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Can DNA-Based Ecosystem Assessments Quantify Species Abundance? Testing Primer Bias and Biomass--Sequence Relationships with an Innovative Metabarcoding Protocol.

ABSTRACT: Metabarcoding is an emerging genetic tool to rapidly assess biodiversity in ecosystems. It involves high-throughput sequencing of a standard gene from an environmental sample and comparison to a reference database. However, no consensus has emerged regarding laboratory pipelines to screen species diversity and infer species abundances from environmental samples. In particular, the effect of primer bias and the detection limit for specimens with a low biomass has not been systematically examined, when processing samples in bulk. We developed and tested a DNA metabarcoding protocol that utilises the standard cytochrome c oxidase subunit I (COI) barcoding fragment to detect freshwater macroinvertebrate taxa. DNA was extracted in bulk, amplified in a single PCR step, and purified, and the libraries were directly sequenced in two independent MiSeq runs (300-bp paired-end reads). Specifically, we assessed the influence of specimen biomass on sequence read abundance by sequencing 31 specimens of a stonefly species with known haplotypes spanning three orders of magnitude in biomass (experiment I). Then, we tested the recovery of 52 different freshwater invertebrate taxa of similar biomass using the same standard barcoding primers (experiment II). Each experiment was replicated ten times to maximise statistical power. The results of both experiments were consistent across replicates. We found a distinct positive correlation between species biomass and resulting numbers of MiSeq reads. Furthermore, we reliably recovered 83% of the 52 taxa used to test primer bias. However, sequence abundance varied by four orders of magnitudes between taxa despite the use of similar amounts of biomass. Our metabarcoding approach yielded reliable results for high-throughput assessments. However, the results indicated that primer efficiency is highly species-specific, which would prevent straightforward assessments of species abundance and biomass in a sample. Thus, PCR-based metabarcoding assessments of biodiversity should rely on presence-absence metrics.

SUBMITTER: Elbrecht V

PROVIDER: S-EPMC4496048 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Can DNA-Based Ecosystem Assessments Quantify Species Abundance? Testing Primer Bias and Biomass--Sequence Relationships with an Innovative Metabarcoding Protocol.

Elbrecht Vasco V Leese Florian F

PloS one 20150708 7

Metabarcoding is an emerging genetic tool to rapidly assess biodiversity in ecosystems. It involves high-throughput sequencing of a standard gene from an environmental sample and comparison to a reference database. However, no consensus has emerged regarding laboratory pipelines to screen species diversity and infer species abundances from environmental samples. In particular, the effect of primer bias and the detection limit for specimens with a low biomass has not been systematically examined, ...[more]

PMID: 26154168

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Project description:This study assessed the cell carbon content and biomass for genera of dinoflagellates and diatoms in the oceanic ecosystem of the Southern Gulf of Mexico. Carbon content estimates were based on biovolume calculations derived from linear dimension measurements of individual cells and the approximate geometric body shape of each genus. Then, biomass assessments were performed for both groups in two gulf regions (Perdido and Coatzacoalcos) using these carbon content factors and cell abundances. After four seasonal cruises, 11,817 cells of dinoflagellates and 3,412 cells of diatoms were analyzed. Diverse body shapes and cell sizes were observed among 46 dinoflagellate genera and 37 diatom genera. Nano-cells of dinoflagellates (68% <20 ?m) and micro-cells of diatoms (77% 20-200 ?m, mostly 50-75 ?m) were predominant. According to this cell-size structure, on average, diatoms contained 40% more carbon per cell than dinoflagellates. Contrasting carbon content estimates were observed within the genera of both microalgae. Large carbon averages (>10,000 pg C cell-1) were attributed to Gonyaulacal and some occasional genera of dinoflagellates (e.g., Pyrocystis and Noctiluca) and centric diatoms. In contrast, values up to 3 orders of magnitude lower were found for Peridinial and Gymnodinial dinoflagellates and pennate diatoms. Based on these carbon content estimates, which can be considered representative for most of this oceanic ecosystem, seasonal and regional differences were found in the biomass assessments conducted for these functional groups. Overall, dinoflagellates (mostly low-carbon Gymnodinales) had larger depth-integrated biomass than diatoms (mainly rich-carbon centric forms) within the euphotic zone. An exception to it was the late-summer cruise at the Coatzacoalcos region when a surface bloom of centric diatoms was observed in stations influenced by river runoff. This work contributes useful reference information for future ecological studies and models for understanding the biogeochemical functioning of this open-ocean ecosystem.

Dataset Information

Can DNA-Based Ecosystem Assessments Quantify Species Abundance? Testing Primer Bias and Biomass--Sequence Relationships with an Innovative Metabarcoding Protocol.

Publications

Can DNA-Based Ecosystem Assessments Quantify Species Abundance? Testing Primer Bias and Biomass--Sequence Relationships with an Innovative Metabarcoding Protocol.

Similar Datasets

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