Pharmaceutical pollution of the world's rivers.

Environmental exposure to active pharmaceutical ingredients (APIs) can have negative effects on the health of ecosystems and humans. While numerous studies have monitored APIs in rivers, these employ different analytical methods, measure different APIs, and have ignored many of the countries of the world. This makes it difficult to quantify the scale of the problem from a global perspective. Furthermore, comparison of the existing data, generated for different studies/regions/continents, is challenging due to the vast differences between the analytical methodologies employed. Here, we present a global-scale study of API pollution in 258 of the world's rivers, representing the environmental influence of 471.4 million people across 137 geographic regions. Samples were obtained from 1,052 locations in 104 countries (representing all continents and 36 countries not previously studied for API contamination) and analyzed for 61 APIs. Highest cumulative API concentrations were observed in sub-Saharan Africa, south Asia, and South America. The most contaminated sites were in low- to middle-income countries and were associated with areas with poor wastewater and waste management infrastructure and pharmaceutical manufacturing. The most frequently detected APIs were carbamazepine, metformin, and caffeine (a compound also arising from lifestyle use), which were detected at over half of the sites monitored. Concentrations of at least one API at 25.7% of the sampling sites were greater than concentrations considered safe for aquatic organisms, or which are of concern in terms of selection for antimicrobial resistance. Therefore, pharmaceutical pollution poses a global threat to environmental and human health, as well as to delivery of the United Nations Sustainable Development Goals.

Metagenomics-Based Microbial Ecological Community Threshold and Indicators of Anthropogenic Disturbances in Estuarine Sediments.

Assessing the impacts of cumulative anthropogenic disturbances on estuarine ecosystem health is challenging. Using spatially distributed sediments from the Pearl River Estuary (PRE) in southern China, which are significantly influenced by anthropogenic activities, we demonstrated that metagenomics-based surveillance of benthic microbial communities is a robust approach to assess anthropogenic impacts on estuarine benthic ecosystems. Correlational and threshold analyses between microbial compositions and environmental conditions indicated that anthropogenic disturbances in the PRE sediments drove the taxonomic and functional variations in the benthic microbial communities. An ecological community threshold of anthropogenic disturbances was identified, which delineated the PRE sediments into two groups (H and L) with distinct taxa and functional traits. Group H, located nearshore and subjected to a higher level of anthropogenic disturbances, was enriched with pollutant degraders, putative human pathogens, fecal pollution indicators, and functional traits related to stress tolerance. In contrast, Group L, located offshore and subjected to a lower level of anthropogenic disturbances, was enriched with halotolerant and oligotrophic taxa and functional traits related to growth and resource acquisition. The machine learning random forest model identified a number of taxonomic and functional indicators that could differentiate PRE sediments between Groups H and L. The identified ecological community threshold and microbial indicators highlight the utility of metagenomics-based microbial surveillance in assessing the adverse impacts of anthropogenic disturbances in estuarine sediments, which can assist environmental management to better protect ecosystem health.

Stereoselective Bioconcentration and Neurotoxicity of Perfluoroethylcyclohexane Sulfonate in Marine Medaka.

Perfluoroethylcyclohexane sulfonate (PFECHS) is an emerging per- and polyfluoroalkyl substance used to replace perfluorooctane sulfonate (PFOS), mainly in aircraft hydraulic fluids. However, previous research indicates the potential neurotoxicity of this replacement chemical. In this study, marine medaka (Oryzias melastigma) was exposed to environmentally relevant concentrations of PFECHS (concentrations: 0, 0.08, 0.26, and 0.91 µg/L) from the embryonic stage for 90 days. After exposure, the brain and eyes of the medaka were collected to investigate the bioconcentration potential of PFECHS stereoisomers and their effects on the nervous systems. The determined bioconcentration factors (BCFs) of PFECHS ranged from 324 ± 97 to 435 ± 89 L/kg and from 454 ± 60 to 576 ± 86 L/kg in the brain and eyes of medaka, respectively. The BCFs of trans-PFECHS were higher than those of cis-PFECHS. PFECHS exposure significantly altered γ-aminobutyric acid (GABA) levels in the medaka brain and disrupted the GABAergic system, as revealed by proteomics, implying that PFECHS can disturb neural signal transduction like PFOS. PFECHS exposure resulted in significant alterations in multiple proteins associated with eye function in medaka. Abnormal locomotion was observed in PFECHS-exposed medaka larvae, which was rescued by adding exogenous GABA, suggesting the involvement of disrupted GABA signaling pathways in PFECHS neurotoxicity.

Aggravating Pollution of Emerging Aryl Organophosphate Esters in Urban Estuarine Sediments of South China.

Emerging aryl organophosphate esters (aryl-OPEs) have been employed as substitutes for organohalogen flame retardants in recent years; however, their environmental occurrence and associated impacts in urban estuarine sediments have not been adequately investigated, impeding regulatory decision-making. Herein, field-based investigations and modeling based on surface sediment and sediment core analysis were employed to uncover the historical pollution and current environmental impacts of aryl-OPEs in the Pearl River Estuary, South China. Our results revealed a substantial increase in aryl-OPE emission, particularly emerging aryl-OPEs, through sediment transport to the estuary since the 2000s. The emerging aryl-OPEs comprised 83% of the total annual input in the past decade, with an average annual input of 155,000 g. Additionally, the emerging-to-traditional aryl-OPE concentration ratios increased with decreasing distance from the shore, peaking in the highly urbanized riverine outlets. These findings indicate that inventories of emerging aryl-OPEs are likely increasing in estuarine sediments and their emissions are surpassing those of traditional aryl-OPEs. Our risk-based priority screening approach indicates that some emerging aryl-OPEs, particularly bisphenol A bis(diphenyl phosphate), can pose a higher environmental risk than traditional aryl-OPEs in estuarine sediments. Overall, our study highlights the importance of recognizing the environmental impacts of emerging aryl-OPEs.

Ocean Stratification Impacts on Dissolved Polycyclic Aromatic Hydrocarbons (PAHs): From Global Observation to Deep Learning.

Ocean stratification plays a crucial role in many biogeochemical processes of dissolved matter, but our understanding of its impact on widespread organic pollutants, such as polycyclic aromatic hydrocarbons (PAHs), remains limited. By analyzing dissolved PAHs collected from global oceans and marginal seas, we found different patterns in vertical distributions of PAHs in relation to ocean primary productivity and stratification index. Notably, a significant positive logarithmic relationship (R2 = 0.50, p < 0.05) was observed between the stratification index and the PAH stock. To further investigate the impact of ocean stratification on PAHs, we developed a deep learning neural network model. This model incorporated input variables determining the state of the seawater or the stock of PAHs. The modeled PAH stocks displayed substantial agreement with the observed values (R2 ≥ 0.92), suggesting that intensified stratification could prompt the accumulation of PAHs in the water column. Given the amplified effect of global warming, it is imperative to give more attention to increased ocean stratification and its impact on the environmental fate of organic pollutants.

Legacy and Emerging Per- and Polyfluoroalkyl Substances in a Subtropical Marine Food Web: Suspect Screening, Isomer Profile, and Identification of Analytical Interference.

The ban/elimination of legacy per- and polyfluoroalkyl substances (PFASs) has led to a dramatic increase in the production and use of various emerging PFASs over the past decade. However, trophodynamics of many emerging PFASs in aquatic food webs remain poorly understood. In this study, samples of seawaters and marine organisms including 15 fish species, 21 crustacean species, and two cetacean species were collected from the northern South China Sea (SCS) to investigate the trophic biomagnification potential of legacy and emerging PFASs. Bis(trifluoromethylsulfonyl)imide was found in seawater via suspect screening (concentration up to 1.50 ng/L) but not in the biota, indicating its negligible bioaccumulation potential. A chlorinated perfluorooctane sulfonate (PFOS) analytical interfering compound was identified with a predicted formula of C14H23O5SCl6- (most abundant at m/z = 514.9373). Significant trophic magnification was observed for 22 PFASs, and the trophic magnification factors of cis- and trans-perfluoroethylcyclohexane sulfonate isomers (1.92 and 2.25, respectively) were reported for the first time. Perfluorohexanoic acid was trophic-magnified, possibly attributed to the PFAS precursor degradation. The hazard index of PFOS was close to 1, implying a potential human health risk via dietary exposure to PFASs in seafood on the premise of continuous PFAS discharge to the SCS.

Insights into the Atmospheric Persistence, Transformation, and Health Implications of Organophosphate Esters in Urban Ambient Air.

Transformation of organophosphate esters (OPEs) in natural ambient air and potential health risks from coexposure to OPEs and their transformation products are largely unclear. Therefore, a novel framework combining field-based investigation, in silico prediction, and target and suspect screening was employed to understand atmospheric persistence and health impacts of OPEs. Alkyl-OPE transformation products ubiquitously occurred in urban ambient air. The transformation ratios of tris(2-butoxyethyl) phosphate were size-dependent, implying that transformation processes may be affected by particle size. Transformation products of chlorinated- and aryl-OPEs were not detected in atmospheric particles, and atmospheric dry deposition might significantly contribute to their removal. Although inhalation risk of coexposure to OPEs and transformation products in urban ambient air was low, health risks related to OPEs may be underestimated as constrained by the identification of plausible transformation products and their toxicity testing in vitro or in vivo at current stage. The present study highlights the significant impact of particle size on the atmospheric persistence of OPEs and suggests that health risk assessments should be conducted with concurrent consideration of both parental compounds and transformation products of OPEs, in view of the nonnegligible abundances of transformation products in the air and their potential toxicity in silico.

Tissue-Specific Uptake, Depuration Kinetics, and Suspected Metabolites of Three Emerging Per- and Polyfluoroalkyl Substances (PFASs) in Marine Medaka.

Restrictions on legacy per- and polyfluoroalkyl substances (PFASs) have led to the widespread use of emerging PFASs. However, their toxicokinetics have rarely been reported. Here, tissue-specific uptake and depuration kinetics of perfluoroethylcyclohexanesulfonate (PFECHS) and 6:2 and 8:2 chlorinated polyfluoroalkyl ether sulfonates (Cl-PFESAs) were studied in marine medaka (Oryzias melastigma). The fish were exposed to these substances for 28 days (0.2 µg/L), followed by a clearance period of 14 days. The depuration constant (kd) of PFECHS [0.103 ± 0.009 day-1 (mean ± standard deviation)] was reported for the first time. Among the six studied tissues, the highest concentrations of 6:2 Cl-PFESA, 8:2 Cl-PFESA, and PFECHS were found in the liver [1540, 1230, and 188 ng (g of wet weight)-1, respectively] on day 28 while the longest residence times were found in the eyes (t1/2 values of 21.7 ± 4.3, 23.9 ± 1.5, and 17.3 ± 0.8 days, respectively). No significant positive correlation was found between the bioconcentration factors of the studied PFASs and the phospholipid or protein contents in different tissues of the studied fish. Potential metabolites of Cl-PFESAs, i.e., their hydrogen-substituted analogues (H-PFESAs), were identified by time-of-flight mass spectrometry. However, the biotransformation rates were low (<0.19%), indicating the poor capacity of marine medaka to metabolize Cl-PFESAs to H-PFESAs.

Occurrence and Trophodynamics of Marine Lipophilic Phycotoxins in a Subtropical Marine Food Web.

Marine lipophilic phycotoxins (MLPs) are produced by toxigenic microalgae and cause foodborne illnesses. However, there is little information on the trophic transfer potential of MLPs in marine food webs. In this study, various food web components including 17 species of mollusks, crustaceans, and fishes were collected for an analysis of 17 representative MLPs, including azaspiracids (AZAs), brevetoxins (BTXs), gymnodimine (GYM), spirolides (SPXs), okadaic acid (OA), dinophysistoxins (DTXs), pectenotoxins (PTXs), yessotoxins (YTXs), and ciguatoxins (CTXs). Among the 17 target MLPs, 12, namely, AZAs1-3, BTX3, GYM, SPX1, OA, DTXs1-2, PTX2, YTX, and the YTX derivative homoYTX, were detected, and the total MLP concentrations ranged from 0.316 to 20.3 ng g-1 wet weight (ww). The mean total MLP concentrations generally decreased as follows: mollusks (8.54 ng g-1, ww) > crustaceans (1.38 ng g-1, ww) > fishes (0.914 ng g-1, ww). OA, DTXs, and YTXs were the predominant MLPs accumulated in the studied biota. Trophic dilution of the total MLPs was observed with a trophic magnification factor of 0.109. The studied MLPs might not pose health risks to residents who consume contaminated seafood; however, their potential risks to the ecosystem can be a cause for concern.

Diversity of gut microbiomes in marine fishes is shaped by host-related factors.

Microorganisms in the gastrointestinal tract of animals play vital roles in food digestion, homeostasis and immune response regulation. Globally, there are 33,700 fish species, representing almost half of all vertebrate diversity and a wide range of physiologies, ecologies and life histories. To investigate gut microbiomes with high coverage, we performed 16S rRNA gene amplicon sequencing with 115 samples of 20 common marine fish species. The fish gut microbiome is a remarkably simple community with low microbial diversity (a maximum of 300 amplicon sequence variants only) and has up to 70% of unknown species in some fish species. The gut microbial community structure was significantly shaped by the combined influence of host-associated factors, including the fish taxon (p < .001, R2  = 0.16, ω2  = 0.04), feeding habit (p < .001, R2  = 0.06, ω2  = 0.02) and trophic level (p < .01, R2  = 0.04, ω2  = 0.01), although the influence was subtle with a small effect size. The core gut microbiomes of different feeding habits were also previously discovered in animal-associated and corresponding habitat samples. Certain energy metabolism pathways were enriched in herbivore/omnivore and zooplanktivore/zoobenthivore fishes, whereas lipid metabolism and glycan metabolism were enriched in zoobenthivore/piscivore fishes. Moreover, substantial taxonomic variability was found between the gut microbiomes of fish and animals, indicated by their low degree of shared microbiota. The data and observations reported herein pave the way for further investigations on the co-evolution of fish gut microbiomes and their hosts, the physiological functions of gut microorganisms and the development of probiotics for improving the nutrition and health of aquaculture fish species.

Acute and chronic toxicity of nickel on freshwater and marine tropical aquatic organisms.

Water quality guidelines and ecological risk assessment of chemical substances like nickel (Ni) in tropical regions such as South East Asia and Melanesia are often based on temperate information as a result of fewer Ni ecotoxicity data available for tropical species. This leaves an unknown margin of uncertainty in the risk assessment in the tropics. In order to fill this data gap, this study was designed to conduct standard toxicity tests on Ni with two freshwater species (acute tests) and three marine species (acute and chronic tests) originated from tropical Hong Kong. All tests were carried out using measured concentrations of Ni with control mortality below 15%. The median lethal concentrations (LC50s) were determined as 2520 (95% confidence interval: 2210, 2860) and 426 (351, 515) µg Ni L-1 for the freshwater gastropods Pomacea lineata (48 h) and Sulcospira hainanensis (96 h), respectively, while 96 h LC50s of 4300 (3610, 5090), 18,200 (6470, 51,200), 62,400 (56,800, 68,500), and 71,700 (68,200, 75,400) µg Ni L-1 were derived for the marine copepod Tigriopus japonicus, the gastropod Monodonta labio, juvenile and adult of the marine fish Oryzias melastigma, respectively. The chronic effect concentration of 10% (EC10) based on the intrinsic rate of increase of the population of T. japonicus was 29 (12, 69) µg Ni L-1. In terms of growth inhibition, the chronic EC10 for M. labio was 34 (17, 67) µg Ni L-1. The results also indicated that T. japonicus in maturation stage (LC10: 484 (349, 919) µg Ni L-1) was less sensitive than its nauplii stage (LC10: 44 (27, 72) µg Ni L-1). This study represents an important addition of high-quality toxicity data to the tropical Ni toxicity database which can be used for future ecological risk assessment of Ni and derivation of its water quality guidelines in tropical regions.

Toxicities of Irgarol 1051 derivatives, M2 and M3, to two marine diatom species.

Irgarol 1051 is highly toxic to marine autotrophs and has been widely used as an antifouling booster biocide. This study tested the toxicities of two s-triazine derivatives of Irgarol, namely M2 (3-[4-tert-butylamino-6-methylthiol-s-triazin-2-ylamino]propionaldehyde) and M3 (2-methylthio-4,6-bis-tert-butylamino-s-triazine) to two marine diatom species, Skeletonema costatum and Thalassiosira pseudonana through standard acute (96h) and chronic (7d) growth inhibition tests. Results showed that both of the two chemicals significantly inhibited the growth of S. costatum (M2: 96h-EC50 = 6789.7 µg L-1, 7d-EC50 = 3503.7 µg L-1; M3: 96h-EC50 = 45193.9 µg L-1, 7d-EC50 = 5330.0 µg L-1) and T. pseudonana (M2: 96h-EC50 = 366.2 µg L-1, 7d-EC50 = 312.5 µg L-1; M3: 96h-EC50 = 2633.4 µg L-1, 7d-EC50 = 710.5 µg L-1), while their toxicity effects were much milder than Irgarol and its major degradation product M1. By comparing with previous findings, the susceptibilities of these s-triazine compounds to two tested species were ranked as: Irgarol > M1 ≫ M2 > M3. This study promotes future research efforts on better understanding of the ecotoxicities of M2 and M3, and incorporating such information to improve the current monitoring, risk assessment and regulation of the use of Irgarol.

Stereoisomer-Specific Trophodynamics of the Chiral Brominated Flame Retardants HBCD and TBECH in a Marine Food Web, with Implications for Human Exposure.

Stereoisomers of 1,2,5,6,9,10-hexabromocyclododecane (HBCD) and 1,2-dibromo-4-(1,2-dibromoethyl)-cyclohexane (TBECH) were determined in sediments and 30 marine species in a marine food web to investigate their trophic transfer. Lipid content was found to affect the bioaccumulation of ΣHBCD and ΣTBECH in these species. Elevated biomagnification of each diastereomer from prey species to marine mammals was observed. For HBCD, biota samples showed a shift from γ- to α-HBCD when compared with sediments and technical mixtures; trophic magnification potential of (-)-α- and (+)-α-HBCD were observed in the food web, with trophic magnification factors (TMFs) of 11.8 and 8.7, respectively. For TBECH, the relative abundance of γ- and δ-TBECH exhibited an increasing trend from abiotic matrices to biota samples; trophic magnification was observed for each diastereomer, with TMFs ranging from 1.9 to 3.5. The enantioselective bioaccumulation of the first eluting enantiomer of δ-TBECH in organisms at higher TLs was consistently observed across samples. This is the first report on the trophic transfer of TBECH in the food web. The estimated daily intake of HBCD for Hong Kong residents was approximately 16-times higher than that for the general population in China, and the health risk to local children was high, based on the relevant available reference dose.

Model for Predicting Toxicities of Metals and Metalloids in Coastal Marine Environments Worldwide.

Metals can pose hazards to marine species and can adversely affect structures and functions of communities of marine species. However, little is known about how structural properties of metal atoms combined with current geographical and climatic conditions affect their toxic potencies. A mathematical model, based on quantitative structure-activity relationships and species sensitivity distributions (QSAR-SSD) was developed by use of acute toxicities of six metals (Cd, Cr, Cu, Hg, Ni, and Zn) to eight marine species and accessory environmental conditions. The model was then used to predict toxicities of 31 metals and metalloids and then to investigate relationships between acute water quality criteria (WQC) and environmental conditions in coastal marine environments. The model was also used to predict WQC in the coastal areas of different countries. Given global climate change, the QSAR-SSD model allows development of WQC for metals that will be protective of marine ecosystems under various conditions related to changes in global climate. This approach could be of enormous benefit in delivering an evidence-based approach to support regulatory decision making in management of metal and metalloids in marine waters.

Tracking Dietary Sources of Short- and Medium-Chain Chlorinated Paraffins in Marine Mammals through a Subtropical Marine Food Web.

Our previous study revealed an elevated accumulation of short-chain chlorinated paraffins (SCCPs) and medium-chain chlorinated paraffins (MCCPs) in marine mammals from Hong Kong waters in the South China Sea. To examine the bioaccumulation potential and biomagnification in these apex predators, we sampled the dietary items of marine mammals and tracked the sources of SCCPs and MCCPs through a marine food web in this region. Sixteen fish species, seven crustacean species, and four mollusk species were collected, and the main prey species were identified for two species of marine mammals. Concentrations of ∑SCCPs and ∑MCCPs in these collected species suggested a moderate pollution level in Hong Kong waters compared to the global range. Lipid content was found to mediate congener-specific bioaccumulation in these marine species. Significantly positive correlations were observed between trophic levels and concentrations of ∑SCCPs or ∑MCCPs (p < 0.05). Trophic magnification factors for ∑SCCPs and ∑MCCPs were 4.29 and 4.79, indicating that both of them have trophic magnification potentials. Elevated biomagnification of SCCPs and MCCPs from prey species to marine mammals was observed. This is the first report of dietary source tracking of SCCPs and MCCPs in marine mammals. The elevated biomagnification between prey and marine mammals raises environmental concerns about these contaminants.

Sub-lethal effects of cadmium and copper on RNA/DNA ratio and energy reserves in the green-lipped mussel Perna viridis.

This study aims to test if RNA/DNA ratio and various energy reserve parameters (i.e., glycogen, lipid, protein content and total energy reserves) are sensitive biomarkers for indicating stresses induced by metal contaminants in the green-lipped mussel Perna viridis, a common organism for biomonitoring in Southeast Asia. This study was, therefore, designed to examine the effects of cadmium (Cd) and copper (Cu) on these potential biomarkers in two major energy storage tissues, adductor muscle (AM) and hepatopancreas (HP), of P. viridis after sub-lethal exposure to either metal for 10 days. The results showed that neither Cd nor Cu treatments affected the RNA/DNA ratio, glycogen and protein contents in AM and HP. As the most sensitive biomarker in P. viridis, the total lipid content in both AM and HP was significantly decreased in the treatment of 5µg Cu/L and 0.01-0.1µgCd/L, respectively. However, soft-tissue body burdens of Cu or Cd did not significantly correlate with each of the four biomarkers regardless of the tissue type. In addition, AM generally stored more glycogen than HP, whereas HP stored more lipids than AM. We proposed that multiple biomarkers may be employed as an integrated diagnostic tool for monitoring the health condition of the mussels.

Toxicity of triphenyltin chloride to the rotifer Brachionus koreanus across different levels of biological organization.

Although triphenyltin (TPT) compounds are ubiquitous pollutants in urbanised coastal environments in Asian regions, their toxicities to marine organisms are still poorly known. This study was designed to investigate the toxicity of triphenyltin chloride (TPTCl) on the rotifer Brachionus koreanus across different levels of biological organisation. Firstly, we concurrently performed a 24 h static-acute toxicity test and a 6-day semi-static multigenerational life-cycle test using the rotifer. Our results demonstrated that the 24-h median lethal concentration of TPTCl for the rotifer was 29.6 µg/L and the 6-day median effect concentration, based on the population growth inhibition, was 3.31 µg/L. Secondly, we examined the expression of 12 heat shock protein (hsp) genes, four glutathione S-transferase (GST) genes, one retinoid X receptor (RXR) gene and 13 cytochrome P450 (CYP) genes in the rotifers after exposure to 20 µg/L TPTCl for 24 h. Among these studied genes, hsp90α2, GST-O and CYP3045C1 were the most significantly up-regulated genes with a relative expression level up to 32.9, 4.4 and 62.6 folds, respectively. The expression of these three genes in the rotifers showed an increasing trend in the first few hours of TPTCl exposure, peaked at 3 h (hsp90α2 and GST-O) and 12 h (CYP3045C1) respectively, and then gradually returned to a lower level at 24 h. Such up-regulations of hsp and GST genes probably offer cellular protection against the TPT-mediated oxidative stress while the accelerated induction of CYP genes possibly facilitates the detoxification of this toxicant in the rotifer.

Temperature-dependent physiological and biochemical responses of the marine medaka Oryzias melastigma with consideration of both low and high thermal extremes.

This study aimed to investigate temperature effect on physiological and biochemical responses of the marine medaka Oryzias melastigma larvae. The fish were subjected to a stepwise temperature change at a rate of 1 °C/h increasing or decreasing from 25 °C (the control) to six target temperatures (12, 13, 15, 20, 28 and 32 °C) respectively, followed by a 7-day thermal acclimation at each target temperature. The fish were fed ad libitum during the experiment. The results showed that cumulative mortalities were significantly increased at low temperatures (12 and 13 °C) and at the highest temperature (32 °C). For the survivors, their growth profile closely followed the left-skewed 'thermal performance curve'. Routine oxygen consumption rates of fish larvae were significantly elevated at 32 °C but suppressed at 13 and 15 °C (due to a high mortality, larvae from 12 °C were not examined). Levels of heat shock proteins and activities of malate dehydrogenase and lactate dehydrogenase were also measured in fish larvae exposed at 15, 25 and 32 °C. The activities of both enzymes were significantly increased at both 15 and 32 °C, where the fish larvae probably suffered from thermal discomfort and increased anaerobic components so as to compensate the mismatch of energy demand and supply at these thermal extremes. Coincidently, heat shock proteins were also up-regulated at both 15 and 32 °C, enabling cellular protection. Moreover, the critical thermal maxima and minima of fish larvae increased significantly with increasing acclimation temperature, implying that the fish could develop some degrees of thermal tolerance through temperature acclimation.

Isoform-specific responses of metallothioneins in a marine pollution biomonitor, the green-lipped mussel Perna viridis, towards different stress stimulations.

Metallothioneins (MTs) are commonly used as biomarker for metal pollution assessment in marine ecosystems. Using integrated genomic and proteomic analyses, this study characterized two types of MT isoform in the digestive gland of a common biomonitor, the green-lipped mussel Perna viridis, towards the challenges of a metal (cadmium; Cd) and a non-metal oxidant (hydrogen peroxide; H2 O2 ) respectively. The two isoforms differed in their deduced protein sequences, with 73 amino acids for MT10-I and 72 for MT10-II (a novel type), but both consisted of a high percentage (27.4 to 29.2%) of cysteine. Two-dimensional gel and Western blot showed that the MT proteins were present in multiple isoform spots, and they were further validated to be MT10-I and MT10-II using MS analysis coupled with unrestricted modifications searching. Expression of mRNA revealed that MT10-I responded promptly to Cd but had a lagged induction to H2 O2 treatments, while MT10-II was exclusively induced by Cd treatment over the course of exposure. Expression of the MT proteins also showed a delayed response to H2 O2 , compared to Cd treatments. This study uncovered the potential different functional roles of various MTs isoforms in P. viridis and thus advances the resolution of using MTs as biomarkers in future applications.