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Funding: Funding for this work was provided by a NERC GW4+ Doctoral Training Partnership studentship (NE/S007504/1) awarded to J.K., an Antarctic Bursary awarded to J.A.S, and NERC grants awarded to L.F.R. (NE/S001743/1; NE/R005117/1; NE/N003861/1). Cruise DY081 was funded by European Research Council starting grant ICY-LAB (Grant Agreement 678371). The concentration of dissolved barium in seawater ([Ba]SW) is influenced by both primary productivity and ocean circulation patterns. Reconstructing past subsurface [Ba]SW can therefore provide important information on processes which regulate global climate. Previous Ba/Ca measurements of scleractinian and bamboo deep-sea coral skeletons exhibit linear relationships with [Ba]SW, acting as archives for past Ba cycling. However, skeletal Ba/Ca ratios of the Stylasteridae – a group of widely distributed, azooxanthellate, hydrozoan coral – have not been previously studied. Here, we present Ba/Ca ratios of modern stylasterid (aragonitic, calcitic and mixed mineralogy) and azooxanthellate scleractinian skeletons, paired with published proximal hydrographic data. We find that [Ba]SW and sample mineralogy are the primary controls on stylasterid Ba/Ca, while seawater temperature exerts a weak secondary control. [Ba]SW also exerts a strong control on azooxanthellate scleractinian Ba/Ca. However, Ba-incorporation into scleractinian skeletons varies between locations and across depth gradients, and we find a more sensitive relationship between scleractinian Ba/Ca and [Ba]SW than previously reported. Paired Sr/Ca measurements suggest that this variability in scleractinian Ba/Ca may result from the influence of varying degrees of Rayleigh fractionation during calcification. We find that these processes exert a smaller influence on Ba-incorporation into stylasterid coral skeletons, a result consistent with other aspects of their skeletal geochemistry. Stylasterid Ba/Ca ratios are therefore a powerful, novel archive of past changes in [Ba]SW, particularly when measured in combination with temperature sensitive tracers such as Li/Mg or Sr/Ca. Indeed, with robust [Ba]SW and temperature proxies now established, stylasterids have the potential to be an important new archive for palaeoceanographic studies. Publisher PDF

Coral growth anomalies (GAs) are tumor-like protrusions that are detrimental to coral health, affecting both the coral skeleton and soft tissues. These lesions are increasingly found throughout the tropics and are commonly associated with high human population density, yet little is known about the molecular pathology of the disease. Here, we investigate the metabolic impacts of GAs through 1H nuclear magnetic resonance (NMR) metabolomics in Porites compressa tissues from a site of high disease prevalence (Coconut Island, Hawaii). We putatively identified 18 metabolites (8.1% of total annotated features) through complementary 1H and 1H–13C heteronuclear single quantum correlation NMR data that increase confidence in pathway analyses and may bolster future coral metabolite annotation efforts. Extract yield was elevated in both GA and unaffected (normal tissue from a diseased colony) compared to reference (normal tissue from GA-free colony) samples, potentially indicating elevated metabolic activity in GA-impacted colonies. Relatively high variation in metabolomic profiles among coral samples of the same treatment (i.e., inter-colony variation) confounded data interpretation, however, analyses of paired GA and unaffected samples identified 73 features that differed between these respective metabolome types. These features were largely annotated as unknowns, but 1-methylnicotinamide and trigonelline were found to be elevated in GA samples, while betaine, glycine, and histamine were lower in GA samples. Pathway analyses indicate decreased choline oxidation in GA samples, making this a pathway of interest for future targeted studies. Collectively, our results provide unique insights into GA pathophysiology by showing these lesions alter both the absolute and relative metabolism of affected colonies and by identifying features (metabolites and unknowns) and metabolic pathways of interest in GA pathophysiology going forward.

Chronically debilitated loggerhead sea turtles (Caretta caretta) (DT) are characterized by emaciation, lethargy, and heavy barnacle coverage. Although histopathological findings associated with this condition have been reported, only limited data is available on health variables with clinical application. The objectives of this study were to 1) to compare morphometrics, clinicopathological variables, and immune functions of DTs to a group of apparently healthy loggerhead turtles to better understand the pathophysiology of the condition and 2) to assess health parameters in live debilitated turtles as they recovered during rehabilitation in order to identify potential prognostic indicators. We examined and sampled 43 DTs stranded from North Carolina to Florida for 47 health variables using standardized protocols to further characterize the condition. DTs were grouped into categories of severity of the condition, and those that survived were sampled at four time points through rehabilitation. All groups and time points were compared among DTs and to clinically healthy loggerhead turtles. Compared to healthy turtles, DTs had significantly lower body condition index, packed cell volume (PCV), total white blood cell (WBC) count, lymphocytes, glucose (Glc), total protein, all protein fractions as determined by electrophoresis, calcium (Ca), phosphorus (P), Ca:P ratio, potassium (K), lymphocyte proliferation, and greater heterophil toxicity and left-shifting, uric acid (UA), aspartate aminotransferase, creatine kinase, lysozyme, and respiratory burst. From admission to recovery, hematology and plasma chemistry data improved as expected. The most informative prognostic indicators, as determined by correlations with a novel severity indicator (based on survival times), were plastron concavity, P, albumin, total solids, UA, lymphocyte proliferation, WBC, K, Glc, Ca:P, and PCV. The results of this study document the wide range and extent of morphometric and metabolic derangements in chronically debilitated turtles. Monitoring morphometrics and clinicopathological variables of these animals is essential for diagnosis, treatment, and prognosis during rehabilitation.

Mercury has been identified as one of the most serious environmental threats to the well-being of wildlife in the southeastern United States (Facemire et al. 1995). The prevalence of Hg in aquatic species has also prompted concerns for the health of subsistence fishermen and the general population who regularly consume fish. This concern is evident in the fact that 76% of fish consumption advisories in the United States in 2003 were due at least in part to Hg, for a total of 5,289,020 hectares of lakes and 1,234,127 river miles [U.S. Environmental Protection Agency (EPA) 2004]. The toxic effects of Hg have been demonstrated in mammals, birds, and fish and include neurotoxicity, impaired growth and development, reduced reproductive success, liver and kidney damage, and immunomodulation (Wiener et al. 2003; Zelikoff et al. 1994). Reptiles are prominent members of ecosystems and often have life history characteristics that make them vulnerable to Hg accumulation (e.g., long life span, high trophic level, aquatic habitat). Despite the fact that many of these reptilian species also have a tenuous conservation status, there are currently few data on the toxicity of Hg in this taxon. One such example is the sea turtle, of which all species are classified as either threatened or endangered. Many anthropogenic factors have been implicated in the decline in sea turtle populations, including directed harvest for food and trade, fisheries bycatch, and degradation of nesting beach habitat (Lutcavage et al. 1997). However, the role of chemical pollutants in marine turtle health is largely unknown. It is therefore important to understand the risk that contaminants pose to the general health and immunologic function of sea turtles because these effects could also impact the survival of their populations. Several studies have measured Hg levels in tissues from juvenile and adult sea turtles (Anan et al. 2001; Davenport and Wrench 1990; Day et al. 2005; Godley et al. 1999; Gordon et al. 1998; Maffucci et al. 2005; Orvik 1997; Presti 1999; Sakai et al. 1995, 2000a, 2000b; Storelli et al. 1998, 2005; Wang 2005). However, relatively few studies have assessed health parameters in sea turtles in relation to environmental contaminants (Keller et al. 2004, 2005, 2006a, 2006b; Lutcavage et al. 1995; Peden-Adams et al. 2002, 2003; Podreka et al. 1998), and these studies have focused primarily on organic contaminants rather than metals. Balazs and Pooley (1991) suggested that environmental contaminants are a possible factor contributing to the development of the viral disease fibropapillomatosis in sea turtles by reducing immune function. Some locations in Florida (Indian River Lagoon and Florida Bay) exhibit up to 70% prevalence of this disease, and these environments also have elevated levels of Hg (Ache et al. 2000; Cantillo et al. 1999; Trocine and Trefry 1996). In the present study we investigated the relationship of blood Hg levels in loggerhead sea turtles (Caretta caretta) off the coasts of South Carolina, Georgia, and northern Florida to various immunologic and general health parameters. Using an integrated approach, correlative field data collection from wild-caught loggerhead sea turtles were combined with laboratory in vitro exposure of loggerhead lymphocytes to methylmercury (MeHg).

Sixty common murre (Uria aalge) and 27 thick-billed murre (Uria lomvia) eggs collected by the Seabird Tissue Archival and Monitoring Project (STAMP) in 1999-2001 from two Gulf of Alaska and three Bering Sea nesting colonies were analyzed for total mercury (Hg) using isotope dilution cold vapor inductively coupled mass spectrometry. Hg concentrations (wet mass) ranged from 0.011 microg/g to 0.357 microg/g (relative standard deviation = 76%), while conspecifics from the same colonies and years had an average relative standard deviation of 33%. Hg levels in eggs from the Gulf of Alaska (0.166 microg/g +/- 0.011 microg/g) were significantly higher (p0.0001) than in the Bering Sea (0.047 microg/g +/- 0.004 microg/g). Within the Bering Sea, Hg was significantly higher (p = 0.0007) in eggs from Little Diomede Island near the arctic than at the two more southern colonies. Although thick-billed and common murres are ecologicallysimilar,there were significant species differences in egg Hg concentrations within each region (p0.0001). In the Bering Sea, eggs from thick-billed murres had higher Hg concentrations than eggs from common murres, while in the Gulf of Alaska, common murre eggs had higher concentrations than those of thick-billed murres. A separate one-way analysis of variance on the only time-trend data currently available for a colony (St. Lazaria Island in the Gulf of Alaska) found significantly lower Hg concentrations in common murre eggs collected in 2001 compared to 1999 (p = 0.017). Results from this study indicate that murre eggs may be effective monitoring units for detecting geographic, species, and temporal patterns of Hg contamination in marine food webs. The relatively small intracolony variation in egg Hg levels and the ability to consistently obtain adequate sample sizes both within and among colonies over a large geographic range means that monitoring efforts using murre eggs will have suitable statistical power for detecting environmental patterns of Hg contamination. The potential influences of trophic effects, physical transport patterns, and biogeochemical processes on these monitoring efforts are discussed, and future plans to investigate the sources of the observed variability are presented.

The validity of using blood samples and keratinized scutes for nonlethal routine monitoring of mercury (Hg) in loggerhead sea turtles, Caretta caretta, is evaluated in the context of how effectively these matrixes predict internal tissue Hg burdens and the different temporal scales of exposure they represent. Total Hg (THg) was measured in blood and scutes collected from live captures (n = 34) and liver, kidney, muscle, spinal cord, blood, and scutes collected from freshly stranded loggerhead turtles (n = 6) along the coast of the southeastern United States. Linear regressions between monitoring compartments and internal tissues from stranded animals were all statistically significant (r20.805, p0.015) but varied in their utility as a predictive tool depending on which tissues were paired. Blood was an effective predictor of THg in muscle (r2 = 0.988, p0.0001) and spinal cord (r2 = 0.988, p0.0001), while scute was the most accurate predictor of THg in liver (r2 = 0.948, p = 0.0010). The strength of the relationship between tissues types is believed to reflect the similarity in the temporal scales they represent and the variability in the fraction of methylmercury present. The stability of Hg in the scute matrix makes this tissue preferable for approximating long-term exposure, while blood Hg levels can be affected by recent changes in Hg intake. THg levels in blood and scutes from live captures were highly correlated (linear regression r2 = 0.926, p0.0001) and increased significantly with body mass (r2 = 0.173, p = 0.016 and r2 = 0.187, p = 0.012 respectively), further supporting thatthere is a component reflecting long-term accumulation of Hg in these matrixes. We also present a novel technique using the residuals from the blood-scute regression as an index of recent exposure (IRE). The interpretation of this value is derived from the comparison between the most recent Hg intake (which contributes to the Hg measured in the blood) relative to the average past intake (which is recorded in the scute). A stepwise multiple regression revealed a significant positive relationship between the IRE and the proximity of the capture site to the nearest major industrial river mouth (p = 0.0102). This suggests that there is an elevation of bioavailable Hg in nearshore habitats where terrestrial influences and anthropogenic impacts are high. Seasonal foraging site fidelity and the variability in environmental Hg may explain the high intraspecific variability and occasional highly contaminated turtle seen in this and previous studies.

cited By 7; International audience; Environmental specimen banks (ESBs) have been a fundamental tool for many nations to monitor contaminant temporal and spatial trends, study fate and transport, and assess the severity and risks of pollution. The specimens archived in ESBs are among the longest time-series, most geographically robust, and highest integrity samples available for performing environmental research. Mercury (Hg) remains one of the world's most ubiquitous environmental contaminants, and ESBs have played a prominent role in Hg research. Historically this has involved measuring concentrations of Hg species in various environmental matrices, but the emerging field of Hg stable isotope research provides a new analytical approach that can augment these traditional techniques. Signatures of Hg isotope fractionation have been effectively used for source apportionment and for elucidating Hg biogeochemical cycling. As the research surrounding Hg stable isotopes continues to mature, ESBs can play a useful role in analytical quality control, provide a robust and economical sample archive to expand and diversify the inventory of Hg isotope measurements, and be used to develop and test hypotheses to evaluate whether broadly prevailing paradigms are supported. Samples archived in ESBs are available for request by external collaborators in order to perform high impact research, and should be utilized more effectively to address emerging global environmental concerns. © 2014 The Royal Society of Chemistry.

cited By 17; International audience; Elevated mercury concentrations ([Hg]) were found in Alaskan murre (Uria spp.) eggs from the coastal embayment of Norton Sound relative to insular colonies in the northern Bering Sea-Bering Strait region. Stable isotopes of Hg, carbon, and nitrogen were measured in the eggs to investigate the source of this enrichment. Lower δ13C values in Norton Sound eggs (-23.3‰ to -20.0‰) relative to eggs from more oceanic colonies (-20.9‰ to -18.7‰) indicated that a significant terrestrial carbon source was associated with the elevated [Hg] in Norton Sound, implicating the Yukon River and smaller Seward Peninsula watersheds as the likely Hg source. The increasing [Hg] gradient extending inshore was accompanied by strong decreasing gradients of δ202Hg and Δ199Hg in eggs, indicating lower degrees of mass-dependent (MDF) and mass-independent Hg fractionation (MIF) (respectively) in the Norton Sound food web. Negative or zero MDF and MIF signatures are typical of geological Hg sources, which suggests murres in Norton Sound integrated Hg from a more recent geological origin that has experienced a relatively limited extent of aquatic fractionation relative to more oceanic colonies. The association of low δ202Hg and Δ199Hg with elevated [Hg] and terrestrial δ13C values suggested that Hg stable isotopes in murre eggs effectively differentiated terrestrial/geogenic Hg sources from oceanic reservoirs. © 2012 American Chemical Society.

A suite of three green tea-containing Standard Reference Materials (SRMs) has been issued by the National Institute of Standards and Technology (NIST): SRM 3254 Camellia sinensis (Green Tea) Leaves, SRM 3255 Camellia sinensis (Green Tea) Extract, and SRM 3256 Green Tea-Containing Solid Oral Dosage Form. The materials are characterized for catechins, xanthine alkaloids, theanine, and toxic elements. As many as five methods were used in assigning certified and reference values to the constituents, with measurements carried out at NIST and at collaborating laboratories. The materials are intended for use in the development and validation of new analytical methods, and for use as control materials as a component in the support of claims of metrological traceability.

The contents from thick-billed murre (Uria lomvia) eggs collected at four Alaskan colonies in 2002 were analyzed for organic contaminants and carbon δ¹³C) and nitrogen (δ¹⁵N) stable isotopes. Contaminant concentrations in the eggs varied from below detection limits to 230 ng g⁻¹ wet mass for 4,4'-DDE in one egg from St Lazaria Island in the Gulf of Alaska. Eggs from this colony generally contained higher levels of contaminants and exhibited significantly different patterns compared to eggs from the Bering and Chukchi seas. Stable isotope values also varied geographically; however, these differences appeared to be related to differences in C and N baselines in the food webs instead of differences in prey. Contaminant and stable isotope correlations were inconclusive, suggesting that better information on regional food web differences and differential offloading of contaminants and stable isotopes to the eggs must be obtained before these kinds of data can be fully incorporated into seabird egg contaminant monitoring programs.

Atmospheric deposition of mercury to remote areas has increased threefold since pre-industrial times. Mercury deposition is particularly pronounced in the Arctic. Following deposition to surface oceans and sea ice, mercury can be converted into methylmercury, a biologically accessible form of the toxin, which biomagnifies along the marine food chain. Mass-independent fractionation of mercury isotopes accompanies the photochemical breakdown of methylmercury to less bioavailable forms in surface waters. Here we examine the isotopic composition of mercury in seabird eggs collected from colonies in the North Pacific Ocean, the Bering Sea and the western Arctic Ocean, to determine geographical variations in methylmercury breakdown at northern latitudes. We find evidence for mass-independent fractionation of mercury isotopes. The degree of mass-independent fractionation declines with latitude. Foraging behaviour and geographic variations in mercury sources and solar radiation fluxes were unable to explain the latitudinal gradient. However, mass-independent fractionation was negatively correlated with sea-ice cover. We conclude that sea-ice cover impedes the photochemical breakdown of methylmercury in surface waters, and suggest that further loss of Arctic sea ice this century will accelerate sunlight-induced breakdown of methylmercury in northern surface waters. Atmospheric deposition of mercury to remote areas has increased threefold since pre-industrial times. Reductions in sea-ice cover accelerate the photodegradation of biologically accessible mercury in Arctic waters, according to an analysis of the isotopic composition of bird eggs in northern latitudes.

An increase in the incidence of debilitated loggerhead sea turtle (Caretta caretta) strandings in the southeastern United States has been observed in recent years. These turtles are characterized by emaciation and heavy burdens of external and internal parasites, and bacterial infections, but the underlying cause of their condition is unknown. To investigate further the causes of these strandings, a health assessment was performed on stranded, debilitated loggerhead turtles, and contaminant concentrations in various tissues were compared to those from healthy turtles. This portion of the study investigated the potential role of mercury (Hg) toxicity in the debilitated condition described above. Hematocrit, total protein, albumin, globulin, glucose, calcium, lymphocyte counts, heterophil:lymphocyte ratios, aspartate aminotransferase, uric acid, sodium, and chloride were altered in debilitated loggerheads relative to healthy animals. However, none of the aforementioned health indicators correlated with Hg concentrations in either red blood cells (RBCs) or plasma. The Hg concentration in RBCs was 129+/-72 (mean+/-standard deviation) times higher than in plasma, causing a significant dilution of Hg in whole blood due to extreme anemia. Mercury concentrations in RBCs (73.7+/-21.2 ng/g) and scutes (455+/-57 ng/g) from debilitated turtles were similar to or lower than those reported for healthy animals, indicating no elevation in Hg exposure before and during the progression of this condition. These findings suggest that Hg toxicity does not play a role in the debilitated loggerhead condition observed in the southeastern United States.

Total mercury concentrations were measured in diamondback terrapin blood and scutes collected from four sites in South Carolina, USA, and at a superfund site in Brunswick, Georgia, USA. There was a strong correlation between mercury concentrations in the two terrapin body compartments (Kendall's tau = 0.79, p < 0.001). Mercury concentrations in terrapin scute and blood and in salt marsh periwinkles, Littoraria irrorata, were significantly higher in Brunswick (scute x = 3810.2 ng/g, blood x = 746.2 ng/g) than from all other sites (scute x = 309.5 ng/g, blood x = 43.2 ng/g, p < 0.001). Seasonal fluctuations of total mercury in the blood and scutes of terrapins collected in the Ashley River, South Carolina, were significantly lower in August than in April, June, or October in blood (p < 0.001); however, scute concentrations did not vary seasonally. Overall, we found higher concentrations of mercury in the scutes of females than males (n = 32, p < 0.05). Larger females may preferentially prey on larger food items, like large periwinkles, which had significantly higher mercury levels in their body tissues than smaller periwinkles (p < 0.001). Methylmercury levels in terrapin scutes were measured, revealing that 90% of the total mercury stored in this compartment was in the organic form. A methylmercury biomagnification factor of 173.5 was calculated from snails to terrapin scutes, and we found that mercury levels in scutes were representative of the mercury levels in other compartments of the ecosystem. These findings show that terrapin scutes are good predictors of mercury pollution and that this species could be used as a bioindicator for assessing mercury contamination of estuarine systems.

The contents from thick-billed murre (Uria lomvia) eggs collected at four Alaskan colonies in 2002 were analyzed for organic contaminants and carbon (δ13C) and nitrogen (δ15N) stable isotopes. Contaminant concentrations in the eggs varied from below detection limits to 230 ng g−1 wet mass for 4,4′-DDE in one egg from St Lazaria Island in the Gulf of Alaska. Eggs from this colony generally contained higher levels of contaminants and exhibited significantly different patterns compared to eggs from the Bering and Chukchi seas. Stable isotope values also varied geographically; however, these differences appeared to be related to differences in C and N baselines in the food webs instead of differences in prey. Contaminant and stable isotope correlations were inconclusive, suggesting that better information on regional food web differences and differential offloading of contaminants and stable isotopes to the eggs must be obtained before these kinds of data can be fully incorporated into seabird egg contaminant monitoring programs.

A highly sensitive and selective method has been developed for the determination of methylmercury in biological specimens and NIST Standard Reference Materials (SRMs). The procedure involves microwave extraction with acetic acid, followed by derivatization and headspace solid-phase microextraction (SPME) with a polydimethylsiloxane (PDMS)-coated silica fiber. Optimization of conditions including gas chromatograph injection temperature, microwave extraction power and microwave extraction time are presented. The identification and quantification (via the method of standard additions) of the extracted compounds is carried out by capillary gas chromatography with inductively coupled plasma mass spectrometric detection (GC-ICP-MS) using a unique heated interface that was designed for this work. The SPME-GC-ICP-MS method was validated for the determination of methylmercury (MeHg) concentrations in a variety of biological Standard Reference Materials (SRMs), ranging from 13.2 ng g−1 in SRM 1566b Oyster Tissue, to 397 ng g−1 in SRM 1946 Lake Superior Fish Tissue. Additionally, this method was applied to the determination of MeHg in seabird eggs (common murres, Uria aalge and thick-billed murres, Uria lomvia) collected from colonies on Little Diomede and Bogoslof islands in the Bering Sea and Saint Lazaria Island in the Gulf of Alaska and cryogenically banked in the Marine Environmental Specimen Bank. The results obtained demonstrate that SPME-GC-ICP-MS is a sensitive technique for the determination of methylmercury at trace and ultra-trace levels in a variety of natural matrices with high reproducibility and accuracy. In all instances, the sample-to-sample variability was typically 2% relative standard deviation (RSD) and the method detection limit for methylmercury was 4.2 pg g−1 (as Hg), based on a 0.5 g tissue sample of SRM 1566b Oyster Tissue.