Digestive Gland Tissues Research Articles

Using the enzymatic and non-enzymatic antioxidant defense system of the land snail Eobania vermiculata as biomarkers of terrestrial heavy metal pollution

The present study aimed to investigate the efficacy of antioxidant defense system of the land snail Eobania vermiculata as biomarkers for terrestrial heavy metal pollution. For this reason, a set of biomarkers was performed on land snails E. vermiculata collected from polluted and non-polluted areas in the field. The biomarkers used were lactate dehydrogenase activity, level of lipid peroxidation, activities of catalase, glutathione peroxidase, glutathione-S-transferase, gamma-glutamyl transferase and content of glutathione, metalothionines, total proteins and total lipid in the digestive gland tissue. The correlation between the bioaccumulation of heavy metal in gland tissue and all the biomarkers in the digestive gland was determined. The results revealed appreciable alterations in the biomarker values in field, accompanied by significant correlations among the biomarkers. Therefore, this study suggests E. vermiculata is a suitable sentinel organism and the selected biomarkers show efficacy for terrestrial heavy metal biomonitoring.

Increased Temperatures Affect Oxidative Stress Markers and Detoxification Response to Benzo[a]Pyrene Exposure in Mussel Mytilus galloprovincialis

The present research work was designed to study mussel's (Mytilus galloprovincialis) digestive gland biotransformation and detoxification responses to benzo[a]pyrene (B[a]P) exposure along with heat stress. Mussels were exposed to a sublethal dose of B[a]P [75 nM (19 μg/L/animal)] + temperature gradient (18, 20, 22, 24 and 26 °C) for 7 days. B[a]P hydroxylase (BPH) and glutathione-S-transferase (GST) activities were assessed in digestive gland tissues as phase I and phase II biotransformation parameters. Catalase (CAT) activity and malonedialdehyde (MDA) were measured as potential biomarkers of oxidative stress and lipid peroxidation. The cholinergic system was evaluated using acetylcholinesterase (AChE) activity. DNA damage was assessed using micronuclei (MN) test. BPH and GST activities showed a decreasing trend in B[a]P-exposed animals at 24 and 26 °C. CAT activity showed a bell-shaped response in B[a]P-exposed and in heat-stressed organisms at a maximum temperature of 22 °C. AChE activity was significantly inhibited in response to B[a]P being more pronounced at a temperature of 26 °C. MN in digestive gland cells suggest that B[a]P exposure induced significant DNA alteration with a maximum response in organisms coexposed to B[a]P and a temperature of 26 °C. Biomarker data are further discussed in relation B[a]P accumulation in mussels digestive gland. These data should be carefully considered in view of the biological effects of organic pollutants, particularly in organisms under the challenging effects of extreme temperature fluctuations.

The interaction of infectious salmon anaemia virus (ISAV) with the blue mussel,Mytilus edulis

Integrated multi-trophic aquaculture (IMTA) is an alternative approach to mono-culture aquaculture that reduces environmental impacts of commercial aquaculture systems by combining the cultivation of fed species with extractive species. Shellfish play a critical role in IMTA systems by filter-feeding particulate-bound organic nutrients. They may also increase or decrease disease risk on farms by serving as reservoirs or barriers for important finfish pathogens such as infectious salmon anaemia virus (ISAV). This study aimed to optimize culture and molecular assays in shellfish tissues and to determine the fate of ISAV in mussels, Mytilus edulis. To determine detection limits, qRT-PCR and culture assays in both CHSE- and ASK cells were optimized in ISAV-inoculated mussel tissue homogenates. Both qRT-PCR and culture assays performed in ASK cells had comparable detection limits of 102.8 TCID50 mL−1. The ISAV RNA genome was consistently detected in digestive gland tissue of ISAV-exposed mussels. Viable ISAV was not detected in mussel tissues by culture analysis in CHSE- and ASK cells. The fact that qRT-PCR analysis resulted in positive cycle threshold (CT) values that corresponded to the detectable range of ISAV in ASK culture assays suggests that little to no viable ISAV particles are present in the mussel tissues.

Benthic community structure and biomarker responses of the clam Scrobicularia plana in a shallow tidal creek affected by fish farm effluents (Rio San Pedro, SW Spain)

The effects of solid organic wastes from a marine fish farm on sediments were tested using benthic community as ecological indicators and biomarkers in native clam (Scrobicularia plana) as biochemical indicators. The benthic fauna and clam samples were collected in the intertidal sediment in October 2010 from five sites of the Rio San Pedro (RSP) creek, following a gradient of contamination from the aquaculture effluent to the control site. Numbers of species, abundance, richness and Shannon diversity were the biodiversity indicators measured in benthic fauna. Morphological and reproduction status of clams were assessed using the condition factor and gonado-somatic index, respectively. Phase I and Phase II detoxification enzymatic activities (ethoxyresorufin O-deethylase (EROD), glutathione S-transferase (GST)), antioxidant enzymatic activities (glutathione peroxidase (GPX), glutathione reductase (GR)) and oxidative stress parameters (Lipid Peroxidation (LPO) and DNA strand breaks) were measured in clams' digestive gland tissues. In parallel, temperature and salinity in the adjacent water, redox potential, pH and organic matter in sediment, and dissolved oxygen in the interstitial water were measured. The results suggested that RSP showed a spatial gradient characterised by hypoxia/anoxia, reduced potential, acidic conditions and high organic enrichment in sediments at the most contaminated sites. Significant (p<0.05) decrease of biodiversity indicators were observed in the areas impacted by the aquaculture discharges. Biomarkers did not show a clear pattern and of all biochemical responses tested, GPX, DNA damage and LPO were the most sensitive ones and showed significant (p<0.05) increase in the polluted sites. Benthic biodiversity indicators were significantly (p<0.05) positively correlated with pH, redox potential and dissolved oxygen and negatively correlated with organic matter. On the contrary, antioxidant enzymatic responses (GPX) and oxidative stress parameters were significantly (p<0.05) negatively correlated with those physico-chemical parameters. It has been demonstrated that effluents from fish aquaculture activities in Río San Pedro creek may produce an alteration of physico-chemical characteristics of seabed and induce oxidative stress and DNA damage in soft-sediment species which may lead to changes of the benthic population structure and health status of the exposed organisms.

Putative Phage Hyperparasite in the Rickettsial Pathogen of Abalone, “Candidatus Xenohaliotis californiensis”

Studies on the ecology of microbial parasites and their hosts are predicated on understanding the assemblage of and relationship among the species present. Changes in organismal morphology and physiology can have profound effects on host-parasite interactions and associated microbial community structure. The marine rickettsial organism, "Candidatus Xenohaliotis californiensis" (WS-RLO), that causes withering syndrome of abalones has had a consistent morphology based on light and electron microscopy. However, a morphological variant of the WS-RLO has recently been observed infecting red abalone from California. We used light and electron microscopy, in situ hybridization and16S rDNA sequence analysis to compare the WS-RLO and the morphologically distinct RLO variant (RLOv). The WS-RLO forms oblong inclusions within the abalone posterior esophagus (PE) and digestive gland (DG) tissues that contain small rod-shaped bacteria; individual bacteria within the light purple inclusions upon hematoxylin and eosin staining cannot be discerned by light microscopy. Like the WS-RLO, the RLOv forms oblong inclusions in the PE and DG but contain large, pleomorphic bacteria that stain dark navy blue with hematoxylin and eosin. Transmission electron microscopy (TEM) examination revealed that the large pleomorphic bacteria within RLOv inclusions were infected with a spherical to icosahedral-shaped putative phage hyperparasite. TEM also revealed the presence of rod-shaped bacteria along the periphery of the RLOv inclusions that were morphologically indistinguishable from the WS-RLO. Binding of the WS-RLO-specific in situ hybridization probe to the RLOv inclusions demonstrated sequence similarity between these RLOs. In addition, sequence analysis revealed 98.9-99.4% similarity between 16S rDNA sequences of the WS-RLO and RLOv. Collectively, these data suggest that both of these RLOs infecting California abalone are "Candidatus Xenohaliotis californiensis," and that the novel variant is infected by a putative phage hyperparasite that induced morphological variation of its RLO host.

Integrated coastal monitoring of a gas processing plant using native and caged mussels

The biological effects of a coastal process water (PW) discharge on native and caged mussels (Mytilus edulis) were assessed. Chemical analyses of mussel tissues and semi permeable membrane devices, along with a suite of biomarkers of different levels of biological complexity were measured. These were lysosomal membrane stability in haemocytes and digestive cells; micronuclei formation in haemocytes; changes in cell-type composition in the digestive gland epithelium; integrity of digestive gland tissue; peroxisome proliferation; and oxidative stress. Additionally the Integrative Biological Response (IBR/n) index was calculated. This integrative biomarker approach distinguished mussels, both native and caged, exhibiting different stress conditions not identified from the contaminant exposure. Mussels exhibiting higher stress responses were found with increased proximity to the PW discharge outlet. However, the biological effects reported could not be entirely attributed to the PW discharge based on the chemicals measured, but were likely due to either other chemicals in the discharge that were not measured, the general impact of the processing plant and or other activities in the local vicinity.

Selection of Target Mussel Tissue for Application of Cellular Energy Allocation as a Physiological Biomarker in Native MusselsMytilus galloprovincialis(Lamarck, 1819)

Three selected mussel tissues (digestive gland, mantle, and gills) were studied to determine which was the most suitable for the potential use of the cellular energy allocation (CEA) methodology in indigenous mussels Mytilus galloprovincialis. In addition, the applicability of CEA in the assessment of natural stress caused by salinity fluctuations in stratified estuary was tested in selected tissues. It was important to identify the mussel gender to reliably assess the changes in organism energy budget. CEA value was calculated as a ratio between available energy (Ea) and energy consumption (Ec). Mantle tissue was under the strongest influence of the differences in protein and lipid content between male and female mussels, and therefore reflected physiological changes in the organism itself, rather than those caused by natural environmental stress. CEA in gills had lower values than in mantle and digestive gland, and was similar at two selected sampling sites, so the changes in CEA caused by natural stress could not be detected in the gill tissue. Greater Ec in mussels from the estuarine site than from the coastal site was detected only in the digestive gland tissue, and can probably be attributed to the energetically costly maintenance of osmotic balance. Last, using digestive gland tissue in CEA analysis demonstrated a clear difference between coastal and estuarine sampling sites, providing the measure of the natural stress posed by variations in salinity.

Seasonal fatty acid profile analysis to trace origin of food sources of four commercially important bivalves

Knowledge of bivalve food sources has great importance in aquaculture areas where cultured species usually occur at higher densities than in natural populations. Except phytoplankton, which is considered as their primary food sources, bivalves can use other food sources such as detritus, bacteria and zooplankton. The objective of this study was to determine food sources of four commercially important bivalve species co-existing in the Mali Ston Bay — most important aquaculture area of the eastern Adriatic Sea. Food sources were investigated by analyzing the fatty acid profiles of digestive gland and muscle tissues of the European flat oyster Ostrea edulis and the black mussel Mytilus galloprovincialis, cultured in the water column, and the bearded horse mussel Modiolus barbatus and Noah's ark shell Arca noae from natural populations. Digestive gland fatty acid analysis revealed that during the period of higher plankton abundance in spring and summer, all species ingested mainly phytoplankton followed by zooplankton and detritus. During the low plankton abundance of the autumn–winter period zooplankton and/or detritus became more important in a bivalve diet. Fatty acid composition of muscles indicated differences in selective incorporation of polyunsaturated fatty acid (PUFA) from their diets. While A. noae and M. barbatus accumulated significantly PUFAs during all and O. edulis during spring–summer period, in M. galloprovincialis this tendency was not observed at all. This suggested better utilization of ingested food in bottom-dwelling species. Because all four species consumed similar food sources they were in competition. However, as investigated species in Mali Ston Bay exist in high density, primary production plus the availability of other food sources in this area was probably sufficient to satisfy demands for all species. Aquaculture in the study area depends wholly on natural spat. This suggests that larviphagy, observed for all species, might constitute a self-regulating restriction to natural populations and/or commercial aquaculture growth.

Biomarkers of petroleum contamination in the digestive gland of Perna viridis (L.)

Oxidative stress responses were evaluated in the digestive gland tissue of Perna viridis exposed to water-accommodated fractions (WAFs) of petrol and diesel at 0.5% and 5% concentrations over 5-, 10-, and 15-day periods. Increased lipid peroxidation, protein carbonyl, DNA integrity, and hydrogen peroxide in WAF-exposed specimens are indicative of oxidative damage due to dissolved petroleum extracts. A consistent increase in superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase, glutathione-S-transferase, reduced glutathione, and ascorbic acid in WAF-exposed specimens signify the importance of antioxidants in protecting the cell against the oxidative damage arising from petroleum contamination. The responses of field specimens collected from petroleum-contaminated sites support the observed laboratory experimental results. This research recommends the use of these oxidative stress indices in the digestive gland tissue of Perna viridis as biomarkers of petroleum contamination.

Potential role of Mytilus edulis in modulating the infectious pressure of Vibrio anguillarum 02β on an integrated multi-trophic aquaculture farm

Marine finfish producers in the Northeast are adopting an integrated multi-trophic aquaculture (IMTA) approach by growing mussels, Mytilus edulis, with marine finfish species. Shellfish play a critical role in an IMTA system by filtering particulate bound nutrients and other seston from surrounding waters. During feeding, mussels bio-accumulate microbes, including fish pathogens, from the water column. This may influence pathogen dynamics on IMTA farms by either reducing infectious pressure or by serving as a reservoir for important finfish pathogens. The first step in evaluating the disease risks or benefits on a mussel-fish IMTA farm is to understand the interaction between mussels and finfish pathogens. We investigated the fate of the cod bacterial pathogen Vibrio anguillarum 02β in the blue mussel Mytilus edulis. Mussels exposed to 10 4 CFU of V. anguillarum ml − 1 concentrated bacteria in digestive gland tissues 2 orders of magnitude above the concentration of bacteria in the surrounding water. To determine if mussels could release concentrated bacteria back into the environment, V. anguillarum-exposed mussels were transferred to clean water and fecal and water samples were analyzed for the presence of bacteria. V. anguillarum was not detected in the water; however mussel feces and pseudofeces had high loads of V. anguillarum (10 7 CFU g − 1 ). Within 72 h of depuration, V. anguillarum was no longer detected in fecal samples. Immersion challenge trials using fecal material from V. anguillarum-exposed mussels resulted in 58–70% mortalities in challenged cod. These results demonstrate that mussels are capable of bio-accumulating V. anguillarum and shedding virulent bacteria through their feces. While this study does not likely reproduce the dynamic conditions on a finfish–mussel farm, results obtained do present valuable information when considering biosecurity and pathogen risk reduction on IMTA farms.

Potential food sources of &lt;em&gt;Glycymeris nummaria&lt;/em&gt; (Mollusca: Bivalvia) during the annual cycle indicated by fatty acid analysis

Seasonal changes of food sources were investigated by analysing the fatty acid profiles of digestive gland and muscle tissues of the naturally occurring clams Glycymeris nummaria in Mali Ston Bay, Croatia. Total lipids in the digestive gland and the adductor muscle showed parallel changes, with a maximum after the main spawning event in September. In the digestive gland saturated fatty acids were highly dominant (up to 82%), indicating detritus as the main food source for this species. This type of food prevailed during the autumn/winter period, in contrast to the spring/summer period when detritus was enriched with phyto- and, to a lesser extent, zooplankton. Fatty acid composition of muscles indicated highly efficient utilization of ingested food through significant retention of polyunsaturated fatty acid from the clams’ diet during the entire period investigated.

Incorporation of in situ exposure and biomarkers response in clams Ruditapes philippinarum for assessment of metal pollution in coastal areas from the Maluan Bay of China

The clams Ruditapes philippinarum were used to assess the impact of metal contaminants when transplanted to seven study sites along the Maluan Bay (China). Metal concentrations in digestive gland tissues of clams after 7-day in situ cage exposure were determined in conjunction with antioxidant enzyme activities. The results showed the importance of specific antioxidant biomarkers to assess complex pollutant mixtures and their good correlations to the pollutant compositions of deployment sites. Multivariate analysis indicated causal relationship between the chemicals at each study site and the biochemical “response” of the caged clams at these sites and demonstrated the potential presence of two different contaminant sources. This study suggested that the incorporation of tissue residue analysis with biomarkers response in caged clams together with factor analysis can be a useful biomonitoring tool for the identification of causal toxic pollutants and the assessment of complex metal pollutions in marine coastal environment.

Subchronic effects of environment-like cadmium levels on the bivalve Anodonta anatina (Linnaeus 1758): I. Bioaccumulation, distribution and effects on calcium metabolism

Cadmium (Cd) bioaccumulation, tissue distribution, and its effects on Ca levels in the freshwater bivalve Anodonta anatina were examined. Mussels were exposed for 35 days to algae- and water-borne 111Cd at an environment-like concentration and were allowed to depurate for posterior 120 days. Body fluids and tissue samples were taken during both phases to determine total Cd, the 114Cd:111Cd isotope ratio, the electrolytes Ca and Mg, and the essential trace metal zinc (Zn). Bioaccumulation and distribution of Cd and its effects on Ca levels were tissue-specific and time-dependent. During exposure, newly incorporated 111Cd increased in all body compartments, with highest levels in the kidney. During depuration, newly incorporated and background Cd both decreased slowly in the digestive gland and others while in the kidney it increased further, indicating mobilization from other body compartments and deposition in this organ. Following the uptake of Cd, Ca concentrations increased in body fluids within a few hours while it decreased in the digestive gland, mantle and other tissues; in the gills and kidney it remained largely unchanged. The derangement of Ca homeostasis, to some extent also of Mg and Zn, appears to be a major component of subchronic Cd toxicity in A. anatina.

Modelling of paralytic shellfish toxin biotransformations in the course of Crassostrea gigas detoxification kinetics

The aim of this study was to evaluate the importance of biotransformation of paralytic shellfish toxins during the detoxification process in contaminated oysters. Mathematical models based upon the detoxification patterns of digestive gland and other tissues were developed. It was demonstrated that biotransformations do not seem to play an important role in digestive gland or other tissue detoxification kinetics with our data set. Moreover, different toxin transfers from digestive gland toward other tissues were investigated. No significant transfer was highlighted in our data set. These first conclusions were drawn after comparing the results obtained from 13 biotransformations and identifiable transfer scenarios. Finally, to determine a more robust model, all 12 states corresponding to toxic compounds and tissues were aggregated into a single state model. The best adjustment was obtained with a simple one-compartment model based on total flesh toxicity with elimination rate expressed by a function depending on initial concentrations of GTX3 and GTX2 (i.e. the two major toxic compounds found in contaminated oysters).

Assessment of Clam Ruditapes philippinarum as Heavy Metal Bioindicators Using NMR‐Based Metabolomics

AbstractThere are mainly distributed three pedigrees (White, Liangdao Red, and Zebra) of Manila clam Ruditapes philippinarum in Yantai population along the Bohai marine and coast. However, the biological differences to environmental stressors have been ignored in toxicology studies, which could lead to the distortion of biological interpretations of toxicological effects induced by environmental contaminants. In this study, we applied a system biology approach, metabolomics to compare the metabolic profiles in digestive gland from three pedigrees of clam and characterize and compare the metabolic responses induced by mercury in clam digestive gland tissues to determine a sensitive pedigree of clam as a preferable bioindicator for metal pollution monitoring and toxicology research. The most abundant metabolites, respectively, included branched‐chain amino acids, alanine, and arginine in White samples, glutamate, dimethylglycine, and glycine in Zebra clams and acetylcholine, betaine, glucose, and glycogen in Liangdao Red clams. After 48 h exposure of 20 µg L−1 Hg2+, the metabolic profiles from the three pedigrees of clams showed differentially significant changes in alanine, glutamate, succinate, taurine, hypotaurine, glycine, arginine, glucose, etc. Our findings indicate the toxicological effects of mercury exposure in Manila clams including the neurotoxicity, disturbances in energetic metabolisms and osmoregulation in the digestive glands and suggest that Liangdao Red pedigree of clam could be a preferable bioindicator for the metal pollution monitoring based on the more sensitive classes of metabolic changes from digestive glands compared with other two (White and Zebra) pedigrees of clams.

Anthropogenic pollution stimulates oxidative stress in soft tissues of mussel Crenomytilus grayanus (Dunker1853)

The digestive gland and gills of the mussel Crenomytilus grayanus extracted from three locations — (i) sampled from a clean and (ii) polluted site and (iii) transplanted from the nonpolluted to polluted site - were analysed for antioxidant enzymes (superoxide dismutase, catalase, glutathione reductase), total oxyradical scavenging capacity and levels of lipid peroxidation products (malondialdehyde, conjugated dienes and lipofuscin). Perturbation of redox status was found in both digestive gland and gill tissues of mussels living in the polluted area. As the activities of superoxide dismutase and catalase were 1.2–3 times higher, the total oxyradical scavenging capacity was lower by 20–35% and the levels of lipid peroxidation products were 2–7 times higher compared to mussels from the reference site. In transplanted mussels, the lipid peroxidation process in both tissues was significantly stimulated (the level of conjugated dienes was increased 1.7–2.5-fold; malondialdehyde and lipofuscin contents were increased 3.5–5-fold) and the total oxyradical scavenging capacity fell by 50–70%. In addition, the transplantation generally resulted in transient and variable responses of antioxidant enzymes for both tissues. Complex response-behaviour of the antioxidant enzymes strongly points to the necessity of employing a combined approach that takes into account activities of antioxidant enzymes and the total oxyradical scavenging capacity, as well as measurement of oxidative damage (e.g., lipid peroxidation) to evaluate the physiological health of molluscs.

What is your diagnosis? Pale yellowish digestive gland and watery tissues in Mediterranean mussels

Veterinary Clinical PathologyVolume 40, Issue 2 p. 273-274 What is your diagnosis? Pale yellowish digestive gland and watery tissues in Mediterranean mussels Francesca Carella, Francesca Carella Departments of Biological SciencesSearch for more papers by this authorSerena Aceto, Serena Aceto Departments of Biological SciencesSearch for more papers by this authorPaola Maiolino, Paola Maiolino Pathology and Animal Health, University of Naples Federico II, Naples, ItalySearch for more papers by this authorGionata De Vico, Gionata De Vico Departments of Biological SciencesSearch for more papers by this author Francesca Carella, Francesca Carella Departments of Biological SciencesSearch for more papers by this authorSerena Aceto, Serena Aceto Departments of Biological SciencesSearch for more papers by this authorPaola Maiolino, Paola Maiolino Pathology and Animal Health, University of Naples Federico II, Naples, ItalySearch for more papers by this authorGionata De Vico, Gionata De Vico Departments of Biological SciencesSearch for more papers by this author First published: 09 May 2011 https://doi.org/10.1111/j.1939-165X.2011.00316.xCitations: 5 Francesca Carella, Department of Biological Sciences, University of Naples Federico II, Via Mezzocannone 8, 80134 Naples, ItalyE-mail: [email protected] Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat No abstract is available for this article.Citing Literature Volume40, Issue2June 2011Pages 273-274 RelatedInformation

Seawater ozonation and formalin disinfection for the larval culture of eastern rock lobster, Jasus ( Sagmariasus) verreauxi, phyllosoma

The effects of seawater disinfection with continuous ozonation or with daily formalin treatment during flow-through culture, on the survival, growth, bacteriology and histology of eastern rock lobster, Jasus ( Sagmariasus) verreauxi, phyllosoma were determined. Survival from hatch to Instar II was highest and bacterial abundance was least in low ozonated seawater (containing 5 ppb ionised bromine, Br). By contrast, at high ozonation (45 ppb Br) and medium ozonation (15 ppb Br), 77% and 69% of larvae, respectively, suffered deformities at the moult to Instar II and starved to death. Histological examination of phyllosoma showed no differences in cuticular epithelium thickness but the digestive gland tissue of moribund deformed phyllosoma had significantly thinner tubule epithelium, the lumen was dilated, B-cells were more abundant and there was greater separation between the cuticular epithelium and distal tip of digestive gland tubules. In unozonated (Control) water, 66% of larvae died during Instar II probably resulting from a possible Vibrio infection. In a second experiment, survival to Instar III was highest at low ozonation or no ozonation without addition of formalin (0 ppm). Bacterial numbers were lowest in low ozonation with 0 ppm formalin treatments, compared to daily treatment with formalin at 10 ppm or above. A concentration of 250 ppm formalin killed all larvae by Instar III. In a final experiment, larval survival between Instar III to VI in low ozonated seawater was approximately 80% at formalin levels of 0 to 80 ppm, whereas all larvae treated with 160 ppm formalin died shortly after Instar V. Disinfection improved survival by minimising bacterial disease for the culture of phyllosoma to Instar VI without interrupting growth and development. The present study established that the optimum treatment for the culture of phyllosoma to Instar VI appeared to be low ozonation (5 ppb Br) with 10 ppm formalin.

Tissue‐specific toxicological effects of cadmium in green mussels (Perna viridis): Nuclear magnetic resonance‐based metabolomics study

Toxicity tests for metals have traditionally focused on selected biomarkers to characterize the biological stress induced by metals in marine organisms. Here nuclear magnetic resonance (NMR)-based metabolomics, a system biology tool, was applied to the marine green mussel, Perna viridis, to investigate the toxicological effects of Cd in both digestive gland and adductor muscle tissues. After Cd exposure for either two or four weeks, there was no significant metabolic change in the mussels exposed to Cd at 2 µg/L. At 20 µg/L, there were major metabolite changes related to amino acids, osmolytes, and energy metabolites. Digestive gland tissue was more sensitive to Cd than adductor muscle tissue. The adductor muscle tissue showed elevated levels of glutamine, glutamate, and lactate, and reduced levels of branched chain amino acids, aspartate, phenylalanine, and tyrosine. Overall, four weeks of Cd exposure produced neurotoxicity and metabolic disturbances and disturbed osmoregulation. These results suggest that the adductor muscle tissue of mussels may be a suitable supplemental biomarker for exposure to toxicants. In addition, the results demonstrate that (1) H-NMR-based metabolomic analysis can provide a systematic view of the toxicological effects of metals on mussels, suggesting that it might be employed to investigate the toxicological effects of other marine pollutants.

The metallothionein genes of Mytilus galloprovincialis: Genomic organization, tissue expression and evolution

Recently, increasing interest has been directed to the study of metallothioneins (MTs), which are small proteins that are able to bind metal ions. The induction of MT synthesis after exposure to metal or other environmental contaminants in a large number of aquatic invertebrates makes these proteins good biomarkers in water monitoring programs. Within bivalves, the species Mytilus galloprovincialis and Mytilus edulis represent model organisms for these types of studies, as well as for molecular studies regarding the expression and characterization of MT encoding genes. In the present paper, we focused on the genomic characterization, evolutionary, and tissue-expression analyses of the MT-10, MT-10 Intronless, and MT-20 genes in M. galloprovincialis. The comparison of the genomic sequences showed the presence of long nucleotide stretches within the introns of the MT genes that are conserved between M. galloprovincialis and M. edulis. These non-coding conserved sequences may contain regulatory motifs. Real-Time RT-PCR experiments revealed that, at the basal conditions, the MT-10 and MT-10 Intronless genes are expressed at levels considerably higher than the MT-20 gene, mainly in the digestive gland and gill tissue. The strong induction of the MT-20 gene expression detected in a field-collected sample is associated with the up-regulation of both the MT-10 and MT-10 Intronless genes. Evolutionary analysis revealed signals of localized positive selection that, together with the tissue-expression data, support a possible functional diversification between the MTs encoded by the MT-10 and MT-10 Intronless genes.