State of the antioxidant enzyme complex in tissues of the Black Sea mollusc <Emphasis Type="Italic">Mytilus galloprovincialis</Emphasis> under natural oxidative stress

The state of the antioxidant (AO) system (AOS) and the content of products of lipid peroxidation (LPO) in hepatopancreas, gill, and foot of the Black Sea mollusc Mytilus galloprovincialis Lam. were studied in norm and under conditions of natural oxidative stress (spawning). It has been established that under conditions of spawning, the AO-protection of hepatopancreas cells is predominantly oriented to inactivation of high concentrations of peroxide compounds with participation of glutathione peroxidase and catalase. In the foot tissue, on the contrary, the main role in AO-processes is played by enzymatic inhibition of superoxide anion radical with aid of superoxide dismutase. The highest level of oxidative stress in the spawning state is revealed in the mollusc gill, which is indicated by the maximal increase of the content of TBA (thiobarbituric acid)-active products.     

Functional states of antioxidant enzymatic complex of tissues of Mytillus galloprovincialis Lam. under conditions of oxidative stress

Materials are summarized on reaction of enzymatic antioxidant (AO) complex of tissues molluscs of the genus Mytilus on oxidative load of different intensity. It is shown that with increase of the level of oxidative stress there is observed a rise of activities of glutathione reductase (GR) and glutathione peroxidase (GP) as well as increases the turnover rate of glutathione (GSH) in tissues. Then the toxic load is compensated by an elevation of activities of enzymes of low affinity to hydroperoxides, specifically of catalase. In the glutathione system it is possible to identify several relatively steady states: processes of low, moderate, and high intensity. In individual tissues, functioning of the AO system seems to be directed not to detoxication, but to generation of reactive oxygen species (O₂ ^·?, superoxide dismutase), which probably is determined by specifics of their running processes (destruction of damaged byssus threads). The AO complex of molluscs is distinguished by high lability and sensitivity both to physiological states (spanning) and to action of factors of anthropogenic nature (cationic detergents). Responses of the mussel AO complex to natural states and toxic action are comparable between each other, which are necessary to be taken into account at diagnostics of the state of the aquatic environment.     

Antioxidant enzyme complex of tissues of the bivalve Mytilus galloprovincialis Lam. under normal and oxidative-stress conditions: a review

The results of studies of tissue specifics of the enzymatic antioxidant complex of the bivalve Mytilus galloprovincialis Lam. are summarized. It is shown that the highest oxidative load is experienced by gills. The antioxidant complex of gills largely depends on environmental conditions than on the mollusk's state, which allows this tissue to be used for ecological diagnostics. A decrease in the content of carotenoids in tissue s suppressed the activities of the key enzymes of antiradical defense-superoxide dismutase (SOD) and catalase-and is accompanied by a decrease in the reduced glutathione (GSH) pool. The state of the antioxidant complex of molluscan tissues under conditions of natural (spawning) and artificial (exposure to a cationic surfactant) oxidative stress was studied.     

Microflora of the Black Sea mussel Mytilus galloprovincialis L

The microbial cenosis in the mantle fluid and stomach of mussels is similar in its biochemical characteristics to the microflora of sea water.     

The activity of antioxidant defence enzymes in the mussel Mytilus galloprovincialis from the Adriatic Sea

The activity of the antioxidant defence enzymes superoxide dismutase (SOD, EC 1.15.1.1), catalase (CAT, EC 1.11.1.6), glutathione peroxidase (GSH-Px, EC 1.11.1.9), glutathione reductase (GR, EC 1.6.4.2) and the phase II biotransformation enzyme glutathione-S-transferase (GST, EC 2.5.1.18) in whole mussels (Mytilus galloprovincialis) were studied. The mussels were collected in winter and in spring at two localities in the Adriatic Sea: Bar Port and Tivat Bay. Our results show that the activities of SOD, GSH-Px and GST were seasonally dependent with higher activities in winter. GR activity was also higher in winter, but only in mussels from Bar Port. In mussels from Tivat Bay, GR activity was lower in winter compared to spring. In addition, a decrease in CAT activity in mussels from Bar Port compared to those from Tivat Bay was found. It can be concluded that seasonal variations should be incorporated into interpretation of biomonitoring studies in mussels.     

Purification and characterization of hyaluronic acid from the mollusc bivalve Mytilus galloprovincialis

Hyaluronan (hyaluronic acid, HA) was for the first time extracted, purified and characterized from the species of mollusc bivalve Mytilus galloprovincialis. HA was characterized by agarose-gel electrophoresis, 13C-NMR, HPLC and normal polarity capillary electrophoresis by evaluating the unsaturated disaccharide, DeltaDiHA (Delta-hexuronic acid-N-acetyl-glucosamine) after treatment with chondroitin ABC lyase, and by separating Delta-tetrasaccharide and Delta-hexasaccharide generated by the specific action of hyaluronate lyase from Streptomyces hyalurolyticus. The weight average molecular weight (M(w)) was found to be about 200 kDa as determined by HPSEC. HA from M. galloprovincialis was not able to interact with aggrecan from bovine cartilage to form high molecular mass aggregate and also had a very low specific viscosity, but it showed the same capacity to inhibit cell proliferation (50 microg per 10(3) human fibroblasts inhibit cell proliferation by about 50%) than high molecular mass HA. HA of M. galloprovincialis could have a physiological role in the regulation of cell functions.     

On Temporal Dynamics of Morphological Variations in the Black Sea Mussel Mytilus galloprovincialis Lam.

Temporal variations in morphological characters of the shells were studied in 12 samples of mussels M. galloprovincialis Lam. from the Black Sea commercial collectors that belong to different generations. The samples were shown to be diverse in every studied character and the degree of characters' variability in the samples was different. The morphological diversity of samples diminished with age. Different generations from the same habitat had specific features of age-related variability expressed as different rates of age-related changes and direction and degree of variability. This effect is considered as an index of the influence of ecological conditions on the shell growth and formation.     

Peculiarities of antioxidant defense system organization of the Black Sea mollusks Mytilus

Antioxidant (AO) system and lipid peroxidation (LP) in tissues of two species of the Black Sea bivalve mollusks Mytilus galloprovincialis and Anadara inaequivalvis were investigated. The activity of superoxide dismutase (SOD, 1.15.1.1), catalase (1.11.1.6), glutathione peroxidase (GP, 1.11.1.9), glutathione reductase (GR, 1.6.4.2), concentrations of reduced glutathione (GSH) and TBA-reactive products were determined in the foot, hepatopancreas and gills of mature mollusks. The characteristics of AO complex and LP products connected with tissue and species specificity of mollusks were found. Hepatopancreas of mussels has been found to have higher values of all characteristics investigated, except GP. The gills and the foot of anadara have been found to be involved in AO defense along with hepatopancreas: maximum activity of GR, catalase and SOD was found in the gills and the highest activity of GP and maximum level of GSH was observed in the foot. Anadara has been shown to have higher antioxidant potential and lower level of oxidative stress because the LP intensity in all tissues examined of the hemoglobin-containing mollusk was twice lower in comparison with the mussel.     

Cadmium accumulation in tissues of the mussel Mytilus galloprovincialis

Studies have been made on cadmium accumulation in tissues of mussels kept within 20-60 days in water artificially enriched by Cd up to 20-100 micrograms/l. Irrespectively of cadmium concentration in the medium, its accumulation in tissues decreases in the following order: mid-gut gland, gills, gonads, mantle, adductor. Maximum concentration of Cd was found in the digestive tubuli of the mid-gut gland by X-ray microanalysis. The increase in S and, to a lower extent, P concentrations in these tubuli was also observed. It is suggested that the latter is due to immobilization of Cd by metal-binding proteins as well as to lyzosomal vesicles involved into detoxication of Cd. The increase in the external cadmium up to 100 micrograms/l did not affect the level of K, Ca and Mg in tissues of the mussel.     

Assessment of environmental pollution at Balearic Islands applying oxidative stress biomarkers in the mussel Mytilus galloprovincialis

The antioxidant enzyme response of the mussel Mytilus galloprovincialis to different degree of pollution was investigated. Antioxidant enzyme activities - catalase (CAT), glutathione peroxidase (GSH-PX), glutathione reductase (GR), superoxide dismutase (SOD) - and malondialdehyde (MDA) concentration were measured in gills and digestive glands of mussels. Mussels from the same origin were transplanted along the Balearic coastal waters in eight stations characterized by a different degree of contamination and human impacts. Antioxidant enzyme activities showed an adaptive response to increase the activities in the more polluted areas. CAT, GR and SOD in gills and CAT and GR in digestive gland presented significant differences between polluted and non-polluted stations. No significant differences were observed in MDA concentration indicating that the antioxidant response is capable to avoid the lipid peroxidation. The use of biomarkers such as CAT and GR in gills and digestive glands of the mussel M. galloprovincialis is a good tool to categorize differences between polluted and non-polluted areas.     

Identification of multiple isoforms of the cAMP-dependent protein kinase catalytic subunit in the bivalve mollusc Mytilus galloprovincialis

Several isoforms of the cAMP-dependent protein kinase catalytic subunit (C-subunit) were separated from the posterior adductor muscle and the mantle tissues of the sea mussel Mytilus galloprovincialis by cation exchange chromatography, and identified by: (a) protein kinase activity; (b) antibody recognition; and (c) peptide mass fingerprinting. Some of the isozymes seemed to be tissue-specific, and all them were phosphorylated at serine and threonine residues and showed slight but significant differences in their apparent molecular mass values, which ranged from 41.3 to 44.5 kDa. The results from the MS analysis suggest that at least some of the mussel C-subunit isoforms arise as a result of alternative splicing events. Furthermore, several peptide sequences from mussel C-subunits, determined by de novo sequencing, showed a high degree of homology with the mammalian Calpha-isoform, and contained some structural motifs that are essential for catalytic function. On the other hand, no significant differences were observed in the kinetic parameters of C-subunit isoforms, determined by using synthetic peptides as substrate and inhibitor. However, the C-subunit isoforms separated from the mantle tissue differed in their ability to phosphorylate in vitro some proteins present in a mantle extract.     

Commensals of Mytilus galloprovincialis in the Black Sea: Urastoma cyprinae (Turbellaria) and Polydora ciliata (Polychaeta)

The intensity of infection of the mussel Mytilus galloprovincialis Lamarck in the Black Sea by the turbellarian Urastoma cyprinae (Graff), which lives on its gills, was found to be higher in larger hosts, reaching a maximum in mussels of 50–70 mm length. Greater numbers occured in mussels inhabiting a silty bottom than in cultivated mussels suspended above the bottom. Over the period 1982–1987, U. cyprinae was most numerous in winter and especially so in years that were colder. The spionid polychaete Polydora ciliata (Johnston) also infects M. galloprovincialis, burrowing into the shell. Young spionids of up to 1 mm length occured in mussels with a shell length of 35 mm. Numbers of this commensal reached a maximum in mussels of intermediate size.     

Age composition and antioxidant enzyme activities in blood of Black Sea teleosts

Age composition and age-related trends of antioxidant enzyme activities superoxide dismutase (SOD), catalase (CAT), peroxidase (PER), glutathione reductase (GR) and glutathione-S-transferase (GST) in the blood of seven Black Sea teleosts (Carangidae, Centracanthidae, Gadidae, Mullidae, Gobiidae and Scorpaenidae) collected in marine coastal area of Sevastopol (Ukraine) were studied. In the catches the animals of 1 – 2 years of age dominated while in the Scorpaena porcus population the number of relatively elder individuals belonging to classes of 3–4 years was the highest. The trends of antioxidant enzyme activities in blood were not uniform. Three types of age-dependent responses were indicated in fish blood: 1. enzymatic activity did not change with age; 2. enzymatic activity decreased with age and 3. enzyme activity increased with age or varied unclearly. The interspecies differences of age-related enzymatic activities associated with the specificity of fish biology and ecology were indicated. Despite no clear evidence of age-related differences between fish species belonging to different ecological groups both benthic forms exhibited similar age-dependent trends of SOD and PER. The correlations between blood antioxidant enzyme activities in fish belonging to suprabenthic and benthic/pelagic groups demonstrated the intermediate values as compared to the benthic and pelagic forms. The results suggest the importance of age trends for biomarkers in fish monitoring studies.     

The formation of a complex habit in Black Sea bottle-nosed dolphins under free-choice conditions]

In experiment situation was created of "free choice", which was characterized by the reinforcement of any succession of bottle-nosed dolphin action on the manipulators, placed in the open-air cage under the condition of pressing three definite levers in the required order independently of the number of "superfluous" reactions before and between necessary actions. In the first series of the experiments with eight above-water levers no succession of actions leading to the reinforcement was formed in the bottle-nosed dolphin [correction of aphaline]. In the second series with eight under-water manipulators--both experimental dolphins regularly reached the reinforcement. Stereotype trajectories of movements in the cage were formed, which included swimming past the levers and successive actions on them. Minimum chain of motor reactions was not formed, though some of "superfluous" levers were not pressed. At solving the task efferent generalization in dolphins in both series of experiments was manifested in two ways: the animals reacted in different ways on one and the same manipulator and pressed different levers.     

Effects of seawater acidification and salinity alterations on metabolic,osmoregulation and oxidative stress markers in Mytilus galloprovincialis

The impacts of seawater acidification and salinity shifts on metabolism, energy reserves, and oxidative status of mussels have been largely neglected. With the aim to increase the current knowledge for the mussel Mytilus galloprovincialis a 28-day chronic test was conducted during which mussels were exposed to two pH (7.8 and 7.3; both at control salinity 28) and three salinity (14, 28 and 35, at control pH, 7.8) levels. After exposure to different conditions, mussels electron transport system activity, energy reserves (protein and glycogen content) carbonic anhydrase activity, antioxidant defences and cellular damage were measured. Results obtained showed that mussels exposed to seawater acidification presented decreased metabolic capacity that may have induced lower energy expenditure (observed in higher glycogen, protein and lipids content at this condition). Low pH condition induced the increase of carbonic anhydrase activity that was related to acid-base balance, while no significant activation of antioxidant defence mechanisms was observed resulting in higher LPO. Regarding the impacts of salinity, the present study showed that at the highest salinity (35) mussels presented lower metabolic activity (also related to lower energetic expenditure) and an opposite response was observed at salinity 14. Carbonic anhydrase slightly increased at stressful salinity conditions, a mechanism of homeostasis maintenance. Lower metabolic activity at the highest salinity, probably related to valves closure, helped to mitigate the increase of LPO in this condition. At low salinity (14), despite an increase of antioxidant enzymes activity, LPO increased, probably as a result of ROS overproduction from higher electron transport system activity. The present findings demonstrated that Mytilus galloprovincialis oxidative status and metabolic capacity were negatively affected by low pH and salinity changes, with alterations that may lead to physiological impairments namely on mussels reproductive output, growth performance and resistance to disease, with ecological and economic implications.Indicators: Physiological and biochemical changes in Mytilus galloprovincialis in response to low pH and salinity changes     

Comparative characteristics of the antioxidant glutathione complex in the black sea molluscs Mytilus galloprovincialis Lam. and Anadara inaequivalvis Br.

The peculiarities in distribution of lipid peroxidation (LPO) products, glutathione (GSH) level, and antioxidant enzymes-glutathione peroxidase (GP) and glutathione reductase (GR)—were studied in tissues of Black Sea bivalve molluscs, Anadata inaequivalvis Br. (anadara) and Mytilus galloprovincialis Lam. (mussel, black morph), as well as their comparative characteristics were presented. The differences were established in organization of the glutathione anti-oxidant system and the LPO intensity in tissues of these mollusc species. In all anadara tissues the intensity of LPO processes was lower than that in Mytilus galloprovincialis. The GP activity in hepatopancreas and gills of mussels was significantly higher than that of anadara. On the contrary, in the foot the GP activity and GSH content in anadara considerably exceeded those in mussel. The revealed differences might reflect the peculiarities in functioning of the glutathione complex and the ratio of its activity with LPO level in tissues of anadara and mussel, as well as be of interest for understanding mechanisms of mollusc adaptation to their habitat conditions.     

Effect of thermal stress on protein expression in the mussel Mytilus galloprovincialis Lmk

The exposure of organisms to stressing agents may affect the level and pattern of protein expression. Certain proteins with an important role in protein homeostasis and in the tolerance to stress, known as stress proteins, are especially affected. Different tissues and cells show a range of sensitivities to stress, depending on the habitat to which organisms have adapted. The response of different tissues and cells from the mussel Mytilus galloprovincialis Lmk. to heat shock has been studied in this work using different exposure times and temperatures. During the assays, protein expression was observed to vary depending on the tissue studied, the temperature or the exposure time used. But maybe the most prominent thing is the different response obtained from the cultured haemocytes and those freshly obtained from stressed mussels, which makes us think that the extraction procedure is the main cause of the response of non-cultured cells, although the haemolymph may contain components that modulate haemocyte response.     

Shell morphometrics and growth rate of the invasive bivalve mollusc Mytilus galloprovincialis in the Knysna estuarine embayment,South Africa

Shell morphometrics of the invasive mussel Mytilus galloprovincialis were compared at five sites and growth rate at four sites (in four seasons) in the Knysna estuarine embayment. Mussels from two sites (The Heads, Leisure Isle) where wave action was present had shells significantly lower for any length when compared with other more sheltered sites. There was no significant difference in shell width of mussels for any given length among sites. Mussels from The Heads had thicker shells than other sites, and those from Leisure Isle thicker shells than three other embayment sites where shells did not differ in thickness. Growth rate of mussels at two embayment sites (Thesen’s Wharf and Thesen Islands Marina) was greatest in autumn and summer whereas at The Heads and Leisure Isle there was little seasonal difference in growth rate. Growth rate of mussels at Thesen’s Wharf and Thesen Islands Marina was mainly greater in all seasons when compared with mussels at The Heads and Leisure Isle. The more rapid growth rate of mussels at the sheltered embayment sites might in part explain why M. galloprovincialis now dominates the mid- to lower intertidal on hard substrata in this region of the Knysna estuary.