DNA damage in gill cells of Corbicula japonica exposed to natural and anthropogenic stressors
Bivalve mollusks are sensitive biomarkers of aquatic ecosystem pollution. The impact of human activities on the environmental is an ongoing and increasing problem. There are many potentially dangerous chemicals that dissolve in water. Aquatic organisms are exposed to these chemicals, which can lead to morphological alterations and change certain physiological processes in their organs. The monitoring of changes in various biochemical parameters at the individual species level of organisms may be useful for identifying and delineating the impact of pollutants. C. japonica have been regarded as the most useful sentinels for chemical pollution studies in the aquatic environment. They are filter-feeding organisms able to bioaccumulate and concentrated most pollutants even if these are present at fairly low concentrations. Several bivalves, such as C. japonica have proven to be potential bioindicators.
Oxidative DNA damage is considered to be one of the crucial steps in ageing and disease. Various factors such as chemical, physical and human activity inducting oxidative stress are known to cause DNA strand breaks through oxygen species and free radical action. The increasing genotoxic risk in marine environmental is motivating intense and continuous ecosystem research in an effort to develop and apply new biomonitoring tools. As a molecular biomarker for genetic effects DNA damage, integrity and repair processes introduce a highly specific and very early signal of genotoxic effects in some organisms. Under the constant influence of various stressors on living organisms, measured DNA damage, as a point in space-time, represent the dynamic state between constant DNA damage that occurs and its repair. In such cases DNA damage could reflect the level of marine pollution or human activity.
The Comet assay or single gel electrophoresis (SCGE) assay is a simple, sensitive and rapid technique for detection of DNA damage in individual cells, and therefore, can be very useful in studies of genetic toxicology, especially ecogenotoxicology.
Genotoxic damage, expressed as DNA strand breaks, was measured in cells isolated from gill from the C. japonica using the single gel electrophoresis (Comet assay). The gill tissue may have shown the highest level of DNA damage as substantial quantity of contaminated water, because of the water interface passed over the gill during the breathing, continuosly exposing the gill membranes. The mollusks were collected from polluted (estuary Razdolnaya river, lagoon Tihaya) and a ‘clean’ areas (estuary Artemovka river, estuary Partizanskaya river) from Peter the Great Bay.
A total of 100 nuclei from each of the two replicates were examined and classified in one of the five damage classes, as described by Mitchelmore (1998), according to the migration distance and the fluorescence rate between the head and the tail of the nucleus: Class 0, intact nucleus with no migrated fragments (<5% fragmented DNA); Class 1, dense nucleus with slight DNA migration forming a small tail (5–20% fragmented DNA); Class 2, tail extending from the nucleus, with a weaker fluorescence than Class 2 (20–40% fragmented DNA); Class 3, comets with a clear tail that may reach the full length (40–75% fragmented DNA); and Class 4, nucleus, when present, is small and completely separated from the tail (>75% fragmented DNA).
The results are presented as the percentage distribution of nuclei in the various damage classes and summarized in an index of DNA integrity or genetic damage index GDI) (Cavas, Kohen, 2008):
The results from our study showed significant level of DNA damage from the C. japonica which were collected from polluted sites. Unpolluted sites were described as with no or minimal DNA strand breaks. Mollusks collected at polluted areas (estuary Razdolnaya river, lagoon Tihaya) showed high levels of DNA damage, GDI is equal to 3.22±0.2 and 3.11±0.7 in gills respectively. C. japonica obtained from a ‘clean’ areas (estuary Artemovka river, estuary Partizanskaya river) demonstrate less high level of DNA damage destruction, GDI is equal to 0,6±0,08 and 0,71±0,12) in gills respectively.
Lipid peroxidation level was assayed measurement of malondialdehyde (MDA), a decomposition product of polyunsaturated fatty acids hydro peroxides were determined by the TBA reaction. The absorbance was read at 532 nm after removal of substances (TBARS) formed was calculated by using an extinction coefficient of 1.56*105 M-1 cm -1 formed per g dry weight.
C. japonica sampled at Artemovka estuary, Partizanskaya estuary, Razdolnaya estuary, Tihaya lagoon showed LPX level (3.46±0.59, 5.62±0.82, 12.85±0.52, 15.32±1.13 nmol TBARS/g dry wt) in gills respectively.
In conclusion, it can be noted that in the course of the experiment we found a clear relationship between the amount of DNA damage and the level of peroxidation products (MDA) in the gills of bivalve C. japonica, collected from sites with varying degrees of anthropogenic load. It should be emphasized that the shellfish that live in polluted areas are likely to be in a state of oxidative stress, which is one of the causes the degradation of DNA.
Acknowledgements
The project was supported by the grant from the Russian Foundation for Basic Research no. 15-04-06526 А.
References
Cavas T., Konen S. In vivo genotoxicity testing of the amnesic shellfish poison (domoic acid) in piscine erythocytes using the micronucleus test and the comet assay // Aquat. toxicol. 2008. Vol. 90. P. 154–159.
Mitchelmore C.L., Birmelin C., Livingstone D.R., Chipman J.K. Detection of DNA strand breaks in isolated mussels (Mytilus edulis) digestive gland cells using the “Comet” assay // Ecotoxicology and Environmental Safety. 1998. Vol. 41. P. 51–58.
Keywords:
Corbicula japonica,
Mollusks,
oxidative DNA damage,
Gills,
Bioindicators
Conference:
ICAW 2015 - 11th International Comet Assay Workshop, Antwerpen, Belgium, 1 Sep - 4 Sep, 2015.
Presentation Type:
Poster Presentation
Topic:
Ecogenotoxicology
Citation:
Slobodskova
VV and
Chelomin
VP
(2015). DNA damage in gill cells of Corbicula japonica exposed to natural and anthropogenic stressors.
Front. Genet.
Conference Abstract:
ICAW 2015 - 11th International Comet Assay Workshop.
doi: 10.3389/conf.fgene.2015.01.00003
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Received:
02 Jun 2015;
Published Online:
23 Jun 2015.
*
Correspondence:
Mrs. Valentina V Slobodskova, POI FEB RAS, Vladivostok, 690041, Russia, slobodskovav@gmail.com