Metal concentrations in tissues of American lobsters (Homarus americanus, Milne-Edwards) with epizootic shell disease.
Article Type: Report
Subject: Anaerobic infections (Risk factors)
Micropollutants (Distribution)
American lobster (Diseases)
Authors: Leblanc, Lawrence A.
Prince, Deanna
Pub Date: 06/01/2012
Publication: Name: Journal of Shellfish Research Publisher: National Shellfisheries Association, Inc. Audience: Academic Format: Magazine/Journal Subject: Biological sciences; Zoology and wildlife conservation Copyright: COPYRIGHT 2012 National Shellfisheries Association, Inc. ISSN: 0730-8000
Issue: Date: June, 2012 Source Volume: 31 Source Issue: 2
Topic: Event Code: 690 Goods & services distribution Advertising Code: 59 Channels of Distribution Computer Subject: Company distribution practices
Geographic: Geographic Scope: United States Geographic Code: 1USA United States
Accession Number: 302109304
Full Text: ABSTRACT Environmental contaminants have been suggested frequently as causative factors in crustacean shell disease. This study explored the relationship of environmental contaminants to epizootic shell disease in the American lobster (Homarus americanus). Trace metal loads were analyzed in lobsters showing signs of shell disease and those without signs of shell disease collected from Narragansett Bay, Rhode Island Sound, and Long Island Sound in 2007. In addition, trace metals were analyzed in lobsters collected in 2008 from a single station in Narragansett Bay and a reference site in Maine for a multiparameter, multiinvestigator study (the "100 Lobsters" project). Analyses focused primarily on hepatopancreas and hemolymph tissues, but muscle, shell, gills, and ovaries were also analyzed to examine the partitioning of metals between various component tissues. Sediment from the "100 Lobsters" collection site was also analyzed. Inductively coupled plasma mass spectrometry (ICP-MS) was used to measure 10 different contaminant elements, including arsenic, cadmium, cobalt, chromium, copper, manganese, molybdenum, nickel, lead, and vanadium. Mercury was determined using a direct mercury analyzer. Overall, there was no systematic difference seen in contaminant metal burdens between those lobsters with signs of shell disease and those without signs, and hepatopancreas concentrations were generally similar to other reported values for field-collected lobsters. Contaminant metals were found to be partitioned between tissues differentially. Hepatopancreas tissue generally contained the greatest concentrations of contaminants, but exoskeleton contained the highest levels of manganese, nickel, and lead. Sediment collected from the "100 Lobster" sampling site showed that concentrations of several metals were similar to or exceeded the suggested sediment quality guidelines, consistent with a site that is moderately impacted by metals. Based on these data, the presence or absence of shell disease cannot be attributed to the magnitude or patterns of metal accumulation and disposition.

KEY WORDS: American lobster, Homarus americanus, trace metals, epizootic shell disease, ESD, lobster, body burden, hepatopancreas

INTRODUCTION

Epizootic lobster shell disease (ESD), characterized by discoloration and degradation of the exoskeleton and, in severe cases, deep lesions in the carapace, is a potent threat to the American lobster, Homarus americanus (Milne Edwards), industry throughout New England. Severe outbreaks of this disease have been reported in southern New England (SNE) and eastern Long Island Sound (LIS) (Cobb & Castro 2006). Studies by Vassiliev (2005) and Vassiliev et al. (2005), and other anecdotal information indicate the presence of ESD at a low level of occurrence in Maine, particularly along the southern coast. Identifying causative agents is essential to designing strategies to prevent the spread of this disease, which is critical to the future of the resource.

A number of studies have pointed to environmental stressors (including contaminants, elevated temperatures, hypoxia, high sulfide and ammonia) as being at least partly responsible for the onset of ESD in lobster populations as well as the mass mortality of lobsters observed in LIS (Laufer et al. 2005, Robohm et al. 2005, Valente & Cuomo 2005). Duboise & Moulton (2005) demonstrated that the bacterial infection alone was not sufficient to explain the spread of the disease, and concluded that there may be complex interactions between the environment, genetics, and microbes causing the disease outbreak.

Metals introduced into aqueous environments may contribute to physiological stress in lobsters and other organisms. Trace metals are known to be toxic to aquatic organisms and can affect a number of organ systems. In lower doses, metals are known to affect immune system functioning and endocrine functions, as well as impart oxidative stress (Correa et al. 2005, Valko et al. 2005). In crustaceans, metals have been shown to disrupt molting, inhibit wound repair and limb regeneration (Weis et al. 1992), and act as immunosuppressants (Henroth et al. 2004, Oweson et al. 2006). Because host susceptibility appears to be an important component of shell disease (Tlusty et al. 2007), one can hypothesize that lobsters with compromised wound repair mechanisms or immune systems would be more susceptible to shell disease.

Evidence of a possible link between contaminant metals and shell disease has been provided in several studies. In a comparison of diseased and nondiseased blue crabs, Callinectes sapidus (Rathbun) from the Pamlico-Albemarle estuaries, Weinstein et al. (1992) reported higher concentrations of some trace metals (aluminum (Al), chromium (Cr), iron (Fe), manganese (Mn), titanium (Ti), uranium (U), and vanadium (V)) in diseased crabs. Differences in the concentration of trace metals in the hepatopancreas of H. americanus were found by Vassiliev (2005) and Vassiliev et al. (2005) between shell-diseased lobsters collected off the Maine coast and Nova Scotia, and nondiseased lobsters from Maine. Initial results showed greater concentrations of mercury (Hg), zinc (Zn), and cadmium (Cd) in the hepatopancreas of shell-diseased lobsters when compared with concentrations in the hepatopancreas from nondiseased lobsters. However, in later results, only Hg was found to be higher in the hepatopancreas of shell-diseased lobsters (Vassiliev 2005).

In 2007 to 2008, as part of a multidisciplinary study to examine shell disease in the American lobster, contaminant metals concentrations in hepatopancreas and hemolymph were surveyed in lobsters with and without signs of the disease. Lobsters from eastern LIS; western LIS; Narragansett Bay, RI; and the coast of Maine were included in this survey. In addition, contaminant metal concentrations were determined for a more focused investigation of a group of lobsters from a single sampling from Narragansett Bay, RI. This group, nicknamed the "100 Lobsters" project (Shields et al. 2012), compared numerous chemical and physiochemical measurements on these lobsters with a reference site off the Maine coast. This article reports the results of multielemental analyses performed on both survey lobsters and "100 Lobster" lobsters. In addition, compartmentalization of elements was examined on 8 lobsters by determining relative metal load in shell, hepatopancreas, muscle tissue, gills, and hemolymph.

MATERIALS AND METHODS

Sample Collection

Lobsters were obtained from 3 sources: Narragansett Bay transects (n = 70, plus 4 large lobsters from Rhode Island Sound), LIS (n = 26), and from the "100 Lobsters" project (n = 65, plus 15 reference samples) collected from 1 location (41.5073[degrees]N and 71.3463[degrees]W; depth, 35 m) in the East Passage of Narragansett Bay. Lobsters from the Narragansett Bay survey were obtained from the Rhode Island Department of Environmental Management's monthly transect. Ten lobsters were obtained (5 with shell disease, 5 not exhibiting the disease) from each of 5 transects during 2007 to 2008. Care was taken to include approximately the same numbers of females and males in each transect. Lobster weights varied between 200 g and 1,000 g wet weight, with an average weight of 470 g. In addition to whole animals, composite samples of hemolymph and hepatopancreas, along with discrete samples of lobster tails from lobsters caught in eastern LIS (offshore from Milton, CT) and western LIS (offshore from Oyster Bay) were obtained from SUNY Stony Brook for chemical analyses. These lobsters were collected in cooperation with the New York Department of Environmental Protection in June and July 2007. Finally, as part of the "100 Lobsters" project, lobsters (mean carapace length, 83 mm) were sampled during summer 2008 and shipped to the laboratory of Dr. Jeffrey Shields, Virginia Institute of Marine Science (VIMS), for coordinated dissection, tissue archival, and storage. Samples of hepatopancreas and muscle tissue were received on ice from VIMS for chemical analysis. In addition, 15 reference lobsters were collected in December 2008 from an offshore site near Mt. Desert Rock, ME, by the Maine State Department of Marine Resources. These lobsters were shipped to VIMS and processed as mentioned.

Sediment was also collected via Smith-McIntyre grab at the "100 Lobsters" site in Narragansett Bay, placed in precleaned pint mason jars, and shipped on ice to the University of Maine. Sediment samples were stored frozen at -20[degrees]C.

Dissection and Sample Preparation

Live lobster samples were processed within 6 h of arrival. Prior to dissection, total weight, carapace length, molt stage, sex, vitality, and disease severity were recorded for each lobster. Several photographs of each lobster were also taken for reference. Hepatopancreas, muscle (primarily from claw), gills, ovary, and exoskeleton (from the crusher claw) were dissected from each lobster. Hemolymph was extracted via syringe prior to removal of other tissues. With the exception of exoskeleton and muscle, each tissue was removed in its entirety, and the remaining viscera were discarded. Hemolymph, hepatopancreas, gills, ovary, and muscle samples were placed in cleaned glass containers, and exoskeleton samples were wrapped in clean aluminum foil. All samples were weighed and stored at 20[degrees]C until analysis.

Homogenization differed with each sample type. Hemolymph homogenization was achieved through the use of a Teflon pestle and glass mortar tissue homogenizer. The pestle was attached to a drill, and hemolymph samples were homogenized on ice for 5 min. Gills were not homogenized, but were cut into smaller pieces with scissors. Exoskeleton was homogenized with mortar and pestle. Hepatopancreas required less physical disruption after the freeze thaw process than other tissues. Samples were stirred with a Teflon spatula to increase sample homogeneity. Muscle tissue, used only for the analysis of Hg, was homogenized using a stainless steel blender after verification of no Hg contamination from blank samples. Moisture and dry matter content were determined for all tissue samples. Samples were initially weighed wet, placed in an oven at 100[degrees]C, and dried to constant weight and reweighed. Frozen sediment was thawed and homogenized by stirring thoroughly in the glass jar container. Moisture content was determined in the same manner as for the tissues.

Sample Digestion

Digests for multielemental trace metal analyses were prepared following the general protocols outlined in EPA Method 3051A (EPA 2007). Each run typically included 10 tissue samples of approximately 0.5 g homogenized wet sample, 1 standard reference material, TORT-2 (lobster hepatopancreas, National Research Council, Canada), 1 reagent blank, and 1 spiked sample. For sediment, 6 replicate analyses of 0.5 g wet weight were performed, along with the marine reference sediment MESS-2 (National Research Council, Canada) and a reagent blank. All samples were initially digested with 10 mL highly purified, metals-free Optima grade nitric acid (Fisher Scientific, Pittsburgh, PA) and 2 mL hydrogen peroxide (Optima grade; Fisher Scientific). To prevent excessive sample venting during microwave digestion, the initial digestion time was increased to 48 h.

Digestion was completed using a CEM MARS Microwave (CEM Corporation, Matthews, NC). The method consisted of 3 stages: (1) 5-min ramp to 100[degrees]C, hold for 3 min; (2) 2-min ramp to 150[degrees]C, hold for 3 min; (3) 2-min ramp to 180[degrees]C, hold for 10 min. After microwaving, the samples were weighed to ensure no significant sample loss was experienced during digestion. The digested samples were then each diluted with deionized water to 40 mL total volume for analysis.

Multielemental Analysis by ICP-MS

Digested lobster and sediment samples were analyzed using a Thermo-Electron Element 2 inductively coupled plasma mass spectrometer (ICP-MS; Thermo-Fisher Scientific, Waltham, MA). A standard quartz cyclonic spray chamber and a self-aspirating Teflon nebulizer were used for sample introduction. A calibration curve was done for each run, and aqueous QC samples (including sample blanks, spiked blanks, and standardized reference tissue) were run along with the digest QC samples. Indium (In) was used as an internal standard for all elements.

Total Mercury Analysis by Direct Mercury Analyzer

Mercury analyses were performed using a Milestone DMA-80 Direct Mercury Analyzer (Milestone, Inc., Shelton, CT). The settings for the instrument allowed for 1.5 min of sample drying at 200[degrees]C, 2 min of decomposition at 750[degrees]C, and 1.5 min of wait time at 750[degrees]C. Quality assurance was maintained by running a system blank (no sample boat), followed by aqueous calibration standards (reagent blank, 1 ppm Hg standard), and TORT-2 and DORM-2 (dogfish muscle; National Research Council, Canada) certified reference materials at the beginning of each sample run. Sample runs generally included a total of l0 samples, with duplicate samples and matrix spikes (a 1-ppm standard) included for each of the 5 samples. Sample runs were concluded with additional quality assurance measurements, which included system blanks, reagent blanks, aqueous standards (0.1 ppm and 1 ppm Hg), and certified reference material as described earlier.

Metal Partitioning and Body Burden Determination

Elemental analyses via ICP-MS of muscle, gill, ovary, and exoskeleton tissues were conducted for 8 individual lobsters from the Narragansett Bay 2007 transects for the determination of partitioning of metals throughout the lobster. For sampling uniformity, muscle tissue was taken from the abdomen, and exoskeletal tissues were taken from the dorsal medial inflation of the crusher claw for these samples.

Determination of body burdens for hepatopancreas, hemolymph, gill, and ovary was made using measured metal concentrations and individual organ weights. Because actual muscle and exoskeleton weights were not available for the 8 lobsters sampled, they were estimated from their proportions of total weight (0.23 and 0.46, respectively) derived from a separate allometric investigation (unpubl. data). Total weight and carapace length were determined for 10 lobsters (5 males, 5 females). Each lobster was dissected carefully, and the weight (both wet and dry) was determined for individual tissues, including hepatopancreas, gills, ovaries, muscle, and carapace. Each weight was expressed as a proportion of the total weight of the lobster. These proportions were then applied to the 8 lobsters for which metal concentrations were determined in the individual tissues, for the purpose of determining the total metal loading in each tissue and the total mass of metal being carried by the lobster.

Data Analysis

Concentrations of metal analytes in hepatopancreas tissue and in sediment were expressed as micrograms per gram dry weight of tissue. Hemolymph concentrations were expressed in micrograms per gram wet weight.

One-way analysis of variance (ANOVA) was performed on transformed data from each geographical location (LIS, Narragansett Bay, and Narragansett Bay "100 Lobsters" site), with disease state (shell disease, no apparent disease, reference) as the independent grouping variable. For each location, the offshore Maine site was used as a reference location. The focus was on the contaminant elements (arsenic (As), Cd, cobalt (Co), Cr, copper (Cu), Hg, Mn, molybdenum (Mo), nickel (Ni), lead (Pb), and V). To correct for multiple testing, the P value was modified by dividing by the number of individual l-way ANOVAs performed (n = 33), resulting in a critical P value of 0.0015. Sigma Stat (Systat Software, Inc., Chicago, IL) statistical software was used. All areas were compared with the Maine reference site. Mathematical transformations were performed on each element separately, and included logarithm (log) base 10, reciprocal, the log base 10 of the reciprocal, square root, and arcsine square root. When transformations did not yield normally distributed data, untransformed data were used.

Data were tested for normality with the Shapiro-Wilk W test (Shapiro & Wilk 1965), and homogeneity of variance was tested using Levene's test (Levene 1952). For normally distributed data, the Holm-Sidak multiple comparison method (Sidak 1967) was used to determine significant differences among mean concentrations from lobsters with signs of disease, lobsters without signs of the disease, or reference lobsters. When assumptions of normality or homogeneity of variance were not met, the analysis was performed on ranked data using the Kruskal-Wallis test (Kruskal & Wallis 1952), and post hoc means testing was conducted using Dunn's test (Dunn 1964).

Mean concentrations of contaminant elements in hemolymph were compared between lobsters with signs of shell disease and those with no apparent signs of the disease from the Narragansett Bay transects. Nonparametric Mann-Whitney rank sum tests (Mann & Whitney 1947) were performed to assess differences between means.

Multivariate analyses were conducted with Systat statistical software (Systat Software, Inc.). Principal component analyses were conducted after mean centering the data, which involved subtraction of the mean concentration for a given element from each individual sample and sample replicate. Discriminant analyses were also performed, using both mean-centered and log-transformed data. Only those data that met the assumption of homogeneity of variance were used. To meet these assumptions, a logarithm base 10 transformation was performed on the "100 Lobsters" data, which produced normality in a subset of the data. For the Narragansett Bay transect data, a combination of log base 10 transformations, reciprocal transformations, and the log base 10 of the reciprocal were used.

RESULTS

Quality Assurance

For ICP-MS, percent recovery of 17 different elements from spiked samples ranged from 84-118% across all analytes, and had an overall mean of 104 [+ or -] 2%. Procedural blanks of contaminant elements were generally low and ranged from 0.01-2% of the sample concentration. However, higher concentrations of some critical elements (Pb, Ni, Cd) during later sample runs necessitated blank subtraction from all samples. A blank was run with every 10 samples, as mentioned earlier, and sample blanks were matched to their respective samples for the purpose of subtraction.

Sample blanks for Hg were very low (<1%) and were not subtracted from samples. Duplicates deviated less than 10% from one another, and standard reference materials were within 90-110% of reported values.

Comparison of Mean Concentrations in Shell Diseased, Nondiseased, and Reference Lobsters

Concentrations of contaminant metals in the hepatopancreas of shell-diseased lobsters collected from different sites in SNE and LIS were characterized by nonnormal distributions and distributions skewed heavily in the positive direction. Mean, SD, and range of concentrations of 11 metals associated with anthropogenic contamination are presented in Tables 1, 2, and 3, along with the results of the statistical comparison of the mean concentrations. Differences were observed between geographical locations and between shell-diseased and nondiseased groups within a geographical location, although patterns were not consistent with a scenario of higher body burdens in lobsters showing signs of shell disease. For the "100 Lobsters" and LIS sites, differences in Cd, Cu, and Pb were seen between lobsters from these sites and those from the Maine reference site, with these metals (along with V and Mn) being higher compared with the offshore Maine reference site (Tables 1 and 3). No differences were seen between lobsters with signs of shell disease and lobsters without signs of shell disease at these sites. In the Narragansett Bay transect data, differences were seen between lobsters in the diseased and nondiseased groups for 4 metals (Cr, Cu, Mn, and Pb). However, mean hepatopancreas concentrations were lower in the lobsters with signs of ESD compared with the lobsters without signs (Table 2). Cu concentrations were significantly different between all 3 groups (diseased, nondiseased, and reference), with the lowest concentrations from reference site lobsters and the highest concentrations found in lobsters not exhibiting signs of shell disease.

Concentrations in Hemolymph

Concentrations of contaminant hemolymph metals are presented in Table 4 for Narragansett Bay lobsters. Data were distributed nonnormally (even after mathematical transformations) and were low in concentration. No consistent trends were apparent, although a couple of statistically significant differences were noted, with Mn being higher in concentration in the lobsters with no apparent signs of shell disease, and As being slightly higher in the hemolymph of lobsters with shell disease.

Multivariate Analysis Investigations

Recognizing the multivariate nature of the data sets generated, data were examined using a number of multivariate analyses, including principal components analysis and discriminant analysis. Running principal component analysis on covariances, using the contaminant metals As, boron (B), Cd, Co, Cr, Mn, Mo, Ni, Pb, V, and Hg produced axes that accounted for 83-95% of the variance on principal component 1, 5-20% on axis 2, and 1-2% on axis 3. Very little to no separation was seen when 2-D plots of eigenvalues from any 2 of the main 3 axes, or 3-dimensional plots of the first 3 axes were examined (Fig. 1). This is consistent with multivariate analyses of variance and discriminant analysis, both performed using contaminant metals. Results of the canonical scores plot from the discriminant analysis are displayed in Figure 2. It can be seen that there was no separation between groups. This is also reflected in the MANOVA test statistics; both Wilk's Lambda and Pillai's Trace showed nonsignificance (P = 0.295 and P = 0.206, respectively) between groups, suggesting that the disease state (shell disease, no disease, reference) had no influence on ordering the data. Similar results were obtained with the Narragansett Bay transect data.

[FIGURE 1 OMITTED]

[FIGURE 2 OMITTED]

Sediment Analysis

Results for the sediment samples obtained from the "100 Lobsters" collection site are given in Table 5. When compared with sediment quality guidelines, Narragansett Bay sediment collected from the "100 Lobsters" sampling site where shell disease is prevalent showed that concentrations of several metals were intermediate between effect range low (ERL) and effect range median (ERM) sediment quality values, which are based on an extensive compilation of sediment toxicity tests. One metal, Ni, exceeded sediment quality guidelines.

Metals Partitioning and Body Burden Determination

Elemental concentrations were not distributed uniformly throughout all tissues, in particular, concentrations of contaminants such as As, Cd, Co, and Cu were highest in hepatopancreatic tissues (Table 6). Other contaminants, such as Cr, Mn, Ni, and Pb, had different distributions, with the highest levels of Cr seen in gills, and the highest levels of Pb, Mn, and Ni found in the exoskeleton. Concentrations of all contaminant metals were generally lowest in hemolymph, although hemolymph Cu concentrations were similar to those found in ovary and gill.

Contaminant metal body burden estimates (i.e., total mass of metal) are shown in Table 7. Hepatopancreas had the greatest burden for Cd (91%) and Cu (64%), whereas the exoskeleton had the greatest burden of Cr (86%), Co (39%), Mn (93%), Ni (93%), and Pb (84%). Muscle tissue contained the highest burden of As (45%; Fig. 3).

DISCUSSION

The results showed that major differences in hepatopancreas contaminant metals did not exist among lobsters showing signs of ESD, not showing signs of ESD, and reference groupings for Rhode Island transects, LIS samples, and the "100 Lobsters" site. The results were not consistent with a scenario of higher body burdens present in diseased versus nondiseased or reference organisms. Thus, one cannot infer that greater body burdens of metals in shell-diseased lobsters were contributing to the onset of the disease, compared with lobsters that were not exhibiting the disease caught in the same location, or in a less impacted reference site. Furthermore, there was no clear pattern to suggest that any singular element or combination of elements was more associated with the disease state than not. Univariate F tests and post hoc analyses revealed differences in mean concentrations of contaminant metals, although, in general, differences were seen between different geographical locations more often than between diseased and nondiseased groups. The multivariate analyses also did not show any significant trends between contaminants and the presence of shell disease.

Like hepatopancreas, hemolymph concentrations of contaminants showed a similar lack of disease-related trends. Although measurable levels of contaminant metals were found, values for all metals were consistently very low in comparison with hepatopancreas, and did not justify the use of hemolymph for metals measurement. As expected, hemolymph Cu concentrations were highest for all contaminant metals. However, Cu content did not appear to have been indicative of disease state, as it was similar to those reported for H. americanus (Engel et al. 2001) and other crustacean species (Norum et al. 2003).

Our analysis of the relationship of shell disease to contaminant metal burdens was undoubtedly complicated by the high degree of variability associated with the samples. Recent literature confirms the challenges of interpreting contaminant levels for crustacean tissue resulting from high degrees of both interand intraspecific variability in concentrations (Wang & Rainbow 2008, Reichmuth et al. 2010). Variability is known to arise from a variety of sources including size, sex, molt stage and season, and geographical origin in many crustacean species (Canli & Furness 1993, Paez-Osuna et al. 1995, Barrento et al. 2009). In H. americanus, variation in metal content has been related to the molt cycle by Engel et al. (2001), who observed that hepatopancreas Cu and Zn concentrations decreased by a factor of 2 at ecdysis. Geographical origin is also known to influence metals concentrations strongly in lobsters such that lobsters from different areas can be identified by their different metals signatures (Chou et al. 2003). In our study, some variability was likely attributable to size (e.g., mean hepatopancreas Cd concentration equaled 41.4 [micro]g/g dry weight for Narragansett Bay/Rhode Island Sound lobsters larger than 600 g), but no definitive relationship for sex and metals content was evident. Molt stage was essentially the same across all lobsters sampled (intermolt), so its influence on sample concentrations could not be assessed. Lobster origin likely represented the largest source of sample variability we experienced, yet metal concentrations in the "100 Lobsters" animals still displayed individual variability despite being collected from the same geographical location. Thus, there may have been other unknown factors, such as routes of exposure and/or geochemical availability (Luoma & Rainbow 2005) that contributed to the metal concentrations found in lobsters.

Concentrations of metal contaminants in lobster hepatopancreas from Narragansett Bay and LIS were similar to levels found in lobsters from environments known to be impacted (Table 8). Only Cu deviated from known ranges for lobster, with values exceeding those recorded prior to this study. Higher Cu concentrations could have reflected anthropogenic influences, as lobsters are known to accumulate Cu according to its availability in the environment (Engel et al. 2001, Yeats & Chou 2001). Alternatively, higher Cu concentrations may have been indicative of enhanced hemocyanin production in response to environmental conditions such as hypoxia, temperature, or food availability (DeFur et al. 1990, Engel et al. 2001).

[FIGURE 3 OMITTED]

Historically, development of shell disease in lobsters and other crustaceans has been associated with polluted or degraded habitat (Young & Pearce 1975, Ziskowski et al. 1996, Vogan et al. 2008). Sediment contaminant concentrations at the "100 Lobsters" sample site were typical of many sediments of northeastern U.S. estuaries, where the regional sediment quality is generally rated "fair to poor" (U.S. Environmental Protection Agency 2008). The levels of As, Cr, Ni, and Pb found at that site suggested that potential adverse biological effects may have occurred in lobsters living in that environment (Table 5). However, it was difficult to predict the likelihood of adverse effects from sediment concentrations alone, because lobsters harvested at this site are not permanent residents, but are migrating through the site; the bioavailability of contaminant metals was not determined as part of this study; and contaminant concentrations in lobster tissues do not always correspond to contaminant concentrations in sediment (Engel et al. 2001, Yeats & Chou 2001). Furthermore, a direct association of sediment contaminant metals levels at the "100 Lobsters" site to shell disease was not evident, given that lobsters inhabit even more highly contaminated sediments throughout their range without exhibiting signs of shell disease (Glenn & Pugh 2006). If sediment metals are important to ESD development, a more complex relationship must exist among the lobster, the environment and the pathogen than was discerned from this study (Tlusty et al. 2007).

Metals Partitioning and Body Burdens

Our measured contaminant metals concentrations for muscle, ovary, gill, and exoskeleton were comparable with previous investigations. For muscle, Barrento et al. (2008) found similar concentrations of As and Pb in H. arnericanus. Cd was an order of magnitude lower in that study, however, and concentrations from the current investigation paralleled concentrations more closely observed in lobsters from polluted environments (Uthe et al. 1987). Muscle tissue concentrations of Cu, Cr, and Ni were also within established ranges for lobster (Reid et al. 1982, Connecticut Department of Environmental Protection 1987 (cited in Mercaldo-Allen & Kuropat 1994)). For ovary, Cu and Mn were higher than previously recorded for H. americanus (Barrento et al. 2009). Information on metal concentrations found in gills and exoskeleton is limited for H. americanus. Eriksson & Baden (1998) found high concentrations (1,560 [micro]g/g dry weight) of Mn in the gills of the Norway lobster Nephrops norvegicus (L.), reflective of exposure to hypoxic conditions. Baden & Eriksson (2006), in a review of Mn in crustaceans, found that concentrations ranging from 0.5-508 [micro]g/g were present in the gills of N. norvegicus collected from pristine areas. Concentrations found in Narragansett Bay lobsters (2.65 [micro]g/g wet weight, approximately 13 [micro]g/g dry weight) were apparently not reflective of a hypoxic environment.

The current study represented the first attempt in the American lobster to document whole body burdens of multiple contaminant metals using a variety of tissues. Earlier investigations were limited to single tissues (Chou et al. 2000). Here, we observed a pattern of contaminant burden by tissue that could be summarized as hepatopancreas/exoskeleton > gills/ gonads > muscle > hemolymph. This pattern approximated that described for other lobster species (Canli & Furness 1993, Paez-Osuna et al. 1995, Morales-Hernandez et al. 2004). Comparisons of percent of body burden between H. americanus and N. norvegicus suggested similar distributions of Cd (91%) and Cu (65%) in the hepatopancreas of the 2 species. And although the percentage of Pb in the exoskeleton was different between the American and the Norwegian lobsters (84% vs. 42%), it was comparable between the American lobster and the spiny lobster Panulirus inflatus (Bouvier) (Paez-Osuna et al. 1995). Body burden percentages for exoskeletal Mn and Ni were also roughly equivalent between H. americanus and P. inflatus (Paez-Osuna et al. 1995). The consequence of the total contaminant body burden is uncertain because field-collected lobsters are known to carry high metal loads with no apparent ill effects (Chou et al. 2002).

The observed intraorganismal partitioning likely reflected the distinct functions of each organ in metals processing, and may have also been indicative of the intake route (diet vs. seawater) (Rainbow 2002). Relatively high hepatopancreas concentrations and burdens of metals were expected given this organ's prominent role in metal absorption, sequestration, and detoxification processes in crustaceans (Ahearn et al. 2004). In H. americanus, the hepatopancreas is known to regulate both essential and toxic metal ions by metallothionein binding (Engel & Brouwer 1986, Chou et al. 1991, Chavez-Crooker et al. 2003). High levels of exoskeletal metals are also typical of decapod crustaceans (Paez-Osuna et al. 1995, Morales-Hernandez et al. 2004). Metals accumulate in the exoskeleton by binding with chitin or other physiological processes (Keteles & Fleeger 2001), and may be subsequently depurated through molting or shifted to soft tissues prior to molting (Bergey & Weis 2007). Crustacean gills are known to represent major sites of metals uptake (Rainbow 2002), intermediate sites of accumulation (Chavez-Crooker et al. 2003), and possibly elimination (Ahearn et al. 2004), whereas hemolymph and hemocytes are responsible for metals binding and transport to various tissues of the body (Chavez-Crooker et al. 2003, Ahearn et al. 2004). Ovary and muscle tissue in metal metabolism are less defined, but metal accumulation is known to occur to some degree in both tissues (Jeckel et al. 1996). Muscle tissue accumulation appears more limited, presumably because it contains low metallothionein levels (Chavez-Crooker et al. 2003).

As a result of inherent sample variability and limited size of this partitioning experiment, it was difficult to determine whether there were disease-associated differences in contaminant concentrations or loads in any tissue. However, notably lower mean ([+ or -] SD) concentrations of calcium (80 [+ or -] 6 mg/g for shell diseased vs. 108 [+ or -] 9 mg/g for nondiseased) and strontium (4.3 [+ or -] 0.6 [micro]g/g for shell diseased vs. 1,280 [+ or -] 970 [micro]g/g for nondiseased) were found in the exoskeletal tissues of diseased lobsters. These results were presumably attributable to the physical loss of mineralized shell material via lesion development, but further investigation of this phenomenon may be warranted to ascertain its relationship to shell disease.

CONCLUSIONS

Contaminant trace metals were detected in the tissues of lobsters showing signs of shell disease collected in Rhode Island and LIS. These concentrations were reflective of environments that are impacted moderately. The concentrations of contaminant metals in sediment collected from the "100 Lobsters" site were consistent with this scenario, with concentrations that were intermediate between ERL and ERM concentrations. The study focused mainly on concentrations in the hepatopancreas, the known site of metal sequestration. The data were highly variable and skewed, which is typical of metal concentrations in aquatic organisms. Although differences in concentration patterns were observed between diseased lobsters and those not exhibiting disease symptoms, no consistent picture emerged that would lead one to conclude that metal burdens were a causative agent in lobster shell disease. Metal concentrations in hemolymph were highly skewed and also did not indicate a relationship to the presence of shell disease.

DOI: 10.2983/035.031.0214

ACKNOWLEDGMENTS

We thank Kathy Castro and Barbara Somers of the University of Rhode Island, Tom Angell of the Rhode Island Department of Environmental Management, Margaret Homerding of Stony Brook University, Jeff Shields of VIMS, and Carl Wilson of the Maine Department of Marine Resources for their assistance in obtaining lobster samples for this study. Gratitude is also extended to Brian Perkins and Denise Skonberg of the University of Maine's Department of Food Science and Human Nutrition, and to John Cangelosi, Clive Devoy, Mike Handley, and Marianne Lagerklint of University of Maine's Sawyer Environmental Chemistry Research Lab for providing laboratory facilities and guidance. This work was supported by the National Marine Fisheries Service as the New England Lobster Research Initiative: Lobster Shell Disease under NOAA grant NA06NMF4720100 to the University of Rhode Island Fisheries Center. The views expressed herein are those of the authors and do not necessarily reflect the views of NOAA or any of its subagencies. The U.S. government is authorized to produce and distribute reprints for government purposes, notwithstanding any copyright notation that may appear hereon.

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LAWRENCE A. LEBLANC * AND DEANNA PRINCE School of Marine Sciences, 5741 Libby Hall, Room 215, School of Marine Sciences, University of Maine, Orono, ME 04469-5741

* Corresponding author. E-mail: lawrence.leblanc@umit.maine.edu
TABLE 1.
Comparison of mean concentrations of contaminant metals * in the
hepatopancreas of diseased and nondiseased lobsters from the "100
Lobster" study.

                          As        Cd ([dagger])       Co
                      ([micro]g/g    ([micro]g/g    ([micro]g/g
                       dry wt.)       dry wt.)       dry wt.)

Shell disease
  Mean                   43.9          11.7 A          1.26
  SD                     50.9           16.6           5.71
  Range                20.0-65.7      2.76-36.7     0.105-0.809
    (n = 32)

None apparent
  Mean                   28.0          9.77 A          0.26
  SD                     10.4           5.47           0.14
  Range                15.1-60.0      2.30-23.0      0.09-0.76
    (n = 32)

Maine reference
  Mean                   35.4          18.8 B          0.29
  SD                     17.4           15.1           0.22
  Range ([section])    16.8-90.5     0.397-74.5     0.110-0.829
    (n = 32)

                          Cr        Cu ([dagger])       Mn
                      ([micro]g/g    ([micro]g/g    ([micro]g/g
                       dry wt.)       dry wt.)       dry wt.)

Shell disease
  Mean                   1.31          2150 A          10.6
  SD                     5.75            912           25.7
  Range               0.049-2.86      197-3730       0.621-164
    (n = 32)

None apparent
  Mean                   0.32          1,780 A         8.33
  SD                     0.26           1,070          4.69
  Range               0.056-1.10      264-3,730      2.57-22.9
    (n = 32)

Maine reference
  Mean                   0.21           189 B          7.42
  SD                     0.29            230           2.22
  Range ([section])   0.027-1.26      16.7-802      4.97-12.63
    (n = 32)

                          Mo            Ni        Pb ([dagger])
                      ([micro]g/g   ([micro]g/g    ([micro]g/g
                       dry wt.)      dry wt.)       dry wt.)

Shell disease
  Mean                   2.33          2.27          0.22 A
  SD                     2.33          2.27           0.51
  Range               0.179-48.2    0.303-34.9     0.010-3.27
    (n = 32)

None apparent
  Mean                   1.53          1.23          0.20 A
  SD                     2.48          1.10           0.18
  Range               0.436-11.5    0.272-5.02     0.039-0.817
    (n = 32)

Maine reference
  Mean                   1.47          1.69          0.05 B
  SD                     0.75          0.79           0.02
  Range ([section])    0.19-3.04    0.781-3.25     0.018-0.105
    (n = 32)

                                    Hg ([double
                           V         dagger])
                      ([micro]g/g   ([micro]g/g
                       dry wt.)      dry wt.)

Shell disease
  Mean                  1.26 A         0.56
  SD                     5.56          0.29
  Range               0.031-31.7    0.226-1.09
    (n = 32)

None apparent
  Mean                  0.33 A         0.36
  SD                     0.35          0.17
  Range               0.085-1.58    0.005-0.733
    (n = 32)

Maine reference
  Mean                  0.87 B         0.26
  SD                     0.57          0.11
  Range ([section])   0.187-2.29    0.181-0.310
    (n = 32)

* All concentrations are in hepatopancreas tissue except for Hg, which
is in muscle tissue. Concentrations are averaged across disease stage
and sex.  Statistically significant differences, tested with either
Holm-Sidak or Dunn's (when the nonparametric Kruskal-Wallis test was
used) are indicated  by letters in bold type.

([dagger]) One-way analysis of variance and post hoc means testing
using Dunn's test performed on rank-transformed data.

([double dagger]) For Hg: diseased, n = 14; no disease, n = 15;
reference, n = 15.

([section]) The range of values for the Maine reference site appears
only in Table 1.

TABLE 2.
Comparison of mean concentrations of contaminant metals * in the
hepatopancreas of diseased and nondiseased lobsters from the
Narragansett Bay transects.

                       As            Cd            Co
                   ([micro]g/g   ([micro]g/g   ([micro]g/g
                    dry wt.)      dry wt.)      dry wt.)
Shell disease
  Mean                28.1          26.3          0.40
  SD                  17.4          28.7          0.35
  Range (n = 14)    5.61-340        28.74         0.35
None apparent
  Mean                48.9          15.4          0.86
  2SD                 24.7           9.0          0.82
  Range (n = 17)    15.1-120      2.3-39.0     0.086-2.87
Maine reference
  Mean                35.4          18.8          0.29
  2SD                 17.4          15.1          0.22

                       Cr            Cu        Mn ([dagger])
                   ([micro]g/g   ([micro]g/g    ([micro]g/g
                    dry wt.)      dry wt.)       dry wt.)
Shell disease
  Mean               0.30 A         816 A         6.83 A
  SD                  0.26          1,040          4.43
  Range (n = 14)      0.26          1,045          4.43
None apparent
  Mean               0.59 B        1,400 B        14.0 B
  2SD                 0.58          1,410           7.9
  Range (n = 17)   0.056-1.96     141-5,270      2.57-38.3
Maine reference
  Mean               0.21 A         189 C         7.42 A
  2SD                 0.23           230           2.22

                       Mo            Ni        Pb ([dagger])
                   ([micro]g/g   ([micro]g/g    ([micro]g/g
                    dry wt.)      dry wt.)       dry wt.)
Shell disease
  Mean                0.94          3.43          0.09 A
  SD                  0.40          9.07           0.07
  Range (n = 14)      0.40          9.07           0.07
None apparent
  Mean                1.77          1.66          0.31 B
  2SD                 0.98          1.04           0.38
  Range (n = 17)    0.43-11.5     0.27-5.02      0.02-1.61
Maine reference
  Mean                1.47          1.69          0.05 A
  2SD                 0.75          0.79           0.02

                                 Hg ([double
                        V         dagger])
                   ([micro]g/g   ([micro]g/g
                    dry wt.)      dry wt.)
Shell disease
  Mean                0.763         0.43
  SD                  1.06          0.23
  Range (n = 14)   0.031-31.7    0.176-1.09
None apparent
  Mean                1.22          3.57
  2SD                 2.31          6.54
  Range (n = 17)    0.08-9.71      ND-23.1
Maine reference
  Mean                0.87          0.26
  2SD                 0.57          0.11

* All concentrations are in hepatopancreas tissue except for Hg, which
is in muscle tissue. Concentrations are averaged across disease stage
and sex.  Statistically significant differences, tested with either
Holm-Sidak or Dunn's (when the nonparametric Kruskal-Wallis test was
used) are indicated  by letters in bold type.

([dagger]) One-way analysis of variance and post hoc means testing
using Dunn's test performed on rank-transformed data.

([double dagger]) For Hg: diseased, n = 15; no disease, n = 17;
reference, n = 15.

ND, not detected.

TABLE 3.
Comparison of mean concentrations of contaminant metals * in the
hepatopancreas of diseased and nondiseased lobsters from Long  Island
Sound.

                      As        Cd ([dagger])       Co
                  ([micro]g/g    ([micro]g/g    ([micro]g/g
                   dry wt.)       dry wt.)       dry wt.)

Shell disease
  Mean              39.9 A         26.2 A         0.68 A
  SD                 13.1            7.4           0.47
  Range            16.9-59.8      11.5-37.1      0.28-1.89
    (n = 9)

None apparent
  Mean              18.7 B         12.1 B         0.53 A
  SD                 11.4            6.5           0.24
  Range            4.56-53.0      1.53-21.0     0.088-0.924
    (n = 17)

Maine reference
  Mean              35.4 A         18.8 B         0.29 B
  SD                 17.4           15.1           0.22

                      Cr            Cu        Mn ([dagger])
                  ([micro]g/g   ([micro]g/g    ([micro]g/g
                   dry wt.)      dry wt.)       dry wt.)

Shell disease
  Mean               0.21          792 A         18.5 A
  SD                 0.11           628           17.1
  Range           0.096-0.396    45.7-2280      2.34-61.4
    (n = 9)

None apparent
  Mean               0.65          977 A         12.9 A
  SD                 1.50          1,000           5.8
  Range           0.090-6.42    10.34-3710      2.42-28.8
    (n = 17)

Maine reference
  Mean               0.21          189 B         7.42 B
  SD                 0.23           230           2.22

                      Mo            Ni        Pb ([dagger])
                  ([micro]g/g   ([micro]g/g    ([micro]g/g
                   dry wt.)      dry wt.)       dry wt.)

Shell disease
  Mean               1.63          1.76          0.19 A
  SD                 0.96          1.03           0.14
  Range           0.805-3.99    0.884-4.22     0.090-0.539
    (n = 9)

None apparent
  Mean               2.60          1.58          0.13 A
  SD                 5.22          1.25           0.08
  Range           0.368-22.7    0.367-6.05     0.044-0.328
    (n = 17)

Maine reference
  Mean               1.47          1.69          0.05 B
  SD                 0.75          0.79           0.02

                                Hg ([double
                       V         dagger])
                  ([micro]g/g   ([micro]g/g
                   dry wt.)      dry wt.)

Shell disease
  Mean               1.18          1.76
  SD                 0.73          1.26
  Range           0.313-2.82    0.546-3.68
    (n = 9)

None apparent
  Mean               1.19          1.15
  SD                 1.73          0.29
  Range           0.246-7.62    0.291-1.54
    (n = 17)

Maine reference
  Mean               0.87          0.26
  SD                 0.57          0.11

* All concentrations are in hepatopancreas tissue except for Hg, which
is in muscle tissue. Concentrations are averaged across disease stage
and sex.  Statistically significant differences, tested with either
Holm-Sidak or Dunn's (when the nonparametric Kruskal-Wallis test was
used) are indicated  by letters in bold type.

([dagger]) One-way analysis of variance and post hoc means testing
using Dunn's test performed on rank-transformed data.

([double dagger]) For Hg: diseased, n = 7; no disease, n = 11;
reference, n = 15.

TABLE 4.
Mean, SD, and minimum and maximum values for selected metal
contaminants in hemolymph from lobsters from the  Narragansett Bay
transects.

              As *            Cd             Co             Cr
          ([micro]g/g)   ([micro]g/g)   ([micro]g/g)   ([micro]g/g)

No apparent shell disease (n = 17)

  Mean        0.41          0.0004         0.009          0.113
  SD          0.72          0.0007         0.005          0.167
  Range    0.05-3.04       0-0.0022     0.004-0.021    0.004-0.579

Shell disease (n = 13)

  Mean        0.58          0.001           0.01           0.52
  SD          0.28          0.002           0.01           1.72
  Range    0.13-1.02       0-0.008      0.003-0.035     0.01-6.93
                          ([dagger])

               Cu            Mn *            Ni             Pb
          ([micro]g/g)   ([micro]g/g)   ([micro]g/g)   ([micro]g/g)

No apparent shell disease (n = 17)

  Mean        46.1          0.774          0.071          0.002
  SD          21.9          0.369          0.067          0.003
  Range    14.0-97.5      0.383-1.7      0.012-0.22    0.0005-0.013

Shell disease (n = 13)

  Mean         59           0.436          0.116          0.003
  SD          23.4          0.568          0.264          0.005
  Range      24-121       0.081-2.2      0.013-1.07       0-0.02

* Statistically significant differences, conducted with nonparametric
Mann-Whitney rank sum tests.

([dagger]) High value is from one very large lobster heavily impacted
by shell disease. Concentrations of cadmium were approximately twice
as high as in any other sample.

TABLE 5.

Comparison of mean trace metal concentrations in sediment collected
from Narragansett Bay, RI, for the "100 Lobster" project.

                 As             Cd             Co
                 ([micro]g/g)   ([micro]g/g)   ([micro]g/g)

Mean (n = 5)            15.3            0.2           25.1
SD               [+ or -]1.3    [+ or -]0.1    [+ or -]2.1
ERL *                    8.2            1.2
ERM ([dagger])          70              9.6

                 Cr             Cu             Mn
                 ([micro]g/g)   ([micro]g/g)   ([micro]g/g)

Mean (n = 5)           104             30.6           815
SD               [+ or -]2.2    [+ or -]2.8    [+ or -]37.7
ERL *                   81             34
ERM ([dagger])         370            270

                 Mo             Ni
                 ([micro]g/g)   ([micro]g/g)

Mean (n = 5)             1.4           420
SD               [+ or -]0.1    [+ or -]24.7
ERL *                                   30
ERM ([dagger])                          50

                 Pb             V
                 ([micro]g/g)   ([micro]g/g)

Mean (n = 5)            106     170
SD               [+ or -]82.4   [+ or -]42.8
ERL *                    47
ERM ([dagger])          218

Sediment quality guidelines for 6 contaminant metals are also
listed (from Long et al. (1995)).

* ERL, effects range low; the 10h percentile of the
concentration range where biological effects were seen for a
given analyte in biological test organisms exposed to sediment.

([dagger]) ERM, effects range median; the 50th percentile value
of the concentration range where biological effects were seen
for a given analyte in biological test organisms exposed to
sediment.

TABLE 6.
Mean concentrations of selected contaminant metals in tissues of
Homarus americanus from Narragansett Bay, RI.

                              As               Cd
                         ([micro]g/g)     ([micro]g/g)
    Tissue                 wet wt.)         wet wt.)

Muscle           Mean        10.5             1.13
(n = 8)          SD     [+ or -] 2.17    [+ or -] 3.12
Gills            Mean        6.82             1.22
(n = 8)          SD     [+ or -] 2.57    [+ or -] 1.20
Exoskeleton      Mean        2.21            0.024
(n = 8)          SD     [+ or -] 1.56    [+ or -] 0.017
Ovary            Mean        14.2            0.059
(n = 5)          SD     [+ or -] 2.82    [+ or -] 0.062
Hemolymph        Mean       0.444            0.002
(n = 8)          SD     [+ or -] 0.306   [+ or -] 0.002
Hepatopancreas   Mean        16.1             15.5
(n = 8)          SD     [+ or -] 6.68    [+ or -] 16.2

                              Co               Cr
                         ([micro]g/g)     ([micro]g/g)
    Tissue                 wet wt.)         wet wt.)

Muscle           Mean       0.005            0.012
(n = 8)          SD     [+ or -] 0.001   [+ or -] 0.006
Gills            Mean       0.103            0.338
(n = 8)          SD     [+ or -] 0.106   [+ or -] 0.259
Exoskeleton      Mean       0.021            0.283
(n = 8)          SD     [+ or -] 0.007   [+ or -] 0.194
Ovary            Mean       0.079            0.052
(n = 5)          SD     [+ or -] 0.033   [+ or -] 0.034
Hemolymph        Mean       0.008            0.073
(n = 8)          SD     [+ or -] 0.003   [+ or -] 0.130
Hepatopancreas   Mean        5.2             0.155
(n = 8)          SD     [+ or -] 0.212   [+ or -] 0.160

                              Cu               Mn
                         ([micro]g/g)     ([micro]g/g)
    Tissue                 wet wt.)         wet wt.)

Muscle           Mean        5.28            0.853
(n = 8)          SD     [+ or -] 2.11    [+ or -] 1.21
Gills            Mean         47              2.65
(n = 8)          SD     [+ or -] 22.4    [+ or -] 1.42
Exoskeleton      Mean       0.944             50.1
(n = 8)          SD     [+ or -] 0.340   [+ or -] 49.9
Ovary            Mean        44.6             4.28
(n = 5)          SD     [+ or -] 22.5    [+ or -] 2.54
Hemolymph        Mean         42             0.448
(n = 8)          SD     [+ or -] 16.9    [+ or -] 0.001
Hepatopancreas   Mean        433              3.63
(n = 8)          SD      [+ or -] 428    [+ or -] 2.69

                              Ni               Pb
                         ([micro]g/g)     ([micro]g/g)
    Tissue                 wet wt.)         wet wt.)

Muscle           Mean        1.25            0.074
(n = 8)          SD     [+ or -] 3.44    [+ or -] 0.199
Gills            Mean       0.474            0.089
(n = 8)          SD     [+ or -] 0.153   [+ or -] 0.074
Exoskeleton      Mean        5.88             0.12
(n = 8)          SD     [+ or -] 0.888   [+ or -] 0.064
Ovary            Mean       0.151            0.009
(n = 5)          SD     [+ or -] 0.040   [+ or -] 0.007
Hemolymph        Mean       0.053            0.002
(n = 8)          SD     [+ or -] 0.387   [+ or -] 0.001
Hepatopancreas   Mean       0.543              0
(n = 8)          SD     [+ or -] 0.265   [+ or -] 0.112

TABLE 7.

Mean burdens of selected contaminant metals [+ or -] SD in tissues of
Homarus americanus from Narragansett Bay, RI.

Tissue                        As                Cd
                             (mg)               (mg)

Muscle             Mean       1,400               70.7
(n=8)              SD     [+ or -]2.17      [+ or -]3.12
Gills              Mean         113                28.8
(n = 8)            SD     [+ or -]2.57      [+ or -]1.20
Exoskeleton        Mean         734                 7.06
(n = 8)            SD     [+ or -]1.56      [+ or -]0.017
Ovary              Mean         248                 1.11
(n = 5)            SD      [+ or -]2.82     [+ or -]0.062
Hemolymph          Mean           17.6              0.063
(n = 8)            SD      [+ or -]0.306    [+ or -]0.002
Hepatopancreas     Mean          592             1140
(11 = 8)           SD      [+ or -]6.68    [+ or -]16.2
Mean body burden               3,100            1,250

Tissue                   Co              Cr             Cu
                        (mg)            (mg)           (mg)

Muscle                     0.614           1.72          771
(n=8)              [+ or -]0.001   [+ or -]0.006   [+ or -]2.11
Gills                      1.06            5.28          687
(n = 8)            [+ or -]0.106   [+ or -]0.259   [+ or -]22.4
Exoskeleton                5.37           86.3           303
(n = 8)            [+ or -]0.007   [+ or -]0.194   [+ or -]0.340
Ovary                      1.35            1.06          695
(n = 5)            [+ or -]0.033   [+ or -]0.034   [+ or -]22.5
Hemolymph                  0.268           2.48         1,430
(n = 8)            [+ or -]0.003   [+ or -]0.130   [+ or -]16.9
Hepatopancreas             5.15            3.69          6,980
(11 = 8)           [+ or -]0.212   [+ or -]0.160   [+ or -]428
Mean body burden          13.8           100           10,800

Tissue                   Mn             Ni              Pb
                        (mg)           (mg)            (mg)

Muscle                    90.4            77.7             4.61
(n=8)              [+ or -]1.21    [+ or -]3.44    [+ or -]0.199
Gills                     29.5             6.73            0.776
(n = 8)            [+ or -]1.42    [+ or -]0.153   [+ or -]0.074
Exoskeleton            8,600           1,500              28.2
(n = 8)            [+ or -]49.9    [+ or -]0.888   [+ or -]0.064
Ovary                      49.5            2.68            0.104
(n = 5)            [+ or -]2.54    [+ or -]0.040   [+ or -]0.007
Hemolymph                 11.7             1.68            0.048
(n = 8)            [+ or -]0.001   [+ or -]0.387   [+ or -]0.001
Hepatopancreas            96.3            17.8             0
(11 = 8)           [+ or -]2.69    [+ or -]0.265   [+ or -]0.112
Mean body burden       8,900           1,600              33.8

TABLE 8.
Comparison of trace metals in 100 lobster hepatopancreas * with
hepatopancreas concentrations found in other studies.

                                     Cd ([micro]g/g dry
Disease State                                 wt.)

Current study
Shell disease (n = 32)                     2.9-100
                                     (11.7 [+ or -] 6.6)
No apparent disease                        2.6-2.3
(n = 25)                             (9.77 [+ or -] 5.47)
Other studies
Boston Harbor([dagger])                    1.2-6.6
Long Island Sound([double dagger])         5.4-50.8
Outfall (GOC)([subsection])              11.5 + 7.5
Maine([paragraph)                          1.8-59.8

                                     Cr ([micro]g/g dry
Disease State                              wt.)

Current study
Shell disease (n = 32)                      0.08-35
                                      (1.31 [+ or -] 5.75)
No apparent disease                         0.08-1.2
(n = 25)                             (0.32 [+ or -] 0.26)
Other studies
Boston Harbor([dagger])                    0.1-2.5
Long Island Sound([double dagger])           0-0.75
Outfall (GOC)([subsection])               0.9 + 0.5
Maine([paragraph)                         0.07-1.3

                                     Cu ([micro]/g dry
Disease State                                wt.)

Current study
Shell disease (n = 32)                     200-4,300
                                     (2150 [+ or -] 912)
No apparent disease                        300-3,700
(n = 25)                             (1,780 [+ or -] 1,070)
Other studies
Boston Harbor([dagger])                     37-640
Long Island Sound([double dagger])         180-3,880
Outfall (GOC)([subsection])               380 + 500
Maine([paragraph)                           12-500

                                     Ni([micro]g/g dry
Disease State                                 wt.)

Current study
Shell disease (n = 32)                     0.32-31
                                     (2.27 [+ or -] 4.83)
No apparent disease                        0.27-5.1
(n = 25)                             (1.28 [+ or -] 1.1)
Other studies
Boston Harbor([dagger])                        --
Long Island Sound([double dagger])         0.90-5.97
Outfall (GOC)([subsection])                 4.4+77
Maine([paragraph)                          0.07-2.4

                                     Pb([micro]g/g dry
Disease State                             wt.)

Current study
Shell disease (n = 32)                    0.04-3.3
                                        (0.22 + 0.51)
No apparent disease                       0.04-0.72
(n = 25)                             (0.21 [+ or -] 0.18)
Other studies
Boston Harbor([dagger])                   0.51-0.85
Long Island Sound([double dagger])         1.2-15
Outfall (GOC)([subsection])               350 + 77
Maine([paragraph)                         0.02-0.46

* Concentrations are the range from minimum to maximum, and
mean [+ or -] SD are included in parentheses.

([dagger]) From Gardner and Pruell (1987).

([double dagger] From Olsen (1989, 1990).

([subsection]) From Morales-Hernandez et al. (2004).

([paragraph] From Maine Department of Environmental Protection, 2007.
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