Key factors to consider in the use of environmental DNA metabarcoding to monitor terrestrial ecological restoration.

Ecological restoration of terrestrial environments is a globally important process to combat the loss of biodiversity and ecosystem services. Holistic monitoring of restored biota and active management of restoration is necessary to improve restoration processes and outcomes, and provide evidence to stakeholders that targets are being achieved. Increasingly, environmental DNA (eDNA) metabarcoding is used as a restoration monitoring tool because it is able to generate biodiversity data rapidly, accurately, non-destructively, and reliably, on a wide breadth of organisms from soil microbes to mammals. The overall objective of this review is to discuss the key factors to consider in the use of environmental DNA for monitoring of restored terrestrial ecosystems, hopefully improving monitoring, and ultimately, restoration outcomes. We identified that the majority of eDNA based studies of ecosystem restoration are currently conducted in Europe, North America, and Australia, and that almost half of total studies were published in 2021-22. Soil was the most popular sample substrate, soil microbial communities the most targeted taxa, and forests the most studied ecosystem. We suggest there is no 'one size fits all' approach to restoration monitoring using eDNA, and discuss survey design. Factors to consider include substrate selection, sample collection and storage, assay selection, and data interpretation, all of which require careful planning to obtain reliable, and accurate information that can be used for restoration monitoring and decision making. We explore future directions for research and argue that eDNA metabarcoding can be a useful tool in the restoration monitoring 'toolkit', but requires informed application and greater accessibility to data by a wide spectrum of stakeholders.