Multilocus DNA metabarcoding diet analyses of small cetaceans: a case study on highly vulnerable humpback dolphins and finless porpoises from the Pearl River Estuary, China

Accurate diet identification of top predators is crucial to fully understand their ecological roles. Compared to terrestrial animals, gathering dietary information from cetaceans is notoriously difficult. Here, we applied a multilocus metabarcoding approach to investigate the diet of vulnerable Indo-Pacific humpback dolphins and Indo-Pacific finless porpoises from the Pearl River Estuary (PRE), China. Our analyses identified 21 prey fish species from the 42 humpback dolphin stomachs, as well as 10 species of fish and 1 species of cephalopod from the 13 finless porpoise stomachs. All of the taxa were assigned to the species level, highlighting that the multimarker approach could facilitate species identification. Most of the prey species were small- and medium-sized fishes that primarily fed on zooplankton. The calculated similarity index revealed a moderated dietary overlap between the 2 cetaceans, presumably due to the feeding of the 2 predators in association with fishing vessels in the PRE. A more diverse diet was observed in humpback dolphins in the closed fishing season compared to the fishing season, implying the influence on the dolphin diet due to the availability of commercial fishery resources. However, according to the results of species rarefaction curves, our findings on the feeding habits of the 2 cetaceans are still limited by insufficient sample size and therefore should be interpreted with caution. This study represents a first attempt to apply the multilocus DNA metabarcoding technique in the diet analysis of small cetaceans, although more efforts are needed to improve this type of analysis.