Environmental DNA (eDNA) metabarcoding of pond water as a tool to survey conservation and management priority mammals

Abstract

23Environmental DNA (eDNA) metabarcoding can identify terrestrial taxa utilising aquatic habitats 24 alongside aquatic communities, but terrestrial species' eDNA dynamics are understudied. We 25 evaluated eDNA metabarcoding for monitoring semi-aquatic and terrestrial mammals, 26 specifically nine species of conservation or management concern, and examined 27 spatiotemporal variation in mammal eDNA signals. We hypothesised eDNA signals would be 28 stronger for semi-aquatic than terrestrial mammals, and at sites where individuals exhibited 29 behaviours. In captivity, we sampled waterbodies at points where behaviours were observed 30 ('directed' sampling) and at equidistant intervals along the shoreline ('stratified' sampling). We 31 surveyed natural ponds (N = 6) where focal species were present using stratified water 32 sampling, camera traps, and field signs. eDNA samples were metabarcoded using vertebrate-33 specific primers. All focal species were detected in captivity. eDNA signal strength did not differ 34 between directed and stratified samples across or within species, between semi-aquatic or 35 terrestrial species, or according to behaviours. eDNA was evenly distributed in artificial 36 waterbodies, but unevenly distributed in natural ponds. Survey methods deployed at natural 37 ponds shared three species detections. Metabarcoding missed badger and red fox recorded by 38 cameras and field signs, but detected small mammals these tools overlooked, e.g. water vole. 39Terrestrial mammal eDNA signals were weaker and detected less frequently than semi-aquatic 40 mammal eDNA signals. eDNA metabarcoding could enhance mammal monitoring through 41 large-scale, multi-species distribution assessment for priority and difficult to survey species, and 42 provide early indication of range expansions or contractions. However, eDNA surveys need high 43 3 spatiotemporal resolution and metabarcoding biases require further investigation before 44 routine implementation. 45 46

Terrestrial mammal surveillance using hybridization capture of environmental DNA from African waterholes

“…Metabarcoding relies on the occurrence of relatively intact DNA for PCR amplification, and the use of degenerate primers to cover multiple species may bias the results (Taberlet, Bonin, Zinger, & Coissac, ). Metabarcoding of terrestrial animals from eDNA has been performed using environmental sample material of animals in captivity (Harper et al, ; Rodgers & Mock, ), and to a lesser extent, also in the wild (Egeter et al, ; Ushio, Fukuda, et al, ). Under natural conditions, however, this approach is typically hampered by the presumably low amount of target DNA in any given environmental substrate compared to the high amount of background DNA.…”

Genetic tools in the management of invasive mammals: recent trends and future perspectives

“…Organisms leave genetic material behind in the surrounding environment (e.g. in water bodies and soil) via excretions and secretions (Harper et al ); this is referred to as environmental DNA ( eDNA ). Single‐species detection from eDNA is possible using PCR, qPCR or droplet digital PCR ( ddPCR ).…”

“…DNA metabarcoding from environmental samples has the potential to be used as an early warning system for the detection of invasive non‐native species, can be used for continuous monitoring programmes, and has been extensively applied for tracking biological invasions in aquatic ecosystems (Deiner et al ). eDNA metabarcoding studies targeting mammalian communities were relatively rare in comparison with other taxonomic groups (Sales et al ), but this may change now that there are established metabarcoding protocols for detecting and monitoring whole communities using vertebrate (Harper et al ) or mammal‐specific primer sets (Ushio et al , Sales et al , b).…”

“…For example, mammals with larger home ranges (e.g. invasive carnivores) have lower probabilities of detection than more abundant group‐living mammals (Harper et al , Sales et al ). Due to the high sensitivity of metabarcoding, contamination is a concern (Sales et al ).…”

“…Another consideration is the existence of gaps in customised or online reference databases for identifying sequences to the appropriate species level in under‐studied geographic regions (Sales et al ). However, with a carefully planned experimental design and the appropriate field and laboratory controls (Zinger et al ), eDNA metabarcoding has the potential to be applied for early detection and ongoing surveillance of invasive mammals (Harper et al , Sales et al ).…”