Weiqun Lu

Physiological energetics of the thick shell mussel Mytilus coruscus exposed to seawater acidification and thermal stress

Anthropogenic CO₂ emissions have caused seawater temperature elevation and ocean acidification. In view of both phenomena are occurring simultaneously, their combined effects on marine species must be experimentally evaluated. The purpose of this study was to estimate the combined effects of seawater acidification and temperature increase on the energy budget of the thick shell mussel Mytilus coruscus. Juvenile mussels were exposed to six combined treatments with three pH levels (8.1, 7.7 and 7.3)×two temperatures (25 °C and 30 °C) for 14 d. We found that clearance rates (CRs), food absorption efficiencies (AEs), respiration rates (RRs), ammonium excretion rates (ER), scope for growth (SFG) and O:N ratios were significantly reduced by elevated temperature sometimes during the whole experiments. Low pH showed significant negative effects on RR and ER, and significantly increased O:N ratios, but showed almost no effects on CR, AE and SFG of M. coruscus. Nevertheless, their interactive effects were observed in RR, ER and O:N ratios. PCA revealed positive relationships among most physiological indicators, especially between SFG and CR under normal temperatures compared to high temperatures. PCA also showed that the high RR was closely correlated to an increasing ER with increasing pH levels. These results suggest that physiological energetics of juvenile M. coruscus are able to acclimate to CO2 acidification with a little physiological effect, but not increased temperatures. Therefore, the negative effects of a temperature increase could potentially impact the ecophysiological responses of M. coruscus and have significant ecological consequences, mainly in those habitats where this species is dominant in terms of abundance and biomass.

Effect of pH and temperature on antioxidant responses of the thick shell mussel Mytilus coruscus.

This study evaluated the combined effects of seawater pH decrease and temperature increase on the activity of antioxidant enzymes in the thick shell mussel Mytilus coruscus, an ecological and economic bivalve species widely distributed along the East China Sea. Mussels were exposed to three pH levels (8.1, 7.7 and 7.3) and two temperatures (25 °C and 30 °C) for 14 days. Activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX), glutathione (GSH), acid phosphatase (ACP), alkaline phosphatase (AKP) and glutamic-pyruvic transaminase (GPT) were measured in gills and digestive glands after 1, 3, 7 and 14 days of exposure. All enzymatic activities were significantly impacted by pH, temperature. Enzymatic activities at the high temperature were significantly higher than those at the low temperature, and the mussels exposed to pH 7.3 showed significantly higher activities than those under higher pH condition for all enzymes except ACP. There was no interaction between temperature and pH in two third of the measured activities suggesting similar mode of action for both drivers. Interaction was only consistently significant for GPX. PCA revealed positive relationships between the measured biochemical indicators in both gills and digestive glands. Overall, our results suggest that decreased pH and increased temperature induce a similar anti-oxidative response in the thick shell mussel.

Immune toxicity of TiO2 under hypoxia in the green-lipped mussel Perna viridis based on flow cytometric analysis of hemocyte parameters

The combined effects of DO and TiO2 (mixed rutile/anatase phase, 7/3) on immune responses in Perna viridis were examined. Mussels were exposed to six combinations of oxygen levels (hypoxia: 1.5 mg O2l(-1), normoxia: 6.0 mg O2 l(-1)) and TiO2 concentrations (0, 2.5 mg l(-1) and 10 mg l(-1)) for 216 h. Mussels were sampled after 24h, 48h, 120 h and 216 h, and immune parameters of hemocytes, including mortality, phagocytosis, non-specific esterase, ROS production, lysosomal content and total hemocyte count were investigated using flow cytometric assay. Hemocyte mortality was higher under hypoxia than normoxia, and increased with TiO2 concentrations, but no interaction was found between DO and TiO2. Phagocytosis was reduced under hypoxia and decreased with TiO2 exposure, and the interactive effect between time and TiO2 was observed. The percentage of hemocytes showing non-specific esterase activity was lower under hypoxia, and decreased as TiO2 concentration increased with the significant interactive effect of DO and TiO2. ROS production and lysosomal content were lower under hypoxia and reduced as concentration of TiO2 increased, and interactive effect of DO and TiO2 on ROS was evident. THC was significantly affected by the interactive effect between TiO2 and DO, with higher values under normoxia in the presence of TiO2. The present study demonstrated that immune functions of P. viridis were influenced by both nano-TiO2 and hypoxia with some synergistic effects between the two stressors. This implies that DO has to be considered in the evaluation of the toxicity of nano-materials to bivalves.

Hemocyte responses of the thick shell mussel Mytilus coruscus exposed to nano-TiO2 and seawater acidification

With increasing production from nanotechnology industries, nanomaterials are inevitably released into the aquatic environment, thereby posing a potential risk to aquatic organisms. Thus, concerns have been raised on the potential ecotoxicological effect of nanoparticle. Furthermore, the ecotoxicological consequences caused by the interaction of nanoparticles with other environmental stresses, such as seawater acidification on marine animals, have not been evaluated. In particular, whether acidification enhances the susceptibility to nanoparticles in bivalves needs to be evaluated. In the present study, we investigated the combined effects of low pH and nanoscale titanium dioxide (nano-TiO2) on some immune parameters of hemocytes in the mussel Mytilus coruscus by flow cytometry under six combinations of two pH values (7.3 and 8.1) and three nano-TiO2 concentrations (0, 2.5, and 10mgL-1) for 14 d. Afterward, the mussels were shifted to normal conditions without nano-TiO2 at pH 8.1 for 7 d further to test their recovery from the multiple stresses. Total hemocyte count (THC), phagocytosis (Pha), esterase (Est), and lysosomal content (Lyso) decreased under low pH and high nano-TiO2 concentration conditions, whereas hemocyte mortality (HM) and reactive oxygen species (ROS) increased with nano-TiO2 concentrations under low pH conditions. The interactive effects between pH and nano-TiO2 were observed at the latter part of the exposure experiment (7 and 14 d) in most hemocyte parameters. Nano-TiO2 influenced the immune functions of mussel more severely than low pH. Slight recovery from the combined stresses was observed for HM, THC, Pha, and Lyso, but significant carry-over effects of nano-TiO2 and low pH were still observed. This study demonstrated that both low pH and high concentration of nano-TiO2 had negative effects on mussels, and these effects still acted for some time even though the mussels were already out of such stressors.

Chronic hypoxia and low salinity impair anti-predatory responses of the green-lipped mussel Perna viridis.

The effects of chronic hypoxia and low salinity on anti-predatory responses of the green-lipped mussel Perna viridis were investigated. Dissolved oxygen concentrations ranged from hypoxic to normoxic (1.5 ± 0.3 mg l(-1), 3.0 ± 0.3 mg l(-1) and 6.0 ± 0.3 mg l(-1)), and salinities were selected within the variation during the wet season in Hong Kong coastal waters (15‰, 20‰, 25‰ and 30‰). The dissolved oxygen and salinity significantly affected some anti-predatory responses of mussel, including byssus production, shell thickness and shell weight, and the adductor diameter was only significantly affected by salinity. Besides, interactive effects of dissolved oxygen and salinity on the byssus production and shell thickness were also observed. In hypoxic and low salinity conditions, P. viridis produced fewer byssal threads, thinner shell and adductor muscle, indicating that hypoxia and low salinity are severe environmental stressors for self-defence of mussel, and their interactive effects further increase the predation risk.

Antioxidant response of the hard shelled mussel Mytilus coruscus exposed to reduced pH and oxygen concentration

Ocean acidification (OA) and hypoxic events are increasing worldwide problems, their interactive effects have not been well clarified, although their co-occurrence is prevalent. The East China Sea (the Yangtze River estuary area) suffers from not only coastal hypoxia but also pH fluctuation, representing an ideal study site to explore the combined effect of OA and hypoxia on marine bivalves. We experimentally evaluated the antioxidant response of the mussel Mytilus coruscus exposed to three pH levels (8.1, 7.7 and 7.3) at two dissolved oxygen (DO) levels (2.0mgL-1 and 6.0mgL-1) for 72h. Activities of superoxide dismutase, catalase, glutathione peroxidase, acid phosphatase, and alkaline phosphatase and levels of malondialdehyde were measured in gills and hemolymph. All enzymatic activities in hemolymph and gills followed a similar pattern throughout the experiment duration. Generally, low DO showed greater effects on enzyme activities than elevated CO2. Significant interactions between DO, pH and time were only observed at superoxide dismutase and catalase in both tissues. PCA revealed positive relationships between most enzyme activities in both gills and hemolymph with the exception of alkaline phosphatase activity and the level of malondialdehyde in the hemolymph. Overall, our results suggested that decreased pH and low DO induced similar antioxidant responses in the hard shelled mussel, and showed an additive effect on most enzyme activities. The evaluation of multiple environmental stressors, a more realistic scenario than single ones, is crucial to predict the effect of future global changes on coastal species and our results supply some insights on the potential combined effects of reduced pH and DO on marine bivalves.

Daily rhythms of urotensin I and II gene expression and hormone secretion in the caudal neurosecretory system of the euryhaline flounder (Platichthys flesus).

The caudal neurosecretory system (CNSS) is a unique neuroendocrine structure for environmental adaptation in fish, and is the major site of expression and secretion of urotensin I (UI) and II (UII). This study examined daily changes in mRNA expression and the secretion profile of UI and UII in the CNSS. Daily rhythms were observed in mRNA level of CNSS UI, urophysis UI, plasma UII, glucose, potassium and sodium. No statistically significant (Cosinor, P>0.05) diel rhythmicity in mRNA level of CNSS UII, urophysis UII, cortisol, lactate, osmolality and chloride were detected. The calculated acrophase of sodium, cortisol, plasma UII, urophysis UII, urophysis UI and mRNA level of CNSS UI rhythms were recorded at 13:04 h, 13:39 h, 14:45 h, 15:27 h, 14:41 h and 14:39 h, respectively and a positive relationship was evident among them. The acrophase of glucose and potassium rhythms were recorded at 18:57 h and 22:35 h, respectively. The glucose levels increased progressively at the onset of the UII surge at 15:00 h and reached peak values at dusk. The results support the hypothesis that the CNSS may play a role in the control of co-ordinated daily changes in energy mobilization, nutritional behavior and osmoregulatory systems in euryhaline flounder. Our findings described for the first time the existence of daily rhythms of CNSS hormone expression and secretion in Platichthys flesus. These results reveal the importance of taking into account the time of day when assessing stress responses and evaluating UI and UII as physiological indicators of stress in this species.

CO2-induced pH reduction increases physiological toxicity of nano-TiO2 in the mussel Mytilus coruscus.

The increasing usage of nanoparticles has caused their considerable release into the aquatic environment. Meanwhile, anthropogenic CO2 emissions have caused a reduction of seawater pH. However, their combined effects on marine species have not been experimentally evaluated. This study estimated the physiological toxicity of nano-TiO2 in the mussel Mytilus coruscus under high pCO2 (2500–2600 μatm). We found that respiration rate (RR), food absorption efficiency (AE), clearance rate (CR), scope for growth (SFG) and O:N ratio were significantly reduced by nano-TiO2, whereas faecal organic weight rate and ammonia excretion rate (ER) were increased under nano-TiO2 conditions. High pCO2 exerted lower effects on CR, RR, ER and O:N ratio than nano-TiO2. Despite this, significant interactions of CO2-induced pH change and nano-TiO2 were found in RR, ER and O:N ratio. PCA showed close relationships among most test parameters, i.e., RR, CR, AE, SFG and O:N ratio. The normal physiological responses were strongly correlated to a positive SFG with normal pH and no/low nano-TiO2 conditions. Our results indicate that physiological functions of M. coruscus are more severely impaired by the combination of nano-TiO2 and high pCO2.

Conflicting Effects of Predator Cue and Ocean Acidification on the Mussel Mytilus coruscus Byssus Production

ABSTRACT Understanding the impact of ocean acidification and warming on communities and ecosystems is a researcher priority. This can only be achieved through a combination of experimental and field approaches that would allow developing a mechanistic understanding of impacts across level of biological organizations. Surprisingly, most published studies are still focusing on single species responses with little consideration for interspecific interactions. In this study, the impacts of a 3 days exposure to three parameters (temperature, pH, and presence/absence of the predator cue of the crab Charybdis japonica) and their interactions on an ecologically important endpoint were evaluated: the byssus production of the mussel Mytilus coruscus. Tested temperatures (25°C and 30°C) were within the present range of natural variability whereas pH (8.1, 7.7, and 7.4) covered present as well as near-future natural variability. As expected, the presence of the crab cue induced an antipredator response in Mytilus coruscus (significant 10% increase in byssus secretion rate, 22% increase in frequency of shed byssus, and 30% longer byssus). Decreased pH but not temperature had a significant negative impact on the same endpoints (up to a 17% decrease in byssus secretion rate, 40% decrease in frequency of shed byssus, and 10% shorter byssus at pH 7.3 as compared with pH 8.1) with no significant interactions between the three tested parameters. In this study, it has been hypothesized that pH and predator cue have different modes of action and lead to conflicting functional responses (escape response versus stronger attachment). Functional consequences for ecosystem dynamics still need to be investigated.

Defense Responses to Short-term Hypoxia and Seawater Acidification in the Thick Shell Mussel Mytilus coruscus

The rising anthropogenic atmospheric CO2 results in the reduction of seawater pH, namely ocean acidification (OA). In East China Sea, the largest coastal hypoxic zone was observed in the world. This region is also strongly impacted by ocean acidification as receiving much nutrient from Changjiang and Qiantangjiang, and organisms can experience great short-term natural variability of DO and pH in this area. In order to evaluate the defense responses of marine mussels under this scenario, the thick shell mussel Mytilus coruscus were exposed to three pH/pCO2 levels (7.3/2800 μatm, 7.7/1020 μatm, 8.1/376 μatm) at two dissolved oxygen concentrations (DO, 2.0, 6.0 mg L−1) for 72 h. Results showed that byssus thread parameters, such as the number, diameter, attachment strength and plaque area were reduced by low DO, and shell-closing strength was significantly weaker under both hypoxia and low pH conditions. Expression patterns of genes related to mussel byssus protein (MBP) were affected by hypoxia. Generally, hypoxia reduced MBP1 and MBP7 expressions, but increased MBP13 expression. In conclusion, both hypoxia and low pH induced negative effects on mussel defense responses, with hypoxia being the main driver of change. In addition, significant interactive effects between pH and DO were observed on shell-closing strength. Therefore, the adverse effects induced by hypoxia on the defense of mussels may be aggravated by low pH in the natural environments.