QUINOXYFEN 1995-2019 IS THE STORY’S END? EVALUATION OF ITS ADVERSE EFFECTS ON HEAD SIZE AND NERVOUS SYSTEM GENES INVOLVED IN SYNAPTIC MATURATION

Quinoxyfen (QXY) is a fungicide belonging to the quinoline family, introduced for the control of powdery mildew. In 2013 QXY was included in the list of priority hazard pollutants of the European Water Framework Directive due to its toxicity to aquatic organisms. The use of products formulated containing QXY was banned from all commercials starting from 27th June of 2019. Nevertheless, the QXY is an organic pollutant with potential persistence and bioaccumulation and its effects on neurological developmental are not still investigated. The aim of the work is to evaluate the toxicity of this compound using a zebrafish (Danio rerio) early- life stage focusing in particular on neurological defects. The study was conducted in 2 steps: in the first one, using the Fish Embryo Acute Toxicity test (FET), the toxicological endpoints as sublethal alterations and abnormal behaviors were investigated and in the second one two sublethal concentrations (0.4 and 0.8 mg/L) were selected to performed molecular investigations and Alcian blue staining. The cyp19a1b, shank3a, gad1b, neurexin1a genes, involved in the development of the nervous system and in the regulation of synaptic transmission, were evaluated. Furthermore, to investigate the craniofacial alterations, morphometric analyses were performed. The results of FET tests showed as the tested concentrations allowed to calculate the lethal concentration 10 (LC10) of 0.9 mg/L, while it was not possible to reach LC20 and LC50 values. Moreover, several sublethal alterations as yolk sac edema, pericardial edema, reduced blood circulation, blood stasis, reduced head size and altered mouth mor- phology were observed d head size and altered mouth morphology were observed. Additionally, at 72 hours post fertilization the larvae exhibited a particular phenotype characterized by abnormal mouth’s gape. Morphometric analysis revealed a decrease head length, an increase of Meckel and palatoquadrate cartilage angle and an increase of Meckel and Ceratohyals cartilage distance. These alterations can be linked to the brain malformations as reported by Raterman et al., 2020. This phenotype was supported by the changes in the expression of crucial genes influencing the normal development of the central nervous system and during the zebrafish embryonic phase, such as cyp 19a1b, shanka, gad1b, and neurexina. At both concentrations QXY led to a decrease in cyp19a1b gene expression suggesting an altered production of estradiol in the brain and a potential role as endocrine disruptor but an increase in gad1b, shanka and neurexina. These results could represent a starting point to study the potential role of QXY on neurodevelopment and possible link with birth defect induced by organism exposure by bioaccumulation effects.