Estimation accuracy and eDNA state
We showed that the use of larger pore size filters could improve the
accuracy of eDNA-based abundance estimation. According to previous
studies, the cellular and molecular structure of larger eDNA particles
derived from intra-cellular DNA, such as cells and tissues, is degraded
into smaller eDNA particles over time (Jo et al., 2019b). Apparent
persistence of such smaller-sized eDNA could thus be longer than that of
larger-sized eDNA, and larger eDNA particles collected using larger pore
size filters are more likely to be recently released and less degraded
than smaller eDNA particles. Such larger-sized ‘fresher’ eDNA is
expected to reflect species presence and abundance at a spatiotemporally
finer scale, consequently improving the accuracy of eDNA-based abundance
estimation. Alternatively, given that filter pore size was positively
correlated with the volume of water samples (see Materials and Methods),
larger volume of water samples might allow to diminish the heterogeneity
of target eDNA quantity across sampling replicates and improve the
estimation accuracy of species abundance. Although individual eDNA
studies could not clearly infer the effects of filter pore size and
water volume on the estimation accuracy (Takahara et al., 2012;
Eichmiller et al., 2016), further empirical studies will be required
which factors substantially contribute to the relationship between eDNA
quantity and species abundance. In any case, our meta-analysis supports
the applicability of larger pore size filters for improved abundance
estimation via eDNA analysis for the first time.
In contrast, some datasets reported high R2 values
using smaller pore size filters (Figure 3), which can collect both
larger-sized fresher eDNA and smaller-sized older eDNA. Thus, these
studies may have collected a higher proportion of larger-sized eDNA
while using smaller pore size filters; in particular, laboratory
experiments with high abundances (e.g., Takahara et al., 2012; Doi et
al., 2015) may have collected large quantities of large-sized fresh
eDNA. Although studies using larger pore size filters (especially
>3 µm) were limited in our dataset, further empirical
studies targeting larger-sized eDNA particles would conceivably
contribute to the robustness of our results and potentially provide a
new approach to improve the accuracy of eDNA-based abundance estimation
in the field.
PCR amplicon size (i.e., DNA fragment length of target eDNA) was not
significantly correlated with R2 values. However,
almost all the studies in our meta-analysis targeted eDNA fragments
(<500 bp) much shorter than overall genome and thus the effect
of PCR amplicon size on the accuracy of eDNA-based abundance estimation
may be underestimated. Owing to higher decay rates, detection of longer
eDNA fragments may mitigate the effect of degraded eDNA and improve
species abundance estimation accuracy (Jo et al., 2017). Nonetheless, Jo
et al. (2017) was conducted in a situation where false-positive
inferences of target individuals could be obvious (i.e., the effect of
fish markets and dead individuals); thus, the performance of longer eDNA
fragments for species abundance estimation in more ordinary situations
is unknown. In addition, some studies reported that there were no
significant differences in eDNA decay rate or persistence time depending
on amplicon size (e.g., Ma et al., 2016; Bylemans et al., 2018). Future
empirical studies targeting longer eDNA fragments are needed to
elucidate the importance of PCR amplicon size on eDNA-based abundance
estimation.