DNA barcoding approach to characterize microalgae isolated from freshwater systems in Ecuador

The use of suitable DNA barcodes and the generation of databases with reference sequences have been considered a promissory approach for the identification of Chlorophyta and Cyanophyta microalgae. In this study, we carried out a molecular characterization and identification of strains isolated from freshwater systems in Ecuador using a dual barcode method. The target sequences for Chlorophyta were 18S rDNA and <i>rbc</i>L genes, and 16S rDNA and 16S–23S rDNA intergenic spacer (ITS) for Cyanophyta. We reported these DNA barcodes for 20 different Molecular Operational Taxonomic Units (MOTUs) for Chlorophyta and 10 for Cyanophyta. Our results show that the 18S V4 hypervariable region (300 bp) is sufficient for differentiating between isolates, but <i>rbc</i>L is a determinant for genus identification in Scenedesmaceae and Chlorellaceae strains. In Cyanophyta, both barcodes enabled the genus-level assignment of 9 out of 10 MOTUs. These results highlight the necessity of a second barcode additional to small ribosomal subunit sequences to improve molecular identification. Furthermore, the present study significantly contributes to the body of Ecuadorian barcode sequences of microalgae that are currently documented, making them available for future comparative diversity studies.

选用合适的DNA条形码(DNA barcode)并构建含参考序列的数据库，被认为是绿藻门(Chlorophyta)与蓝藻门(Cyanophyta)微藻鉴定的极具应用前景的技术方案。本研究采用双条形码法，对从厄瓜多尔淡水生境中分离得到的菌株进行分子特征分析与鉴定。其中，绿藻门的靶标序列为18S核糖体DNA(18S rDNA)与核酮糖二磷酸羧化酶大亚基基因(rbcL)，蓝藻门的靶标序列为16S核糖体DNA(16S rDNA)及16S-23S核糖体DNA基因间区(ITS)。本研究共报道了20个绿藻门分子操作分类单元(Molecular Operational Taxonomic Units, MOTUs)与10个蓝藻门分子操作分类单元的DNA条形码序列。结果表明，18S rDNA的V4高变区（长度约300 bp）足以区分不同分离菌株，但rbcL基因是栅藻科(Scenedesmaceae)与小球藻科(Chlorellaceae)菌株属级鉴定的决定性标记。对于蓝藻门，两种条形码均可实现10个分子操作分类单元中的9个的属水平分类鉴定。本研究结果凸显了在核糖体小亚基序列(small ribosomal subunit sequences)之外增设第二条形码以优化分子鉴定准确性的必要性。此外，本研究显著扩充了当前已收录的厄瓜多尔微藻DNA条形码序列数据集，可为后续的多样性比较研究提供数据支持。