Supplementary material and data for ''Interactive effects of sedimentary turbidity and elevated water temperature on the Pugnose Shiner (<i>Miniellus anogenus</i>), a Threatened freshwater fish''
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This supplementary material and data is associated with: Fortin-Hamel, L and Chapman, LJ ''Interactive effects of turbidity and elevated water temperature on the Pugnose Shiner (<i>Miniellus anogenus</i>), a Threatened freshwater fish'' (<i>under revision</i>).See our methods for detailed information on the collection of each dataset. Please contact Liana Fortin-Hamel (liana.fortin-hamel[at]mail.mcgill.ca) for any questions or comments.<b>Abstract</b>High turbidity and elevated water temperature are environmental stressors that can co-occur in freshwater ecosystems such as when deforestation increases solar radiation and sedimentary runoff. However, we have limited knowledge about their combined impacts on fish behaviour and physiology. We explored independent and interactive effects of sedimentary turbidity and temperature on the swimming activity and both thermal- and hypoxia tolerance of the Pugnose Shiner (<i>Miniellus anogenus</i>, formerly<i> Notropis anogenus</i>), a small leuciscid fish listed as Threatened under Canada’s Species at Risk Act (SARA). Fish underwent a 15-week acclimation to two temperatures (16 °C or 25 °C) crossed with two turbidities (~0 NTU or 8.5 NTU). Swimming activity was measured during the first 8 weeks of acclimation. Fish in warm water were more active compared to those in cold water, but turbidity had no effect on activity. Behavioural response to hypoxia was measured after 12 weeks of acclimation, as the oxygen level at which fish used aquatic surface respiration (ASR). Fish in warm water engaged in ASR behaviour at higher oxygen thresholds, indicating less tolerance to hypoxia. Turbidity had no effect on ASR thresholds. Finally, thermal tolerance was measured as the critical thermal maximum (CT<sub>max</sub>) after 13 to 15 weeks of acclimation. Acclimation to warm water increased fish CT<sub>max</sub> and T<sub>ag</sub> (agitation temperature) but reduced the agitation window (°C difference between T<sub>ag</sub> and CT<sub>max</sub>) and thermal safety margin (°C difference between acclimation temperature and CT<sub>max</sub>). Furthermore, fish in warm, turbid water had a lower CT<sub>max</sub> and smaller thermal safety margin than fish in warm, clear water, indicating an interaction between turbidity and temperature. This reduced thermal tolerance observed in Pugnose Shiner in warm, turbid water highlights the importance of quantifying independent and interactive effects of multiple stressors when evaluating habitat suitability and conservation strategies for imperiled species.<b>Code/Software</b>R scripts and codes used for statistical analysis are provided in our supplementary material (R Core Team, 2022; v.4.2.1). Essential packages for statistical analysis include:<i>lme4</i> v.1.1.30<i>car</i> v.3.1-0<i>emmeans</i> v.1.8.1-1<br>
本附属材料及配套数据关联以下研究：Fortin-Hamel, L与Chapman, LJ《浊度（turbidity）与水温升高（elevated water temperature）对扁吻鲦（Pugnose Shiner，*Miniellus anogenus*，一种受胁淡水鱼类）的交互影响》（已投稿待刊）。有关各数据集的采集细节，请参阅本附属材料中的方法章节。如有任何疑问或建议，请联系Liana Fortin-Hamel（邮箱：liana.fortin-hamel[at]mail.mcgill.ca）。 <b>摘要</b>高浊度与水温升高是淡水生态系统（freshwater ecosystems）中可协同出现的环境胁迫因子（environmental stressors），例如森林砍伐可增加太阳辐射入射量并引发沉积物径流。目前学界对二者对鱼类行为与生理的联合影响认知仍较为有限。本研究旨在探讨沉积物浊度（sedimentary turbidity）与温度对扁吻鲦（*Miniellus anogenus*，曾用名*Notropis anogenus*，隶属于雅罗鱼科的小型鱼类，根据《加拿大濒危物种法案》（Species at Risk Act, SARA）被列为受胁物种）的游泳活性（swimming activity）、热耐受性（thermal tolerance）及缺氧耐受性（hypoxia tolerance）的独立与交互效应。 实验鱼类先接受为期15周的驯化（acclimation）处理，设置两个温度梯度（16 ℃与25 ℃），并分别交叉搭配两种浊度条件（约0 NTU与8.5 NTU）。游泳活性于驯化阶段的前8周内完成测定。结果显示，温水环境中的鱼类活性显著高于冷水环境个体，但浊度对游泳活性无显著影响。 驯化12周后，测定鱼类的缺氧行为响应，以鱼类启动水生表面呼吸（aquatic surface respiration, ASR）时的氧浓度作为评价指标。温水环境中的鱼类在更高氧浓度阈值下即启动ASR行为，表明其缺氧耐受性更低；而浊度对ASR阈值无显著影响。 驯化13至15周后，通过测定临界热最大值（critical thermal maximum, CT<sub>max</sub>）评估鱼类热耐受性。温水驯化可提升鱼类的CT<sub>max</sub>与搅拌温度（agitation temperature, T<sub>ag</sub>），但会缩短搅拌窗口（T<sub>ag</sub>与CT<sub>max</sub>的温差）及热安全裕度（thermal safety margin，即驯化温度与CT<sub>max</sub>的温差）。进一步分析显示，处于温水浊水环境中的鱼类，其CT<sub>max</sub>与热安全裕度均低于温水清水环境个体，表明浊度与温度存在显著交互效应。本研究中，扁吻鲦在温水浊水环境中表现出的热耐受性下降，凸显了在评估濒危物种的栖息地适宜性与保护策略时，量化多重胁迫因子独立与交互效应的重要性。 <b>代码/软件</b>本附属材料中提供了统计分析所用的R脚本与代码（R Core Team, 2022；版本4.2.1）。统计分析所需的核心R包包括： <i>lme4</i> 版本1.1.30 <i>car</i> 版本3.1-0 <i>emmeans</i> 版本1.8.1-1
