Effects of field-contaminated sediments and related water soluble components on haemocyte function and Perkinsus marinus susceptibility and expression in oysters

This paper reviews and discusses our recent findings on the effects of contaminated sediments (CSs) and related water-soluble fractions (WSFs) on haemocyte function/activity and the onset and progression of an infectious disease caused by the protozoan parasite, Perkinsus marinus (Dermo) in the eastern oyster, Crassostrea virginica. Sediments used to generate WSFs and sediments used for the whole CS exposure experiments were collected in different areas of the southern branch of the Elizabeth River, a heavily polluted sub-estuary of the Chesapeake Bay, USA. The WSFs were dominated by low molecular weight polycyclic aromatic hydrocarbons (PAHs). The CSs used for whole CS exposure experiment had elevated concentrations of high molecular weight PAHs. Polychlorinated biphenyls (PCBs) and metals were also present in the CSs. No PCBs were detected in the WSFs. In vitro exposure of haemocytes to WSFs derived from CSs reduced to haemocytes' chemotaxic, phagocytic, and chemiluminescent responses to some extent. Exposure of oysters to suspended CSs stimulated neutral red uptake, mitochondrial dehydrogenase production and ³H-leucine incorporation in haemocytes. Exposure of oysters to 0, 15, 30% WSFs increased the oysters' susceptibility to laboratory-induced infection caused by P. marinus. Exposure of oysters to 15, and 30% dilutions of WSFs for 33 days or to 1.0, 1.5, and 2.0g CSs for 30 days significantly elevated the expression/progression of latent P. marinus infection in oysters in a dose-dependent manner.     

The chemiluminescence response of bivalve haemocytes: utility in screening for immunomodulators and as a biomarker

Resistance to infectious diseases in bivalves depends primarily on the vigour and efficacy of haemocyte-dependent antimicrobial defence mechanisms. Like other phagocytes, haemocytes seem to rely on oxygen-independent (lysosomal hydrolases, lysozyme) and oxygen-dependent (reactive oxygen species) mechanisms to destroy ingested microorganisms. The generation of cytotoxic oxyradicals by haemocytes can be precisely quantified by means of a simple chemiluminescence (CL) assay using luminol or other CL probes. Tributyltin (TBT), and other environmental contaminants, at sublethal levels, will produce dose-dependent suppression of CL activity in haemocytes exposed in short-term, in vitro assays. Presumably, this suppression would find expression as impaired host defence capability. In fact, TBT has been shown to exacerbate progression and lethality of Perkinsus marinus infections in the oyster, Crassostrea virginica. This suggests that CL assays on haemocytes exposed in vitro to single agents or complex mixtures might be useful in screening for aquatic immunomodulators. Statistically significant alterations in CL responses of haemocytes withdrawn from bivalves exposed to xenobiotics in the laboratory or field are more difficult to identify because of high inter-animal variation; however, the use of haemocyte CL as a biomarker of effect merits further investigation.     

Genotoxic damage in Solea senegalensis exposed to sediments from the Sado Estuary (Portugal): Effects of metallic and organic contaminants

Juvenile Solea senegalensis (Senegalese sole) were exposed to freshly collected sediments from three sites of the Sado Estuary (West-Portuguese coast) in 28-day laboratory assays in order to assess the ecological risk from sediment contaminants, by measuring two genotoxicity biomarkers in peripheral blood: the percentage of Erythrocyte Nuclear Abnormalities (ENA) by use of an adaptation of the micronucleus test, and the percentage of DNA strand-breakage (DNA-SB) with the Comet assay. Sediments were surveyed for metallic (Cr, Ni, Cu, Zn, As, Cd and Pb) and organic (PAHs (polycyclic aromatic hydrocarbons), PCBs (polychlorinated biphenyls) and DDTs (dichloro-diphenyl-trichloroethane)) contaminants. Sediments from site A (farthest from hotspots of contamination) were found to be the least contaminated and weaker inducers of genotoxic damage, whereas sediments from sites B (urban influence) and C (affected by industrial effluents and agricultural runoffs) were responsible for a very significant increase in both ENA and DNA-SB, site B being most contaminated with metals and site C mainly with organic pollutants, especially PAHs and PCBs . Analysis of genotoxic effects showed a strong correlation between the concentrations of PAHs and PCBs and both biomarkers at sampling times T₁₄ and T₂₈, while the amounts of Cu, As, Cd and Pb were less strongly correlated, and at T₂₈ only, with ENA and DNA-SB. These results show that organic contaminants in sediment are stronger and faster acting genotoxic stressors. The results also suggest that metals may have an inhibitory effect on genotoxicity when interacting with organic contaminants, at least during early exposure. ENA and DNA-SB do not show a linear relationship, but a strong correlation exists between the overall increase in genotoxicity caused by exposure to sediment, confirming that they are different, and possibly non-linked effects that respond similarly to exposure. Although the Comet assay showed enhanced sensitivity, the two analyses are complementary and suitable for the biomonitoring of sediment contaminants in a benthic species like S. senegalensis.     

Landscape-Level Variation in Disease Susceptibility Related to Shallow-Water Hypoxia

Diel-cycling hypoxia is widespread in shallow portions of estuaries and lagoons, especially in systems with high nutrient loads resulting from human activities. Far less is known about the effects of this form of hypoxia than deeper-water seasonal or persistent low dissolved oxygen. We examined field patterns of diel-cycling hypoxia and used field and laboratory experiments to test its effects on acquisition and progression of Perkinsus marinus infections in the eastern oyster, Crassostrea virginica, as well as on oyster growth and filtration. P. marinus infections cause the disease known as Dermo, have been responsible for declines in oyster populations, and have limited success of oyster restoration efforts. The severity of diel-cycling hypoxia varied among shallow monitored sites in Chesapeake Bay, and average daily minimum dissolved oxygen was positively correlated with average daily minimum pH. In both field and laboratory experiments, diel-cycling hypoxia increased acquisition and progression of infections, with stronger results found for younger (1-year-old) than older (2-3-year-old) oysters, and more pronounced effects on both infections and growth found in the field than in the laboratory. Filtration by oysters was reduced during brief periods of exposure to severe hypoxia. This should have reduced exposure to waterborne P. marinus, and contributed to the negative relationship found between hypoxia frequency and oyster growth. Negative effects of hypoxia on the host immune response is, therefore, the likely mechanism leading to elevated infections in oysters exposed to hypoxia relative to control treatments. Because there is considerable spatial variation in the frequency and severity of hypoxia, diel-cycling hypoxia may contribute to landscape-level spatial variation in disease dynamics within and among estuarine systems.     

In situ phytoremediation of PAH- and PCB-contaminated marine sediments with eelgrass (Zostera marina)

In view of the fact that there are presently no cost-effective in situ treatment technologies for contaminated sediments, a 60-week-long phytoremediation feasibility study was conducted in seawater-supplied outdoor ponds to determine whether eelgrass (Zostera marina) is capable of removing polynuclear aromatic hydrocarbons (PAHs) and polychlorinated biphenyls (PCBs) from submerged marine sediments. It was determined that all PAHs and PCBs, independent of the number of aromatic rings and degree of chlorination, respectively, were removed to a much larger extent in planted sediments compared to unplanted controls. After 60 weeks of treatment, the concentration of total PAHs decreased by 73% in planted sediments but only 25% in unplanted controls. Similarly, total PCBs declined by 60% in the presence of plants while none were removed in the unplanted sediment. Overall, the apparent PAH and PCB biodegradation was greatest in the sediment layer that contained most of the eelgrass roots. Abiotic desorption tests conducted at week 32 confirmed that the phytoremediation process was not controlled by mass-transfer or bioavailability limitations since all PAHs and PCBs desorbed rapidly and to a large extent from the sediment. PAHs were detected in both roots and shoots, with root and shoot bioaccumulation factors for total PAHs amounting to approximately 3 and 1, respectively, after 60 weeks of phytoremediation treatment. Similarly, the root bioaccumulation factor for total PCBs was around 4, while no PCBs were detected in the eelgrass leaves at the end of the experiment. The total mass fraction of PAHs and PCBs absorbed and translocated by plant biomass during the 60-week period was insignificant, amounting to less than 0.5% of the total mass of PAHs and PCBs which was initially present in the sediment. Finally, the number of total heterotrophic bacteria and hydrocarbon degraders was slightly but not statistically significantly greater in planted sediments than in unplanted controls. After ruling out contaminant loss to the water column or absorption and transformation within plant cells, it is most likely that the presence of eelgrass stimulated the microbial biodegradation of PAHs and PCBs in the rhizosphere by releasing root exudates, plant enzymes, or even oxygen. Additional research is needed to further elucidate these potential phytoremediation mechanisms.     

Preparation of chondroitin sulfate libraries containing disulfated disaccharide units and inhibition of thrombin by these chondroitin sulfates

Chondroitin sulfate (CS) containing GlcA-GalNAc(4,6-SO₄) (E unit) and CS containing GlcA(2SO₄)-GalNAc(6SO₄) (D unit) have been implicated in various physiological functions. However, it has been poorly understood how the structure and contents of disulfated disaccharide units in CS contribute to these functions. We prepared CS libraries containing E unit or D unit in various proportions by in vitro enzymatic reactions using recombinant GalNAc 4-sulfate 6-O-sulfotransferase and uronosyl 2-O-sulfotransferase, and examined their inhibitory activity toward thrombin. The in vitro sulfated CSs containing disulfated disaccharide units showed concentration-dependent direct inhibition of thrombin when the proportion of E unit or D unit in the CSs was above 15–17%. The CSs containing both E unit and D unit exhibited higher inhibitory activity toward thrombin than the CSs containing either E unit or D unit alone, if the proportion of the total disulfated disaccharide units of these CSs was comparable. The thrombin-catalyzed degradation of fibrinogen, a physiological substrate for thrombin, was also inhibited by the CS containing both E unit and D unit. These observations indicate that the enzymatically prepared CS libraries containing various amounts of disulfated disaccharide units appear to be useful for elucidating the physiological function of disulfated disaccharide units in CS.     

The effects ofPerkinsus marinusextracellular products and purified proteases on oyster defence parametersin vitro

The effects of extracellular products (ECP) and purified proteases from the protozoan parasitePerkinsus marinuson three host defence parameters (haemocyte motility, lysozyme and haemagglutinin) of the eastern oyster,Crassostrea virginica, were investigated. ECP with high proteolytic activities, as well as purified proteases, significantly decreased the random migration of haemocytes through micro-porous filters in Boyden chambers. Stimulation of haemocyte migration byP. marinuscells orP. marinuscell lysate was also dramatically reduced by ECP and purified proteases. Incubation of oyster plasma with ECP and purified proteases caused a significant decrease in lysozyme activity and also appeared to reduce haemagglutinin titres. These data suggest thatP. marinusECP, as well as the proteolytic fraction of the ECP, can modulate some defence parameters of oystersin vitro.     

Perkinsus marinus in pleasure oyster Crassostrea corteziensis from Nayarit, Pacific coast of México

Culture of the pleasure oyster Crassostrea corteziensis is emerging as an alternative to the Pacific oyster (Crassostrea gigas) for oyster producers, who face severe mortalities since 1997 in Northwest México. For determining the health status of this species, we conducted a histopathological analysis of cultured populations from two estuaries in the Pacific coast of México. Macroscopical analysis revealed animals with transparent and retracted mantle. Histopathological analysis of these specimens showed tissue alterations and parasitic forms consistent with Perkinsus sp. infection. Stages of the parasite identified included tomont and trophozoites with an eccentric vacuole characteristic of Perkinsus spp. Pieces of tissues of infected oysters were incubated in Fluid Thioglycollate Medium (FTM) resulting in blue–black hypnospores after incubation. The identity of the parasite was confirmed by species specific PCR-based assay in DNA samples from oysters, tissue fractions from FTM cultures, and deparaffined samples with Perkinsus-like parasite detected by histology. Sequencing of positive amplified fragments (307 bp) showed a sequence similar to Perkinsus marinus strain TXsc NTS ribosomal RNA gene (100% coverage and 98% identity, GenBank Accession No. AF497479.1) and to P. marinus, Genomic DNA, (100% coverage and 97% identity, GenBank Accession No. S78416.1). The prevalence of P. marinus varied from 1 to 5% in Boca del Camichín and from 1 to 6% in Pozo Chino. In general, the intensity of infection was moderate. The infection was observed in oysters from 31 to 110 mm of shell length. This is the first record of P. marinus in oysters from the North America Pacific coast and the first record in C. corteziensis. The origin of this parasite in the area is unknown, but it may be associated to introductions of Crassostrea virginica from the East coast of United States of America or Gulf of México.     

Spatial and temporal distributions of contaminant body burden and disease in Gulf of Mexico oyster populations: The role of local and large-scale climatic controls

As part of NOAA's Status and Trends Program, oysters were sampled from 43 sites throughout the Gulf of Mexico from Brownsville, Texas, to the Florida Everglades from 1986 to 1989. Oysters were analysed for body burden of a suite of metals and petroleum aromatic hydrocarbons (PAHs), the prevalence and intensity of the oyster pathogen,Perkinsus marinus, and condition index. The contaminants fell into two groups based on the spatial distribution of body burden throughout the Gulf. Arsenic, selenium, mercury and cadmium were characterized by clinal reduction in similarity with distance reminiscent of that followed by mean monthly temperature and precipitation. Zinc, copper, PAHs and silver showed no consistent geographic trend. Within local regions, industrial and agricultural and use andP. marinus prevalence and infection intensity frequently correlated with body burden. Contaminants and biological attributes followed one of three temporal trends. Zinc, copper and PAHs showed concordant shifts over 4 years throughout the eastern and southern Gulf. Mercury and cadmium showed concordant shifts in the northwestern Gulf. Selenium, arsenic, length, condition index andP. marinus prevalence and infection intensity showed concordant shifts throughout most of the entire Gulf. Concordant shifts suggest that climatic factors, the El Niño/Southern Oscillation being one example, exert a strong influence on biological attributes and contaminant body burdens in the Gulf. Correlative factors are those that probably affect or indicate the rate of tissue turnover and the frequency of reproduction; namely, temperature, disease intensity, condition index and length.     

Development of a sediment bioassay to determine bioavailability of PAHs to fish

Recent work has shown that MFO induction and loss of control of steroid hormone production occurs in fish after exposure to pulp mill effluents, PCBs, PAHs, and some pesticides. We had recently developed laboratory assays to evaluate the effluents on these responses, but were lacking a protocol for a sediment assay. This paper describes the development of a sediment test capable of demonstrating MFO induction in fish. MFO responses were evident in rainbow trout within 4 days of exposure to contaminated sediments. Further testing showed that fish were responding to chemicals from the sediments, but not from bottom water, and a survey of sediment from thirteen contaminated areas showed that MFO induction more closely paralleled PAH levels in the sediments than the observed PCB concentrations. The sites showing MFO induction were also the sites where sediment toxicity was demonstrated with laboratory bioassays using Daphnia magna and Hyalella azteca. The protocol has been further refined to describe the quantity of sediment required and duration of testing. This test will enable us to study the biochemical effects of exposure to contaminated sediments. The protocol could also be used to prioritize areas of contamination and to evaluate dredging impacts and remediation success.     

Hydrothermal synthesis, characterization, and KOH activation of carbon spheres from glucose

Carbon spheres (CSs) with controllable sizes and rich in oxygen-containing groups were fabricated using a simple hydrothermal treatment of glucose. The effects of the hydrothermal parameters, including the concentration of glucose, reaction temperature, duration, and the second hydrothermal treatment were investigated. The obtained CSs were then activated using KOH for the eventual preparation of porous carbon spheres. A scanning electron microscope was used to characterize the morphology and size of the CSs. Fourier-transform infrared spectroscopy and X-ray photoelectron spectroscopy were used to analyze the functional surface groups. N₂ adsorption–desorption isotherms were used to analyze the porous structure of the CS. The results revealed that the morphologies and size distribution of the CSs can be controlled by adjusting the experimental parameters. A hydrothermal temperature between 180 and 190 °C over 4–5 h was suitable for CS formation. Under these conditions, the size of the CS increased with the concentration of glucose. Mono-dispersed CSs with good morphologies and large numbers of oxygen-containing functional groups (primarily –OH and CO) can be obtained using a 0.3 mol/L glucose solution that is hydrothermally treated at 190 °C for 4 h. The resulting CSs sizes were about 350 nm in diameter. After a second hydrothermal treatment, the sizes of CSs grew nearly 250 nm without damage to its morphology or broadening of their size distribution. Porous CSs with perfectly spherical shapes and fully developed structures (S_BET = 1282.8 m²/g, V_micro = 0.44 cm³/g) could then be obtained via KOH activation.     

The behavioural response of adult Petromyzon marinus to damage‐released alarm and predator cues

Using semi‐natural enclosures, this study investigated (1) whether adult sea lamprey Petromyzon marinus show avoidance of damage‐released conspecific cues, damage‐released heterospecific cues and predator cues and (2) whether this is a general response to injured heterospecific fishes or a specific response to injured P. marinus. Ten replicate groups of 10 adult P. marinus, separated by sex, were exposed to one of the following nine stimuli: deionized water (control), extracts prepared from adult P. marinus, decayed adult P. marinus (conspecific stimuli), sympatric white sucker Catostomus commersonii, Amazon sailfin catfish Pterygoplichthys pardalis (heterospecific stimuli), 2‐phenylethylamine (PEA HCl) solution, northern water snake Nerodia sipedon washing, human saliva (predator cues) and an adult P. marinus extract and human saliva combination (a damage‐released conspecific cue and a predator cue). Adult P. marinus showed a significant avoidance response to the adult P. marinus extract as well as to C. commersonii, human saliva, PEA and the adult P. marinus extract and human saliva combination. For mobile P. marinus, the N. sipedon washing induced behaviour consistent with predator inspection. Exposure to the P. pardalis extract did not induce a significant avoidance response during the stimulus release period. Mobile adult female P. marinus showed a stronger avoidance behaviour than mobile adult male P. marinus in response to the adult P. marinus extract and the adult P. marinus extract and human saliva combination. The findings support the continued investigation of natural damage‐released alarm cue and predator‐based repellents for the behavioural manipulation of P. marinus populations in the Laurentian Great Lakes.     

Establishing the Causal Relationship between Polycyclic Aromatic Hydrocarbon (PAH) Exposure and Hepatic Neoplasms and Neoplasia-Related Liver Lesions in English Sole (Pleuronectes vetulus)

For almost 25 years our laboratory has studied the impact of PAHs and related industrial contaminants on benthic fish, following an interdisciplinary approach involving chemical exposure assessment linked to synoptic detection of various effects at several levels of biological organization. These data demonstrate a cause-and-effect relationship between neoplastic and neoplasia-related liver lesions in English sole, and exposure to PAHs, and to a lesser degree, chlorinated hydrocarbons such as PCBs. In statistical analyses of data from multiple field studies conducted since 1978, exposure to PAHs measured in various compartments has consistently been identified as a highly significant, major risk factor for neoplasms and related lesions in this species, with PCB exposure shown to be a significant, but less consistent and less strong risk factor for these lesions. A cause-and-effect relationship between PAHs and toxicopathic liver lesions in this species is further supported by the experimental induction of toxicopathic lesions identical to those observed in field-collected fish, in sole exposed in the laboratory to model carcinogenic PAHs such as BaP or to PAH-rich extracts of sediments from Eagle Harbor, a severely PAH-contaminated site in Puget Sound. More recent field studies have identified significant associations between hepatic cytochrome P4501A (CYP1A) induction and xenobiotic-DNA adduct formation, and hepatic lesion prevalences in wild subadult English sole. Field studies in Eagle Harbor subsequent to capping of the most PAH-contaminated region of this harbor with clean dredge spoils have shown a decline in exposure to PAHs as assessed by biliary fluorescent aromatic compounds (FACs) and hepatic xenobiotic-DNA adducts. This decline in PAH exposure has been accompanied by a dramatic decline in risk of occurrence of toxicopathic hepatic lesions in English sole from Eagle Harbor. Further, laboratory studies have induced lesions in English sole by injections of extracts from PAHcontaminated sediments. Overall, these findings relating to exposure to PAHs and chlorinated hydrocarbons and the occurrence of hepatic neoplasms and neoplasiarelated lesions in English sole fulfill the classic criteria for causality in epizootiological or ecological risk assessment studies, including: (1) strength of association, (2) consistency of association, (3) specificity of association, (4) toxicological and biological plausibility, (5) temporal sequence/timing (i.e., exposure precedes disease, effect decreases when the cause is decreased or removed), (6) dose-response or biological gradient, and (7) supportive experimental evidence.     

Attractiveness of the maleAcheta domestica calling song to females

Summary FemaleAcheta domestica did not discriminate between pairs of model calling songs (CSs) which differed only in syllable period (SP; Fig. 1). The females selected the louder CS (Fig. 2) or the CS with a faster chirp rate (CR; Fig. 3) when presented with pairs of otherwise identical CSs. A CS with an SP of 50 ms (modal for the male's CS) was preferred when it was 5 dB louder than one with a 60-ms SP while a CS with a 60-ms SP was only consistently chosen when it was 10 dB louder than a CS with a 50-ms SP (Fig. 4). A more intense CS was preferred by the females regardless of whether its CR was faster or slower than that of the CS produced at a lower intensity (Fig. 6). When CSs with SPs of 50 or 60 ms had several different CRs, the females that made a significant choice preferred a CS with a 50-ms SP regardless of whether it was produced at a faster or slower CR (Figs. 7, 8). No significant selection between CSs with 40- and 50-ms SPs resulted when they were produced at different intensities (Fig. 5) or CRs (Fig. 9). Females only significantly chose a CS with a 50-ms SP over those with 40 ms SPs when the 50-ms-SP CS was louder and produced at a different CR (Fig. 10). From these results, it was apparent that SP, intensity, and CR all influenced a female's choice of a CS, and thus the male producing it. However, our results indicate that SP was the most important feature influencing the female's choice and that intensity was more effective than CR.Abbreviations CR chirp rate - CS calling song - POD polar orientation diagram - SP syllable period     

Influence of some environmental variables on the ascorbic acid status of striped mullet, Mugil cephalus Linn., tissues. III. Effects of exposure to oil

Exposure to water-soluble fractions (WSF) of a crude and two fuel oils altered the ascorbic acid (AsA) content of several striped mullet, Mugil cephalus , tissues. Exposure to sublethal concentrations of all three WSFs caused a depletion of AsA reserves in brain, gill, kidney and liver tissues, but not in muscle. There was a marked decline in AsA stores in kidney and gill tissues after only one day of exposure to WSFs of both crude and fuel oils. Liver AsA concentrations were significantly depleted after one week of oil exposure. Brain AsA content was only significantly depleted during chronic exposure to the highest oil concentration (20% WSF). A dose-dependent depletion of AsA reserves in the liver but not in the other tissues was observed one week after a single exposure to 2–20% WSFs of a No. 2 fuel oil. Exposure to 20% WSF of the No. 2 fuel oil caused a 47% decrease in liver AsA content one week later. Hepatic concentrations were still significantly depleted after 15 days, but had returned to control levels 20 days after the initial exposure. The data suggest that the depletion of tissue AsA reserves in fish inhabiting oil-contaminated environments could be sufficient on occasions to lead to AsA deficiency.     

Assessing the Mutagenic Potential of Surface Sediments from Beijing Guanting Reservoir to Salmonella typhimurium

Mutagenicity of organic extracts from Beijing Guanting Reservoir sediments was investigated with TA98 and TA100 of Salmonella typhimurium. TA98 and TA100 were employed to detect frameshift mutation and base-pair substitution mutation, respectively. For TA100, no positive result was found, while TA98 was more sensitive and pro-mutagenic frameshift mutagens were mainly detected in sediments. Sediments from northern and southern Guanting Reservoir were at potential mutagenic risk. No mutagenicity was found in the sediments from the entrance of the tributaries, but strong mutagenicity was observed in the sediments from the outlet of the reservoir. Chemical analysis was also performed, and poor correlation was found between mutagenicity and organochlorine pesticides (OCPs). However, significant positive correlation was found for polycyclic aromatic hydrocarbons (PAHs) (r = 0.603–0.946), which showed that PAHs were dominated for the tested mutagenicity in the sediments. Polychlorinated biphenyls (PCBs) might induce mutagenicity or promote the mutagenicity of other substances.     

The uptake of persistent organic pollutants by plants

In a field experiment, the transfer of polychlorinated biphenyls (PCBs) and polycyclic aromatic hydrocarbons (PAHs) from contaminated soil to maize (Zea mays L.), sunflower (Helianthus annuus), poplar (Populus nigra × P. maximowiczii) and willow (Salix × smithiana) and the distribution of PCB congeners in maize and sunflower was investigated. The former waste incinerator in Hradec Králové (Czech Republic) was chosen for the experiment. Results of plot screening showed heterogenous contamination by PCBs and PAHs. PCB soil contamination was evidently caused by Delor 106 or Aroclor 1260 stocking and PAH contamination by chemicals containing fluoranthene, benzo/b/fluoranthene, phenanthrene and pyrene. Tested plants were planted on a contaminated field site, in soil contaminated with 1530 μg/kg of total PCBs and 0.138 and 3.42 mg/kg of total PAHs. The results show that maize and sunflower roots accumulated the most PCBs from soil. These plants accumulated hexa- and heptachlorobiphenyl congeners more than tri-, tetra-, and pentachlorobiphenyl congeners. Total concentrations of PAHs in tested plants ranged from 0.096 to 1.34 mg/kg. The highest phenanthrene concentration was found in aboveground biomass of sunflower and the highest concentration of pyrene, in maize roots.     

A comparative study of chemical contamination in sediments of East River (Dongjiang) and selected reservoirs in Hong Kong

Abstract

The levels of TKN, orthophosphate, heavy metals and trace organics including total PCBs and PAHs in the sediments of the East River (Dongjiang) and various reservoirs in Hong Kong were chemically analyzed. It was found that the sediments of various sections of Dongjing and the Shenzhen Reservoir were contaminated by heavy metals at slight to medium levels. Generally speaking, the highest levels of Cd, Cu, Ni, Pb and Zn were recorded in the middle and lower tributaries of Dongjaing, particularly those near Tangxia and Guanlan. Slight amount of PCBs, PAHs were also detected in the lower sections of Dongjiang, indicating industrial contamination due to economic growth in the 1980–90s. Comparatively, only slight amounts of heavy metals were detected in the sediments of some Hong Kong reservoirs that store imported Dongjiang water. Moreover, trace organics including PCBs and PAHs were rarely detected in the reservoirs of Hong Kong. Nevertheless, the high acidity in the sediments of Hong Kong reservoirs is noteworthy. Results revealed that the Dongjiang waters were purified to a certain extent before being imported to and stored at the Hong Kong reservoirs. At present, health risks from the drinking water of Hong Kong are not apparent. However, the long-term trend of organic and inorganic contamination in Dongjiang should not be overlooked and remedial actions should be taken as early as possible. Collaboration between the governments of Hong Kong and Mainland China in tackling chemical pollution in drinking water is an example of cross-border cooperation and control measures should be stepped up as soon as possible.     

In vitro interactions between several species of harmful algae and haemocytes of bivalve molluscs

Harmful algal blooms (HABs) can have both lethal and sublethal impacts on shellfish. To understand the possible roles of haemocytes in bivalve immune responses to HABs and how the algae are affected by these cells (haemocytes), in vitro tests between cultured harmful algal species and haemocytes of the northern quahog (= hard clam) Mercenaria mercenaria, the soft-shell clam Mya arenaria, the eastern and Pacific oysters Crassostrea virginica and Crassostrea gigas and the Manila clam Ruditapes philippinarum were carried out. Within their respective ranges of distribution, these shellfish species can experience blooms of several HAB species, including Prorocentrum minimum, Heterosigma akashiwo, Alexandrium fundyense, Alexandrium minutum and Karenia spp.; thus, these algal species were chosen for testing. Possible differences in haemocyte variables attributable to harmful algae and also effects of haemolymph and haemocytes on the algae themselves were measured. Using microscopic and flow cytometric observations, changes were measured in haemocytes, including cell morphology, mortality, phagocytosis, adhesion and reactive oxygen species (ROS) production, as well as changes in the physiology and the characteristics of the algal cells, including mortality, size, internal complexity and chlorophyll fluorescence. These experiments suggest different effects of the several species of harmful algae upon bivalve haemocytes. Some harmful algae act as immunostimulants, whereas others are immunosuppressive. P. minimum appears to activate haemocytes, but the other harmful algal species tested seem to cause a suppression of immune functions, generally consisting of decreases in phagocytosis, production of ROS and cell adhesion and besides cause an increase in the percentage of dead haemocytes, which could be attributable to the action of chemical toxins. Microalgal cells exposed to shellfish haemolymph generally showed evidence of algal degradation, e.g. loss of chlorophyll fluorescence and modification of cell shape. Thus, in vitro tests allow a better understanding of the role of the haemocytes and the haemolymph in the defence mechanisms protecting molluscan shellfish from harmful algal cells and could also be further developed to estimate the effects of HABs on bivalve molluscs in vivo.