Projects presented in this paper were funded by Severn Trent Water and Brunel University London. The authors would like to thank Alan Henshaw and John Churchley for providing field and laboratory assistance. T.J.C. thanks the Heinz Endowments for support. M.R.M. thanks the Steinbrenner Institute for a Steinbrenner Doctoral Fellowship and Carnegie Mellon University for a Presidential Fellowship.

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The authors have nothing to disclose.

废水处理厂是用EDC地表水污染的主要途径。去除常规的，先进的或新出现的治疗过程的内分泌活性的效力的评估需要使用的各种化学和生物测定的。用非靶向和靶向分析化学分析提供了关于除去各组分的功效的定性或定量的数据，因此，允许对环境质量标准生产或预测分析的化合物或化合物的混合物无影响浓度的评估。 从下面的治疗和未知的生物活性成分的废水中存在的物质的不完全矿化所得转化产物的产生限制单独化学测试是有用的。 A 在体内和体外生物测定结合分析化学家组合RY筛选提供了一个有用的工具箱新兴废水处理过程，以确定的EDC去除的效力。这些测试时，除了传统的水质参数和其他毒理学和微生物学终点进行允许现有和新兴的废水处理技术的一个重要的评价。 要注意的是酵母基于雌激素的屏幕（ 例如 ，是）是很重要的并不是唯一的体外测定，以确定化学品和废水的雌激素效能。一些稳定转染基于哺乳动物细胞的测定法，已经开发过，对ER-CALUX 27和与人乳腺癌细胞或子宫颈肿瘤细胞hERα-的HeLa-9903 28分别为例子。的是已经相比类似哺乳动物基于细胞的测定法，并已发现具有可重复性的一个可比较的水平高，真阳性和真阴性雌激素的识别率29本书虽然啊它有时被认为是稍微较不敏感的27。基于酵母报告分析的一个优点是，在不与哺乳动物细胞培养显著经验的实验室的是，可以更容易地通过，因为它需要较少的严格的生物控制措施和无菌技术（可置于工作台顶部，如果必要，可以进行是） 。人细胞为基础的分析也需要的 CO 2孵化器和光度计相比，在是使用的标准培养箱和酶标仪。两个基于酵母的雌激素记者分析（YES， 酿酒酵母和A-YES，Arxula adeninivorans）的ISO 19040的认证目前正在实验室间的小径"水质-水和废水的雌激素潜在的测定"突出产业在这些技术的兴趣。 有一些所描述的方法的局限性，其中包括的潜在的污染的采样，样品存储和分析来自现场或实验室环境或人类污染（ 例如，增塑剂，表面活性剂，个人护理产品）始发雌激素作用物质中的样品。这种类型中的YES测定（或其他基于细胞的报告分析）污染将提升的背景和影响使用该测定的。存储在塑料瓶的水样或溶剂极易造成假阳性。假阴性也备受关注既是LCMS / MS和YES检测要求SPE到雌激素到集中检测水平。基质中，SPE吸附剂和洗脱溶剂的选择可影响萃取效率和类型洗脱化合物。使用C18 SPE柱，使用在此协议中描述的条件可以产生负偏压的提取，作为高极性和碱性化合物会通过吸附剂进行很差保留。此外，该协议要求从甲烷洗脱YES洗脱液重建醇通过蒸发乙醇至干产生挥发性化合物的损失出资者氮。其结果是该协议可以提供低估测试的样品的雌激素活性。考虑到是检测未知的或意外的化合物可能会被错过，因为他们没有被提取或他们因蒸发而丢失时，这些限制是特别重要的。此外，LCMS / MS技术利用了内标来校正恢复;这种方法不能与YES测定中使用。 体内流出物的测试显著局限性包括相比体外方法进行评估所需的高成本和时间。目前使用的鱼胚胎试验检测雌激素活性是有限的。然而，已经有了一些成功的生产雌激素反应转基因发光斑马鱼胚胎30，这可能会对未来的应用。黑头呆鱼（在此protoc使用醇）是一种常见的实验室物种和VTG诱导雄性鱼是雌激素暴露的证据充分的生物标志物和废水流出物雌激素22或其它的雌激素化合物或其混合物31的量化度量。对内分泌干扰物质的OECD测试规定已使用成人黑头呆鱼，青鳉日本和斑马鱼32,33验证，与VTG是在所有三种雌激素暴露的敏感生物标志物。然而，VTG感应不直接相关的生殖障碍，因此废水曝光的生态后果，如严重雌雄同体蟑螂3所示。在另一方面，蟑螂是不是生态毒理学研究的一个经典的"实验室物种，由于其大尺寸，长的生成时间（2 - 3年才能达到性成熟），生殖风格;组产卵（繁殖）发生一年一次，难识别从女性的男性（比在其他产卵季节）。然而，这通常雌雄异体物种已被很好的研究在英国，这是因为雌激素废水排放的下游，雄鱼展出扰动他们的内分泌（ 例如 ，在他们的血液女性特有的卵黄蛋白原的存在）和组织病理学（ovotestes发现-开发鸡蛋睾丸和/或女性生殖管道）5,6。因此，这些协议的应用前景，罗奇（或类似物质）可能是一个有益的野生物种定点显示，如果废水质量（并降低雌激素）真正的改善河流接受先进的污水处理被看见。它们也可以在配管系统的端部来监视从试验规模的工厂7技术上改进的废水。当考虑在体内废水评估要使用的物种有相对快速和受控的测试用实验室物种之间的折衷相比长场为主，但更多的环境有关​​，使用本地物种进行测试。然而，这种在体内的评价是成本高，只应视为最后一组使用以下化学分析评估，试验和体外试验。 所述的协议中的关键步骤包括制备和样品和玻璃器皿的处理（ 即，瓶和取样设备应用适当的表面活性清洁剂预先处理），以避免从环境污染物包括限制样品接触与塑料和样品的污染能产生误报的其他材料。设计和构建水族馆鱼曝光系统时，这是非常重要的。理想的情况是水族馆（住宅存量和期间暴露）应从材料建成具有低吸附32以最小的污染风险。不锈钢可以用于流出物或水储水槽。而一个玻璃结构的坦克是优选的鱼缸（因为这也提供了鱼便于观察）。使用低等级塑料管或管道的，应避免32，PVC 34和ABS可以当&#39;适当调味&#39;被使用， 即，留在运行稀释水中至少12小时，使用前以浸出任何污染物。医用级硅管已经在我们的蠕动泵输送化学品和废水/稀释水的坦克工厂成功应用。除了考虑建筑雌激素污染和养殖系统的运行，它考虑鱼的饮食也很重要;许多鱼类礼仪食品已被认为是雌激素对鱼类。因此它来测试任何食物为活动（ 例如 ，在酵母雌激素屏幕，查看贝雷斯福德等人 14）在这些类型的研究使用它们之前是重要的。 故障排除的描述，如果质量保证样本，包括多个行程，实验室和溶剂空白分析一起阳性对照和实际样品，以消除假阳性和假阴性结果是简化了化学分析或YES测定协议。正（ 例如 ，EE2）和负极（稀释仅水）控制应也总是在体内测定可用于证实预期生物标志物或端点（ 即 ，VTG或组织病理学）的灵敏度，并且允许任何意外污染待检测（ 例如，从实验装置，饮食，或稀释水域）。在协议的任何修改之前应进行任何研究来验证。 与进入通过污水处理厂的污水对环境的雌激素化合物的更严格的法规它是设想更有效的污水处理技术将需要开发。在这个手稿中描述的测试电池恭维生态毒理学和化​​学评估测试通常用于污水处理厂的污水排放。因此，这种类型的测试应使废水技术开发商与设备操作员，整体的电池的应用前景实行最生态安全设计考虑的最佳方法同时删除特定的调节雌激素类化学和整体的生物活性。

关于内分泌干扰物对野生动物的生殖健康产生不利影响的担忧已导致欧盟在水框架指令（WFD）"观察名单"上放置两个雌激素物质（雌二醇和炔雌醇）。 EDC包括各种化学类，包括天然和合成的类固醇的雌激素，药物，杀虫剂和工业化学品和消费产品上的野生动物已知的不利影响的成分的。这些化合物的一些可能潜在地影响人类健康1。 有研究表明，污水处理厂的污水是雌激素对鱼类2并因此很多受纳水体也雌激素对鱼类3。这是首先通过在英国的全国调查表明，野生雄鱼的血液和前期高位表明卵黄蛋白原增加浓度（女卵黄蛋白前体4）阴阳的价（发展雄鱼睾丸鸡蛋和/或女性生殖管道）的正常雌雄异体鱼种5,6。 传统的污水处理通常是由经过初步筛选后一级和二级处理从而消除溶解和悬浮有机物的一个三阶段的过程。除去个别的EDC的功效依赖于物质的物理化学性质和所采用的治疗方法的有效性。对于通过吸附和生物降解许多EDC去除可以显著，但不完整的。三级处理，如砂滤，可在采用先进的氧化（例如臭氧）增加，而先进的治疗EDC拆除7有效或活性炭能有效地接近完全去除7实现。 任何新技术的污水处理NEE评估DS确定EDC除去所提出的方法的功效。测试，包括有针对性的化学分析毒理学一起测试， 使用体内和体外生物测定的电池，为此提供了全面的数据。虽然监管是非常有用的，有针对性的化学分析只能提供有关监测化合物（和具体的代谢物）数据。生物测定另外允许的副产品代谢物和治疗产生的废水变换的不利影响，否则将被未被发现-8,9-"检测"。本文介绍了使用化学和生态毒性实验室检测的电池的评估除去原油的雌激素效力和处理后的污水和受纳水体的一些先进和新兴的污水处理工艺的功效。

use of a battery of chemical and ecotoxicological methods for the assessment of the efficacy of wastewater treatment processes to remove estrogenic potency

Endocrine Disrupting Compounds pose a substantial risk to the aquatic environment. Ethinylestradiol (EE2) and estrone (E1) have recently been included in a watch list of environmental pollutants under the European Water Framework Directive. Municipal wastewater treatment plants are major contributors to the estrogenic potency of surface waters. Much of the estrogenic potency of wastewater treatment plant (WWTP) effluents can be attributed to the discharge of steroid estrogens including estradiol (E2), EE2 and E1 due to incomplete removal of these substances at the treatment plant. An evaluation of the efficacy of wastewater treatment processes requires the quantitative determination of individual substances most often undertaken using chemical analysis methods. Most frequently used methods include Gas Chromatography-Mass Spectrometry (GCMS/MS) or Liquid Chromatography-Mass Spectrometry (LCMS/MS) using multiple reaction monitoring (MRM). Although very useful for regulatory purposes, targeted chemical analysis can only provide data on the compounds (and specific metabolites) monitored. Ecotoxicology methods additionally ensure that any by-products produced or unknown estrogenic compounds present are also assessed via measurement of their biological activity. A number of in vitro bioassays including the Yeast Estrogen Screen (YES) are available to measure the estrogenic activity of wastewater samples. Chemical analysis in conjunction with in vivo and in vitro bioassays provides a useful toolbox for assessment of the efficacy and suitability of wastewater treatment processes with respect to estrogenic endocrine disrupting compounds. This paper utilizes a battery of chemical and ecotoxicology tests to assess conventional, advanced and emerging wastewater treatment processes in laboratory and field studies.

试图了解的改进废水处理过程的影响或确定最合适的技术在现有污水处理厂相对于除去排出废水的内分泌干扰活性的功效加装设备作为三级处理，不仅需要键化学的测定即进入工作，但需要分解产物也可以具有内分泌干扰活性的分析组件。在生活污水流出物，目前最雌激素物质是类固醇激素，雌酮（E1），17β雌二醇（E2）和17α炔雌醇（EE2）5,8。类固醇的雌激素主要从体内排出作为非活性结合物16,17的混合物。这些结合雌激素基本上在污水处理系统由细菌活动deconjugated和污水处理厂发生进一步退化。该deconjugated AR类固醇e。通过吸附从废水流中去除污泥或导致形成二次处理过程中降解，首先改造的副产品，并最终完全矿化可能发生的初始​​活性组分。的流出流中的所有单个化合物的化学分析将是困难的，耗时的和昂贵的，并且不会涵盖存在于样品中未知的活性成分。此外，各成分的雌激素贡献的总和将只提供所分析的化合物的样品的累积雌激素效力的指示。这是转化过程产生未知的雌激素物质或者其中流入是工业来源的风险。 在体内 和体外生态毒理学生物测定组合化学分析提供了在混合物如处理过的污水中的溶液到未知的雌激素组分的存在， 在体外测定，如酵母雌血肌酐EEN（YES），已被广泛用于确定污水流出物的雌激素活性，并帮助识别处理的样品8,18,19的活性成分。然而， 在体内的体外试验比较可以显著11，新工艺相对于治疗内分泌干扰效能的综合评估，需要化学和生态毒理学测试电池。 可使用随后的样品的提取，浓缩和分析前提取物的清理化学分析来实现的个别处理厂或过程是否从废水流中除去活性化合物A的确定，使用LCMS（/ MS）或GCMS最多进行（/ MS）的方法。从化学分析获得的数据可以被用来确定是否符合个别预测无影响浓度（PNEC）20或环境质量标准S（EQS）特定个人化合物21，因此这种方法对于法规遵从数据至关重要。此外，定位或非靶向化学分析方法允许用生物学方法，它提供了一个总的响应相比单个化合物或异构体的鉴定和定量。因此化学分析方法允许进行离散化合物的评估，以满足和解决个别处理厂的基础上，这些废水处理的挑战。有研究表明，虽然除去合成激素的EE2趋于较少有效的常规废水处理（ 例如，活性污泥处理厂）可能是在去除天然类固醇激素的高度有效的。使用利用的技术，如臭氧先进的治疗领域的研究中，颗粒活性炭（GAC）和膜已经证实，尽管成本高，它们可以作为尾管的溶液以除去EE2至低于预计值icted效果水平和低于检测限。 图3显示了EE2的使用GAC在中试规模的城市污水处理厂的拆除。在中试规模在用管GAC治疗结束市政污水处理厂进行的研究还显示以下的GAC在雌激素效力减少使用如图4中可见的酵母雌激素屏幕（是）进行测定。 <img alt="图3" src="/files/ftp_upload/54243/54243fig3.jpg" /> 图3.实例字段数据显示以下高级三级处理去除炔雌醇（A）样品是从污水处理厂以下为以下保存样品描述的步骤常规（活性污泥）处理收集。 （B）样品固相萃取，净化后用正相SPE去除干扰物质提取，凝胶渗透色谱法。 （C）的清洁浓缩提取物浓缩至小体积并在MRM模式使用负离子电LCMS / MS进行分析。结果是使用采用同位素标记的内标内部标准化计算。在所示的例子中，EE2是在0.1纳克/升的预测没有影响水平（PNEC）以上的浓度存在于最终的ASP流出物并用活性炭和臭氧去除（O 3），以对环境安全的浓度。 <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/54243/54243fig3large.jpg" target="_blank">请按此查看该图的放大版本。</a> <img alt="图4" src="/files/ftp_upload/54243/54243fig4.jpg" /> 图4.照片一个酵母雌激素屏幕（是）试验板（A）中示出从黄色到红色的颜色变化，与该样品的雌激素活性。从YES分析板显示创建的图形校正吸光度（540雌二醇标准（B）的纳米），活性污泥处理的流出物（ASP）和粒状活性炭（GAC）处理过的废水流出物的样品（C）将每个样品一式两份进行测试。 ASP和GAC废水提取和使用第1节中列出的SPE方法集中<a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/54243/54243fig4large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> 臭氧也是从常规治疗的污水处理厂去除类固醇雌激素和雌激素活性高效。臭氧是能够氧化多种有机污染物和溶解的有机物质的废水样品中，并提供消毒性能。臭氧化的有效性取决于水的特性如pH，有机物量和臭氧的施用剂量。由常规处理除去不良的雌激素可以是从废水0.8和2毫克O 3 /毫克之间DOC去除剂量。臭氧是一种选择性的氧化剂，其与富电子网站（不饱和碳 - 碳键，芳族化合物，包括芳族醇），这使得适用于一个号码的EDC的击穿臭氧反应。然而，单独的化合物的消除并不必然导致对原始化合物的完全矿化。有机物质以下臭氧可能的副产物，其中包括一些低分子量的转化产生的中间体或转化氧化，极性类型的化合物如醛，酮，羧酸，酮基羧酸，和溴化的化合物。实例包括溴酸盐，甲醛，乙醛和羧酸。 采用体内 和体外生物测定已经表明，尽管臭氧仅部分氧化一些化学物质，将得到的主要转变产物具有较低estrogENIC效力并因此臭氧在一个较高的去除雌激素活性的适当剂量的结果的应用程序。 一个废水处理额外的主要优点是在受纳水雄鱼女性化的减少;产生不利影响，可导致减少的生育3。使用鱼体内研究（ 例如 ，蟑螂或黑头呆鱼）暴露于废水显示雌性生殖细胞或卵母细胞的雄性鱼的精巢（ 例如，如在图5中看到）。阴阳或男性VTG缺席或鱼类以下先进的治疗，如海关总署7或臭氧22显著减少。这些研究表明，臭氧化过程中产生的变换的产品没有雌激素，然而，这并没有解决所产生的流出物的毒性。此问题已在其他研究中得到解决，例如通过马格德堡等的研究。 <s达&gt; 23这表明副产品臭氧氧化是有毒的虹鳟鱼，但这种毒性可以通过下游砂滤以下臭氧被删除。 <img alt="图5" src="/files/ftp_upload/54243/54243fig5.jpg" /> 图5.正常男性（A）和阴阳的显微照片（B，C）从成年罗奇（ 拟鲤 ）性腺在基于实地评估。显微照片-A 暴露于废水 ，描绘了正常男性睾丸的组织切片。显微照片-B和-C，描绘了阴阳公鱼的组织切片，被暴露在活性污泥法处理废水排放六个月。箭头指示存在于睾丸组织卵母细胞。比例尺为100μm在每个显微照片。 <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/54243/54243fig5large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> 12,24在有机微双氧水氧化- 26。 TAML与H 2 O 2活化剂，有效地降解EE2和其他类固醇雌激素在纯实验室用水，以及在城市污水处理厂的污水和尖刺尿样12英寸实验室的研究，示出TAML / H 2 O 2的处理提供了高类固醇的雌激素去除包括EE2去除和实质上降低使用是生物测定和质体外测定雌激素活性LLY减少鱼女性化在使用VTG生物测定（ 图1和图6）测量体内 。 <img alt="图6" src="/files/ftp_upload/54243/54243fig6.jpg" /> 图6.平均EE2浓度和雌激素活性处理和未处理的罐水域（A）和基线血浆卵黄蛋白原并暴露雄鱼（B）的A）EE2浓度（ng / L，深蓝色条）通过LCMS / MS测量，雌激素活性（EE2当量纳克/升，深绿色条）通过体外酵母雌激素屏幕测量（YES）。B）血浆卵黄蛋白原（毫微克/毫升，淡蓝色的条）浓度男性黑头呆鱼通过定量酶测定 - 连接的免疫吸附测定（ELISA）。报告为EE2化学分析结果&lt;0.03纳克/升EE2（ 即，高于检测限（LOD）低级）被视为具有一半的LOD（即 </青霉&gt; 0.015纳克/升EE2），用于在平均值，标准误差和统计分析的计算中使用。 EE2和雌激素活性是测得的平均浓度在取样21天的暴露。等离子VTG是之前曝光（基线），并在21天后曝光测量。治疗制度包括的;阴性对照组（仅稀释水），EE2 + H 2 O 2 + TAML，EE2 + H 2 O 2和EE2只。在图-A误差棒代表平均值的标准误差，在图-B误差棒代表标准偏差。以上图-B条字母代表统计相似。这个数字已经从米尔斯等人修改。12 <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/54243/54243fig6large.jpg" target="_blank">，请点击这里查看本图的放大版本。</a>

Watch this Scientific Journal Video about Use of a Battery of Chemical and Ecotoxicological Methods for the Assessment of the Efficacy of Wastewater Treatment Processes to Remove Estrogenic Potency at

Use of a Battery of Chemical and Ecotoxicological Methods for the Assessment of the Efficacy of Wastewater Treatment Processes to Remove Estrogenic Potency

内分泌干​​扰物（EDC）构成重大风险到水生环境。城市污水处理厂的主要贡献者，以地表水的雌激素效力。本文提供的方法允许疗效和污水处理工艺的适用性相对于EDC去除的评估。

对污水处理工艺的疗效评估利用化学和生态毒理学方法的电池，以消除雌激素潜能

Endocrine Disrupting Compounds pose a substantial risk to the aquatic environment. Ethinylestradiol (EE2) and estrone (E1) have recently been included in ...

use-battery-chemical-ecotoxicological-methods-for-assessment-efficacy

Nanyang Technological University

Research

JoVE Journal

Environment

14.0K 次观看.  Brunel University London. 内分泌干​​扰物（EDC）构成重大风险到水生环境。城市污水处理厂的主要贡献者，以地表水的雌激素效力。本文提供的方法允许疗效和污水处理工艺的适用性相对于EDC去除的评估。 

视频: Use of a Battery of Chemical and Ecotoxicological Methods for the Assessment of the Efficacy of Wastewater Treatment Processes to Remove Estrogenic Potency

Physical, Chemical and Biological Characterization of Six Biochars Produced for the Remediation of Contaminated Sites

The physical and chemical properties of biochar vary based on feedstock sources and production conditions, making it possible to engineer biochars with specific functions (e.g. carbon sequestration, soil quality improvements, or contaminant sorption). In 2013, the International Biochar Initiative (IBI) made publically available their Standardized Product Definition and Product Testing Guidelines (Version 1.1) which set standards for physical and chemical characteristics for biochar. Six biochars made from three different feedstocks and at two temperatures were analyzed for characteristics related to their use as a soil amendment. The protocol describes analyses of the feedstocks and biochars and includes: cation exchange capacity (CEC), specific surface area (SSA), organic carbon (OC) and moisture percentage, pH, particle size distribution, and proximate and ultimate analysis. Also described in the protocol are the analyses of the feedstocks and biochars for contaminants including polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs), metals and mercury as well as nutrients (phosphorous, nitrite and nitrate and ammonium as nitrogen). The protocol also includes the biological testing procedures, earthworm avoidance and germination assays. Based on the quality assurance / quality control (QA/QC) results of blanks, duplicates, standards and reference materials, all methods were determined adequate for use with biochar and feedstock materials. All biochars and feedstocks were well within the criterion set by the IBI and there were little differences among biochars, except in the case of the biochar produced from construction waste materials. This biochar (referred to as Old biochar) was determined to have elevated levels of arsenic, chromium, copper, and lead, and failed the earthworm avoidance and germination assays. Based on these results, Old biochar would not be appropriate for use as a soil amendment for carbon sequestration, substrate quality improvements or remediation.

Biochar is a carbon-rich material used as a soil amendment with the ability to sustainably sequester carbon, improve substrate quality and sorb contaminants. This protocol describes the 17 analytical methods used for the characterization of biochar, which is required prior to large scale implementation of these amendments in the environment.

物理，化学和六Biochars生产的生物表征污染场地修复的

The physical and chemical properties of biochar vary based on feedstock sources and production conditions, making it possible to engineer biochars with ...

科研

环境科学

Characterization and Application of Passive Samplers for Monitoring of Pesticides in Water

Five different water passive samplers were calibrated under laboratory conditions for measurement of 124 legacy and current used pesticides. This study provides a protocol for the passive sampler preparation, calibration, extraction method and instrumental analysis. Sampling rates (RS) and passive sampler-water partition coefficients (KPW) were calculated for silicone rubber, polar organic chemical integrative sampler POCIS-A, POCIS-B, SDB-RPS and C18 disk. The uptake of the selected compounds depended on their physicochemical properties, i.e., silicone rubber showed a better uptake for more hydrophobic compounds (log octanol-water partition coefficient (KOW) &#62; 5.3), whereas POCIS-A, POCIS-B and SDB-RPS disk were more suitable for hydrophilic compounds (log KOW &#60; 0.70).

A protocol about the characterization and application of five different passive sampling devices is presented.

表征和被动采样的应用在农药水监测

Five different water passive samplers were calibrated under laboratory conditions for measurement of 124 legacy and current used pesticides. This study ...

Methods of Soil Resampling to Monitor Changes in the Chemical Concentrations of Forest Soils

Recent soils research has shown that important chemical soil characteristics can change in less than a decade, often the result of broad environmental changes. Repeated sampling to monitor these changes in forest soils is a relatively new practice that is not well documented in the literature and has only recently been broadly embraced by the scientific community. The objective of this protocol is therefore to synthesize the latest information on methods of soil resampling in a format that can be used to design and implement a soil monitoring program. Successful monitoring of forest soils requires that a study unit be defined within an area of forested land that can be characterized with replicate sampling locations. A resampling interval of 5 years is recommended, but if monitoring is done to evaluate a specific environmental driver, the rate of change expected in that driver should be taken into consideration. Here, we show that the sampling of the profile can be done by horizon where boundaries can be clearly identified and horizons are sufficiently thick to remove soil without contamination from horizons above or below. Otherwise, sampling can be done by depth interval. Archiving of sample for future reanalysis is a key step in avoiding analytical bias and providing the opportunity for additional analyses as new questions arise.

Repeated soil sampling has recently been shown to be an effective way to monitor forest soil change over years and decades. To support its use, a protocol is presented that synthesizes the latest information on soil resampling methods to aid in the design and implementation of successful soil monitoring programs.

土壤重采样的方法监控森林土壤的化学浓度的变化

Recent soils research has shown that important chemical soil characteristics can change in less than a decade, often the result of broad environmental ...

Ecotoxicological Method with Marine Bacteria Vibrio anguillarum to Evaluate the Acute Toxicity of Environmental Contaminants

Bacteria are an important component of the ecosystem, and microbial community alterations can have a significant effect on biogeochemical cycling and food webs. Toxicity testing based on microorganisms are widely used because they are relatively quick, reproducible, cheap, and are not associated with ethical issues. Here, we describe an ecotoxicological method to evaluate the biological response of the marine bacterium Vibrio anguillarum. This method assesses the acute toxicity of chemical compounds, including new contaminants such as nanoparticles, as well as environmental samples. The endpoint is the reduction of bacterial culturability (i.e., the capability to replicate and form colonies) due to exposure to a toxicant. This reduction can be generally referred to as mortality. The test allows for the determination of the LC50, the concentration that causes a 50% decrease of bacteria actively replicating and forming colonies, after a 6 h exposure. The culturable bacteria are counted in terms of colony forming units (CFU), and the &#34;mortality&#34; is evaluated and compared to the control. In this work, the toxicity of copper sulphate (CuSO4) was evaluated. A clear dose-response relationship was observed, with a mean LC50 of 1.13 mg/L, after three independent tests. This protocol, compared to existing methods with microorganisms, is applicable in a wider range of salinity and has no limitations for colored/turbid samples. It uses saline solution as the exposure medium, avoiding any possible interferences of growth medium with the investigated contaminants. The LC50 calculation facilitates comparisons with other bioassays commonly applied to ecotoxicological assessments of the marine environment.

Ecotoxicological Method with Marine Bacteria Vibrio anguillarum to Evaluate the Acute Toxicity of Environmental Contaminants

This work describes a new protocol to assess the ecotoxicity of pollutants, including emerging contaminants such as nanomaterials, using the marine bacterium Vibrio anguillarum. This method allows for the determination of the LC50, or mortality, the concentration that causes a 50% decrease in bacteria culturability, after a 6 h exposure.

生物毒理学方法与海洋细菌<em&gt;鳗弧菌</em&gt;评估环境污染物的急性毒性

Bacteria are an important component of the ecosystem, and microbial community alterations can have a significant effect on biogeochemical cycling and food ...

Comparison of Scale in a Photosynthetic Reactor System for Algal Remediation of Wastewater

An experimental methodology is presented to compare the performance of two different sized reactors designed for wastewater treatment. In this study, ammonia removal, nitrogen removal and algal growth are compared over an 8-week period in paired sets of small (100 L) and large (1,000 L) reactors designed for algal remediation of landfill wastewater. Contents of the small and large scale reactors were mixed before the beginning of each weekly testing interval to maintain equivalent initial conditions across the two scales. System characteristics, including surface area to volume ratio, retention time, biomass density, and wastewater feed concentrations, can be adjusted to better equalize conditions occurring at both scales. During the short 8-week representative time period, starting ammonia and total nitrogen concentrations ranged from 3.1-14 mg NH3-N/L, and 8.1-20.1 mg N/L, respectively. The performance of the treatment system was evaluated based on its ability to remove ammonia and total nitrogen and to produce algal biomass. Mean &#177; standard deviation of ammonia removal, total nitrogen removal and biomass growth rates were 0.95&#177;0.3 mg NH3-N/L/day, 0.89&#177;0.3 mg N/L/day, and 0.02&#177;0.03 g biomass/L/day, respectively. All vessels showed a positive relationship between the initial ammonia concentration and ammonia removal rate (R2=0.76). Comparison of process efficiencies and production values measured in reactors of different scale may be useful in determining if lab-scale experimental data is appropriate for prediction of commercial-scale production values.

An experimental methodology is presented to compare the performance of small (100 L) and large (1,000 L) scale reactors designed for algae remediation of landfill wastewater. System characteristics, including surface area to volume ratio, retention time, biomass density, and wastewater feed concentrations, can be adjusted based on application.

规模在光合反应器系统对污水的整治藻类比较

An experimental methodology is presented to compare the performance of two different sized reactors designed for wastewater treatment. In this study, ...

Development of New Methods for Quantifying Fish Density Using Underwater Stereo-video Tools

The use of video camera systems in ecological studies of fish continues to gain traction as a viable, non-extractive method of measuring fish lengths and estimating fish abundance. We developed and implemented a rotating stereo-video camera tool that covers a full 360 degrees of sampling, which maximizes sampling effort compared to stationary camera tools. A variety of studies have detailed the ability of static, stereo-camera systems to obtain highly accurate and precise measurements of fish; the focus here was on the development of methodological approaches to quantify fish density using rotating camera systems. The first approach was to develop a modification of the metric MaxN, which typically is a conservative count of the minimum number of fish observed on a given camera survey. We redefine MaxN to be the maximum number of fish observed in any given rotation of the camera system. When precautions are taken to avoid double counting, this method for MaxN may more accurately reflect true abundance than that obtained from a fixed camera. Secondly, because stereo-video allows fish to be mapped in three-dimensional space, precise estimates of the distance-from-camera can be obtained for each fish. By using the 95% percentile of the observed distance from camera to establish species-specific areas surveyed, we account for differences in detectability among species while avoiding diluting density estimates by using the maximum distance a species was observed. Accounting for this range of detectability is critical to accurately estimate fish abundances. This methodology will facilitate the integration of rotating stereo-video tools in both applied science and management contexts.

We describe a new method for counting fishes, and estimating relative abundance (MaxN) and fish density using rotating stereo-video camera systems. We also demonstrate how to use distance from camera (Z distance) to estimate species-specific detectability.

利用水下立体视频工具量化鱼类密度的新方法研究进展

The use of video camera systems in ecological studies of fish continues to gain traction as a viable, non-extractive method of measuring fish lengths and ...

Protocol for Acute and Chronic Ecotoxicity Testing of the Turquoise Killifish Nothobranchius furzeri

The killifish Nothobranchius furzeri is an emerging model organism in the field of ecotoxicology and its applicability in acute and chronic ecotoxicity testing has been demonstrated. Overall, the sensitivity of the species to toxic compounds is in the range with, or higher than, that of other model species.
This work describes protocols for acute, chronic, and multigenerational bioassays of single and combined stressor effects on N. furzeri. Due to its short maturation time and life-cycle, this vertebrate model allows the study of endpoints such as maturation time and fecundity within four months. Transgenerational full life-cycle exposure trials can be performed in as little as 8 months. Since this species produces eggs that are drought-resistant and remain viable for years, the on-site culture of the species is not needed but individuals can be recruited when required. The protocols are designed to measure life-history traits (mortality, growth, fecundity, weight) and critical thermal maximum.

Protocol for Acute and Chronic Ecotoxicity Testing of the Turquoise Killifish Nothobranchius furzeri

In this work, we describe an acute, chronic and multigenerational bioassay to study the effects of single and combined stressors on the Turquoise killifish Nothobranchius furzeri. This protocol is designed to study life-history traits (mortality, growth, fecundity, weight) and critical thermal maximum.

绿松石鳉鱼的急性和慢性毒性测试协议Nothobranchius furzeri

The killifish Nothobranchius furzeri is an emerging model organism in the field of ecotoxicology and its applicability in acute and chronic ecotoxicity ...

Visualizing Efficacy of Pesticides Against Disease Vector Mosquitoes in the Field

Efficacy of public health pesticides targeting nuisance and disease-vector insects such as mosquitoes, sand flies, and filth-breeding flies is not uniform across ecological zones. To best protect public and veterinary health from these insects, the environmental limitations of pesticides need to be investigated to inform effective use of the most appropriate pesticide formulations and techniques. We have developed a research program to evaluate combinations of pesticides, pesticide application equipment, and application techniques in hot-arid desert, hot-humid tropical, warm and cool temperate, and urban locations to derive pesticide use guidelines specific to target insect and environment. To these ends we designed a system of protocols to support efficient, cost-effective, portable, and standardized evaluation of a diverse range of pesticides and equipment across multiple environments. At the core of these protocols is the use of an array of small cages with colony-reared sentinel mosquitoes (adults and immatures) and sand flies (adults), strategically arranged in natural habitats and exposed to pesticide spray. Spatial and temporal patterns of pesticide efficacy are derived from percent mortality in sentinel cages, then mapped and visualized in a geographic information system. Maps of sentinel mortality data may be statistically compared to evaluate relative efficacy of a pesticide across multiple environments, or to study multiple pesticides in a single environment. Protocols may be modified to accommodate a variety of scenarios, including, for example, the vertical orientation of sentinels in canopy habitats or simultaneous testing of ground and aerial application methods.

The efficacy of public health pesticides targeting nuisance and disease-vector insects is not uniform across different ecological zones. Here we present a system of techniques using captive vector insects as sentinels for pesticide efficacy to derive electronic maps supporting the standard evaluation of pesticides across multiple environments.

农药在田间对病媒蚊子的影响

Efficacy of public health pesticides targeting nuisance and disease-vector insects such as mosquitoes, sand flies, and filth-breeding flies is not uniform ...

Composition and Distribution Analysis of Bioaerosols Under Different Environmental Conditions

Variable microorganisms in particulate matter (PM) under different environmental conditions may have significant impacts on human health. In this study, we described a protocol for multiple analyses of the biological compositions in environmental PM. Five experiments are presented: (1) PM number monitoring by using a laser particle counter; (2) PM collection by using a cyclonic aerosol sampler; (3) PM collection by using a high-volume air sampler with filters; (4) culturable microbes collection by the Andersen six-stage sampler; and (5) detection of biological composition of environmental PM by bacterial 16SrDNA and fungal ITS region sequencing. We selected hazy days and a livestock farm as two typical examples of the application in this protocol. In this study, these two sampling methods, cyclonic aerosol sampler and filter sampler, showed different sampling efficiency. The cyclonic aerosol sampler performed much better in terms of collecting bacteria, while these two methods showed the same efficiency in collecting fungi. Filter samplers can work under low temperature conditions while cyclonic aerosol samplers have a sampling limitation for temperature. A solid impacting sampler, such as an Andersen six-stage sampler, can be used to sample bioaerosols directly into the culture medium, which increases the survival rate of culturable microorganisms. However, this method mainly relies on culture while more than 99% of microbes cannot be cultured. DNA extracted from the culturable bacteria collected by the Andersen six-stage sampler and samples collected by cyclonic aerosol sampler and filter sampler were detected by bacterial 16S rDNA and fungal ITS region sequencing.All the methods above may have wide application in many fields of study, such as environmental monitoring and airborne pathogen detection. From these results, we can conclude that these methods can be used under different conditions and may help other researchers further explore the health impacts of environmental bioaerosols.

Understanding the biological composition of environmental particulate matter is important for the study of its significant impacts on human health and disease spread. Here, we used three types of bioaerosol sampling methods and a biological analysis of airborne microbes to better explore airborne microbial communities under different environmental conditions.

不同环境条件下生物气溶胶的组成及分布分析

Variable microorganisms in particulate matter (PM) under different environmental conditions may have significant impacts on human health. In this study, ...

A High-throughput Assay for the Prediction of Chemical Toxicity by Automated Phenotypic Profiling of Caenorhabditis elegans

Applying toxicity testing of chemicals in higher order organisms, such as mice or rats, is time-consuming and expensive, due to their long lifespan and maintenance issues. On the contrary, the nematode Caenorhabditis elegans (C. elegans) has advantages to make it an ideal choice for toxicity testing: a short lifespan, easy cultivation, and efficient reproduction. Here, we describe a protocol for the automatic phenotypic profiling of C. elegans in a 384-well plate. The nematode worms are cultured in a 384-well plate with liquid medium and chemical treatment, and videos are taken of each well to quantify the chemical influence on 33 worm features. Experimental results demonstrate that the quantified phenotype features can classify and predict the acute toxicity for different chemical compounds and establish a priority list for further traditional chemical toxicity assessment tests in a rodent model.

A High-throughput Assay for the Prediction of Chemical Toxicity by Automated Phenotypic Profiling of Caenorhabditis elegans

A quantitative method has been developed to identify and predict the acute toxicity of chemicals by automatically analyzing the phenotypic profiling of Caenorhabditis elegans. This protocol describes how to treat worms with chemicals in a 384-well plate, capture videos, and quantify toxicological related phenotypes.

一种用自动表型分析预测线虫化学毒性的高通量测定

Applying toxicity testing of chemicals in higher order organisms, such as mice or rats, is time-consuming and expensive, due to their long lifespan and ...