Desiccation to DNA: Bulk DNA Metabarcoding of Macroinvertebrate Egg Banks via Sugar-Flotation Enhances Biodiversity Surveys of Dry Salt Lakes.

Ephemerally inundated salt lakes in arid regions support unique aquatic macroinvertebrate communities that typically persist through dry periods as resistant resting-stages (e.g., spores, eggs). Salt lakes represent a large proportion of inland lentic habitats on the Australian continent, many occurring in remote arid areas where filling events are infrequent and unpredictable. Currently, biodiversity surveys of dry salt lakes rely on artificial rehydration of sediments to stimulate emergence of resident macroinvertebrate taxa. However, egg banks contain cryptic assemblages with poorly understood hatching requirements that may fail to emerge under artificial rehydration conditions and underestimate biodiversity. DNA metabarcoding of resting eggs may offer a complementary approach by enabling the detection of taxa independent of emergence. We conducted two experiments using sediments from an ephemeral salt lake in central Western Australia, applying two metabarcoding assays (fwh and Crust16S) targeting aquatic invertebrates. Experiment 1 compared a sugar-flotation pre-extraction method versus a standard sediment DNA extraction protocol. Experiment 2 investigated how sediment weight (50, 250, 500 and 1000 g) subjected to sugar-flotation influenced the biodiversity detected. Sugar-flotation vastly outperformed standard DNA extraction, with DNA sequences from target taxa almost exclusively recovered when the sugar-flotation step was applied. Species richness increased significantly from 50 to 250 g and continued increasing until 1000 g although pairwise comparisons among the 250, 500 and 1000 g samples were not significant. DNA metabarcoding results from the 1000 g treatment were compared with collections of taxa recovered from a contemporaneous rehydration trial. DNA metabarcoding detected more taxa, including seven unique taxa, whereas only one taxon was exclusive to rehydration trials. Our findings indicate that bulk metabarcoding of egg banks isolated by sugar-flotation is an effective method for documenting aquatic macroinvertebrate communities in dry salt lakes. This represents a valuable addition to monitoring programmes, enhancing effective management and conservation of these important ecosystems.