Under the karst: detecting hidden subterranean assemblages using eDNA metabarcoding in the caves of Christmas Island, Australia

Katrina M West,Michael Bunce,Alicia Grealy,Euan S Harvey,Zoe T Richards,Robert Susac

doi:10.1038/s41598-020-78525-6

Katrina M West, Michael Bunce + Show 4 more

Open Access

https://doi.org/10.1038/s41598-020-78525-6

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Abstract

Subterranean ecosystems are understudied and challenging to conventionally survey given the inaccessibility of underground voids and networks. In this study, we conducted a eukaryotic environmental DNA (eDNA) metabarcoding survey across the karst landscape of Christmas Island, (Indian Ocean, Australia) to evaluate the utility of this non-invasive technique to detect subterranean aquatic ‘stygofauna’ assemblages. Three metabarcoding assays targeting the mitochondrial 16S rRNA and nuclear 18S genes were applied to 159 water and sediment samples collected from 23 caves and springs across the island. Taken together, our assays detected a wide diversity of chordates, cnidarians, porifera, arthropods, molluscs, annelids and bryozoans from 71 families across 60 orders. We report a high level of variation between cave and spring subterranean community compositions which are significantly influenced by varying levels of salinity. Additionally, we show that dissolved oxygen and longitudinal gradients significantly affect biotic assemblages within cave communities. Lastly, we combined eDNA-derived community composition and environmental (water quality) data to predict potential underground interconnectivity across Christmas Island. We identified three cave and spring groups that showed a high degree of biotic and abiotic similarity indicating likely local connectivity. This study demonstrates the applicability of eDNA metabarcoding to detect subterranean eukaryotic communities and explore underground interconnectivity.

Highlights

Whilst capture-based sampling is fundamental to biospeleological research, it can be hindered in subterranean environments that present difficult to access underground voids and networks
Preliminary research has demonstrated the viability of Environmental DNA (eDNA) metabarcoding in detecting multi-species compositions from underground water s amples[15,16,17,18,19,20], these have largely focused on microbial communities
We omitted all amplicon sequence variants (ASV) for taxa outside of our study scope of subterranean macrofauna; this included any ASV in the domain Bacteria, hairybacks, the kingdom Fungi, ciliates, nematodes, microscopic flatworms, plants and algae

Summary

Introduction

Preliminary research has demonstrated the viability of eDNA metabarcoding in detecting multi-species compositions from underground water s amples[15,16,17,18,19,20], these have largely focused on microbial communities. There is great potential to expand the use of eDNA metabarcoding, for the detection and monitoring of both described and undescribed eukaryotes in subterranean environments, and to investigate community assemblages across trophic levels and haloclines, the evolution and population diversity of stygofauna, and the interconnectivity of underground ecosystems. The diversity and distribution of stygofauna, in particular anchialine fauna, across CI’s extensive underground networks requires further r esearch[25] This is of particular importance given the use of the karstic landscape for phosphate mining and as a water supply for local households and businesses. We seek to evaluate the applicability of eDNA metabarcoding as a non-invasive, highly-sensitive tool for biospeleological assessment

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