Impacts of oil exposure during early life developmental stages in Sheepshead minnows (Cyprinodon variegatus) under different environmental factors

The purpose of this research was to determine windows of increased sensitivity to combined oil and hypoxic exposure under varying salinity regimes during early life stage development of sheepshead minnows. The early life development was divided into three stages: embryonic (< 15 hpf), post-hatch (<10 hph), and post larval (96 hph). Immediately after each transition period sheepshead minnow larvae were exposed to four concentrations of HEWAF (6.25%, 12.5%, 50% and 100% of 1g/L stock solution) under different oxic (2.5 mg/L and >5.0 mg/L) and salinity (10 and 30 ppt) regimes, while temperature was maintained at 30°C. After the 48 h exposure period, larvae were transferred to normoxic clean water to monitor growth and mortality for up to 10 days. The experiments were static experiments with water renewals every 48 hours. Water quality and organism performance were monitored and recorded daily. A sample of 3.5 mL of exposure water was collected from each test chamber during water quality monitoring at 0 h, 24 h, 48 h, and 72 h for fluorescence analysis to measure PAH concentrations. Whole body tissue samples were collected directly after the 48 h exposure period measure relative gene expression and investigate pathway inhibition between the aryl hydrocarbon and hypoxia inducible factor 1-α signaling pathways. Target genes included Cytochrome P450 1A1 (CYP1A1), Erythropoietin (EPO), and Aryl hydrocarbon receptor nuclear translocator (ARNT).

本研究旨在确定羊头鲦鱼早期发育阶段在不同盐度条件下，对石油与低氧联合暴露的敏感窗口期。其早期发育分为三个阶段：胚胎期（<15小时受精后，hpf）、孵化后期（<10小时孵化后，hph）及幼后期（96小时孵化后，hph）。每个过渡期结束后，立即将羊头鲦鱼幼虫暴露于四种HEWAF浓度（1g/L储备液的6.25%、12.5%、50%及100%），同时设置不同氧浓度（2.5 mg/L和>5.0 mg/L）与盐度（10和30千分比，ppt）条件，温度维持在30℃。暴露48小时后，幼虫被转移至常氧清水环境，监测其生长与死亡率长达10天。实验采用静态模式，每48小时更换一次试验用水。每日监测并记录水质参数及生物表现。在暴露0、24、48及72小时时，从各试验舱采集3.5 mL暴露水样，通过荧光分析测定多环芳烃（Polycyclic Aromatic Hydrocarbons，PAH）浓度。暴露48小时后，直接收集幼虫全组织样本，用于检测相对基因表达水平，并探究芳烃受体与低氧诱导因子1-α信号通路之间的抑制作用。目标基因包括细胞色素P450 1A1（Cytochrome P450 1A1，CYP1A1）、促红细胞生成素（Erythropoietin，EPO）及芳烃受体核转位蛋白（Aryl Hydrocarbon Receptor Nuclear Translocator，ARNT）。