Evaluation of parental and transgenerational effects of clotrimazole in Daphnia magna - A multi-parametric approach

Abstract

Azole antifungals inhibit the cytochrome P450 complex, decreasing the production of ergosterol in fungi, and compromising the biosynthesis of ecdysteroids in crustaceans, which are hormones regulating reproduction and ecdysis. The azole antifungal clotrimazole (CLO) raises environmental concerns due its toxicity. This work evaluated the effects on the number of moults, feeding rate, growth, reproduction, transgenerational reproductive effects on two different generations (F0, parental generation; and F1, organisms born from F0), and energetic balance in Daphnia magna. Neonates (<24 h) were exposed to sublethal concentrations (0, 2.7, and 3.4 mg/L) of CLO, to assess its effects on the moulting process. Neonates were also exposed to environmentally realistic concentrations of CLO (0, 30, 150, 750, and 3750 ng/L) for 24 and 96 h, to assess adverse effects on their feeding behaviour. Effects on energy reserves (fatty acids, glycogen, and protein levels) were also measured in animals exposed to CLO. A reproduction test was carried out to evaluate the amount and size of neonates from F0 and F1 generations. CLO exposure decreased the number of moults, and the size of organisms, but did not alter the feeding pattern of 5 days old individuals. However, neonates (<24 h) exposed to CLO had a significant decrease in their feeding pattern. CLO decreased the fatty acids content in exposed animals, but did not change glycogen and protein. CLO also decreased the size of adult daphnids from the third brood, born from animals exposed in F0; in F1 animals, the size of neonates from the third brood was decreased. This study evidenced the toxic effects caused by CLO on growth, feeding and reproduction of D. magna. Nevertheless, it is not possible to conclude whether the effects are due to the inhibition of cytochrome P450 enzymes, or to unspecific effects caused by general toxic stress and decreased nutrition.

Keywords: Azole fungicides; Clotrimazole; Daphnia magna; Fatty acids; Feeding; Reproduction.