Avoiding quantification bias in metabarcoding: Application of a cell biovolume correction factor in diatom molecular biomonitoring
Avoiding quantification bias in metabarcoding: Application of a cell biovolume correction factor in diatom molecular biomonitoring
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避免元条形码中的量化偏差：细胞生物体积校正因子在硅藻分子生物监测中的应用
DOI：
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发表时间：
2017
期刊：
影响因子：
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通讯作者：
I. Domaizon
中科院分区：
文献类型：
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作者：
V. Vasselon;A. Bouchez;F. Rimet;S. Jacquet;R. Trobajo;M. Corniquel;K. Tapolczai;I. Domaizon
近年来，环境DNA元条形码研究取得了显著进展。然而，其量化物种相对丰度的能力仍然不确定，限制了其在生物监测中的应用。在硅藻，虽然rbcL基因似乎是一个合适的条形码硅藻，提供相关的定性数据来描述分类组成，改进的物种量化仍然是必需的。在这里，我们假设rbcL拷贝数与硅藻细胞生物体积相关（如前所述的18S基因），并应应用基于细胞生物体积的校正因子（CF）来提高分类群的定量。我们使用具有对比细胞生物体积的八种硅藻物种的纯培养物进行了实验室实验，以（1）验证rbcL拷贝数（通过qPCR估计）与硅藻细胞生物体积之间的关系，以及（2）定义潜在的CF。为了评估CF效率，通过混合来自八个物种的不同量的DNA来创建五个模拟群落，并使用HTS并靶向相同的rbcL条形码进行测序。正如预期的那样，通过CF校正DNA读数比例提高了形态学和分子库存之间的一致性。CF的最后验证获得环境样品（从80底栖生物膜的metabarcoding数据），CF的应用程序允许分子和形态水质指数之间的差异减少47%。总体而言，我们的结果强调了应用CF因子的有用性，该因子可有效减少对高生物量物种的高估，纠正硅藻元巴编码研究中的定量偏差并改善最终水质评估。
In recent years, remarkable progress has been made in developing environmental DNA metabarcoding. However, its ability to quantify species relative abundance remains uncertain, limiting its application for biomonitoring. In diatoms, although the rbcL gene appears to be a suitable barcode for diatoms, providing relevant qualitative data to describe taxonomic composition, improvement of species quantification is still required. Here, we hypothesized that rbcL copy number is correlated with diatom cell biovolume (as previously described for the 18S gene) and that a correction factor (CF) based on cell biovolume should be applied to improve taxa quantification. We carried out a laboratory experiment using pure cultures of eight diatom species with contrasted cell biovolumes in order to (1) verify the relationship between rbcL copy numbers (estimated by qPCR) and diatom cell biovolumes and (2) define a potential CF. In order to evaluate CF efficiency, five mock communities were created by mixing different amounts of DNA from the eight species, and were sequenced using HTS and targeting the same rbcL barcode. As expected, the correction of DNA reads proportions by the CF improved the congruence between morphological and molecular inventories. Final validation of the CF was obtained on environmental samples (metabarcoding data from 80 benthic biofilms) for which the application of CF allowed differences between molecular and morphological water quality indices to be reduced by 47%. Overall, our results highlight the usefulness of applying a CF factor, which is effective in reducing over‐estimation of high biovolume species, correcting quantitative biases in diatom metabarcoding studies and improving final water quality assessment.
影响因子：
7.7
作者：
Zimmermann, Jonas;Gloeckner, Gernot;Gemeinholzer, Birgit
通讯作者：
Gemeinholzer, Birgit
影响因子：
12.3
作者：
Lommer M;Specht M;Roy AS;Kraemer L;Andreson R;Gutowska MA;Wolf J;Bergner SV;Schilhabel MB;Klostermeier UC;Beiko RG;Rosenstiel P;Hippler M;LaRoche J
通讯作者：
LaRoche J