Esterases as pesticide biomarkers in crayfish (Procambarus clarkii, Crustacea): tissue distribution, sensitivity to model compounds and recovery from inactivation.

The specific activities of acetyl- and butyrylcholinesterase and carboxylesterase were assayed in the digestive gland and in nervous and muscle tissues of the crayfish Procambarus clarkii. Since acetylcholinesterase prevails in nervous tissue and carboxylesterase in digestive gland, they are proposed as biomarkers. Muscle had negligible activities of all esterases, and all tissues had a low butyrylcholinesterase activity. Esterases were mostly cytosolic in digestive gland and muscle, but membrane-bound in nervous tissue; use of Triton X-100 is not recommended due to its widely diverging effects in esterase assays. Phenylmethylsulphonylfluoride inhibited acetyl- and butyrylcholinesterase in extracts from all tissues, and in digestive gland only carboxylesterase. In digestive gland, tetra[monoisopropyl]-pyrophosphorotetramide inhibited all esterases with different sensitivities, while in muscle and nervous tissue it only partially inhibited all esterases. Carbamates inhibited 100-fold more strongly than organophosphates acetyl- and butyrylcholinesterase activities. Carboxylesterase was inhibited by carbaryl and chlorpyrifos, but not by eserine and malathion. In vitro conditions to evaluate recovery from inactivation of esterases by model pesticides were established for acetylcholinesterase and carboxylesterase. The new reactivation protocol could be useful as a biomarker of pesticide exposure to differentiate between dilution-reversible inhibitions, indicating carbamate exposure, from dilution-irreversible effect, attributed to organophosphate exposure.

Doñana National Park survey using crayfish (Procambarus clarkii) as bioindicator: esterase inhibition and pollutant levels.

Utility of carboxylesterase and acetylcholinesterase inhibition as pesticide exposure biomarker was studied at Doñana National Park (SW Spain) in crayfish (Procambarus clarkii). Activities were measured in animals from reference sites or potentially exposed to pesticides, and their reactivation studied after dilution or 2-PAM treatment. Crayfish from affected sites had significantly less carboxylesterase and acetylcholinesterase activity than reference ones. No significant differences were found after dilution or 2-PAM treatment, showing that inhibition was irreversible. High pesticide levels were found in water and/or soil at rice growing sites, and lower levels at other affected places. High metal levels existed at rice growing sites and lower at other affected and at both reference sites. A combined effect on esterase inhibition of pesticides and metals is proposed. This field study suggest that the rice growing areas near Guadiamar stream are most polluted, followed by strawberry and citrics growing zones near Partido and Rocina streams. However, no correlation exist between the pesticide concentration at different sites and the extent of esterase inhibition, indicating that other factors could affect esterase response of animals from polluted sites.