Biological Oceanography, Leibniz Institute of Marine Sciences (IFM-GEOMAR), Kiel, Germany.
Comp Biochem Physiol A Mol Integr Physiol. 2011 Nov;160(3):320-30. doi: 10.1016/j.cbpa.2011.06.023. Epub 2011 Jul 1.
Extensive use of fossil fuels is leading to increasing CO(2) concentrations in the atmosphere and causes changes in the carbonate chemistry of the oceans which represents a major sink for anthropogenic CO(2). As a result, the oceans' surface pH is expected to decrease by ca. 0.4 units by the year 2100, a major change with potentially negative consequences for some marine species. Because of their carbonate skeleton, sea urchins and their larval stages are regarded as likely to be one of the more sensitive taxa. In order to investigate sensitivity of pre-feeding (2 days post-fertilization) and feeding (4 and 7 days post-fertilization) pluteus larvae, we raised Strongylocentrotus purpuratus embryos in control (pH 8.1 and pCO(2) 41 Pa e.g. 399 μatm) and CO(2) acidified seawater with pH of 7.7 (pCO(2) 134 Pa e.g. 1318 μatm) and investigated growth, calcification and survival. At three time points (day 2, day 4 and day 7 post-fertilization), we measured the expression of 26 representative genes important for metabolism, calcification and ion regulation using RT-qPCR. After one week of development, we observed a significant difference in growth. Maximum differences in size were detected at day 4 (ca. 10% reduction in body length). A comparison of gene expression patterns using PCA and ANOSIM clearly distinguished between the different age groups (two-way ANOSIM: Global R=1) while acidification effects were less pronounced (Global R=0.518). Significant differences in gene expression patterns (ANOSIM R=0.938, SIMPER: 4.3% difference) were also detected at day 4 leading to the hypothesis that differences between CO(2) treatments could reflect patterns of expression seen in control experiments of a younger larva and thus a developmental artifact rather than a direct CO(2) effect. We found an up regulation of metabolic genes (between 10%and 20% in ATP-synthase, citrate synthase, pyruvate kinase and thiolase at day 4) and down regulation of calcification related genes (between 23% and 36% in msp130, SM30B, and SM50 at day 4). Ion regulation was mainly impacted by up regulation of Na(+)/K(+)-ATPase at day 4 (15%) and down regulation of NHE3 at day 4 (45%). We conclude that in studies in which a stressor induces an alteration in the speed of development, it is crucial to employ experimental designs with a high time resolution in order to correct for developmental artifacts. This helps prevent misinterpretation of stressor effects on organism physiology.
化石燃料的大量使用导致大气中二氧化碳浓度不断增加，改变了海洋的碳酸盐化学平衡，而海洋是人为二氧化碳的主要吸收汇。因此，预计到 2100 年，海洋表面的 pH 值将下降约 0.4 个单位，这是一个重大变化，可能对一些海洋物种产生负面影响。由于它们的碳酸盐骨架，海胆及其幼虫阶段被认为是可能更敏感的分类群之一。为了研究未进食（受精后 2 天）和进食（受精后 4 天和 7 天）的幼海胆浮游幼虫的敏感性，我们在对照条件下（pH8.1 和 pCO241 Pa，例如 399 μatm）和 CO2酸化海水中培养强壮真海胆胚胎，pH 为 7.7（pCO2134 Pa，例如 1318 μatm），并研究了生长、钙化和存活情况。在受精后 2 天、4 天和 7 天三个时间点，我们使用 RT-qPCR 测量了 26 个与代谢、钙化和离子调节相关的代表性基因的表达。经过一周的发育，我们观察到生长有明显差异。在第 4 天（体长减少约 10%）检测到最大的差异。主成分分析（PCA）和非参数多变量方差分析（ANOSIM）的比较清楚地区分了不同的年龄组（双向 ANOSIM：全局 R=1），而酸化作用的影响较小（全局 R=0.518）。在第 4 天还检测到基因表达模式的显著差异（ANOSIM R=0.938，SIMPER：差异 4.3%），这导致了这样的假设，即 CO2 处理之间的差异可能反映了年轻幼虫对照实验中的表达模式，因此是一种发育性假象，而不是直接的 CO2 效应。我们发现代谢基因的上调（第 4 天 ATP 合酶、柠檬酸合酶、丙酮酸激酶和硫解酶上调 10%至 20%）和钙化相关基因的下调（第 4 天 msp130、SM30B 和 SM50 下调 23%至 36%）。离子调节主要受第 4 天 Na+/K+-ATPase 上调（15%）和第 4 天 NHE3 下调（45%）的影响。我们得出的结论是，在研究胁迫因子会改变发育速度的情况下，采用具有高时间分辨率的实验设计至关重要，以便纠正发育性假象。这有助于防止对胁迫因子对生物体生理影响的错误解释。
