Assessment of the toxic effects exerted by chromium in soils combining standardized OECD tests and cell-based approaches in earthworms (Eisenia fetida)

Abstract

Chromium (Cr) is widely used in a variety of industrial processes which lead to releases into the different environmental compartments. Inputs of Cr into terrestrial environments could modify soil community and affect soil health. The standardized earthworm Acute Toxicity Test (ATT) (OECD-207) with Eisenia fetida provides information about which concentrations of Cr pose effects on growth and mortality in earthworms. Complementarily, parameters measured in earthworms immune cells (coelomocytes), could be an accurate tool to assess the toxic effects exerted by Cr in these soil dwelling organisms. OECD soils (OECD-207, 1984) were contaminated with 10 - 150 mg Cr/kg (Cr was added as K2Cr2O7) and left stabilizing for 3 weeks. Healthy and clitellated E. fetida earthworms of similar size (400 ± 100 mg fresh weigh) were weight in tens and introduced in the test soils, maintained at 19 °C, in constant humidity and light conditions. For the ATT mortality and weight loss were measured (14 d) in order to obtain LC50 and EC50 values.. Cr was quantified in soils (EPA 3051A method), lixiviates (soluble fraction, DIN 38414-4) and earthworms (n=15) by AAS (SGIker General Service; UPV/EHU). Coelomocytes were extruded from exposed earthworms (3 pools of 5 individuals per treatment) and their number and cell viability were calculated. Cr concentrations in lixiviates were very low and suggest the formation of Cr-soil complexes while Cr was accumulated in earthworms showing an increasing gradient from 40 mg Cr/kg onwards. Mortality and weight loss followed a dose response fashion. Exposure to the highest doses (130 - 150 mg Cr/kg) produced 100 % mortality. The LC50 value was calculated as 97 mg Cr/kg, which corresponded to 20 mg Cr/kg soluble fraction. The EC50 was 115 mg Cr/kg and corresponded to 22 mg Cr/kg (soluble fraction). Regression analyses showed a significant reduction of cell number with exposure concentrations (R2 = 0.97, p<0.05). Coelomocytes decrease could be a first sign of more severe toxicological effects at higher levels of biological complexity. The NR uptake capacity of coelomocytes appeared to be affected at concentrations higher than 40 mg Cr/kg. Effects reported in the present work could be related to responses at lower levels of biological complexity such as oxidative stress, DNA damage and apoptosis. Acknowledgements - Funded by Basque Government (Consolidated Research Groups, IT810-13) and BEC.AR (Program of scholarships for Argentine professionals).