Effects of freshwater release on oyster reef density, reproduction, and disease in a highly modified estuary.

Few estuaries remain unaffected by water management and altered freshwater deliveries. The Caloosahatchee River Estuary is a perfect case study for assessing the impact of altered hydrology on natural oyster reef (Crassostrea virginica) populations. The watershed has been highly modified and greatly enlarged by an artificial connection to Lake Okeechobee. Accordingly, to generate data to support water management recommendations, this study monitored various oyster biometrics over 15 years along the primary salinity gradient. Oyster reef densities were significantly affected by both prolonged high volume freshwater releases creating hyposaline conditions at upstream sites and by a lack of freshwater input creating hypersaline conditions at downstream sites. Low freshwater input led to an increase in disease caused by Perkinsus marinus and predation. Moderate (< 2000 cfs) and properly timed (winter/spring) freshets benefited oysters with increased gametogenesis, good larval mixing, and a reprieve from disease. If high volume freshets occurred in the late summer, extensive mortality occurred at the upstream site due to low salinity. These findings suggest freshwater releases in the late summer, when reproductive stress is at its peak and pelagic larvae are most vulnerable, should be limited to < 2000 cfs, but that longer freshets (1-3 weeks) in the winter and early spring (e.g., December-April) benefit oysters by reducing salinity and lessening disease intensity. Similar strategies can be employed in other managed systems, and patterns regarding the timing of high volume flows are applicable to all estuaries where the management of healthy oyster reefs is a priority.

Evaluation of toxicity of Deepwater Horizon slick oil on spat of the oyster Crassostrea virginica.

The 2010 explosion of the Deepwater Horizon (DWH) oil rig generated the largest marine oil spill in US history with millions of barrels of crude oil released in the Gulf of Mexico (GoM). The eastern oyster, Crassostrea virginica, is an ecologically and economically important species in the northern GoM. Due to its biological characteristics (sessile, filter feeding), juvenile oysters may have been affected. This study investigated the effects of surface-collected DWH oil prepared as high-energy water-accommodated fraction (HEWAF) on the survival of 2-month-old oyster spat, and evaluated the potential impacts of HEWAF on particle clearance rate and spat tissue. Exposure of oysters to a range of oil/HEWAF (0-7-66-147-908-3450 µg tPAH50 (sum of 50 polycyclic aromatic hydrocarbons) L-1) resulted in non-dose-dependent mortalities and reduced clearance rates of algal food (Tisochrysis lutea). A morphometric study of the digestive tubules (DGTs) indicated a dose-dependent response to oil exposure on lumen dilation, on epithelium thinning of the DGT, and a significant change in DGT synchrony (LOEC = 66 µg tPAH50 L-1). This finding suggests that structural changes occurred in the digestive gland of exposed oysters most likely due to an oil-related stress. In addition, histological observations showed that tissues in contact with HEWAF (gills, palp, connective tissue, digestive gland) were adversely impacted at ≥ 7 µg tPAH50 L-1, and exhibited pathological symptoms typical of an inflammatory response (e.g., hemocyte diapedesis and infiltration, syncytia, epithelium sloughing).

Potential impacts of blooms of the toxic dinoflagellate Karenia brevis on the growth, survival and juvenile recruitment of the non-native green mussel Perna viridis in southeastern United States.

Red tide blooms formed by Karenia brevis are frequent along the Gulf coast of Florida and it is unclear what tolerance the green mussel Perna viridis, a recently introduced species to coastal waters, has toward these events. Established populations of P. viridis were monitored along the coastal waters of Estero Bay, Florida before, during and following two consecutive red tide blooms to assess the potential effects on growth, survival and juvenile recruitment. Upon onset of the bloom, growth rates fell from 6 to 10 mm month(-1) (March 2011-November 2011) to less than 3 mm month(-1). In the succeeding years, K. brevis blooms were present, and average growth of individually tagged mussels remained below 3 mm month(-1). During growth monitoring the use of calcein as an internal marker was tested with positive staining results and no observed effect on growth or survival. In March 2012, following the first red tide bloom, a population-wide mortality event was observed. Following this event, increased mortality rates were observed with peaks during onset of the bloom in the fall of 2012 and 2013. Juvenile recruitment was also limited during years in which blooms persisted into the spring spawning period suggesting gamete and/or larval sensitivity to K. brevis. Although it cannot be conclusively determined that the cause of reduced growth and survival is due to red tide events, the parallels observed suggest that K. brevis is a factor in the observed changes in population structure.

Effects of field and laboratory exposure to the toxic dinoflagellate Karenia brevis on the reproduction of the eastern oyster, Crassostrea virginica, and subsequent development of offspring.

Blooms of the brevetoxin-producing dinoflagellate, Karenia brevis, are a recurrent and sometimes devastating phenomenon in the Gulf of Mexico. The eastern oyster, Crassostrea virginica, is exposed regularly to these blooms, yet little is known about the impacts of K. brevis upon this important species. The present study considered the effects of exposure to both a natural bloom and cultured K. brevis on the reproductive development of C. virginica. Oysters had been exposed to a bloom of K. brevis that occurred in Lee County, Florida, from September 2012 through May 2013, during a period of gametogenesis and gamete ripening. Ripe adult oysters were collected from this bloom-exposed site and from a site 200 miles north which was not exposed to any bloom. In addition, responses to two 10-day laboratory exposures of either unripe or ripe adult oysters to whole cells of K. brevis at high bloom concentrations (1000 and 5000cellsmL-1) were determined. Both field- and laboratory-exposed adult oysters accumulated PbTx (attaining ∼22×103ngg-1 and 922ngg-1 PbTx-3 equivalents in the laboratory and the field, respectively), and significant mucal, edematous, and inflammatory features, indicative of a defense response, were recorded in adult tissues in direct contact with K. brevis cells. Laboratory-exposed oysters also showed an increase in the total number of circulating hemocytes suggesting that: (1) new hemocytes may be moving to sites of tissue inflammation, or, (2) hemocytes are released into the circulatory system from inflamed tissues where they may be produced. The area of oyster tissue occupied by gonad (representative of reproductive effort) and reactive oxygen species production in the spermatozoa of oysters exposed to the natural bloom of K. brevis were significantly lower compared to oysters that were not exposed to K. brevis. Additionally, following 10-day exposure of ripe oysters, a significant, 46% reduction in the prevalence of individuals with ripe gametes was obtained in the 5000cellsmL-1K. brevis treatment. Brevetoxin (PbTx) was recorded within the spermatozoa and oocytes of naturally exposed oysters and was estimated to be 18 and 26% of the adult PbTx load, respectively. Larvae derived from gametes containing PbTx showed significantly higher mortalities and attained a smaller larval size for the first 6 days post-fertilization. These negative effects on larval development may be due to the presence of PbTx in the lipid droplets of the oocytes, which is mobilized by the larvae during embryonic and lecithotrophic larval development. Provision of a non-contaminated food source to larvae however, appeared to mitigate the early negative effects of this neonatal PbTx exposure. Results herein show that adult eastern oysters and their offspring are susceptible to exposure to K. brevis. Caution should therefore be exercised when identifying oyster reef restoration areas and in efforts to establish aquaculture in areas prone to red tides.

Susceptibility of gametes and embryos of the eastern oyster, Crassostrea virginica, to Karenia brevis and its toxins.

The bivalve mollusc, Crassostrea virginica, is frequently exposed to blooms of Karenia brevis along the west coast of Florida during periods of spawning and early larval development. A continuous 4-day exposure of gametes and 2-4 cell stage embryos of C. virginica to whole-cell and culture filtrate of K. brevis at 500 and 5000 cells mL(-1), was followed by a 4-day 'recovery' period. Larval growth, percent of normal, abnormal and dead larvae, and the presence of food in the larval gut were measured throughout the exposure period. Results suggest that negative effects mainly occur during embryogenesis and early development. Damage to feeding apparatus/gut may occur during embryonic development or exposure to toxins may act as a feeding deterrent on non-toxic algae. Following 2-h in vitro exposure of gametes, differences in oocyte and sperm cell parameters were investigated using flow cytometry. The reduced sperm viability in the whole-cell 5000 cells mL(-1) treatment suggests the involvement of extracellular brevetoxins (PbTx) and perhaps other harmful, uncharacterized compounds associated with the K. brevis cell membrane. The cumulative effects of reduced sperm viability, fertilization success, embryonic and larval survival, and the near-annual exposure to blooms of K. brevis could cause significant bottlenecks on oyster recruitment.

Uptake and elimination of brevetoxin in the invasive green mussel, Perna viridis, during natural Karenia brevis blooms in southwest Florida.

Perna viridis is a recently introduced species to US coastal waters and have vigorously spread throughout the southeastern seaboard since their invasion. Little information regarding their response to local environmental factors has been reported including responses to the local HAB species, Karenia brevis. This study monitored the tissue toxin concentration of brevetoxins in P. viridis from existing populations throughout two consecutive natural K. brevis blooms. The results showed P. viridis to rapidly accumulate PbTx upon exposure to the bloom, far exceeding the peak tissue concentrations of oysters, Crassostrea virginica, sampled during the same period, 57,653 ± 15,937 and 33,462 ± 10,391 ng g(-1) PbTx-3 equivalent, respectively. Further, P. viridis retained high PbTx concentrations in their tissues post bloom remaining above the regulatory limit for human consumption for 4-5 months, significantly longer than the depuration time of 2-8 weeks for native oyster and clam species. In the second year, the bloom persisted at high cell concentrations resulting in prolonged exposure and higher PbTx tissue concentrations indicating increased bioaccumulation in green mussels. While this species is not currently harvested for human consumption, the threat for post bloom trophic transfer could pose negative impacts on other important fisheries and higher food web implications.

Effects of the red tide dinoflagellate, Karenia brevis, on early development of the eastern oyster Crassostrea virginica and northern quahog Mercenaria mercenaria.

The brevetoxin-producing dinoflagellate, Karenia brevis, adversely affects many shellfish species including the commercially and ecologically important bivalve molluscs, the northern quahog (=hard clam) Mercenaria mercenaria and eastern oyster Crassostrea virginica, in the Gulf of Mexico, USA. This study assessed the effects of exposure of these bivalves to K. brevis during their early development. In separate experiments, embryos of 2-4 cell stage of M. mercenaria and C. virginica were exposed to both whole and lysed K. brevis cells isolated from Manasota Key, Florida. Low bloom concentrations of 500 to 3000 cells mL(-1) were simulated for 96 h. Shell length, percent abnormality (and normality), and percent mortality of resulting larvae were measured. Percentages were recorded after 6, 24, and 96 h of exposure; larval shell length was measured at 24 and 96 h. For both quahogs and oysters, the effects of exposing embryos to K. brevis on all larval responses were generally dose- and time-dependent. Percent mortalities and abnormalities of both clam and oyster embryos increased significantly after only 6h of exposure to whole cells of K. brevis. For clams, these parameters were significantly higher in whole and lysed treatments (at 3000 cells mL(-1)) than in controls. Percent mortalities of oysters were significantly higher in the whole-cell treatment (3000 cells mL(-1)) than under control conditions. After 24h of exposure, mean larval shell length of both bivalve species was significantly reduced relative to controls. This was evident for clam larvae in both the lysed treatment at 1500 cells mL(-1) and in whole and lysed treatments at 3000 cells mL(-1), and for oyster larvae in the lysed treatment at 3000 cells mL(-1). After 96 h, both species exposed to the lysed cell treatment at 3000 cells mL(-1) had significantly smaller larvae compared to those in the control. Overall, lysed cells of K. brevis had a more pronounced effect on shell length, percent abnormality, and mortality in both clams and oysters than did whole cells. Given the fact that blooms of K. brevis overlap with the spawning periods of these two bivalves, and that cells of this naked dinoflagellate are readily lysed by wave action, these results suggest that exposure to K. brevis during the early life history stages of clams and oysters could adversely affect their population recruitment. Further, the presence of whole or lysed cells of K. brevis in hatcheries could have a major negative impact on production.

Flow cytometric characterization of hemocytes of the sunray venus clam Macrocallista nimbosa and influence of salinity variation.

Sunray venus clam Macrocallista nimbosa is a native bivalve mollusc of Florida, USA, currently evaluated as a potential new aquaculture species. Very little is known about the physiology and hemocyte characteristics of this species. Bivalve hemocytes are generally involved in various physiological functions including nutrition, tissue repair, detoxification and immune defense. Understanding hemocytes of M. nimbosa and their response to environmental variations is crucial. In estuarine Florida areas, salinity is probably the most important factor potentially affecting clams physiology since wide variations can occur within few days. In the present work, using flow cytometry, hemocyte types and cellular parameters (oxidative activity, lysosomal content, phagocytosis capacity) were first characterized in sunray venus clams, in relation with endogenous variables (i.e., size, body weight, gender). Clams were then transferred from salinity 30 psu to 18, 21, 25, 30, 35 and 38 psu. After 7 days, impact of salinity variations was determined on hemocyte parameters, along with estimation of physiological status of clams (mortality, valve closure, filtration activity). Hemocytes of sunray venus clam appeared as a unique population, both in terms of morphology (FSC vs. SSC) and intracellular parameters, but displayed high inter-individual variability. Allometric relationship was only described for intracellular oxidative activity. Transfer of clams to 18 psu and, at lower extent, 21 psu resulted in valve closure, mortality and decreased filtration activity. Low salinities resulted in reduction of the number of circulating hemocytes, potentially reflecting infiltration in tissues as part of an inflammatory response or to optimize nutrient distribution. Low salinities also highly impacted hemocytes as depicted by increased cell and lysosomal compartment volumes, decreased phagocytosis capacity as well as increased oxidative stress and mortality. Salinity drops depress physiology and immune defense capacities of sunray venus clams, potentially threatening survival in case of concomitant pathogen encounter or secondary stress.

Bioaccumulation and depuration of brevetoxins in the eastern oyster (Crassostrea virginica) and the northern quahog (= hard clam, Mercenaria mercenaria).

The eastern oyster (Crassostrea virginica) and northern quahog (= hard clam, Mercenaria mercenaria) are two species of economic and ecological significance in east coast waters of the United States and the Gulf of Mexico. Commercial industries for these species, especially within the state of Florida, are significant. The current study was undertaken to build upon the already established body of knowledge surrounding effects of the toxic dinoflagellate Karenia brevis on shellfish, to provide an understanding of the kinetics of brevetoxins within shellfish tissues, and to provide an estimate of brevetoxin retention times in these shellfish after a bloom event. Individual clams and oysters were exposed to the toxic dinoflagellate, K. brevis at a bloom concentration of 5 × 10(5) cells·L(-1) for eight days and then transferred to filtered water for depuration. Individuals were sampled periodically to determine depuration rates. Concentrations of brevetoxins (and/or their metabolites measured as PbTx-3 equivalent) in tissues were determined using an Enzyme Linked Immunosorbent Assay (ELISA). After five days of exposure, brevetoxin levels in tissues of both species reached concentrations well above the regulatory limit of 800 ng g(-1) (Pb-TX3 equivalent). Averaged concentration of brevetoxins in clams was 1000 ng g(-1), while the oysters averaged 1986 ng g(-1). After two weeks of depuration, tissue concentrations in both species were below regulatory levels with clams averaging ~204 ng g(-1) and oysters averaging ~437 ng g(-1). Toxins (or their metabolities) remained detectable in both clams (139 days) and oysters (82 days) for the duration of the experiment.

Reactive oxygen species in unstimulated hemocytes of the pacific oyster Crassostrea gigas: a mitochondrial involvement.

The Pacific oyster Crassostrea gigas is a sessile bivalve mollusc whose homeostasis relies, at least partially, upon cells circulating in hemolymph and referred to as hemocytes. Oyster's hemocytes have been reported to produce reactive oxygen species (ROS), even in absence of stimulation. Although ROS production in bivalve molluscs is mostly studied for its defence involvement, ROS may also be involved in cellular and tissue homeostasis. ROS sources have not yet been described in oyster hemocytes. The objective of the present work was to characterize the ROS sources in unstimulated hemocytes. We studied the effects of chemical inhibitors on the ROS production and the mitochondrial membrane potential (Δψ(m)) of hemocytes. First, this work confirmed the specificity of JC-10 probe to measure Δψ(m) in oyster hemocytes, without being affected by ΔpH, as reported in mammalian cells. Second, results show that ROS production in unstimulated hemocytes does not originate from cytoplasmic NADPH-oxidase, nitric oxide synthase or myeloperoxidase, but from mitochondria. In contrast to mammalian cells, incubation of hemocytes with rotenone (complex I inhibitor) had no effect on ROS production. Incubation with antimycin A (complex III inhibitor) resulted in a dose-dependent ROS production decrease while an over-production is usually reported in vertebrates. In hemocytes of C. gigas, the production of ROS seems similarly dependent on both Δψ(m) and ΔpH. These findings point out differences between mammalian models and bivalve cells, which warrant further investigation about the fine characterization of the electron transfer chain and the respective involvement of mitochondrial complexes in ROS production in hemocytes of bivalve molluscs.

Functional and metabolic characterization of hemocytes of the green mussel, Perna viridis: in vitro impacts of temperature.

The green mussel, Perna viridis, is a bivalve mollusk native to Asia and was recently introduced to Florida, USA. Since its first observation in 1999 in Tampa Bay, Florida, green mussel population has expanded considerably, to reach the Atlantic coast of Florida, Georgia and South Carolina. Most of currently available studies about the ecology and biology of green mussels were performed in the Indian and Pacific oceans. Very recently, it has been suggested that due to a weak low temperature resistance, green mussels might have already reached the Northern edge of their distribution in the USA. However, there is currently an obvious lack of data about the adaptation capacities of Perna viridis to environmental conditions in Florida, especially at the physiological and cellular levels. In the present work, we determined and characterized the populations of circulating hemocytes, and the cellular components of hemolymph involved in various physiological functions, including immunity. Two main populations were characterized, hyalinocytes and granulocytes. Granulocytes accounted for 60% of circulating cells, and displayed higher phagocytic capacities, lysosomal content and basal oxidative metabolism than hyalinocytes. Hemocyte parameters were not influenced by the size of green mussels. In addition, hemocytes were subjected to acute temperature challenges (10, 20 and 30 °C) and their immune-related functions and metabolism analyzed. Our results showed that 10 °C represent a stressful condition for the Floridian green mussels, as depicted by a low phagocytosis capacity and an increase of oxidative metabolism.

Effects of salinity, heavy metals and pesticides on health and physiology of oysters in the Caloosahatchee Estuary, Florida.

The Caloosahatchee Estuary has been exhibiting signs of impaired ecological health due to the extensive hydrological alteration, agricultural land use, and increasing watershed development. This project investigated the responses of the American oyster, Crassostrea virginica at five locations in the Caloosahatchee River in relation to salinity changes, levels of heavy metals, pesticides, and PCBs in the water as well as in the oyster tissue. Individual heavy metal and organochlorine pesticide concentrations in oysters varied significantly between sampling locations and sampling months. PCB concentrations in oyster tissues were below detection limits as were the metal, pesticide and PCB concentrations in water at all the sampling locations. Both heavy metal and pesticide concentrations decreased with increasing distance downstream indicating upstream source of contaminants. The highly pathogenic oyster parasite Perkinsus marinus infection intensity (level) and prevalence (% infected oysters), condition index, spat recruitment, and gonadal index showed a seasonal trend varying with spawning activity and increased downstream. However, juvenile oyster growth was higher at upstream estuarine locations. Oyster responses varied more with seasonal programming (salinity), rather than due to contaminant levels. While significant correlations were noted between some oyster responses and metal concentrations in oyster tissues, overall metal concentrations were low compared to national averages. It appears that oyster health in the Caloosahatchee River is influenced more by freshwater inflow and resulting salinity fluctuations, rather than due to the measured contaminants.

Influence of salinity on prevalence of the parasite Loxothylacus panopaei in the xanthid Panopeus obesus in SW Florida.

This study was conducted to examine the potential influence of salinity, a proxy for freshwater inflow, on the prevalence of the castrator parasite Loxothylacus panopaei on saltmarsh mud crabs Panopeus obesus on SW Florida oyster reefs. Spatial and seasonal patterns of the presence of potential host crabs and the prevalence of the parasite were assessed in the Caloosahatchee, Estero, and Faka Union estuaries. Lift nets (1 m2) containing 5 1 of oyster clusters were deployed on intertidal reefs at 3 sites along the salinity gradient of each estuary. Nets were deployed during 3 seasonally dry and 3 seasonally wet months for a period of 30 d. P. obesus densities tended to increase downstream in higher salinity waters, with crabs being absent from the upper station in the Caloosahatchee during both seasons and absent from the upper station of the Faka Union during wet months. Parasite prevalence was reduced upstream in each estuary during wet months compared to dry months, and for those estuaries that experienced higher relative levels of freshwater inflow. Furthermore, parasite prevalence was positively correlated with the mean salinity of capture of host crabs. Based on the distribution of P. obesus and the above patterns related to salinity, it appears that freshwater inflow and seasonal rains might regulate the prevalence of this parasite in SW Florida by creating spatiotemporal, low salinity refuges for its host.

Purification and characterization of lysozyme from plasma of the eastern oyster (Crassostrea virginica).

Lysozyme was purified from the plasma of eastern oysters (Crassostrea virginica) using a combination of ion exchange and gel filtration chromatographies. The molecular mass of purified lysozyme was estimated at 18.4 kDa by SDS-PAGE, and its isoelectric point was greater than 10. Mass spectrometric analysis of the purified enzyme revealed a high-sequence homology with i-type lysozymes. No similarity was found however between the N-terminal sequence of oyster plasma lysozyme and N-terminal sequences of other i-type lysozymes, suggesting that the N-terminal sequences of the i-type lysozymes may vary to a greater extent between species than reported in earlier studies. The optimal ionic strength, pH, cation concentrations, sea salt concentrations, and temperature for activity of the purified lysozyme were determined, as well as its temperature and pH stability. Purified oyster plasma lysozyme inhibited the growth of Gram-positive bacteria (e.g., Lactococcus garvieae, Enterococcus sp.) and Gram-negative bacteria (e.g., Escherichia coli, Vibrio vulnificus). This is a first report of a lysozyme purified from an oyster species and from the plasma of a bivalve mollusc.

Greater hemocyte bactericidal activity in oysters (Crassostrea virginica) from a relatively contaminated site in Pensacola Bay, Florida.

Bivalve mollusks such as Crassostrea virginica inhabiting polluted estuaries and coastal areas may bioaccumulate high concentrations of contaminants without apparent ill effects. However, changes in putative internal defense activities have been associated with contaminant accumulation in both experimental and long-term field exposures. In an effort to elucidate these relationships, 40 oysters were collected from Bayou Chico (BC) and East Bay (EB) in Pensacola Bay, FL, two estuaries known to differ in the type and magnitude of chemical contaminants present. Oyster tissue concentrations of metals, tri- and dibutyltin (TBT, DBT), polycyclic aromatic hydrocarbons (PAHs) and polychlorinated biphenyls (PCBs) were measured in individual oysters, as were hemocyte counts (HCs), hemocyte bacterial killing indices (KI), serum lysozyme (LYS) and serum protein (PRO) levels. Average HC, KI, LYS and PRO were significantly higher in BC oysters, which also had significantly higher tissue concentrations of total trace metals, butyltins (BTs), PAHs, PCBs, pesticides, and Mn, Cu, Zn and Sn. EB oysters had low organic contaminant levels and no detectable BTs, but significantly higher concentrations of Al, Cr, Fe, Ag, Cd, and Hg. Simple correlation analysis between specific defense measurements and specific chemical analytes showed specific positive relationships that corroborated previous findings in other FL estuaries. Canonical correlation analysis was used to examine relationships between defense measurements and tissue metals using linearly combined sets of variables. Results were also consistent with previous findings-the highest possible canonical correlation was positive: r=0.864, P<0.0019 among canonical variables composed of HC, KI and LYS for defense, and Fe, Cu, Ag, Cd, Sb, Sn, Ni, Pb and Hg for metals.

Stimulation of defense factors for oysters deployed to contaminated sites in Pensacola Bay, Florida.

A positive association between chemical contaminants and defense factors has been established for eastern oysters (Crassostrea virginica) from Florida, but it is unknown whether such factors can be stimulated through short-term exposure to contaminants in the field. Hatchery oysters were deployed at two contaminated sites and one reference site near Pensacola, Florida, during spring and summer in 1998. Putative defense measurements, notably hemocyte count and bactericidal activity, were significantly elevated after 12-week deployment during summer at the most contaminated site. This site exhibited a dramatic increase in chemical concentrations in oyster tissue relative to both the initial concentrations in hatchery oysters and to oysters deployed at the reference site. Hemocyte activity was not stimulated after 16-week deployment of hatchery oysters in spring, despite similar increases in tissue chemical concentrations, so defense activation by short-term exposure may covary with other unmeasured environmental or physiological parameters. Using the converse approach, Pensacola Bay oysters were collected from two contaminated sites and deployed at the reference site for 16 weeks during spring. Results from this converse deployment were ambiguous; serum lysozyme concentrations were reduced for oysters transplanted from both sites, but hemocyte activities were not significantly changed. The principal outcome from this study was the demonstration of enhanced defense activities for oysters upon short-term summer deployment at a contaminated site.

Cellular responses and disease expression in oysters (Crassostrea virginica) exposed to suspended field contaminated sediments.

Exposure of oysters to water soluble fractions derived from field-contaminated sediments (FCS) containing predominantly lower molecular weight organic aromatic compounds, has been previously demonstrated to enhance pre-existing infections caused by the protozoan parasite, Perkinsus marinus (Dermo), and the prevalence of experimentally induced infections. To further explore the role of pollution on the onset and progression of disease, effects of suspended FCS from an estuarine creek in Virginia, USA, dominated by higher molecular weight polycyclic aromatic hydrocarbons (PAHs) on cellular responses and Dermo disease expression in oysters (Crassostrea virginica) were examined. Sediments were collected from a PAH polluted estuarine creek in Virginia, USA. To test effects on cellular response, oysters from Maine were exposed daily to 0, 1.0, 1.5, or 2.0 g suspended FCS (corresponding to 0, 70.2, 105, or 140 microg PAHs, respectively) for 5, 10, 20, and 40 days. Hemocyte activities and plasma lipid, protein and lactate dehydrogenase (LDH) levels were then measured. Exposure stimulated neutral red uptake, MTT reduction, and 3H-leucine incorporation in oyster hemocytes at various exposure times, but did not affect the plasma protein, lipid and LDH levels. To test effects on Dermo expression, oysters from a Dermo enzootic area, with an initial estimated infection prevalence of 39%, were exposed daily to 0, 1.0, 1.5, or 2.0 g suspended FCS (corresponding to 0, 75.0, 113, or 150 microg PAHs, respectively) for 30 days. Exposure enhanced disease expression in oysters. However, no significant change was noted in any measured cellular or humoral parameters.