Connectivity assessment of fish communities in Welsh Rivers using Environmental DNA metabarcoding Teja Petra Muha¹, Carlos Garcia De Leaniz¹, Sonia Consuegra del Olmo¹ Bioscience department, Swansea University, Singleton Park, Swansea, SA2 8PP, Wales, UK t.p.muha@swansea.ac.uk Goals To define how natural and artificial barriers limit the spread of invasive fish species; To assess the connectivity of native fish species in Welsh rivers using a novel environmental DNA metabarcoding technique. Introduction Aquatic invasive species (AIS) are one of the major causes for worldwide decline in biodiversity. Limiting the impact of AIS requires an appropriate management strategy, focused on effective early detection. Invasive species in freshwater lotic systems are limited in dispersal downstream or upstream from a point of colonization by natural or artificial barriers. A powerful genetic technique based on multiple species detection (metabarcoding) through environmental DNA (eDNA) have been developed. Metabarcoding can be described as a method of biodiversity assessment that combines two technologies: eDNA based identification and high-throughput DNA sequencing. Figure 1: Courtesy: Fishbio.com Methods The process involves environmental DNA collection of water samples, filtration, DNA extraction, amplification, sequencing and bioinformatics analysis for taxonomic assignment, which is based on Molecular Operational Taxonomic Unit (MOTU)s and on reference databases. . Within a single drop of water… Sampling activities ..There is hidden information about all species in the area! Results Environmental DNA metabarcoding method provides much more accurate, reliable, quicker and cheaper way of species detection compared to traditional sampling techniques. Take home Message Future directions A perfectly designed universal fish primer set, which will be as species specific on one hand as well as supporting wide species coverage on the other hand. DNA sequence databases is far from finished and continues to be updated with each individual research like this one. Further optimisation of each individual step for environmental DNA matabarcoding pipeline is needed. Acknowledgments Would like to thank to my mentors for all the calm support, the very productive team of Aquainvad-ed project, to the Natural Resources Wales for the full support on field work at Teifi river, and to anglers’ societies who offered helpful hand at sampling in Tawe and Afan river. Last but not least would like to offer my gratitude to the EU Horizon 2020 for the Aquainvad-ed project funding support. Figure 3- Environmental DNA metabarcoding workflow: The sampling strategy involves collecting the water samples and variety of filtration techniques used (on the top scheme), afterwards eDNA extraction and universal Primer optimisation takes place (in the middle ), whereas library preparation for MiSeq Illumina high-throughput sequencing, sequencing itself and identification of fish species using bioinformatics tools takes place (at the bottom). Figure 4- Optimisation of universal set of primers testing the detection efficiency on eDNA water samples using standard PCR and gel electrophoresis. Figure 2: Sampling locations at Welsh rivers : Teifi, Tawe and Afan. This project has received funding from the European Union’s Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant agreement No 642197.