Different approaches to processing environmental DNA samples in turbid waters have distinct effects for fish, bacterial and archaea communities

Coastal lagoons are an important habitat for endemic and threatened species in California that have suffered impacts from urbanization and increased drought. Environmental DNA has been promoted as a way to aid in the monitoring of biological communities, but much remains to be understood on the biases introduced by different protocols meant to overcome challenges presented by unique systems under study. Turbid water is one methodologic challenge to eDNA recovery in these systems as it quickly clogs filters, preventing timely processing of samples. We investigated biases in community composition produced by two solutions to overcome slow filtration due to turbidity: freezing of water prior to filtration (for storage purposes and long-term processing), and use of sediment (as opposed to water samples). Bias assessments of community composition in downstream eDNA analysis was conducted for two sets of primers, 12S (fish) and 16S (bacteria and archaea). Our results show that freezing water ..., A sterilized water jug was used to collect a single water sample in the lagoon, at a mid-point between the mouth margin and the road bridge (Fig. 1). The sample was then placed on ice and brought to the laboratory (~1 hr car ride). This method of “grab-and-hold” has proven to be similarly effective as on-site filtration in a previous study (Pilliod et al., 2013). Once in the laboratory, the total volume was divided for two separate protocols: centrifugation; and filtration.
The centrifugation protocol followed Doi et al. (2017) using five replicates of 50 mL tubes (with 27 mL water samples each). Besides extracting the pellet, we also included a filtration step of the supernatant using a 0.45 µm filter. For the filtration protocol, water was separated into ten 500 mL bottles (Fig. 2) and these were used in two separate protocols. Half (i.e. five bottles) were frozen in the -20°C for three days before thawing for filtration (hereafter referred to ‘pre-freezing (PF) protocol’), and half w..., For processing reads into taxonomic tables, please refer to the Anacapa Toolkit (Curd, Gomer, et al., 2018). All other bioinformatic analyses can be performed using R v.3.6.2 (R Core Team, 2018) in RStudio v.1.2.1335 (RStudio Team, 2020).