4.4 No Holy Grail for macrobenthos metabarcodes?
The heterogeneous performance of the different metabarcodes highlights
the complexity of the identification of the best markers. No primer
showed the best performance for all the considered metrics, as the most
”universal” marker (Euka02) showed a generally poor resolution, while
the marker with the highest resolution (BF1_BR2-COI) did not
successfully amplify many taxa. The selection of metabarcodes for
biomonitoring is therefore a trade-off, depending on the aims of
studies. Euka02 can allow a good assessment of overall biodiversity, but
it is unable to tease apart closely related taxa, thus it might be not
enough to define the ecological status of environments. Furthermore, the
poor resolution would hamper the comparison with historical data for
most of taxa. Conversely, the excellent resolution of BF1_BR2-COI could
allow species-level identification, and might have more power to
distinguish different communities. However, this comes at a cost. Many
taxa did not amplify either because the level of DNA degradation
compromises the amplification of a relatively long metabarcode, or
because the poor match of the primer(s) with their target. In fact, the
relatively long amplified metabarcode could limit its usefulness for
application with environmental DNA extracted from water. Finally, Inse01
showed a generally good performance, but it is not appropriate for many
non-insect taxa.
Given these limitations, it is unlikely that one single metabarcode will
be able to fully replace the traditional biomonitoring using
macrobenthic invertebrates. Nevertheless, the data obtained through
multiple metabarcodes can be integrated for a more comprehensive and
accurate understanding of ecological impacts on freshwater biodiversity.
For instance, highly universal markers, providing a complete but coarse
picture of animal biodiversity (e.g. Euka02) can be combined with
markers providing a specific focus on key taxa (e.g. Inse01) or a
high-resolution level (e.g. BF1_BR2-COI). The integration of
multiple metabarcodes certainly increases the cost and complexity of
studies, still it has the potential to provide an unprecedented amount
of data, thus opening unexplored avenues to biodiversity assessment.