Reviewing Metabarcoding Guidelines and Understanding Sources of Error

The overall project aim is to contribute to greater use of DNA-based methods for monitoring species and ecosystems by environmental public bodies. This project will focus particularly on metabarcoding, and aims to improve end user understanding and awareness of: • What guidance documents and standards for using metabarcoding in environmental exist and what the strengths and weaknesses of these documents are • Why metabarcoding results can differ from conventional methods, and potential approaches to mitigation where these differences may arise from error or bias in metabarcoding. DNA-based methods for detecting and identifying species have many potential applications within the Defra group of organisations (APHA, Defra, CEFAS, Environment Agency, Forest Research, JNCC, Kew, Marine Management Organisation, Natural England). These include surveillance for invasive or rare species, characterising communities to assess the condition of protected sites, understanding the effects of management interventions, and describing ecosystem functioning and resilience. However, operational use by public bodies faces several challenges. To contribute to addressing these challenges, Defra have established a Centre of Excellence (CoE) to support projects that will help facilitate greater uptake of DNA-based methods for monitoring species and ecosystems. The CoE brings together expertise and interests from across the Defra group of organisations to identify and tackle shared priorities through a series of projects. As with all environmental monitoring carried out by public bodies, practical implementation of DNA-based methods requires agreed guidelines and protocols that give confidence the results are robust and are spatially and temporally comparable. However, whilst CoE organisations are aware of and have contributed to several existing guidelines and protocols for using DNA in monitoring, there is less knowledge of documents developed by other organisations - particularly in relation to metabarcoding. A better understanding amongst end users of what metabarcoding guidelines and standards exist would therefore be beneficial for several reasons: • Brings together key resources to make it easier to find relevant documents • Provides opportunities to learn from existing guidelines • Avoids duplicating effort or producing conflicting documents • Allows new guidance documents to be targeted to key remaining gaps

A related important knowledge gap for several CoE organisations is understanding how, when, and why metabarcoding results can differ from conventional methods. Understanding the sources of error and bias in metabarcoding, and being able to distinguish these from situations in which metabarcoding and conventional methods give different results because each samples a different portion of the community, is particularly important for end user interpretation of results from DNA-based methods.