Rosie Dowell

and 4 more

1. Marine sponges as natural samplers of environmental DNA (eDNA) are receiving growing attention as an untapped source of biodiversity data. However, little is known about the state of the DNA that is represented by these samples and how this compares to conventional aquatic eDNA samples. 2. Here, we present an artificial spiking experiment where DNA in cellular and extracellular states was added into tanks containing two sponge species. Aquatic eDNA samples and sponge (natural sampler DNA (nsDNA)) samples were collected over 7 days and DNA from the two states was quantified using qPCR. 3. We found that there was a significant interaction between DNA state and sample type (eDNA and nsDNA), with lower detection and concentration of extracellular DNA, compared to cellular DNA, found in nsDNA samples. We also found that detection probability and concentration of DNA were significantly lower in nsDNA than eDNA overall. 4. During methodological testing, PCR inhibition was observed in both sponge species; this was prohibitive in one of the species. Further work to investigate the degree of PCR inhibition during nsDNA metabarcoding is important to understand it’s impact on the communities resolved using these nsDNA methods. 5. Synthesis and applications. We show that nsDNA may originate from a subset of the DNA present in environmental media, potentially resolving a more stable picture of local communities. Natural samplers provide a promising option for hard-to-reach environments and for analysing archived samples; however, further work and optimisation is required to understand what is and isn’t well represented by this sample type compared to widely applied aquatic eDNA approaches.