New environmental metabarcodes for analysing soil DNA: potential for studying past and present ecosystems

Abstract

Metabarcoding approaches use total and typically degraded DNA from environmental samples to analyse biotic assemblages and can potentially be carried out for any kinds of organisms in an ecosystem. These analyses rely on specific markers, here called metabarcodes, which should be optimized for taxonomic resolution, minimal bias in amplification of the target organism group and short sequence length. Using bioinformatic tools, we developed metabarcodes for several groups of organisms: fungi, bryophytes, enchytraeids, beetles and birds. The ability of these metabarcodes to amplify the target groups was systematically evaluated by (i) in silico PCRs using all standard sequences in the EMBL public database as templates, (ii) in vitro PCRs of DNA extracts from surface soil samples from a site in Varanger, northern Norway and (iii) in vitro PCRs of DNA extracts from permanently frozen sediment samples of late‐Pleistocene age (∼16 000–50 000 years bp) from two Siberian sites, Duvanny Yar and Main River. Comparison of the results from the in silico PCR with those obtained in vitro showed that the in silico approach offered a reliable estimate of the suitability of a marker. All target groups were detected in the environmental DNA, but we found large variation in the level of detection among the groups and between modern and ancient samples. Success rates for the Pleistocene samples were highest for fungal DNA, whereas bryophyte, beetle and bird sequences could also be retrieved, but to a much lesser degree. The metabarcoding approach has considerable potential for biodiversity screening of modern samples and also as a palaeoecological tool.

Lake sediment multi-taxon DNA from North Greenland records early post-glacial appearance of vascular plants and accurately tracks environmental changes

“…Vorren et al, 2013). However, detection of ancient bryophyte DNA has been reported to be difficult in previous studies (Suyama et al, 2008;Epp et al, 2012), and the late start of bryophyte DNA detection (at 225 cm) in our core is in line with this. This sample is nearly 8000 cal yr BP old, but we would have expected bryophytes to be present earlier in the record.…”

“…Extraction of total DNA was carried out using the PowerMax™ Soil DNA Isolation Kit (MOBIO) with protocol modifications described in Epp et al (2012), and included one extraction blank containing only the chemicals. PCRs were set up in the dedicated aDNA laboratory at the Natural History Museum in Oslo.…”

Sedimentary DNA versus morphology in the analysis of diatom-environment relationships

“…ribosomal, mitochondrial and chloroplast markers (e.g. 18S rRNA, cox1 and rbcL) (Evans et al 2007;Medinger et al 2010;Theriot et al 2010;Hamsher et al 2011;Zimmermann et al 2011;Epp et al 2012;Kermarrec et al 2013;StoofLeichsenring et al 2014). In our study, a short and variable sequence of 76 bp of the chloroplast rbcL gene (rbcL_76 amplicon) was used because of its specificity to the amplification of diatom DNA from sedimentary records, and especially ancient sedimentary DNA.…”

High-throughput sequencing of African chikanda cake highlights conservation challenges in orchids

“…Several metabarcoding studies have been published that use nrITS1 or nrITS2 for the identification of fungi (Epp et al 2012;Blaalid et al 2013;Schmidt et al 2013;De Beeck et al 2014). So far the only plant studies to use nrITS in metabarcoding have used nrITS2 to determine the taxonomic composition of pollen collected by honey bees (Galimberti et al 2014;Richardson et al 2015) and species composition in herbal medicines (Cheng et al 2014;Ivanova et al 2016).…”