Current laboratory protocols for detecting fish species with environmental DNA optimize sensitivity and reproducibility, especially for more abundant populations

Abstract Analysing environmental DNA (eDNA) in seawater can aid monitoring marine fish populations. However, the extent to which current methods optimize eDNA detection from water samples is unknown. Here, we test modifications laboratory components of an metabarcoding protocol targeting finfish. As compared baseline methods, amplifying a smaller proportion extracted yielded fewer species, and, conversely, larger identified more taxa. Higher-read species were amplified reproducibly and with less variation read number than lower-read species. Among pooled samples, 20-fold deeper sequencing recovered one additional out total 63 No benefit was observed PCR cycles, alternative primer concentrations, or fish-selective primers. Experiments using exogenous standard assess absolute concentration suggested that, for given sample, are near maximally sensitive. Our results support unofficial collection volume liter assessment commonly encountered We conclude that rarity poses main challenge methods.