using dna based methods for monitoring - eurosite€¦ · dna barcodes= short regions of a genome,...

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Using DNA based methods for monitoring

Andy Nisbet

Why use DNA based methods for biological monitoring?

Which species? IdentificationWhere? Location/distributionHow much? Abundance

Sampling bottleneck and low detection rate

Why use DNA based methods for biological monitoring?

Which species? IdentificationWhere? Location/distributionHow much? Abundance

Identification bottleneck and taxonomic inconsistency

Why use DNA based methods?

Why use DNA based methods

DNA based methods

DNA barcoding or targeted assays• What species is this?

• Is species X present?

PCR/qPCRSingle or few targets

Presence absence or semi-quantification of genes (qPCR)

High throughput, low/medium cost

DNA barcodes = short regions of a genome, usually mitochondrial egCytochrome Oxidase 1 (CO1)

DNA based methods

DNA metabarcoding = high throughput, can analyse mixed samples• What species are here?

High throughput sequencing Many targetsRelative quantification (possibly)High throughputMedium/high costs

• Linked to reference database to identify organism (eg Barcode of Life Database (BOLD))

DNA based methods

Environmental (eDNA)

Community

Targeted assays for Protected and Invasive species

Great Crested Newt (Triturus cristatus)Annexes II and IVStandard qPCR technique in UK

Targeted assays for Protected and Invasive species

Native white clawed crayfish (Austropotamobius pallipes) – Annexes II and IV

Invasive signal crayfish(Pacifastacus leniusculus)

A metabarcoding approach for Protected Species

Nene Washes SPA and SACAnnex II species – Spined Loach, Cobitis Taenia

Spined loach in the Nene Washes

Metabarcoding invertebrate communities

Vane trapping2 sites in Lincolnshire and Staffordshire

Riparian pitfall trapsRiver Frome, Dorset

Metabarcoding invertebrate communities –River Frome by family

Using DNA based methods

Pros• Does not rely on limited number of taxonomic specialists• Can be automated and standardized• Scalable to 1000s of samples• Can identify cryptic species, species that are difficult to detect (including at

different stages in life cycle)• Less prone to error?

Cons• Costs and protocols still in flux, including field sampling• Less cost effective for small sample• Reference databases need to be improved• Need to understand transport and persistence in water and sediment• Limited information on abundance

Where are we now?

• Use for vertebrates is well developed. eDNA is very effective for surveying fish and standard protocols are being developed for fish in lakes.

• Targeted assays are being developed for Protected and Invasive species.• DNAquanet are working to standardise approaches across Europe to using

eDNA for Water Framework Directive monitoring.• Metabarcoding can be used to characterising trap caught terrestrial

invertebrate samples (but understand the limitations).• More work is needed on using metabarcoding of invertebrates from water

and sediment samples.• Huge potential – protected species, characterising communities, INNS,

pests and pathogens.• Takes time and resources to move from proof of concept to routine

operational use.• Will still need ecologists to interpret results.