National Institute for Agro-Environmental Sciences, Tsukuba, Ibaraki, Japan.
Ecotoxicol Environ Saf. 2013 Aug;94:37-44. doi: 10.1016/j.ecoenv.2013.04.020. Epub 2013 May 21.
Although riverine periphytic algae attached to riverbed gravel are dominant species in flowing rivers, there is limited toxicity data on them because of the difficulty in cell culture and assays. Moreover, it is well known that sensitivity to pesticides differ markedly among species, and therefore the toxicity data for multiple species need to be efficiently obtained. In this study, we investigated the use of fluorometric microplate toxicity assay for testing periphytic algal species. We selected five candidate test algal species Desmodesmus subspicatus, Achnanthidium minutissimum, Navicula pelliculosa, Nitzschia palea, and Pseudanabaena galeata. The selected species are dominant in the river, include a wide range of taxon, and represent actual species composition. Other additional species were also used to compare the sensitivity and suitability of the microplate assay. A 96-well microplate was used as a test chamber and algal growth was measured by in-vivo fluorescence. Assay conditions using microplate and fluorometric measurement were established, and sensitivities of 3,5-dichlorophenol as a reference substance were assayed. The 50 percent effect concentrations (EC50s) obtained by fluorometric microplate assay and those obtained by conventional Erlenmeyer flask assay conducted in this study were consistent. Moreover, the EC50 values of 3,5-dichlorophenol were within the reported confidence intervals in literature. These results supported the validity of our microplate assay. Species sensitivity distribution (SSD) analysis was conducted using the EC50s of five species. The SSD was found to be similar to the SSD obtained using additional tested species, suggesting that SSD using the five species largely represents algal sensitivity. Our results provide a useful and efficient method for high-tier probabilistic ecological risk assessment of pesticides.
尽管附着在河床砾石上的河流周生藻类是流动河流中的优势物种，但由于细胞培养和测定的困难，对它们的毒性数据有限。此外，众所周知，不同物种对农药的敏感性差异很大，因此需要有效地获得多种物种的毒性数据。在这项研究中，我们调查了使用荧光微板毒性测定法测试周生藻类物种的情况。我们选择了五个候选测试藻类物种：束丝藻、脆杆藻、舟形藻、脆杆藻和伪鱼腥藻。所选物种在河流中占优势，包括广泛的分类群，并代表实际的物种组成。还使用了其他附加物种来比较微板测定法的敏感性和适用性。96 孔微孔板用作测试室，通过体内荧光测量藻类生长。建立了使用微孔板和荧光测量的测定条件，并测定了 3,5-二氯苯酚作为参考物质的敏感性。通过荧光微板测定法获得的 50%效应浓度（EC50）和本研究中通过常规摇瓶测定法获得的 EC50 一致。此外，3,5-二氯苯酚的 EC50 值在文献中报道的置信区间内。这些结果支持了我们的微板测定法的有效性。使用五个物种的 EC50 进行了物种敏感性分布（SSD）分析。发现 SSD 与使用其他测试物种获得的 SSD 相似，表明使用五个物种的 SSD 在很大程度上代表了藻类的敏感性。我们的结果为农药的高等级概率生态风险评估提供了一种有用且高效的方法。
