DNA metabarcoding data characterizing insectivorous diet of purple martins (Progne subis subis) using two COI primer sets (ANML and ZBJ)

DNA metabarcoding is a molecular technique frequently used to characterize
diet composition of insectivorous birds. However, results are sensitive to
methodological decisions made during sample processing, with primer
selection being one of the most critical. The most frequently used DNA
metabarcoding primer set for avian insectivores is ZBJ. However, recent
studies have found that ZBJ produces significant biases in prey
classification that likely influence our understanding of foraging
ecology. A new primer set, ANML, has shown promise for characterizing
insectivorous bat diets with fewer taxonomic biases than ZBJ, but ANML is
not yet widely used to study insectivorous birds. Here, we evaluate the
ANML primer set for use in metabarcoding of avian insectivore diets
through comparison with the more commonly used ZBJ primer set. Fecal
samples were collected from both adult and nestling Purple Martins (Progne
subis subis) at two sites in the USA and one site in Canada to maximize
variation in diet composition and to determine if primer selection impacts
our understanding of diet variation among sites. In total, we detected 71
arthropod prey species, 39 families, and 10 orders. Of these, 40 species
were uniquely detected by ANML, whereas only 11 were uniquely detected by
ZBJ. We were able to classify 54.8% of exact sequence variants from ANML
libraries to species compared to 33.3% from ZBJ libraries. We found that
ANML outperformed ZBJ for PCR efficacy, taxonomic coverage, and
specificity of classification, but that using both primer sets together
produced the most comprehensive characterizations of diet composition.
Significant variation in both alpha- and beta-diversity between sites was
found using each primer set separately and in combination. To our
knowledge, this is the first published metabarcoding study to directly
compare avian diet characterizations produced with both ANML and ZBJ
primer sets.