal marine environment in Nova Scotia, Canada, Marine Pollution Bulletin (2016), http://dx.doi.org/10.1016/j.marpolbul.2016.08.056

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K. Doe et al. / Marine Pollution Bulletin xxx (2016) xxx–xxx

Table 1 Location of sampling stations, and results for arsenic and mercury in sediment samples (mg/kg dry weight), 2004 and 2005 surveys. Site name

GPS: North

GPS West

As

Hg

2004 Seal Harbour 1 Seal Harbour 2 Seal Harbour 2B Seal Harbour 2C Seal Harbour 3 Seal Harbour 4

45° 09′ 36.9″ 45° 09′ 37.7″ 45° 09′ 37.9″ 45° 09′ 38.9″ 45° 09′ 38.3″ 45° 09′ 36.6″

061° 34′ 45.3″ 061° 34′ 44.8″ 061° 34′ 44.4″ 061° 34′ 45.2″ 061° 34′ 40.4″ 061° 34′ 34.4″

688 684 767 457 568 608

0.21 0.30 0.49 0.33 0.41 0.32

45° 09.618′ 45° 09.509′ 45° 09′ 30.7″ 45° 09′ 06.8″ 45° 09′ 36.4″ 45° 09′ 39.0″ 45° 09′ 43.6″ 45° 09′ 42.2″ 45° 09′ 47.0″ 45° 04.710′ 45° 04.651′ 45° 04′ 25.4″ 45° 04′ 25.6″ 44° 49.315′ 44° 55.759’ 44° 04.240′

061° 34.717′ 061° 34.894′ 061° 34′ 49.4″ 061° 34′ 35.8″ 061° 34′ 34.5″ 061° 34′ 34.2″ 061° 34′ 25.8″ 061° 34′ 13.8″ 061° 34′ 07.4″ 061° 49.551′ 061° 49.584′ 061° 50′ 47.6″ 061° 51′ 46.7″ 062° 38.057′ 062° 17.609′ 061° 58.787

44° 54.681′ 44° 38′ 57.4″ 44° 36′ 50.8″ 44° 36′ 50.8″

062° 23.052′ 063° 21′ 33.5″ 063° 26′ 14.2″ 063° 26′ 12.3″

45° 10.930′

061° 27.529′

680 140 47 12 74 19 21 18 10 860 96 300 650 16 400 59 39 31 3.3 b2.0 2.6 22 b2.0

0.47 0.97 0.09 b0.02 0.07 0.02 0.06 0.03 0.03 0.05 0.18 2.5 3.97 b0.02 6.47 0.09 0.07 0.02 b0.02 b0.02 b0.02 b0.02 b0.02

2005 Seal Harbour 1 (mud flats) Seal Harbour 2 A (Long Cove) Seal Harbour 2 (Long Cove) Seal Harbour 3 Seal Harbour K #1 Seal Harbour K #2 Seal Harbour K #3 Seal Harbour K #4 Seal Harbour K #5 Wine Harbour #1 Wine Harbour #2 Wine Harbour # 3 A Wine Harbour # 3B Pope′s Harbour Harrigan Cove Gegogan #1 Gegogan #3 Port Dufferin Lawrencetown Cow Bay 1 Cow Bay 2 Gold River New Harbour (reference site)

All samples reported as below detection limit were recorded as one half of the method detection limit for calculation purposes. 3. Results and discussion 3.1. 2004 study

Underlined and bold values exceed the CCME (1999) Probable Effects Level (41.6 mg/kg for arsenic, and 0.7 mg/kg for mercury). Underlined values exceed the Interim Sediment Quality Guideline (7.24 mg/kg for arsenic, and 0.13 mg/kg for mercury) but are less than the Probable Effects Level. b = below the detection limit.

2.2.2. Statistical analysis Regression analyses were performed on the 2005 Seal Harbour samples using the data from both east and west of West Brook (source of tailings) to determine if there was a relationship between distance from source and concentration. Separate regression analyses were performed for sites located east and west of West Brook. Statistical analyses were performed using Minitab 14 software. All samples reported as below detection limit was recorded as half of the detection limit for statistical purposes. Pearson's r correlation was used to test the strength of the relationships. All statistical relationships were considered significant at P b 0.05. The relationship between arsenic and mercury in sediment, water, clam tissue and mussel tissue was examined by Pearson Product Moment Correlations using the computer program Sigmastat (Systat Software Inc., 2004). 2.2.3. Calculation of bioaccumulation factors Bioaccumulation factors are useful in assessing the varying degrees of accumulation of a contaminant by species with different modes of living (Ruus et al., 2005), and to determine if the contaminant is entering the food web. Bioaccumulation factors provide a ratio between the concentration found in the environmental medium and the concentration found in the organism. BAF's were calculated as reported in Meador et al. (2004) and USEPA (2000). The formulae for BAF's are as follows;

BAF from sediment ¼

BAF from water
total arsenic ðdry weightÞ in the tissue of the organism Ë cg=kg
¼ total arsenic in the water Ë cg=L

total arsenic ðdry weightÞ in the tissue of the organism ðmg=kgÞ total arsenic ðdry weightÞ in the sediment ðmg=kgÞ

3.1.1. Arsenic and mercury in sediments, and mollusc tissues Results of the analysis of arsenic in intertidal sediment samples from Seal Harbour collected near the mouth of West Brook are presented in Table 1, and shown in Fig. 2. The measured concentrations were compared with the Canadian Council of Ministers of the Environment (CCME) Sediment Quality Guidelines (CCME, 1999). CCME published sediment quality guidelines for use as benchmarks for evaluating the potential for adverse biological effects in aquatic ecosystems. Two guideline values have been published. The lower value is the Interim Sediment Quality Guideline (ISQG), below which adverse biological effects are expected to occur rarely. The upper value is the Probable Effects Level (PEL), above which adverse biological effects are expected to occur frequently. For marine sediments, the arsenic ISQG is 7.24 mg/ kg, and the PEL is 41.6 mg/kg. Arsenic in all six sediment samples exceeded the PEL by a factor of 11.0 to 18.4 times, indicating that the sediments pose a high risk to cause adverse biological effects. MartınDıaz et al. (2008) studied the relationship between chemical concentration in sediments and histopathological lesions, and used the results to suggest sediment quality guidelines for As for the clam Ruditapes philippinarum as follows: ‘not hazardous’ As b 272.78 mg/kg dry weight, and ‘hazardous’ As N532.27 mg/kg dry weight. Arsenic in intertidal sediment samples from Seal Harbour collected near the mouth of West Brook are in the ‘hazardous’ range as suggested by these authors (Table 1). Concentrations of mercury in sediment are presented in Table 1, and shown in Fig. 3. For mercury in marine sediments, the ISQG is 0.13 mg/ kg, and the PEL is 0.70 mg/kg (CCME, 1999). All six sediment samples exceeded the ISQG value, but all were below the PEL, indicating that the mercury concentrations in sediments pose a low to moderate risk to cause adverse biological effects. The concentrations of arsenic in the soft-shell clam tissues are presented in Table 2. The mean arsenic value for the Seal Harbour clams was 1200 ± 565 mg/kg (±SD, range 508 to 2325, n = 10, dry weight). For the five reference clams, purchased at a Moncton, New Brunswick food store, mean value was 12.7 ± 6.69 mg/kg (± SD, range 7.7 to 24.3, n = 5, dry weight). The arsenic in the Seal Harbour clams was approximately 100 times higher than in the commercially available control clams sold for human consumption. There is no Health Canada Canadian standard or guideline for acceptable arsenic concentrations in molluscs. The concentrations of arsenic in soft-shell clam tissues reported in this study are higher than tissue concentrations previously reported in the scientific literature (Table 3). The mean mercury value for the Seal Harbour clams is presented in Table 2 and was 0.098 ± 0.028 mg/kg (±SD, range 0.07 to 0.15, N = 10, wet weight). For the five reference clams, purchased at a local food store, the mercury tissue concentrations were non-detectable (b0.02 mg/kg, wet weight). The mercury in the Seal Harbour clams was ≥ 5 times higher than in the commercially available reference clams, but still well below the Australian food standard for mercury in molluscs of 0.5 mg/kg, and the Danish rejection limit for crustaceans/molluscs of 0.3 mg/kg (Rayment, 1991), and within the mid-range of mercury in shellfish reported by other studies (Table 3). There is no Health Canada Canadian standard or guideline for acceptable mercury concentrations in molluscs.

Please cite this article as: Doe, K., et al., Biological effects of gold mine tailings on the intertidal marine environment in Nova Scotia, Canada, Marine Pollution Bulletin (2016), http://dx.doi.org/10.1016/j.marpolbul.2016.08.056

K. Doe et al. / Marine Pollution Bulletin xxx (2016) xxx–xxx

5

900 800 700

As (mg/kg)

600 500

400 300 200 100 0 Stn #1

Stn #2

Stn #2 B

Stn #2 C

Stn #3

Stn #4

= CCME PEL (41.6 mg/kg)

Sampling Locations

Fig. 2. Arsenic (mg/kg dry weight) in intertidal marine sediment in Seal Harbour, 2004.

3.1.2. Bioaccumulation factors of arsenic from sediments to clam tissues For the 2004 samples collected at the mouth of West Brook, the bioaccumulation factor for arsenic from sediment to clam tissues was calculated to be 2.6 (Table 2). The bioaccumulation factor for mercury from sediment to clam tissues was calculated to be 2.0 (Table 2). A bioaccumulation factor of N 1 indicates that the metal(oid) in the sediment was bioavailable to the clams, and that the clams were concentrating the arsenic in their tissues to a concentration higher than present in the sediment they live in. This will be discussed further in the discussion of the 2005 results.

3.2. 2005 study 3.2.1. Arsenic and mercury in sediments, water, and mollusc tissues Mercury concentrations in intertidal sediments from the nine study locations and from the reference location along the southern and eastern shores of Nova Scotia are presented in Table 1 and Fig. 4. One sampling station in Seal Harbour, as well as sediments from Wine Harbour

and Harrigan Cove exceeded the Probable Effects Level (PEL) for mercury, indicating risk of adverse biological effects in these areas. Similar high mercury concentrations in marine sediments from Wine Harbour were reported by Little et al. (2015). Using different sampling stations from our study, they found that the mercury burdens in sediments from Wine Harbour are particularly high, in agreement with our findings. They suggested that this reflected the continuous use of Hg amalgamation for the duration of gold production in the Wine Harbour district. Walker and Grant (2015) found mercury concentrations in sediments in Isaacs Harbour (immediately West of Seal Harbour) near historical land-based gold mining activities that ranged from b 0.05 to 0.16 mg/kg, similar to the concentrations that we found in the lower contaminated stations K1 to K5 of Seal Harbour, as well as in Port Dufferin, Pope's Harbour, Gold River and Gegogan study locations, but much lower than the concentrations that we found in Wine Harbour, Harrigan Cove, and the more contaminated stations in Sea Harbour. Mercury concentrations in sediments collected at the Mouth of West Brook in 2005 was 0.47 mg/kg dry weight, within the range of values collected from this location in 2004 (0.21 to 0.49 mg/kg dry weight). We found that

0.8 0.7

Hg (mg/kg)

0.6 0.5 0.4 0.3 0.2

Sampling Locations

STN # 4

STN # 3

STN # 2C

STN # 2B

STN # 2

0

STN # 1

0.1

= CCME PEL (0.7 mg/kg)

Fig. 3. Mercury (mg/kg dry weight) in intertidal marine sediment in Seal Harbour, 2004.

Please cite this article as: Doe, K., et al., Biological effects of gold mine tailings on the intertidal marine environment in Nova Scotia, Canada, Marine Pollution Bulletin (2016), http://dx.doi.org/10.1016/j.marpolbul.2016.08.056

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K. Doe et al. / Marine Pollution Bulletin xxx (2016) xxx–xxx

Table 2 The concentrations of arsenic and mercury in soft-shell clam and in blue mussel tissue (mg/kg wet weight), 2004 and 2005 surveys, and calculated bioaccumulation factors from sediment and water. Site name

Soft-shell clams As

2004 Seal Harbour near mouth of West Mean = 160 (range 67.4 to 309) (As dry weight, Brook, N = 10 mean = 1200; range 508 to 2325) Control clams, N = 5 Mean = 1.68 (range 1.06 to 3.19) (As dry weight, mean = 12.7; range 7.7 to 24.3) 2005 Seal Harbour 1 (mud flats) Seal Harbour 2A (Long Cove) Seal Harbour 2 (Long Cove) Seal Harbour 3 Seal Harbour K #1 Seal Harbour K #2 Seal Harbour K #3 Seal Harbour K #4 Seal Harbour K #5 Wine Harbour #1 Wine Harbour #2 Wine Harbour #3A Wine Harbour #3B Pope's Harbour Harrigan Cove Gegogan #1 Gegogan #3 Port Dufferin Lawrencetown Cow Bay 1 Cow Bay 2 Gold River New Harbour (reference site)

259 32.3 – 9.10 32.6 21.1 13.5 22.0 10.1 – 6.79 – – – 1.85 45.8 9.18 – 2.47 – – – 1.62

Mussels As sediment BAFa

As water BAFb

Hg

Hg sediment BAFa

As

Hg

2.6

NC

2.2

–

–

NC

NC

0.098 ± 0.028 (range 0.07 to 0.15) b0.02 ± 0.0 (no range)

NC

–

–

2.8 1.7

5220 NC

2.0 0.77

5.73 3.3 8.4 4.9 9.2 7.6

6870 24,500 5680 1670 4580 961

0.53

5110

0.03 5.8 1.8

178 NC NC

5.6

1860

N2.0

1210

0.13 0.10 – 0.09 0.11 0.13 0.12 0.12 0.10 – 0.06 – – – 0.11 b0.05 b0.05 – 0.04 – – – 0.05

6.14 – 6.35 4.88 5.72 7.72 7.85 5.83 4.55 7.26 – 6.94 – 2.07 – 3.66 – 2.44 1.82 – – 10.6 1.76

0.09 – 0.08 0.07 0.1 0.09 0.09 0.10 0.10 0.14 – 0.13 – b0.05 – b0.05 – 0.06 b0.05 – – b0.05 0.07

68 11.9 49 15 30 25 45

0.1 NC NC 30

2.1

– = no sample collected; NC = cannot be calculated. a Bioaccumulation factor for As and Hg from sediment to tissue. b Bioaccumulation factor for As from water to clam tissue.

the maximum exceedance of the mercury PEL was at one station at Harrigan Cove, an exceedance by a factor of 9.2 times. Arsenic concentrations in intertidal sediments from the nine study locations and from the reference location along the southern and eastern shores of Nova Scotia are presented in Table 1 and Fig. 5. The PEL for arsenic was exceeded at several stations in Seal Harbour, all stations at Wine Harbour, and at Harrigan Cove, with an exceedance of 10 times or more at one or more sampling sites at each of these locations indicating a high risk of adverse biological effects in these areas. The maximum exceedance of the arsenic PEL was at Wine Harbour, where the samples exceeded the guideline by a factor of 42 times. Arsenic in intertidal sediment samples from Seal Harbour collected near the mouth of West Brook and from two stations in Wine Harbour are in the ‘hazardous’ range as suggested by Martın-Dıaz et al. (2008). Loring et al. (1996) reported that background concentrations of arsenic in sediments of coastal embayments of Nova Scotia were approximately 20 mg/kg. Concentrations found in the present study ranged from b 2.0 mg/kg at the reference location (New Harbour) to 860 mg/kg at one location in Wine Harbour. Similar findings of high arsenic concentrations in tailings-contaminated marine sediments in Wine Harbour (range 4–1500 mg/kg) were made by Little et al. (2015). Walker and Grant (2015) found that arsenic concentrations in sediments in Isaacs Harbour (immediately West of Seal Harbour) near historical land-based gold mining activities ranged from 5 to 40 mg/kg, similar to the concentrations that we found in the lower contaminated stations K2 to K5 of Seal Harbour, as well as in Port Dufferin, Pope's Harbour and Gold River mining areas. We found that the concentrations of arsenic in sediment at several sampling stations in Seal Harbour, Wine Harbour, and Harrigan Cove exceeded the concentration found in the New Harbour reference location by a factor of N 50 times, indicating the presence of mine tailings

in the intertidal marine environment, though some of the arsenic may have originated from other processes such as bio-deposition, solubilisation and subsequent bioaccumulation, or other complexing processes. Arsenic concentrations in sediments collected at the Mouth of West Brook in 2005 was 680 mg/kg dry weight, within the range of values collected from this location in 2004 (457 to 787 mg/kg dry weight). Concentrations of arsenic and mercury in unfiltered water samples from the nine study locations and the reference location along the southern and eastern shores of Nova Scotia are presented in Table 4. Arsenic concentrations in several samples collected from stations in Seal Harbour, Wine Harbour, and Harrigan Cove exceeded the CCME (1999) Guideline for the Protection of Marine Aquatic Life (12.5 μg/L), indicating potential risk of adverse biological effects to aquatic organisms in these areas. Mercury was not detected in any of the water samples except at one station in Wine Harbour, where it exceeded the CCME (1999) Guideline for the Protection of Marine Aquatic Life (0.016 μg/L), indicating potential risk of adverse biological effects to aquatic organisms in that area. It should be noted that the detection limit of the analytical methods used for arsenic and mercury water sample analyses were too high (As = 20 μg/L; Hg = 0.05 μg/L) to detect small exceedances of the guideline value. Clams could only be collected at five of the nine study locations in 2005. Arsenic concentrations in soft-shell clam tissue from these five study locations ranged from 1.85 to 259 mg/kg wet weight, while the arsenic concentration in soft-shell clam tissue from the reference location was 1.62 mg/kg wet weight (Table 2). The arsenic in the clam tissue from the study locations was up to 160 times higher than arsenic concentrations in clam tissue from the reference location. Arsenic in clam tissue from five of the eight Seal Harbour sampling stations where

Please cite this article as: Doe, K., et al., Biological effects of gold mine tailings on the intertidal marine environment in Nova Scotia, Canada, Marine Pollution Bulletin (2016), http://dx.doi.org/10.1016/j.marpolbul.2016.08.056

K. Doe et al. / Marine Pollution Bulletin xxx (2016) xxx–xxx

7

Table 3 Summary of reported concentrations of arsenic and mercury in whole tissue of bivalve molluscs from sites around the world. Species

Concentration of arsenic and mercury reported in tissues

Site

Reference

Mytilus edulis Mya arenaria Mytilus edulis

Range of As: 0.918–2.33 μg/g WW Range of As: 0.385–1.05 μg/g WW Average of As: 17 mg/kg DW

New York-New Jersey Harbor Estuary, USA

Skinner et al. (1997) Veinott et al. (2003)

Mya arenaria

Average of As: 0.82 μg/g WW Average of Hg: 0.01 μg/g WW Range of As: 5.8–11.7 μg/g DW Range of As: 5.4–11.5 μg/g DW Range of As: 7.1–13.7 μg/g DW Range of Hg: 0.10–0.063 μg/g DW Average of Hg at three stations: 660, 323, 412 ng/g DW Range of As: 3.1–3.4 mg/kg DW Range of Hg: 0.0004–0.014 mg/kg DW Range of Hg: 0.01–0.05 μg/g WW Mean of As: 25.4 μg/g DW Highest As: 66 μg/g DW Range of As: 3 to 30 μg/g DW

Little Bay Arm, Newfoundland and Labrador, Canada St. Lawrence River Lower Estuary, Canada

Gagnon et al. (2004)

San Francisco Bay, USA

Johns and Luoma (1990)

Galicia, NW Iberian Peninsula, Spain

Beiras et al. (2003)

Pialassa Baiona, Italy

Cattani et al. (1999)

King County Beaches, Washington State, USA

Stark (1998)

Port Curtis, Scotland Southeast Atlantic and eastern Gulf of Mexico Coasts (1986–1992) 10 Pacific Coast (USA) sites, 6 in California and 4 in Alaska. 2 locations in Passamaquoddy Bay, Maine, USA

Davies (1981) Valetee-Silver et al. (1999)

Macoma balithica Corbicula sp. Mytilus galloprovincialis Mytilus galloprovincialis Native and Japanese littleneck clams Periwinkle Oysters Crassostrea virginica Several species of clams Soft-shell clam Three species of bivalve molluscs 192 samples of bivalves and snails Mytilus edulis Mytilus edulis Mytilus edulis Mytilus galloprovincialis Manila clam Ruditapes philippinarum Mytilus galloprovincialis Mytilus galloprovincialis Mactra ovalina Mytilus edulis Scrobicularia plana Bivalve (Ostrea sp.) Mya arenaria

Mytilus edulis

Range of As: 2.9 to 8.1 μg/g DW Range of Hg: 0.11–0.33 μg/g DW Range of As: ND to 225 mg/kg DW Range of As: ND to 0.13 mg/kg DW Range of As: b0.6 to 533 μg/g DW Range of As: 5.0 to 14.3 μg/g DW Range of As: majority b3.0 μg/g, maximum 13.8 μg/g WW Range of As: 1.29–1.97; Mean ± SD: 1.66 ± 0.24 Range of As: 2.2 to 3.1 μg/g WW Range of As: 0.46–11.95 mg/kg DW

Meador et al. (2004)

7 locations in Sarawak, Malaysia

Environment Department of the Passamaquoddy Tribe at Pleasant Point (2001) Lau et al. (1998)

Worldwide (literature review)

Neff (1997)

10 locations in New Brunswick and NS, Canada 175 samples, various locations along Norwegian coastline. 6 northern big coastal cities of China

Engel et al. (2006) Sloth and Julshamn (2008) Li and Gao (2014)

2 stations over 2 years in Izmir Bay, Turkey 9 sampling sites along the coast of China.

Kucuksezgin et al. (2014) Yang et al. (2014)

Range of As: 14.7 ± 1.3, N = 3 to 29.9 ± 0.6, N = 3 mg/kg DW Range of As: 6.39–17.2 mg/kg DW

3 stations, over 3 seasons, Gulf of Trieste, Slovenia 3 surveys over 2 decades at sampling sites on the N-NW Spanish Coast Range of As: 0.7 to 2.6 μg/g DW Intertidal areas of the Dar es Salaam coast, Tanzania Range of As: 34 to 109 μg/g DW 4 sites in Seal Harbour and 1 in Coddles Harbour, NS Average of As at each station: 24, 27, 23, 18, 5 sites in Ria de Aveiro, Portugal and 13 mg/kg DW 15 stations from Rosetta beach, Mediterranean Average: 0.759 (0.006–2.496 μg/g)a Sea, Egypt Range of As: 1.62 to 309 mg/kg WW Ten locations along the Atlantic Coast of NS, Range of As: 12.1 to 2325 mg/kg DW Canada Range of Hg: b0.02 to 0.15 mg/kg WW Range of As: 1.76 to 10.6 mg/kg WW Range of Hg: b0.05 to 0.15 mg/kg WW

Kristan et al. (2014) Besada et al. (2014) Rumisha et al. (2012) Whaley-Martin et al. (2012) Ereira et al. (2015) El-Sorogy and Attiah (2015) This study

Notes: WW indicates results expressed as wet weight; DW indicates results expressed as dry weight; ND means below the detection limit. a Wet or dry weight not specified.

clams were collected was N 10 times higher than arsenic concentrations in clam tissue from the reference location. Similar concentrations of arsenic in molluscs from the Seal Harbour area were reported by Koch et al. (2007) in clams (M. arenaria) and by Whaley-Martin et al. (2013) in periwinkles (Littorina littorea). Arsenic in the clam tissue collected at the Mouth of West Brook in 2005 was 259 mg/kg wet weight, within the range of values collected from this location in 2004 (67.2 to 309 mg/ kg wet weight). Blue mussels could only be collected at seven of the nine study locations in 2005. Arsenic concentrations in blue mussel tissue from these seven study locations ranged from 1.82 to 10.6 mg/kg wet weight, while arsenic concentrations in blue mussel tissue from the reference location was 1.76 mg/kg wet weight (Table 2). Walker and Grant (2015) found that arsenic concentrations in blue mussel tissue from Isaacs Harbour (immediately West of Seal Harbour) near historical land-based gold mining activities ranged from 1.3 to 2.0 mg/kg, similar to the concentrations that we found in blue mussel tissue from

Lawrencetown and Harrigan Cove study locations, but much lower than the concentrations that we found in the other study locations. The arsenic in the mussel tissue from the study locations was up to 6.0 times higher than arsenic concentrations in mussel tissue from the reference location, but much lower than concentrations found in soft-shell clams. Similar concentrations of arsenic in blue mussels (M. edulis) from several stations in Seal Harbour, NS were reported by Whaley-Martin et al. (2012). Mercury concentrations in soft-shell clam tissue from the study locations ranged from 0.04 to 0.13 mg/kg wet weight, while mercury concentrations in soft-shell clam tissue from the reference location were 0.05 mg/kg wet weight (Table 2). Mercury concentrations in blue mussel tissue from the study locations ranged from b 0.05 to 0.14 mg/kg wet weight, while mercury concentrations in blue mussel tissue from the reference location was 0.07 mg/kg wet weight (Table 2). Walker and Grant (2015) found that mercury concentrations in blue mussel tissue from Isaacs Harbour (immediately West of Seal Harbour) near historical

Please cite this article as: Doe, K., et al., Biological effects of gold mine tailings on the intertidal marine environment in Nova Scotia, Canada, Marine Pollution Bulletin (2016), http://dx.doi.org/10.1016/j.marpolbul.2016.08.056

K. Doe et al. / Marine Pollution Bulletin xxx (2016) xxx–xxx

Hg (mg/kg)

8

Sampling Locations

= CCME PEL (0.7 mg/kg)

Fig. 4. Mercury (mg/kg dry weight) in intertidal marine sediment from 2005 study sites.

Pearson Product Moment Correlations were performed on data from all sampling locations to examine the relationship between arsenic and mercury in sediment and water samples with arsenic and mercury in mollusc tissues from the two species using the computer program SigmaStat Version 3.1 (Systat Software Inc., 2004). Highly significant correlations were found between arsenic in sediment and arsenic in clams (r2 = 0.824, P = 0.00099, N = 12), arsenic in water and arsenic in clams (r2 = 0.952, P = 0.000006, N = 11), and arsenic in sediment and arsenic in water (r2 = 0.883, P = 0.0003, N = 11). These relationships are shown in Fig. 6. Arsenic in mussels was not significantly correlated with arsenic in sediment or with arsenic in water. Mercury in sediment was not significantly correlated with mercury in clams or in mussels. Most water samples had mercury concentrations below the analytical detection limits and so no correlations were attempted with mercury in water. 3.2.2. Bioaccumulation factors of arsenic from sediments and water to clam tissues Because there was a strong correlation between arsenic in clam tissues and arsenic in sediment and water for the 2005 samples, it was decided to calculate bioaccumulation factors (BAFs) for arsenic from sediment to clam tissues and from water to clam tissues (Table 2). A

As (mg/kg)

land-based gold mining activities ranged from 0.02 to 0.05 mg/kg, similar to the concentrations that we found in blue mussel tissue from the Pope's Harbour, Gegogan, Port Dufferin, Lawrencetown and Gold River study locations, but lower than the concentrations that we found at the Seal Harbour and Wine Harbour mining areas. We found that the mercury in mussel tissue from the Seal Harbour and Wine Harbour study locations was up to 2.6 times higher than mercury concentrations in tissue from the reference locations, but all locations were well below the Australian food standard for mercury in molluscs of 0.5 mg/kg, the Danish rejection limit for crustaceans/molluscs of 0.3 mg/kg (Rayment, 1991), and within the mid-range of mercury in shellfish reported by other studies (Table 3). The additional sampling stations in Seal Harbour in 2005 were added at increasing distances from the mouth of West Brook (source of tailings material to Seal Harbour). We examined the relationship between total arsenic and mercury in sediment, clams, and mussels versus distance from source. Regression analysis showed no statistically significant relationships at P = 0.05, but we found a general pattern of decreasing concentrations with an increase of distance from West Brook (Tables 1 and 2). The strongest patterns are seen with the concentration of total arsenic in sediment and soft-shelled clams collected east from West Brook (Seal Harbour stations 1, K#1, K#2, K#3, K#4, and K#5).

Sampling Locations

= CCME PEL (41.6 mg/kg)

Fig. 5. Arsenic (mg/kg dry weight) in intertidal marine sediment from 2005 study sites.

Please cite this article as: Doe, K., et al., Biological effects of gold mine tailings on the intertidal marine environment in Nova Scotia, Canada, Marine Pollution Bulletin (2016), http://dx.doi.org/10.1016/j.marpolbul.2016.08.056

K. Doe et al. / Marine Pollution Bulletin xxx (2016) xxx–xxx Table 4 Results for arsenic and mercury in water samples (μg/L), 2005 survey. Site name

Hg

As

Seal Harbour Stn 1 — mud flats Seal Harbour Stn 1 — streamb Seal Harbour Stn 2 Seal Harbour Stn 3 Seal Harbour K #1 Seal Harbour K #2 Seal Harbour K #3 Seal Harbour K #4 Seal Harbour K #5 Wine Harbour #1 Wine Harbour #2 Wine Harbour #3A Pope's Harbour Harrigan Cove Port Dufferin Lawrencetown Cow Bay Gold River Gold River — duplicate test New Harbour (reference site)

b0.05 b0.05 b0.05 b0.05 b0.05 b0.05 b0.05 b0.05 b0.05 b0.05 b0.05 0.62 b0.05 b0.05 b0.05 b0.05 b0.05 b0.05 b0.05 b0.05

370.0a 380.0 35.0 b20 b20 28.0 61.0 36.0 79.0 37.0 b20 130.0 b20 78.0 b20 b20 b20 b20 b20 b20

b = below the detection limit. a Underlined values exceed the CCME (1999) Guideline for the Protection of Marine Aquatic Life (12.5 μg/L for arsenic, and 0.016 μg/L for mercury). b Fresh water discharge to intertidal sediments.

bioaccumulation factor of N 1 indicates that the arsenic in the sediment is bioavailable to the clams, and that the clams are concentrating the arsenic in their tissues. BAFs from sediment to clams for all locations were

9

N1 (range 1.7 at Seal Harbour station 2A, to 9.2 at Seal Harbour station K5), except at Wine Harbour (BAF = 0.53) and Harrigan Cove (BAF = 0.03). Velez et al. (2015) found a similar range of BAF for As for three species of clams in sediments from the Ria de Aveiro lagoon, Portugal (range of BAF 0.35 to 5.41), with 15 of 21 BAFs for As N 1.0, similar to our study (12 of 14 BAFs for As N1). Velez et al. (2015) found that clams from the most contaminated areas contained higher concentrations of contaminants than clams from the less contaminated areas, while clams from the less contaminated areas tend to have higher BAF values than clams from the most contaminated ones, findings that are in agreement with our present study, as well as that of McGeer et al. (2003). For the 2004 samples collected at the mouth of West Brook, the bioaccumulation factor for arsenic from sediment to clam tissues was calculated to be 2.6, similar to the BAF for the 2005 samples collected in the same area (BAF 2.8). In a study to assess the extent of trace metal pollution and its influence on the community structure of soft bottom molluscs in intertidal areas of the Dar es Salaam coast, Tanzania, Rumisha et al. (2012) reported biota-sediment bioaccumulation factors of 4 to 19 in the bivalve Mactra ovalina, higher than our present study. Liu et al. (2007) reported BAF from sediment that ranged from 0.35 to 0.74, for the freshwater clams (Meretrix lusoria), lower than our present study. Bioaccumulation factors (BAFs) from water to clams for all locations (Table 2) ranged from approximately 1000 to approximately 25,000, except at Harrigan Cove (BAF = 178). Liu et al. (2007) reported BAF from water that ranged from 165 to 364 for the freshwater clams (Meretrix lusoria). The reason for the low BAF from both sediment and water at Harrigan Cove is not known, but indicates low bioavailability of arsenic from both water and sediments at this site compared to the

Scatter Matrix

Hg in sediment

As in sediment

Hg in clams

As in clams

As in water

Fig. 6. Correlations between arsenic and mercury in sediment, arsenic in water, and arsenic and mercury in clam tissues. Highly significant correlations were found between arsenic in sediment and arsenic in clams (r2 = 0.824, P = 0.00099), arsenic in water and arsenic in clams (r2 = 0.952, P = 0.000006), and arsenic in sediment and arsenic in water (r2 = 0.883, P = 0.0003). Other relationships were not significantly correlated.

Please cite this article as: Doe, K., et al., Biological effects of gold mine tailings on the intertidal marine environment in Nova Scotia, Canada, Marine Pollution Bulletin (2016), http://dx.doi.org/10.1016/j.marpolbul.2016.08.056

10

K. Doe et al. / Marine Pollution Bulletin xxx (2016) xxx–xxx

other study sites. The High BAFs from water to clam tissue are not really meaningful as discussed below. Because of the higher concentrations of arsenic and mercury in softshell clam tissue than in blue mussel tissue, the better correlation of clam tissue arsenic concentrations with arsenic in water and sediment when compared with blue mussels, and because of the high bioaccumulation factors for arsenic in clams, it is logical to suggest that soft-shell clams are a better indicator species for assessing contaminants from gold mine tailings than blue mussels. This can be attributed to their mode of living as they live directly within the sediment in the intertidal area that has acted to some extent as the depositional environment for gold mine tailings at the study sites. Clams within the sediment feed by extending their siphon to the boundary of where the water meets the sediment, drawing in organic matter, detritus, and fine-grained sediment. Branched cilia strain out suspended particles as small as 2 μm in diameter excreting water and substances through their external siphon (Abraham, 1986). Consequently, contaminants found bound to small particles within the sediment and in the water are ingested and can be absorbed by the tissues of clams. Milligan and Law (2013) showed that in Seal Harbour, metal(oid)s are enriched in loosely packed aggregates of particulate material called flocs as compared to metal(oid) concentrations in the underlying sediment. These flocs allow contaminants to accumulate at the sediment-water interface and package them in a form that is readily available for ingestion by filter-feeding organisms, and form a pathway for the uptake of metals by clams. Mussels are not sediment infauna, living attached to substrates, and so do not as readily come in direct contact with sediment. Mussels are largely influenced by contaminants in the water column, feeding predominately on phytoplankton (Newell, 1989). Therefore a very high BAF for clams from water are not really meaningful since the tissue concentrations are most undoubtedly due to sediment concentrations, while the reverse is true for mussels. This is in agreement with Bryan and Langston (1992), who reported that field and laboratory studies have demonstrated that sediments are the major source of arsenic to a number of infaunal invertebrates including clams. Consequently, As concentrations in these species generally reflect sediment contamination (measured as total As in sediments). 3.2.3. Arsenic speciation in soft-shell clam and blue mussel tissue Much of the bioaccumulated arsenic in marine organisms is present in relatively nontoxic organic forms, such as arsenobetane, while inorganic arsenic species are considered more toxic than organic forms (Bryan and Langston, 1992; Cullen and Reimer, 1989; Francesconi and Kuehnelt, 2002; Phillips and Depledge, 1985, Sharma and Sohn, 2009). Therefore inorganic arsenic is a better indicator of risk to consumers of bivalve molluscs than total arsenic. Soft-shell clams and blue mussels

were analyzed for As (III) and As (V) to determine the amount of inorganic arsenic that has accumulated in the tissues, and these results are shown in Figs. 7 and 8 respectively. Clams accumulated higher concentrations of both inorganic species of arsenic than mussels, at locations where both species were found. Soft-shell clams accumulated more As (V) than As (III), while blue mussels accumulated more As (III) than As (V) at most locations. Whaley-Martin et al. (2012) also found that blue mussels accumulated more As (III) than As (V). The reason for this difference is not clear, but may be related to different uptake routes for the two species of molluscs. The highest concentration of As (III) and total arsenic found in mussel tissue from any study site was from the Gold River location, and was unexpected based on the relatively low arsenic concentration found in water and sediment samples from this location. This finding is unexplained and worth further study. For inorganic arsenic, salts with arsenic in the pentavalent state are less toxic than salts with arsenic in the trivalent state. However, studies suggested that some reduction of arsenate to arsenite might occur within the mammalian body therefore, the eventual toxicity of inorganic arsenic in either valency state should not be disregarded (Edmonds and Francesconi, 1993). Overall, inorganic arsenic accounted for a much greater percent of total arsenic in clams (mean = 17.0%, range = 6.5– 45.4%, n = 12) than in mussels (mean = 6.7%, range = 1.2–17.8%, n = 15). The higher percentages of inorganic arsenic found in this study were among the highest reported in the literature, though recently percentages of inorganic arsenic of up to 42% were reported in bivalves from Norwegian fiords (Sloth and Julshamn, 2008), and as high as 42% in blue mussels (M. edulis) from Seal Harbour, NS (WhaleyMartin et al., 2012). Those authors found significantly higher proportions of inorganic arsenic in marine samples with total arsenic concentrations exceeding 3 mg/kg wet weight than in samples with total arsenic concentrations b3 mg/kg wet weight. Koch et al. (2007) found that the bioaccessible arsenic in the Seal Harbour clam samples was essentially all inorganic arsenic (III and V), ranging from an average of 98 ± 10% for the gastric phase only (Phase 1), to an average of 86 ± 42% for the gastric plus intestinal phase (Phase 2). Greene and Crecelius (2008) found much lower percentage of inorganic arsenic with a mean percent in hard clams from the Delaware Estuary of 1.1%. Le (2002) stated that the predominant form of arsenic found in shellfish is the organic arsenic compound arsenobetaine, which is believed to have low or negligible toxicity. Arsenosugars are also present in seaweed and in molluscs (Francesconi and Kuehnelt, 2002). Le (2002) found that little work has been done on the toxicity of arsenosugars to humans but it is understood that humans may metabolize arsenosugars, and so they may be toxicologically relevant. The very high concentrations of inorganic arsenic in the clams from the Seal Harbour (Canada) study site (this study, and Koch et al., 2007),

As (ug/kg) (wet wt.)

25000 = As III = As V

20000

15000

10000

5000

0 New Hbr

L-1

WH-1

HC-1

SH-4

SH-9 SH-10 SH-8

SH-6

SH-3

SH-5

SH-1

Sampling Locations Fig. 7. As (III) and As (V) in clam tissue collected from 2005 study sites.

Please cite this article as: Doe, K., et al., Biological effects of gold mine tailings on the intertidal marine environment in Nova Scotia, Canada, Marine Pollution Bulletin (2016), http://dx.doi.org/10.1016/j.marpolbul.2016.08.056

K. Doe et al. / Marine Pollution Bulletin xxx (2016) xxx–xxx

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1800 As (ug/kg) (wet wt.)

1600

= As III = As V

1400 1200 1000 800 600 400 200 0 NH-2

L-2

PH-1

PB-1 SH-18 SH-13 SH-14 SH-17 SH-11 SH-12 WH-3 WH-2 SH-15 SH-16 GR-1

Sampling Locations Fig. 8. As (III) and As (V) in mussel tissue collected from 2005 study sites.

pose a risk to predators such as humans, sea gulls, ducks, raccoons, river otters, northern moon snails, green crabs, ribbon worms, blood worms, and sand worms (Commito, 1982; Department of Marine Resources, 2001; Jacques Whitford Limited, 2008; Newell and Hidu, 1986), and indicate that clams from this area should not be used for human consumption. The concentrations of inorganic arsenic in the clam and mussel tissues from the 2005 study (Figs. 7 and 8) were used in a sitespecific human health risk assessment by Health Canada (2006). The results of this risk assessment led to a shellfish harvesting closure at the mouth of West Brook by the Canadian Department of Fisheries and Oceans, and consumption advisories for clams and mussels applied to all other areas in Seal Harbour as a precaution because of potential risks to human health (Environment Canada, 2007). 3.2.4. Ecological risk assessment An Ecological Risk Assessment for Wine and Seal Harbours, NS, using data generated during the present study indicated that there is a potential for adverse chronic sublethal effects of arsenic on molluscivores feeding at these sites (Jacques Whitford Limited, 2008). This is in agreement with a study by Little et al. (2015) who found elevated arsenic and mercury burdens in the tailings and intertidal zone samples at Wine Harbour and suggested that these high concentrations could present a risk to human and ecosystem health. 3.2.5. QA/QC checks on arsenic in clam tissue sample results from two different laboratories The results of analyses for arsenic in thoroughly homogenized and split samples of tissue (n = 4) and sediment (n = 7) are presented in Tables 5 and 6, respectively. There is good overall agreement in the results reported by the two independent laboratories in each table. This indicates that the data from 2004 are comparable with the data from Table 5 QA/QC checks on total arsenic in clam tissue samples (mg/kg wet weight) (n = 4) from two different laboratories.

Site name Harrigan Cove Seal Harbour #1 New Harbour reference Wine Harbour — 2

Arsenic total, EC Moncton Laboratory

Arsenic total, CFIA Dartmouth Laboratory

3.81 226 0.99

1.85 259 1.62

4.19

6.79

2005, and that the high concentrations reported in this study are repeatable. 4. Conclusions The results from this study identified Seal Harbour, Wine Harbour and Harrigan Cove as areas highly contaminated by arsenic and mercury from gold mine tailings. Concentrations of arsenic and mercury in intertidal sediments and water from the study sites were compared with various environmental quality guidelines (CCME, 1999; Martın-Dıaz et al., 2008) and indicated a high risk of adverse biological effects in the intertidal environments of Seal Harbour, Wine Harbour and Harrigan Cove. Arsenic in intertidal sediments and water at several stations in Seal Harbour is bioavailable and is present in higher concentrations in the soft-shell clam tissues than in surrounding water and sediments. High concentrations of arsenic in soft-shell clam (M. arenaria) tissue samples from Seal Harbour were found to be higher than tissue concentrations previously reported in the scientific literature for molluscs. The results from this study on arsenic in clams were used by Health Canada and the Department of Fisheries and Oceans to close the area at the mouth of West Brook to shellfish harvesting. Consumption advisories for clams and mussels were applied to all other areas in Seal Harbour. This study generated useful insights that could be applied to other areas potentially contaminated by historic gold mining practices. The results have also raised many questions on the cycling and fate of arsenic and mercury through marine ecosystems. For example, further research should explore the reason why clams accumulate more As (V) while mussels accumulate more As (III), and why bioavailability of arsenic was lower at Harrigan Cove than at Seal Harbour. Table 6 QA/QC checks on arsenic in sediment samples (mg/kg dry weight) (n = 7) from two different laboratories.

Site name Seal Harbour #3 Seal Harbour K #1 Seal Harbour K #2 Seal Harbour K #3 Seal Harbour K #4 Seal Harbour K #5 New Harbour Reference

Arsenic, EC Moncton Laboratory

Arsenic, Maxxam Laboratory

11.4 56.5 16.3 20.4 18.2 11.1 1.1

12.0 74 19.0 21.0 18.0 10.0 b2.0

Please cite this article as: Doe, K., et al., Biological effects of gold mine tailings on the intertidal marine environment in Nova Scotia, Canada, Marine Pollution Bulletin (2016), http://dx.doi.org/10.1016/j.marpolbul.2016.08.056

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In order to fully understand the impacts of mine tailings on the intertidal area, a quantitative analysis of arsenic and mercury accumulation at all trophic levels in the food chain needs to be completed. From our studies, it is apparent that intertidal organisms from Seal Harbour, Wine Harbour and Harrigan Cove are living within highly contaminated environments and that arsenic and mercury, especially arsenic, is bioavailable to molluscs. For Seal Harbour, arsenic contamination appears to represent more of an environmental risk than mercury contamination. Whether or not the contaminants reduce the fitness, survival, or adversely affect population levels of these organisms is not presently known and warrants further investigation. Acknowledgements This paper is dedicated to the memory of André Gauthier of Environment Canada, Dartmouth, Nova Scotia who initiated our participation in the study of abandoned gold mine tailings in Nova Scotia. The authors would like to acknowledge the helpful assistance and advice provided by Dr. Michael B. Parsons, Research Scientist, Natural Resources Canada, Dartmouth, Nova Scotia who coordinated the study of abandoned gold mine tailings in Nova Scotia and assisted us in so many ways. Scott Lewis, Environment Canada, Vancouver, British Columbia, created the maps of the study sites. Jeff Corkum, Chris Roberts, Paul Klaamas, Don Walter and Eric Hundert of Environment Canada, Dartmouth, Nova Scotia, assisted in project planning. Finally, the authors thank the anonymous reviewers who provided valuable comments on our manuscript. We feel the paper has benefited from their input. References Abraham, B.J., 1986. Softshell clam. Species Profiles: Life Histories and Environmental Requirements of Coastal Fishes and Invertebrates (Mid-Atlantic)Biological Report Vol. 82(11.68). US Fish and Wildlife Service. Agency for Toxic Substances and Disease Registry, 2007. Toxicological Profile for Arsenic. U.S. Department of Health and Human Services. Public Health Service Agency for Toxic Substances and Disease Registry, 1600 Clifton Road, NE, Mailstop F-32, Atlanta, Georgia, 30333 559 pp. Alpers, C.N., Hunerlach, M.P., 2000. Mercury contamination from historic gold mining in California. United States Geological Survey, Fact Sheet Fs-061-00 Retrieved 16 March, 2016 from http://pubs.usgs.gov/fs/2005/3014/fs2005_3014_v1.1.pdf. Bates, J.L.E., 1987. Gold in Nova Scotia. Nova Scotia Department of Natural Resources, Mineral Resources Branch, Information Series ME 13 Retrieved on March 22, 2016. http:// novascotia.ca/natr/meb/pdf/is13.asp. Beiras, R., Bellas, J., Fernandez, N., Lorenzo, J.I., Cobelo-Garcia, N., 2003. Assessment of coastal marine pollution in Galicia (NW Iberian Peninsula); metal concentrations in seawater, sediments and mussels (Mytilus galloprovincialis) versus embryo-larval bioassays using Paracentrotus lividus and Ciona intestinalis. Mar. Environ. Res. 56 (4), 531–553. Besada, V., Sericano, J.L., Schultze, F., 2014. An assessment of two decades of trace metals monitoring in wild mussels from the Northwest Atlantic and Cantabrian coastal areas of Spain, 1991–2011. Environ. Int. 71, 1–12. Bryan, G.W., Langston, W.J., 1992. Bioavailability, accumulation and effects of heavy metals in sediments with special reference to United Kingdom estuaries: a review. Environ. Pollut. 76, 89–131. Cattani, O., Fabbri, D., Salvati, M., Trombini, C., Claudio, I., 1999. Biomonitoring of mercury pollution in a wetland near Ravenna, Italy by translocated bivalves (Mytilus galloprovincialis). Environ. Toxicol. Chem. 18 (8), 1801–1805. CCME, 1999. Canadian Environmental Quality Guidelines. Canadian Council of Ministers of the Environment, Winnipeg, Canada Available at: http://www.ccme.ca/en/about/ index.html Accessed 10 March, 2016. Commito, J.A., 1982. Effects of Lunatia heros predation on the population dynamics of Mya arenaria and Macoma baithica in Maine, USA. Mar. Biol. 69, 187–193. Cullen, W.R., Reimer, K.J., 1989. Arsenic speciation in the environment. Chem. Rev. 89 (4), 713–764. Davies, I.M., 1981. Survey of Trace Elements in Fish and Shellfish Landed at Scottish Ports 1975–1976. Marine Laboratory Aberdeen, Scotland 28 pp. Accessed 16 March, 2016 from http://www.gov.scot/Uploads/Documents/No%2019.pdf. Department of Marine Resources, 2001. Soft-shell clams (Mya arenaria). Prepared by the Gulf of Maine Aquarium. January 10, 2001 http://www.maine.gov/dmr/research/ soft_shell_clams.htm#predators Accessed 3 March, 2016. Dixon, D.R., Wilson, J.T., 2000. Genetics and marine pollution. Hydrobiologia 420 (1–3), 29–43. Edmonds, J.S., Francesconi, K.A., 1993. Arsenic in seafoods: human health aspects and regulations. Mar. Pollut. Bull. 27 (12), 665–674. Eisler, R., 2004. Biogeochemical, Health, and Ecotoxicological Perspectives on Gold and Gold Mining. CRC Press, Boca Raton, Florida 355 pp.

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Please cite this article as: Doe, K., et al., Biological effects of gold mine tailings on the intertidal marine environment in Nova Scotia, Canada, Marine Pollution Bulletin (2016), http://dx.doi.org/10.1016/j.marpolbul.2016.08.056