Developmental toxicity of the fungicide Boscalid and primary metabolite M510F01 in zebrafish embryos
Succinate Dehydrogenase Inhibitors (SDHIs) have been used for decades, with new formulations cropping up as fungi evolve. Boscalid was introduced to United States agriculture in 2003 and was determined to be effective for fungal management and safe for aquatic discharge and human consumption. Recent studies have found Boscalid's primary metabolite, M510F01, on crops and in irrigation runoff as well. However, limited information is available about the aquatic and developmental toxicity of M510F01. The goal of our study was to compare and contrast the developmental toxicity of Boscalid with its metabolite, M510F01. Zebrafish (AB, wild-type strain) embryos were exposed to Boscalid or M510F01 from 3-96 hours post fertilization (hpf) at concentrations of 0.03, 0.3, or 3 µg/L. Embryos were examined for aberrant morphologies at 96 hpf using microscopy. Boscalid exposures at concentrations of 3 µg/L increased incidence of blue sac disease by 72%, craniofacial malformation by 97%, pericardial edema by 69%, delayed swim bladder inflation by 35%, and yolk edema by 83% (p<0.001 for all morphologies). 3 µg/L M510F01 increased the incidence of blue sac disease by 25% and yolk edema by 17% (p=0.005). Additionally, Boscalid exposures at 3 µg/L indicated a significant effect on fish length at 96 hpf (p<0.05).To assess whether Boscalid and M510F01 impact Cytochrome P450 1a (Cyp1a) activity, embryos were exposed to Boscalid or M510F01 (0.03 µg/L) from 96-100 hpf concurrently to 7-ethoxyresorufin (EROD)—a fluorometric indicator of Cyp1a activity. There is no significant change in acute EROD activity for Boscalid or M510F01; however, chronic exposure of Boscalid and M510F01 at 0.03 µg/L indicated a significant decrease in Cyp1a activity in the gut and M510F01 additionally reflected a significant decrease in Cyp1a activity in the liver at 0.03 µg/L. RNA sequencing and pathway analysis were performed to assess the mechanisms of xenobiotic response to Boscalid and M510F01 exposures (0.03 µg/L). Boscalid significantly upregulated pathways involved in ATP production including oxidative phosphorylation, glycolysis, and TCA cycle. Overall, this research suggests that M510F01 exhibits reduced aquatic and developmental toxicity when compared to Boscalid. Overall design: Zebrafish embryos were exposed to Boscalid or M510F01 from 3-96 hpf at concentrations of 0.03, 0.3, or 3 µg/L, with exposures renewed daily. Embryos were monitored for survival and hatching, and imaged to assess structural anomalies and malformations. RNA was isolated and sequenced at 96 hpf.