Using Metabarcoding Techniques to Map Multiple Dung Beetle-Vertebrate iDNA Networks in a Southeast Asian Forest

Given the high levels of endemism, diversity, and increasing anthropogenic impacts in tropical regions, studies on species interactions and ecological networks are urgently needed to understand community responses to environmental change. Increasingly, molecular methods are used to monitor biodiversity and identify species interactions. In this study, we developed an optimised DNA metabarcoding protocol to construct quantitative dung beetle-vertebrate trophic networks in a heterogenous tropical forest. We show that while the highest probability of detection is within 3 h of feeding, vertebrate DNA can still be detected in most beetles after 24 h, including around 50% of beetles with visually empty guts. Using the vertebrate taxa detected from DNA in the gut contents, we constructed dung beetle-vertebrate networks across 50 forest sites. Our networks displayed high trophic generalism and nestedness. By using group-specific primers, we documented interactions between dung beetles and multiple vertebrate species, including amphibians, birds, and rare mammals, such as the Sunda slow loris and Sunda pangolin. Our optimised molecular techniques contribute towards the development of new methods for comprehensive biodiversity assessments and ecological network studies that are urgently needed to monitor biodiversity in the hyper-diverse tropics.