5 Prospects
The implementation of eDNA methodologies for amphibian surveillance
encompasses a sequence of five primary stages: sample acquisition, DNA
extraction, PCR amplification, sequencing, and data analysis (Figure 1).
It is evident from this review that the heterogeneous array of research
objectives often engenders considerable divergence within the procedural
framework. Noteworthy variables encompass the volumetric scale of
aquatic samples, ranging from 15 mL to 10 L, the selection of filter
pore sizes spanning the interval from 0.22 μm to 1 μm, and the
deliberate designation of genetic loci such as 12S, 16S, and Cytochrome
b, each of which shows pronounced heterogeneity. This intricate
landscape precludes the facile derivation of a universally ’omnipotent’
experimental protocol. It should be noted that attempts have been made
to propagate standardized methodologies for eDNA techniques (Bruce et
al., 2021; De Brauwer et al., 2022; Minamoto et al., 2021), however
these so far have not encountered widespread acceptance or practical
implementation (Takahashi et al., 2023).
The multifaceted paradigm underlying eDNA application, as discussed in
section 2 of this review, illustrates the causes for the limited
applicability of ’standardization guidelines’ across diverse contexts.
The unique physicochemical characteristics of different environments
make it necessary to carry out iterative adjustments to the experimental
approach blueprint and the methodology, as suitable to varied research
goals (Pawlowski, Apothéloz-Perret-Gentil, et al., 2020; Pawlowski,
Apothéloz‐Perret‐Gentil, et al., 2020; Taberlet et al., 2018; Takahashi
et al., 2023).
Because spatial and temporal differences have a large and unavoidable
effect on eDNA capture, it may be possible to improve the accuracy of
eDNA methods for investigating species diversity or richness of plants
and animals by combining them with distribution or occupancy models.
A high-quality and accurate reference database is required for eDNA
metabarcoding, and the information base must be sufficient to cover all
species in that experimental region. Errors or gaps in the database will
lead to a decrease in the accuracy of the findings (Abad et al., 2016;
Šigut et al., 2017; Yang et al., 2017). Continued enrichment and
improvement of DNA barcode databases in the future are therefore
desirable to improve their accuracy and credibility.
Despite the convenience, accuracy and low cost of the eDNA method, it
has many disadvantages compared to traditional surveys, such as the
inability to directly observe the life stages and disease conditions of
surveyed organisms or to measure organism indicators. eDNA metabarcoding
cannot completely replace traditional survey methods, and choice of
survey methodology should be weighed against the advantages and
limitations of both types of approach, as well as specific research
objectives. Complementary use is most likely to achieve the most
desirable research results.