Large-volume sampling enhances eDNA monitoring of endangered marine species

Environmental DNA (eDNA) has emerged as a powerful tool for detecting marine biodiversity, particularly for monitoring elusive or endangered species. However, traditional eDNA sampling protocols often rely on small water volumes (≤2 L), which can limit detection sensitivity, especially in vast and dynamic marine environments. Here, we evaluated the effectiveness of large-volume eDNA enrichment (5–50 L) for improving the detection probability of endangered marine species, using metabarcoding and quantitative PCR (qPCR) designed for Balaenoptera brydei (Bryde’s Whale). Results showed that the capsule sampler, equipped with a highly folded filter membrane, can filter over 50 L of seawater, significantly increasing eDNA yield and species detection rates compared to standard small-volume sampling (1 L). For metabarcoding results, the large-volume sampling also yielded more total eDNA sequence reads from marine vertebrates and from the endangered Beibu Gulf population of Bryde’s Whale, enabling more robust detection of endangered species. Furthermore, combining metabarcoding and qPCR provided complementary detection capabilities for monitoring endangered marine mammals, with metabarcoding enabling broad-spectrum screening and qPCR providing target-specific quantification. Additionally, eDNA samples containing fecal matter were analyzed to explore candidate prey species identification. This result, the detection of eight fish species as candidate prey-associated taxa via large-volume filtration, shows that eDNA enrichment offers a practical, scalable approach. It effectively enhances the sensitivity of marine biodiversity monitoring and provides a valuable tool for conservation programs focused on endangered marine fauna.