Transcriptomic analysis reveals the endocrine toxicity of tributyltin and triphenyltin on the whelk Reishia clavigera and mechanisms of imposex formation.

Organotin compounds (OTs) are endocrine disruptors that induce imposex in hundreds of gastropods, but little is known about their underlying molecular mechanisms. This study aimed to investigate the endocrine toxicity and molecular responses to tributyltin (TBT) and triphenyltin (TPT) exposure in the whelk Reishia clavigera, which often serves as a biomonitor for OT contamination. Over a 120-day exposure to environmentally relevant concentrations of TBT (1000 ng L-1) and TPT (500 ng L-1), we observed a significant increase in penis length in both male and female whelks. Notably, TPT exhibited a stronger potency in inducing pseudo-penis development and female sterility, even at a half dose of TBT. Bioaccumulation analysis also revealed higher persistence and accumulation of TPT in whelk tissues compared to TBT. Differential expression analysis identified a substantial number of differentially expressed genes (DEGs), with TPT exposure eliciting more DEGs than TBT. Our results demonstrated that OTs induced xenobiotic metabolism and metabolic dysregulation in the digestive gland, impaired multiple cellular functions and triggered neurotoxicity in the nervous system, and disrupted lipid homeostasis and oxidative stress in the gonads. Furthermore, imposex was possibly associated with disturbances in retinoic acid metabolism, nuclear receptor signaling, and neuropeptide activity. When compared to TBT, TPT exhibited a more pronounced endocrine-disrupting effect, attributable to its higher bioaccumulation and substantial interruption of transcriptional regulation, OT detoxification, and biosynthesis of retinoic acids in R. clavigera. Our results, therefore, highlight the importance of considering the differences in bioaccumulation and molecular toxicity between TBT and TPT in future risk assessments of these contaminants. Overall, our study provided molecular insights into the toxicity and transcriptome profiles in R. clavigera exposed to TBT and TPT, shedding light on the endocrine-disrupting effects and reproductive impairment in female gastropods.

Joint effects of temperature and copper exposure on developmental and gene-expression responses of the marine copepod Tigriopus japonicus.

There is growing contamination of copper (Cu) in the marine environment, particularly after the ban of organotin compounds and the increase of the use of Cu-based antifouling paints. Although there are increasing research interests in temperature-dependent chemical toxicity to aquatic organisms, most existing studies focused on acute impacts of chemicals at high concentrations. This study aimed to investigate the interacting effect of temperature and copper exposure at environmentally relevant concentrations on survival and development in the marine copepod Tigriopus japonicus with a partial life-cycle toxicity test. Expressions of five stress response genes in the copepod, namely two glutathione S-transferases (GST-S and GST-O), two heat shock proteins (HSP70 and HSP90), and glutathione reductase (GR) were also investigated. The copepod's survival was significantly impaired at 15 °C after development to adult stage, while its developmental time reduced significantly with increasing temperature. Copper at the two environmentally relevant test concentrations had no significant impacts on these apical endpoints whereas the interaction between Cu and temperature was more significant in modulating gene expressions. GST-S, GST-O and HSP90 genes in copepods exposed to 100 µg Cu L-1 were significantly upregulated at 20 °C. At 32 °C, most genes were either insignificantly expressed or down-regulated, compared to the control, likely suggesting that thermal stress inhibited the copepod's antioxidative defense system. Overall, the results revealed that the joint Cu and thermal stresses have significantly elicited antioxidative system in the copepods. It clearly demonstrated the need for more fundamental studies about potential impacts of different environmental factors such as temperature on chemical toxicity under realistic scenario of marine pollution.

Extreme cold or warm events can potentially exacerbate chemical toxicity to the marine medaka fish Oryzias melastigma.

Marine ecosystems are currently subjected to dual stresses of chemical pollution and climate change. Through a series of laboratory experiments, this study investigated the impact of exposure to chemical contaminant such as DDT or copper (Cu), in combination with cold or warm temperature extremes on the marine medaka fish Oryzias melastigma. The results showed that extreme seawater temperatures (i.e., 15 and 32 °C in sub-tropical Hong Kong) exacerbated adverse chemical impacts on the growth performance of O. melastigma, in particular at the high thermal extreme. This was likely associated with an interruption of oxygen consumption and aerobic scope. Most importantly, the results of acclimation experiments, as reflected by thermal tolerance polygons, showed that chemical exposure substantially narrowed the thermal tolerance of the medaka, making them more vulnerable to temperature changes and extreme thermal events. Under dual stresses of thermal extremes and chemical exposure, the medaka switched their metabolic pathway to anaerobic respiration that might deplete their energy reserve for chemical detoxification. Although stress proteins such as heat shock proteins (HSP90) were up-regulated for cellular protection in the fish, such a defensive mechanism was repressed with intensifying dual stresses at high temperature and high chemical concentration. Bioconcentration of DDT or Cu generally increased with increasing temperature and its exposure concentration. Overall, these complex chemical-temperature interactions concomitantly exerted a concerted adverse impact to O. melastigma. The temperature-dependent toxicity of DDT or Cu shown in this study clearly demonstrated the potential challenge brought by the risk of chemical pollution under the impact of global climate change.

Littoral Water in Hong Kong as a Potential Transient Habitat for Juveniles of a Temperate Deepwater Gnomefish, Scombrops boops (Acropomatiformes: Scombropidae).

A total of 40 juveniles belonging to a temperate deepwater gnomefish species, Scombrops boops, were sampled from littoral habitats (2-5 m depth) of eastern Hong Kong waters in April and May 2017 and March 2019. The presence of gnomefish juveniles in subtropical southern China is reported for the first time at a record low latitude of 22°11'-22°21'N. The specimens were identified based on the COI gene sequence. The genetic composition between Japan and Hong Kong gnomefish populations were compared by sequencing the mitochondrial Cytb gene, which showed no genetic differentiation. The juveniles ranged from 3.5-10.1 cm (n = 40) in total length, with 35 individuals caught from Sargassum beds and five from rocky reefs. Our findings highlighted that the littoral habitats in Hong Kong waters, in particular the seasonal Sargassum beds, are important for small juveniles of S. boops.

Transcriptomics reveal triphenyltin-induced molecular toxicity in the marine mussel Perna viridis.

Triphenyltin (TPT) is widely used as an active ingredient in antifouling paints and fungicides, and continuous release of this highly toxic endocrine disruptor has caused serious pollution to coastal marine ecosystems and organisms worldwide. Using bioassays and transcriptome sequencing, this study comprehensively investigated the molecular toxicity of TPT chloride (TPTCl) to the marine mussel Perna viridis which is a commercially important species and a common biomonitor for marine pollution in Southeast Asia. Our results indicated that TPTCl was highly toxic to adult P. viridis, with a 96-h LC10 and a 96-h EC10 at 18.7 µg/L and 2.7 µg/L, respectively. A 21-day chronic exposure to 2.7 µg/L TPTCl revealed a strong bioaccumulation of TPT in gills (up to 36.48 µg/g dry weight) and hepatopancreas (71.19 µg/g dry weight) of P. viridis. Transcriptome analysis indicated a time course dependent gene expression pattern in both gills and hepatopancreas. Higher numbers of differentially expressed genes were detected at Day 21 (gills: 1686 genes; hepatopancreas: 1450 genes) and at Day 28 (gills: 628 genes; hepatopancreas: 238 genes) when compared with that at Day 7 (gills: 104 genes, hepatopancreas: 112 genes). Exposure to TPT strongly impaired the endocrine system through targeting on nuclear receptors and putative steroid metabolic genes. Moreover, TPT widely disrupted cellular functions, including lipid metabolism, xenobiotic detoxification, immune response and endoplasmic-reticulum-associated degradation expression, which might have caused the bioaccumulation of TPT in the tissues and aggregation of peptides and proteins in cells that further activated the apoptosis process in P. viridis. Overall, this study has advanced our understanding on both ecotoxicity and molecular toxic mechanisms of TPT to marine mussels, and contributed empirical toxicity data for risk assessment and management of TPT contamination.

Toxicity effects of hydrophilic algal lysates from Coolia tropicalis on marine medaka larvae (Oryzias melastigma).

Coolia tropicalis is a species of benthic and epiphytic toxic algae, which can produce phycotoxins that intoxicate marine fauna. In this study, the potential toxic effects of C. tropicalis on fish were investigated using larval marine medaka (Oryzias melastigma) as a model to evaluate fish behavior, physiological performance, and stress-induced molecular responses to exposure to two sublethal concentrations (LC10 and LC20) of hydrophilic algal lysates. Exposure to C. tropicalis lysates inhibited swimming activity, activated spontaneous undirected locomotion, altered nerve length ration, and induced early development abnormalities, such as shorter eye diameter, body as well as axon length. Consistent with these abnormalities, changes in the expression of genes associated with apoptosis (CASPASE-3 and BCL-2), the inflammatory response (IL-1ß and COX-2), oxidative stress (SOD), and energy metabolism (ACHE and VHA), were also observed. This study advances our understanding of the mechanisms of C. tropicalis toxicity in marine fish in the early life stages and contributes to future ecological risk assessments of toxic benthic dinoflagellates.

Classification of marine microalgae using low-resolution Mueller matrix images and convolutional neural network.

In this paper, we used a convolutional neural network to study the classification of marine microalgae by using low-resolution Mueller matrix images. Mueller matrix images of 12 species of algae from 5 families were measured by a Mueller matrix microscopy with an LED light source at 514 nm wavelength. The data sets of seven resolution levels were generated by the bicubic interpolation algorithm. We conducted two groups of classification experiments; one group classified the algae into 12 classes according to species category, and the other group classified the algae into 5 classes according to family category. In each group of classification experiments, we compared the classification results of the Mueller matrix images with those of the first element (M11) images. The classification accuracy of Mueller matrix images declines gently with the decrease of image resolution, while the accuracy of M11 images declines sharply. The classification accuracy of Mueller matrix images is higher than that of M11 images at each resolution level. At the lowest resolution level, the accuracy of 12-class classification and 5-class classification of full Mueller matrix images is 29.89% and 35.83% higher than those of M11 images, respectively. In addition, we also found that the polarization information of different species had different contributions to the classification. These results show that the polarization information can greatly improve the classification accuracy of low-resolution microalgal images.

The effect of temperature on physiology, toxicity and toxin content of the benthic dinoflagellate Coolia malayensis from a seasonal tropical region.

Coolia malayensis is one of the commonly found benthic dinoflagellates in Hong Kong which can produce biotoxins and threaten the early life stages of marine invertebrates. Seawater temperature has been recognized as one of the primary environmental factors that affect the formation of harmful algal blooms. The present study evaluated the responses of C. malayensis, including growth, toxicity and toxin content (putative analogues of okadaic acid and azaspiracids), after exposure to a range of seven different temperatures (i.e., 16°C, 18°C, 20°C, 22°C, 24°C, 26°C, and 28°C). The highest algal density and specific growth rate were recorded at 24°C. Significantly higher Fv/Fm (maximum quantum yield of PSII) and total phaeo-pigment values were observed in the exponential growth phase at 28°C. The toxicity of the algal extract, which was assessed by the lethality rate of Artemia larvae, increased with temperature. The highest toxin content was detected at the second highest temperature treatment, i.e., 26°C. Overall, temperature had significant effects on the physiological activities and toxicity of C. malayensis. This study has raised attention to the potentially increasing risks posed by toxic benthic dinoflagellates during heat waves in coastal waters.

The role of 44-methylgambierone in ciguatera fish poisoning: Acute toxicity, production by marine microalgae and its potential as a biomarker for Gambierdiscus spp.

Ciguatera fish poisoning (CFP) is prevalent around the tropical and sub-tropical latitudes of the world and impacts many Pacific island communities intrinsically linked to the reef system for sustenance and trade. While the genus Gambierdiscus has been linked with CFP, it is commonly found on tropical reef systems in microalgal assemblages with other genera of toxin-producing, epiphytic and/or benthic dinoflagellates - Amphidinium, Coolia, Fukuyoa, Ostreopsis and Prorocentrum. Identifying a biomarker compound that can be used for the early detection of Gambierdiscus blooms, specifically in a mixed microalgal community, is paramount in enabling the development of management and mitigation strategies. Following on from the recent structural elucidation of 44-methylgambierone, its potential to contribute to CFP intoxication events and applicability as a biomarker compound for Gambierdiscus spp. was investigated. The acute toxicity of this secondary metabolite was determined by intraperitoneal injection using mice, which showed it to be of low toxicity, with an LD50 between 20 and 38 mg kg-1. The production of 44-methylgambierone by 252 marine microalgal isolates consisting of 90 species from 32 genera across seven classes, was assessed by liquid chromatography-tandem mass spectrometry. It was discovered that the production of this secondary metabolite was ubiquitous to the eight Gambierdiscus species tested, however not all isolates of G. carpenteri, and some species/isolates of Coolia and Fukuyoa.

Effects of dietary exposure to ciguatoxin P-CTX-1 on the reproductive performance in marine medaka (Oryzias melastigma).

Ciguatoxins are natural compounds produced by benthic dinoflagellates Gambierdiscus and Fukuyoa spp., which cause fish intoxication by ciguatera fish poisoning. This study aimed to assess the dietary exposure effects of ciguatoxin P-CTX-1 on the reproductive performance in marine medaka (Oryzias melastigma). Fish which ingested >1.16 pg·day-1 for 21 days exhibited abnormal behaviors including diarrhea, abnormal swimming, loss of appetite and decreased egg production. After 7-day exposure to P-CTX-1 at a dose of 1.93 pg·day-1, significant gender-specific differences in reproductive performance and decreased hatching rate of the offspring were observed. Chemical analysis of P-CTX-1 showed that the P-CTX-1 accumulation rates were 24.1 ± 1.4% in female fish and 9.9 ± 0.4% in male fish, and 0.05 pg·egg-1 was detected. The results illustrate that dietary exposure to P-CTX-1 affected the reproductive performance and survival of offspring, and caused bioaccumulation and maternal transfer of P-CTX-1 in marine medaka.

Hemolysis associated toxicities of benthic dinoflagellates from Hong Kong waters.

Benthic dinoflagellates produce a diverse range of phycotoxins, which are responsible for intoxication events in marine fauna. This study assessed the hemolysis associated toxicities of six species of benthic dinoflagellates from the genera Coolia, Fukuyoa, Amphidinium and Prorocentrum. Results demonstrated that Amphidinium carterae, Coolia tropicalis and Fukuyoa ruetzleri were the three most toxic species, while Prorocentrum cf. lima did not have significant hemolytic effect. Grouper samples (Cephalopholis boenak) were more tolerant to the hemolytic algae than the blackhead seabream (Acanthopagrus schlegelii), with decreased heart rate and blood flow being observed in medaka larvae after exposure to toxic algal extracts. LC-MS/MS analysis detected a gambierone analogue called 44-methylgambierone produced by the C. tropicalis isolate. This analogue was also detected in the F. ruetzleri isolate. This study provided new information on the hemolysis associated toxicities of local toxic benthic dinoflagellates, which contributes to better understanding of their emerging threats to marine fauna and reef systems in Hong Kong.

Sunscreens containing zinc oxide nanoparticles can trigger oxidative stress and toxicity to the marine copepod Tigriopus japonicus.

The study, for the first time, evaluated the leaching rate of zinc oxide nanoparticles (nZnO) from human skins which were applied with three commercial sunscreens containing nZnO as an active ingredient. The leaching rate of nZnO varied greatly among the sunscreens, with a range of 8-72% (mean ± SD: 45% ± 33%). We further investigated their toxicities to the marine copepod Tigriopus japonicus. We found that 96-h median lethal concentrations of the three sunscreens to T. japonicus were > 5000, 230.6, and 43.0 mg chemical L-1, respectively, equivalent to Zn2+ concentrations at >82.5, 3.2, and 1.2 mg Zn L-1, respectively. Exposure to the individual sunscreens at environmentally realistic concentrations for 96 h led to up-regulation of antioxidant genes in T. japonicus, while they triggered the release of reactive oxygen species based on the results of in vivo assays. Evidently, these nZnO-included sunscreens can cause oxidative stress and hence pose risk to marine organisms.

Population Genetic Structure of A Marine Pelagic Egg Producer and Popular Marine Aquarium Species, the Mandarinfish Synchiropus splendidus.

The mandarinfish Synchiropus splendidus is extensively collected in Southeast Asia (mainly in the Philippines) and highly favoured for the marine aquarium trade. Males are more popular than females for their large first dorsal fins and the fishery is not managed. To examine possible population replenishment dynamics arising as a result of selective fishing, the effects of sex-selective fishing on sex ratios and population connectivity were considered. This study determined the sex ratios and analyzed the population genetic structure from mandarinfish collected at six locations: one from Palau, where the species is not exploited, and five from Bohol in the Philippines, where the species has long been heavily fished. The findings reported very low male to female ratios (0.12 to 0.30) from four of the five locations in Bohol, with relatively more males to females in the specimens collected from Palau (2.3). The analyses from allozymes (43 alleles from 10 loci) and microsatellites (118 alleles from 5 loci) revealed that genetic connectivity was high among the five locations in the Philippines as well as with the specimens collected from the more-distant Palau. The genetic homogeneity observed across the geographical range considered is inconsistent with the hypothesized limited dispersal ability of the species and could be explained by recent species range expansion associated with sea level rise in the region. The results suggest that the present genetic structure, at least in the geographic region considered, may not be determined by current patterns of gene flow, but may, instead, be driven by recent sea-level changes associated with periods of glaciation. Caution is suggested to ensure that heavily localized fishing does not produce excessively biased adult sex ratios.

Polarimetric learning: a Siamese approach to learning distance metrics of algal Mueller matrix images.

Polarimetric measurements are becoming increasingly accurate and fast to perform in modern applications. However, analysis on the polarimetric data usually suffers from its high-dimensional nature spatially, temporally, or spectrally. This paper associates polarimetric techniques with metric learning algorithms, namely, polarimetric learning, by introducing a distance metric learning method called Siamese network that aims to learn good distance metrics of algal Mueller matrix images in low-dimensional feature spaces. As an experimental example, 12,162 Mueller matrix images of eight algal species are measured via a forward Mueller matrix microscope. Eight classical metric learning algorithms, including principle component analysis, multidimensional scaling, isometric feature mapping, t-distributed stochastic neighbor embedding, Laplacian eigenmaps, locally linear embedding, linear discriminant analysis, and metric learning to rank, are considered, by which the algal Mueller matrix images are mapped to two-dimensional (2D) feature spaces with different distance metrics. Support-vector-machine-based holdout sample classification accuracies of the 2D feature vectors are provided in a supervised manner for quantitative comparisons of the low-dimensional distance metrics, including the results of the eight metric learning algorithms and 16 Siamese architectures with varying convolution, inception, and full connection modules. This study shows that the Siamese approach is an effective metric learning algorithm that can adaptively extract features exhibiting empirical correlations with the fast-axis-orientation-dependent and spatially variant algal retardance induced by the algal microstructures.

Phylogeny, morphology and toxicity of benthic dinoflagellates of the genus Fukuyoa (Goniodomataceae, Dinophyceae) from a subtropical reef ecosystem in the South China Sea.

Species of Fukuyoa, recently revised from the globular Gambierdiscus, are toxic benthic dinoflagellates associated with ciguatera. In this study, a total of ten strains of Fukuyoa collected from Hong Kong waters were characterized using morphological and phylogenetic analyses. Results from both analyses showed that one of the strains is a putative new species, namely Fukuyoa sp. HK Type 1 (plate formula Po, 3', 7″, 6c, 7s, 5‴, 1p and 2'‴ with a distinctive small and narrow cell shape, narrow Po plate, high Po pore density, large and broad Plate 1' but small and round Po pore size, small and narrow Plate 2', long and narrow Plates 2'‴ and 1p), and the others were F. ruetzleri. This is the first report of these two species of Fukuyoa in the South China Sea and Asia-Pacific region. Phylogenies on 18S, 28S D1/D3 and D8/D10 ribosomal DNA sequences strongly support that Fukuyoa sp. HK Type 1 is currently the most divergent species in the genus Fukuyoa. The diagrammatic plots on the p-distance matrices of 18S, 28S D1/D3 and D8/D10 and ITS regions resolved that the species of Fukuyoa were separated into three main groups, i.e., Fukuyoa sp. HK Type 1, F. paulensis and a group consisting of F. ruetzleri, F. yasumotoi and F. cf. yasumotoi, while Fukuyoa sp. HK Type 1 was always the most distant from the other two groups. Additionally, the pairwise p-distance values calculated based on the ITS region have always been the highest for pairs between Fukuyoa sp. HK Type 1 and other Fukuyoa species, ranging from 0.142 to 0.150. Our molecular results suggested that Fukuyoa sp. HK Type 1 is a putative new species. Both morphological and molecular data of more strains from different localities should be, however, collected to address its intraspecific variability and further evaluate its taxonomic status. A bioassay analysis demonstrated that algal lysates extracted from F. ruetzleri and Fukuyoa sp. HK Type 1 were lethal to brine shrimp larvae, indicating that both species were toxic. Bulk cultures were tested for Pacific ciguatoxins (P-CTXs) and maitotoxins (MTXs) by liquid chromatography-tandem mass spectrometry (LC-MS/MS). All isolates of Fukuyoa produced neither P-CTXs nor MTX-1, but isolates of F. ruetzleri produced a compound putatively assigned as MTX-3. This study has updated the current biodiversity and distribution of the toxic benthic dinoflagellates Fukuyoa, and thus contributes to the understanding of their emerging threats to the sub-tropical reef systems locally and regionally.

Classification of morphologically similar algae and cyanobacteria using Mueller matrix imaging and convolutional neural networks.

We present the Mueller matrix imaging system to classify morphologically similar algae based on convolutional neural networks (CNNs). The algae and cyanobacteria data set contains 10,463 Mueller matrices from eight species of algae and one species of cyanobacteria, belonging to four phyla, the shapes of which are mostly randomly oriented spheres, ovals, wheels, or rods. The CNN serves as an automatic machine with learning ability to help in extracting features from the Mueller matrix, and trains a classifier to achieve a 97% classification accuracy. We compare the performance in two ways. One way is to compare the performance of five CNNs that differ in the number of convolution layers as well as the classical principle component analysis (PCA) plus the support vector machine (SVM) method; the other way is to quantify the differences of scores between full Mueller matrix and the first matrix element m11, which does not contain polarization information under the same conditions. As the results show, deeper CNNs perform better, the best of which outperforms the conventional PCA plus SVM method by 19.66% in accuracy, and using the full Mueller matrix earns 6.56% increase of accuracy than using m11. It demonstrates that the coupling of Mueller matrix imaging and CNN may be a promising and efficient solution for the automatic classification of morphologically similar algae.

Photosynthetic and transcriptional responses of the marine diatom Thalassiosira pseudonana to the combined effect of temperature stress and copper exposure.

A 96-h exposure experiment was conducted to elucidate the toxicity responses of the marine diatom Thalassiosira pseudonana upon exposure to different temperatures and copper (Cu) concentrations. Three Cu treatments (seawater control; 200µg/L Cu, EC50 for the yield at 25°C; and 1000µg/L Cu, EC50 for growth inhibition at 25°C) were conducted against four temperatures (10°C, 15°C, 25°C and 30°C). Growth rate and photosynthetic responses showed a significant interacting thermal-chemical effect with strong synergistic responses observed at 30°C treatments. Expression of heat shock protein (hsp) was positively modulated by increasing temperatures. Hsp 90, hsp90-2 and sit1 (related to silica shell formation) were highly expressed at 30°C under 1000µg/L Cu, while the genes encoding light harvesting proteins (3HfcpA and 3HfcpB) and silaffin precursor sil3 were significantly up-regulated at 15°C under 200µg/L Cu. Our results indicated an increase Cu toxicity to T. pseudonana under high temperature and Cu dose.

Molecular phylogeny and toxicity of harmful benthic dinoflagellates Coolia (Ostreopsidaceae, Dinophyceae) in a sub-tropical marine ecosystem: The first record from Hong Kong.

Coolia are marine benthic dinoflagellates which are globally distributed and potentially toxic. This study provides the first investigation of species diversity and toxicity assessment of Coolia in Hong Kong waters. Fifty-one strains of four Coolia species, including C. malayensis, C. canariensis, C. tropicalis, and C. palmyrensis, were isolated from twelve sub-tidal habitats, and identified phylogenetically using 28S rDNA sequences. Exposure experiments (48-hour) demonstrated that the algal lysates extracted from the four Coolia species exhibited different toxic effects on the lethality and abnormality of two invertebrate larvae, i.e., brine shrimp Artemia franciscana and sea urchin Heliocidaris crassispina. Heliocidaris crassispina was more sensitive to the toxic effects of Coolia species than A. franciscana. Toxicity tests from both larvae revealed that C. malayensis was generally more toxic, and caused higher mortality rates when compared with the other three species. The emerging threat of harmful benthic dinoflagellates to marine environments and sensitive biota is discussed.

Developmental toxicity and molecular responses of marine medaka (Oryzias melastigma) embryos to ciguatoxin P-CTX-1 exposure.

Ciguatoxins are produced by toxic benthic dinoflagellates and cause ciguatera fish poisoning worldwide, but the toxic effects on developing marine fish have not been well investigated. The Pacific ciguatoxin (P-CTX-1), is a potent sodium channel agonist, which is one of the most toxic members among all CTXs. This study evaluated the toxic effects of microinjecting purified Pacific ciguatoxin-1 (P-CTX-1) on embryonic development of marine medaka Oryzias melastigma. A lower 96h-LD50 value was estimated for eleuthero-embryos (1.32ngg-1) than that for embryos (1.71ngg-1), indicating that P-CTX-1 is more lethal to newly hatched medaka larvae. P-CTX-1 induced detrimental effects during embryonic development, including hatching failure, abnormalities in physical development (caudal fin malformation and spinal deformities), internal damage (green coloration of the gall bladder and hemorrhaging), immune dysfunction, and altered muscle physiology (bradycardia and hyperkinetic twitching). The results of a transcriptional expression analysis of genes related to the stress/immune responses, cardiac and bone development, and apoptosis supported the observed developmental abnormalities. This study advanced the understanding of P-CTX-1 mediated toxic mechanisms in the development of early life stages of a fish, and thus contributed to the toxicity assessment of CTXs in marine ecosystems.

De novo transcriptome assembly of the marine gastropod Reishia clavigera for supporting toxic mechanism studies.

The intertidal whelk Reishia clavigera is commonly used as a biomonitor of chemical contamination in the marine environment along Western Pacific region, and as a model for mechanistic studies of organotin-mediated imposex development. However, limited genomic resources of R. clavigera have restricted its role for the investigation of molecular mechanisms of such endocrine disruptions. This study, therefore, aimed to establish tissue-specific transcriptomes of the digestive gland, gonad, head ganglia, penis and the remaining body part of the male and female R. clavigera. By combining the results, a global transcriptome was obtained. A total of 578,134,720 high-quality filtered reads were obtained using Illumina sequencing. The R. clavigera transcriptome comprised of 38,466 transcripts and 32,798 unigenes with predicted open reading frames. The average length of transcripts was 1,709bp with N50 of 2,236bp. Based on sequence similarity searches against public databases, 28,657 transcripts and 24,403 unigenes had at least one BLAST hit. There were 17,530 transcripts and 14,897 unigenes annotated with at least one Gene Ontology (GO) term. Moreover, 5,776 transcripts and 5,137 unigenes were associated with 333 Kyoto Encyclopaedia of Genes and Genomes (KEGG) pathways. The numbers of unigenes were similar among the five target tissues and between sexes, but tissue-specific expression profiles were revealed by multivariate analyses. Based on the functional annotation, putative steroid hormone-associated unigenes were identified. In particular, we highlighted the presence of steroid hormone receptor homologues that could be the targets for mechanistic studies of the organotin-mediated imposex development in marine gastropods. This newly generated transcriptome assembly of R. clavigera provides a valuable molecular resource for ecotoxicological and environmental genomic studies.