Benthic diatoms have traditionally been used as bioindicators of aquatic ecosystems. Because diatom-based monitoring of water quality is required by European legislation, molecular-based methods had emerged as useful alternatives to classical methods based on morphological identification using light microscopy. The aim of this study was to test the reliability of DNA metabarcoding combined with High-Throughput Sequencing (HTS) techniques in the bioassessment of the trophic status of 22 Mediterranean shallow ponds in NW Spain. For each pond, the Trophic Diatom Index (TDI) was calculated from inventories obtained by identification using light microscopy (LM) followed by high-throughput sequencing (HTS) at the molecular level. Ponds were subsequently classified into five water quality classes. The results showed a good correspondence between both methods, especially after applying a correction factor that depended on the biovolume of the cells. This correspondence led to the assignment to the same quality class in 59% of the ponds. The determination and quantification of valves or DNA sequences was one of the main pitfalls, which mainly included those related to the variability in the relative abundances of some species. Accordingly, ponds with similar relative abundances for the dominant species were assigned to the same quality class. Moreover, other difficulties leading the discrepancies were the misidentification of some species due to the presence of semi-cryptic taxa, the incompleteness of the reference database and the bioinformatic protocol. Thus, the validation of DNA-based methods for the identification of freshwater diatoms represents an important goal, as an alternative to using traditional methods in Mediterranean shallow ponds.

中文翻译：

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地中海池塘中基于附生硅藻的生物监测：传统显微镜与元条形码方法

传统上，底栖硅藻被用作水生生态系统的生物指标。由于欧洲法规要求基于硅藻的水质监测，基于分子的方法已成为基于光学显微镜形态识别的经典方法的有用替代方法。这项研究的目的是测试结合西班牙高通量测序（HTS）技术的DNA元条形码在西班牙西北部22个地中海浅水池营养状况生物评价中的可靠性。对于每个池塘，从通过使用光学显微镜（LM）进行鉴定，然后在分子水平进行高通量测序（HTS）进行鉴定而获得的清单中计算出营养硅藻指数（TDI）。池塘随后被分为五个水质等级。结果显示两种方法之间具有良好的对应性，尤其是在应用取决于细胞生物量的校正因子之后。这种对应关系导致在59％的池塘中分配了相同的质量等级。瓣膜或DNA序列的确定和定量是主要陷阱之一，其中主要包括与某些物种相对丰度变化有关的那些陷阱。因此，具有相对优势度的优势种相似的池塘被指定为相同的质量等级。此外，导致差异的其他困难是由于半隐式分类单元的存在，某些物种的错误识别，参考数据库的不完整和生物信息学方案。因此，