DNA Damage Caused by Waterborne Metals and their Accumulation in Fish Body

Abstract

In the present project “DNA damage caused by waterborne metals and their accumulation in fish body” acute toxicity tests and chronic sub-lethal exposure effects of metals and their mixtures on DNA damage was investigated in the fingerlings (110 days old) of major carps viz. Catla catla, Cirrhina mrigala and Labeo rohita. Acute toxicity of cobalt (Co), chromium (Cr), lead (Pb), cobalt+chromium (Co+Cr), cobalt+lead (Co+Pb), chromium+lead (Cr+Pb) and cobalt+chromium+lead (Co+Cr+Pb) was determined in terms of 96-hr LC50 and lethal concentrations. Accumulation of metals due to exposure of individual metals and their mixtures was also studied during acute toxicity experiments. Fish were separately exposed to five sub-lethal concentrations viz. SLC-I (1/3rd of LC50), SLC-II (1/4th of LC50), SLC-III (1/5th of LC50), SLC-IV (1/6th of LC50) and SLC-V (1/7th of LC50) of their respective individual metals and metals mixtures 96-hr LC50 values for a period of 84 days to evaluate their sub-lethal effects on time- and concentration dependent accumulation and genotoxicity of various individual metals/metals mixtures to the fish. After the chronic sub-lethal exposure of individual metals and metals mixtures the selected organs of fish viz. liver, kidney, heart, gills, skin and muscles were isolated and examined for the determination accumulation patterns of respective exposure metals. Fish peripheral blood erythrocytes were also collected after 14, 28, 42, 56, 70 and 84 days of sub-lethal metals exposure and an extent of DNA damage was measured by using Comet assay in terms of following parameters viz. cumulative tail lengths (µm), damaged nuclei (%) and genetic damage index values. The tolerance of three selected fish species, in terms of both LC50 and lethal concentrations (mgL-1), against metals and their mixtures varied significantly. Among the three fish species, Cirrhina mrigala exhibited least sensitivity against all the exposed metals and their mixtures than that of Labeo rohita and Catla catla. The overall toxicity pattern of metals/metals mixtures followed the order: Co+Cr+Pb > Co+Pb > Cr+Pb > Pb > Co+Cr > Cr > Co, in terms of 96-hr LC50, while for lethal concentrations this order was: Co+Cr+Pb > Cr+Pb > Co+Pb > Pb > Co+Cr > Cr > Co. The exposure of metals/metals mixtures at acute toxicity concentrations caused significant accumulation in the organs of the selected fish species. Cirrhina mrigala exhibited significantly (p<0.05) higher tendency to concentrate all the metals in its organ while amassing of metals was minimum in Catla catla. In general, metallic ion distribution in the fish tissues predicted its tolerance against specific metal and patterns of its accumulation in different tissues. Overall abilities of metals to concentrate in the fish followed the order: Co > Cr > Pb > Co+Pb > Co+Cr > Cr+Pb > Co+Cr+Pb. The amassing of metals in the fish was in the following order: kidney > liver > gills > heart > skin > muscle. Fish species also exhibited variable differences in their tendencies to amass metals due to 84 days exposure of selected concentrations (SLC-I, SLC-II, SLC-III, SLC-IV and SLC-V). The amassing of metals in the fish body increased concomitantly with the exposure duration of various sub-lethal concentrations of individual metals/metals mixtures. Among all the treatments, significantly higher accumulation of metallic ions in the fish body was observed due to exposure of Co+Cr+Pb mixture while the control fish had significantly least metals in their bodies. The overall accumulation pattern of individual metals/metals mixtures in the fish was: Co+Cr+Pb > Co+Cr > Co+Pb > Cr+Pb > Co > Cr > Pb > control. Among the selected fish species, Cirrhina mrigala exhibited higher (significant at p<0.05) ability to concentrate metallic ions than that of Labeo rohita and Catla catla. The accumulations pattern of metals in the selected organs of fish was: liver > kidney > gills > heart > skin > muscle. It seems that interactions among various metals, in a mixture form, were related to their competitive uptake from the exposed media to cause significantly variable accumulation and ultimately resulted into additive, synergistic or antagonistic effects in the fish body as observed during this investigation. Significantly variable DNA damage to the fish erythrocytes, measured in terms of cumulative tail lengths (CTL), damaged nuclei (%) and genetic damage index (GDI) values was observed in all the selected fish species The exposure of metals/metals mixtures caused significantly (p<0.05) variable damage to the fish erythrocyte nuclei in all the selected fish species. Among the fishs, Labeo rohita showed significantly higher tail lengths of comets in their peripheral erythrocytes, followed by that of Catla catla and Cirrhina mrigala. Labeo rohita also exhibited higher percentages of damaged nuclei while it was minimum for Cirrhina mrigala. Regarding overall sensitivities of three fish species towards genetic damage indices due to exposure of metals/metals mixtures, both Labeo rohita and Catla calta appeared significantly more sensitive than that of Cirrhina mrigala (1.34±0.23). Significantly variable cumulative tail lengths, damaged nuclei (%) and genetic damage index values were observed among various treatments (metals and metals mixtures). Comparison of treatment means revealed that Co+Cr+Pb toxicity induced significantly maximum damage to the fish erythrocyte nuclei while the same was significantly minimum induced by the exposure of Co to the all three fish species. Significant dependence of DNA damage was observed upon exposure of various concentrations of Co, Cr, Pb, Co+Cr, Co+Pb, Cr+Pb and Co+Cr+Pb). The cumulative tail length (µm), damaged nuclei (%), and genetic damage index values showed concomitant increase with the exposure duration of metallic ions. The cumulative tail lengths, damaged nuclei (%), GDI values and were observed maximum after 56 days exposures while the same were least during the first 14 days of exposure of metals/metals mixture. The duration based decrease in DNA damage after 56 days of exposure of metals/metals mixtures may be due to repairing of damaged DNA. Therefore, it is concluded to quantify the DNA damage in the erythrocytes of fish exposed to metals, comet assay has successfully been applied and can also be used as a sensitive biomarker for the determination of genotoxic effects of various toxicants in the aquatic organisms.