1. DNA Barcoding and Biodiversity Conservation Eduardo Eizirik Centro de Biologia Genômica e Molecular, PUCRS, Brasil Instituto Pró-Carnívoros, Brasil Laboratory of Genomic Diversity, NIH, EUA

2. Images (partial): www.tolweb.org Biodiversity: - Species, ecosystems, evolutionary lineages, genetic diversity, cultural diversity. - Why is it important ? - Intrinsic value - Practical (anthropocentric) value: - Ecosystem services. - Source of food, materials, substances, innovation.

3. The Biodiversity Crisis

4. Time No. of species Species known to science Presently living species Species diversity

5. The Biodiversity Crisis Urgent needs: - To inventory, document, map and characterize the world´s biodiversity; - To understand and monitor current threats to this biodiversity, as well as their impacts in various contexts; - To eliminate or limit the magnitude of these threats, and/or to minimize their impacts through conservation and management actions.

6. Applications of DNA Barcoding in Biodiversity Conservation 1.Gathering data on components of native biodiversity; 1.1. Baseline data; 1.2. Monitoring of biodiversity in impacted areas; 2.Gathering data on threats to native biotas; - e.g. invasive species, pathogens, wildlife trafficking. 3.Helping to enforce actions aimed at curbing threats to biodiversity. - e.g. wildlife forensic analyses.

7. Applications of DNA Barcoding in Biodiversity Conservation 1.Gathering data on components of native biodiversity 1.1. Baseline data: 1.1.1. Thoroughly inventorying all of the Earth’s ecosystems -Discovery of new species; -Improved knowledge on geographic ranges of species. -Analysis of diverse sources/kinds of biological specimens (including environmental samples); -Close interaction with morphology-based taxonomy and reference collections; -Link to in-depth studies (e.g. phylogeographic) – DNA banks

8. Applications of DNA Barcoding in Biodiversity Conservation 1.Gathering data on components of native biodiversity 1.1. Baseline data: 1.1.2. Improving knowledge on species’ ecology and life history -Identification of morphologically distinct genders and life history stages; -Diet (e.g. using fecal DNA); -Interactions with parasites, symbionts and pathogens; -Spatial and temporal patterns of species occurrence, including habitat association.

9. Applications of DNA Barcoding in Biodiversity Conservation 1.Gathering data on components of native biodiversity 1.2. Monitoring of biodiversity in impacted areas: (i.e. almost everywhere...) -Surveys of species occurrence in areas suffering varying degrees and kinds of disturbance: -Information on susceptibility vs. resilience of each organism; -Caution with sampling procedures. -Links to DNA barcoding efforts targeting agricultural and epidemiological (e.g. emerging diseases) aspects.

10. Applications of DNA Barcoding in Biodiversity Conservation 2. Gathering data on threats to native biotas: -Mapping and monitoring the spread of invasive species; -Identifying vectors and reservoirs of pathogens that threaten wildlife populations (as well as addressing potential emerging diseases for humans); -Understanding the dynamics of wildlife trafficking activities; DNA barcodes that also identify geographic provenance.

11. Applications of DNA Barcoding in Biodiversity Conservation 3. Helping to enforce actions aimed at curbing threats to biodiversity. - Wildlife forensic analyses: - Species-level identification (and geographic origin when possible) of confiscated materials (e.g. meat products, eggs, pelts, bones, wood, etc.); - Example: whale meat in Japanese and Korean markets (DNA surveillance initiative); - Barcoding for Species Conservation (BSC) initiative (CBoL): - Bushmeat trade in Africa, international pet trade, trade in tiger parts in China, overexploitation of sharks, etc.

12. Examples of ongoing and planned research fronts in Brazil - Species identification in carnivores; - Fecal DNA: ecological and biogeographic studies (large- scale assessment of current geographic ranges); - Study on rabies virus in wild canids. - Large-scale inventory of Brazilian biodiversity - Multi-center project designed in 2005 to boost taxonomic research in Brazil using the DNA barcode concept as a catalyst to integrate field collections, museum research, genome center networks and bioinformatic advances.

13. Central portion of the “standard” COI barcode (~400 bp) Building a carnivore barcode database: COI

14. Selection of a Carnivore-optimized mini-barcode for fecal DNA: - COI barcode (658 bp) -Sliding window approach; - Data set with 45 carnivore species representing several families

15. 16 Kb 12.5 Kb ATP6 (125 bp) Panthera Numt mtDNA COI 1560 bp BARCODE 648 bp MINI-COI (186 bp) Development of additional segments for carnivore non- invasive DNA barcoding: -Special interest in identifying jaguar vs. puma scats; - Nuclear mtDNA insertion in the genus Panthera (numt); - Use of a segment of the ATP6 gene

16. Species identification of field- collected scats from large felids using the ATP6 segment. 0.02 bPon56 (fezes Panthera onca em álcool) bPon55 (fezes de Panthera onca em DET) bPon306 (fezes de Panthera onca) bPon307 (fezes de Panthera onca) bPon56 (fezes Panthera onca - 98 mg) bPon32 (sangue de Panthera onca) bPon56 (fezes de Panthera onca em DET) P3-16 (pele de Panthera onca) bPon305 (fezes de Panthera onca) P3-2 (pêlos de Panthera onca) bPon56 (fezes de Panthera onca - 48 mg) bPon24 (sangue de Panthera onca) P31-1 (pêlos de Panthera onca) F2-636 (fezes de felino grande) Puma concolor (sangue) L.V.(fezes de felino grande) F57-1C (fezes de felino grande) F2-635 (fezes de felino grande) Felis catus 100 99

17. DNA identification of field-collected scats – ATP6 (105 bp)

18. 10/FOX/BA/04 77/FOX/BA/05 65/FOX/BA/05 66/FOX/BA/05 35/FOX/PE/05 33/FOX/PE/05 43/FOX/PE/05 45/FOX/PE/05 38/FOX/PE/05 40/FOX/PE/05 55/FOX/PI/05 37/FOX/PE/05 28/FOX/BA/04 4/FOX/BA/04 64/FOX/BA/05 41/FOX/PE/05 Cerd bcth184 PE 46/FOX/PE/05 36/FOX/PE/05 Cerd bcth228 MA 23/FOX/PI/05 42/FOX/PE/05 44/FOX/PE/05 Cerd LG212 MT 25/FOX/PI/05 Cerd bcth198 MG 2/FOX/BA/04 Lgymno LG166 Lgymno LG45(RS) Lvetulus LG282 Lvetulus LG185 MG Lvetulus LG184 BA 26/FOX/PI/05 59/FOX/PI/05 50/DOG/PE/05 51/DOG/PE/05 48/DOG/PE/05 34/DOG/PE/05 49/DOG/PE/05 81 99 80 99 77 71 96 97 93 89 47 72 53 51 62 0.02 Rabies virus in Brazilian wild canids (P. Carnieli Jr. – Inst. Pasteur – Brazil) [control region]

19. Phylogeography of the crab-eating fox (Cerdocyon thous) Northern Clade C14 C4 C3 C6 C2 C5 C15 C11 C9 C10 C17 C21 C16 C13 C12 C18 C8 C7 C19 C20 C1 C29 C25 C35 C22 C24 C26 C27 C28 C23 C30 C33 C34 C32 C31 P.vetulus P.gymnocercus 0.005 substitutions/site 99/70/100/100 85/ */62/53 99/94/99/100 Southern Clade mtDNA CR Tchaicka et al. 2007

20. Large-scale inventorying of Brazilian biodiversity (2005) Sampling strategy 15 sites: -10,000 samples/site: - fish - amphibians - reptiles (incl. birds) - mammals - spiders - Leguminosae

21. Large-scale inventorying of Brazilian biodiversity 6 museums - Involved in coordinating collection trips, curation of sampled specimens and their morphology-based analysis 14 Centers of Molecular Biodiversity: - Network of genome centers and molecular biology laboratories performing high-throughput sequencing of DNA barcodes from 150,000 samples; Bioinformatics tools: online management of data collection, analysis and integration (e.g. morphology, DNA barcodes, geography) Training activities: courses, workshops.

22. Paulo B. Chaves Vanessa G. Graeff Taiana Haag Ligia Tchaicka Tatiane C. Trigo Eunice Matte Paulo Prates Jr. Cristine S. Trinca Daniela Copetti Manoel Rodrigues Gabriel Macedo Luana Cardoso Henrique Figueiró Flávia Tirelli Anelisie S. Santos Ana Carolina G. Escobar Sandro L. Bonatto Brazilian DNA barcode network Oliver Ryder Pedro Carnieli Jr. Dênis Sana Flávio Rodrigues Cibele Indrusiak Ronaldo Morato Tadeu G. Oliveira Laury Cullen Jr. Rodrigo Jorge Leonardo Vianna Carlos De Angelo Mario S. Di Bitetti Support: CNPq, CAPES, FNMA/MMA, CESP, NIH-USA, PUCRS ACKNOWLEDGEMENTS: