Aleksandras Rybakovas

Environmental genotoxicity and cytotoxicity in the offshore zones of the Baltic and the North Seas

Micronuclei (MN), nuclear buds (NB) and fragmented-apoptotic cells (FA) were analyzed in mature peripheral blood and immature cephalic kidney erythrocytes of flounder (Platichthys flesus), dab (Limanda limanda) and cod (Gadus morhua) from 12 offshore sites in the Baltic Sea (479 specimens) and 11 sites in the North Sea (291 specimens), which were collected during three research vessel cruises in December 2002, 2003 and in September 2004. The highest levels of environmental genotoxicity (frequencies of MN up to 0.5 per thousand, NB - up to 0.75 per thousand) and cytotoxicity (FA - up to 0.53 per thousand) were observed in flatfishes from areas close to oil and gas platforms in the North Sea and in zones related to the extensive shipping and potentially influenced by contamination from large European Rivers (Elbe, Vistula, Oder). In dab from the offshore zones of the North Sea, the levels of nuclear abnormalities were higher as compared to those in dab from the Baltic Sea. Responses in immature kidney erythrocytes were higher than in mature erythrocytes from peripheral blood. MN frequency lower than 0.05 per thousand (the Baltic Sea) and lower than 0.1 per thousand (the North Sea) could be suspected as a reference level in the peripheral blood erythrocytes of flatfish.

Biomarker responses in flounder Platichthys flesus from the Polish coastal area of the Baltic Sea and applications in biomonitoring

The objective of this study was to investigate the pattern of enzymatic activities, environmental genotoxicity and cytotoxicity in flounder, Platichthys flesus, from the Polish coastal area of the Baltic Sea. Fish were sampled in different contaminated sites in the Gulf of Gdansk and in a reference area outside the gulf. The activities of acetylcholinesterase (AChE), glutathione s-transferase (GST), catalase (CAT), alanine aminotransferase (ALT), aspartate aminotransferase (AST), creatine kinase (CK), lactate dehydrogenase (LDH), were studied, as well as the frequency of micronuclei, nuclear buds and fragmented-apoptotic cells. A higher mean activity level of muscular AChE and a lower activity level of hepatic GST were evident in samples taken from the reference site, relative to those found in the gulf. Modeled CAT activity (in both liver and gill tissue), blood plasma LDH and CK activities were all significantly higher in flounder collected at locations within the Gulf of Gdansk than at the reference site. No statistically significant alterations were observed in the activities of ALT and AST in the blood plasma of flounder in this study. Fish collected from a location at the mouth of the Vistula River showed the highest hepatic GST and CAT, the highest gill CAT activity, and the highest frequency of blood micronuclei, nuclear buds and fragmented-apoptotic cell inductions, as well as the lowest level of blood plasma CK. The present study confirms that compared to fish from the reference area, flounder from the Gulf of Gdansk clearly demonstrate a different enzyme activity, genotoxicity and cytotoxicity biomarker response pattern.

Risk of environmental genotoxicity in the Baltic Sea over the period of 2009-2011 assessed by micronuclei frequencies in blood erythrocytes of flounder (Platichthys flesus), herring (Clupea harengus) and eelpout (Zoarces viviparus).

Environmental genotoxicity was investigated at 82 locations encompassing different regions of the Baltic Sea. Micronuclei (MN) analysis was performed in erythrocytes of 1892 specimens of flounder Platichthys flesus, herring Clupea harengus and eelpout Zoarces viviparus, three of the most common native fish species of the Baltic Sea collected in 2009-2011. MN background levels in fish were determined using data obtained in 2001-2011 from 107 Baltic sites. Extremely high genotoxicity risk zones were found for flounder at 11 stations out of 16 in 2009 and 33 stations of 41 in 2010-2011, for herring, at 5 of 18 stations in 2009 and 20 of 43 stations in 2010-2011, in eelpout only at one out of 29 stations. The sampling stations were restricted mainly to the southern and eastern Baltic Sea offshore zones and in most of them, MN frequencies in flounder and herring significantly exceeded the reference and background levels of micronuclei. This is a first attempt to evaluate the background MN responses, as well as low, high and extremely high genotoxicity risk levels for native fish species.

Environmental genotoxicity and cytotoxicity levels in fish from the North Sea offshore region and Atlantic coastal waters.

In the framework of the ICON project, environmental genotoxicity and cytotoxicity levels were assessed in blood erythrocytes of dab (Limanda limanda) and haddock (Melanogrammus aeglefinus) collected at 25 stations in the North Sea and near the coast of Iceland in August-October 2008. Micronuclei, nuclear buds and bi-nucleated cells with nucleoplasmic bridges were assessed as environmental genotoxicity biomarkers, and the frequency of fragmented-apoptotic and bi-nucleated erythrocytes were assessed as environmental cytotoxicity biomarkers. The lowest frequencies of genotoxic and cytotoxic abnormalities were detected in fish from the Icelandic study stations. The highest frequencies of abnormalities were recorded in dab from the Dogger Bank and the German Bight, in haddock from the Egersund Bank and from an area off the Firth of Forth (North Sea). In fish from the Icelandic reference area, frequencies of genotoxicity and cytotoxicity responses were significantly lower than in fish from most areas of the North Sea.

Environmental Mutagenesis in different zones of the Baltic Sea

Environmental mutagenesis was assessed in fish from the selected coastal areas of the Baltic SeanKvadofjarden and the Stockholm archipelago (Sweden), the KlaipodanBstingo zone (Lithua- nia), the Gulf of Gdansk (Poland) and the Wismar Bay (Germany). The frequency of micronuclei, as markers of cytogenetic damage, was evaluated in erythrocytes of 911 flounder (Platichthys flesus), 332 perch (Perca fluviatilis) and 458 eelpout (Zoarces viviparus) specimens. Fish was collected du- ring bi-annual sampling campaigns performed in 2001 and 2002. Statistically significant differences (p < 0.0001) regarding season and sampling location were recorded. The highest values of cytogenetic damage were registered in flounder from the Lithuanian coast near Palanga (autumn 2001) and from the Wismar Bay (spring 2001), in perch from the Lithuanian coast near Nemirseta (autumn 2001) and in eelpout from the Wismar Bay (autumn 2001). These values are 5n10-fold higher than the baseline level of micronucleus incidence. In general, the decrease in environmental mutagenesis was observed in 2002. Nevertheless a significant increase in micronucleus levels was determined in fish after the oil spill in the Bstingo oil terminal.

Genotoxic and cytotoxic effects in the bivalve mollusks Macoma balthica and Mytilus edulis from the Baltic Sea@@@Genotoksinio ir citotoksinio poveikio įvertinimas Baltijos jūros dvigeldžių moliuskų Macoma balthica ir Mytilus edulis ląstelėse

Micronuclei (MN), nuclear buds (NB), fragmented-apoptotic (FA) and bi-nucleated (BN) cells were analysed in gills of bivalve mollusks Macoma balthica and Mytilus edulis collected from fi ve study locations in Lithuanian territorial waters of the Baltic Sea. Th e frequency of micronuclei varied from 1.28 to 3.63‰ (MN/1000 cells), of nuclear buds – from 0.50 to 1.49‰, fragmented-apoptotic cells from 0.53 to 1.72‰ and of bi-nucleated cells from 1.51 to 2.23‰. Th e highest level of MN and bi-nucleated cells was determined in mollusks from the off shore Station 65, nuclear buds and fragmented-apoptotic cells from Station 1B located close to the Būtingė oil terminal. A comparatively high frequency of MN was observed in bivalves from the Būtingė oil terminal area – 3.38‰ in Station 1B, 2.85‰ in Station B06 in June and in mollusks from Station N-4 (2.66‰) located nearby the oil platform D-6.

Cytogenetic and cytotoxic Effects in Gill Cells of Blue Mussels (Mytilus Edulis) from the Baltic Coast After 1–3 Day maintenance in Laboratory Conditions

Frequency of micronuclei (MN) and other nuclear abnormalities (nuclear buds, bi-nucleated and fragmented-apoptotic cells) were analysed in gill cells of blue mussels Mytilus edulis. One group of mussels were dissected immediately after collection from the Baltic Sea (Lithuanian coast near Palanga and Būtingė) and other groups were kept in laboratory tanks for 1–3 days without aeration and feeding. Each study group consisted of 10 mussel individuals. Study results showed that after maintenance of mussels in the laboratory, frequencies of MN and bi-nucleated cells did not increase significantly. Nevertheless, a significant increase in fragmented-apoptotic cells was recorded in mussels kept in the laboratory for 2, 2.5 and 3 days. Decreased values of nuclear buds were observed in mussels after 1–3-day maintenance in the laboratory, whilst statistical significance was reached only in the Būtingė group after 1-day exposure.

Biological effects of environmental pollution in fish and mussels inhabiting Klaipėda-Būtingė area (Baltic Sea)

In June and September 2001 and 2002, acetyl cholinesterase activity, metallothioneins level, bile PAH-metabolites, lysosomal membrane stability, DNA adducts, micronuclei, other nuclear abnormalities, immunological response and histopathological incidences were measured in blue mussel (Mytilus edulis) and flounder (Platichthys flesus) collected along a suspected pollution gradient in the Lithuanian coastal zone (near Nemirseta, Būtingė and Palanga). The biomarkers in flounder from the Klaipėda offshore (reference site) zone were analyzed in December 2001 and 2002. The study results showed the lowest level of genotoxicity and cytotoxicity in organisms inhabiting offshore and Palanga localities in spring 2001. The Palanga zone was originally considered as a coastal reference site, whilst an oil spill occurred in the Būtingė oil terminal in November 2001, causing contamination of this area. After the oil spill the significant increase in genotoxicity and cytotoxicity was detected in mussels and flounder from the Palanga and Būtingė localities. The levels of PAH metabolites, lysosomal membrane instability, as well as neoplastic lesions increased also significantly in flounder after the oil spill. Further elevation of cytogenetic damage caused by the oil spill in mussels was also observed in June 2003. Nevertheless, recovery of flounder was shown by some biomarker responses 10 months after the oil spill.