Enhancing environmental DNA metabarcoding from marine ecosystems: Impact of filter type, storage method, and storage time on the assessment of fish alpha and beta diversity

Abstract

The collection of environmental DNA (eDNA) and subsequent metabarcoding are useful tools for assessing marine fish biodiversity noninvasively. It is of particular importance to evaluate biodiversity in regions that are hard to access and thus less well studied. Sampling and preservation methods tailored to the specific circumstances are required. Aquatic eDNA is often captured on filters made of different materials and pore sizes, and subsequently stored under divergent conditions for varying periods of time. Previous studies on multispecies detection in marine systems have primarily focused on capture and extraction effects. Our study, in contrast, examined the effects of filter type, storage method, and storage time on DNA yield, alpha (i.e., ZOTU richness) and beta diversity (i.e., ZOTU composition) recovered from a marine ecosystem in Shark Bay, Western Australia. We compared two different filter types (cellulose-nitrate filters with pore sizes of 0.45 μm; glass-fiber filters with pore sizes of 0.1 μm), two storage methods (preservation in Longmire's solution and drying, respectively), various storage times (30–68 days) on two metabarcoding assays using different fish-specific primers. Our results showed that storage time decreased DNA yield and affected alpha and beta diversity estimates. Cellulose-nitrate filters stored in Longmire's solution proved to be the best combination with the smallest decrease in DNA yield, no effect on alpha diversity and consistent community compositions. Storing glass-fiber filters in Longmire's solution led to a decrease in eDNA yield and alpha diversity estimates with increasing storage time. Furthermore, the largest change in beta diversity for each metabarcode was found for glass-fiber filters regardless of storage method. Our results highlight the importance of considering storage time and interactions between storage method and filter when analyzing eDNA results, especially when storing samples for an extended time period or comparison of samples stored for different durations.