Assessment of Leaf-Litter Invertebrate Biodiversity Using High Throughput Sequencing

Abstract Leaf litter ecosystems and their fauna are largely understudied, despite their critical ecological roles. Here, we investigate challenges associated with estimating biodiversity in terrestrial leaf litter. Current methodologies for biodiversity assessment are fraught with limitations, amongst the most significant is a decline in taxonomic expertise, complicating the process of species identification and the significant costs associated with species-level morphological identifications. DNA barcoding employs the mitochondrial gene cytochrome c oxidase I (COI) to identify animal species, and DNA metabarcoding facilitates the identification of multiple species without necessitating taxonomic expertise. Recent studies indicate that environmental DNA (eDNA) may exhibit greater sensitivity compared to traditional methods. To test whether these methods work in a real-world application, we sampled leaf litter across a temperate forest/field ecotone. Leaf litter was dried, ground and processed to extract environmental DNA. We evaluated the DNA extraction protocols to test their relative efficacy. We found that the Qiagen Blood and Tissue Kit was the most effective at recovering invertebrate diversity and that there were notable differences in biodiversity between forest and field habitats. Temperature emerged as a significant factor influencing the composition of the communities observed. Our methodology is applicable across various environments for efficient biodiversity assessment and might be particularly beneficial for monitoring pests and invasive species. Our approach offers a cost-effective and timely alternative to conventional biodiversity assessment methods and underscores the significance of accurate assessment methodologies for leaf litter communities. Competing Interest Statement The authors have declared no competing interest.