<scp>DNA</scp> State Influences the Uptake and Persistence of Environmental <scp>DNA</scp> by Marine Sponge Natural Samplers

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 DNA (e.g., cellular or extracellular) that is captured by these samples and how this compares to conventional aquatic eDNA samples. 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 in each sample using quantitative real-time PCR (qPCR). 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 rate and concentration of DNA were significantly lower in nsDNA than in eDNA overall. 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 its impact on the communities resolved using nsDNA methods. <i>Synthesis and applications</i>. We show that nsDNA may originate from a subset of the DNA present in environmental media, potentially providing a more stable picture of local communities. Natural samplers provide a promising option for hard-to-reach environments and for retrieving biodiversity data from archived samples; however, further work and optimization are required to understand what is and is not well represented by this sample type compared to widely applied aquatic eDNA approaches.