Pitfalls in relative abundance estimation using edna metabarcoding

Abstract

Metabarcoding provides unparalleled information of overall ecosystem ecology and diversity, from occurrence to relative abundances, from micro- to macro-organisms, including cryptic species and larval stages that would otherwise be impossible to identify. PCR is still the most cost-effective and routinely used method to extract information from environmental samples coupled with high-throughput sequencing (HTS). Nichols et al. (2018) metabarcoded nine soil environmental samples from Alaska and six mixtures of synthetic oligonucleotides, using a short fragment of the chloroplast trnL gene. The authors tested the effect of PCR replicates, different polymerase mixes and amplicon GC content on diversity estimates. Results show that occurrence data was not biased for any of the parameters tested and only insufficient PCR replicates could impair the detection of rare taxa. For abundance data, polymerases were found to differentially bias estimates of the relative amplicon abundances and this depended on GC content.