Recurring Themes
Besides the insights made in each of the major theme areas above, there were several recurring themes that emerged from multiple studies:
Importance of museum & associated reference collections . Museums play a key role in metabarcoding approaches. First, while many insights can be gained from molecular sequences alone, the availability of a reference collection, ie, molecular barcodes for identified specimens, adds unprecedented dimensionality to the data. The availability of a reference collection allows us to infer the functional traits and morphological attributes of every taxon in a sample, whether it is native or not at a given site, its overall distribution and trophic relationships. Moreover, it is critical that the identity of the specimen has been thoroughly confirmed, as misidentification can lead to flawed interpretations. Thus, rather than diminishing any role of natural history museums in such approaches, the vast data that have been generated through molecular profiling approaches have increasingly highlighted the fundamental importance of barcodes from reliably identified species and populations (Valdivia‐Carrillo, Rocha‐Olivares, Reyes‐Bonilla, Domínguez‐Contreras, & Munguia‐Vega, 2021). The importance of a reference collection is highlighted by Lue et al. (2022) which describes the importance of a vetted and curated reference library for biological control studies. Likewise, Lu et al. (2022) emphasize the limitations of inference without a reference database, and introduce a fungal rRNA operon database (FRODO) with 1116 linked to taxonomically identified species.
A second role of museum specimens in these approaches is that they can provide historic samples of past environments. For example, metabarcoding of pollen loads from museum bee specimens has provided key insights into environmental change over decadal scales, both in the availability of plants, and changes in interaction networks (Bell et al., 2022; Gous, Swanevelder, Eardley, & Willows‐Munro, 2019). This work adds to the increasing body of research that shows how metabarcoding of museum specimens can provide information on changes in interactions through time, including diet and microbiome (Heindler et al., 2018) and parasite-host interactions (Greiman et al., 2018).
Insights from clustering at different levels : Early metabarcoding studies used clustering approaches and generally grouped ASVs into Operational Taxonomic Units (OTUs), assumed to correspond to species. The purpose of this step was to remove the known noise in the data, while also grouping taxa into species. However, new denoising approaches have presented the opportunity of analyzing ASVs and hence gaining insights into population-level patterns (Noguerales et al., 2023). The most important aspect of the ability to look at different levels of genetic clustering is that the comparison can be tremendously informative into the processes that govern species assembly.
Incorporation of Deep Learning / Artificial Intelligence approaches applied to image analysis to study arthropod biodiversity (Emerson et al., 2022). Supervised learning was used to make predictions of sediment sample proximity to shipwrecks based on frequency of microbial taxa (Hampel et al., 2022).