Assessing the degradation of environmental DNA and RNA based on genomic origin in a metabarcoding context

Abstract

Molecular tools of species identification based on eNAs (environmental nucleic acids; environmental DNA [eDNA] and environmental RNA [eRNA]) have the potential to greatly transform biodiversity science. However, the ability of eNAs to obtain “real‐time” biodiversity estimates may be complicated by the differential persistence and degradation dynamics of the molecular template (eDNA or eRNA) and the barcode marker used. Here, we collected water samples over a 28‐day period to comparatively assess species detection using eDNA and eRNA metabarcoding of two distinct barcode markers—a mitochondrial mRNA marker (COI) and a nuclear rRNA marker (18S)—following complete removal of Arthropoda taxa in a semi‐natural freshwater system. Our findings demonstrate that Arthropoda community composition was largely influenced by marker choice, rather than molecular template, individual microcosm, or sampling time point. Furthermore, although eRNA may capture similar species diversity as the established eDNA method, this finding may be marker‐dependent. Although we found little to no difference in decay rates observed among sample groups (COI eDNA, COI eRNA, 18S eDNA, 18S eRNA), this result is likely due to limitations in the ability of eNA‐based metabarcoding to provide a strong correlation between true eNA copy numbers present in the environment and final read counts obtained (following the metabarcoding workflow). Collectively, our findings provide further support for the use of multi‐marker assessments in metabarcoding surveys to unravel the broadest taxonomic diversity possible, highlight the limitations of eNA metabarcoding methods in providing accurate decay rate estimates, as well as establish the need for further comparative studies using both metabarcoding and single‐species detection methods to assess the persistence and degradation dynamics of eNAs for a diverse range of taxa.

Marine invasive alien species in Europe: 9 years after the IAS Regulation

“…Environmental RNA (eRNA), similar to eDNA, is shed by metazoans or exists in the form of whole live or dead individuals of smaller organisms, making it a potential monitoring target (Lejzerowicz et al, 2015;Keeley et al, 2018). eRNA has the disadvantage of lower stability in the environment (Kagzi et al, 2023) and requires stricter sample contamination and preservation protocols but is likely a better reflection of the presence of live organisms (Pochon et al, 2017).…”

Beyond species detection—leveraging environmental DNA and environmental RNA to push beyond presence/absence applications

“…Nevertheless, they demonstrate that eRNA was readily detectable and that the ratio of eDNA to eRNA increased over time, implying that eRNA may be a promising method for quantifying "living" biotic assemblages. Kagzi et al (2023) similarly employ metabarcoding to quantify Arthropoda community diversity after the removal of live organisms in seminatural mesocosms using eDNA and eRNA from both nuclear and mitochondrial markers. Contrary to Jo et al (2023), they do not observe a consistent difference in degradation between eDNA and eRNA markers, although degradation was highly variability across eNA type and genomic source.…”

Metabarcoding advances agricultural invertebrate biomonitoring by enhancing resolution, increasing throughput, and facilitating network inference

“…DNA degradation presents another significant challenge for metabarcoding: the more intact DNA is, the better the chances of successfully detecting it. Degradation occurs naturally over time in the environment, in traps and even in storage, but can be exacerbated by factors like UV light (Gomes et al, 2009) and temperature (Ballari and Martin, 2013;Kagzi et al, 2023). Suboptimal storage of samples can lead to a greater rate of DNA degradation.…”

“…To our knowledge, bulk sample community metabarcoding has not yet been used for sticky traps, potentially due to prolonged exposure to conditions which degrade DNA, such as sunlight (UV; Gomes et al, 2009), and high temperatures (Ballari and Martin, 2013;Butterwort et al, 2022;Kagzi et al, 2023); although, metabarcoding has successfully detected the diet of sticky trapped coccinellids (Ammann et al, 2020).…”