DNA Barcoding of Pacific Invasive and Pest Species

Pacific Science Congress Kuala Lumpur

David E. Schindel, Executive SecretaryNational Museum of Natural History

Smithsonian InstitutionSchindelD@si.edu; http://www.barcoding.si.edu

202/633-0812; fax 202/633-2938

Today’s GoalsShare information on barcoding, invasives

Share information on projects and organizations in Pacific

Discuss potential regional cooperation; and

Discuss possible formation of a PSA Working Group on DNA barcoding of invasive/pest species:– Participants (individuals, labs, institutes, agencies)– Activities (training, workshops, collecting, producing

data)– Deliverables (data, publications, websites)

Existing ActivitiesNational quarantine agencies (NPPOs)

Regional agencies and initiatives (RPPOs, Quads, QBOL)

Global Initiatives (IPPC, CABI, GISP)

BioNET INTERNATIONAL LOOP PaciNET

PBIF: Pacific Node of GBIF

The DNA Barcoding Initiative

Barcoding is becoming a global standard for species identification

Rapidly expanding by region, taxa, applications

The Barcoding Initiative is global with participants in 50+ countries

CBD, IPPC, Global Taxonomy Initiative, Census of Marine Life, others involved

Government agencies: USDA, FDA, NOAA

Species Identification MattersBasic research on evolution, ecology

Invasive species (e.g., in ballast water)Agricultural pests/beneficial speciesEndangered/protected species Disease vectors/pathogensEnvironmental quality indicatorsManaging for sustainable harvestingConsumer protection, ensuring food qualityFidelity of seedbanks, culture collections

6

A DNA barcode is a short gene sequence

taken from standardized portions

of the genome, used to identify species

An Internal ID System for All Animals

Typical Animal Cell

Mitochondrion

DNA

mtDNA

D-Loop

ND5

H-strand

ND4

ND4L

ND3COIII

L-strand

ND6

ND2

ND1

COII

Small ribosomal RNA

ATPase subunit 8

ATPase subunit 6

Cytochrome b

COICOI

The Mitochondrial Genome

Associating Life Stages, Processed Parts, Dimorphic Genders

Non-COI regions for other taxaLand plants:– Chloroplast matK and rbcL approved Nov 09

– 70-75% resolving ability, higher in angiosperms– Non-coding plastid and nuclear regions being

explored

Fungi:– CBOL Working Group met this week in Amsterdam– Agreed to recommend ITS; 72% effective

Protists:– CBOL Working Group July meeting, Berlin

How Barcoding Works

PHASE 1: Build a barcode reference library:– Well-identified specimen– Tissue subsample– DNA extraction, PCR amplification– DNA sequencing– Data submission to GenBank

PHASE 2: Identify unknowns:– Any unidentified juvenile, adult, fragment, product– Tissue sample, DNA, sequencing– Comparison with sequences in reference library

Current Norm: High throughputLarge labs, hundreds of samples per day

ABI 3100 capillary

automated sequencer

Large capacity PCR and

sequencing reactions

● US$100-150K purchase ● 2-3 hours processing time● 150-500 samples per day ● US$3-5 per sample

Technology Development Partnership Goal

The DNA Sequencing

Lab of 2013?

NBII, 25 February 2009

BOLD System Workbench in Canada

USER

/GenBank

Key

Mirroring

Update Channel

Private Records

BARCODE Record Flow Chart

Barcode Sequence

Voucher Specimen

Species Name

Specimen Metadata

Literature(link to content or

citation)

BARCODE Records in INSDC

Indices - Catalogue of Life - GBIF/ECAT

Nomenclators - Zoo Record - IPNI - NameBank

Publication links - New species

GeoreferenceHabitat

Character setsImages

BehaviorOther genes

Trace filesOther

DatabasesPhylogenetic

Pop’n GeneticsEcological

Primers

Databases - Provisional sp.

Linkout from GenBank to BOLD

ISBER: 13 May 2009

Linkout from GenBank to Taxonomy

ISBER: 13 May 2009

Link from GenBank to Museums

Darwin Core TripletStructured Link to Vouchers

Institutional Acronym

Collection Code

Catalog ID

: :

Structured Link to Vouchers

NHM LEP 123456: :

personal DHJanzen SRNP12345: :

NCBI’s Biorepository List

Compiled from Index Herbariorum, literature sources, GenBank submissions

6,936 records

1,177 records with non-unique acronyms

517 homonymous acronyms

374 shared by two records

143 shared by three records

AMNHIcelandic Institute of Natural History, Akureyri Division Akureyri Iceland

AMNH American Museum of Natural History New York USA

UNL Universidad Autónoma de Nuevo León Monterrey, Nuevo León Mexico

UNL University of Nebraska State Museum Lincoln, Nebraska USA

UNLCentro de Estratigrafia e Paleobiologia da Universidade Nova de Lisboa Monte de Caparica Portugal

ZMK Zoological Musem, Kristiania Oslo Norway

ZMK Zoologisches Museum der Universität Kiel Kiel Germany

ZMK Zoological Museum, Copenhagen Copenhagen Denmark

CBOL/GBIF/NCBI Registry of Biorepositories

www.biorepositories.org

31 Malaysian Biorepositories Recorded10 Confirmed, 21 Unconfirmed

Producing Barcode Data: 201?Barcode data anywhere, instantly

Data in seconds to minutes

Pennies per sample

Link to reference database

A taxonomic GPS

Usable by non-specialists

• Promote barcoding as a global standard

• Build participation• Working Groups• BARCODE standard• International

Conferences• Increase production

of public BARCODE records

Networks, Projects, Organizations

Barcode of Life Community1,264,000 specimens already barcoded from 104,500 species

The International Barcode of Life Project (iBOL)

5 Million specimens, 500,000 species in 5 years$150 million with core funding from Genome Canada

iBOL website, University of Guelph, Ontario: www.ibol.org

iBOL Theme 1: DNA Barcode Library

WG 1.1 VertebratesWG 1.2 Land PlantsWG 1.3 FungiWG 1.4 Human Pathogens and ZoonosesWG 1.5 Agricultural and Forestry Pest and ParasitoidsWG 1.6 PollinatorsWG 1.7 Freshwater Bio-SurveillanceWG 1.8 Marine Bio-SurveillanceWG 1.9 Terrestrial Bio-SurveillanceWG 1.10 Polar Life

• 200+ Member organizations, 50 countries

• 35+ Member organizations from 20+ developing countries

Consortium for the Barcode of Life (CBOL)CBOL

Member Organizations: 2010

Outreach ActivitiesCape Town, South Africa, April 2006, SANBI– Scale insects in African agriculture

Nairobi, Kenya, October 2006 – Commercial fisheries in Rift Valley lakes

Brazil, March 2007– Hardwood tree species– Endangered mammals, reptiles, amphibians

Taiwan, September 2007

Nigeria, October 2008

Beijing, May 2009

India, November 2010

Adoption by RegulatorsInternational Plant Protection Commission– CBOL and APHIS to host Diagnostic Protocol Panel

meeting, July 2010Federal Aviation Administration – $500K for birdsEnvironmental Protection Agency– $250K pilot test, water quality bioassessment

Food and Drug Administration – Reference barcodes for commercial fish

NOAA/NMFS– $100K for Gulf of Maine pilot project

CITES, National Agencies, Conservation NGOs

ConclusionsBarcoding is a cost-effective system for rapid identification

Barcode reference libraries are being constructed for several endangered groups

CBOL and iBOL provide a global network of specialists capable of constructing barcode reference libraries on selected groups

Partnerships with national and regional groups and regulatory agencies are the critical missing components