Comparative reproductive toxicity of 6:2 FTS and PFOS in female zebrafish (Danio rerio): From histopathology to molecular response.

6:2 Fluorotelomer sulfonate (6:2 FTS), a substitute for perfluorooctane sulfonate (PFOS), is frequently detected in aquatic environments, yet its reproductive toxicity remains incompletely understood. In this study, 480 adult female zebrafish were randomly assigned to five groups: DMSO control, PFOS (0.25 and 5 μg/L), and 6:2 FTS (0.25 and 5 μg/L), with 96 fish per group and three replicate tanks. After 42 days of exposure, PFAS bioaccumulation was assessed using three fish per group; separate sets of six fish per group were used for ovarian oxidative stress, HPG-axis-related hormones, HPG-axis-related gene expression, and reproductive performance; ovarian histopathology was examined using four fish per group; offspring development was assessed using 120 embryos per group; and integrated biomarker response version 2 (IBRv2) was applied. Data are presented as mean ± SEM, with significance set at p < 0.05. Compared with PFOS, 6:2 FTS showed lower bioaccumulation in the maternal liver, ovary, brain, and offspring, but still exhibited relative accumulation in ovarian tissue. Moreover, the ovarian oxidative stress induced by 6:2 FTS was relatively mild. Both compounds disrupted HPG-axis function, increasing gonadotropin releasing hormone (GnRH) and luteinizing hormone (LH) levels while decreasing follicle-stimulating hormone (FSH), testosterone (T), and estradiol (E2) levels. Consistently, gnrh-family genes and lhβ were upregulated, whereas fshβ and 17β-hsd were downregulated. These endocrine disturbances were associated with abnormal oocyte development, increased follicular atresia, reduced spawning output, and higher offspring malformation risk. IBRv2 analysis indicated that although 6:2 FTS caused lower overall toxicity than PFOS, it still impaired reproduction mainly through HPG-axis disruption. These findings demonstrate the reproductive risk of 6:2 FTS and support careful safety evaluation of PFAS alternatives.