The combined effects of oxygen availability and salinity on physiological responses and scope for growth in the green-lipped mussel Perna viridis

Mussels were maintained for 4 weeks under different combinations of dissolved oxygen concentration (1.5, 3.0 and 6.0 mg O2 l(-1)) and salinity (15, 20, 25 and 30) in a 3×4 factorial design experiment. Clearance rate (CR), absorption efficiency (AE), respiration rate (RR) and scope for growth (SFG) decreased with decreasing salinity and dissolved oxygen concentration (DO), while excretion rate (ER) increased with decreasing salinity and increasing DO. The O:N ratio was <10 at salinities of 15 and 20, irrespective of DO levels. SFG was negative in most of the treatments, except for those under 6.0 mg O2 l(-1) or at a salinity of 30 when DO was lower. The results may help explain the distribution pattern of Perna viridis in Hong Kong waters and provide guidelines for mussel culture site selection.     

Immune responses to combined effect of hypoxia and high temperature in the green-lipped mussel Perna viridis

Flow cytometry was used to examine immune responses in haemocytes of the green-lipped mussel Perna viridis under six combinations of oxygen level (1.5 mg O2 l(-1), 6.0 mg O2 l(-1)) and temperature (20 °C, 25 °C and 30 °C) at 24 h, 48 h, 96 h and 168 h. The mussels were then transferred to normoxic condition (6.0 mg O2 l(-1)) at 20 °C for further 24 h to study their recovery from the combined hypoxic and temperature stress. Esterase (Est), reactive oxygen species (ROS), lysosome content (Lyso) and phagocytosis (Pha) were reduced at high temperatures, whereas hypoxia resulted in higher haemocyte mortality (HM) and reduced phagocytosis. For HM and Pha, changes were observed after being exposed to the stresses for 96 h, whereas only a 24 h period was required for ROS and Lyso, and a 48 h one for Est. Recovery from the stresses was observed for HM and Pha but not other immune responses.     

Immune responses to combined effect of hypoxia and high temperature in the green-lipped mussel Perna viridis

Flow cytometry was used to examine immune responses in haemocytes of the green-lipped mussel Perna viridis under six combinations of oxygen level (1.5 mg O₂ l⁻¹, 6.0 mg O₂ l⁻¹) and temperature (20 °C, 25 °C and 30 °C) at 24 h, 48 h, 96 h and 168 h. The mussels were then transferred to normoxic condition (6.0 mg O₂ l⁻¹) at 20 °C for further 24 h to study their recovery from the combined hypoxic and temperature stress. Esterase (Est), reactive oxygen species (ROS), lysosome content (Lyso) and phagocytosis (Pha) were reduced at high temperatures, whereas hypoxia resulted in higher haemocyte mortality (HM) and reduced phagocytosis. For HM and Pha, changes were observed after being exposed to the stresses for 96 h, whereas only a 24 h period was required for ROS and Lyso, and a 48 h one for Est. Recovery from the stresses was observed for HM and Pha but not other immune responses.     

Size effects of suspended particles on gill damage in green-lipped mussel Perna viridis

The green-lipped mussels Perna viridis were exposed to <500 μm suspended solids (SS) with concentrations of 0 (control), 250, 500, 750 and 1000 mg/L for 14 days, followed by transferring to clean, filtered seawater for 28 days. Results of scanning microscopy showed significantly higher damages to the cilia on the frontal surface of the gill filaments than that on the abfrontal surface in both demibranchs. Percent ciliary depletion varied with SS concentrations and time. No sign of recovery of the gill filaments was observed after the mussels were transferred to clean seawater. In a second experiment, mussels were exposed to SS with size range from <63, >125–<250 and >250–<500 μm at 600 mg/L, together with a control (0 mg/L) for 14 days, followed by transferring to clean, filtered seawater for 28 days. Results of scanning microscopy showed significant ciliary damages in both the ascending and descending lamellae under the three particle size groups as compared with the control. Percent depletion of frontal cilia was most serious for the >250–<500 μm size group and least for the <63 μm size group. However, percent depletion of abfrontal cilia was most serious for the >125–<250 μm size group and least for the <63 μm size group. No recovery of ciliary damages was observed. The effects of particle size of suspended sediments on the morphological damages of gill filaments in the green-lipped mussels were discussed.     

Induction, adaptation and recovery of lysosomal integrity in green-lipped mussel Perna viridis

Biomarkers are generally applied to detect pollution in environmental monitoring. Such biological responses should accurately reflect the stress over time in a quantitative manner. As such, the initial and maximum responses induced by stress, as well as adaptation and recovery of these biomarkers, need to be fully understood or else erroneous false-negative or false-positive may be arrived. However, most of the biomarker studies only provided information on initially induced responses under different concentrations of toxicants, while biological adaptation and recovery were poorly known. In this study, the time required for induction, adaptation and recovery of lysosomal integrity in green-lipped mussel Perna viridis upon exposure to benzo[a]pyrene was investigated over a period of 62 days. Maximum induction occurred on day 6 when lysosomal integrity was significantly reduced by 51%, and no further change or adaptation was detected thereafter. When mussels were depurated in clean seawater after 18 days of exposure to benzo[a]pyrene, a gradual recovery was observed, with lysosomal integrity returning to its background level and showing a complete recovery after 20 days of depuration. Lysosomal integrity was significantly correlated with the body burden concentrations of benzo[a]pyrene and condition index of the mussels. The relatively fast induction (6 days) and recovery (20 days) without apparent adaptation suggested that lysosomal integrity in P. viridis can serve as a good biomarker in biomonitoring, as its response is not likely to generate both false-negative and false-positive results.     

Responses of the green-lipped mussel Perna viridis (L.) to suspended solids

Laboratory experiments were conducted to investigate the lethal and sublethal effects of suspended solids on the survival and physiological, behavourial and morphological changes of the green-lipped mussel Perna viridis collected from Tolo Harbour, Hong Kong. Results showed that P. viridis survived in all test conditions of suspended solids from 0 to 1200 mg/l over a period of 96 h. Physiological responses of the green-lipped mussel under 14-d exposure of suspended solids from 0 to 600 mg/l, followed by 14-d recovery in natural seawater, revealed no significant changes (p>0.05) in oxygen consumption and dry gonosomatic index for treatments in different concentrations of suspended solids and exposure time. Changes in clearance rate were only found to be significant (p<0.001) with exposure time. Responses in behavourial and morphological changes of the green-lipped mussel were also studied under similar experimental treatments and exposure time. Byssus production was significantly (p<0.001) related to exposure time. Gill damage, however, was significantly greater in treatments (p<0.001). Present findings suggested that P. viridis could tolerate a high level of suspended solids in the laboratory. There were dose-dependent effects of suspended solids on morphology of gill filaments. Implications of survival and responses of the green-lipped mussel to suspended solids in the marine environment are discussed.     

Effect of size, tissue parts and location on six biochemical markers in the green-lipped mussel, Perna viridis

The effect of size and tissue parts as well as site locations on a suite of six anti-oxidative biochemical markers, viz. the enzyme degrading neural transmitters acetylcholinesterase (AChE), the Phase II transforming enzyme glutathione-S-transferase (GST), the anti-oxidative enzymes superoxide dismutase (SOD) and catalase (CAT), the lipid metabolite malondialdehyde (MDA) and the ratio of CAT/MDA, in the bivalve mollusc, Perna viridis, was studied. Field mussels were collected from the piers of University of Science and Technology (UST), Pak Sha Wan or Hebe Haven (PS) and Sai Kung (SK) in Port Shelter, Hong Kong. Statistically, ANOVA showed that the activity of all six biochemical markers were generally tissue dependent, size independent and location dependent. Tukey multi comparison test indicated that the biochemical activities fell into two groups, with SK the highest activity group in most cases. The significance of these findings in relation to long-term coastal water biomonitoring is discussed.     

Influences of ammonia-nitrogen and dissolved oxygen on lysosomal integrity in green-lipped mussel Perna viridis: laboratory evaluation and field validation in Victoria Harbour, Hong Kong

Lysosomal integrity in mussels has been applied as a biomarker to detect the pollution of trace organics and metals in the natural environments. However, few studies have examined the effects of water quality on the response of lysosomal integrity, in particular total ammonia-nitrogen (TAN) and dissolved oxygen (DO). This study demonstrated that high level of TAN (2.0 mg/l) and low DO (2.5 mg O₂/l) could significantly reduce the lysosomal integrity in green-lipped mussel Perna viridis, respectively by 33% and 38%, whereas the mussel lysosomal integrity decreased by 70% in the combined treatment of TAN and low DO under laboratory conditions after one week. The mussel lysosomal integrity of all treatment groups could return to the control level after a three week recovery period. In the field validation in Victoria Harbour, Hong Kong during an one-year study period, lysosomal integrity in P. viridis identified the cleanest site east to the harbour, where the lowest TAN and highest DO concentrations were found. While lysosomal integrity in mussels seemed not affected by seasonal changes, approximately 40% of the variation of this biomarker could be attributable to the changes in TAN and DO in seawater. In conclusion, the response of the mussel lysosomal integrity can be confounded by both TAN and DO prevailing in the natural environments and thus caution must be exercised in relating the observed changes in lysosomal integrity to any specific pollutant in coastal water quality monitoring studies.     

Biomonitoring of heavy metals and trace organics using the intertidal mussel Perna viridis in Hong Kong coastal waters

This paper presents the results of a 6-year (1998–2003) survey of trace toxics in the intertidal mussel Perna viridis conducted by the Hong Kong Environmental Protection Department. Concentrations of heavy metals and trace organics were measured in the soft bodies of P. viridis collected from five sites in Hong Kong waters, i.e. Wu Kai Sha (Tolo Harbour), Ma Wan (Northwest), Tsim Sha Tsui (Victoria Harbour), Tai Tam (Hong Kong South) and Lamma Island (Southern Waters) in order to establish the spatial patterns of contaminants in mussels. Among the metals analysed, Cd showed a significant concentration gradient in Hong Kong waters. The levels of Cd in P. viridis were significantly higher at Ma Wan as compared to the other sites studied. Ma Wan also had relatively higher concentrations of Pb. Mn concentrations were particularly prominent at Wu Kai Sha. Significantly higher concentrations of Hg and Cu were recorded at Tai Tam and Tsim Sha Tsui. Tai Tam and Wu Kai Sha had higher levels of V; whereas higher Ni concentrations were recorded at Lamma Island and Tai Tam. No clear spatial patterns for Al, As, Cr, Fe and Zn were observed.

Higher concentrations of PAHs in P. viridis were observed around urban centres impacted by sewage discharges (e.g. Tsim Sha Tsui); whereas higher PCB levels were found not only in Tsim Sha Tsui but also in less urbanised areas such as Lamma Island and Tai Tam, suggesting that these may be due to non-sewage related inputs. The study also shows that Northwest and Southern waters are subject to a higher degree of DDT pollution compared with other sites. Of the 17 dioxin compounds analysed, positive data were mostly recorded for two compounds which are of low toxicity (i.e. OCDD and 1,2,3,4,6,7,8-HpCDD) whereas the most toxic congeners (i.e. 2,3,7,8-TCDD and 1,2,3,7,8-PeCDD) were not detected in the 6 years of monitoring. In general, the levels of OCDD in P. viridis were found to be higher in Tai Tam and Lamma Island in Southern Waters of Hong Kong.

This study found that the levels of some highly toxic heavy metals (i.e. Cd, Hg and Pb) in the mussel P. viridis did not exceed the recommended limits for shellfish as food in Hong Kong (i.e. Cd: 2.0 ppm; Hg: 0.5 ppm; Pb: 6.0 ppm wet weight). The levels of As and Ni in P. viridis were also well below the action limits set by the US FDA (i.e. As: 86 ppm; Ni: 80 ppm wet weight). DDT and PCB contaminations in P. viridis were below the concentrations of concern. Compared with data obtained in the 1980s, the current levels of DDTs in P. viridis were 4–16 times lower; whereas Pb concentrations recorded in Tsim Sha Tsui have also been lowered significantly. This is mainly related to reduction in local and regional pollution sources in the past 20 years.     

The use of selected genotoxicity assays in green-lipped mussels (Perna viridis): a validation study in Hong Kong coastal waters

In Hong Kong, discharges of domestic and industrial wastewaters and disposal of contaminated muds into coastal waters have resulted in high levels of persistent organic contaminants in the water column, biota and bottom sediments. Although the acute toxicity of effluents has been tested by government regulators using mortality-based bioassays, the sublethal effects of many of the pollutants, including environmental carcinogens, have not been well studied. The objective of the present work was to examine the efficiency and efficacy of using: (1) DNA strand breaks (comet assay and alkaline agarose gel electrophoresis assay); (2) oxidative DNA damage (levels of 8-hydroxy-2-deoxyguanosine in cellular DNA) and (3) micronuclei (MN) formation as early warning toxicological indicators of potential environmental carcinogen exposure. Green-lipped mussels (Perna viridis) were transplanted to study sites and retrieved after 4, 8, 12, 16 and 30 days. The bioaccumulation of organic pollutants, including the polycyclic aromatic hydrocarbons (PAHs), organochlorine pesticides (OC pesticides) and polychlorinated biphenyls (PCBs) was observed, and compared with biomarker assays. Some significant dose- and time-dependent inductions of the genotoxic biomarkers, including 8-OHdG, DNA strand breaks and micronuclei were observed. In particular, several very strong (r>0.9) and significantly positive relationships were found at all sites during days 0-12. In addition, significant strong relationships between certain organic contaminants (carcinogenic PAHs and 4 common OC pesticides) and micronuclei were detected at the polluted sites, Tsim Sha Tsui and Sai Wan Ho. The results suggest that the use of micronuclei as a biomarker is potentially effective, especially at the more polluted sites. Nonetheless, further research is required to validate their effectiveness under varying field conditions, especially to elucidate responses under lower contaminant loads, interactions between the environmental and physiological factors affecting organisms in differing coastal environments, and changes in biotransformation and DNA repair enzymes in test organisms.     

Comparative effects of the blue green algae Nodularia spumigena and a lysed extract on detoxification and antioxidant enzymes in the green lipped mussel (Perna viridis)

Nodularia spumigena periodically proliferates to cause toxic algal blooms with some aquatic animals enduring and consuming high densities of the blue green algae or toxic lysis. N. spumigena contains toxic compounds such as nodularin and lipopolysaccharides. This current work investigates physiological effects of exposure from bloom conditions of N. spumigena cells and a post-bloom lysis. Biochemical and antioxidative biomarkers were comparatively studied over an acute 3-day exposure. In general, a post-bloom N. spumigena lysis caused opposite physiological responses to bloom densities of N. spumigena. Specifically, increases in glutathione (GSH) and glutathione peroxidase (GPx) and decreases in glutathione S-transferase (GST) were observed from the N. spumigena lysis. In contrast, N. spumigena cell densities decreased GSH and increased GST and lipid peroxidation (LPO) in mussels. Findings also suggest that at different stages of a toxic bloom, exposure may result in toxic stress to specific organs in the mussel.     

Size effects of suspended particles on gill damage in green-lipped mussel Perna viridis

The green-lipped mussels Perna viridis were exposed to <500 μm suspended solids (SS) with concentrations of 0 (control), 250, 500, 750 and 1000 mg/L for 14 days, followed by transferring to clean, filtered seawater for 28 days. Results of scanning microscopy showed significantly higher damages to the cilia on the frontal surface of the gill filaments than that on the abfrontal surface in both demibranchs. Percent ciliary depletion varied with SS concentrations and time. No sign of recovery of the gill filaments was observed after the mussels were transferred to clean seawater. In a second experiment, mussels were exposed to SS with size range from <63, >125–<250 and >250–<500 μm at 600 mg/L, together with a control (0 mg/L) for 14 days, followed by transferring to clean, filtered seawater for 28 days. Results of scanning microscopy showed significant ciliary damages in both the ascending and descending lamellae under the three particle size groups as compared with the control. Percent depletion of frontal cilia was most serious for the >250–<500 μm size group and least for the <63 μm size group. However, percent depletion of abfrontal cilia was most serious for the >125–<250 μm size group and least for the <63 μm size group. No recovery of ciliary damages was observed. The effects of particle size of suspended sediments on the morphological damages of gill filaments in the green-lipped mussels were discussed.     

Okadaic acid, a causative toxin of diarrhetic shellfish poisoning, in green-lipped mussels Perna viridis from Hong Kong fish culture zones: method development and monitoring

Green-lipped mussels (Perna viridis) were collected from seven fish culture zones (FCZs) in Hong Kong and analyzed for okadaic acid (OA). A conventional HPLC method was modified by incorporating a proteinase K digestion step. Results suggest that a higher recovery (2.5 times higher) of OA was obtained from spiked samples after the addition of 1.08 mg proteinase K in comparison with samples incubated without the proteolytic enzyme. For the hepatopancreas (HP) of individual field-collected mussels, the additional digestion step can enhance OA extraction by 3.1 times. Spatial and temporal variations in OA concentrations in the mussels from various FCZs were investigated. The highest concentration of OA in mussel HP samples was 1164.9 ng/g HP wet wt. With respect to OA concentrations in whole mussel tissues from seven sites and four seasonal samplings, the concentrations were between 70.0 and 131.0 ng/g wet wt., which did not exceed the generally recognized international regulatory criteria (>200 ng/g) for OA.     

The larvae of congeneric gastropods showed differential responses to the combined effects of ocean acidification,temperature and salinity

The tolerance and physiological responses of the larvae of two congeneric gastropods, the intertidal Nassarius festivus and subtidal Nassarius conoidalis, to the combined effects of ocean acidification (pCO₂ at 380, 950, 1250 ppm), temperature (15, 30 °C) and salinity (10, 30 psu) were compared. Results of three-way ANOVA on cumulative mortality after 72-h exposure showed significant interactive effects in which mortality increased with pCO₂ and temperature, but reduced at higher salinity for both species, with higher mortality being obtained for N. conoidalis. Similarly, respiration rate of the larvae increased with temperature and pCO₂ level for both species, with a larger percentage increase for N. conoidalis. Larval swimming speed increased with temperature and salinity for both species whereas higher pCO₂ reduced swimming speed in N. conoidalis but not N. festivus. The present findings indicated that subtidal congeneric species are more sensitive than their intertidal counterparts to the combined effects of these stressors.     

Evaluation of the mussel Perna perna as a biomonitor of polycyclic aromatic hydrocarbon (PAH) exposure and effects

The mussel Perna perna was used to monitor and assess areas suspected of oil contamination by transplanting animals from unimpacted to impacted sites and vice versa. The deployment lasted for three months during which total PAH were measured in seawater samples collected every 15 days. PAH concentrations were determined in mussel tissue monthly. The lysosomal neutral red assay and the condition index were used as biomarkers of the adverse biological effects. The PAH concentration in mussels at unimpacted sites, prior to deployment was ca. 100 microg kg(-1) but increased to 300 microg kg(-1) after three months at the contaminated site. Mussels transplanted to the uncontaminated site showed a decrease from 380 to 80 microg kg(-1). Significant changes in lysosomal stability occurred within one month of the beginning of the transposition; in contrast condition indexes were not correlated with the observed changes in PAH concentration.     

Exposure and time dependent DNA strand breakage in hepatopancreas of green-lipped mussels (Perna viridis) exposed to Aroclor 1254, and mixtures of B[a]P and Aroclor 1254

Green-lipped mussels (Perna viridis) were exposed to Aroclor 1254 (0.5, 5 and 50 microgl(-1)) and a mixture of benzo[a]pyrene (B[a]P) and Aroclor 1254 (0.3+0.5 and 3+5 microgl(-1)) for 12 days. On day 0, 1, 3, 6 and 12, the levels of DNA strand breaks in the mussel hepatopancreas were monitored using an alkaline unwinding assay. The results were compared to the findings of a previous study in which the levels of DNA strand breakage in the same species were measured following exposure to various concentrations of B[a]P (0.3, 3 and 30 microgl(-1)). The results indicated that Aroclor 1254 at a concentration 相似文献   

Effects of trace metal and exposure to air on serotonin and dopamine levels in tissues of the mussel Perna perna

We evaluated levels of serotonin (5HT) and dopamine (DOPA) in muscle and digestive glands of the mussel, Perna perna, collected at different times of day; exposed to air for 24 h, followed by re-submersion; and after exposure to different metals. Mussels collected at different periods of day showed little oscillation in 5HT and DOPA levels. Mussels exposed to metals showed significant changes in 5HT and DOPA levels in digestive gland and muscle, as did mussels exposed to air. Our data suggest that analyses of 5HT and DOPA in tissues of mussels could serve as a tool to evaluate the presence and effects of heavy metal contamination in mussels. Care in data interpretation is required, however, since other environmental factors such as exposure of mussels to air (i.e. at low tides) can also cause changes in DOPA and 5HT levels. Additional research is necessary to separate such natural environmental effects from effects of contaminants.