Quantitative monitoring of diverse fish communities on a large scale combining eDNA metabarcoding and qPCR

eDNA metabarcoding is an effective method for studying fish communities
but allows only an estimation of relative species abundance (density /
biomass). Here, we combine metabarcoding with an estimation of the total
abundance of eDNA amplified by our universal marker (teleo) using a qPCR
approach to infer the absolute abundance of fish species. We carried out a
2,850 km eDNA survey within the Danube catchment using a spatial
integrative sampling protocol coupled with traditional electrofishing for
fish biomass and density estimation. Total fish eDNA concentrations and
total fish abundance were highly correlated. The correlation between eDNA
concentrations per taxon and absolute specific abundance was of comparable
strength when all sites were pooled and remained significant when the
sites were considered separately. Furthermore, a non-linear mixed model
showed that species richness was underestimated when the amount of
teleo-DNA extracted from a sample was below a threshold of 0.65.106 copies
of eDNA. This result, combined with the decrease in teleo-DNA
concentration by several orders of magnitude with river size, highlights
the need to increase sampling effort in large rivers. Our results show a
comprehensive description of longitudinal changes in fish communities and
underline our combined metabarcoding/qPCR approach for biomonitoring and
bioassessment surveys when a rough estimate of absolute species abundance
is sufficient.

环境DNA宏条形码（eDNA metabarcoding）是研究鱼类群落的有效手段，但仅能估算物种相对丰度（密度/生物量）。本研究将宏条形码测序与通过通用标记（teleo）扩增的eDNA总丰度估算（采用定量聚合酶链式反应（qPCR）方法）相结合，以此推断鱼类物种的绝对丰度。研究团队在多瑙河流域开展了总长2850公里的eDNA调查，采用空间整合采样方案，并结合传统电捕鱼法对鱼类生物量与密度进行估算。鱼类总eDNA浓度与鱼类总丰度呈高度相关。当合并所有采样点分析时，各分类单元的eDNA浓度与物种绝对丰度之间的相关强度相当；且在单独分析各采样点时，该相关性仍具有统计学显著性。此外，非线性混合模型分析结果显示，当样本中提取的teleo-DNA总量低于0.65×10^6个eDNA拷贝的阈值时，物种丰富度会被低估。该结果结合“teleo-DNA浓度随河流规模增大而降低数个数量级”的现象，凸显了在大型河流中提升采样强度的必要性。本研究结果全面阐明了鱼类群落的沿程变化特征，并证实：当仅需对物种绝对丰度进行粗略估算时，本研究结合宏条形码测序与qPCR的方法可有效应用于生物监测与生物评估调查。