Metabarcoding primers for Indo-Pacific fishes

Environmental DNA (eDNA) metabarcoding offers an effective solution to
determine fish species compositions in communities across diverse
environments. However, it is not clear how different metabarcoding primers
perform in terms of recovering fish species and community diversity in the
Indo-Pacific bioregion. In this study, we compared the relative
performance of five metabarcoding primers (Berry 16S, Riaz 12S, MiFish E
12S, MiFish U 12S, and Leray CO1) in recovering Indo-Pacific fish taxa. We
tested the primers using template DNA from three different environments:
(1) a controlled mock community composed of tissue-based DNA extractions
from 96 species, (2) a semi-controlled Indo-Pacific reef fish community
from a public aquarium tank, and (3) a natural tropical coral reef lagoon.
In the mock community sample, each primer recovered a distinct subset of
the community, and no single primer recovered all taxa. Of the 65 distinct
genera included in the mock community, all but six were recovered by at
least one primer, representing 91% of genera. Fifty-nine of the 96
included species (61%) were identified to species-level using at least one
primer set. From the aquarium community, 17 of the 20 known genera were
recovered (85%), and 13 out of 24 (54%) censused species were identified
by at least one primer. In the coral reef lagoon 48 genera were
identified, and 47 species-level identifications were made, including 87%
endemic and established species. Overall, we find Riaz 12S performed
better than other fish-specific markers although there were differences in
the specific fish taxa recovered. While all markers performed well in the
mock community in terms of the relative proportion of fish sequences
recovered, this did not accurately predict how they would perform under
natural conditions. Caution is therefore urged in using a mock community
alone to evaluate metabarcoding primer performance for studies in natural
environments.