Aminotransferases in the bivalve mollusc Mytilus edulis L. and short term effects of crude oil in brackish water

Transaminases are among the crucial enzymes in amino acid metabolism, which in aquatic organisms is known to be affected by exposure to oil hydrocarbons. The transamination reactions in Mytilus edulis L. were studied to estimate their adequacy to indicate short term oil exposure in mussels. The transamination reactions were measured using paper chromatography and spectrophotometry. A high degree of transamination was observed between 2 oxoglutarate and alanine, aspartate and ornithine. A slight degree of transamination was shown with methionine, leucine, isoleucine, phenylalanine, serine, tryptophan, threonine, tyrosine and valine. No transamination was observed between 2 oxoglutarate and glycine, arginine, histidine, lysine, proline, citrulline and alanine. The effect of the water accommodated fraction WAF of crude oil on selected transaminase reactions was measured. The highest changes during the WAF exposure were mostly observed in the gills and mantle. Alanine aminotransferase EC 2.6.1.2 activity in the mantle was, at its highest, 55 over the control. Aspartate aminotransferase EC 2.6.1.1 activity increased in the gills by 52 . For ornithine transamination, in the gills the highest increase was by 75 and in the mantle by 50 . The metabolic pathways involved in the alterations of aminotransferase activities are discussed. It is concluded that ornithine transamination in gills is a potential indicator for short term crude oil exposure in Mytilus edulis. More studies are needed to evaluate the effects of other organic pollutants on ornithine transamination.     

Pharmacological Induction of Larval Settlement and Metamorphosis in the Blue Mussel Mytilus edulis L.

The blue mussel Mytilus edulis L. is an important aquaculture and fouling species in northern seas. Although the general role of chemical cues for settlement of larvae of the blue mussel has been proposed, few studies have focused on induction of settlement and metamorphosis by pharmacological agents. In this study, the induction of larval settlement of the blue mussel by pharmacological compounds was investigated through a series of laboratory experiments with an aim of identifying artificial cues for laboratory bioassay systems in fouling and antifouling research. Gamma-aminobutiric acid (GABA), dihydroxyphenyl L-alanine (DOPA), isobutyl methylxanthine (IBMX) and acetylcholine chloride (ACH) at 10^{m 7}-10^{m 2} M as well as KCl at 10-40 mM K⁺ in excess of the level in normal seawater were tested for their inductive effect on larval settlement. In filtered seawater (FSW) <9% of the larvae settled after 48 h. Elevated K⁺ and GABA levels had no effect on larval settlement and metamorphosis. DOPA at 10^{m 5} M and IBMX at 10^{m 6}-10^{m 4} M induced 41-83% larval settlement and ACH at 10^{m 7}-10^{m 5} M induced < 40% larval settlement. While the highest settlement rates were observed after 48 h exposure to the chemicals, most of the larvae settled within 24 h. Compounds at concentrations of 10^{m 3}-10^{m 2} M were either toxic to larvae or retarded the growth of the post-larvae shell. Juveniles resulting from induction by lower concentrations of chemicals had a very high survival rate, completed metamorphosis and grew as well as the juveniles that metamorphosed spontaneously. IBMX at 10^{m 6}-10^{m 4} M and L-DOPA at 10^{m 5} M are effective agents for induction of settlement and metamorphosis for future studies using juvenile M. edulis.     

Assessment of a glutathione S-transferase and related proteins in the gill and digestive gland of Mytilus edulis (L.), as potential organic pollution biomarkers

The response of the glutathione S-transferase (GST, EC 2.5.1.18) and related proteins of Mytilus edulis to environmental pollution load was assessed. Mussels were reciprocally transplanted between an industrial estuary (Douglas), a rural estuary (Youghal) and a m arine site (Bantry). In addition, m ussels were sam pled along a pollution gradient in an estuary receiving leather tannery effluent (Colligan). These latter m ussels were previously shown to be subject to oxidative stress resulting from the discharges. GST specific activity of cytosolic extracts from the gill and digestive gland tissues was determ ined for all anim als. Specific activity was shown to vary significantly in anim als from different sites, with highest specific activity always observed in sam ples (local and transplanted) taken from the industrial site. By com parison, the m ussels exposed to tannery discharges displayed no significant alteration in GST specific activity. Total intracellular glutathione (GSH) was also determ ined for sam ples taken from the Douglas and Youghal estuaries but no correlation with pollution load was observed. Using FPLC analysis, we observed no specific effect on the relative levels of the GST and the individual GST related proteins in gill or digestive gland sam ples from local or reciprocally transplanted anim als from Douglas or Youghal. The increase in GST specific activity observed in samples from the industrial estuary are indicative of a possible, specific inductive agent at this site. The results from the tannery site, by com parison, indicate that general oxidative stress does not result in elevated G ST specific activity in M. edulis.     

A metabolic model for the determination of shell composition in the bivalve mollusc, Mytilus edulis

Rosenberg. G. D. & Hughes, W. W. 1991 01 15: A metabolic model for the determination of shell composition in the bivalve mollusc, Mytilus edulis. Lethaia, Vol. 24. pp. 83–96. Oslo. ISSN 0024–1164. This research describes compositional variations within the shell of the extant mussel Mytilur edulis and proposes that they are produced by metabolic gradients within the shell-secreting mantle. Because we have previously proposed that the same metabolic gradients are responsible for variations in shell form (curvature), we establish here a model for molluscan shell growth integrating. for the first time. shell form and composition with mantle metabolism. The electron microprobe was used to measure the distribution of Mg. S, and Ca in the outer calcitic shell layer of sectioned. polished, and either A1- or C-coated shell. Mg/Ca and S/Ca ratios in the outer shell are respectively 1.25 and 1.40 times higher along slow-growing, commissure-umbo axes of high shell curvature and high metabolic activity than along rapidly growing axes of low curvature and low metabolic activity. The ratios within the inner surface of the calcitic shell layer decline most rapidly along commissure-umbo axes where mantle metabolic activity also declines rapidly. We reject the null hypothesis, generally at high levels of significance (1-tests. F-tests. regression analyses, and discriminant analysis. with p 4 0.01) that there is no difference in either Mg or S concentration in sections of the calcitic shell layer that differ in shell curvature and mantle metabolic activity. We conclude that calcium (mineral)-rich portions of shells are energctically less costly to produce than matrix or minor element-rich portions. in agreement with the proposal that natural selection favors mineral-rich shells because they are more efficient to produce than matrix-rich shells. Among-specimen differences are also highly significant (mixed model ANOVA). This confirms our assertion that paleontologists need to describe variations in skeletal composition among populations and throughout ontogeny as systematically as classical taxonomists describe morphology. if ever the environmental and the genetic influences on skeletal composition are to be distinguished. Bivalves. biomineralization, shell composition. magnesium, sulfur, calcium, metabolism, growth. Mytillus edulis     

Use of microstereology and quantitative cytochemistry to determine the effects of crude oil-derived aromatic hydrocarbons on lysosomal structure and function in a marine bivalve mollusc, Mytilus edulis

Summary The marine bivalve mollusc,Mytilus edulis (blue mussel), is a noted accumulator of many environmental pollutants and is increasingly used for the chemical and biological assessment of environmental impact. The toxic effects of crude oil-derived aromatic hydrocarbons (30 g/l total hydrocarbons) on the lysosomal-vacuolar system of the digestive cells have been investigated in cryostat sections of hexane-frozen digestive glands. Exposure to aromatic hydrocarbons reduced the cytochemically determined latency of lysosomal -N-acetylhexosaminidase; lysosomal volume density and surface density increased while the numerical density decreased. Experimental exposure resulted in the formation of very large lysosomes which are believed to be largely autophagic in function and these results indicate a significant structural and functional disturbance of digestive cell lysosomes in response to hydrocarbons.     

Alternate pathogenesis of systemic neoplasia in the bivalve mollusc Mytilus.

The proliferative disease systemic neoplasia, also termed hemic neoplasia or disseminated sarcoma, was studied in four Puget Sound, Washington populations of the bay mussel (Mytilus sp.). Using flow cytometric measurement of DAPI-stained cells withdrawn from the hemolymph, DNA content frequency histograms were generated for 73 individuals affected by the disease. The cells manifesting systemic neoplasia were found to exist as either of two separate types, characterized by G0G1 phase nuclear DNA contents of either approximately 4.9 x haploid (pentaploid form) or approximately 3.8 x haploid (tetraploid form). The two disease forms were found to coexist in all four mussel populations sampled, with overall relative prevalences of 66% pentaploid form, 29% tetraploid form, and 5% exhibiting both disease forms simultaneously. These findings represent the first unequivocal demonstration of multiple cell types in a bivalve neoplasia. The two forms appear to represent separate pathogenetic processes rather than sequential stages of a single pathogenesis. Two cell cycling parameters associated with proliferative activity were employed to compare the alternate forms: (i) the percentage of cells assigned to the DNA Synthesis (S) phase of the neoplastic cell cycle, and (ii) the proportion of neoplastic cell mitotic figures in hemocytological preparations. Mean values for both parameters were significantly higher for mussels with the tetraploid form of the disease, suggesting a higher rate of proliferation relative to the pentaploid form. Qualitatively, cells of the tetraploid form contained slightly lower nuclear and cytoplasmic volumes compared to those of the pentaploid form. An observed wide variation in neoplastic cell nuclear size within either disease form may reflect the distribution of cells in the G0G1, S, and G2M phases of the cell cycle. Potential etiologic relationships between the two forms are discussed.     

Introgression patterns in the mosaic hybrid zone between Mytilus edulis and M. galloprovincialis

Hybrid zones are fascinating systems to investigate the structure of genetic barriers. Marine hybrid zones deserve more investigation because of the generally high dispersion potential of planktonic larvae which allows migration on scales unrivalled by terrestrial species. Here we analyse the genetic structure of the mosaic hybrid zone between the marine mussels Mytilus edulis and M. galloprovincialis, using three length-polymorphic PCR loci as neutral and diagnostic markers on 32 samples along the Atlantic coast of Europe. Instead of a single genetic gradient from M. galloprovincialis on the Iberian Peninsula to M. edulis populations in the North Sea, three successive transitions were observed in France. From South to North, the frequency of alleles typical of M. galloprovincialis first decreases in the southern Bay of Biscay, remains low in Charente, then increases in South Brittany, remains high in most of Brittany, and finally decreases again in South Normandy. The two enclosed patches observed in the midst of the mosaic hybrid zone in Charente and Brittany, although predominantly M. edulis-like and M. galloprovincialis-like, respectively, are genetically original in two respects. First, considering only the various alleles typical of one species, the patches show differentiated frequencies compared to the reference external populations. Second, each patch is partly introgressed by alleles of the other species. When introgression is taken into account, linkage disequilibria appear close to their maximum possible values, indicating a strong genetic barrier within all transition zones. Some pre- or postzygotic isolation mechanisms (habitat specialization, spawning asynchrony, assortative fertilization and hybrid depression) have been documented in previous studies, although their relative importance remains to be evaluated. We also provided evidence for a recent migratory 'short-cut' connecting M. edulis-like populations of the Charente patch to an external M. edulis population in Normandy and thought to reflect artificial transfer of spat for aquaculture.     

Innervation of the anterior byssal retractor muscle in Mytilus edulis L.

Summary Studies on the intrinsic innervation of the anterior byssal retractor muscle (ABRM) in Mytilus edulis L. were continued at the ultrastructural level. Electron micrographs show nerve processes ensheathed by glio-interstitial cells running between muscle fibers. The glio-interstitial cells may represent all the types of osmiophilic cells previously described by the light microscopic ZIO technique in the anterior byssal retractor muscle.     

Contamination of a dredged material disposal site La Rochelle Bay,France .The use of the acetylcholinesterase activity of Mytilus edulis L. as a biomarker of pesticides: the need for a critical approach

A single aspect of the toxic impact of a dredged material disposal site located near a mussel-farming zone was followed for eight months. Acetylcholinesterase activity (AChE) of Mytilus edulis was investigated as a biomarker for possible contamination by neurotoxic compounds (carbamates and/or organophosphorous pesticides). Our observations showed that the enzymatic activities (including AChE) of these harbour mussels were decreased in sites directly and indirectly influenced (according to hydrodynamic conditions) by the dumping of dredged sediments, suggesting possible contamination by pesticides. The strong correlations observed between AChE activity and growth parameters (length and weight) seems to show, however, that the enzyme activity is also indirectly controlled through growth restriction, which may imply limitation of the development of the nervous system in juveniles. The concentration of total proteins, as well as the spawning process also seem to disturb the assessment of AChE activity. These field observations clearly indicate that the use of this enzyme activity as a biomarker should proceed with caution. For example, the seasonal variability of such activity should be taken into account in a biomonitoring programme.     

Assortative fertilization and selection at larval stage in the mussels Mytilus edulis and M. galloprovincialis

Assortative mating (prezygotic isolation) and reduced hybrid fitness (postzygotic isolation) are typically invoked to explain the stability of hybrid zones. In the tension zone model, these factors work in opposition to migration, which promotes genetic homogeneity. Many marine animals migrate over long distances through a planktonic larval stage. Therefore, strong reproductive isolation is needed to maintain stable marine hybrid zones. However, surprisingly little is known about mating preferences and hybrid fitness in marine organisms. Smooth-shelled mussels (Mytilus spp.) form a well-known species complex, with hybridization over extensive areas such as the contact zone of M. edulis and M. galloprovincialis around European Atlantic coasts. This paper reports direct experimental evidence of assortative fertilization, hybrid larval inviability, and early heterosis for growth rate in M. edulis and M. galloprovincialis. Four crosses between pure M. edulis and M. galloprovincialis were analyzed with a new polymerase-chain-reaction-based diagnostic marker. Gamete competition between taxa was allowed in two out of the four crosses. Genotype frequencies observed at an early stage (36 h after fertilization) unambiguously revealed assortative fertilization when gamete competition was allowed. A significant reduction in hybrid viability was subsequently observed during the larval stage. At the same stage an antagonistic effect, heterosis, was observed on growth rate. However, even if heterosis is observed in the F1, it is expected to vanish in subsequent hybrid generations. Although specialization for different habitats and asynchronous spawning have been mentioned as factors contributing to the maintenance of the blue mussel hybrid zone in Europe, we argue that assortative fertilization and reduced hybrid fitness are important factors that also contribute to the stabilization of this zone. These results emphasize that multiple factors may act concomitantly in a barrier to gene flow, especially in complex life cycles. Furthermore, they show that assortative mating through gamete preference, as already demonstrated for sea urchins, may play a role in speciation processes taking place in the sea.     

Electrophoretic investigation of systematic relationships in the marine mussels Modiolus modiolus L., Mytilus edulis L., and Mytilus galloprovincialis Lmk. (Mytilidae; Mollusca)

Genetic variation was assayed electrophoretically at 13–16 loci in Modiolus modiolus, Mytilus edulis, and Mytilus galloprovincialis. High genetic distance ( D ) values were observed between Modiolus modiolus and Mytilus edulis (1.516 ± 0.523) and between Modiolus modiolus and Mytilus galloprovincialis (1.564 ± 0.539), whereas the distance between Mytilus edulis and M. galloprovincialis (0.167 ± 0.118) was rather low. The systematic status ot Mytilus edulis and M. galloprovincialis is discussed in relation to these lindings and the genetic distance values are used to estimate divergence times which in turn are compared with paleontological estimates. The observations of high average heterozygosity in Modiolus modiolus, and high correlations of locus heterozygosity between taxa are discussed briefly.     

Oxygen consumption,ammonia excretion,and filtration rate of the marine bivalve Mytilus edulis exposed to methamidophos and omethoate

In this study, the oxygen consumption, ammonia excretion, and filtration rate were monitored in Mytilus edulis in response to administration of the two pesticides, methamidophos and omethoate. Five sublethal concentrations (1, 10, 50, 100, and 200 µgL^?1) were administered over 96 h. Oxygen consumption rates increased following administration of all concentrations of methamidophos for 96 h and to the lower concentrations of omethoate (1, 10, and 50 µg L^?1) from 6 to 24 h. Over 24 h, oxygen consumption decreased significantly. Ammonia excretion rates were higher than the control after 36 h of exposure to methamidophos, while reduction was observed when M. edulis was exposed to omethoate. The O:N ratios increased when the animals were first exposed to these two pesticides and then decreased at all the concentrations. At the concentrations of 100 and 200 µgL^?1, the O:N ratio was below 30 after 72 h. The filtration rate of M. edulis decreased with the increasing concentration of pesticide exposure.     

Influence of the antimitotic naphthoquinone on the cleavage of the eggs of the mollusc,Mytilus edulis L.

Summary 1. A brief account is given of the cleavage ofMytilus edulis L.2.Naphthoquinone in concentrations above 1 million produces cytolysis of the newly fertilised eggs. It causes some blockage of cleavage at the single cell stage in concentrations down to 140 M. The main area of its antimitotic activity is in the range between 1 M and 16 M. Resistance to the substance increases as cleavage proceeds. 3. Some abnormalities of cleavage are found at all antimitotic concentrations. The types of abnormality are described. They do not appear to fit easily into the hypothesis of Huber (1947) that the effect of this substance is to enhance the contraction and inhibit the expansion of the cell cortex.     

Biodeterioration of crude oil and oil derived products: a review

Biodeterioration of crude oil and oil fuels is a serious economic and an environmental problem all over the world. It is impossible to prevent penetration of microorganisms in oil and fuels both stored in tanks or in oilfields after drilling. Both aerobic and anaerobic microorganisms tend to colonise oil pipelines and oil and fuel storage installations. Complex microbial communities consisting of both hydrocarbon oxidizing microorganisms and bacteria using the metabolites of the former form an ecological niche where they thrive. The accumulation of water at the bottom of storage tanks and in oil pipelines is a primary prerequisite for development of microorganisms in fuels and oil and their subsequent biological fouling. Ability of microorganisms to grow both in a water phase and on inter-phase of water/hydrocarbon as well as the generation of products of their metabolism worsen the physical and chemical properties of oils and fuels. This activity also increases the amount of suspended solids, leads to the formation of slimes and creates a variety of operational problems. Nowadays various test-systems are utilized for microbial monitoring in crude oils and fuels; thus allowing an express determination of both the species and the quantities of microorganisms present. To suppress microbial growth in oils and fuels, both physico-mechanical and chemical methods are applied. Among chemical methods, the preference is given to substances such as biocides, additives, the anti-freezing agents etc that do not deteriorate the quality of oil and fuels and are environmentally friendly. This review is devoted to the analysis of the present knowledge in the field of microbial fouling of crude oils and oil products. The methods utilized for monitoring of microbial contamination and prevention of their undesirable activities are also evaluated. The special focus is given to Russian scientific literature devoted to crude oil and oil products biodeterioration.     

Growth and longevity of Mytilus edulis (L.) from northeast Europe

The growth of mussels, Mytilus edulis (L.), was studied in most of the northeastern part of their distribution. The growth, longevity and maximal size of molluscs from 24 wild populations and one cultured population located in the White Sea and the southeast part of the Barents Sea were compared. The 25 studied populations were combined in six clusters. The maximal longevity and the size of the mussels varied between 7 and 18 years and 25.5 and 77.7 mm, respectively. The geographical location of the population within the studied region did not affect either maximal longevity or maximal size, or the growth rate of mussels. However, these parameters were influenced by local habitat conditions, primarily connected with the position within intertidal and subtidal zones. Animals inhabiting the intertidal zone were characterized by relatively low growth performance, a short life span and a small size. The longest life span was typical for deep subtidal mussels, whereas the highest growth rate was recorded in the cultured population and in the upper subtidal habitats. Growth patterns of Mytilus edulis in the subarctic White and the Arctic southeast Barents seas are similar to those reported from other parts of the area of distribution. Therefore, growth was mostly determined by local environmental factors, including those related to vertical zonation, rather than by latitude/longitude and related temperature effects.     

Turnover rate of metallothionein and cadmium in Mytilus edulis

The results demonstrate the first attempt to determine metallothionein turnover in the whole soft tissues of mussels Mytilus edulis exposed to cadmium. Half-lives for metallothionein and cadmium are 25 and 300 days, respectively. As metallothionein degrades the released cadmium induces further synthesis of the protein, to which the metal becomes resequestered. The slow metallothionein turnover rates (compared with mammals) and the lack of significant cadmium excretion testify to the relatively stable nature of the cadmium-metallothionein complex in these invertebrates and supports the view of a detoxifying role for metallothionein in the mussels.     

On the ultrastructure of polypeptide hormone-producing cells in the gut of the ascidian,Ciona intestinalis L. and the Bivalve,Mytilus edulis L.

Summary Ultrastructural evidence has been found for the presence of polypeptide hormone-producing cells in the gut of Ciona intestinalis L. and Mytilus edulis L. which do not appear to have been described before. Due to their localization and ultrastructural characteristics, it is suggested that the cells in Mytilus edulis probably produce an insulin-like substance and that some of these cells in Ciona intestinalis may produce 5-HT (5-Hydroxytryptamine). In each species only one granulated cell type can be observed. The granules, which are electron dense and membrane bound, also show a halo. The average diameter of the granules is 100–200 nm for Ciona and 200–400 nm for Mytilus.I thank Mr. G. Bargsten, M.A., Dept. of Marine Zoology, University of Kiel, for the supply of the animals