Bílý和 Ondřej. 西蒙得到了捷克生命科学大学的赠款的支持 [环境科学学院的内部赠款机构, CULS 布拉格 (42110 1312 3175 (20164236))]。对 Douda 的支持来自捷克科学基金会 (13-05872S)。在捷克共和国自然保护局管理的捷克淡水珍珠贻贝行动计划执行期间, 收集了关于珍珠贻贝的 bioindication 和目前发生情况的数据, 由政府资助捷克共和国, 可在

1. 网笼
注: 见图 1。
<ol><li>准备材料
	<ol>
<li>为实验部分准备材料: 1-2 L 每网笼水, 网笼 (1 主要塑料体, 2 塑料盖, 2 张特殊技术筛子与340µm 毛孔, 4 个螺栓和4坚果每笼), 钳子, 扳手, 巴斯德吸管, 过滤器, 数码相机, 三目照相解剖变焦立体显微镜, 校准网格 (显微镜设备), 5 培养皿50毫米直径, 烧杯, 2 塑料碟 (~ 25 厘米 x 15 厘米 x 3-5 厘米), 和一个塑料盒。</li>
		<li>要执行 hyporheal 安装, 请准备一个橡胶软管和一个100µm 孔网, 和一个喷瓶。有关设备的构造, 请参阅辅助文件 1: S. 1。网笼施工。</li>
	</ol></li>
	<li>组装网笼的底部和中央部分。装配容纳个人的笼子部分。首先插入一个塑料盖子, 然后一张塑料筛子, 最后主体在上面。使用四螺栓来保护它。</li>
	<li>制备生物材料
	<ol>
<li>将网笼放入含河水的塑料盘中。确保会议厅已满一半。采取 FWPM 少年 (参见补充文件 1: S. 6。生物材料) 出热绝缘盒, 并把它们在培养皿。 注: 确保突然温度变化不超过摄氏2摄氏度。</li>
		<li>使用喷瓶和过滤器, 筛过的青少年, 以清除碎石。</li>
	</ol></li>
	<li>设置显微镜和照相机。对仪器进行校准 (见补充 File1: S. 5.显微镜和 phototechnics)。在显微镜下放置一个含有少许水的培养皿。</li>
	<li>把幼仔放进笼子里 (实验室工作)
	<ol>
<li>使用巴斯德吸管从培养皿中除去一个人, 并小心地将其放在显微镜下的培养皿中。</li>
		<li>检查个人的健身通过看目镜 (~ 40 x 放大)。 注意: "好" 的健身意味着个体移动, 从一侧到另一侧旋转, 将脚从壳中推出.用巴斯德吸管除去死或低的健身者, 并将它们放在单独的培养皿中 (FWPM 打开贝壳, 不动, 脚不拔出, 碎片壳, 少年谁不控制地漂浮在水中, 一个可见的分解壳, 部分脱钙)。</li>
		<li>拍两张 FWPM 的照片, 用恒定的放大倍数 80 x 来显示良好的身体素质。请参阅补充文件 1: S. 5。显微镜和 phototechnics。保存照片。 注意: 为了很好的测量它的长度, 必须将幼鱼纵向铺设 (侧面视图)。主要的目标是采取一个高品质的图片的最大壳长度足够好, 使图片分析后。</li>
		<li>拍摄完照片后, 将幼兽插入笼子中适当的房间。记录图片和房间的数量。</li>
		<li>对网笼中所有使用的腔室重复此步骤。 注: 见补充文件 1: S. 1。网笼施工。</li>
		<li>一旦所有使用的房间都有珍珠贻贝, 把塑料筛子放在笼子上, 然后轻轻地把塑料盖上, 并确保所有部件连同坚果。</li>
		<li>在安装到交错区域的情况下, 通过其中一个管腔并将其固定在该位置, 然后采取防堵塞网格并将其绑定到底部 (参见辅助文件 1: S. 1。网笼施工)。</li>
	</ol></li>
	<li>商店少年
	<ol>
<li>用河水把笼子放进塑料盒里, 使幼兽完全浸入, 并保存在 thermobox 中。安装前, 让幼仔适应原位河水温度在安装场所 (逐渐冷却, 最大5°c 24 小时)。</li>
	</ol></li>
	<li>安装网格笼
	<ol>
<li>准备现场材料, 包括与幼鱼, 钢钉, 螺栓和金属螺母, 扳手, 场温度 dataloggers (见材料表和补充文件 1: S. 4.2 的网笼。水测量), 一个字符串, 一个相机, 现场协议, 锤子, 和铁锹。</li>
		<li>将 FWPM 幼鱼运到现场 thermobox (隔热箱), 保持稳定的水温与变化 < 2 °c。将 thermobox 与网笼放进河中, 让幼仔适应当地环境条件 (pH 值、电导率等)。</li>
		<li>安装网格笼。<ol>
<li>从字段 thermobox 中移除网格笼。为它提供两个钢钉和固定的领域数据存储器。将笼子锚定在研究区 FWPMs 典型的条件下 (例如, 在主水流的边缘, 而不是直接水流, 而不是在直水中, 而不是在阳光直射下)。<ol>
<li>对于开阔的水, 使用一对钢钉, 固定笼到河底;把它放在它的侧面和水平与河底, 下游在一个角度的45°到河流流, 向中心的河流。较低水平的边缘应约 10-15 厘米以上的河流底部表面。保持在一个地方的每个笼子之间的最低距离2米 (见补充文件 1: S. 4。网箱保养)。</li>
				<li>对于交错区, 在垂直于水流的竖直景观位置, 将网箱挖入河底, 使笼的上水平边缘平行于河底表面, 而分室位于交错应测试的深度。在实验过程中, 取出底部表面的橡胶软管的上端, 以进行水取样的可能性 (见补充文件 1: S. 4.2。水的测量)。 注: 建议在网箱上进行定期检查和维护 (参见补充文件 1: S. 4。网箱保养)。</li>
			</ol>
</li>
		</ol>
</li>
	</ol></li>
	<li>在暴露后卸载笼子并运送幼仔。为此, 把笼子从水中拉出, 清除它们的细泥沙以及漂过的物质, 把它们放进 thermobox 水中的田野里。立即将笼子运送到实验室, 开始死亡率和增长率评估。 注: 见补充文件 1: S. 3。曝光时间。在笼子和实验室环境之间的温度差超过5摄氏度的情况下, 首先必须让温度相等。</li>
	<li>通过检查每个青少年的生活/健康状况 (见步骤1.5.2 和 1.5.3), 并在一个培养皿中使用2张图像, 每只活的少年用一个恒定的放大倍数 80 x 来评估。记录的健身和数字的图片和房间。</li>
	<li>完成实验 (所有方法通用)
	<ol>
<li>在图像分析软件中进行测量。在输入图像 (步骤 1.5.3) 和输出图像 (步骤 1.9) 中, 使用图像分析软件来确定每个被评估的青少年的身体大小。使用两张照片中记录的最大总壳长度作为输入和输出中的体大小值。</li>
		<li>将测量值插入表处理器中, 并计算每个幸存少年的增长增量 (%)。</li>
		<li>利用存活个体数与网笼中所有实验个体的比值, 估计每个网笼的存活率 (%)。 注: 实验结束后, 将幸存者送回育种计划 (请参阅补充文件 1: S. 6。生物材料)。</li>
	</ol></li>
</ol>2. 砂笼
注: 见图 2。
<ol><li>准备材料
	<ol>
<li>为实验的实验室部分准备材料: 2 培养皿 (直径 ~ 8.5 厘米), 巴斯德吸管, 过滤器, 25 L 河水, 塑料盒, 筛子 (网格大小1和2毫米), 一个大塑料盒 (25 L), 一个沙笼 (参见补充文件1: S. 2。沙笼建设), 数码相机, 三目照相解剖变焦立体显微镜, 校准网格 (显微镜设备), 排序河沙从研究区 (见步骤 2.1.3), 和协议。见材料表和补充文件 1: S. 2。沙笼建筑。</li>
		<li>准备隔离过程的材料: 圆形容器 (每笼1加 1)、2培养皿 (直径 ~ 14 厘米)、巴斯德吸管、放大镜和河水 1 L。</li>
		<li>筛河砂通过一个2毫米的筛子, 然后通过一个1毫米的筛子得到一个粒度的 1-2 毫米. 干燥的沙子, 并保存在干燥的形式, 直到需要。</li>
	</ol></li>
	<li>采取少年 (参见补充文件 1: S. 6。生物材料) 出的 thermobox, 并把他们在培养皿。使用喷瓶和过滤器, 筛过的青少年, 以清除碎石。</li>
	<li>设置显微镜和照相机 (参见辅助文件 1: S. 5.显微镜和 phototechnics)。</li>
	<li>把幼仔放进笼子里 (实验室工作)
	<ol>
<li>把沙笼放在塑料盒里。分散被排序的沙子 (参见步 2.1.3) 1/3 高度的沙笼。把水倒入盒子里。确保沙子表面在水位以下10毫米。将沙笼插入 25 L 箱水中, 并将其暴露在与少年 FWPMs 相同的温度下 (见补充文件 1: S. 6.2。生物材料的贮存) 12 小时, 避免任何暴露在阳光下的沙子。</li>
		<li>将培养皿与准备好的 FWPM 幼仔一起服用。</li>
		<li>检查个人的健身通过查看目镜 (见步骤 1.5.2)。</li>
		<li>按如下方式执行照片文档。拍一张所有发现的人的照片 (见步骤 1.5.3), 选择10最大的个体。或者, 用低放大倍数 (~ 40 x) 对所有青少年拍照, 进行批量评估, 选择10个最大的个体。保存所有的照片, 并记录他们的数字。</li>
		<li>使用喷瓶, 将 FWPM 的幼兽移入准备好的沙笼中。</li>
	</ol></li>
	<li>商店少年
	<ol>
<li>把笼子放进有河水的大塑料箱里, 使笼子完全浸入并保存在 thermobox 中。在安装前, 让幼仔适应原位河水温 (逐渐冷却, 最大5°c 24 小时)。</li>
	</ol></li>
	<li>安装沙笼
	<ol>
<li>准备现场安装材料: 砂笼, a ~ 25 L 场 thermobox, 扁石 (最小重量1公斤), 网 (网格尺寸 10 x 10 毫米), 喷瓶, 场温度 dataloggers (见材料表和补充文件 1:S. 4.2. 水的测量), 一锹, 和现场协议。</li>
		<li>将笼子与幼仔运到现场 thermobox, 保持水温稳定 (2 摄氏度变化)。将田间 thermobox 与沙笼放在田间的河道中, 让 FWPM 少年适应当地环境条件 (pH 值、电导率等)。</li>
		<li>将沙笼安装到生境中, 条件典型为 FWPMs (例如, 在主水流的边缘, 在蜿蜒的水流中, 而不是直接的水流, 而不是在直接的阳光下)。<ol>
<li>对于开阔的水域, 用网将沙笼固定在平坦的石头上, 并将其放在河底。确保笼的较大一侧与流形成一个45°角。</li>
			<li>对于 Hyporheal, 将笼层与水的水流垂直地挖入河底, 使笼盖与河底表面保持水平。 注: 建议在网箱上进行定期检查和维护 (参见补充文件 1: S. 4。1. 现场检查)。</li>
		</ol>
</li>
	</ol></li>
	<li>在暴露后卸载笼子和运送幼仔 注: 见补充文件 1: S. 3。曝光时间。<ol>
<li>把笼子从水里拉出来, 把它们弄干净, 把它们放进田野里, thermobox 水。</li>
		<li>将笼子运送到实验室, 开始死亡率和增长率评估。 注: 在笼子和实验室环境之间的温度差超过5摄氏度的情况下, 必须让温度均衡。</li>
	</ol></li>
	<li>从沙子中分离 FWPM 幼仔
	<ol>
<li>准备一个圆形容器, 水深为50毫米 (分别为每个笼) 和一个额外的圆形容器。把沙子从笼子里转移到圆形容器里。用旋转的运动把打火机的微粒洗净到一个额外的容器里。</li>
		<li>逐步从这个容器中取样, 用巴斯德吸管和放大镜搜索幼仔。使用巴斯德吸管将幼仔放入培养皿中。重复此步骤, 直到找到最后一个少年, 然后在第一个阴性发现之后再10x。每洗一步后, 用沙子将清水添加到原容器中。 注意: 特别是在第一次洗完后, 适当检查其内容, 并清洁它的压载物, 如细泥沙和其他冲积层。</li>
	</ol></li>
	<li>评估实验
	<ol>
<li>检查每个青少年的健康状况 (见步骤2.4.3 和 1.5.2) 并计算幸存者人数。</li>
		<li>单独拍一张照片 (见步骤 2.4.4), 虽然这意味着每个个体都没有明确的身份。或者, 拍散照片, 从最终结果中选择10个最好的个体的子集。 注意: 两种可能性都具有类似的报告值。可能性1有一个更高的工作量的限制, 而且最高的相片放大率的好处并且因而也更高的准确性。</li>
	</ol></li>
	<li>完成实验
	<ol>
<li>在图像分析软件中执行测量。完成在网格笼中完成的实验 (参见步骤 1.10), 有以下例外: 不评估每个青少年的生长速率 (%), 但在沙笼实验中评估整个群体。 注: 实验结束后, 幸存者应返回育种计划 (请参阅补充文件6.1。生物材料的硒经文)。</li>
	</ol></li>
</ol>

<ol>
	<li>Goldberg, E. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+mussel+watch-a+first+step+in+global+marine+monitoring.">The mussel watch-a first step in global marine monitoring.</a> Marine Pollution Bulletin. 6 (7), 111-114 (1975).</li><li>Vaughn, C. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ecosystem+services+provided+by+freshwater+mussels.">Ecosystem services provided by freshwater mussels.</a> Hydrobiologia. , (2017).</li><li>Lopes-Lima, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Conservation+status+of+freshwater+mussels+in+Europe:+state+of+the+art+and+future+challenges.">Conservation status of freshwater mussels in Europe: state of the art and future challenges.</a> Biological Reviews. 92 (1), 572-607 (2017).</li><li>Strayer, D. L., Malcom, H. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Causes+of+recruitment+failure+in+freshwater+mussel+populations+in+southeastern+New+York.">Causes of recruitment failure in freshwater mussel populations in southeastern New York.</a> Ecological Applications. 22 (6), 1780-1790 (2012).</li><li>Douda, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+nitrate+nitrogen+pollution+on+Central+European+unionid+bivalves+revealed+by+distributional+data+and+acute+toxicity+testing.">Effects of nitrate nitrogen pollution on Central European unionid bivalves revealed by distributional data and acute toxicity testing.</a> Aquatic Conservation: Marine and Freshwater Ecosystems. 20 (2), 189-197 (2010).</li><li>Modesto, V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fish+and+mussels:+importance+of+fish+for+freshwater+mussel+conservation.">Fish and mussels: importance of fish for freshwater mussel conservation.</a> Fish and Fisheries. , (2017).</li><li>Bauer, G., W&#228;chtler, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Ecology and evolution of the freshwater mussels Unionoida. 145, 1-394 (2001).</li><li>Neves, R. J., Widlak, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Habitat+ecology+of+juvenile+fresh-water+mussels+(Bivalvia,+Unionidae)+in+a+headwater+stream+in+Virginia.">Habitat ecology of juvenile fresh-water mussels (Bivalvia, Unionidae) in a headwater stream in Virginia.</a> American Malacological Bulletin. 5, 1-7 (1987).</li><li>&#352;vanyga, J., Simon, O. P., Min&#225;rikov&#225;, T., Spisar, O., B&#237;l&#253;, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Z&#225;chrann&#253; program pro perlorodku &#345;&#237;&#269;n&#237; v &#268;R (Action plan for the endangered freshwater pearl mussel in the Czech Republic). , (2013).</li><li>Schmidt, C., Vandr&#233;, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ten+years+of+experience+in+the+rearing+of+young+freshwater+pearl+mussels+(Margaritifera+margaritifera).">Ten years of experience in the rearing of young freshwater pearl mussels (Margaritifera margaritifera).</a> Aquatic Conservation: Marine and Freshwater Ecosystems. 20 (7), 735-747 (2010).</li><li>Gum, B., Lange, M., Geist, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+critical+reflection+on+the+success+of+rearing+and+culturing+juvenile+freshwater+mussels+with+a+focus+on+the+endangered+freshwater+pearl+mussel+(Margaritifera+margaritifera+L.).">A critical reflection on the success of rearing and culturing juvenile freshwater mussels with a focus on the endangered freshwater pearl mussel (Margaritifera margaritifera L.).</a> Aquatic Conservation: Marine and Freshwater Ecosystems. 21 (7), 743-751 (2011).</li><li>Denic, M., Taeubert, J. E., Lange, M., Thielen, F., Scheder, C., Gumpinger, C., Geist, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Influence+of+stock+origin+and+environmental+conditions+on+the+survival+and+growth+of+juvenile+freshwater+pearl+mussels+(Margaritifera+margaritifera)+in+a+cross-exposure+experiment.">Influence of stock origin and environmental conditions on the survival and growth of juvenile freshwater pearl mussels (Margaritifera margaritifera) in a cross-exposure experiment.</a> Limnologica. 50, 67-74 (2015).</li><li>&#268;ern&#225;, M., Simon, O. P., B&#237;l&#253;, M., Douda, K., Dort, B., Galov&#225;, M., Volfov&#225;, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Within-river+variation+in+growth+and+survival+of+juvenile+freshwater+pearl+mussels+assessed+by+in+situ+exposure+methods.">Within-river variation in growth and survival of juvenile freshwater pearl mussels assessed by in situ exposure methods.</a> Hydrobiologia. , (2017).</li><li>Denic, M., Taeubert, J. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trophic+relationships+between+the+larvae+of+two+freshwater+mussels+and+their+fish+hosts.">Trophic relationships between the larvae of two freshwater mussels and their fish hosts.</a> Invertebrate Biology. 134 (2), 129-135 (2015).</li><li>Douda, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Host-dependent+vitality+of+juvenile+freshwater+mussels:+implications+for+breeding+programs+and+host+evaluation.">Host-dependent vitality of juvenile freshwater mussels: implications for breeding programs and host evaluation.</a> Aquaculture. 445, 5-10 (2015).</li><li>Buddensiek, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+culture+of+juvenile+freshwater+pearl+mussels+Margaritifera+margaritifera+L.+in+cages:+a+contribution+to+conservation+programmes+and+the+knowledge+of+habitat+requirements.">The culture of juvenile freshwater pearl mussels Margaritifera margaritifera L. in cages: a contribution to conservation programmes and the knowledge of habitat requirements.</a> Biological Conservation. 74 (1), 33-40 (1995).</li><li>Hru&#353;ka, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experience+of+semi-natural+breeding+program+of+freshwater+pearl+mussel+in+the+Czech+Republic.">Experience of semi-natural breeding program of freshwater pearl mussel in the Czech Republic.</a> Die Flussperlmuschel in Europa: Bestandssituation und Schutzma&#223;nahmen. , 69-75 (2001).</li><li>Barnhart, M. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Buckets+of+muckets:+a+compact+system+for+rearing+juvenile+freshwater+mussels.">Buckets of muckets: a compact system for rearing juvenile freshwater mussels.</a> Aquaculture. 254 (1), 227-233 (2006).</li><li>Simon, O. P., Van&#237;&#269;kov&#225;, I., B&#237;l&#253;, M., Douda, K., Patzenhauerov&#225;, H., Hru&#353;ka, J., Pelt&#225;nov&#225;, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+status+of+freshwater+pearl+mussel+in+the+Czech+Republic:+several+successfully+rejuvenated+populations+but+the+absence+of+natural+reproduction.">The status of freshwater pearl mussel in the Czech Republic: several successfully rejuvenated populations but the absence of natural reproduction.</a> Limnologica. 50, 11-20 (2015).</li><li>. A language and environment for statistical computing Available from: <a target="_blank" href="https://www.r-project.org/">https://www.r-project.org/</a> (2013)</li><li>Hastie, L. C., Yang, M. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Conservation of the freshwater pearl mussel I: captive breeding techniques. 2, (2003).</li><li>Hru&#353;ka, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nahrungsanspr&#252;che+der+Flu&#223;perlmuschel+und+deren+halbnat&#252;rliche+Aufzucht+in+der+Tschechischen+Republik.">Nahrungsanspr&#252;che der Flu&#223;perlmuschel und deren halbnat&#252;rliche Aufzucht in der Tschechischen Republik.</a> Heldia. 4 (6), 69-79 (1999).</li><li>Scheder, C., Lerchegger, B., Jung, M., Csar, D., Gumpinger, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Practical+experience+in+the+rearing+of+freshwater+pearl+mussels+(Margaritifera+margaritifera):+advantages+of+a+work-saving+infection+approach,+survival,+and+growth+of+early+life+stages.">Practical experience in the rearing of freshwater pearl mussels (Margaritifera margaritifera): advantages of a work-saving infection approach, survival, and growth of early life stages.</a> Hydrobiologia. 735 (1), 203-212 (2014).</li><li>Braun, A., Auerswald, K., Geist, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drivers+and+spatio-temporal+extent+of+hyporheic+patch+variation:+implications+for+sampling.">Drivers and spatio-temporal extent of hyporheic patch variation: implications for sampling.</a> PLoS ONE. 7 (7), e42046 (2012).</li><li>Franken, R. J. M., Storey, R. G., Williams, D. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biological,+chemical+and+physical+characteristics+of+downwelling+and+upwelling+zones+in+the+hyporheic+zone+of+a+north-temperate+stream.">Biological, chemical and physical characteristics of downwelling and upwelling zones in the hyporheic zone of a north-temperate stream.</a> Hydrobiologia. , 183-195 (2001).</li><li>Roley, S. S., Tank, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pore+water+physicochemical+constraints+on+the+endangered+clubshell+mussel+(Pleurobema+clava).">Pore water physicochemical constraints on the endangered clubshell mussel (Pleurobema clava).</a> Canadian Journal of Fisheries and Aquatic Sciences. 73 (12), 1712-1722 (2016).</li><li>Larson, J. H., Eckert, N. L., Bartsch, M. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intrinsic+variability+in+shell+and+soft+tissue+growth+of+the+freshwater+mussel+Lampsilis+siliquoidea.">Intrinsic variability in shell and soft tissue growth of the freshwater mussel Lampsilis siliquoidea.</a> PLoS ONE. 9 (11), e112252 (2014).</li><li>Lavictoire, L., Moorkens, E., Ramsey, A. D., Sinclair, W., Sweeting, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+substrate+size+and+cleaning+regime+on+growth+and+survival+of+captive-bred+juvenile+freshwater+pearl+mussels,+Margaritifera+(Linnaeus,+1758).">Effects of substrate size and cleaning regime on growth and survival of captive-bred juvenile freshwater pearl mussels, Margaritifera (Linnaeus, 1758).</a> Hydrobiologia. 766 (1), 89-102 (2015).</li><li>Hru&#353;ka, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experience+of+semi-natural+breeding+programme+of+freshwater+pearl+mussel+in+the+Czech+Republic.">Experience of semi-natural breeding programme of freshwater pearl mussel in the Czech Republic.</a> Die Flussperlmuschel in Europa: Bestandssituation und Schutzmassnahmen. , 69-75 (2000).</li><li>Bayne, B. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Physiological+components+of+growth+differences+between+individual+oysters+(Crassostrea+gigas)+and+a+comparison+with+Saccostrea+commercialis.">Physiological components of growth differences between individual oysters (Crassostrea gigas) and a comparison with Saccostrea commercialis.</a> Physiological and Biochemical Zoology. 72 (6), 705-713 (1999).</li><li>Tamayo, D., Azpeitia, K., Markaide, P., Navarro, E., Ibarrola, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Food+regime+modulates+physiological+processes+underlying+size+differentiation+in+juvenile+intertidal+mussels+Mytilus+galloprovincialis.">Food regime modulates physiological processes underlying size differentiation in juvenile intertidal mussels Mytilus galloprovincialis.</a> Marine Biology. 163 (6), (2016).</li></ol>

作者没有什么可透露的。

曝光时间:
即使是一个月的裸露的网笼显示了明显的增长, 反映了现场之间的差异 (图 3), 所以它们是非常有用的快速和容易检测的现场描述。然而, 结果的相关性取决于条件的短期状态, 可以振荡。特别是, 短期降雨事件可以起到作用。相比之下, 不可预知的偶发污染可能并不总是被记录下来。在现场 V (图 4A), 水化学分析检测到一波强?...

实验生物体的现场暴露和随后对其病情的评估是获取环境质量信息和 (特别是) 物种适宜性的一种可能方法。在动物体内, 这种 bioindication 主要适用于能够生活在有限的有界空间中的小无脊椎动物。双壳贝类的年轻阶段 (双壳类) 是一个这样适当的有机体小组1。
家庭 Unionidae 的双壳贝类是水生生态系统的一个非常重要的组成部分2。然而, 这些物种往往严重濒危, 特别是在溪流和河流。其中一些被描绘成 "伞种", 其保护与整个溪流群落生境的养护密切相关, 需要采取综合办法3。这些动物的生命周期与许多环境成分有关, 从水化学4,5到鱼类种群的变化, 作为贻贝幼虫宿主6。因为贻贝幼鱼通常代表贻贝生命周期的关键阶段, 所以在这个阶段适合它们发展的地点对于一个地区成功的物种种群发展至关重要。
淡水珍珠贻贝 (FWPM,母母;Unionida, 双壳类) 是一种濒危的双壳贝类, 发生在贫营养泥炭欧洲溪流中。他们的人数在20世纪的整个发生地区急剧下降。在大多数中欧人口中, 目前物种繁殖的下降似乎主要是由于青少年在生命的头几年中生存率极低至零。它假设, 少年 FWPMs 居住在浅交错区7年, 其中条件和他们的可变性仍然没有被描述的很好。此外, 直到他们的第二年的生活, 幼仔只有一个维度约1毫米, 所以他们很难找到在大量的泥沙下自然条件8。因此, 对圈养幼仔进行实验是研究其生态学的必要条件。
在捷克9淡水珍珠贻贝行动计划中, 每年有数以千计的青少年从半自然育种计划中崛起。然而, 有一个问题是, 哪些地方和生境适合这些青少年成功的人口支持, 或最终物种重新引进。在原位bioindications 现在找到答案的方法。
尽管在早先的一些著作中观察到幼贻贝在生物指标10的适宜性, 但最近的一些研究证实, 在暴露笼中, 幼贻贝的存活率不一致。青少年暴露方法在水质检测中的适用性11、12、13。此外, 还证明了在解释这些特定研究的结果时需要考虑的几个因素, 例如14的股票来源和幼虫条件15的持续影响。
问题在于如何在测试现场安装实验性青少年, 以及如何最有效地评估其状况。首次严格应用的现场曝光方法与青少年 FWPMs 发表了 Buddensiek16。青少年 FWPM 个体被保存在板笼中, 暴露在溪流的自由流动的水中, 在几个星期的暴露后, 它们的生存和生长被量化。该方法最初是作为一种半人工育种方法开发的, 但同时也强调了其对生境需求和水质评价的适用性。虽然 FWPM 少年的存活率自然非常低, 在一个月/年的规模, 只有很小数量的动物将生存, 生存率可以是一个良好的标志的环境影响的16周的规模。经过多年的研究, 进一步发展了暴露方法, 以保持实验性的幼贻贝在溪流中栖息, 并评估其生长和成活率;这些包括沙盒17, 贻贝筒仓基于一个上升的原则18, 和各种其他接触笼 (由牙龈和同事总结)11。由于青少年自然发生在浅交错区7, 实验装置在流底的应用是非常可取的。
在本文中, 我们描述了使用两个曝光装置的 FWPMs: i) 修改 Buddensiek 板笼 ("网笼") 也使 bioindication 测试 hyporheal 条件;和 ii) Hruška 沙盒 ("沙笼")。该议定书描述了这两种方法在开阔水域和交错条件下的应用 (即四种暴露的变体)。这些方法在捷克淡水珍珠贻贝9号行动计划中的应用已有15年的时间逐步修改和扩大, 并经过了一系列实验验证。

<table><tbody><tr><td>&lt;strong&gt;biological material maintenance and care&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Freshwater pearl mussel juveniles</td><td>any</td><td>NA</td><td>from a FWPM breeding programme</td></tr><tr><td>plastic boxes</td><td>any</td><td>NA</td><td></td></tr><tr><td>thermobox</td><td>MERCI</td><td>212,070,600,030</td><td>There are many possibilities. This is one example only.</td></tr><tr><td>field thermobox (ca25 l)</td><td>any</td><td>NA</td><td>cold box (insulated box) commonly used for food transport</td></tr><tr><td>river water</td><td>any</td><td>NA</td><td></td></tr><tr><td>Petri dishes</td><td>any</td><td>NA</td><td></td></tr><tr><td>plastic Pasteur pipettes with balloon bulb (droppers)</td><td>any</td><td>NA</td><td>hole diameter 1 mm</td></tr><tr><td>hydrogen peroxide</td><td>any</td><td>NA</td><td></td></tr><tr><td>plastic container (ca 50 l) for river water</td><td>any</td><td>NA</td><td></td></tr><tr><td>plastic tea strainer</td><td>any</td><td>NA</td><td>commonly used in kitchen</td></tr><tr><td>&lt;strong&gt;mesh cages construction&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>main plastic bodies</td><td>any</td><td>NA</td><td></td></tr><tr><td>plactic covers</td><td>any</td><td>NA</td><td></td></tr><tr><td>special technical sieves 340 &amp;micro;m</td><td>Silk &amp;amp;Progress</td><td>UHELON 20 T</td><td></td></tr><tr><td>special technical sieves 100 &amp;micro;m</td><td>Silk &amp;amp;Progress</td><td>UHELON 67 M</td><td></td></tr><tr><td>rubber hose (diameter 5.5 mm)</td><td>any</td><td>NA</td><td></td></tr><tr><td>steel bolts</td><td>any</td><td>NA</td><td></td></tr><tr><td>steel nuts</td><td>any</td><td>NA</td><td></td></tr><tr><td>spanner</td><td>any</td><td>NA</td><td></td></tr><tr><td>steel spikes</td><td>any</td><td>NA</td><td></td></tr><tr><td>pliers</td><td>any</td><td>NA</td><td></td></tr><tr><td>beakers</td><td>any</td><td>NA</td><td></td></tr><tr><td>plastic dishes (ca. 25x15x3-5cm)</td><td>any</td><td>NA</td><td></td></tr><tr><td>squirt bottle</td><td>any</td><td>NA</td><td></td></tr><tr><td>field protocols</td><td>any</td><td>NA</td><td></td></tr><tr><td>stationery</td><td>any</td><td>NA</td><td></td></tr><tr><td>plastic container</td><td>any</td><td>NA</td><td></td></tr><tr><td>string</td><td>any</td><td>NA</td><td></td></tr><tr><td>hammer</td><td>any</td><td>NA</td><td></td></tr><tr><td>&lt;strong&gt;sandy cages construction and use&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>sieve 1 mm</td><td>any</td><td>NA</td><td></td></tr><tr><td>sieve 2 mm</td><td>any</td><td>NA</td><td></td></tr><tr><td>special technical sieves 340 &amp;micro;m</td><td>Silk &amp;amp;Progress</td><td>UHELON 20 T</td><td></td></tr><tr><td>plastic boxes with tight-fitting lid</td><td>any</td><td>NA</td><td></td></tr><tr><td>hot melt adhesive</td><td>any</td><td>NA</td><td></td></tr><tr><td>plastic box (ca 250 x 150 x 100 cm)</td><td></td><td></td><td></td></tr><tr><td>big plastic box (ca 25 l)</td><td>any</td><td>NA</td><td></td></tr><tr><td>flat stone</td><td>any</td><td>NA</td><td></td></tr><tr><td>net</td><td>any</td><td>NA</td><td></td></tr><tr><td>river sand</td><td>any</td><td>NA</td><td></td></tr><tr><td>round containers</td><td>any</td><td>NA</td><td></td></tr><tr><td>magnifying glasses</td><td>Carson</td><td>Carson CP 60</td><td>There ar many possibilities. This is one example only</td></tr><tr><td>&lt;strong&gt;cages installation and maintenance&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>field temperature dataloggers</td><td>ONSET</td><td>UA-001-64</td><td>http://www.onsetcomp.com/products/data-loggers/ua-001-64</td></tr><tr><td>spade</td><td>any</td><td>NA</td><td></td></tr><tr><td>toothbrush</td><td>any</td><td>NA</td><td></td></tr><tr><td>&lt;strong&gt;experiment evaluation&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>trinocular dissecting zoom stereo microscope</td><td>Bresser optic</td><td>ICD 10x-160x</td><td>There are many possibilities. This is one example only.</td></tr><tr><td>digital camera/ electronic eyepiece</td><td>Bresser optic</td><td>MikroCamLab 5M</td><td>There are many possibilities. This is one example only.</td></tr><tr><td>Calibration gird</td><td>Am Scope</td><td>SKU: MR100</td><td>There are many possibilities. This is one example only.</td></tr><tr><td>external power source with two movable light guides</td><td>Arsenal</td><td>K1309010150021</td><td>There are many possibilities. This is one example only.</td></tr><tr><td>Image software</td><td>ImageJ software</td><td></td><td>There are many possibilities. This is one example only.</td></tr><tr><td>table processor</td><td>MS excel</td><td></td><td>There are many possibilities. This is one example only.</td></tr></tbody></table>

bioindication testing of stream environment suitability for young freshwater pearl mussels using in situ exposure methods

对淡水贻贝生境适宜性的认识是保护濒危物种群体的重要一步。我们描述了一个协议,在贫营养泥炭河流域内进行现场青少年暴露试验, 为期一个月和三月的时间。两种方法 (在两种修改中) 都是为了评估青少年的生长和生存率。方法和修改在价值为地方 bioindication 不同, 并且每个有它的好处并且制约。沙笼法适用于一大组个体, 但只有部分个体被测量, 结果被大量评估。在网格笼法中, 分别对个体进行了单独的保存和测量, 但计算出了低个体数。公开的水接触改性比较容易应用;它显示了青少年的生长潜力, 也可以有效的水毒性试验。在床内暴露的修改需要很高的工作量, 但更接近自然的青少年环境的条件, 它是更好地报告的实际适宜性的地方。另一方面, 由于其高交错的环境变异性, 此修改需要更多的复制。

采用四种 bioindication 方法 (开放水沙笼、床内砂笼、露天网箱、内网网箱), 研究了上伏尔塔瓦河河流域 FWPMs 的环境条件适宜性 (捷克波希米亚森林共和国)。这条河代表一 FWPM 残余地方在中欧之内19。在这里, 我们提出了一组特别选定的结果, 说明了四方法的最重要的方面。进一步的细节被描述在一个全面的研究Černá等。13。
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Watch this Scientific Journal Video about Bioindication Testing of Stream Environment Suitability for Young Freshwater Pearl Mussels Using In Situ Exposure Methods at JoVE.com

Bioindication Testing of Stream Environment Suitability for Young Freshwater Pearl Mussels Using In Situ Exposure Methods

原位bioindications 能够确定环境对濒危贻贝种类的适宜性。我们描述了两种方法基于青少年暴露的淡水珍珠贻贝在网箱贫营养泥炭河生境。这两种方法都是在开放水和交错水环境的变型中实现的。

用原位曝光法测定淡水珍珠贻贝 Bioindication 的流环境适宜性

Knowledge of habitat suitability for freshwater mussels is an important step in the conservation of this endangered species group. We describe a protocol ...

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Bioindication Testing of Stream Environment Suitability for Young Freshwater Pearl Mussels Using In Situ Exposure Methods

7.3K 次观看.  Czech University of Life Sciences Prague. 原位bioindications 能够确定环境对濒危贻贝种类的适宜性。我们描述了两种方法基于青少年暴露的淡水珍珠贻贝在网箱贫营养泥炭河生境。这两种方法都是在开放水和交错水环境的变型中实现的。

视频: Bioindication Testing of Stream Environment Suitability for Young Freshwater Pearl Mussels Using In Situ Exposure Methods

Use of Chironomidae (Diptera) Surface-Floating Pupal Exuviae as a Rapid Bioassessment Protocol for Water Bodies

Rapid bioassessment protocols using benthic macroinvertebrate assemblages have been successfully used to assess human impacts on water quality. Unfortunately, traditional benthic larval sampling methods, such as the dip-net, can be time-consuming and expensive. An alternative protocol involves collection of Chironomidae surface-floating pupal exuviae (SFPE). Chironomidae is a species-rich family of flies (Diptera) whose immature stages typically occur in aquatic habitats. Adult chironomids emerge from the water, leaving their pupal skins, or exuviae, floating on the water&#8217;s surface. Exuviae often accumulate along banks or behind obstructions by action of the wind or water current, where they can be collected to assess chironomid diversity and richness. Chironomids can be used as important biological indicators, since some species are more tolerant to pollution than others. Therefore, the relative abundance and species composition of collected SFPE reflect changes in water quality. Here, methods associated with field collection, laboratory processing, slide mounting, and identification of chironomid SFPE are described in detail. Advantages of the SFPE method include minimal disturbance at a sampling area, efficient and economical sample collection and laboratory processing, ease of identification, applicability in nearly all aquatic environments, and a potentially more sensitive measure of ecosystem stress. Limitations include the inability to determine larval microhabitat use and inability to identify pupal exuviae to species if they have not been associated with adult males.

Use of Chironomidae Diptera Surface-Floating Pupal Exuviae as a Rapid Bioassessment Protocol for Water Bodies

Rapid bioassessment protocols using benthic macroinvertebrates are often used to monitor and assess water quality. An efficient protocol involves collections of Chironomidae surface-floating pupal exuviae (SFPE). Here, techniques for field collection, laboratory processing, slide mounting, and identification of Chironomidae SFPE are described.

使用摇蚊科（双翅目）的表面浮蛹蜕为快速生物议定书水体

Rapid bioassessment protocols using benthic macroinvertebrate assemblages have been successfully used to assess human impacts on water quality. ...

科研

环境科学

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.

Endocrine Disrupting Compounds (EDC) pose a substantial risk to the aquatic environment. Municipal wastewater treatment plants are major contributors to the estrogenic potency of surface waters. The methodology provided in this paper allows for an assessment of the efficacy and suitability of wastewater treatment processes with respect to EDC removal.

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 ...

Salinity-dependent Toxicity Assay of Silver Nanocolloids Using Medaka Eggs

Salinity is an important characteristic of the aquatic environment. For aquatic organisms it defines the habitats of freshwater, brackish water, and seawater. Tests of the toxicity of chemicals and assessments of their ecological risks to aquatic organisms are frequently performed in freshwater, but the toxicity of chemicals to aquatic organisms depends on pH, temperature, and salinity. There is no method, however, for testing the salinity dependence of toxicity to aquatic organisms. Here, we used medaka (Oryzias latipes) because they can adapt to freshwater, brackish water, and seawater. Different concentrations of embryo-rearing medium (ERM) (1x, 5x, 10x, 15x, 20x, and 30x) were employed to test the toxicity of silver nanocolloidal particles (SNCs) to medaka eggs (1x ERM and 30x ERM have osmotic pressures equivalent to freshwater and seawater, respectively). In six-well plastic plates, 15 medaka eggs in triplicate were exposed to SNCs at 10 mg/L&#8722;1 in different concentrations of ERM at pH 7 and 25 &#176;C in the dark.
We used a dissecting microscope and a micrometer to measure heart rate per 15 sec and eye diameter on day 6 and full body length of the larvae on hatching day (section 4). The embryos were observed until hatching or day 14; we then counted the hatching rate every day for 14 days (section 4). To see silver accumulation in embryos, we used inductively coupled plasma mass spectrometry to measure the silver concentration of test solutions (section 5) and dechorionated embryos (section 6).The toxicity of the SNCs to medaka embryos obviously increased with increasing salinity. This new method allows us to test the toxicity of chemicals in different salinities.

Embryonic stages are the most susceptible to xenobiotics. Although chemical toxicity depends on salinity, no method exists to test the salinity dependence of toxicity to aquatic organisms. Here, we describe a new and high-throughput method for determining the salinity dependence of toxicity to aquatic embryos.

白银纳米胶体的盐度依赖性毒力测定使用青鳉鸡蛋

Salinity is an important characteristic of the aquatic environment. For aquatic organisms it defines the habitats of freshwater, brackish water, and ...

发育生物学

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 ...

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 ...

Design and Use of an Apparatus for Quantifying Bivalve Suspension Feeding at Sea

As shellfish aquaculture moves from coastal embayments and estuaries to offshore locations, the need to quantify ecosystem interactions of farmed bivalves (i.e., mussels, oysters, and clams) presents new challenges. Quantitative data on the feeding behavior of suspension-feeding mollusks is necessary to determine important ecosystem interactions of offshore shellfish farms, including their carrying capacity, the competition with the zooplankton community, the availability of trophic resources at different depths, and the deposition to the benthos. The biodeposition method is used to quantify feeding variables in suspension-feeding bivalves in a natural setting and represents a more realistic proxy than laboratory experiments. This method, however, relies upon a stable platform to satisfy the requirements that water flow rates supplied to the shellfish remain constant and the bivalves are undisturbed. A flow-through device and process for using the biodeposition method to quantify the feeding of bivalve mollusks were modified from a land-based format for shipboard use by building a two-dimensional gimbal table around the device. Planimeter data reveal a minimal pitch and yaw of the chambers containing the test shellfish despite boat motion, the flow rates within the chambers remain constant, and operators are able to collect the biodeposits (feces and pseudofeces) with sufficient consistency to obtain accurate measurements of the bivalve clearance, filtration, selection, ingestion, rejection, and absorption at offshore shellfish aquaculture sites.

A flow-through device for using the biodeposition method to quantify filtration and feeding behavior of bivalve mollusks was modified for shipboard use. A two-dimensional gimbal table built around the device isolates the apparatus from boat motion, thereby allowing the accurate quantification of bivalve filtration variables at offshore shellfish aquaculture sites.

海上双壳贝类悬浮饲料定量装置的设计与应用

As shellfish aquaculture moves from coastal embayments and estuaries to offshore locations, the need to quantify ecosystem interactions of farmed bivalves ...

Development and Testing of Species-specific Quantitative PCR Assays for Environmental DNA Applications

New, non-invasive methods for detecting and monitoring species presence are being developed to aid in fisheries and wildlife conservation management. The use of environmental DNA (eDNA) samples for detecting macrobiota is one such group of methods that is rapidly becoming popular and being implemented in national management programs. Here we focus on the development of species-specific targeted assays for probe-based quantitative PCR (qPCR) applications. Using probe-based qPCR offers greater specificity than is possible with primers alone. Furthermore, the ability to quantify the amount of DNA in a sample can be useful in our understanding of the ecology of eDNA and the interpretation of eDNA detection patterns in the field. Careful consideration is needed in the development and testing of these assays to ensure the sensitivity and specificity of detecting the target species from an environmental sample. In this protocol we will delineate the steps needed to design and test probe-based assays for the detection of a target species; including creation of sequence databases, assay design, assay selection and optimization, testing assay performance, and field validation. Following these steps will help achieve an efficient, sensitive, and specific assay that can be used with confidence. We demonstrate this process with our assay designed for populations of the mucket (Actinonaias ligamentina), a freshwater mussel species found in the Clinch River, USA.

Environmental DNA assays require rigorous design, testing, optimization and validation before the collection of field data can begin. Here, we present a protocol to take users through each step of designing a species-specific, probe-based qPCR assay for the detection and quantification of a target species DNA from environmental samples.

开发和测试针对物种的定量PCR检测环境DNA应用

New, non-invasive methods for detecting and monitoring species presence are being developed to aid in fisheries and wildlife conservation management. The ...

Hellgrammite as a Non-Conventional Biomonitor for Surface Water and Environmental Health Assessment

The larvae of dobsonflies from the genus Corydalus, commonly known as Hellgrammites, are characterized by their notable size, extensive range of occurrence, and extended period of immaturity, which can last up to one year. Hellgrammites are clearly known to exhibit sensitivity to pollution and habitat structure impacts. Given these unique features, using Corydalus texanus larvae is highly suitable as reliable biomonitoring agents to assess the ecological integrity of aquatic ecosystems. This protocol aims to provide the necessary tools for C. texanus assessment and demonstrate their efficacy through a case study. Research findings have practical implications, indicating that C. texanus larvae exhibit early-warning responses to mining pollution, bioaccumulating high amounts of heavy metals such as Zn, Fe, and Al. The presence or absence of C. texanus populations may serve as a helpful indicator for identifying potential issues related to ecosystem health. The unconventional approach has shown early warnings of pollution in mining-impacted sites, highlighting the need for timely action to protect the environment. Given their unique traits, the use of C. texanus larvae is highly suggested as a reliable non-conventional bioindicator.

This protocol describes the use of biomarkers for the early detection of deleterious impacts in aquatic ecosystems. The biomarkers are closely related to sentinel traits, and their changes aid in detecting early-warning damages.

Hellgrammite 作为地表水和环境健康评估的非常规生物监测器

The larvae of dobsonflies from the genus Corydalus, commonly known as Hellgrammites, are characterized by their notable size, extensive range of ...

Standardizing a Non-Lethal Method for Characterizing the Reproductive Status and Larval Development of Freshwater Mussels (Bivalvia: Unionida)

Actively monitoring the timing, development, and reproductive patterns of endangered species is critical when managing for population recovery. Freshwater mussels are among the most imperiled organisms in the world, but information about early larval (glochidial) development and brooding periods is still lacking for many species. Previous studies have focused on the complex life history stage when female mussels are ready to parasitize host fish, but few studies have focused on the brooding period and timing of larval development. The protocol described here allows researchers to non-lethally evaluate the state of gravidity for female mussels. The results of this study show that this method does not affect a female mussel&rsquo;s ability to stay gravid or become gravid again after sampling has been performed. The advantage of this method may permit its use on federally threatened or endangered species or other populations of high conservation concern. This protocol can be adapted for use on both preserved or live individuals and was tested on a variety of mussel species. The database provided is a repository for a breadth of information on timing of reproductive habits and will facilitate future freshwater mussel research, conservation, and recovery efforts.

Standardizing a Non-Lethal Method for Characterizing the Reproductive Status and Larval Development of Freshwater Mussels Bivalvia: Unionida

Freshwater mussel conservation is dependent on monitoring reproductive patterns and processes of species. This study standardizes a non-lethal protocol for sampling gill contents, characterizing larval development, and providing a digital repository for data collected. This protocol-database package will be an important tool for mussel researchers in recovery of imperiled species.

标准化一种非致命方法，以描述淡水贝类的生殖状况和幼虫发育（比瓦尔维亚:联合达）

Actively monitoring the timing, development, and reproductive patterns of endangered species is critical when managing for population recovery. Freshwater ...

生物学

A Toxicological and Ecotoxicological Assay Based on Mussel (Mytilus galloprovincialis) Hemocytes Motility

Hemocytes are the circulating immune-competent cells in bivalve mollusks and play a key role in several important functions of cell-mediated innate immunity. During the early stages of the immune response, hemocytes actively migrate to the site of infection. This inherent motility is a fundamental characteristic of these cells. It represents a key cellular function that integrates multiple processes, such as cell adhesion, cell signaling, cytoskeletal dynamics, and changes in cell volume. Therefore, alterations in cell motility following exposure to drugs or pollutants can serve as a useful toxicological endpoint. Despite the fundamental role of cell motility in cellular physiology, it has been poorly investigated from a toxicological perspective. This work proposes a novel in vitro method for the rapid and sensitive assessment of the toxicity and ecotoxicity of pollutants, based on evaluating the hemocyte motility of Mytilus galloprovincialis. We developed a cell motility assay on hemocytes adhering to the bottom of a 96-well polystyrene microplate. Following exposure to increasing concentrations of drugs, cell trajectories, and velocities were quantified by cell tracking under time-lapse microscopy, allowing us to measure the effects on hemocyte motility. Due to the ease of hemocyte collection from the animals in a relatively non-invasive manner, the proposed method offers an alternative test for screening the effects and mechanisms of action of pollutants and drugs. It aligns with the 3Rs (Replacement, Reduction, and Refinement) criteria, addressing ethical concerns and contributing to the reduction of vertebrate in vivo animal testing.

The article proposes a novel in vitro method for the rapid and sensitive assessment of the toxicity and ecotoxicity of pollutants, based on the motility of Mytilus galloprovincialis hemocytes. The method aims to contribute to the development of more ethical and sensitive toxicological and ecotoxicological exposure tests.

基于贻贝 （Mytilus galloprovincialis） 血细胞运动的毒理学和生态毒理学测定

Hemocytes are the circulating immune-competent cells in bivalve mollusks and play a key role in several important functions of cell-mediated innate ...

用原位曝光法测定淡水珍珠贻贝 Bioindication 的流环境适宜性

摘要

探索更多视频

使用摇蚊科（双翅目）的表面浮蛹蜕为快速生物议定书水体

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

白银纳米胶体的盐度依赖性毒力测定使用青鳉鸡蛋

生物毒理学方法与海洋细菌

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

海上双壳贝类悬浮饲料定量装置的设计与应用

开发和测试针对物种的定量PCR检测环境DNA应用

Hellgrammite 作为地表水和环境健康评估的非常规生物监测器

标准化一种非致命方法，以描述淡水贝类的生殖状况和幼虫发育（比瓦尔维亚:联合达）

基于贻贝（Mytilus galloprovincialis）血细胞运动的毒理学和生态毒理学测定

用原位曝光法测定淡水珍珠贻贝 Bioindication 的流环境适宜性

摘要

探索更多视频

使用摇蚊科（双翅目）的表面浮蛹蜕为快速生物议定书水体

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

白银纳米胶体的盐度依赖性毒力测定使用青鳉鸡蛋

生物毒理学方法与海洋细菌

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

海上双壳贝类悬浮饲料定量装置的设计与应用

开发和测试针对物种的定量PCR检测环境DNA应用

Hellgrammite 作为地表水和环境健康评估的非常规生物监测器

标准化一种非致命方法，以描述淡水贝类的生殖状况和幼虫发育（比瓦尔维亚:联合达）

基于贻贝 （Mytilus galloprovincialis） 血细胞运动的毒理学和生态毒理学测定

基于贻贝（Mytilus galloprovincialis）血细胞运动的毒理学和生态毒理学测定