Second International Barcode of Life Conference 18-20 September 2007 Conference Abstracts 1 Welcome

Dear Conference Participants, The Local Organizing Committee and the Consortium for the Barcode of Life welcome you to Taipei, to Academia Sinica, and to the Second International Barcode of Life Conference. The First Barcoding Conference was held more than two years ago at the Natural History Museum in London and momentum and participation in the Barcode of Life Initiative has increased remarkably since then. At the time of this writing, more than 350 people from more than 45 countries have registered for the conference. Over the course of the three-day conference, you will receive presentations from 34 speakers from 14 countries and see 125 poster displays presented by researchers from 38 countries. These numbers confirm that DNA barcoding has become a truly global enterprise. The Scientific Program Committee has designed the agenda to provide a broad overview of the latest developments in DNA barcoding science and technology. We will learn the “state-of-the-art” of barcoding lab protocols, data management, and data analysis. Presenters will bring us up to date on their progress on global barcoding projects and the application of barcoding to applied problems that face society. Panelists will discuss the future of barcoding and its relation to other fields of research. We hope that each of you will learn more about what interests you, and will become interested in the new things you will learn. The conference will include four types of presenters: Keynote Speakers (their presentations will be 30 minutes each), Invited Speakers (20 minutes each), Contributed Speakers selected through committee evaluation (15 minutes each), and Discussion Panelists. Almost every session includes time for discussion and questions from the audience. The sessions with Panel Discussions have been organized to stimulate discussion with the conference participants. We hope that you will take advantage of these opportunities for interaction with the speakers and each other. You will also see that the agenda includes 90-minute lunch breaks and 20-30 minute coffee breaks each morning and afternoon. We hope that you will use these opportunities to visit the poster presentations and exhibit booths and to interact with the presenters. The latest software for DNA barcoding will be demonstrated at several of the exhibit booths and we urge you to take a look. The Local Organizing Committee has arranged for meeting staff to be available at the registration desk throughout the meeting if you have any questions. You will also see Conference Assistants circulating around the meeting and we hope that you will ask them for help if you need anything. Thank you all for coming to Taipei to take part in this event. We hope that you find the conference stimulating and productive!

David E. Schindel Kwang-Tsao Shao Scott E. Miller Executive Secretary, Conference Chair CBOL Chair CBOL Academia Sinica Smithsonian Institution

Second International Barcode of Life Conference 18-20 September 2007 Conference Abstracts 2 Table of Contents

Welcome Letter …2

Sponsors…4

Agenda…5

Plenary Session Abstracts…8

Posters by Subject…62

Posters Floorplan…67

Poster Abstracts…68

Index…155

Second International Barcode of Life Conference 18-20 September 2007 Conference Abstracts 3 Sponsors

We would like to thank the following sponsors who have made this Conference possible:

Academia Sinica

National Science Council

Forestry Bureau, Council of Agriculture

Fisheries Agency, Council of Agriculture

Bureau of Animal and Plant Health Inspection and Quarantine, Council of Agriculture

Taipei City Government

National Taiwan Museum

Ministry of the Interior

Ichthyological Society Taiwan

Taiwan Forestry Research Institute

National Sun Yat-Sen University

National Museum of Natural Science

Taiwan Seed Improvemnet and Propagation Station

Consortium for the Barcode of Life

Smithsonian Institution

Genome Canada

BioNET International

Swiss Agency for Development and Cooperation

The Alfred P. Sloan Foundation

Special Thanks to Local Organizing Committee General Innovation Service (GIS) CBOL Secretariat

Second International Barcode of Life Conference 18-20 September 2007 Conference Abstracts 4 Agenda DAY ONE: TUESDAY 18 SEPTEMBER 2007

SESSION 1: GLOBAL ACTIVITIES OF THE BARCODE OF LIFE INITIATIVE (BOLI) 8:00 - 9:00 Conference Registration 9:00-9:30 Conference Opening - Kwang-Tsao Shao, Conference Chair, Academia Sinica 9:30 - 10:00 Keynote I -DNA Barcoding in the Genome Era - Ya-Ping Zhang, Kuming Institute of Zoology, Chinese Academy of Science 10:00 - 10:30 Keynote II – Biodiversity Research and DNA Barcoding in Korea - Won Kim, Seoul National University 10:30 - 11:00 Keynote III: Barcoding in Applied Taxonomic Challenges- Helida Oyieke, National Museums of Kenya 11:00 - 11:30 COFFEE BREAK SESSION 2: DIVERSE APPLICATIONS OF DNA BARCODING 11:30 - 11:45 Progress by CBOL since London Conference - Scott Miller, Smithsonian Institution (CBOL) 11:45 - 12:45 Brief presentations and panel discussion of major barcoding projects • Yvonne Linton, Natural History Museum London, CBOL Demonstrator Project 1: Mosquito Barcoding Initiative • Marc De Meyer, Royal Museum of Central Africa, Belgium, CBOL Demonstrator Project 2: Tephritid Barcode Initiative • Mark Stoeckle, The Rockefeller University, All Birds Barcoding Initiative (ABBI) Update • Robert Hanner, University of Guelph, The Fish Barcode of Life (FISH-BOL) Campaign • Mark Bagley, US Environmental Protection Agency, DNA Barcodes for Assessment of the Biological Integrity of Aquatic Ecosystems • Ann Bucklin, University of Connecticut, DNA Barcoding of the Global Marine Zooplankton Assemblage 12:45 - 1:00 Open discussion between audience and panel 1:00 - 2:30 LUNCH SESSION 3: HOW DOES BARCODING WORK AND HOW WELL DOES IT WORK? 2:30 - 3:00 Keynote Address- Toward a Barcoded World- Paul Hebert, University of Guelph 3:00 - 3:15 The BARCODE Data Standard: Enabling Molecular Diagnostics for Biodiversity-Bob Hanner, University of Guelph 3:15 - 3:30 Barcoding in an all-species inventory – The Moorea Biocode Project- Chris Meyer, University of California Berkeley 3:30 - 3:45 Barcodes bridge the old and the new: use of museum specimens to identify molecular operational taxonomic units in larvae of scarab beetles (Coleoptera: Scarabaeoidea) Andrew Mitchell, NSW Department of Primary Industries, Australia 3:45 - 4:00 DNA barcode, type specimens and species delimitation in the genus Eumunida Sarah Samadi, Museum National d'Histoire Naturel, Paris 4:00 - 4:30 COFFEE BREAK SESSION 4: HOW CLOSE ARE WE TO HAND-HELD, FIELD-FRIENDLY, OR TABLE-TOP BARCORDERS? 4:30 - 4:45 In vitro repair enhances amplicon recovery and accuracy from damaged DNA -Tom Evans, New England Biolabs 4:45 - 5:00 Express Barcodes: Racing from Bugs to Identifications -Natalia Ivanova, University of Guelph 5:00 - 5:15 Development of a DNA barcode-based DNA chip for identification of marine organisms in the East Sea of Korea -Youn-Ho Lee, Korea Ocean Research and Development Institute 5:15 - 5:30 Fish and Chips: microarray-based DNA-barcoding of European Marine Fishes - Kochzius Marc, University of Bremen 5:30 - 5:45 Portable Pyrosequencing device for DNA sequencing -Mostafa Ronaghi, Stanford University 5:45 - 6:00 Open discussion between audience and speakers

Second International Barcode of Life Conference 18-20 September 2007 Conference Abstracts 5 Agenda DAY 2: WEDNESDAY, 19 SEPTEMBER 2007

SESSION 5: ANALYTICAL METHODS FOR BARCODE DATA. 8:30 – 8:45 Introduction-Michel Veuille, Museum National d’Histoire Naturel Paris 8:45 - 9:00 Comparing phylogenetic and statistical classification methods for DNA barcoding – Frederic Austerlitz, CNRS/AgroParisTech/Universite Paris Sud 9:00 - 9:15 Fast Barcode-Based Species Identification Using String Kernels –Vladimir Pavlovic, Rutgers University 9:15 - 9:30 Species Classification with Optimized Logic Formulas – Giovanni Felici, Consiglio Nazionale delle Ricerche 9:30 - 9:45 DNA Barcode sequence identification incorporating taxonomic hierarchy and within taxon variability – Damon Little, The New York Botanical Garden 9:45 - 10:00 A Comparison of Algorithms for Species Identification based on DNA barcodes – Bogdan Pasaniuc, University of Connecticut 10:00 - 10:30 Open discussion between audience and speakers 10:30 - 11:00 COFFEE BREAK SESSION 6: CASE STUDIES I. INTEGRATIVE TAXONOMIC STUDIES USING DNA BARCODING 11:00 - 11:20 Barcoding, Biogeography and Evolution in the Tropics of the New World – Eldredge Bermingham, STRI, Panama 11:20 - 11:40 The role of DNA Barcoding in the broader context of Integrative Taxonomy - Benoit Dayrat, University of California, Merced 11:40 - 12:00 Sphingids and Barcodes- The New Taxonomy- Rodolphe Rougerie, University of Guelph 12:00 - 12:20 Bamboozled by bloodsuckers: barcoding backs biodiversity- Mark Siddall, American Museum of Natural History 12:10 - 12:30 Open discussion between audience and speakers 12:30 - 2:00 LUNCH/ Group Photo at Front Gate SESSION 7: WHAT NEW SCIENCE WILL WE SEE AT THE THIRD BARCODE CONFERENCE? 2:00 - 2:20 Barcoding and biodiversity research - Les Christidis, Australian Museum 2:20 - 2:40 "Ecological inventory with a barcorder, the ecological taxascope: who eats what in a complex tropical forest" - Dan Janzen, University of Pennsylvania 2:40 - 3:40 Panel discussion: What new research can we do with a MILLION Barcode records? • Paul De Barro, CSIRO, Australia • Peter Ng, National University of Singapore • Axel Meyer, University of Konstanz • Robert Andersen, Bigelow Laboratory for Ocean Sciences 3:40 - 4:00 COFFEE BREAK SESSION 8: CASE STUDIES II. DNA BARCODING IN DEVELOPING COUNTRIES. 4:00 - 4:20 DNA Barcoding: considerations for vectors of neglected diseases - Daniel Masiga, ICIPE, Nairobi, Kenya 4:20 - 4:40 All Birds Barcoding Initiative in the Neotropics: Identifying and discovering bird species in the richest avifauna of the World - Pablo Tubaro, National Museum of Natural History, Argentina 4:40 - 5:00 FISH-BOL and Barcoding in India - Wazir Lakra, National Bureau of Fish Genetic Resources, India 5:00 - 5:15 Transforming taxonomy for effective biodiversity assessment of arthropods in Madagascar -Brian Fisher, California Academy of Sciences 5:15 - 5:30 Construction of A DNA Barcode System for Amphibians and Reptiles in Taiwan – Si-Min Lin, Chinese Culture University, Taiwan 5:30 - 5:45 Open discussion between audience and speakers

Second International Barcode of Life Conference 18-20 September 2007 Conference Abstracts 6 Agenda

DAY 3: THURSDAY, 20 SEPTEMBER 2007

SESSION 9: BARCODING IN THE WIDER EVOLUTIONARY CONTEXT 8:30 - 9:30 Panel discussion: Barcoding and other initiatives in evolutionary research • Michael Donoghue, Yale University, Barcoding and the Tree of Life • Masaki Miya, Natural History Museum, Chiba University, Evolutionary History of Fishes: An overview based on >1000 whole mitochondrial genome sequences • Dan Faith, The Australian Museum, How large-scale barcoding promotes large-scale biodiversity assessment • Richard Lane, Natural History Museum London, Barcoding and the practice of systematics 9:30 - 10:00 Open discussion between audience and panel 10:00 - 10:30 COFFEE BREAK SESSION 10: STATE-OF-THE-ART PRACTICES: HOW TO MANAGE YOUR BARCODE DATA. 10:30 - 10:50 BOLD, the Barcoding Workbench - Sujeevan Ratnasingham, University of Guelph 10:50 - 11:50 Panel discussion: How are the BAROCDE data standards being implemented? • Scott Federhen, NCBI/NIH, BARCODE records in GenBank • Simon Tillier, Museum National d’Histoire Naturel, Paris, BARCODE data, museum catalogs and GBIF • Andrew Polaszek, ICZN, London, Zoobank- the open-access animal name registry-working with CBOL towards the new taxonomy 11:50 - 12:10 The Encyclopedia of Life: A website for every species - James Edwards, Smithsonian Institution (EOL) 12:10 - 12:30 Open discussion between audience and speakers 12:30 - 2:00 LUNCH

SESSION 11: STATE-OF-THE-ART PRACTICES: HOW DO THE BEST BARCODE LABS DO THEIR WORK? 2:00 - 2:20 Technology Transfer, an efficient DNA barcoding workflow: how we do it and more importantly to you, how we can help you do it- Lee Weigt, Chair, CBOL DNA Working Group, Smithsonian Institution 2:20 - 2:40 African Wildlife Tissue Banking - Paul Bartels, BioBank, South Africa 2:40 - 3:00 Recovering historic DNA (HDNA) and ancient DNA (ADNA): Implications for DNA barcoding - David Lambert, Allan Wilson Centre, New Zealand 3:00 - 3:20 Exploring archival and environmental samples through minimalist barcodes- Mehrdad Hajibabaei, University of Guelph 3:20 - 3:30 Open discussion between audience and speakers 3:30 - 4:00 COFFEE BREAK SESSION 12: BARCODING WITH NON-COI GENE REGIONS. 4:00 - 4:20 Optimising selection of DNA barcode regions and CBOL’s guidelines for non-CO1 selection Freek Bakker, University of Wageningen, Netherlands Herbarium 4:20 - 4:40 Standardized land plant barcoding requires a multi-loci approach - Robyn Cowan, Royal Botanic Gardens, Kew 4:40 - 5:00 Using DNA barcodes to test the identity and purity of plant-based medicines and herbals - John Kress, Smithsonian Institution 5:00 - 5:20 DNA-barcoding on a different scale: Challenges within Coccoidea - Lyn Cook, University of Queensland 5:20 - 5:40 Canadian plant barcoding results with some global implications - Sean Graham, University of British Columbia 5:40 - 6:00 Progress toward DNA barcoding the vast diversity of fungi- Amy Rossman,USDA Beltsville 6:00 Closing remarks and adjournment - Kwang-Tsao Shao, Conference Chair, Academia Sinica

Second International Barcode of Life Conference 18-20 September 2007 Conference Abstracts 7 Plenary Session Abstracts (By Speaker Order)

Session 1: Global activities of the Barcode of Life Iniative (BOLI)

KEYNOTE ADDRESS

DNA BARCODING IN NEW GENOME ERA

YA-PING ZHANG

State Key Laboratory of Genetic Resource and Evolution, Kunming Institute of Zoology, Chinese Academy of Science, Kunming 650223 Laboratory for Conservation and Utilization of Bioresource, Yunnan University, Kunming 650091

Both the identification and the knowledge of evolutionary position of species are very important for all biological studies. The new identification systems based on DNA sequence diversity have been developed in recent years. The ideal core barcode DNA fragments should not only enable accurate, rapid and cost-effective identification of species, but can also provide high phylogenetic resolution. A 648-bp fragment of the cytochrome c oxidase I (COI) gene has been selected as the core barcode DNA marker for animals. We sequenced the entire mitochondrial genome of the Ursidae family, which represents a typical example of rapid evolutionary radiation, to evaluate the individual mitochondrial gene performance in supporting the complete mtDNA-based phylogeny. Among single gene trees, only the CYTB and ND5 trees have the same branching order as that from the combined all gene analysis. With more and more genome sequence data from animals, we will be able to evaluate the effect of mitochondrial pseudogenes on DNA barcoding, and develop additional markers for DNA barcoding.

Zhang Ya-Ping

Dr. Zhang Ya-Ping held a B.S. in Biochemistry from Fudan University in 1986 and a Ph.D. in Genetics Zoology in 1991 from Kunming Institute of Zoology (KIZ), Chinese Academy of Sciences. He completed post-doctoral studies at the Center for Research of Endangered Species (CRES), the Zoological Society of San Diego during 1992-1995. He has been working in KIZ Since 1995. He was appointed as an Academician of CAS in 2003 and the director of KIZ in 2005.

His major research interests are Conservation genetics, molecular evolution, and population genetics. The Laboratory’s current research interests can be summarized into following areas: (1) adaptive evolution, (2) molecular phylogeny of animals, (3) population genetics and conservation genetics of animal species in China, (3) genetic diversity and origin of domesticated animals, (4) genetic diversity and evolution history of human populations in East Asia, (5) genome evolution.

The results of all above fields have been published more than 170 papers in peer review journals, including Nature, Science, PNAS, Mol. Biol. Evol., Mol. Phylogen.Evol., Am. J. Hum. Genet., Genetics, Genomics and et al.

Second International Barcode of Life Conference 18-20 September 2007 8 Conference Abstracts Plenary Session Abstracts

KEYNOTE ADDRESS

BIODIVERSITY RESEARCH AND DNA BARCODING IN KOREA

WON KIM

School of Biological Sciences, Seoul National University

Biodiversity research in Korea has been far from comprehensive. Even the biota has not been systematically well surveyed or documented owing to a lack of taxonomic specialists and research funds. Nevertheless the major taxonomic works have been published in “Illustrated Encyclopedia of Fauna & Flora of Korea” since 1959 (1959-2006; vol.1-vol.41). After the Rio meeting in 1992, numbers of systematists have consistently emphasized the importance of systematics along with biological resources in public. On years of efforts, “National Institute of Biological Resources, Korea” is scheduled to be open in October, 2007. A master plan of the tentatively named “National Institute of Marine Biological Resources, Korea” is also being promoted. These two institutes will be the central units managing biological resources in Korea. The molecular systematics was first introduced to Korea in 1989. Since then, the majority of Korean systematists have adopted molecular methods, with developments of technology, in their researches. While a few systematists have individually conducted DNA barcoding researches, the necessity of national undertaking DNA barcording project has been invoked through several symposiums. A three year DNA barcode research project on all major groups of animals and plants, sponsored by Korea Institute of Environmental Science and Technology, has been finally launched this year. We hope for this project to be continued.

Won Kim

Dr. Won Kim received a B.S. and a M.S. from the Dept. of Zoology, Seoul National University in 1977 and 1981, respectively. He received a Ph.D. from the Dept. of Biological Sciences, Florida State University in 1985 and worked as a research associate at the same department during 1986-1989. He has been working in Seoul National University since 1989. He served as a chairperson of the Dept. of Molecular Biology during 1995-1999 and an associate dean of College of Natural Science during 1999-2000. He is a member of the Korean Academy of Science and Technology since 2004. His major research interests are classical taxonomy, molecular systematics, molecular ecology and biogeography, and molecular evolution and bioinformatics.

The current research interests can be summarized into following areas: (1) classical taxonomy of various marine invertebrates, (2) molecular systematic studies of selected invertebrates, (3) molecular identification based on various molecular markers and DNA barcoding, (4) microsatellite search and studying population structure, (5) evolution of genes based on nucleotide analysis, (6) pattern search for finding candidate genes. About 150 papers have been published in various journals, including Smithoni. Contrib. Zool., J. Crustacean. Biol., Korean J. Syst. Zool., Mol. Biol. Evol., Mol. Phylogen. Evol., J. Mol. Evol., Nature, Mol. Ecol. Notes, Computat. Biol. Chem., etc.

Second International Barcode of Life Conference 18-20 September 2007 9 Conference Abstracts Plenary Session Abstracts

KEYNOTE ADDRESS

BARCODING IN APPLIED TAXONOMIC CHALLENGES

HELIDA OYIEKE

National Museums of Kenya

Taxonomy plays a very important function in the scientific and socio-economic arenas by facilitating species classification and identification. Millions of species have been discovered in both aquatic and terrestrial systems while many more are yet to be. Traditional taxonomy has in the past met human needs in biodiversity conservation and management, agricultural production and human health. However, as the demand for taxonomic services increases, traditional taxonomy is becoming inadequate in coping with the demand. The issue is aggravated by the fact that the number of taxonomists is significantly getting reduced globally. It is currently clear that DNA barcoding technique is increasingly complimenting the efforts made by taxonomists. It is becoming a tool that could settle conflicts arising among taxonomists on species identities. The technique becomes handy in cases where only parts of specimens are available such as in the identification of wildlife and their products in illegal trade. Barcoding can be used as a tool for flagging new species that have not yet been described as well as cryptic species. The neotropical skipper butterfly, Astrapes fulgerator, acryptic species complex found in north-western Costa Rica, was found to consist of 10 different species through DNA barcoding by Herbet et al. Associating all life history stages of a specific species have been a challenge to traditional taxonomy and there is great potential in surmounting this huddle through barcoding. Much as DNA barcoding technique is becoming a possible solution to problems that traditional taxonomy has delayed or failed to solve in good time, a lot of mileage will be gained if experts from the two divide work together to complement one another.

Dr. Helida A. Oyieke

Dr. Helida Oyieke is the Director for Research and Collections at the National Museums of Kenya, a natural history based museum centered in Nairobi, where she oversees and provides general leadership on research programmes in the institutions. She holds a B.Ed, M.Sc and Ph.D degrees in biological sciences all from the University of Nairobi and is specialized in the Marine Botanical Sciences. Helida started her research career in 1984 at the Kenyan Marine and Fisheries Research Institute in Mombasa where she rose to the rank of Senior Research Scientist. In 1994 she joined her current institution as a Senior Research Scientist, rose to Principal Scientist position before finally being appointed to her current position in 2004.

Throughout her research career she has mobilized grants for research in the fields of taxonomy, ecology, biodiversity conservation and collection management, and serves in several technical national and regional committees. Dr. Oyieke is currently serving as a member of CBOL Executive Committee, representing Africa, and also chairs the steering committee for the eastern Africa Barcode Initiative where she is making efforts to popularize barcoding technology as a tool for taxonomy in the region.

Second International Barcode of Life Conference 18-20 September 2007 10 Conference Abstracts Plenary Session Abstracts

Session 2: Diverse Applications of DNA Barcoding

PROGRESS SINCE THE 2005 LONDON DNA BARCODE OF LIFE CONFERENCE

SCOTT E. MILLER

Smithsonian Institution, Washington D.C., USA

In February 2005, 240 people from 44 countries participated in the First International Barcode of Life Conference at the Natural History Museum in London. At that time, the Consortium for the Barcode of Life had 42 Member Organizations from 18 countries. There were 33,000 barcode records representing 12,700 species in BOLD, the Barcode of Life Data Systems at the University of Guelph in Canada. CBOL now has about 160 Member Organizations from more than 50 countries and there are almost 300,000 barcode records from 31,300 species in BOLD, and these records are starting to flow into GenBank. The BARCODE data standard has been adopted by GenBank, EMBL and DDBJ and the standards are being implemented, in partnership with GBIF, GenBank and other informatics initiatives. Global barcoding projects on birds, fish, agricultural pests, mosquitoes, Lepidoptera, fungi and other taxa have been launched and others are in the planning stages. There is a regular flow of publications documenting and expanding the use of DNA barcodes in taxonomy, ecology, biogeography, and applications such as forensics. Government agencies in several countries are testing barcoding for formal adoption as a tool for regulation and enforcement. As part of its effort to promote global participation in barcoding, CBOL has held outreach meetings to southern and eastern Africa and to South/Central America. CBOL has also organized workshops to build collaboration with marine scientists, molecular evolutionists, phylogenetics projects, and bioinformatics projects. Most recently, CBOL has initiated a global network of “Leading Labs” that are sharing and disseminating information on barcoding and will be offering training opportunities.

The Taipei Barcode of Life Conference is an opportunity to see the tremendous progress made by the Barcode of Life Initiative since February 2005, and to plan for the next few years of work. CBOL invites participants in the conference to share their ideas for the goals we should set for the Third International Barcode Conference in 2009.

Scott E. Miller

Scott E. Miller is Chair of the Executive Committee of the Consortium for the Barcode of Life, and has an active research program applying DNA barcodes to the systematics and ecology of moths. He is Senior Program Officer in the Office of the Under Secretary for Science of the Smithsonian Institution, and Curator of Entomology at the Smithsonian National Museum of Natural History. He has a Ph.D. from Harvard University and has published over 145 research publications in systematics, biogeography, and ecology. At the Smithsonian he has previously served as Associate Director for Science at the National Zoological Park, and Chairman of the Departments of Entomology and Systematic Biology in National Museum of Natural History. Before coming to the Smithsonian, he led the Natural Science Department and Hawaii Biological Survey at Bishop Museum (Hawaii) and the Biodiversity and Conservation Programme at International Centre of Insect Ecology and Physiology (Kenya). He is committed to applying biodiversity information from biodiversity collections and systematics research to conservation and sustainable development, has participated in major reviews of biodiversity related to conservation planning, and catalyzed an Integrated Conservation Development Project in Kenya. He co-edited the books "Papua New Guinea Biological Diversity Country Study," "The origin and evolution of Pacific island biotas, New Guinea to Eastern Polynesia: Patterns and processes" and "Arthropods of tropical forests: Spatio-temporal dynamics and resource use in the canopy."

Second International Barcode of Life Conference 18-20 September 2007 11 Conference Abstracts Plenary Session Abstracts

CBOL DEMONSTRATOR PROJECT 2: MBI (MOSQUITO BARCODING INITIATIVE)

YVONNE-MARIE LINTON

The Natural History Museum, Cromwell Road, London SW7 5BD, England.

The requirement for correct species identification in mosquitoes needs little introduction. Malaria alone kills more than 1 million people each year, and infects between 300-500 million worldwide, yet this is only on of many socio-economically devastating diseases transmitted by mosquitoes. Key to disease management is vector control, yet these efforts consistently hampered by out-dated morphological keys which perpetuate species misidentification. Only few expert mosquito taxonomists remain, and they struggle to keep up with the rapid discovery of new taxa, particularly cryptic species, detected by molecular techniques and revisionary taxonomic works. The Mosquito Barcoding Initiative was established to tackle this global taxonomic impediment and provides properly identified specimens, with associated voucher material, to populate the BOLD and ultimately GenBank databases with high quality sequence data. This will serve as a taxonomic framework for systematic studies in the future, and data will be freely accessible.

There are currently 3,489 recognized species in the Culicidae. The MBI aims to sequence a minimum of 5 specimens per species, for 80% of all known mosquitoes within two years. To facilitate this coverage in this time frame, the MBI relies heavily on archive specimens in museums across the globe. DNA retrieved from these specimens is often degraded and/or sheared and herein we will discuss the methodologies adapted to overcome these issues and produce high quality sequence chromatograms. Six months in, we will present the progress of the MBI, discussing in detail the utility of the COI barcode region in species identification across eighteen mosquito genera, and discussing the limitations of the COI region for reliable identification of subspecific taxa and in some cryptic species complexes.

Yvonne Linton

Dr Yvonne-Marie Linton works in the Department of Entomology at the Natural History Museum in London, where she heads a team of molecular systematists interested in mosquitoes and the diseases they transmit. Her scientific interests include medical entomology, mosquito and midge systematics and vector biology. Her particular passion is teasing out component members of species complexes. Dr Yvonne-Marie Linton trained as a Zoologist and holds a B.Sc. (Hons.) in Zoology (Environmental Physiology) and a PhD in Molecular Entomology from the University of Aberdeen in Scotland. After finishing her PhD on Culicoides biting midges in 1998, Yvonne started working on Mosquitoes at the Natural History Museum in London. She currently holds honorary research positions in the University of Antioquia, Colombia, is the chairperson of MOTAX (Mosquito Taxonomy Group), Editor of the European Mosquito Bulletin (Journal of the European Mosquito Control Association), and is project leader of the Mosquito Barcoding Initiative.

Second International Barcode of Life Conference 18-20 September 2007 12 Conference Abstracts Plenary Session Abstracts

CBOL DEMONSTRATOR PROJECT 2: TBI (TEPHRITID BARCODING INITIATIVE)

MARC DE MEYER1, Bruce McPheron2 and Allen Norrbom3

1Royal Museum for Central Africa, Tervuren, Belgium 2Pennsylvania State University, University Park, PA, USA 3Smithsonian Institution, Washington D.C., USA

TBI aims at creating a system to allow identification of any specimen from an agricultural pest group worldwide, using fruit flies as a subject group. A steering committee was composed in April 2006 and detailed strategy, protocol and timeline developed. Because of delay in obtaining start-up funds, activities only started early 2007. Subsequent efforts have led to agreement from the key museums housing specimens to participate in the project and laboratory confirmation of repeatable DNA extraction and PCR amplification from museum specimens using an invasive, non- destructive protocol. The work, through regional centers, focuses on: COI database including all species of economic importance, their closest relatives, and a representative selection of other tephritid clades; a follow-on research agenda, material for a DNA ‘bank’ to include extractions and amplification products, and evaluation of the demonstrator as a practical, cost-effective pest identification system. Because TBI recognizes the fact that the limited number of tephritid taxonomists is decreasing, training in tephritid systematics is envisaged. Currently, DNA extraction and PCR amplification methods are largely in place and part of the tephritid specimens in museums has been requested. Actual routine DNA extraction, amplification and sequencing is at its initial stages with the main emphasis on relatively recent material of those taxa of economic significance and congeners.

Marc De Meyer

Dr. Marc De Meyer is the Head of the Entomology Section at the Royal Museum for Central Africa at Tervuren (RMCA, Belgium). The RMCA is a Belgian Federal Research Institute and Museum, focusing on natural and human sciences in Africa. The Entomology Section holds a collection of more than 6 million insect specimens.

Dr De Meyer was trained as a zoologist at the University of Antwerp, with post-graduate in tropical medical biology at the Institute for Tropical Medicine (both in Belgium). He obtained a Ph.D. in zoology at the former institution. Prior to his appointment as Head of the Entomology Section, he worked for 10 years in Africa (Botswana, and Kenya).

His research focuses on the systematics, phylogenetics and biogeography of Diptera, in particular Pipunculidae and Tephritidae. He is also in charge of the curation of the Coleoptera collection at the RMCA. He is one of the coordinators of the Tephritid Barcoding Initiative (TBI), one of the demonstrator projects of CBOL.

Second International Barcode of Life Conference 18-20 September 2007 13 Conference Abstracts Plenary Session Abstracts

ALL BIRDS BARCODING INITIATIVE (ABBI) UPDATE

MARK STOECKLE

Program for the Human Environment, The Rockefeller University, New York, NY

Genetic surveys including with DNA barcoding suggest there are hundreds of as yet undescribed bird species. The All Birds Barcoding Initiative (ABBI), launched in September 2005, aims to collect DNA barcodes from the approximately 10,000 species of world birds. ABBI will help speed discovery of new species and provide a practical tool to confirm identifications, including of live specimens in banding studies. As of July 2007, ABBI researchers have deposited over 9000 barcodes representing more than 1800 species in the Barcode of Life Database (BOLD), an electronic library that links barcodes, reference specimens in collections, and associated collection data. BOLD has barcodes from most all (95%) of northern Nearctic species, and growing representation of Neotropical and Palearctic avifauna. A challenge for ABBI will be to obtain barcodes for the approximately 3000 world birds which are not represented in tissue collections, particularly Afrotropical and Indomalayan avifauna. Depending on cost, one option will be to analyze DNA from museum skin collections, which contain millions of specimens representing nearly all world birds. Avian barcode data supports the emerging view that selective sweeps prune mitochondrial diversity.

Marc Stoeckle

Mark Stoeckle is an Adjunct Faculty Member in the Program for the Human Environment at The Rockefeller University. Since joining the Program in January 2003, he co-authored a DNA barcoding protocol for the Census of Marine Life, and helped organize the DNA barcoding conference held at Cold Spring Harbor Laboratory in September 2003, and the inaugural meeting of the Consortium for the Barcode of Life at Smithsonian Institution, National Museum of Natural History in May 2004. Dr. Stoeckle co-authored two recent DNA barcoding studies which uncovered 15 possible new bird species in North America. Dr. Stoeckle helped organize the inaugural workshop for the All Birds Barcoding Initiative held at Museum of Comparative Zoology in September 2005. As ABBI Coordinator he works with an international group of ornithologists to collect barcodes for the approximately 10,000 species of world birds.

Dr. Stoeckle is a graduate of Harvard University and Harvard Medical School and completed his medical training in Internal Medicine and Infectious Diseases at The New York-Presbyterian Hospital-Weill Medical College. During 1984-1989 he was a Research Associate at The Rockefeller University. He is a Clinical Associate Professor of Medicine and Associate Attending Physician at The New York-Presbyterian Hospital. In addition to scientific interests, Dr. Stoeckle is an accomplished graphic artist and award-winning nature photographer.

Second International Barcode of Life Conference 18-20 September 2007 14 Conference Abstracts Plenary Session Abstracts

THE FISH BARCODE OF LIFE (FISH-BOL) CAMPAIGN

ROBERT HANNER

Biodiversity Institute of Ontario and Department of Integrative Biology, University of Guelph, Guelph, ON N1G 2W1 Canada

The Fish Barcode of Life Campaign (FISH-BOL; www.fishbol.org) aims to assemble a reference sequence library for all cartilaginous and bony fishes to better characterize and broadly identify species.Launched in May 2005, FISH-BOL currently includes over 160 researchers. Ten Regional Working Groups have been established to cover all of the FAO fishing areas of the world, using species checklists drawn from FishBase. DNA sequences are derived from expert-identified voucher specimens and are integrated using the Barcode of Life Data System (BOLD; www.Boldsystems.org), where they can be querried by the public without charge. The database contains DNA barcodes, images, and geospatial coordinates of examined specimens, providing information on species distributions.

More than 20,000 specimens have been analyzed to date, covering over 4,000 described species. The FISH-BOL initiative represents the first effort to assemble a global sequence library for such a diverse group of organisms and benefits from organizational support provided by the Consortium for the Barcode of Life.

Robert Hanner

Dr. Robert Hanner is the Associate Director of the Canadian Barcode of Life Network, headquartered at the Biodiversity Institute of Ontario on the University of Guelph campus, where Dr. Hanner also holds a faculty position in the Department of Integrative Biology. The Network includes over 40 university, museum and government scientists dedicated to barcoding the flora and fauna of Canada.

Dr. Hanner holds a B.S. in Biology from Eastern Michigan University and a Ph.D. in Biological Sciences from the University of Oregon. Dr. Hanner was a Kalbfleisch Postdoctoral Fellow at the American Museum of Natural History (AMNH) from 1998 to 1999. He was hired as Curatorial Associate of the Ambrose Monell Collection for Molecular and Microbial Research at the AMNH, where he spearheaded development of the Museum’s new cryogenic collection unit from 2000-2003. Dr. Hanner joined the faculty of the Coriell Institute for Medical Research, where he served as Scientific Program Director of the Coriell Cell Repositoires from 2003-2005. Dr Hanner served as President of the International Society for Biological and Environmental Repositories from 2002-2005. He is currently Chair of the CBOL Database Working Group and Campaign Coordinator for FISH-BOL.

Second International Barcode of Life Conference 18-20 September 2007 15 Conference Abstracts Plenary Session Abstracts

DNA BARCODES FOR ASSESSMENT OF THE BIOLOGICAL INTEGRITY OF AQUATIC ECOSYSTEMS

MARK BAGLEY

United States Environmental Protection Agency, Cincinnati, OH, USA

Water quality regulations and aquatic ecosystem monitoring increasingly rely on direct assessments of biological integrity. Because these aquatic “bioassessments” evaluate the incidence and abundance of sensitive aquatic species, they are able to measure cumulative ecosystem effects of chronic or intermittent pollutants in a way that water chemistry assays cannot. Reliance on morphological identifications has constrained greater adoption of bioassessment protocols due to limited availability of trained taxonomists, difficulty or inability to identify cryptic larval forms with high taxonomic resolution, and sample size limitations imposed by the time and cost of morphological analysis. Integration of DNA barcodes into aquatic monitoring and assessment programs provides a viable solution to each of these problems. We are comparing the accuracy, precision, and cost of taxonomic identifications based on DNA barcode and morphological approaches for an existing state (Maryland, USA) bioassessment program, focusing on aquatic insects in the families Ephemeroptera, Trichoptera, and Plecoptera. Importantly, morphologically cryptic aquatic larvae collected in streams are linked via DNA barcodes to taxonomically resolved adults vouchered at the Smithsonian Institution. Our results will be used to recommend strategies to integrate DNA and morphological approaches into bioassessment programs that optimally leverage available resources for specimen collection, taxonomic assessment, and data analysis.

Mark Bagley

Dr. Mark Bagley is a research biologist and acting chief of the Molecular Ecology Research Branch of the United States Environmental Protection Agency (US EPA). This branch of the US EPA uses molecular tools to assess the ecological integrity of aquatic ecosystems and to evaluate the efficacy of U.S. environmental regulations.

Dr. Bagley received his Ph.D. in Genetics from the University of California at Davis. He also holds an M.S. degree in Animal Science (UC Davis) and B.A. in Ecology, Behavior and Evolution (UC San Diego). Dr. Bagley joined the US EPA in 1999 where his work has focused on analyses of population genetic methods in environmental assessments, focusing on fish and aquatic macroinvertebrates. He has used similar methods to assess assumptions behind insect resistance management models that form the basis of US EPA’s biotechnology risk assessments. He is co-lead on a multi-agency (US EPA, Smithsonian Institution, Maryland Department of Natural Resources) project to assess the use of DNA barcodes in assessment of the integrity of aquatic ecosystems.

Second International Barcode of Life Conference 18-20 September 2007 16 Conference Abstracts Plenary Session Abstracts

DNA BARCODING OF THE GLOBAL MARINE ZOOPLANKTON ASSEMBLAGE

ANN C. BUCKLIN

Department of Marine Sciences, University of Connecticut, 1080 Shennecossett Road, Groton, CT 06340 USA

The marine holozooplankton assemblage includes ~7,000 described species in 15 phyla. As part of a global survey of biodiversity, the Census of Marine Zooplankton (CMarZ) has determined DNA barcodes for more than 1,500 zooplankton species and seeks to barcode all described species by 2010. The growing database of DNA barcodes will serve the function of a ‘Rosetta Stone’ for the zooplankton community. Uses of the database include: identifying individual specimens, revealing cryptic species, describing biogeographical distributions, discovering new species, and characterizing species diversity through environmental sequencing. In the not-too-distant future, DNA barcodes may be used in automated taxonomic analysis of known species in zooplankton samples, thus freeing experts to focus on the identification and description of new species. Eventually, DNA barcodes may be used for remote and autonomous characterization of species diversity and biogeography throughout the world oceans.

Ann C. Bucklin

Ann Bucklin is a professor and head of the Department of Marine Sciences and Director of the Marine Sciences and Technology Center at the University of Connecticut. Prior to joining the University of Connecticut in 2005, she was a professor at the University of New Hampshire. Dr. Bucklin was Program Manager of the Oceanic Biology Program at the Office of Naval Research (1988 - 1991). She was a Fulbright Senior Scholar in Norway (1992 - 1993) and was elected Fellow of the American Association for the Advancement for Science in 1995. She served as the director of the New Hampshire Sea Grant Program during 1993 - 2005. She is currently completing a 3-year term as the USA Academic Delegate to the International Council for the Exploration of the Sea (ICES). Dr. Bucklin received her B.A. in biology from Oberlin College and her Ph.D. in zoology from the University of California, Berkeley. The theme underlying her research interest—spatial and temporal patterns of molecular genetic variation in marine organisms—has developed from her early interest in sea anemones to current work on planktonic crustaceans. She leads the Census of Marine Zooplankton (CMarZ), a Census of Marine Life ocean realm field project launched in 2004 and is a member of the International Committee for the Ocean Biogeographical Information System (OBIS).

Second International Barcode of Life Conference 18-20 September 2007 17 Conference Abstracts Plenary Session Abstracts

Session 3: How does barcoding work and how well does it work?

KEYNOTE ADDRESS

TOWARDS A BARCODED WORLD

PAUL HEBERT

Biodiversity Institute of Ontario, University of Guelph, Guelph, ON N1G 2W1

All-species barcode libraries are under construction for several animal groups and the International Barcode of Life Project plans coverage for 500K species within 5 years. Because of the fractal nature of genomes, this work will yield insights which are both taxonomically comprehensive and genomically representative. By probing biotas rather than small assemblages of species, barcoding should allow traditional case studies to gain context. Are the youthful African Rift Lake cichlids exceptional or common? Further, by merging industrial-scale sampling with massively parallelized sequencing, barcode reference libraries should revolutionize our capacity to map the distributions of life. However, these advances depend upon two factors - the ability of barcodes to deliver species identifications and the capacity for rule sets developed from investigations on well-studied groups to illuminate taxonomic terra incognita. Results from the first 30K animal species provide cause for optimism. Species resolution is high and rule sets are remarkably robust across groups with strident differences in biology. Further, early investigations suggest these conclusions are extensible to the other domains of eukaryotic life. As a consequence, barcoding the world seems irresistible.

Paul Hebert

A native of Kingston (Ontario), Dr. Paul Hebert completed his undergraduate work in biology at Queen’s University, his PhD in genetics at Cambridge University and postdoctoral fellowships at the University of Sydney and at the Natural History Museum in London. He took up a faculty position at the University of Windsor in 1976 where he remained until repositioning to the University of Guelph in 1990 where he holds a Canada Research Chair in Molecular Biodiversity. Over his career, Dr. Hebert has served as Director of the Great Lakes Institute at Windsor, as Chair of the Department of Zoology at Guelph, and as Chairman of the Huntsman Marine Science Centre in St. Andrews.

He is currently Director of the Biodiversity Institute of Ontario and of the Canadian Barcode of Life Network. Dr. Hebert has authored 275 papers, most employing molecular markers to probe issues related to biological diversity. He has supervised more than 70 graduate students and postdoctoral fellows over his career, has received varied national and international scientific awards and is a Fellow of the Academy of Sciences of Canada.

Second International Barcode of Life Conference 18-20 September 2007 18 Conference Abstracts Plenary Session Abstracts

THE BARCODE DATA STANDARD: ENABLING MOLECULAR DIAGNOSTICS FOR BIODIVERSITY

ROBERT HANNER and the CBOL Database Working Group

Biodiversity Institute of Ontario, University of Guelph, Guelph, ON N1G 2W1

The Consortium for the Barcode of Life (CBOL) promotes DNA barcoding as a tool for the identification of species. The adoption of standard gene targets is a primary objective of barcoding. However, additional requirements are necessary to enable a robust molecular-diagnostic platform capable of supporting species-level identifications. While data couplets (consisting of paired "species name" & "sequence contig assembly" elements) form the core look-up table used for the barcode identification of unknowns, both elements of these couplets are inferences. Access to the underlying evidence supporting each inference is crucial. The voucher specimens examined must be available for secondary inspection and the species names used must be valid and consistently applied. Similarly, the underlying molecular data used to generate the sequence contig assembly must also be available to gauge the reliability of diagnostic molecular characters. These considerations led to the formation of a community consensus-based BARCODE Data Standard to differentiate barcode records from typical genetic database entries. Key requirements include: a minimum of 500 basepairs of bidirectional sequence with fewer than 1% ambiguous base calls, archiving of the electropherogram trace files with notation of the PCR primer sequences used to generate them, and explicit reference to a voucher specimen. The Barcode of Life Data System (BOLD) supports these requirements, facilitating the assembly of barcode records. CBOL, GBIF and GenBank are constructing an online registration system for repository identifiers that are essential for linking barcode records to archived voucher specimens. The International Nucleotide Sequence Database Collaboration (INSDC; DDBJ/EMBL/GenBank) is the open archive for published sequence data and working in collaboration with CBOL, the INSDC agrees to annotate barcode compliant data submissions with the reserved keyword BARCODE. A new barcode submission tool developed by GenBank aids this process. These developments facilitate a deeper integration of genomics and biodiversity science by creating a robust molecular diagnostic interface between them.

Robert Hanner

Second International Barcode of Life Conference 18-20 September 2007 19 Conference Abstracts Plenary Session Abstracts

BARCODING IN AN ALL SPECIES INVENTORY – THE MOOREA BIOCODE PROJECT

CHRISTOPHER MEYER

University of California Berkeley

The Moorea Biocode Project (MBP) is a multi-national, multi-institutional project to digitally characterize the biodiversity for the entire ecosystem of a tropical island in French Polynesia. A key unifying goal of the MBP is to barcode all macrobiota for the island, animals, plants, algae and fungi, from the highest mountain peaks to the limits of diving depths. In the first year of the project, a pilot study targeted a number of taxonomic groups to develop the necessary informatics for data capture, accessibility and connectivity. In doing so, we intend to maximize field efficiency.

These initial efforts to barcode fishes, marine invertebrates, lizards, ferns, and insects helped to identify bottlenecks other ‘All Taxa Biotic Inventories’ will have to overcome for full implementation. Challenges occur at all levels of the process chain from permitting and access, collection and specimen metadata capture, tissue selection and subsampling dynamics, DNA extraction, primer development and marker choice, and linkages to data repositories including museums, BOLD and Genbank. With field collectors working in real-time with informatics specialists and programmers, we have sought to ensure that solutions are user-driven and practical. Infrastructure developed during the MBP is intended to be shared with other biological surveys, All Taxa Biotic Inventories, and utilized by ecologists, and natural resource managers.

Christopher Meyer

Chris Meyer is currently a Research Zoologist at the Smithsonian Institution’s Museum of Natural History. For the past two years he was the Program Manager for the Moorea Biocode Project (MBP) based at the Berkeley Natural History Museums at the University of California, Berkeley. Supported by a grant from the Gordon and Betty Moore Foundation, the Moorea Biocode project is pioneering an integrated approach to biodiversity science that bridges the gap between ecology, genomics, and systematics. A key unifying goal of the MBP is to genetically barcode every species on Moorea.

Dr. Meyer received his Ph.D. in Integrative Biology in 1998 at the University of California at Berkeley where he also received a Masters in Paleontology in 1992. His research generally concerns speciation patterns and processes in marine systems, especially tropical reef communities. To do this, it is important to understand the diversification currency which has led to his interest and involvement in DNA barcoding.

Second International Barcode of Life Conference 18-20 September 2007 20 Conference Abstracts Plenary Session Abstracts

BARCODES BRIDGE THE OLD AND THE NEW: USE OF MUSEUM SPECIMENS TO IDENTIFY MOLECULAR OPERATIONAL TAXONOMIC UNITS IN LARVAE OF SCARAB BEETLES (COLEOPTERA: SCARABAEOIDEA).

ANDREW MITCHELL1, K.A. Rigg1,2, A.J. Campbell1, T.A. Weir3 & A. Raman2

1NSW Department of Primary Industries, Wagga Wagga Agricultural Institute, Private Mail Bag, Wagga Wagga NSW 2650, AUSTRALIA 2Charles Sturt University, Leeds Parade, Orange NSW 2800, AUSTRALIA 3Australian National Insect Collection, CSIRO Entomology, Clunies Ross Street, Black Mountain ACT 2601, AUSTRALIA

Scarab beetles are serious though sporadic agricultural pests in Australia. It is most often the larval stages that inflict most damage, but species identification usually is not possible for scarab larvae as they lack diagnostic morphological characters. We investigate the utility of DNA barcodes for overcoming this problem. We barcoded >200 scarab larvae from across New South Wales (NSW) and identified approximately 30 putative species, or molecular operational taxonomic units (MOTU). In order to identify MOTUs we also barcoded identified adult specimens from museum collections. It has largely been assumed that the DNA of museum specimens more than a few years old will be too degraded for barcoding. We demonstrate that barcodes can be derived from specimens up to 40 years old by PCR amplifying the barcode region in smaller overlapping fragments, although PCR success rates remain low relative to fresh specimens. We verify the utility of combining barcoding of field-collected larvae with targeted barcoding of identified adult beetles, producing species diagnostics and beginning the process of identifying the key scarab pest species in NSW. Linking larvae with identified adults also could facilitate identification of morphological characters that could be used for diagnosis of larvae.

DNA BARCODE, TYPE SPECIMENS AND SPECIES DELIMITATION IN THE GENUS EUMUNIDA

SARAH SAMADI, Nicolas Puillandre, Enrique Macpherson, Josie Lambourdiëre, Marie- Catherine Boisselier.

Muséum National d'Histoire Naturel

The constitution of a DNA barcode dataset on a given set of organisms must cover as well as possible the range of characters variability, the geographic distribution and the most closely related taxa. To link specimens to species names we moreover need to determine the DNA barcode of type specimens. For the genus Eumunida (Decapoda) we obtained COI and 28S data for several specimens from several locations including type specimens for a large part of the described species. We also compared the distribution of morphological characters used in the determination key over the identified genetic clusters. The alpha-taxonomy of the genus was enhanced by bring up one cryptic species but also by revealing polymorphic morphological characters that leaded morphologists to oversplitting. We also revealed that the morphological characters considered as invariable are only invariable if considering only adult specimens. This last point revealed that although identification keys are efficient identification tools with adult specimens, the DNA barcode identification is fare more efficient in the identification of sub-adults or juveniles. This case study reveals that the taxonomic expertise based on DNA barcode would be as much more effective that it will be included in the procedures of an integrative taxonomic research.

Second International Barcode of Life Conference 18-20 September 2007 21 Conference Abstracts Plenary Session Abstracts

Session 4: How close are we to hand-held, field-friendly, or table-top barcoders?

IN VITRO REPAIR ENHANCES AMPLICON RECOVERY AND ACCURACY FROM DAMAGED DNA.

TOM EVANS

New England Biolabs, Inc.

Genetic information is being used to identify species or individuals within a species in the biological and forensic sciences. Despite the theoretical utility, a common practical limitation is that the samples are often degraded; yielding low levels of DNA, damaged DNA, and co-purified PCR inhibitors. Therefore, improved techniques for DNA extraction and DNA repair may permit a broader range of samples to be analyzed. We have optimized a cocktail of 7 enzymes to perform concerted DNA repair in vitro. This cocktail, termed the PreCR Repair Mix, enhanced the yield and accuracy of PCR from DNA templates subjected to UV light, enhanced depurination, oxidation, and heat. As expected, at extremes in damage the repair cocktail was not detectably effective. Importantly, undamaged templates were not adversely affected by the PreCR treatment and no post-repair clean-up was necessary, facilitating the practical application of the cocktail. In vitro DNA repair is complementary to advances in DNA extraction and amplification that together should increase the range of degraded samples that can be analyzed.

EXPRESS BARCODES: RACING FROM BUGS TO IDENTIFICATIONS

NATALIA V. IVANOVA, Paul D.N. Hebert

Canadian Centre for DNA Barcoding, Biodiversity Institute of Ontario, University of Guelph, 579 Gordon Street, Guelph, ON, Canada, N1G 2W1

Some applications of DNA barcoding, such as those involving interceptions of biological materials at ports of entry, are very time sensitive. Although devices combining microfluidic and advanced sequencing technologies promise a future where it will be possible to generate DNA barcodes in minutes, what of the present? In this study, we describe simple protocols that enable the move from organism to barcode-based identification in less than 2 hours. Some elements of our analytical train may aid the future development of a hand-held barcode device. Our protocols use frozen or lyophilized reagents to minimize analytical errors and they can be provided in ‘kits’ that support barcode analysis across the animal kingdom. Our analytical regime allows 5 minutes for DNA extraction, 25 minutes for PCR amplification of the barcode region, 25 minutes for the sequencing reaction, 10 minutes for cleanup, 45 minutes for capillary sequencing and 5 minutes for analysis of the barcode record to generate an identification. Although the race from organism to identification is no sprint, it is already shorter than any marathon.

Second International Barcode of Life Conference 18-20 September 2007 22 Conference Abstracts Plenary Session Abstracts

DEVELOPMENT OF A DNA BARCODE-BASED DNA CHIP FOR IDENTIFICATION OF MARINE ORGANISMS IN THE EAST SEA OF KOREA

YOUN-HO LEE1, Dageum Jeong1, Goeun Kim1, Choongon Kim1, Heungsik Park1, Sejin Pae1, Jinwook Chung2, Hyunkye Yoon2, Seongyong Hwang3

1Korea Ocean Research and Development Institute, Ansan 426-744, 2GenoCheck Inc., Ansan 426-791 3Department of Molecular Biology, Hanyang University, Ansan 426-791, Korea

As a tool for rapid identification of marine organisms, a DNA chip with oligonucleotide probes were developed based on the DNA barcode information. Approximately 650 nucleotide long DNA barcode sequences were obtained from Osteichthyes (Scomber japonicus, Gadus macrocephalus, Stephanolepis cirrhifer, Thamnaconus modestus), Molluscs (Todarodes pacificus, Batillus cornutus, Pomaulax japonicus, Crassostrea nippona, Crassostrea gigas), and Crustaceans (Chionoecetes opilio, Chionoecetes japonica) which are some of the dominant fisheries species in the East Sea of Korea. The sequences were compared with those of congeneric species deposited in the GenBank and species specific nucleotides were identified. Then, 21 to 33 nucleotide long probes were designed, synthesized, and spotted on a glass slide. To test the usefulness of the DNA chip, fragments of the COI gene were amplified from marine samples and hybridized with the probes. Distinct hybridization patterns appeared on the DNA chip dependent on the species. Such DNA chips were also made for Salmonid and Rajiformes fishes and tested useful. As one of the utilizations of the DNA barcode information, the DNA chip method provides a rapid, reliable, and objective measure for identification of marine organisms of interest.

Second International Barcode of Life Conference 18-20 September 2007 23 Conference Abstracts Plenary Session Abstracts

FISH AND CHIPS: MICROARRAY-BASED DNA-BARCODING OF EUROPEAN MARINE FISHES

MARC KOCHZIUS1, Antoniou, A.2, Botla, S.1, Campo Falgueras, D.3, Garcia Vazquez, E.3, Hauschild, J.1, Hervet, C.4, Hjörleifsdottir, S.5, Hreggvidsson, G.O.5, Kappel, K.1, Landi, M.6, Magoulas, A.2, Marteinsson, V.5, Nölte, M.7, Planes, S.4, Seidel, C.1, Silkenbeumer, N.1, Tinti, F.6, Turan, C.8, Weber, H.1, Blohm, D.1

1Centre for Applied Gene Sensor Technology (CAG), University of Bremen, Leobener Strasse UFT, 28359 Bremen, Germany; 2Institute of Marine Biology and Genetics, Hellenic Centre for Marine Research, P.O. Box 2214, 71003 Iraklio, Greece; 3Universidad de Oviedo, C/ Julian Claveria, s/n, 33006 Oviedo, Spain; 4UMR CNRS EPHE 8046, UniversitÈ de Perpignan, 66860 Perpignan Cedex, France; 5Prokaria, Gylfaflöt 5, 112 Reykjavik, Iceland; 6University of Bologna, Via Sant'Alberto 163, 48100 Bologna, Italy; 7Zentrum f¸r Technomathematik (ZeTeM), University of Bremen, 28359 Bremen, Germany; 8Mustafa Kemal University, College of Fisheries and Aquaculture, 31040 Antakya, Turkey

A fast and reliable identification of fishes is important in ichthyoplankton research as well as in seafood and fisheries control. Here, we develop a microarray-based DNA-barcoding technology for the genetic identification of 50 marine European fishes that are ecologically and economically important. Genetic markers are fragments of the mitochondrial cytochrome oxidase I (COI), cytochrome b (cyt b), and 16S rRNA (16S) genes. Oligonucleotide probes have been designed based on 470 COI, 281 cyt b, and 404 16S sequences. The designed probes have been tested in silico for specificity against more than 1000 non-target sequences of the three genes from other marine European fishes. Experiments have shown e.g. clear true-positive signals for the 16S probes in single-target as well as in multiple-target hybridizations. In contrast to conventional Sanger DNA sequencing, microarray hybridizations only require one PCR and purification step, allowing easier handling. These results show that DNA microarrays are a feasible technology that has the potential to make the identification by DNA-barcodes faster, cheaper, more portable, more reliable, and more accessible to non-specialists in the future.

PORTABLE PYROSEQUENCING DEVICE FOR DNA SEQUENCING

MOSTAFA RONAGHI

Stanford Genome Technology Center

We are developing a device for long read Pyrosequencing. The device will enable large scale analysis of DNA amplicons or genome sequencing. The device has integrated a CMOS image sensor with microfluidics enabling a portable solution. We will demonstrate recent results achieved from this integrated device. Challenges and opportunities based on this technique will be reviewed.

Second International Barcode of Life Conference 18-20 September 2007 24 Conference Abstracts Plenary Session Abstracts

Session 5: Analytical methods for barcode data

COMPARING PHYLOGENETIC AND STATISTICAL CLASSIFICATION METHODS FOR DNA BARCODING

FREDERIC AUSTERLITZ1, Olivier David2, Brigitte Schaeffer2, Kevin Bleakley4, Madalina Olteanu2, Raphael Leblois3, Michel Veuille3, Catherine Laredo2

1Laboratoire Ecologie, Systematique et Evolution, U.M.R. C.N.R.S./U.P.S./E.N.G.R.E.F. 8079, Universite Paris-Sud, Batiment 360, F-91405 Orsay cedex, France. 2INRA, Laboratoire de Biometrie, Centre de Recherches de Jouy-en-Josas, 78352 JOUY-EN-JOSAS, France. 3EPHE, Museum National d'Histoire Naturelle, 16 rue Buffon, 75005 Paris, France. 4Institut de Mathematiques et de Modelisation de Montpellier, UMR CNRS 5149, Equipe de Probabilites et Statistique, Universite Montpellier II, CC 051, Place Eugene Bataillon, 34095 Montpellier Cedex 5, France

The barcode is a mitochondrial gene fragment, and we know that mitochondrial DNA evolution is a genealogical process. Does this mean that the identification of species using barcodes must follow a phylogenetic rationale? Or, since we need only to recognize species, irrespective of their phylogeny, is it better to run an efficient classification method on barcode data? The issue is one of efficiency. We compared the ability of genealogical methods (neighbor-joining and PhyML) and of statistical classification methods (CART and Random Forest, Support Vector Machines and Kernel Methods, Neuronal Classification Methods) to correctly identify the species of an individual solely from its barcode. All these methods are implemented using learning data sets, which allow us to develop attuned methods for species identification. To this end, we used both data from empirical studies and simulated mtDNA data sets generated to follow predetermined speciation models. The data sets were partitioned into two subsets: i) a reference sample consisting of individuals already assigned to known species, and ii) query individuals known only from their barcode sequence. Our results show that supervised classification methods perform better than phylogenetic methods when the reference sample per species is large (n≥10) and the number of nucleotide substitutions is low, whereas phylogenetic methods are best in the converse case. Adding information from independent loci (that is, from nuclear loci) increases the power of discrimination of the barcode in some conditions, when several loci are used.

Second International Barcode of Life Conference 18-20 September 2007 25 Conference Abstracts Plenary Session Abstracts

FAST BARCODE-BASED SPECIES IDENTIFICATION USING STRING KERNELS

Pavel P. Kuksa and VLADIMIR PAVLOVIC

Department of Computer Science, Rutgers University, Piscataway, NJ 08854

In this work we study kernel methods for barcode-based species-level identification and propose a framework for fast string algorithms suitable for large and increasingly growing barcode datasets. We present efficient sorting-based algorithms and divide-and-conquer techniques for k-mer kernel computations that improve currently known time bounds for these computations. To further reduce computational cost and facilitate interpretability, we introduce the string kernel problem with feature selection, and present efficient algorithms for it. We also consider string kernels that include spatial information, which show state-of-the-art performance and can easily be evaluated using proposed computational framework. Crucial benefits of the approach are its computational speed and high prediction accuracy for species recognition tasks (error rates of 1-2%). On three datasets, the Astraptes fulgerator, Hesperiidae, and fish larvae, proposed methods considerably improve accuracy compared to prior results. Our experimental results indicate that, for string kernels with mismatches, kernel matrices can be computed 100-200 times faster than traditional approaches. String kernels with feature selection demonstrate competitive classification performance with substantially fewer computations. Our results show that the string kernel-based DNA barcoding can successfully identify specimens by examining only few sequence features, resulting in increased scalability and interpretability of current computational approaches to barcoding.

SPECIES CLASSIFICATION WITH OPTIMIZED LOGIC FORMULAS

Paola Bertolazzi, GIOVANNI FELICI

IASI-CNR, Istituto di Analisi dei Sistemi ed Informatica, Consiglio Nazionale delle Ricerche, viale Manzoni 30 00185 Roma, Italy

DNA Barcode has emerged as a standard for distinguishing species using short, standardized sequences. Barcode is used to assign unidentified specimens to known species, or to better classify species. We study the problem of identifying in a barcode a subset of nucleotides able to classify individuals in one of several species. Such task is related to issues in the analysis of barcode: classification, species discovery, barcode shrinking. We adopt Integer Programming for Feature Selection (FS), and Logic Mining (LM) for Classification. FS is used to identify, in the complete barcode, a subset of nucleotides that carry sufficient information to tell one class from the other. FS is formulated as an optimization problem (Set Covering). Such approach suffers of computational complexity when the data set are too large, so an approximated Set Covering model is adopted, reducing the complexity. The LM tool Lsquare is then used to build a logic system that combines the selected features in compact formulas to explain the differences among individuals of different classes and the similarities among individuals of the same class. We test the model on different data sets and show that the method determines very compact logic formulas for each species and performs well on crossvalidation data.

Second International Barcode of Life Conference 18-20 September 2007 26 Conference Abstracts Plenary Session Abstracts

DNA BARCODE SEQUENCE IDENTIFICATION INCORPORATING TAXONOMIC HIERARCHY AND WITHIN TAXON VARIABILITY

DAMON P. LITTLE

Lewis B. and Dorothy Cullman Program for Molecular Systematics Studies, The New York Botanical Garden, Bronx, New York 10458-5126

For DNA barcoding to succeed as a scientific endeavor an accurate and expeditious query sequence identification method is needed. Although a global alignment can be generated for some barcoding loci (e.g. COI), not all proposed loci are structurally conserved. Thus, algorithms that depend on global alignments are not universally applicable. Sequence identification methods that use local alignments (e.g., BLAST) are unable to accurately differentiate between highly similar sequences and are not designed to cope with hierarchic relationships or within taxon variability. Here, I present a new alignment-free query sequence identification algorithm that accounts for observed within taxon variability and hierarchic relationships between taxa. The method identifies short variable segments and corresponding invariant flanking regions in the reference database. These flanking regions are used to score variable regions in the query sequence without the production of a local or a global alignment. By incorporating observed within taxon variability into the scoring procedure misidentifications arising from shared alleles are minimized. An explicit treatment of the taxonomic hierarchy allows for separate identifications and corresponding measurements of certainly to be made for each level of the hierarchy.

A COMPARISON OF ALGORITHMS FOR SPECIES IDENTIFICATION BASED ON DNA BARCODES

BOGDAN PASANIUC, Alexander Gusev, Sotirios Kentros, James Lindsay, and Ion Mandoiu

Computer Science and Engineering Dept., University of Connecticut

The main use of DNA barcodes is to assign unidentified specimens to species for which barcodes have been already been catalogued. Numerous studies have established DNA barcoding as a standard method for rapid species identification and have lead to the development of large public barcode repositories such as the Barcode of Life Data Systems. As the size of these repositories continues to exponentially increase there is an urgent need for scalable algorithms for species identification capable of handling millions of barcodes, while maintaining high accuracy and providing meaningful confidence measures of their assignments. Several methods for species identification have been proposed in the literature, ranging from using simple distances between barcode sequences to constructing evolutionary trees for these short genomic regions. However, to date there is no agreed upon measure of assignment accuracy and no direct comparison on standardized benchmarks. In this paper we attempt to fill this gap by proposing a principled comparison methodology and performing a comprehensive study of several of the proposed methods, including distance, tree, and statistical model based methods. Besides assessing the accuracy and scalability of individual methods on both simulated and real datasets, we also study the effect that the number of species in the repository and number of sampled specimens per species has on identification accuracy.

Second International Barcode of Life Conference 18-20 September 2007 27 Conference Abstracts Plenary Session Abstracts

Session 6: Case Studies I. Integrative taxonomic studies using DNA barcoding

BARCODING, BIOGEOGRAPHY AND EVOLUTION IN THE TROPICS OF THE NEW WORLD

ELDREDGE BERMINGHAM

Smithsonian Tropical Research Institute, Panamá

I will present the COI barcode results and analysis of one completed study on the avifauna of the Caribbean, and preliminary barcode results on the avifauna and ichthyofauna of Panamá, and the flora of a 50 ha forest dynamics plot in Panamá. My objectives are three-fold. First, we have published a series of articles that explore the evolutionary history of small land birds within the West Indies and now provide COI barcode data to address general principles that govern dispersal, differentiation, and extinction of populations in a geographically structured setting. We chose an island archipelago because islands provide a discrete, replicated geographical reference for the partitioning of variation. Second, we use preliminary COI barcode data for the birds of Panamá to undertake a community phylogenetic comparison between birds in the West Indies and the birds of Panamá. Third, we use preliminary barcode data for the fishes of Panamá and tree species on the 50 ha forest dynamics plot on Barro Colorado Island, Panamá to extend our community phylogenetic analyses and establish a challenge regarding the barcode characterization of regional faunas and floras.

Eldredge Bermingham

Dr. Eldredge Bermingham works at the Smithsonian Tropical Research Institute. His lab carries out studies in the areas of evolutionary biology, genetics and molecular systematics. The lab’s principal research interest is biogeography and history-based analysis of community assembly, diversification and maintenance. They have focused their attention on two regional community assemblages: 1) the birds of the Caribbean; 2) the Neotropical freshwater fishes of lower Mesoamerica. The lab uses the tools of molecular biology to infer the phylogenetic and demographic histories of the taxa comprising these avian and freshwater fish communities. Because they are interested in the tempo as well as the mode of evolutionary change in these communities, the lab also studies the behavior of molecular clocks. Their molecular clock studies have tended to focus on the molecular divergence between marine species pairs found on either side of the Isthmus of Panama.

STRI is part of the Naos Laboratories located near the Pacific entrance of the Panama Canal in the Republic of Panamá. The lab is part of STRI's Molecular Systematics and Evolution Group along with the labs of Haris Lessios, Nancy Knowlton and a visiting scientist’s laboratory.

Bermingham also serves as the regional barcode chair or co-chair for Neotropical fishes (FISH-BOL) and Neotropical birds (ABBI).

Second International Barcode of Life Conference 18-20 September 2007 28 Conference Abstracts Plenary Session Abstracts

THE ROLE OF DNA BARCODING IN THE BROADER CONTEXT OF INTEGRATIVE TAXONOMY

BENOÎT DAYRAT

School of Natural Sciences, University of California, PO box 2039, Merced, CA 95344, USA

Today is a wonderful time for taxonomists! For the first time since the emergence of the ‘population thinking’ in the beginning of the 20th century, something is finally happening in species-level (alpha) taxonomy. Deep discussions about the goals of taxonomy and its place in biological sciences started in the mid 1990s with papers on the ‘taxonomy crisis’ mentioning the lack of funds for alpha-taxonomy and the lack of experts, followed more recently by papers on DNA barcoding. All those discussions make it appear clearly that taxonomy needs to re-invent itself in order to survive, which is a fantastic opportunity but also a challenging responsibility. I recently argued that the future of taxonomy is to become integrative. By ‘Integrative Taxonomy,’ I wish to emphasize that the study of biodiversity units should be based on different kinds of biological data (anatomy, DNA sequences, behavior, etc.). As I view it, the question is not whether taxonomy should become integrative, but how we can make this to happen. In this communication, I shall discuss some of the barriers (e.g., scientific, practical, sociological) to overcome in order to build a 21st-century Integrative Taxonomy, and some aspects of the role of DNA barcoding within this broader context.

Benoît Dayrat

Dr. Benoît Dayrat, Assistant Professor at the University of California, Merced, holds a M.S. (1996) in Molecular Biology from the Ecole Normale Supérieure, Paris, and a Ph.D. (2000) in Systematics and Evolution from the University of Paris VII (his research actually took place at the Muséum National d’Histoire Naturelle). He spent several years at the California Academy of Sciences, funded as a postdoctoral fellow through the NSF PEET (Partnership for Enhancing Expertise in Taxonomy) program. Dr. Dayrat’s post-doctoral experience yielded, among other publications, a Taxonomic monograph on discodorid sea slugs (450 pp., 370 plates).

Dr. Dayrat, a specialist of gastropod systematics and evolution, is the author of empirical systematic studies, contributions on the history and philosophy of evolutionary and systematic biology, and a book on the history of the discovery of the French Flora (2003). Dr. Dayrat attempts to use his empirical studies to address issues of general interest in taxonomy: e.g., he participated in the development of the ‘Species Article’ recently added to the PhyloCode; he also proposed a series of practical guidelines that could help moving towards what he defined as “Integrative Taxonomy.”

Second International Barcode of Life Conference 18-20 September 2007 29 Conference Abstracts Plenary Session Abstracts

SPHINGIDS AND BARCODES - THE NEW TAXONOMY

RODOLPHE ROUGERIE1, Jean Haxaire2, Ian J. Kitching3, Thierry Vaglia4, Paul D. N. Hebert1

1Canadian Centre for DNA Barcoding, Biodiversity Institute of Ontario, University of Guelph, 50 Stone Rd., Guelph, Ontario, N1G2W1 Canada 2Le Roc, F-47310 Laplume, France 3The Natural History Museum, Department of Entomology, Cromwell Road, London SW7 5BD, U.K. 4287, 62eme Avenue, Chomedey, Quebec, H7V2G4, Canada

Projects which seek planetary coverage for particular taxonomic groups provide an important way to advance the All-Life barcoding initiative. This presentation focuses on a global barcode survey on the Sphingidae (Hawkmoths), a family of Lepidoptera with some 1350 recognized species. Although work was activated just a year ago, a comprehensive barcode library now exists for more than 90% of these species including images, collection data, sequences and trace files. This rapid progress reflects the involvement of a broadly international team of taxonomists who provided critically identified specimens, and who aided in the interpretation of barcode data. This combination of taxonomic expertise with the overview of sequence diversity delivered by barcoding proved a powerful tool in revealing overlooked species, hybrid taxa and cases of synonymy. We believe that the sphingid campaign serves as a good model for a new taxonomy whose primary deliverables include (1) a global capacity to identify specimens (2) accelerated biodiversity exploration and species descriptions and (3) a foundation for genomic, phylogeographic, and ecological studies. The present study shows further that this new taxonomy can be delivered with modest incremental support when there is a standing taxonomic capacity. Where this is not the case, it will need to be created.

Rodolphe Rougerie

Dr. Rodolphe Rougerie is the supervisor of two global barcoding campaigns started on August 2006 and aiming at assembling barcodes and associated data for the world species of two families of moths – the Sphingidae and Saturniidae – representing about 4000 species. The resulting reference database is built in close collaboration with the community of expert taxonomists. This project is funded by a NSERC Discovery grant and by the Gordon and Betty Moore Foundation. Rodolphe completed his PhD in Systematics and Evolution in October 2005 at the National Natural History Museum of Paris under the supervision of Prof. Joel Minet. His doctoral research focused on the phylogeny and biogeographical history of the saturniid moths (wild silkmoths), using both morphology and DNA data. Rodolphe's postgraduate research diploma, completed in October 2000, dealt with microevolution in Collembola, chiefly hybrid zones structure and related taxonomic issues. He has been passionate about collecting and rearing insects since childhood and has specialized in the taxonomy of saturniid moths as well as in the study of their life-history. Rodolphe also enjoys communicating science to the public, and in spring 2006 published a book for kids on insects.

Second International Barcode of Life Conference 18-20 September 2007 30 Conference Abstracts Plenary Session Abstracts

BAMBOOZLED BY BLOODSUCKERS: BARCODING BACKS BIODIVERSITY

MARK E. SIDDALL

American Museum of Natural History

European medicinal leeches have direct application in modern surgical medicine. Dozens of protease inhibitors purified from leech saliva hold the promise of therapeutic application and these leeches are widely used in flap and replantation surgery. Hirudo medicinalis has been approved by the United States Food and Drug Administration as a prescription medical device; a designation requiring proper labelling. Similarly, accurate annotation of bioactive compounds relies on precise species determination. A a model organism in neurobiology, nterpretations of developmental and neurophysiological characteristics also presuppose uniformity in laboratory settings. DNA Barcoding indicates that that there are at least three species of European medicinal leech, and that leeches marketed as H. medicinalis are actually Hirudo verbana. These results are corroborated by nuclear microsatellite markers. Beyond the obvious need for reconsideration of decades of biomedical research on this widely used model organism, these findings impact regulatory statutes and raise concerns for the conservation status of European medicinal leeches.

Mark E. Siddall

Dr. Mark Siddall is Curator of Invertebrates at the American Museum of natural history in New York City. His work on the biodiversity and evolution of annelid worms has been funded in part by the Richard Lounsbery Foundation, particularly as it relates to barcoding medicinal leeches.

Dr. Siddall began his scientific career in protozoon parasitology, earning his PhD from the University of Toronto in 1994. He arrived at AMNH after a postdoctoral period at the College of William and Mary and an Assistant Professorship at the University of Michigan.

Dr. Siddall's systematic work, funded mostly through PEET and REVSYS grants from the US National Science Foundation, has included extensive field work covering all continents where he and his students gather leeches by wading through swamps and trekking through damp forests to attract their specimens. As well, his lab is actively engaged in determining the relationships of various endosymbiotic microbes found in association with invertebrates. Dr. Siddall is curator of the Hall of Ocean Life at AMNH, is a recipient of the Henry Baldwin Ward Medal from the American Society of Parasitologists, is Treasurer of the Willi Hennig Society and serves in several journal editorial roles pertaining to systematics of invertebrates and parasites.

Second International Barcode of Life Conference 18-20 September 2007 31 Conference Abstracts Plenary Session Abstracts

Session 7: What new science will we see at the third barcode conference?

BARCODING AND BIODIVERSITY RESEARCH

LES CHRISTIDIS

Australian Museum

The use of DNA data in determining taxonomic identifications and establishing phylogenetic relationships is well established. The DNA barcode approach to use a small section of DNA as a “species identifier” has generated much controversy, much of it centring on examples where the Barcode DNA region has failed to identify “established species”. Given that in biology no method is 100% accurate and that some of these “established species” may not be separate species but rather morphological variants, this type of debate is deflecting attention from more important issues. These include whether the Barcode approach should be largely aimed at making it easier to identify described species, to identify new biodiversity, or a combination of the two. It is acknowledged that biodiversity is disappearing before it is described and that Barcoding is one way of speeding up identification of new species. How useful is a DNA Barcode if it is not linked to a morphological description? The importance of taxonomic expertise is implicit in most DNA Barcode projects and yet there is a shortage of researchers willing to pursue careers in taxonomy, due in part to lack of employment opportunities. Can Barcoding provide an avenue to encourage new participants into taxonomy, and if so how?

Les Christidis

Dr Les Christidis has been the Assistant Director of the Australian Museum in Sydney and Head of the Division of Research and Collections since 2004. The Division includes over 120 staff and students and undertakes research and collection development in zoology, geology, palaeontology, anthropology, molecular biology, ecology and materials conservation.

Dr Christidis undertook a PhD at the Australian National University from 1981 to 1984 where he focussed on the evolutionary genetics of Australian finches. He went on to conduct research on the origins of Australia’s songbirds, first as a CSIRO Post-doctoral Fellow and then as the recipient of a Queen Elizabeth II Fellowship. From 1987 to 1996 he was Senior Curator of Ornithology at Museum Victoria and in 1997 he became the Science Program Director. As Program Director (1997 to 2000) he led the teams that developed four major exhibitions for Melbourne Museum and Australia’s first Digital Planetarium. From 2001 to 2004 he was Head of the newly created Department of Sciences.

Dr Christidis has maintained an active research career, has written over 90 scientific papers, including 3 books, and supervised numerous graduate students. He is an Associate Professor at two universities and sits on several national and international committees.

Second International Barcode of Life Conference 18-20 September 2007 32 Conference Abstracts Plenary Session Abstracts

ECOLOGICAL INVENTORY WITH A BARCORDER, THE ECOLOGICAL TAXASCOPE: WHO EATS WHAT IN A COMPLEX TROPICAL FOREST?

DANIEL H. JANZEN1, Winnie Hallwachs1, Mehrdad Hajibabaei2, John M. Burns3, Alex Smith2, Josephine Rodriguez4, Monty Wood5, Tanya Dapkey1, Paul Hebert2

1Department of Biology, University of Pennsylvania, Philadelphia, PA 19104 2Biodiversity Institute of Ontario, University of Guelph, Guelph, Ontario N1G 2W1 3Department of Entomology, Smithsonian Institution, Washington, D.C. 20560 4Department of Entomology, University of Illinois, Urbana, IL 61801 5Diptera Unit, Canadian National Collection of Insects, Agriculture and Agrifood Canada Ottawa, Ontario K1A OC6, Canada

Twenty-nine years of biodiversity inventory of the caterpillars, and their parasitoids and foodplants, of the dry forest, rain forest and cloud forest of Area de Conservacion Guanacaste (ACG), northwestern Costa Rica, has yielded more than 100,000 dried and ETOH-preserved voucher specimens of more than 5,000 insect species, a very substantial number of which are undescribed, very similar, or otherwise difficult to identify. The past four years of DNA barcoding more than 60,000 specimens from this complex and taxonomically murky package has demonstrated unambiguously that at least for one very complex tropical site - 85 X 30 km in area and 0-1500 m elevation - DNA barcoding serves extremely well in species-level identification of specimens and in exposing overlooked species, a process that is corroborated by matching DNA barcodes with parasitoid and food plant records, and with microgeographic and ecological distributions. The resultant taxonomic clarification and species discovery has greatly increased the accuracy of statements about the trophic structure of plant-caterpillar-parasitoid food webs in ACG, by revealing and confirming very tight interactions among species, and between species and their physical environment. Furthermore, the first steps have been taken towards largely machine-based identification of any ACG species and the known ACG species-richness has been significantly increased.

Daniel Janzen

Daniel Janzen is the DiMaura Professor of Conservation Biology at the University of Pennsylvania, Philadelphia, and the technical advisor to Area de Conservación Gaunacaste, in northwestern Costa Rica.

While focused on tropical animal-plant relationships (1963-1985), Janzen has more recently conducted an inventory of tropical caterpillars, their parasites, and their food plants, and on the conservation of tropical biodiversity through its non-damaging development (see ). His 428 publications encapsulate much of this. He and his biologist wife, Winnie Hallwachs, are architects of Area de Conservación Guanacaste (ACG) in northwestern Costa Rica (http://www.acguanacaste.ac.cr), decreed a UNESCO World Heritage Site in 1999. Janzen received the first Crafoord Prize in biology from the Swedish Royal Academy of Science (1984), and the Kyoto Prize in Basic Biology (1997). A member of the US National Academy of Sciences (1992) and honorary member of the Costa Rican National Park Service, he promotes the relevance and potential of conservation of tropical wildland biodiversity for global understanding, national sustainable development, and individual quality of life. His current focus is finding the funds to endow the entire national park system of Costa Rica, and facilitating global bioliteracy through the emergence of the ability by all people to be able to identify any organism anywhere anytime through DNA barcoding.

WHAT NEW RESEARCH CAN WE DO WITH A MILLION BARCODE RECORDS?

Second International Barcode of Life Conference 18-20 September 2007 33 Conference Abstracts Plenary Session Abstracts

PAUL DE BARRO

CSIRO, Australia

One big science question where 1 million barcodes code make a considerable difference is the generation of fundamental advances in our understanding of the relationship between biodiversity and ecosystem function (BD-EF). By extending the existing BD-EF paradigm to explore changes in diversity across multiple trophic levels and incorporating realistic levels diversity and complexity, we can consider the processes of community assembly and changes in species diversity and abundance. In so doing we can directly link this to the concept of ecosystem resilience. The application of barcodes using high throughput technology will enable the exploration of this on a scale that involves replication through both space and time. One can imagine that through this application of barcodes, different patches of ecosystems could be compared over time and an index applied which measures changes in the relative proportions of indigenous and exotic species. Over time this will enable different habitats to be monitored with changes in the number and proportion of different haplotypes providing an index of their health in response to factors such as global climate change and biological invasion. This data can then be used to provide a more rigorous underpinning to our understanding of relative ecosystem vulnerability.

Paul J. De Barro

Dr. Paul De Barro is a member of the Science Advisory Board of the Consortium for the Barcode of Life (CBOL). He trained as an entomologist at the University of Adelaide where he completed a BAgSc (Hons) in 1985 before working for three years with the South Australian Department of Primary Industries and then the Bureau of Sugar Experiment Stations in Queensland. In late 1988 he returned to the University of Adelaide to complete his PhD and was subsequently awarded the University’s George Murray Fellowship which he used this to undertake a postdoctoral fellowship at the University of Southampton. Here he contributed to the development of the use of DNA fingerprinting as tool for studying insect ecology. In 1994 he moved to Canberra, Australia to begin his career with CSIRO Entomology focusing on a range of research projects involving the invasive species Bemisia tabaci. Tired of the rather chilling Canberra winters, Dr. De Barro relocated to CSIRO’s laboratories in tropical Brisbane, Queensland in 1999 to continue his research on invasive insect species. Dr. De Barro is currently the Stream Leader for Preparedness as well as the Program Leader for the Cooperative Research Centre for National Plant Biosecurity.

Second International Barcode of Life Conference 18-20 September 2007 34 Conference Abstracts Plenary Session Abstracts

WHAT NEW RESEARCH CAN WE DO WITH A MILLION BARCODE RECORDS?

PETER NG

National University of Singapore

A million things! Species and genera boundaries can be tested. The barcode datasets can be integrated with morphology, and an even better understanding of life on Earth can be elucidated – not just how many species and what they are – but how they came about, their biogeography etc. Data is always valuable – and the more data we have – the better we understand our natural world.

Peter K. L. Ng

Dr. Peter Ng is a member of the Scientific Advisory Board (SAB) of the Barcode of Life (CBOL), an international initiative hosted by the Smithsonian Institution’s Museum of Natural History and supported by a grant from the Sloan Foundation. Dr. Ng is currently a professor of biological sciences at the Department of Biological Sciences at the National University of Singapore; and until recently, was head of the biodiversity program there. He is also the director of the Raffles Museum of Biodiversity Research, a faculty institute who holds one of the largest and oldest collections of animals in Singapore, Malaysia and SE Asia, dating back over 150 years. He is also a member of the International Commission for Zoological Nomenclature which oversees all nomenclatural problems in zoology. Dr. Ng’s forte is the systematics and ecology of crabs and other decapod crustaceans, and has published widely on them. He has also been active in freshwater fish systematics, deep sea diversity, conservation biology; and broader interests in molecular systematics and education. He sits on the editorial boards of some two dozen journals and has been actively involved in many regional and international projects involving biodiversity surveys and systematics.

Second International Barcode of Life Conference 18-20 September 2007 35 Conference Abstracts Plenary Session Abstracts

WHAT NEW RESEARCH CAN WE DO WITH A MILLION BARCODE RECORDS?

AXEL MEYER

Department of Biology, University of Konstanz, Germany

The answer to this question depends on several factors among those are: the extent and completeness of taxon sampling and which molecular marker(s) are used. Since the advent of PCR, about 20 years ago, and the design of “universal” primers molecular systematics and molecular taxonomy were, for the first time, based on a universal metric – i.e. orthologous DNA sequences – that could be collected with technical ease and across a wide taxonomic range. This facilitated a revolution in evolutionary biology and allowed to conduct sequence comparisons and analyses of tempo and mode of evolution of molecules and evolutionary lineages. Allan Wilson called this development the “democratization of the genetic code”. The power that derived from these events can be illustrated by the fact that the first paper to report universal primers (Kocher et al. 1989 PNAS) has been cited over 2000 times and mitochondrial control region, cytochrome b, 12SrRNA and 16SrRNA gene fragments – for which the first universal primers were available - have been determined hundreds of thousands of times. While these universal metrics have been immensely useful, no molecular marker will provide the best answers to all questions (Meyer 1995 TREE). This will apply to COI sequences as well. While for many taxa this gene offers enough variation to be taxonomically informative for others (e.g., East African cichlid fish) it is going to be too slowly evolving or too short to be able to distinguish extremely young species. For this particular question, and presumably others that involve radiations of extremely young taxa, additional marker systems will have to be developed to reach a resolution that approaches barcoding quality. The analysis of correlations and possibly cause-effect relationships of molecular and phenotypic diversification is one potential novel direction of research that will be facilitated by millions of barcode records.

Axel Meyer

Axel Meyer received his BA Degree from the University of Marburg in 1982, and studied for one more semester at the University of Kiel in Germany before he continued his education the US. Sponsored by a Fulbright Fellowship he first studied for one semester at the University of Miami before he moved to the University of California at Berkeley in 1983. He received his M.A. in 1984 and his Ph.D. in 1988 from the Zoology Department at Berkeley. The academic year 1986/7 he spent at the OEB Department at Harvard University. Funded by a fellowship from the Sloan Foundation in molecular evolution he was a postdoc in the lab of Allan C. Wilson in the Biochemistry Department at Berkeley from 1988 to 1990. He then moved to the Department of Ecology and Evolution at Stony Brook where he was an Assistant Professor from 1990- 93 and Associate Professor from 1993 to 1997 until he returned to Germany where he currently holds a Professorship in Zoology and Evolutionary Biology at the University of Konstanz. Most of the research focuses on the evolution of adaptive radiations of cichlid fishes in Eastern Africa, but we have also conducted projects on many other (mostly) vertebrate taxa, dealing with molecular phylogenetic, phylogeographic and population genetic issues. Several current projects deal the genetics and genomics of speciation and adaptation.

Second International Barcode of Life Conference 18-20 September 2007 36 Conference Abstracts Plenary Session Abstracts

WHAT NEW RESEARCH CAN WE DO WITH A MILLION BARCODE RECORDS?

ROBERT A. ANDERSEN

Provasoli-Guillard National Center for Culture of Marine Phytoplankton, Bigelow Laboratory for Ocean Sciences, West Boothbay Harbor, Maine USA

Protists are often difficult to identify - single cells have limited morphology compared to vascular plants, insects or mammals. We cannot conduct rapid surveys of protists because of their small size, cryptic morphologies, and incredible cell numbers. A liter of seawater may have one million protistan cells, which equal one million individuals representing many species. A flower or an insect may have one million cells but there is only one individual. Now, barcodes and 454 pyrosequencing offer the possibility of accurately identifying large numbers of individual organisms from field samples, e.g., plankton tows, benthic mats or seaweed collections. This is currently impossible with traditional methods. Taxonomic, ecological and biogeographical studies will increase rapidly using protistan barcodes. Some protists cause diseases (e.g., malaria, sleeping sickness, toxoplasmosis), with malaria killing more than 1 million humans annually. New, rapid detection in humans, vectors and water can be achieved with protist barcodes. Parasitologists claim that every insect species has at least one sporozoan parasite that is unique to that insect species, implying huge cryptic diversity, but it is impossible to thoroughly test this with today’s techniques. Barcodes are powerful tools for taxonomic studies, and conservation of our natural world depends upon this taxonomy.

Robert A. Andersen

Dr. Robert Andersen is the Director of the Provasoli-Guillard National Center for Culture of Marine Phytoplankton, the world’s largest culture collection of marine algae. The collection maintains approximately 2500 strains from around the world, and it annually distributes about 2500 strains worldwide. The collection is part of Bigelow Laboratory for Ocean Sciences, located in Boothbay Harbor, Maine, USA.

Dr. Andersen was trained as a botanist specializing in algal research. He holds a B.S. in Botany from North Dakota State University, a M.S. in Aquatic Biology from St. Cloud State College, and a Ph.D. in Botany (Phycology) from the University of Arkansas. Dr. Andersen was a faculty member at DePaul University from 1980 to 1989. During a sabbatical at Melbourne University, Australia, during 1988, he became interested in marine phytoplankton, and in 1989 he accepted his current position as Director of the CCMP. He has carried out research as a visiting scientist at Bristol University, U.K. and the National Institute of Environmental Studies, Japan. Dr. Andersen has served as the President of the International Society of Evolutionary Protistology and Secretary of the International Phycological Society; he is currently the Vice President – President-elect of the Phycological Society of America.

Second International Barcode of Life Conference 18-20 September 2007 37 Conference Abstracts Plenary Session Abstracts

Session 8: Case Studies II. DNA barcoding in developing countries

DNA BARCODING: CONSIDERATIONS FOR VECTORS OF NEGLECTED DISEASES

DANIEL MASIGA1, Johnson Ouma2

1Molecular Biology and Biotechnology Department, ICIPE, P.O. Box 30772, Nairobi 00100 2Trypanosomiasis Research Centre, Kenya Agricultural Research Institute, P.O. Box 362, Kikuyu 00902.

The study of the broad field of biology relies extensively on the ability to classify and identify individuals to the lowest taxonomic level. Although arthropod vectors of human and animal diseases are an enduring threat to health and productivity, their taxonomy is limited to morphological descriptions, often in regions where such expertise is limited. This is unfortunate, because good quality studies on vector ecology, disease epidemiology, predicting potential success of control methods that are based on vector ecology and behaviors, such as trapping, are compromised. Kenya has many vectors of pathogens, which include mosquitoes that transmit various pathogens (viruses, worms and plasmodium), tsetse flies (Diptera: Glossinidae; vectors of African trypanosomes) and phlebotomine sand flies, which are vectors of leishmaniasis. Our focus in this initial stage of the barcoding activities in Kenya is on the genus Glossina, species of which transmit sleeping sickness (or Human African Trypanosomiasis), and nagana in livestock. Eight species and sub-species are recognized in the 3 sub-genera of tsetse flies that have been identified in Kenya. We sought to generate barcodes to aid taxonomy and investigate the integrity of the current taxonomic status among members of this genus. Using universal primers described by Folmer et al (1994), barcodes were obtained from all 8 taxa of tsetse (G. fuscipes fuscipes, G. pallidipes, G. swynnetoni, G. morsitans morsitans, G. m. centralis, G. m. submorsitans, G. brevipalpis and G. longipennis). This marks the beginning of a barcoding effort for all species of Glossina in Africa, the only continent where this genus is found. We also examine the potential application of DNA barcoding to studies of the epidemiology of trypanosomes, which are transmitted by tsetse flies.

Daniel Masiga

Dr. Daniel Masiga is a Research Scientist and head of the Molecular Biology and Biotechnology Department at the International Centre of Insect Physiology and Ecology (icipe) in Kenya. The institute carries out research on a broad range of insects, including disease vectors (medical veterinary and plant), pests and insects of commercial interest, such as pollinators, honeybees and silk moths. Dr. Masiga has a special interest in vectors and the pathogens they transmit. Dr. Masiga studied Biochemistry and Zoology at the University of Nairobi, before undertaking an MSc in Biochemistry at University College London. He obtained a PhD in Molecular Parasitology at the University of Bristol, UK in 1994, where his focus was on developing methodologies for species identification of African trypanosomes. He then worked at the Kenya Trypanosomiasis Research Institute in the field of trypanosome genetics and epidemiology. Between 1997 and 2000, he was a Postdoctoral Research Fellowship in trypanosome genetics at Glasgow University, before retuning to Kenya, where he worked as a Senior Lecturer at Kenyatta University, prior to joining icipe. He is a member of the Scientific Advisory Board of the Consortium for the Barcode of Life (CBOL).

Second International Barcode of Life Conference 18-20 September 2007 38 Conference Abstracts Plenary Session Abstracts

ALL BIRDS BARCODING INITIATIVE IN THE NEOTROPICS: IDENTIFYING AND DISCOVERING BIRD SPECIES IN THE RICHEST AVIFAUNA OF THE WORLD

PABLO TUBARO

MACN, Buenos Aires, Argentina

About a third of the known species of birds live in the Neotropical Region (4069 species with 3121 endemics) and several more species are probably waiting to be described. This set includes extensive radiations of both non-passerine and passerine families where plumage similarity in some cases and non-concordant patterns of trait variation in others cause problems in species identification. Thus, Neotropical avifauna provides a particularly interesting ground to test the effectiveness of DNA barcoding. Up to now, progress in the barcoding of the birds of the Neotropics includes most of the species in the Caribbean region and Panama, and more than 50% of the species from Argentina. Other efforts in the region include the barcode of the Psittaciforms, Formicariids, Tyrannids and Charadriiforms. Based on the results from the birds of Argentina, interespecific levels of divergence in COI sequence are similar to those of the birds in North America. In 3% of the cases we also found significant levels of intraspecific divergence in COI sequences suggesting the possible existence of cryptic species or geographical races that deserve a species status.

Pablo Tubaro

Pablo L. Tubaro is an Independent Researcher at the National Research Council of Argentina (CONICET), Curator of Birds and Vice-director of the National Museum of Natural Sciences of that country. He received his Ph.D. in Biology from the University of Buenos Aires in 1990. Since 2005 he has been a member of the Scientific Advisory Board of the Consortium for the Barcode of Life and has been appointed as Chair of the Steering Committee of the All Birds Barcoding Initiative in 2006. He has taught behavioral ecology, systematics and evolution of vertebrates at the University of Buenos Aires for more than 15 years. His main research interest deals with the evolutionary biology of birds including: 1) systematics and phylogeography of Neotropical birds, 2) hybridization and speciation, and 3) comparative studies about the evolution of morphological and behavioral traits.

Second International Barcode of Life Conference 18-20 September 2007 39 Conference Abstracts Plenary Session Abstracts

FISH-BOL AND BARCODING IN INDIA

WAZIR S. LAKRA

National Bureau of Fish Genetic Resources, Indian Council of Agricultural Research, Canal Ring Road, P.O. Dilkusha, Lucknow-226002, India

DNA Barcoding is a new science technique and global standard for the identification of biological species using a short gene sequence from standardized position in the genome Dr. Paul Hebert and his colleagues at University of Guelph in Ontario, Canada first proposed and initiated DNA Barcoding of life forms based on mitochondrial cytochrome c oxidase I (CO1) in 2003. Consequently, several workers have evaluated the effectiveness of DNA barcodes in fish, insects, birds and some plant species from America, Europe and Asia. In this endeavor, the Consortium for Barcode of Life (CBOL) has played a pivotal role towards the promotion of efforts and networking. In 2005, CBOL launched the Fish Barcode of Life campaign (FISH-BOL) to create a global reference library of all species of fish. India ranks as eighth mega biodiversity centre of the world and harbours around 2200 finfish species with taxonomic ambiguity reported from several genera and groups. The bounty of marine biodiversity which is exploited from 2.02 million square kilometer of the Exclusive Economic Zone (EEZ) of India constitutes one of the largest heritage resources. The National Bureau of Fish Genetic Resources of the Indian Council of Agricultural Research at Lucknow initiated a national program on DNA Barcoding of Fish in 2005 as first barcoding project in the country. The Institute has emerged as a Lead Centre for South Asia in terms of HRD and generation of data for barcoding of aquatic species of the region. In addition, India has recently initiated efforts to barcode butterflies, anurans and some important taxa of plants.

Wazir S. Lakra

Dr. Wazir S. Lakra is the Director of the National Bureau of Fish Genetic Resources (NBFGR), a premier Institute of the Indian Council of Agricultural Research at Lucknow, India. The NBFGR has state-of-art facilities and a national mandate to catalogue, classify and conserve the aquatic genetic resources of the country.

Dr. Lakra holds a Masters Degree in Zoology and a Ph.D. in Fish Biology. He was Professor and Chair, Department of Fish Genetics and Biotechnology at Central Institute of Fisheries Education, Mumbai from 1998-2005 and guided research in fish genetics, biodiversity and biotechnology. He has published over 100 research papers and 20 books and bulletins and been conferred several awards for his contributions to Fishery Science Dr. Lakra was trained at CSIRO Marine Research, Hobart, Australia in Fish Molecular Genetics as FAO Fellow and is the Regional Chair of FISH-BOL for South Asia.

Second International Barcode of Life Conference 18-20 September 2007 40 Conference Abstracts Plenary Session Abstracts

TRANSFORMING TAXONOMY FOR EFFECTIVE BIODIVERSITY ASSESSMENT OF ARTHROPODS IN MADAGASCAR.

BRIAN L. FISHER1, M. Alex Smith2,

1Department of Entomology, California Academy of Sciences 2Biodiversity Institute of Ontario, University of Guelph

Madagascar has received immense interest in the arthropod taxonomic community over the last 20 years. Despite the enormous efforts to inventory the arthropods on the island, there have been few taxonomic products. Millions of specimens have been collected but relatively few species have been described. The slowness of the current taxonomic process has had a huge impact in the role insects play in conservation. Madagascar’s President, Marc Ravalomanana, has formally committed his government to a massive expansion of the protected areas network, more than tripling the total area. Several million hectares will be designated for inclusion in this new Madagascar System of Protected Areas over the next few years. Unfortunately, insects are not a key player in the analysis of priority areas. If conservation planning efforts focus only on well- known and relatively species-poor taxa, then we risk overlooking significant ecological and evolutionary components of biodiversity. If monitoring efforts overlook insects, we will fail to hear the early warning signs from these canaries in the coal mine. We evaluate the use COI barcoding to accelerate both the delimitation and the identification of ants across the island in a study that includes over 6,000 specimens sequenced.

Second International Barcode of Life Conference 18-20 September 2007 41 Conference Abstracts Plenary Session Abstracts

CONSTRUCTION OF A DNA BARCODE SYSTEM FOR AMPHIBIANS AND REPTILES IN TAIWAN

SI-MIN LIN1, Hsin-Yun Chao1, Hsuan-Yi Ho1, Ying-Rong Chen1, and Kuang-Yang Lue2

1Department of Life Science, Chinese Culture University 2 Department of Life Science, National Taiwan Normal University

Although highly organized and efficient barcoding proposals are now in progress in the field of ichthyology and ornithology, much less efforts are paid in constructing a DNA barcode system for amphibians and reptiles. In the last two years, our laboratory has been dedicated to constructing a DNA barcode system for amphibians and reptiles in Taiwan. COI sequences have been obtained from more than 85% of native species. Considerable divergences are observed among different species, indicating COI sequence as a suitable marker for species recognition. However, phylogenetic trees revealed obvious contradiction to traditional taxonomy, showing unreliability of these sequences in phylogenetic reconstruction. Saturation in point mutations might be the main reason for such unreliability: as in the case of the snakes, divergence between species within the same genus reaches to 0.20, relevant to that of inter-genera or inter-family levels. As in the case of the lizards, the average inter-family divergence reaches to 0.34. Such high divergence caused operational difficulties in primer design and PCR amplification. Obviously, a large proportion of species could not be amplified by so-called “university primers”. In a case study from a Sphenomorphus skink, COI sequences showed extremely high intra-specific variation up to 12%. Owing to their sympatric distribution and with no diagnostic morphological character, we are not able to do further taxonomic treatments in this case. On the other hand, cryptic species, such as two Lacertid lizards (Takydromus spp.), and two Hynobiids (Hynobius spp.), were discovered through DNA barcodes. These results showed that the biodiversity of amphibians and reptiles in Taiwan is still under estimated. Finally, we sincerely call for a better organized activity in international cooperation on DNA barcoding for amphibians and reptiles.

Second International Barcode of Life Conference 18-20 September 2007 42 Conference Abstracts Plenary Session Abstracts

Session 9: Barcoding in the wider evolutionary context

BARCODING AND THE TREE OF LIFE

MICHAEL J. DONOGHUE

Peabody Museum of Natural History, Yale University, New Haven, Connecticut, USA.

These two international mega-science projects should be better coordinated, both to avoid duplication and the re-collection of crucial biological materials, but also to capitalize on possible synergies. As a united front, adopting a common ethical framework, we will make greater progress in obtaining the necessary materials for study. Efforts should be undertaken immediately to share specimens and to store tissues and DNA extracts for long-term use by both communities. Likewise, databasing efforts should be coordinated to ensure seamless integration across projects. Such connectivity will enhance the ability to quickly identify unknowns, but will also allow new scientific research at the intersection of micro- and macro-evolutionary biology. As research progresses in both realms there may be important feedbacks and adjustments. For example, a special concern evident in phylogenetic studies of some organisms is the pervasiveness of recent hybridization. In such cases accurate identifications may require the use of markers drawn from several genomes.

Michael J. Donoghue

Michael Donoghue joined Yale University in 2000 as the G. Evelyn Hutchinson Professor of Ecology and Evolutionary Biology. He served as Chair of the Ecology and Evolutionary Biology Department in 2001-02, and was appointed Director of the Peabody Museum of Natural History in January 2003. Professor Donoghue earned his undergraduate degree from Michigan State University in 1976 and his Ph.D. in Biology from Harvard University in 1982. He served on the faculty of San Diego State University (1982-1985), the University of Arizona (1985-1992), and Harvard University (1992-2000), and as a Visiting Professor at Stanford University (1998-1999). He was the Director of the Harvard University Herbaria from 1995-1999.

Professor Donoghue was elected a Fellow of the American Association for the Advancement of Science in 1997, and as a member of the U. S. the National Academy of Sciences in 2005. His research has focused on understanding the diversity and evolution of flowering plants, phylogenetic systematics, and historical biogeography. He has published over 170 papers, co-authored two books on plant diversity, and served as a mentor to 23 postdoctoral associates and 20 graduate students.

Second International Barcode of Life Conference 18-20 September 2007 43 Conference Abstracts Plenary Session Abstracts

EVOLUTIONARY HISTORY OF FISHES: AN OVERVIEW BASED ON >1000 WHOLE MITOCHONDRIAL GENOME SEQUENCES

MASAKI MIYA1 and Mutsumi Nishida2

1 Natural History Museum and Institute, Chiba 2 Ocean Research Institute, University of Tokyo

The COI barcoding is a standardized approach to identifying species by DNA, having helped resolve the “leaves” on the tree of life. The mitochondrial genome (mitogenome) project in fishes, on the other hand, is a phylogenomic approach to resolving the “branches” of the tree based on exhaustive sampling from higher taxa at the levels of orders, families, and possibly genera. Such comprehensive character and taxon samplings have become feasible through the development of a PCR-based approach for sequencing whole mitogenomes using many fish-versatile primers. With this technique, we have determined mitogenomic sequences for over >1000 species of fishes and published many novel, well supported, but previously unanticipated phylogenies. Correct estimation of the current diversity and how it has been attained through a long history of fishes are both complementary and can be achieved through exhaustive taxon samplings (at the different levels) in the DNA barcoding and mitogenomic approaches, respectively.

Masaki Miya

Dr. Masaki Miya is Curator of Fishes at Natural History Museum and Institute, Chiba, and Adjunct Associate Professor at Graduate School, Chiba University. Dr. Miya was trained as a marine biologist and holds a Ph.D. in Fisheries Sciences from The University of Tokyo. He started his career as an ecologist of deep-sea fishes, investigating life histories and vertical distributions of Cyclothone, numerically most abundant vertebrate in the world. Comparisons of ecological traits of these fishes have aroused his interests in phylogenetics and evolutionary biology. Subsequently he acquired techniques in molecular biology in Prof. Mutsumi Nishida’s lab and developed a PCR-based approach for sequencing whole mitochondrial genome (mitogenome) sequences (ca. 16,500 bp) . Using this technique, Dr. Miya’s research group has determined the mitogenome sequences for > 950 species that encompass a whole spectrum of fish diversity and published 54 papers on phylogeny and evolution of various fish groups during 1999–2007. Currently his research is focused on large-scale analyses of evolution in modern ray-finned fishes based on 327 mitogenome sequences.

Second International Barcode of Life Conference 18-20 September 2007 44 Conference Abstracts Plenary Session Abstracts

HOW LARGE-SCALE BARCODING PROMOTES LARGE-SCALE BIODIVERSITY ASSESSMENT

DANIEL P. FAITH1, Andrew Baker2 and Steffen Klaere3

1The Australian Museum, Australia 2Queensland University of Technology, Australia 3MFPL - Center for Integrative Bioinformatics, Vienna, Austria

Biodiversity goals such as the widely-accepted 2010 biodiversity target of a “significant reduction in the rate of loss of biodiversity” desperately need information covering many places and many taxa - including “surrogates” information for the extensive biodiversity still unknown to science. The expected large increase in biodiversity information from CBOL can be integrated with environmental and other information to assist such assessments. These strategies may use species- level data, or use the approximate phylogenetic information from DNA barcoding through application of a phylogenetic diversity measure, PD (the amount of evolutionary history spanned by a set of taxa; Faith, 1992). An example for freshwater invertebrates suggests that the PD approach not only may side-step the species-designation problem, but also may boost surrogacy in reflecting historical relationships among areas. An example application of PD to geographic and phylogenetic information from BoLD illustrates regional conservation planning assessments for the 2010 target. Our developing web-based PD analyses link to the Newick output trees from the BoLD Taxon-ID Tree module and provide the foundation for barcoding-based “systematic conservation planning”.

Faith & Williams. How Large-scale DNA Barcoding Programs Can Boost Biodiversity Conservation Planning: Linking Phylogenetic Diversity (PD) Analyses to the Barcode of Life Database (BoLD). Abstract. www.amonline.net.au/systematics/pdf/invert-conf-abs2.pdf

Daniel P. Faith

Dr. Faith is a Principal Research Scientist at the Australian Museum in Sydney. Dr. Faith obtained a BA in Mathematics from the University of Chicago and then earned a PhD in Ecology and Evolution from SUNY Stony Brook. Dr. Faith was a research scientist at the CSIRO from 1979 to 1999, before moving to the Australian Museum. His research covers a broad range of topics in biodiversity assessment. Recent research has focused on conservation applications of a “phylogenetic diversity” measure, on novel approaches to achieving the 2010 biodiversity target, and on the philosophy of biodiversity (for the Stanford Encyclopedia of Philosophy). Special emphasis has been given to the best-possible use of museum collections in regional biodiversity assessment, and to the links from biodiversity assessment to sustainability and economics. Applied biodiversity research also includes work on methods for detecting environmental impacts.

Dr. Faith is on the Scientific Committee for a new Core Project within DIVERSITAS, “bioGENESIS”, which promotes links from systematics and evolution to other areas of biodiversity science. He is a long- serving Associate Editor of Systematic Biology and also serves on the editorial board for Global Environmental Change: Human and Policy Dimensions.

Second International Barcode of Life Conference 18-20 September 2007 45 Conference Abstracts Plenary Session Abstracts

BARCODING AND THE PRACTICE OF SYSTEMATICS

RICHARD LANE

Natural History Museum, London

DNA barcoding is beginning to establish its position in contemporary systematics practice after a rather tumultuous gestation period. However, three areas remain either contentious or open to further resolution: the ability of barcoding to distinguish all species (i.e. is specific and sensitive), the potential to divert resource from other areas of systematics and, finally, whether it is introducing a reductionist approach to defining species. The first is a scientific and testable issue tractable by empirical data, the second a sociological phenomenon observable and analysable by qualitative social science methodology, although it contains a value-based element not tractable to analysis, and the third is a philosophical matter which is becoming less relevant to the debate as the objectives of barcoding are refined.

Richard Lane

Prof Lane is Director of Science, at the Natural History Museum, London responsible for setting the strategy for science at the Museum, ensuring that research is relevant and and of high quality. He is also responsible for making sure that the research collection of some 70 million items from all parts of the world operates as a major scientific infrastructure, develops and is made accessible.

Prof. Lane was educated at Imperial College, London and was a senior lecturer at the London School of Hygiene & Tropical Medicine (1985-92).He was then head of the Entomology Department at the NHM, London (1992-97) before moving to the Wellcome Trust (1997-2003) to head International Programmes where he was responsible for funding major research programmes in biomedical and social sciences and forging alliances and collaborations with international organisations and governments.

Prof Lane’s research has been on various aspects of insect vectors of disease. As a consequence of this and other activities he has traveled extensively in the developing world. He has been a consultant to a number of governments and on external review bodies for several research organisations in the UK and internationally. The Natural History Museum, London was a founding member of the Consortium for the Barcode of Life and Prof Lane hosted the first International Barcode of Life Conference in London in 2005.

Second International Barcode of Life Conference 18-20 September 2007 46 Conference Abstracts Plenary Session Abstracts

Session 10: State-of-the-Art Practices: How to manage your barcode data

BOLD, THE BARCODING WORKBENCH

SUJEEVAN RATNASINGHAM

Biodiversity Institute of Ontario & Department of Integrative Biology, University of Guelph, Guelph, ON N1G 2W1, Canada

The Barcode of Life Data System (BOLD) is an informatics workbench aiding the acquisition, storage, analysis and publication of DNA barcode records.By assembling molecular, morphological and distributional data, it bridges a traditional bioinformatics chasm. By providing specialized services, it aids the assembly of records that meet the standards needed to gain BARCODE designation in global sequence databases. Because of its web-based delivery and flexible data security model, it is also well positioned to support projects that involve broad research alliances.

Sujeevan Ratnasingham

Sujeevan Ratnasingham is the Informatics Lead at the Biodiversity Institute of Ontario, University of Guelph, Canada and the Chief Architect of the Barcode of Life Data System (BOLD). He currently leads a team of programmers and data managers to extend and maintain the Barcode of Life Data System. Sujeevan has a background in Computer Science from the University of Guelph with a focus on high performance computing and database analysis. He joined Paul Hebert¹s lab as a bioinformatican in 2002 to support early work on assessing the utility of a single gene barcode approach to delineate species boundaries. He has since been involved in the development of informatics tools, workflows, and infrastructure that support high throughput DNA barcoding.

Second International Barcode of Life Conference 18-20 September 2007 47 Conference Abstracts Plenary Session Abstracts

BARCODE RECORDS IN GENBANK

SCOTT FEDERHEN

NCBI, Nationanl Institutes of Health, Bethesda MD

GenBank, the INSDC (International Sequence Database Collaboration) and the NCBI (National Center for Biotechnology Information) have provided ongoing support for the Barcoding initiative, and are working actively with CBOL (Consortium for the Barcode of Life) and BOLD (Barcode of Life Data Systems) to improve the submission, validation, update and exchange of barcode data. We introduced the BARCODE keyword in GenBank, a barcode data type in the Trace Archive, and a new set of source feature qualifiers to support barcode data requirements (which have proved useful for many other classes of sequence submissions). We have developed a customized web submission tool for barcode submissions (the BarSTool) which has served as the model for the next generation of GenBank sequence submission tools. We have collected and established a database of ~7000 biological specimen repository acronyms (museums, herbaria, culture collections, stock centers, seed banks &c.) that we will curate in collaboration with CBOL and GBIF. We are using this collections database to support the implementation of structured specimen voucher qualifiers in GenBank entries (/specimen_voucher, /culture_collection and /bio_material) that will complement the ongoing use of our LinkOut facility for linking sequence entries with specimen data.

Scott Federhen

Mr. Federhen has a background in mathematics and in computer science/artificial intelligence (MIT, 1975 & Maryland, 1981). While he was working on the formal (denotational) semantics of AI programming languages, he became interested in control structures of biological systems, and returned to graduate school to study developmental genetics of the nematode in the Horvitz lab (MIT, 1979). Mr. Federhen joined the National Center for Biotechnology Information (NCBI) shortly after its inception, and found the perfect place to combine his various interests. His first significant contribution at the NCBI was the development (in collaboration with John Wootton) of the seg low-complexity filter for nucleotide and protein sequences. Since that time, his primary responsibility has been the development of the NCBI/GenBank taxonomy database and related applications, which have been accepted as the international standard for the nucleotide and protein sequence databases. In addition to these responsibilities, Mr. Federhen has made significant contributions to the development of the Entrez system, and to GenBank annotation policies and guidelines. He is one of the principle architects of the highly successful INSD (GenBank/EMBL/DDBJ) sequence database collaboration. Mr. Federhen helped to organize the first of the Banbury Barcode meetings, and has been an active and enthusiastic supporter of the project from the onset.

Second International Barcode of Life Conference 18-20 September 2007 48 Conference Abstracts Plenary Session Abstracts

BARCODE DATA, MUSEUM CATALOGS AND GBIF

SIMON TILLIER

EDIT and Museum National d’Histoire Naturel, Paris

Access to voucher specimens is crucial for successful long term implementation of the Barcode concept. The latter indeed implies a permanent link between the virtual sequences and the real objects which bear the species name in order to maintain the relationship between a sequence and changing species concepts. Although there is so far no formal linking between TDWG, Genbank and Zoobank, discussing these issues seems absolutely necessary to eventually implement data standards allowing easy exchange of barcode data and related specimen information; however it is worth noting that the ABCD standard already includes a section on sequences which allows linking specimens to Genbank. From Museums point of view, the real challenge – in addition to databasing all their biodiversity specimens collections, which is far from being achieved globally – lies in curation of fragmented objects, ie DNA extracts and the specimens from which DNA has been extracted. This implies both standards for DNA extracts collections, and an organisation of the curatorial workflow which is presently not part of the culture of the community. In parallel to collective reflexion on database standards which is already well developed, collective reflexion on curatorial standards and their implementation has to be developed by networks and consortia of collections and museums, such as SYNTHESYS and EDIT.

Simon Tillier

Pr. Simon Tillier is the leader of EDIT, the consortium of large Taxonomic Institutions aiming at integrating taxonomic research and capacities at the European Union level and beyond with support from the European Commission.

Pr. Tillier was trained as a geologist and invertebrate paleontologist in the Université Paris Sud and holds a PhD in Paleontology and a DSc in Zoology from the Université Pierre et Marie Curie. He was the curator of land and freshwater molluscs at the Muséum national d’Histoire naturelle from 1976 to 1990 and collected invertebrates in various regions of the world, particularly in New Caledonia where he had annual field work and worked permanently in 1986-1987. From 1990 Pr. Tillier turned to molecular systematics and phylogeny, and set up and directed the molecular systematics facility of the Museum until 2002. In parallel he got engaged in organization of research in systematics at International, National and Museum levels: he participated in committees in the ESF network “Systematics biology” (1994-1997), in the Consortium of European Taxonomic Facilities (1995-2002), in the Coordination mechanism of the Global taxonomic Initiative (2000-2002), in CBoL, and chaired the scientific subcommittee on digitization of biodiversity data of the GBIF (2000-2002). In France he coordinated the National network of Biosystematics (1994 -1997), and set up and directed the CNRS Institut de Systématique and the Museum department of Systematics en Evolution from 1998 to 2002.

Second International Barcode of Life Conference 18-20 September 2007 49 Conference Abstracts Plenary Session Abstracts

ZOOBANK - THE OPEN-ACCESS ANIMAL NAME REGISTRY - WORKING WITH CBOL TOWARDS THE NEW TAXONOMY

ANDREW POLASZEK

International Commission on Zoological Nomenclature

The ZooBank proposal for an open-access register for the scientific names of all animal species, genera, higher taxonomic categories and nomenclatural acts, is now just over one year old. The prototype was released in August 2006. Our next goal is for ZooBank to develop into a complete register of all nomenclatural and taxonomic data that are governed and affected by the current and future codes of zoological nomenclature. This involves amendments to the present code in order to facilitate electronic publication and registration of names, as well as the precise nature of name- bearing type specimens. With the CBOL initiative an opportunity presents itself to regulate the naming of animal taxa based either partially or entirely on gene sequences in a way that would be compliant with an amended International Code of Zoological Nomenclature.

Andrew Polaszek

Dr Andrew Polaszek is the Executive Secretary of the International Commission on Zoological Nomenclature, an organisation founded in 1895, and based originally at the Smithsonian Institution until its move to the Natural History Museum in London in 1936, where it remains today. The ICZN Secretariat is funded through sales of publications, including the International Code of Zoological Nomenclature, now in its 4th (1999) edition. The Commission's Mission is to achieve "standards, sense and stability in the scientific naming of animals". Dr Polaszek was trained as an applied invertebrate systematist and holds a BSc in Agricultural Zoology from the University of Newcastle and a PhD in Zoology from Imperial College London. He worked for 15 years for the Commonwealth Institute of Entomology (later CABI Bioscience) studying parasitoid wasps, particularly in the context of tropical biological pest control. As well as running the ICZN Secretariat, he still continues research into the systematics and biology of a range of Hymenoptera, investigating their species relationships and phylogenies using combined morphological and molecular analyses.

Second International Barcode of Life Conference 18-20 September 2007 50 Conference Abstracts Plenary Session Abstracts

THE ENCYCLOPEDIA OF LIFE: A WEBSITE FOR EVERY SPECIES

JAMES EDWARDS

Encyclopedia of Life, Smithsonian Institution, USA

The idea for the Encyclopedia of Life (EOL) is deceptively simple: construct a web site for each of the approximately 1.8 million species now known to be present on Earth, and make them all accessible through a single portal. The centerpiece for each site is a species page that provides, in a format common to all the pages, significant information about that species, including the scientific and common names (in several languages) by which the species is known and has been known in the past, an illustration, a range map, habitat and natural history, molecular data (including barcodes), conservation status and human uses, and how to identify it. In addition, each species site will contain a wide variety of more specialized pages, including ones for taxonomists, molecular biologists, geneticists, phylogeneticists, specimen and observational databases, etc. Biodiversity literature will be available, too, through the EOL’s partnership with the Biodiversity Heritage Library (BHL) consortium. The EOL is thus eminently suited to serve as the species database for barcode users.

The EOL is being developed via partnerships with a wide range of biodiversity organizations and institutions from around the world, including CBOL. Further information about the EOL, including examples of draft species pages and a list of FAQs, can be found at www.eol.org.

James Edwards

Dr. James Edwards is first Executive Director of the Encyclopedia of Life. His major role is to oversee the management and coordination of all aspects of the project. In addition, he is responsible for working with the scientific community and general public to identify the material to be included in the EOL species pages and for engaging the scientific community to authenticate that material.

From 1982 to 2007, Dr. Edwards served in the Directorate for Biological Sciences of the U.S. National Science Foundation (NSF). From November 2001 to May 2007, NSF detailed him to Copenhagen to be the Executive Secretary of the Global Biodiversity Information Facility (GBIF).GBIF is an intergovernmental organization devoted to providing Internet access to two major kinds of information: specimens in natural history collections (museums, botanical gardens, living stock centers) and observations of organisms where a specimen was not collected (such as the Audubon Society’s Christmas Bird Counts). These data document the presence of plants, animals, and microorganisms in both space and time and therefore provide a comprehensive view of the world’s contemporary and historical biodiversity. GBIF and the EOL are preparing a Memorandum of Understanding to allow GBIF-mediated data to be made available through the EOL.

Second International Barcode of Life Conference 18-20 September 2007 51 Conference Abstracts Plenary Session Abstracts

Session 11: State-of-the-Art Practices: How do the best barcode labs do their work?

TECHNOLOGY TRANSFER. AN EFFICIENT DNA BARCODING WORKFLOW: HOW WE DO IT, AND MORE IMPORTANTLY TO YOU, HOW WE CAN HELP YOU DO IT.

LEE WEIGT, Amy Driskell, Jeff Hunt, Andrea Ormos

L.A.B., Smithsonian Institution, USA

Technology transfer: DNA barcoding is possible due to the technological revolution brought about by the Human Genome Project and the resultant DNA sequencing technologies we are currently using. Likewise, DNA Barcoding efforts at labs like CCDB (Guelph) and L.A.B. (Smithsonian) are investing much time and effort to produce methods, protocols and workflows that can be readily adopted. There will be a very brief portion on how the major centers with major infrastructure investment can throughput tens/hundreds of thousands of samples per year; but these centers are not located where the biodiversity is, and that biodiversity is sometimes difficult to transport due to logistical or legal constraints. So we will then focus on some strategies to help anyone generate DNA extracts or barcodes from their own biodiversity, either at their facilities or at those in their region. These will include networks of information dissemination and expertise, potential for on- site workshops, new methods to enable better sample capture at the point of collection, and plans for laboratory “blitzes” where we bring the lab capacity, consumables and personnel to your area and train a group of individuals on hundreds to thousands of their own samples in their own region.

Lee A. Weigt

Lee is the Director of the Laboratories of Analytical Biology (L.A.B.) at the Smithsonian Institution’s National Museum of Natural History. The L.A.B. is the hub of the molecular biotechnology infrastructure at NMNH and as some of its components, has the SI’s DNA Barcoding Initiative, the Instrumentation Core, and remote management and assistance to other satellite molecular facilities and projects at the SI.

Lee was trained as a herpetologist specializing in rates of molecular evolution, speciation and population genetics at Miami University in Oxford, Ohio. He then went to Panama in 1988 to build a multiple user molecular laboratory at STRI (the Smithsonian Tropical Research Institute), where he then stayed on to run that facility until 1996. He was the Manager of the Pritzker Lab at the Field Museum of Natural History from 1996 to 1998, and then Director of the Core Facilities at the Virginia Bioinformatics Institute (at Virginia Tech) from 1998 to 2001. He came to the NMNH labs in 2001, and took over the directorship of those labs in 2003, along with co-writing the first proposals for DNA barcoding at the institution, and obtained the institution’s first permanent funding for DNA barcoding infrastructure in 2005.

Second International Barcode of Life Conference 18-20 September 2007 52 Conference Abstracts Plenary Session Abstracts

SAMPLING WILDLIFE IN AFRICA

PAUL BARTELS

Wildlife Biological Resource Centre, National Zoological Gardens of SA, NRF, South Africa. P.O Box 582, Pretoria, 0001. South Africa.

Africa has one of the world’s richest biodiversity heritages and hence a natural advantage that presents a unique opportunity to generate a strong mass of scientific expertise. Wildlife resources are under threat from a variety of quarters, including disease, habitat destruction, population fragmentation and species hybridisation. With the growing global market in biomaterials and biodiversity informatics, developing countries, particularly those that are recognised as ‘mega- diverse countries’, face the enormous challenges of setting up systems to govern access to biodiversity and sustainable utilisation of it’s biodiversity heritage. DNA barcoding promises an efficient and standardised system of cataloguing Africa’s biodiversity, discovering new species and thus broadens the opportunity for developing innovative biodiversity management and technology tools to the benefit of society. Issues of biodiversity concern to Africa include Access and Benefit Sharing (ABS), Intellectual Property Rights (IPR) and Indigenous Knowledge (IK), which relate directly to the basics of species identification and exploitation. DNA barcoding not only supports education and training within the discipline of genomics, but also facilitates national, regional and global multi-disciplinary scientific cooperation. Securing good quality African wildlife specimens for DNA barcoding represents a challenge that is being addressed by scientists within Africa.

Paul Bartels

Dr. Paul Bartels is the head of the Wildlife Biodiversity Resources Centre (wBRC) of the National Zoological Gardens of SA, National Research Foundation. Dr. Bartels also heads up BioBankSA, a membership consortium devoted to Biodiversity Resource Banking (BRB) in southern Africa / Africa. At a CBOL workshop held in South Africa in 2006, Dr. Bartels was nominated as the African Wildlife CBOL representative in South Africa.

Dr. Bartels was trained as a veterinarian at the University of Pretoria and in addition holds a MSc in Zoology from the same university. He was a staff member of the Tompi Seleka Agricultural College for 10 years before joining the Endangered Wildlife Trust (a wildlife NGO) in 1997 as director of the wBRC, a program that he had established while still working at the college. In 2004 the wBRC’s biomaterials bank was recognized as a national asset by the government and after a due diligence process, its operations transferred to the National Zoo, National Research Foundation. Dr Bartels directs the systematic collection, processing, banking, use and distribution of wildlife biomaterials derived from Africa’s tremendous animal biodiversity resources for biodiversity conservation and biotechnology development. The national program includes cooperation with SADC and NEPAD countries where gene-banks are being developed as knowledge-hubs to the benefit of society.

Second International Barcode of Life Conference 18-20 September 2007 53 Conference Abstracts Plenary Session Abstracts

RECOVERING HISTORIC (HDNA) AND ANCIENT DNA (ADNA): IMPLICATIONS FOR DNA BARCODING

DAVID LAMBERT1 and Craig Millar2

1 New Zealand Institute of Advanced Studies, Massey University, Private Bag 102 904, Auckland, New Zealand 2 School of Biological Sciences, University of Auckland, Private Bag 92019, Auckland, New Zealand

Generally, the success of the DNA Barcoding enterprise will be depend on the recovery of barcodes from voucher specimens, typically those held in museums. In the case of avian barcoding, the vast majority of such specimens will be study skins or mounted birds. The recovery of DNA from these samples is difficult because of the properties of historical and ancient DNA. We will review the general characteristics of both hDNA and aDNA and outline the methods that can be used for the accurate recovery of barcodes from such material. For avian species we report conserved primer sequences that enable the amplification of the 648 bp COI region of the mitochondrial genome in either one, three or five fragments. These primers are efficient across a wide phylogenetic range of avian groups. We have used these in our work to barcode the birds of New Zealand and the Antarctic. We have currently barcoded approximately one third of the extant and extinct faunas of these two regions and many samples are from such voucher specimens.

David Lambert

Prof David Lambert is a Distinguished Professor at Massey University in New Zealand and a principal investigator in New Zealand’s Allan Wilson Centre for Molecular Ecology and Evolution. He is also a professor in Massey University’s Institute of Molecular BioSciences. He graduated from the University of Queensland with BSc and MSc degrees in Zoology and Genetics. Later, he obtained a PhD from the University of the Witwatersrand, working in the laboratory of Prof Hugh Paterson. Prof Lambert’s research has focused on aspects of evolutionary theory and evolutionary genetics. He has published extensively in these areas, particularly in relation to species theory, the nature of Darwinian biology and ancient DNA. His research group has pioneered approaches to the estimation of evolutionary rates, as measured by changes in ancient DNA over time.

Prof Lambert’s research group has current programmes on Adélie penguins from the Antarctic, ancient DNA from species of extinct moa and the use of DNA barcoding to determine the efficacy of this approach to the identification of species of ancient life. He is currently a James Cook Fellow working as part of an international programme aimed at the determination of modes of repeat DNA evolution using ancient Sacred Ibis mummies from Egypt.

Second International Barcode of Life Conference 18-20 September 2007 54 Conference Abstracts Plenary Session Abstracts

EXPLORING ARCHIVAL AND ENVIRONMENTAL SAMPLES THROUGH MINIMALIST BARCODES

MEHRDAD HAJIBABAEI

Canadian Centre for DNA Barcoding, Biodiversity Institute of Ontario, University of Guelph

DNA barcoding has proven a powerful approach for probing species diversity in large taxonomic groups and in diverse geographical settings. However, barcode recovery is currently constrained in two important contexts: archival specimens and environmental samples. In the case of archival material, problems derive from the fact that DNA degradation limits the recovery of large amplicons. In the case of environmental samples, massively parallel sequencing is required and such platforms are only capable of delivering up to 200 base sequence reads. These barriers raise a simple question: How much does barcode size matter? Our in silico tests indicate that a 100 base fragment in any region within the cytochrome c oxidase I barcode can provide resolution closely comparable to full-length (650 base) barcodes. In addition, such short regions can be reliably amplified from type specimens of up to two centuries old using universal primers. The same mini- barcode fragment is effective in species recovery from DNA mixtures using FLX 454 parallel pyrosequencing. Minimalist barcodes can therefore broaden the application of barcoding in archival and environmental samples.

Mehrdad Hajibabaei

Dr. Mehrdad Hajibabaei is a Research Scientist at the Canadian Centre for DNA Barcoding, Biodiversity Institute of Ontario, University of Guelph, Canada. He obtained his B.Sc. in microbiology from Tehran Azad University and his PhD from the University of Ottawa in molecular evolution and phylogenetics. During his PhD he studied molecular evolution of the RNA polymerase gene family and applied these genes in resolving a complex phylogenetic problem: seed plant phylogeny and the position of Gnetales. Dr. Hajibabaei joined Dr. Paul Hebert’s lab at Guelph in September 2003, as the first postdoctoral fellow on DNA barcoding. During his postdoctoral research he pioneered the development of high throughput DNA barcoding and worked on DNA barcoding tropical Lepidoptera--in collaboration with Dan Janzen--as well as barcode recovery from archival specimens.

Dr. Hajibabaei is a founding member of the Canadian Centre for DNA Barcoding where he continues his research as an investigator (since April 2006) on various aspects of DNA barcoding. He has published numerous articles on barcoding and holds a patent on a novel bioinformatics search algorithm for barcode sequences. He also serves as the Research Program Coordinator for plants, protists, and fungi in the Canadian Barcode of Life Network.

Second International Barcode of Life Conference 18-20 September 2007 55 Conference Abstracts Plenary Session Abstracts

Session 12: Barcoding with non-COI gene regions

OPTIMISING SELECTION OF DNA BARCODE REGIONS AND CBOL’S GUIDELINES FOR ‘NON-CO1’ SELECTION

FREEK T. BAKKER

Nationaal Herbarium Nederland, Wageningen University branch, The Netherlands

Although mitochondrial encoded cox1 has been the region of choice in most DNA barcoding approaches so far (vertebrates, Lepidoptera) it is becoming clear that alternative and additional regions may be needed for DNA barcoding in other clades. For instance, fungal and nematode DNA barcoding is largely based on rDNA, whereas for land plants multiple regions from the plastid DNA will be used for effective DNA barcoding, and for various protest clades region selection is in its infancy. Of course practical and empirical considerations in region selection are paramount: (1) ease and feasibility of PCR amplification, especially from degraded material, (2) species-level DNA sequence divergence encountered, and (3) the universality of primers used to amplify the barcodes. However, factors relating to structural aspects of the region proposed, to reproductive biology of its target clades, as well as its possibility of matching 'unknowns' to a growing reference library will influence region selection too. Several non-COI DNA barcode regions are expected to be proposed to CBOL’s Science Advisory Board soon. In this presentation, guidelines, protocols and procedures for non-COI region selection as used by CBOL will be outlined.

Freek T. Bakker

Dr. Freek Bakker chairs CBOL’s Scientific Advisory Board, currently consisting of thirteen experts in the fields of systematics, taxonomy, molecular evolution, population genetics, databasing, DNA protocols and DNA barcode data analysis, at the same time including the chairs of all CBOL’s Working Groups. The SAB serves as a think tank as and provides advice to CBOL’s Executive Committee, for instance on DNA barcode projects or on ‘non-CO1’ DNA barcode region proposals.

Dr. Bakker was trained as a biochemist/population geneticist and holds a PhD in seaweed systematics at the University of Groningen, The Netherlands. He did a 5-year postdoc on angiosperm phylogenetics at the University of Reading and The Natural History Museum, London (UK), studying South African Cape Pelargonium. From 2000 Dr. Bakker has been at the Nationaal Herbarium Nederland at the Wageningen University branch, where he is an Assistant Professor, focusing on evolution and systematics of African plants. He joined CBOL in 2005.

Second International Barcode of Life Conference 18-20 September 2007 56 Conference Abstracts Plenary Session Abstracts

STANDARDIZED LAND PLANT BARCODING REQUIRES A MULTI-LOCI APPROACH

ROBYN COWAN

Jodrell Laboratory, Royal Botanic Gardens, Kew

To fulfill the envisaged range of applications of DNA barcoding in land plants, the barcode region would ideally be both sufficiently variable to obtain species level identifications and universally applicable so that even a sample with no morphological characters, ie: a complete unknown, can be identified. Because of the unsuitability of mitochondrial regions, and presently nuclear regions, for use as a DNA barcode in land plants, the challenge has been to identify plastid regions that would serve this purpose. Using in silico screening of the Nicotiana plastid genome to identify potentially suitable regions we then went through a process of initial screening for universality and variability using approximately fifty closely related species pairs from all major land plant lineages to identify the regions with the most potential as barcodes. From this six regions were selected and further screened on a range of plant groups with near complete species level sampling. The results of this study showed that no one region performed well on both variability and universality. However by combining up to three regions the species level variability score is improved significantly, and including a slightly less variable but more universal region provides an ‘anchor’ for initial placement of a complete unknown to its major plant group.

Robyn Cowan

Following an early career in biomedical sciences, Robyn Cowan has spent the last eight years at Royal Botanic Gardens, Kew as a conservation geneticist. Her main research is in the development and use of molecular techniques to study species delimitations and population dynamics. Much, although not all of this work, is carried out in collaboration with the UK statutory conservation agencies and the resulting data are used to inform Species and Biodiversity Action Plans, applications for protected areas, and ‘on the ground’ management of species. Lately a main focus of her work has been establishing and implementing a standard plant DNA barcode for land plants in an externally funded collaborative research programme. This project was established under the auspices of the Plant Working Group of the Consortium for the Barcode of Life, of which she is presently chairperson, and involved a collaboration of 11 institutes.

Second International Barcode of Life Conference 18-20 September 2007 57 Conference Abstracts Plenary Session Abstracts

USING DNA BARCODES TO TEST THE IDENTITY AND PURITY OF PLANT-BASED MEDICINES AND HERBALS

JOHN W. KRESS and David L. Erickson

Department of Botany, MRC-166, National Museum of Natural History, Smithsonian Institution, P.O. Box 37012, Washington, DC 20013-7012, USA

Barcoding is emerging as one of the many important tools on the modern taxonomist’s work bench as an accurate genetic identifier in taxonomic and other applied studies. Discovery of a DNA barcode for land plants has been limited by intrinsically lower rates of sequence evolution in plant genomes than rates observed in animals. This low rate has complicated the trade-off in finding a locus that is universal and readily sequenced yet has sufficiently high sequence divergence at the species-level. We have shown that a combination of the rbcL coding region and the trnH-psbA non-coding region of the plastid genome provides the necessary universality and species discrimination necessary for an effective plant barcode. We apply this two-locus plant barcode to a data set of 689 species of vascular plants (in 103 families) listed as the major medicinal plants in World Economic Plants: A Standard Reference. This set of plant barcodes can now be used to screen the content and quality of natural plant products employed as medicines and herbals in both commercial and non-commercial applications.

John Kress

John Kress was born in Illinois and received his education at Harvard University (B. A., 1975) and Duke University (Ph. D. 1981) where he studied tropical biology, ethnobotany, and plant systematics and evolution. Before coming to the Smithsonian Dr. Kress was the Director of Research at the Marie Selby Botanical Gardens in Sarasota, Florida, from 1984 to 1988.

Currently he is Curator and Research Scientist as well as former Chairman of the Department of Botany at the National Museum of Natural History, Smithsonian Institution. He is an Adjunct Professor of Biology at George Washington University and at Xishuangbanna Tropical Botanical Garden (Chinese Academy of Sciences), a Fellow of the American Association for the Advancement of Science, and Executive Director of the Association for Tropical Biology and Conservation. His research studies are concentrated on the evolution of breeding and pollination systems in plants, systematics and phylogeny of the Zingiberales, genetic variation and speciation in tropical angiosperms, new tools for taxonomy, and conservation of tropical ecosystems.

Second International Barcode of Life Conference 18-20 September 2007 58 Conference Abstracts Plenary Session Abstracts

DNA-BARCODING ON A DIFFERENT SCALE: CHALLENGES WITHIN THE COCCOIDEA

LYN COOK

The University of Queensland

Anyone who has been charged with the identification of a scale insect to species will appreciate that this is a group that could well do with a DNA barcode database. However, this has proved more difficult for scale insects than for most other insect groups examined to date, with world-wide efforts so far failing to amplify the COI barcode region reliably for any but a handful of species. Sequences from other regions of the mitochondrion of scale insects reveal extreme base composition bias within and among taxa, suggesting that a "universal" primer set for the COI barcode region will be difficult (or impossible?) to develop. In the interim, other regions have been explored for use as an alternative barcode region. The most reliable thus far appears to be the D2 (and D3) expansion region of nuclear large subunit rRNA (28S). Universal primers readily amplify a fragment for all taxa yet tested, but length variation and high sequence divergence may hinder its utility.

Lyn Cook

Dr Lyn Cook specializes in scale insect molecular systematics and is a member of the Scale Insect Barcode Initiative (SIBI). She has a PhD in Evolution, Ecology and Systematics from the Australian National University, and is currently a faculty member in the School of Integrative Biology at the University of Queensland. Dr Cook's research also extends to the molecular systematics, biogeography and evolution of plants (especially those that are hosts to scale insects!), and characterizing the microbial endosymbionts of scale insects.

Second International Barcode of Life Conference 18-20 September 2007 59 Conference Abstracts Plenary Session Abstracts

CANADIAN PLANT BARCODING RESULTS WITH SOME GLOBAL IMPLICATIONS

SEAN W. GRAHAM1, Steve Newmaster2, Brian C Husband2 and Spencer CH Barrett3

1UBC Botanical Garden & Centre for Plant Research, and Dept. of Botany, University of British Columbia 2Department of Integrative Biology, University of Guelph 3Department of Ecology & Evolutionary Biology, University of Toronto

The Canadian Plant Barcoding Working Group is addressing the effectiveness of a variety of plastid (chloroplast) regions as barcoding markers, primarily from the perspective of the land-plant flora of Canada. Our initial efforts have focused on quantifying how well barcoding regions work for a regional flora, and on assessing the utility of barcoding markers for belowground ecological surveys. Although the Canadian flora is relatively depauperate (some 5,000 species, of which ~80% are flowering plants), it includes a diverse array of taxonomically complex groups, such as grasses, sedges and willows. Individual plant species may often not have well-understood or well- behaved boundaries; for example, a substantial fraction of all plant species are thought to be paraphyletic. We quantify the impact of species fuzziness on barcoding success in a broad survey of Canadian grasses (Poaceae) that includes multi-population sampling from regional to continental scales.

Sean W. Graham

Dr. Sean Graham is an Associate Professor and Research Director at the University of British Columbia (UBC) Botanical Garden & Centre for Plant Research. His plant molecular biodiversity research program includes efforts to reconstruct land-plant deep phylogeny, funded through Canadian and US federal granting agencies (NSERC, NSF). He is a founding member of the Canadian University Biodiversity Consortium (funded by the Canadian Foundation for Innovation, CFI), whose goal is to develop a nationally networked database of university biodiversity collections. He is a principle investigator in the plant theme of the Canadian Barcode of Life Network, whose goal is to develop and apply barcoding markers to the study of the Canadian land-plant flora. Dr. Graham was trained in plant systematics and evolution and holds a B.Sc. in Genetics from the University of St. Andrews, and a Ph.D. in Botany from the University of Toronto. He was a research associate in plant molecular systematics at the University of Washington from 1996-1998, and an Assistant Professor at the University of Alberta Department of Biological Sciences, where he served as curator of the vascular plant herbarium from 1999-2003. Dr. Graham’s position at UBC is a cross-appointment between the Faculty of Land and Food Systems, and the Dept. of Botany.

Second International Barcode of Life Conference 18-20 September 2007 60 Conference Abstracts Plenary Session Abstracts

PROGRESS TOWARD DNA BARCODING THE VAST DIVERSITY OF FUNGI

AMY ROSSMAN

Systematic Botany & Mycology Laboratory, USDA-ARS, Beltsville, MD USA

The Fungi are a group of organisms essential for life on Earth. With 90-99% of the fungi still unknown, a great deal of diversity would be revealed through DNA barcoding. Experiences with use of the CO1 gene as a barcode have produced mixed results. Although the CO1 gene works for the non-fungal Oomycota and a few groups of true Fungi, problems in using this gene as a DNA barcode for most true Fungi involve amplification because of length variation in the mitochondrial genomes that results in introns of differing lengths. Other issues concern the presence of multiple copies of the gene and the difficulty of designing primers with a broad taxonomic coverage. The entire ITS region of the nuclear rDNA is considered the most appropriate gene for DNA barcoding of true Fungi. Fungal-specific ITS primers exist, have been widely applied, and are known to work well across the major groups of fungi. Potential DNA barcoding projects of fungi include fungi in the air including mycotoxin producers, fungi associated with global trade, fungi of boreal forests, and medically important fungi. Those most excited about this technology are ecologists working with environmental samples and those concerned with biosecurity and agricultural quarantine issues.

Amy Rossman

Dr. Amy Rossman is the Research Leader of the Systematic Mycology and Microbiology Laboratory (SMML), U.S. Department of Agriculture, Agricultural Research Service (ARS), Beltsville, Maryland, USA. In addition, she is the Director of the U.S. National Fungus Collections, the “Smithsonian” for fungi with one-million fungal reference specimens. The SMML conducts research on plant-associated fungi of importance to agriculture and natural resources as plant pathogens including invasive species and as agents of biological control.

Dr. Rossman was trained as a systematic mycologist and holds a B.S. in Biology from Grinnell College and a Ph.D. in Botany (Mycology) from Oregon State University. She was a postdoctoral associate at Cornell University, Ithaca, New York, and the New York Botanical Garden, Bronx, New York. She served as the plant quarantine mycologist from 1980-1983 before joining the ARS as Research Leader. She had conducted systematic research on a wide range of fungi concentrating on microfungi in the Diaporthales and Hypocreales including their asexual states publishing over 120 scientific papers. She was the leader in completing the comprehensive reference book, Fungi on Plants and Plant Products on the United States, including 13,000 species of fungi. This resource now exists as part of a database of fungi on plants worldwide with over 72,000 species of fungi. She also published a book on Protocols for an All Taxa Biodiversity Inventory of Fungi in a Costa Rican Conservation Area. She is a Fellow of both the American Academy for the Advancement of Science and the Mycological Society of America.

Second International Barcode of Life Conference 18-20 September 2007 61 Conference Abstracts Posters – By Subject

PRESENTER BOOTH BIRDS

HEBERT, P. E1 Barcoding Birds: progress towards a global perspective YOO, H. E2 DNA barcode analysis for Korean birds

FISH BAILLY, N. D1 Biodiversity information systems and Barcode of Life: Crosslinks CHEN, I-S. D2 Mitochondrial DNA barcoding identification employing on larval fish community for gobioid fishes in the estuary of Langyang river basin, Taiwan DìAZ DE ASTARLOA, J. D3 New species and novel biological insights discovered via barcode assisted taxonomy of Argentinean marine fishes GWO, J. D4 Genetic relationship among four subspecies of Oncorhynchus masou examined using AFLP HANNER, R. D5 Barcoding deep sea fishes from the northern Mid Atlantic HSU, K. D6 Comparative performance of the COI and cyt b genes in DNA barcoding of fishes KAPOOR, S. D7 DNA barcoding endemic freshwater finfish of India KARTAVTSEV, Y D8 An experience of a participation in the barcoding of far eastern fish species of Russia on cytochrome oxidase 1 gene sequence data LIN, S. D9 Evaluation of species diversity of the South American armored fishes (Siluriformes: loricariidae) in the aquarium markets through DNA barcodes SAMADI, S. D10 The barcoding of Antarctic fishes of Terre Adélie. SHAO, K.T. D11 Applying DNA barcode on identification of larval fish can improve community analysis SHY, H. D12 DNA barcoding of Taiwanese fish species STEINKE, D. D13 DNA barcoding New Zealand marine fishes: proof of principle and applications SWARTZ, E D14 Targets and challenges for the Fish Barcode of Life Initiative (FISH- BOL) to barcode all African fish YANCY , H. D15 The potential use of DNA barcodes in regulatory science

FUNGI CHEN, W. J1 Development of a DNA array using signature oligonucleotides from CO1 barcodes for identification and detection of species of Penicillium Subgenus Penicillium HAJIBABAEI, M. J2 High throughput barcoding of fungal species: a feasibility analysis LIANG, Y. J3 Barcodes of mycorrhizal fungi in subtropical forest ecosystems: applications in species identification and biodiversity estimation MRIDHA, M J4 Fungal diversity in Bangladesh SANTAMARIA, M. J5 Mitochondrial barcode in Ascomycota: testing an intron-free gene ZARE, R. J6 Gibellulopsis, a suitable genus for Verticillium nigrescens, and Musicillium, a new genus for V. theobromae

INITIATIVES ATOYEBI, O.J. A1 The need for barcoding applications to plant genetic resources conservation program in Nigeria JINBO, U. A2 The Japanese Barcode of Life Initiative (JBOLI) ZEIN, M. A3 Initiative DNA barcoding research in Indonesia

Second International Barcode of Life Conference 18-20 September 2007 62 Conference Abstracts Posters – By Subject

INSECTS ARMSTRONG, K. I1 Filling diagnostic gaps from the border: exotic species identification by DNA barcoding BALL, S. I2 DNA barcodes contribute to elucidation of keystone species taxonomy for the New Zealand Sooty Beech Scale Insect BALL, S. I3 Using DNA barcodes to assess New Zealand tachind fly biodiversity BALL, S. I4 Molecular identification of Thrips obscuratus (Thysanoptera) using DNA barcodes BOAKYE, D. I5 Defining the taxonomic status of Anopheles gambiae s.s. through DNA barcoding CHANDRASEKHAR, N. I6 Molecular systematics and barcoding of Hyblaea puera (Teak Defoliator) based on mitochondrial and nuclear genes CHO, S. I7 Adoxophyes orana species complex (Lepidoptera: Tortricidae), a conflict between barcodes and pheromones? CYWINSKA, A. I8 DNA barcoding of biting insects from North America and beyond. DEWAARD, J. I9 Monitoring moth diversity in Canadaís forests: the application of DNA barcoding FLOYD, R. I10 DNA barcoding of pests and invasive species: aphids, adelgids and scales GIBBS, J. I11 DNA barcode phylogeny identifies dual origins of cleptoparasitic behaviour in the bee subgenus Dialictus (Halictidae: Lasioglossum) HAJIBABAEI, M. I12 Unmasking hidden butterfly diversity with DNA barcodes: fifteen putative species of Nymphalidae. HAJIBABAEI, M. I13 Barcoding Ephemeroptera, Plecoptera, and Trichoptera of North America HUANG, D. I14 Multigenes match mass extraordinary sexual dimorphism and male polymorphism of twelve fig wasps from small fig Syconia JOSHI, R. I15 Application of DNA barcoding to invasive populations of leafminers, Liriomyza spp. (Diptera: Agromyzidae) in the Philippines KARUNARATNE, I. I16 Barcoding of bees of Sri Lanka KUMAR, N. I17 DNA Barcodes and sibling species complexes of Anopheline mosquitoes in India (Diptera: Culicidae) LU, K.H. I18 Development of a biochip for rapid discrimination of the species of four tephritid fruit flies (Diptera: Tephritidae) MALEWSKI, T. I19 DNA barcoding in forensic entomology SHOUCHE, Y. I20 DNA barcoding to distinguish species of butterflies from Western Ghat, India SMITH, A. I21 A CO1 DNA barcode is a sufficient lens to begin to focus species discovery: case studies in hyper-diverse parasitoid assemblages SUTOU, M. I22 DNA barcodes of the genus Sciara (Insecta, Diptera, Sciaridae): Application to the study of "armyworms" SUTRISNO, H. I23 Molecular Phylogeny of Indo-Australian Glyphodes and its allied genera (Lepidoptera: Crambidae) TSO, W. I24 DNA-Based recognition of Papilioniodea from Taiwan VIRGILIO, M. I25 DNA barcoding reveals incongruence between molecular characters and nominal species in the Ceratitis fasciventris, C. anonae, C. rosa complex (Diptera: Tephritidae) YASSIN, A. I26 Mitochondrial DNA (CO-I + CO-II) discovers two cryptic species and two independent geographic radiations in Zaprionus indianus, a recent cosmopolitan Drosophilid YOSHITAKE, H. I27 All-Ceram-JPN: a barcoding project on longicorn beetles of Japan

Second International Barcode of Life Conference 18-20 September 2007 63 Conference Abstracts Posters – By Subject

INVERTEBRATES ARITUA, V. H1 Molecular DNA barcoding of nematode species in Uganda CHANG, C. H2 Barcoding of earthworms: a case study of the Pheretima complex (Clitellata: Megascolecidae) LEE, Y-C. H3 Molecular phylogeny suggests polyphyly of both Cyclophorinae and Aycaeinae (Mollusca: Cyclophoridae) KANG, S.H H4 Molecular identification of the perinereis species (nereididae: polychaeta) from indo-west pacific shores MAMMALS HEBERT, P. F1 Barcoding bats: global campaigns and regional focuses TOMMASEO, M. F2 Evolutionary history of the genus Sus in Citochrome b sequences YAO, C-J. F3 Identification of cetacean meat in the markets of Taiwan based on mtDNA markers MARINE BRIX, S. C1 Barcoding CeDAMar Isopoda BUCKLIN, A. C2 DNA barcoding of the marine holozooplankton assemblage: sequencing- at-sea during oceanographic biodiversity surveys. CHITIPOTHU, S. C3 Towards microarray-based DNA-barcoding of marine invertebrates CHUANG, Y. C4 Mitochondria data of ancient genus Siderastrea implied the importance of taxonomy revision in Scleractinia ERPENBECK, D. C5 Detection of marine bioinvaders in a Mediterranean industrial port by DNA barcoding. ERPENBECK, D. C6 Cleaning up the mess - applied DNA taxonomy of sponges with the Sponge Barcoding Database (SBD, www.spongebarcoding.org) ERPENBECK, D. C7 The Sponge Barcoding Project (SBP) and the Sponge Barcoding Database (SBD) - aiding in the identification and description of poriferan taxa GAONKAR, C. C8 Molecular probes: A tool to identify marine invertebrate larvae HSIAO,S.T. C9 Identification of important crabs of Taiwan using mitochondrial COI gene sequence and PCR-RFLP analysis KIM, C-G. C10 DNA barcode of the Korean marine invertebrates LAI, J. C11 The union of barcoding and taxonomy: A case study in the commercially valuable Blue Swimming Crab, Portunus pelagicus species complex LIN, C-W C12 The taxonomic status of the morphological groups in the deep-sea squat lobstergenus Munidopsis (Crustacea: Decapoda: Galatheidae) inferred by molecular anaylsis. LIN, M. C13 Evolutionary mitogenomics of the hexacorallian anthozoans (Cnidaria; Anthozoa): gene order, intron evolution, and phylogeny MOOTHIEN PILLAY, K. C14 Delineation of species boundaries in Pavona spp. PLAISANCE, L. C15 Exploring coral reef biodiversity using DNA barcoding PONTIN, D. C16 Molecular analysis of the genus Physalia (Cnidaria: Siphonophora) in New Zealand PUILLANDRE, N. C17 DNA barcodes and Molecular alpha-taxonomy of a poorly explore but hyperdiverse group of marine gastropods : the Turrinae (Gastropoda). SAMADI, S. C18 A Barcode inventory of deep-sea fishes from the SANTO 2006 expedition. STEINKE, D. C19 The Census of Antarctic Marine Life (CAML) Barcoding Initiative TSANG, L-M. C20 What DNA Barcode tells us about hidden biodiversity: a story of intertidal acorn barnacles WANG, W. C21 A phylogenetic analysis of brine shrimp in China by DNA barcoding WU, S-P. C22 Detecting new and cryptic species of Camaenids land snails from Taiwan with evidences via traditional and molecular methods, and the phylogenetic analysis with different molecular markers YANG, C.H. C23 Revision of the Heterocarpus gibbosus Bate, 1888 group (Crurstacea: Decapoda: Pandalidae) by inferring to partial mitochondrial DNA sequence analysis

Second International Barcode of Life Conference 18-20 September 2007 64 Conference Abstracts Posters – By Subject

METHODS CHU, K.H. B1 Rapid tree construction for DNA barcoding using a simple correlation analysis without sequence alignment HANNER, R. B2 Species identification in cell culture: A two-pronged molecular approach. HEBERT, P. B3 Differentiating between error and information in DNA sequence data IVANOVA, N. B4 Methods for streamlining high throughput DNA barcoding MASSARI, J. B5 Species classification in DNA barcode data RODERICK, G. B6 The Moorea BIOCODE Project: enabling a model ecosystem approach to conservation science SARKAR, N. B7 Deriving and Using Barcode Based Diagnostics to Facilitate Information Retrieval SAVERIO, V. B8 Testing the efficiency of mt barcode markers across large stretches of taxonomic diversity SINGER, G. B9 GoogleGene: a visual, web-based tool for linking DNA barcode sequences to species information. STEINKE, D. B10 Accelerated biodiversity surveys through DNA barcoding VAUGHAN, R. B11 BARCODE data submission and retrieval from the EMBL Nucleotide database YEH, W-B B12 Microplate-based method for DNA barcoding YU, C.R. B13 Classification and clustering for DNA barcode data ZIMMERMAN, J. B14 DNA damage in preserved specimens and tissue samples: A molecular assessment

METHODS COMPUTER DEMONSTRATION BOOTHS HAJIBABAEI, M. B15 ibarcode.org - Analytical and visual tools for DNA barcoding LOU, M. B16 Fingerprint: visual depiction of variation in multiple sequence alignments LU, H.S. B17 Information Visualization and Fusion for Barcodes of Life in Environment and Society RATNASINGHAM, S. B18 BOLD, The barcoding workbench SAVERIO, V. B19 Exploring more phylogenetic explicit protocols for species assignment to be implemented in a barcode server STONES, R. B20 DNAPrints - Bionformatics software for VNTR Analysis/Visualisation

Second International Barcode of Life Conference 18-20 September 2007 65 Conference Abstracts Posters – By Subject

PLANTS ALMEIDA-da-SILVA, R. K1 Phylogenetic relationships within the Scilla species complex (Hyacinthaceae) based on plastid barcoding-DNA sequences AMEKA, G. K2 Barcoding of Podostemaceae from Africa AUSTERLITZ, F. K3 A practical use of the DNA Barcode: differentiation of the common ash (Fraxinus excelsior L.) and the narrow-leaved ash (F. angustifolia Vahl) for commercial purposes. BARCENAS, R. K4 Barcoding cacti: a tool in the battle against illegal trade BIENIEK, W. K5 Testing of matK, rpoB, rpoC1, trnH-psbA sequences for plant DNA barcoding CAMERON, K. K6 Alternative genes for plant DNA narcoding: a floristic approach CRAYN, D. K7 From banks to barcodes: barcoding a species rich, historically important and geographically constrained flora from Australia GUO, H. K8 Identification of traditional Chinese medicine Radix Astragali by DNA barcoding JAMES, K. K9 Barcoding Darwin’s meadow: a 21st century botanical inventory at historic Down House KIM, K-J. K10 Phylogeny of Panax (Araliaceae) based on the complete chloroplast DNA sequences and the development of DNA barcoding makers at species level. LIU, C. K11 Identification of multiple markers for phylogenetic and DNA barcoding studies for medicinal plants MADRIAN, S. K12 Order out of chaos: DNA barcoding in the Lauraceae PASHA, M. K13 Construction of DNA barcode of Pteridophytes by using rbcL gene and its flanking region SHAW, P-C. K14 Construction of a DNA database for barcoding Chinese medicinal material SPOUGE, J. K15 A generalized ROC analysis for comparing the effectiveness of various loci and distances for plant barcoding VAN DER BANK, M. K16 DNA barcoding of the flora of the Kruger National Park (South Africa) VISCHI, M. K17 Application of DNA barcoding in endagered grasslands of Italy WARNER, J. K18 DNA barcoding of Meso-American orchids for biodiversity research and conservation YAO, H. K19 Applying DNA barcodes to identify medicinal plants

PROTISTS FERRELL, J. G1 The evaluation of DNA barcoding for identification of dinoflagellates: a test using Prorocentrum MCDEVIT, D. G2 DNA barcoding reveals substantial cryptic diversity among macroalgae in Canada's three oceans SHERWOOD, A. G3 DNA barcoding the Hawaiian red algal flora SIDDIKI, A. G4 The DNA barcoding of Cryptosporidium - Bangladesh perspective

Second International Barcode of Life Conference 18-20 September 2007 66 Conference Abstracts Posters - Floorplan

Second International Barcode of Life Conference 18-20 September 2007 67 Conference Abstracts Poster Abstracts – By Presenter

PHYLOGENETIC RELATIONSHIPS WITHIN THE SCILLA SPECIES COMPLEX (HYACINTHACEAE) BASED ON PLASTID BARCODING-DNA SEQUENCES

Castro Araújo, C.1, Sequeira, F.2, Ferrand, N.1, ALMEIDA-DA-SILVA, RUBIM1

1CIBIO/UP, Centro de Investigação em Biodiversidade e Recursos Genéticos, Campus Agrário de Vairão, 4485-661 Vairão, Portugal & Departamento de Botânica, Faculdade de Ciências, Universidade do Porto, Rua Campo Alegre 1191, 4150 -181 Porto, Portugal

2CIBIO/UP, Centro de Investigação em Biodiversidade e Recursos Genéticos, Campus Agrário de Vairão, 4485-661 Vairão, Portugal

The Scilla complex comprises approximately 70 recognised species distributed across Europe, Africa and Asia. In this study, we examined the potential applicability of three barcoding plastid genes (accD, ropC and ropB) for phylogenetic analysis within the Scilla species complex, focusing on the Mediterranean Basin. We amplified multiple sequences from each of those genes (encompassing a total of ≈ 1000 bp) corresponding to 15 Scilla species and one outgroup (Brimeura sp). Our preliminary results showed evidence for two well-supported clades; one comprising essentially the Iberian species, and a second grouping those that occur mostly in North Africa and Madeira Island. The results also showed that S. verna is paraphyletic, suggesting that populations from North Africa could be highly differentiated, thus deserving further studies. The congruence between phylogenetic position and geographic distribution as well as taxonomic implications of this study are discussed.

Subject: Plants

BARCODING OF PODOSTEMACEAE FROM AFRICA

GABRIEL AMEKA1, and Mike Chase2

1Department of Botany, University of Ghana, Legon, Ghana 2Jodrell laboratory, Royal Botanic Gardens, Kew, London, UK

The Podostemaceae are the largest family of strictly aquatic flowering plants and are found mainly in the tropics and subtropics. The plants grow attached to rocks in rapid river habitats and waterfalls. There are c. 270 species of Podostemaceae world-wide with c. 79 species in 16 genera occurring in Africa. The Podostemaceae may be used as indicators of river health. For this concept to have a wider application, however, the identification of the species is important. Presently the taxonomy, including the identification, of the Podostemaceae in Africa needs updating. Some work is in progress to address this concern. DNA barcoding of species of Podostemaceae in Africa will enhance their identification. DNA barcoding can be used by non-specialists to assign unidentified specimens to their species. The project, on barcoding of Podostemaceae of Africa, will start in October 2007 and will be carried out in the Jodrell laboratory in RBG, Kew. Freshly collected plant materials, dried in silica gel, will be prepared and run through a sequencer. We propose to use portions of three plastid genes, rpoCI, rpoB and matK. The results of the project will be widely disseminated through publications in scientific journals.

Subject: Plants

Second International Barcode of Life Conference 18-20 September 2007 68 Conference Abstracts Poster Abstracts

MOLECULAR DNA BARCODING OF NEMATODE SPECIES IN UGANDA

ARITUA, VALENTE1, Nakachwa, R.1, Namaganda, J.1, Lilley J. C.2, Nabulya, G.1, Tushemereirwe, W.1, and Atkinson, J. H.2

1National Agricultural Biotechnology Center, Kawanda Agricultural Research Institute, P. O Box 7065, Kampala, Uganda 2University of Leeds, LS2 9JT, Leeds, UK.

Molecular fingerprinting approaches to characterise nematodes are numerous and are commonly employed together to provide a polyphasic data. Definitive information on species relatedness in nematodes is often obtained by DNA sequencing studies using ribosomal internal transcribed spacer (ITS) loci. However, the level of diversity in the ITS loci has proven insufficient for reliable species identification of nematodes. Recently, the sequences of small fragments of the small subunit nuclear ribosomal DNA (SSU or 18S-rDNA) was shown to be useful in identification of nematodes and other small organisms in an approach known as DNA taxonomy (Floyd et al., 2002; Blaxter 2004). In this report, we provide a preliminary data on our first steps towards use of this system to i) conduct a diagnostic survey and barcode nematode species in Uganda, and ii) to produce baseline data on nematodes for biosafety analysis of banana bearing transgenes for resistance to parasitic nematodes of banana. Using primers designed from the 5’ third of the SSU gene (Floyd et al., 2002), we sequenced individual nematodes sampled from banana plantations at Kawanda Agricultural Research Institute. From this initial screening, search for relatedness in the available public data base genome sequences using the Basic local alignment search tool (BLAST) identified the nematodes as Rhadopholus similis, Helicotylenchus multicinctus and Helicotylenchus dihyslera. This was found congruent with morphological identification. However, for four other nematodes, no relatedness could be found in the database although multiple sequence alignment revealed closeness to Helicotylenchus species. From the DNAs of these nematodes and using the universal primers identified by (Folmer et al. 1994), we are currently amplifying and sequencing the 650-bp fragment of the mitochondrial Cytochrome C Oxidase subunit I (COI) gene, the barcoding region recommended by the Consortium for the barcode of Life (CBOL, www.barcoding.si.edu). In the work that follows, we will further analyse nematode collections made from more banana plantations and other ecosystems in Uganda.

Subject: Invertebrates

Second International Barcode of Life Conference 18-20 September 2007 69 Conference Abstracts Poster Abstracts

IMPROVING SPECIES IDENTIFICATION FOR BORDER BIOSECURITY WITH DNA BARCODES

KAREN ARMSTRONG1, Lalitha Karuneratne1, Carol Muir2, David Voice2, and Alan Flynn2

1Bio-Protection Centre, Lincoln University, New Zealand 2Investigation & Diagnostic Centre, MAF Biosecurity New Zealand

Biosecurity New Zealand (BNZ) entomologists undertake the routine morphological identification of invertebrate specimens intercepted at the countries border. These specimens are associated with various pathways, from commercially imported host material to hitchhikers on inanimate objects, and are important indicators of potential species threats to New Zealand’s primary industries and natural ecosystems. Unfortunately, the variety of life stages and conditions of specimens means that many cannot be readily identified to species. A proof of concept programme is therefore underway to determine if DNA barcode-based identifications can assist. A ‘snapshot’ collection of 313 specimens representing two arthropod classes, including 19 orders of insects, were provided by BNZ. The majority were Araneae, Coleoptera, Diptera and Lepidoptera; in total 51% had been identifiable to species. Focusing on dipteran and lepidopteran taxa, for which a reasonable reference COI barcode dataset exists in BOLD (to June 2007), their identification rate was increased from 60% to 74%. This as-yet small improvement is anticipated to grow as reference COI data continues to be accrued, and the specimens here will be used as a means of targeting taxa to achieve that. The process highlights the potential to capitalize on international and unrelated efforts to improve diagnostic procedures for New Zealand’s biosecurity.

Subject: Insects

Second International Barcode of Life Conference 18-20 September 2007 70 Conference Abstracts Poster Abstracts

THE NEED FOR BARCODING APPLICATIONS TO PLANT GENETIC RESOURCES CONSERVATION PROGRAM IN NIGERIA

ATOYEBI, O. JOHN1, Odofin, W.T1, Aladele, S.E1, Solomon, B.2, Onyia, C.2, Adetunji, O.A.2, Oyedapo, O.O.3

1National Centre for Genetic Resources and Biotechnology, Moor-Plantation, Ibadan, Nigeria 2National Biotechnology Development Agency, Abuja, Nigeria 3Obafemi Awolowo University, Ile-Ife, Nigeria

The National Centre for Genetic Resources and Biotechnology, a centre working with the National Biotechnology Development Agency, has a total collection of approximately 12,500 accessions consisting of both indigenous and exotic germplasm, mainly of food crops, vegetables, tuber, fodder, industrial, medicinal and forest plants. In the seed genebank, there are 7,000 accessions of 216 crop species (i.e. economic and food crops) being kept for short and long term uses. The field genebank covers an area of about 12 hectares of farmland, hosting about 4,000 accessions of various plant and animal species, like the fishery unit in its living collections. Generally, the country’s flora is rich in indigenous medicinal, aromatic and pesticidal (MAP) plants with genetic characteristics of great potential to pharmacological industry. Identification and conservation efforts (which lack application of modern techniques) are associated with scientific and technical problems, and sometimes authenticity. Application of molecular tools and the introduction of barcoding to plant (taxonomy) species identification and biodiversity conservation is still in infancy in the country. Future Biosecurity work Nigeria on Plant Genetic Resources will include the use of DNA barcoding techniques towards solving the problem of species identification among these germplasm collections.

Subject: Initiatives

Second International Barcode of Life Conference 18-20 September 2007 71 Conference Abstracts Poster Abstracts

A PRACTICAL USE OF THE DNA BARCODE: DIFFERENTIATION OF THE COMMON ASH (FRAXINUS EXCELSIOR L.) AND THE NARROW-LEAVED ASH (F. ANGUSTIFOLIA VAHL) FOR COMMERCIAL PURPOSES.

Arca, M.1, Hinsinger D.D.1, Fernandez-Manjarres J.F.1, AUSTERLITZ FREDERIC.1, Gaudeul M.2, Frascaria-Lacoste N.1,

1Laboratoire Ecologie, Systématique, Evolution, UMR AgroParisTech-UPXI-CNRS 8079, Bât, 360, Université Paris-Sud, 91405 Orsay Cedex, France. 2UMR CNRS-MNHN 5202, 45, rue Buffon 75005 Paris, France

Common ash (Fraxinus excelsior L.) and narrow-leaved ash (F. angustifolia Vahl.) are the two ash species that are widely distributed in Western Europe. Since only common ash has major commercial value, contamination of seed by crossing or hybridisation with narrow-leaved ash is strongly undesirable. However the discrimination of seeds, seedlings and juvenile plants is very complicated because of their phenotypic similarity and the widespread hybridisation between the two species that precludes detecting diagnostic markers. This has, in the past, caused a serious commercial dispute between France and Ireland over seed purity.

The present work tests the utility of a DNA Barcode as a tool for differentiating these two species. We have examined the genetic variability of matK, rpoB, rpoC1 and trnH-psbA in a large population sample of Fraxinus excelsior and Fraxinus angustifolia from throughout Europe. The homogeneity between Europeans ashes led us to test the DNA barcode approach at the genus level. Results of our study suggest that the proposed genes cannot be used as discriminating characters between the two species. Therefore for more variable markers are needed to discriminate close species of the genus Fraxinus.

Subject: Plants

Second International Barcode of Life Conference 18-20 September 2007 72 Conference Abstracts Poster Abstracts

BIODIVERSITY INFORMATION SYSTEMS AND BARCODE OF LIFE: CROSSLINKS

NICOLAS BAILLY

WorldFish Center, Natural Resource Management, Philippine Office

FishBase is the provider of the taxonomic and nomenclatural backbone for Fish-BoL using information from Catalog of Fishes, Fishes of the World, and ITIS. Other biodiversity information systems are available for other taxonomic groups, the most important being Catalogue of Life aiming at completing the list of all valid names for the 1.8 million species described by 2011 (one million species were reached in March 2007).

The data structure, accessibility, validation, and updates are crucial to maintain the quality of the taxonomic backbone. Alternatives are presented for data exchanges using semantic web technologies, in particular ontologies and web services.

Two major global programs, Encyclopedia of Life and SpeciesBase, are being developed to realize the Global Species Information System that the G8+5 requested from the scientific community in their Potsdam initiative in March 2007: create a global portal with information on all species on earth. Clearly, Barcode of Life results must be integrated in these efforts. Also, they will have important impacts on taxonomy that will be reflected in the taxonomic backbones themselves.

Crosslinks between Barcode of Life and biodiversity information systems are thus discussed in the GBIF framework to insure the reliability, completeness, and timeliness of data at species level.

Subject: Fish

MOLECULAR IDENTIFICATION OF THRIPS OBSCURATUS (THYSANOPTERA) USING DNA BARCODES

Katrin Schmidt, SHELLEY L. BALL, Steve D. Wratten, Marlene V. Jaspers

Bio-protection Centre, Lincoln University, Canterbury, New Zealand

DNA barcodes were used to develop a molecular identification system for the New Zealand endemic insect, Thrips obscuratus. In New Zealand, this endemic species has successfully extended its host range to many introduced crops and has been classed as a quarantine pest on some export produce. Although it has not been classified as a grapevine pest, there is increasing concern over possible feeding damage on grapes and the potential for it to act as a vector of the fungal pathogen, Botrytis cinerea. Given the increasing importance of the wine industry to New Zealand´s economy the ability to rapidly and accurately identify this species and distinguish it from others is critical. We tested whether DNA barcodes could reliably distinguish this thrips species from several exotic thrips species. All species were successfully discriminated by their COI barcode sequences. Mean divergence between congeneric species was 22.4 %. Overall, mean intraspecific sequence divergence was 1.1% for all species, except T. obscuratus and T. tabaci, which showed higher mean intraspecific sequence divergences (2.5% and 2.4 respectively). We conclude that DNA barcoding, in combination with traditional morphological identification, is a promising tool for identifying thrips species.

Subject: Insects

Second International Barcode of Life Conference 18-20 September 2007 73 Conference Abstracts Poster Abstracts

DNA BARCODES CONTRIBUTE TO THE ELUCIDATION OF KEYSTONE SPECIES TAXONOMY FOR THE NEW ZEALAND SOOTY BEECH SCALE INSECT

SHELLEY L. BALL1, Roger Dungan2, Alison Evans3, David Kelly2, Rosa Henderson4, Karen F. Armstrong1

1 Molecular Diagnostic Laboratory, Bio-protection Centre, Lincoln University, Canterbury, New Zealand 2 Department of Biology, University of Canterbury, Canterbury, New Zealand 3 Department of Conservation, Christchurch, New Zealand 4 Landcare Research, Auckland, New Zealand

Many species play vital roles in ecology by performing important ecosystem services, such as pollination, parasitism, and nutrient cycling. We have developed DNA barcode data to assist in taxonomic investigations of a poorly understood New Zealand native scale insect which is a keystone species in the southern beech (Nothofagus) forests. The New Zealand sooty beech scale insect (Ultracoelostoma sp.) are phloem-feeding insects that excrete ‘honeydew’ - an important food source for native birds and vital to sooty mould growth which is a key element in the breakdown of organic matter and the nutrient cycling. The original morphologically variable species, Ultracoelostoma assimile, was split into two species, U. assimile and U. brittini, but the morphological keys cannot consistently discriminate them. DNA barcodes supported the presence of two very distinct groups differing by ~ 13% COI divergence. These data are assisting in a review of the diagnostic morphological characters for these two species. We are also currently expanding our population sampling to determine the geographic distribution of these species and to test for any other morphologically cryptic species.

Subject: Insects

USING DNA BARCODES TO ASSESS NEW ZEALAND TACHIND FLY BIODIVERSITY

SHELLEY BALL1, Monty Wood2, Karen Armstrong1

1Molecular Diagnostic Laboratory, Bio-protection Centre, Lincoln University, Canterbury, New Zealand. 2Agriculture and Agri-food Canada, Ottawa, Ontario, Canada.

The tachinids are one of the most speciose families of flies, with roughly 10,000 species having been described worldwide. In New Zealand the tachinds are poorly described and their role as natural enemies in both agricultural and natural ecosystems is poorly understood. A few native species are used as biocontrol agents of lepidopteran pests, however, the full potential of the native tachinid fauna as biocontrol agents is unknown. We used DNA barcoding to investigate the taxonomic diversity of New Zealand tachinid flies. DNA barcode data suggests that New Zealand tachinids are a diverse taxon. Concomitant morphological examination is being undertaken to match barcodes to known species, to examine genetic diversity within species, and to identify putative new species.

Subject: Insects

Second International Barcode of Life Conference 18-20 September 2007 74 Conference Abstracts Poster Abstracts

BARCODING CACTI: A TOOL IN THE BATTLE AGAINST ILLEGAL TRADE

ROLANDO T. BARCENAS1, Julie A. Hawkins2

1Facultad de Ciencias Naturales, Universidad Autonoma de Queretaro, Juriquilla, Queretaro, Mexico, CP. 76230. 2School of Biological Sciences, The University of Reading, Berkshire, U.K. RG6 6AS.

Mexico is home to more cacti than any other country in the world. Many of these cactus species are traded, often illegally. High levels of endemism, low densities within sparse populations and the high levels of horticultural interest make the cactus family one of the most endangered plant families in the world. The Mexican government is fighting a continuous battle to tackle illegal trade in cacti but it is hard for law enforcement officials to identify many of the species of cacti and certainly impossible with conventional mechanisms to determine the provenance of individuals. Sequence-based identification of cacti has become a priority for Mexico. We describe an ongoing binational project for the Certificaction of Cacti in Mexico from Aridlands (CCMA, www.uaq.mx/ccma) which aims to test the utility of proposed barcoding regions in the identification of cacti species. We discuss the potential application of sequence-based methods in tracking and controlling illegal trade in cacti and in encouraging legal trade. We outline some of the limitations of barcoding in this context, and describe a complementary approach using barcoding with other DNA sequence-based species identification systems which might be applicable at the species and population levels.

Subject: Plants

Second International Barcode of Life Conference 18-20 September 2007 75 Conference Abstracts Poster Abstracts

TESTING OF MATK, RPOB, RPOC, RBCL AND TRNH-PSBA SEQUENCES FOR PLANT DNA BARCODING

Zbigniew Mirek, WOJCIECH BIENIEK, Agnieszka Sztorc

W. Szafer Institute of Botany, Polish Academy of Sciences, Krakow

Lack of the standard sequences is so far the key question in DNA barcoding of plants. To test the genome domains suggested by the Plant Working Group (CBOL) a set of species representing various systematic groups and differing in respect to age, type of geographical distribution (narrow endemics and the widespread species), biotops occupied and mode of reproduction has been selected. Results of sequencing the above mentioned groups of species are presented and discussed.

Subject: Plants

DEFINING THE TAXONOMIC STATUS OF ANOPHELES GAMBIAE S.S. THROUGH DNA BARCODING

Dziedzom K. de Souza, Charles A. Brown, Michael D. Wilson, and DANIEL A. BOAKYE

Parasitology Department, Noguchi Memorial Institute for Medical Research, University of Ghana. P.O.Box LG 581, Legon-Accra, Ghana.

Of the Anopheles gambiae complex, Anopheles gambiae s.s. is the most important vector of malaria and lymphatic filariasis. Although considered as one species, a huge biodiversity has been described within the Anopheles gambiae s.s. These are five chromosomal forms, with a non- Linnean nomenclature; “Bamako”, “Mopti”, “Savanna”, “Forest” and “Bissau”, and two molecular forms namely; M and S forms. However, the correspondence between molecular and chromosomal forms does not exist in all areas. Furthermore, the presence of the kdr mutation leading to resistance to DDT and pyrethroids in the S form and not M form indicates possible isolation mechanisms. Whether all these forms in West Africa can be classified as the same species is therefore debatable. In this study, we aim at defining operational taxonomic units in the Anopheles gambiae s.s. from different ecological zones in Ghana, West Africa, by analyzing the sequence variations in the cytochrome oxidase subunit 1 gene. Also, the spatial dimensions of genetic variability will be exploited in a simple modeling approach, with recourse to Geographic Information System.

Subject: Instects

Second International Barcode of Life Conference 18-20 September 2007 76 Conference Abstracts Poster Abstracts

BARCODING CeDAMar ISOPODA

SASKIA BRIX

Senckenberg - German Centre for Marine Biodiversity Research (DZMB), c/o Biocentrum Grindel, Martin-Luther-King- Platz 3, 20146 Hamburg, Germany

The CoML project CeDAMar studies the diversity of abyssal fauna. Isopoda (Crustacea) are one of the most important macrofaunal groups in the deep sea. A high number of undescribed species with a low abundance is typical for abyssal plains. This makes taxonomic work time consuming and global biodiversity comparisons a difficult task. The difficulty is to obtain non degraded DNA samples from the deep sea. In CeDAMar we had to overcome this impediment by optimizing sampling gears and protocols. Our project is attempting DNA barcoding in deep-sea isopods for the first time using COI. Recently, the use of COI sequences for species discrimination of Crustacea has been shown. Additionally, we want to test other mitochondrial genes for their applicability in speciesdifferenciation. The main aim is to link morphological descriptions with barcodes, establish a storage of DNA together with voucher specimens. Barcoding and other alternative forms of molecular systematics need to be used alongside traditional taxonomic methods. We want to use the CeDAMar facilities like exchange programs to engage interest in the analysis of barcode data of taxonomists working with material and to develop and disseminate the most effective analytical procedures and display tools for barcode data gained from deep-sea Isopoda.

Subject: Marine

DNA BARCODING OF THE MARINE HOLOZOOPLANKTON ASSEMBLAGE: SEQUENCING-AT-SEA DURING OCEANOGRAPHIC BIODIVERSITY SURVEYS

ANN BUCKLIN1, Robert M. Jennings1, Brian D. Ortman1, Lisa M. Nigro1, Nancy J. Copley2, Peter H. Wiebe2,

1Department of Marine Sciences, University of Connecticut, Groton, CT 06340 USA 2Department of Biology, Woods Hole Oceanographic Institution, Woods Hole, MA 02543 USA

DNA barcodes for the marine holozooplankton assemblage, including ~7,000 described species in fifteen phyla, are being determined in association with a global survey of biodiversity, the Census of Marine Zooplankton (CMarZ). Barcoding is an integral goal of CMarZ, since taxonomic analysis of samples is time-consuming and identification of difficult taxa requires the attention of experts. DNA sequencing is done at sea during CMarZ cruises, where taxonomists place identified specimens in a barcoding queue for immediate analysis. Currently, barcodes for more than 1,500 species of holozooplankton have been determined. The DNA barcode database is used to identify individual specimens, and can be used to identify species in unsorted samples. Future applications of DNA barcodes include automated - and perhaps remote - taxonomic analysis of zooplankton samples, resulting in accurate and complete descriptions of species diversity and distribution throughout the world’s oceans.

Subject: Marine

Second International Barcode of Life Conference 18-20 September 2007 77 Conference Abstracts Poster Abstracts

ALTERNATIVE GENES FOR PLANT DNA BARCODING: A FLORISTIC APPROACH

KENNETH M. CAMERON

The Lewis B. and Dorothy Cullman Program for Molecular Systematics Studies, The New York Botanical Garden, Bronx, New York 10458 USA

A floristically-oriented project was initiated in order to test alternative gene regions for their applicability in serving as universal plant DNA barcodes. The project was designed to be conducted in parallel with and to complement a phylogenetically-oriented project undertaken by an international subset of CBOL’s Plant Working Group. New collections were made and sequenced for every native/naturalized species of vascular plant within the 50-acre forest of The New York Botanical Garden, Bronx, New York. As a “real world” test of the candidate genes, blind samples were collected and sequenced. The identity of these unknowns was only revealed after the DNA barcode libraries were created. Two alternative search methods were employed to make identifications: a dendrogram building method based on ClustalX alignments with neighbor joining, and a BLASTn search method. Of the five coding genes studied, matK exhibited the greatest level of sequence variation, and this was more than sufficient to correctly identify most of the unknown samples. In fact, this was true for all of the test genes. Whereas DNA barcoding of particular taxonomic groups (e.g., palms) may be problematic, this proof-of-principal project demonstrates that floristic approaches to DNA barcoding hold great promise as an alternative means of plant identification.

Subject: Plants

MOLECULAR SYSTEMATICS AND BARCODING OF HYBLAEA PUERA (TEAK DEFOLIATOR) BASED ON MITOCHONDRIAL AND NUCLEAR GENES

CHANDRASEKHAR, NATARAJAN, Neetha N.V., L Vaidyan, L.K., and Banerjee, M.

Rajiv Gandhi Centre for Biotechnology, Thiruvananthapuram, Kerala, India, 695 014

Hyblaea puera is a pest moth of teak woodlands in India and other tropical regions. It was originally described in family Noctuidae later moved to superfamily Pyraloidea due to its strong morphological resemblance. The inter- and intraorder relationship of Hyblaeoidea superfamily with highly diverse superfamilies like Pyraloidea, Bombycoidea, Noctuoidea, Geometroidea, Torticoidea and others are not well understood. Barcoding activities with a potential to develop a digitized library of the genome has opened up wide opportunities. Therefore the aim of the present study was to understand the precise barcoding region and its ability to address the issues on taxonomic debate, species diversity, species and ecotype identification using multigene and ‘folmer region’ of mitochondrial genome. We compared H. puera with 42 species spanning 12 orders in class Insecta. We further compared the barcoding potential of the “folmer region’ for Hyblaea at the intrageneric and ecotype level. The site properties indicate that 12S and 16S Mt genes are better suited for assessing the interorder variability and barcoding application while Cytochrome complex genes are more suited for intraorder variability. While intrageneric and ecotype comparisons of Hyblaea indicate that H. puera is the most primitive species of superfamily Hyblaeoidea. The ‘folmer region’ is able to identify ecotypes of H. puera from across the tropical world.

Subject: Insects

Second International Barcode of Life Conference 18-20 September 2007 78 Conference Abstracts Poster Abstracts

BARCODING OF EARTHWORMS: A CASE STUDY OF THE PHERETIMA COMPLEX (CLITELLATA: MEGASCOLECIDAE)

CHIH-HAN CHANG1, Samuel W. James2, Yong Hong3, Huei-Ping Shen1,4, Jiun-Hong Chen1

1Institute of Zoology and Department of Life Science, National Taiwan University, Taipei, Taiwan 2Natural History Museum and Biodiversity Research Center, Kansas State University, Lawrence, Kansas, USA 3Institute of Agricultural Science, Sangju National University, Sangju, Korea 4Habitat and Ecosystem Division, Taiwan Endemic Species Research Institute, Chichi, Nantou, Taiwan

The Pheretima complex is a group of earthworms in the Megascolecidae. Its distribution is in northeast Australia, Pacific islands, and East and Southeast Asia, including Indonesia and the Philippines, with approximately 30 cosmopolitan species, and more than 900 nominal species within 12 genera. In this study, we reviewed published works of molecular phylogenetic analyses on earthworms, and discussed the feasibility of the mitochondrial cytochrome c oxidase subunit 1 (COI) gene as the barcode of species among these animals. We then made an analysis of the Pheretima complex using 110 species/subspecies collected from various localities in East and Southeast Asia. In this analysis, the monophyly of most described species was supported. Levels of sequence variation are appropriate for species separation. In addition, some incorrectly identified specimens were revealed, new species were discovered, and some previously proposed synonyms were supported or rejected. Conclusively, the results support future earthworm barcode analyses based on COI sequences.

Subject: Invertebrates

Second International Barcode of Life Conference 18-20 September 2007 79 Conference Abstracts Poster Abstracts

MITOCHONDRIAL DNA BARCODING IDENTIFICATION EMPLOYING ON LARVAL FISH COMMUNITY FOR GOBIOID FISHES IN THE ESTUARY OF LANGYANG RIVER BASIN, TAIWAN

Jar-Hong Shih1, Kwang-Tsao Shao1,2 and I-SHIUNG CHEN1

1Institute of Marine Biology, National Taiwan Ocean University, Keelung 202, Taiwan, ROC. 2Research Center for Biodiversity, Academia Sinica, Taipei, Taiwan, ROC.

The larvae of gobioid fishes are as the most important group for estuarine fish resources. The rapid evolving group of gobioid fishes with high species diversity, the necessity of employing high variable genetic marker may be more applicable for revealing the real species diversity and seasonal change for larval fish community. Totally 24 morphotypes have been identified for different fish species with varieties of different developmental stages. Among them, totally 16 recognizable species of larval gobioids can be assigned into 10 species based the comparison of COI and D-loop sequences from extant species with adult. The larvae and juveniles of Rhinogobius maculafasciatus has four different morphotypes; Awaous sp., Periophthalmus modestus, Eleotris acanthopoma, Oligolepis acutipennis have two types; and Eleotris fusca, Scartelaos histophorus, Eleotris sp., Sicyopterus japonicus and Sicyopterus lagocephalus has only one type. So far, a few species of gobies still could not be successfully amplified and sequenced for COI or D-loop because of primer problem. Our study results shown that most dominant goby species in Lanyang estuary is Rhinogobius maculafasciatus which comprised 67.9% of total abundance of 2004-2006. Their migration period to the upstream is in early April every year. The 2nd dominant amphidromous species is Sicyopterus japonicus which occupied 24%. The remaining 5 species of gobioid larval fishes can not be found either freshwater or brackish region in adult, which may be a coastal species in other specific habitat. Current research reveals that great utility of mtDNA sequences for recognizing the real diversity of larval fish community with great morphological difficulty and it strongly suggests the high possibility for underestimating the real larval fish diversity compared to adult gobioid fauna.

Subject: Fish

Second International Barcode of Life Conference 18-20 September 2007 80 Conference Abstracts Poster Abstracts

DEVELOPMENT OF A DNA ARRAY USING SIGNATURE OLIGONUCLEOTIDES FROM COI BARCODES FOR IDENTIFICATION AND DETECTION OF SPECIES OF PENICILLIUM SUBGENUS PENICILLIUM

WEN CHEN, Keith A. Seifert, C. André Lévesque

Biodiversity, Agriculture and Agri-Food Canada; Department of Biology, Carleton University

Membrane-based oligonucleotide DNA arrays are a powerful technology for detecting individual species in complex environmental samples. Here, we report on the development of a CO1 DNA array, based on a previous study database of 358 sequences at the 5' end of CO1, representing 58 species of Penicillium subgenus Pencillium and 12 allied species (Seifert et al., 2007. PNAS 104:3901-6). The current project aims to employ species and clade specific oligonucleotides derived from this database for the development of a DNA array to detect and identify Penicillium species in food, feed, indoor air and soil. 182 perfect match oligos and 175 mismatch oligos (20-41 bp in length) were selected by using two softwares, Sigoli and Array Designer, respectively. The BLAST analyses confirmed that the perfect match oligos were unique within each target clade. All synthesized oligonucleotides were spotted on membranes with a microarrayer to produce the first CO1 array. The corresponding CO1 region was amplified and digoxigenin-labeled from DNA extracted from either pure culture for validation or from ‘unknown’ mixtures from environmental or food samples. These PCR labeled products are being hybridized to the array for direct detection of these species from complex samples without culturing.

Subject: Fungi

Second International Barcode of Life Conference 18-20 September 2007 81 Conference Abstracts Poster Abstracts

TOWARDS MICROARRAY-BASED DNA-BARCODING OF MARINE INVERTEBRATES

SRUJANA CHITIPOTHU1, A. Cariani2, M.N.Venugopal3, M. Landi2, M. T. M.5, J. Jaeger5, M. Kochzius1, C. Arvanitidis4, H. Weber5, M. Noelte5, F. Tinti2, A. Magoulas4, D. Blohm1

1CAG, University of Bremen, 28359 Bremen, Germany 2Interdept. Centre of Research for Environmental Sciences, University of Bologna, Italy 3College of Fisheries, Mangalore, India 4Hellenic Centre for Marine Research, Crete, Greece 5Institute for Technomathematics, University of Bremen, Germany.

Species differentiation of marine invertebrates necessary for sea food quality control and marine biodiversity research is often difficult, cumbersome and requires highly experienced specialists. To support these endeavours the suitability of microarray-based DNA-barcoding was tested for the identification of Nephtys hombergii, Lentidium mediterraneum, Alpheus glaber and 12 other difficult to classify species of annelida (4), crustacea (7) and mollusca (4) from European seas by developing a prototype of an “invertebrate-chip”. Standard kits often failed to extract high quality DNA from these animals. Among different protocols used, FTA Elute paper (Whatmann) turned out to be efficient, cost-effective and convenient for fieldwork. A combination of conserved 16S rDNA sequences and gene information of the highly variable CO1 barcoding sequence was used to design specific microarray captures by a proprietary computer programme. The resulting capture probes were experimentally evaluated for their specificity and sensitivity. Results obtained so far show that microarray-based DNA-analysis using captures designed from sequences of both genes is capable to identify at least the limited number of marine invertebrate species investigated. The limits of detection if one organism needs to be identified in the presence of large amounts of others are presently under investigation. This work was supported by the EC-FP6 grant “Fish-and-Chips” (Nr 505491)

Subject: Marine

Second International Barcode of Life Conference 18-20 September 2007 82 Conference Abstracts Poster Abstracts

ADOXOPHYES ORANA SPECIES COMPLEX (LEPIDOPTERA: TORTRICIDAE), A CONFLICT BETWEEN BARCODES AND PHEROMONES?

Lim, Eun Ji, Ik Ju Park, Hyungjin Park, and SOOWON CHO

Department of Plant Medicine, Chungbuk National University, Republic of Korea

Adoxophyes orana (Lepidoptera: Tortricidae) is famous not only in its notorious pest status but also in its species complexity due to the morphological similarities with its adjacent species. In Japan, many studies of the sex pheromones, morphology and ecology of Adoxophyes have been done for the last several decades, and currently, the species in apple orchards is classified as A. orana while that in tea plantations is classified as A. honmai plus a third species, A. dubia (Yasuda, 1998). However, the specific distinctions and definitions within the complex have still been obscure.

In 2005, the Korean Adoxophyes species was studied and concluded that there are at least two species, A. orana and A. honmai (Lee, 2005). In the meantime, on the same year, a pheromonal study on Adoxophyes found a third species possible, similar to a Taiwanese species, based on the pheromone component ratio difference (Yang, 2005). Here we used COI barcoding region to compare the Adoxophyes species and found A. honmai is same as A. dubia. In addition, A. orana and the third species, suggested by the pheromonal study, are the same species, too. This is a conflict between the barcode and pheromone, and the problems and solutions are discussed.

Subject: Insects

Second International Barcode of Life Conference 18-20 September 2007 83 Conference Abstracts Poster Abstracts

MITOCHONDRIA DATA OF ANCIENT GENUS-SIDERASTREA IMPLIED THE IMPORTANCE OF TAXONOMY REVISION IN SCLERACTINIA

YAOYANG CHUANG1, Ching-Chi Tseng, Hironobu Fukami, David Obura, Michel Claereboudt, Chang-Feng Dai, Nancy Knowlton, Chaolun Allen Chen

1 Research Centre for Biodiversity, Academia Sinica, Nangang, Taipei, 115, Taiwan

Taxonomy is an important issue in conservation of marine biota. Previous studies indicated that conventional taxonomy may contribute to the misleading of corals species and obscure the truly evolutionary history of corals. In this study, we surveyed the species phylogeny and evolution of the global-distributed corals genus-Siderastrea as a model to highlight the problems of molecular evolution and species identification in Siderastrea.

Siderastrea, an ancient genus that has existed in the Atlantic and Indo-West Pacific (IWP) Oceans at least 54 million years, has five species sporadically spreading over the world according to previous taxonomy. The results of mitochondrian phylogeny implied that 3 major lineages which connecting to the geographic distribution of 3 oceans province were inferred among the whole genus and rare divergence within lineages. Relatively deep divergences were identified in Pacific lineage to Indian and to Atlantic lineages (with cytB distance as 6.75% to Indian and 5.97% to Atlantic). The results inferred that the 3 lineages have an ancient origin in the evolution history. Closed relationship between Pacific Siderastrea and Pseudosiderastrea also suggested that the taxonomy in these two genera needs revision further.

Subject: Marine

Second International Barcode of Life Conference 18-20 September 2007 84 Conference Abstracts Poster Abstracts

RAPID TREE CONSTRUCTION FOR DNA BARCODING USING A SIMPLE CORRELATION ANALYSIS WITHOUT SEQUENCE ALIGNMENT

KA HOU CHU1, 2, Minli Xu2, Chi Pang Li1

1Department of Biology, The Chinese University of Hong Kong 2Molecular Biotechnology Programme, The Chinese University of Hong Kong

Sequence alignment is the rate-limiting step in constructing profile trees for DNA barcoding purpose. In a recent study, we demonstrated the feasibility of using unaligned rRNA sequences as barcodes based on a compositional vector (CV) approach without sequence alignment (Bioinformatics 22:1690). Here, we further explored the grouping effectiveness of the CV method in large barcode datasets (18S, 16S and COI) from a variety of organisms, including birds, fishes, nematodes, and crustaceans. Groupings of taxa at genus/species levels based on the CV method are always in good agreement with trees generated by traditional approaches, although in some cases the relationships among groups might differ. Moreover, the CV method could reduce analyzing time by over one hundred fold, which is critical in analyzing large barcode datasets. For instance, alignment of 754 COI sequences (average length of 649 bp) from fishes took more than ten hours to complete, while the whole process using the CV method required no more than five minutes on the same computer. We conclude that CV method is a fast and reliable method for analyzing large datasets for barcoding purpose.

This work was fully supported by a grant (CUHK4419/04M) from the Research Grants Council of HKSAR, China.

Subject: Methods

FROM BANKS TO BARCODES: BARCODING A SPECIES RICH, HISTORICALLY IMPORTANT AND GEOGRAPHICALLY CONSTRAINED FLORA FROM AUSTRALIA

DARREN CRAYN and Michael Whitehead

National Herbarium of New South Wales, Botanic Gardens Trust, Sydney 2000 Australia

Plant DNA barcoding efforts lag those for other groups. To date only one published study has barcoded a geographically constrained flora. Here we report on an ongoing study of the Kurnell Peninsula flora, a rich, diverse flora of c. 340 native vascular species (193 genera, 81 families) located within metropolitan Sydney, Australia. The flora of Kurnell’s c. 450 hectares of coastal dune scrub, heath, wetland, rainforest and sclerophyll woodland is well-known, of manageable size for barcoding and well represented in herbaria (for assessing preserved versus fresh material). Also, Kurnell’s was the first Australian flora studied scientifically by Europeans, namely Joseph Banks and Daniel Solander, botanists on Cook’s first voyage (in 1770). To date, c. 100 vascular species have been provisionally barcoded for 3 loci (accD, rpoB, rpoC1), with another 60+ barcoded for one or two loci. The other four loci being investigated by the Plant Working Group of CBOL (trnH-psbA spacer, matK, ycf5 and ndhJ), showed relatively poor amplification success in early, limited trials. These problems are probably related to PCR optimisation and primer specificity. Preliminary analysis of these barcode data, as well as prospects for developing plant DNA barcoding in Australia will be discussed.

Subject: Plants

Second International Barcode of Life Conference 18-20 September 2007 85 Conference Abstracts Poster Abstracts

DNA BARCODING OF BITING INSECTS FROM NORTH AMERICA AND BEYOND

ALINA CYWINSKA1, Fiona. F. Hunter1, and Doug Currie2

1Dept. of Biological Sciences, Brock University, Ontario, Canada, 2Royal Ontario Museum, Toronto, Canada.

Systematic COI barcoding of biting insects was initiated in 2005 as a part of the Canadian Barcoding of Life Network - the first national programme dedicated to large-scale DNA species identification. Biting insects are well-known worldwide pests of birds and mammals, vectors of diseases and stress factors to their hosts through their bloodsucking activities and resultant allergic reactions. Currently we are focusing on groups of biting flies with relatively well established taxonomy, such as, mosquitoes, black flies, and tabanids. So far, 154 black fly species in 12 genera, represented by almost 1600 individuals have been barcoded. This is 60% of 255 simuliid species known from N. America and 9% of 1777 formally described species from all over the world. Samples come from N. America, S. America, Europe, Iceland, and New Zealand. The N. American mosquito is represented in our barcoding data base by around 500 sequences from 72 species and 13 genera. Additionally, 75 barcodes for 29 species of deer flies, horse flies and stable flies have been obtained. Mitochondrial COI works well for discrimination of most biting insect species, including closely related taxa. Previous research indicates that the origin of cytoforms in simuliids as a result of adaptation to different ecological conditions of larval habitats could ultimately lead to speciation. We are using a combination of the barcoding method of species identification along with morphological and cytological analyses of the giant polytene chromosomes found in simuliid larvae to revise taxonomy of certain species in black fly complexes. We are also developing a method that would allow us to extract genomic DNA from tissues preserved in Carnoy's fixative for karyotyping.

Subject: Insects

Second International Barcode of Life Conference 18-20 September 2007 86 Conference Abstracts Poster Abstracts

MONITORING MOTH DIVERSITY IN CANADA’S FORESTS: THE APPLICATION OF DNA BARCODING

JEREMY R. DEWAARD1, and Leland M. Humble1,2

1Pacific Forestry Centre, Canadian Forest Service, Natural Resources Canada Victoria, BC, Canada 2Department of Forest Science, Faculty of Forestry, University of British Columbia, Vancouver, BC, Canada

Monitoring biodiversity hinges on determining what taxa exist in a given location and detecting changes over space and time. Unfortunately, the subtle morphological characters that separate species often demand expert interpretation, forcing studies to limit their taxonomic scope, or to identify specimens to higher taxonomic categories only. DNA barcoding can circumvent these limits by transforming the often lengthy and tedious chore of identifying specimens to a rapid, accurate and unbiased task. Moreover, in addition to allowing the discrimination of species and quantification of richness and abundance, the sequence data can also provide valuable measures for other levels of biodiversity often neglected, namely genetic and phylogenetic diversity. DNA barcoding thus opens the door to a more holistic approach to measuring biodiversity. We describe an ongoing project that is combining a barcode library, a multi-gene phylogeny, and planned field trials to monitor three levels of geometrid moth diversity in British Columbia, Canada. While our current work is focussed on assessing the community effects of natural and anthropogenic forest modifications, we will also discuss how our approach can be applied to other facets of forest conservation and management, as well as its amenability to arrays and emerging technology.

Subject: Insects

Second International Barcode of Life Conference 18-20 September 2007 87 Conference Abstracts Poster Abstracts

NEW SPECIES AND NOVEL BIOLOGICAL INSIGHTS DISCOVERED VIA BARCODE ASSISTED TAXONOMY OF ARGENTINEAN MARINE FISHES

JUAN M. DIAZ DE ASTARLOA

Departamento de Ciencias Marinas, Facultad de Ciencias Exactas y Naturales, Universidad Nacional de Mar del Plata. Funes 3350. B7602AYL, Mar del Plata, Argentina.

Stimulated by an invitation to join the inaugural FISH-BOL workshop in 2005 [at the University of Guelph, a barcode sample protocol was subsequently adopted, resulting in the collection of over 350 voucher specimens that were accompanied by tissue samples, photographs and geospatial coordinates. From these, a total of 327 unambiguous bidirectional barcode sequence records were generated for a total of 87 species of marine fishes from Argentinean waters. The material has been collected from several research cruises along the Argentinean continental shelf, considering the Magellan Zoogeographic Province, characterized by the incursion of cold and cold-temperate waters from the south with high productivity, and the Argentine Zoogeographic Province which is characterized by warm and warm-temperate waters. All the surveys have been conducted by the INIDEP, the National Fisheries Institute in Argentina, which its main purpose is the formulation and execution of research programs on basic and applied investigation related to fisheries resources and their sustainable exploitation. The results demonstrate that DNA barcodes are effective in discriminating all species surveyed. The molecular assisted recognition of cryptic diversity (highlighted by the existence of deep genetic divergences within taxa), fueled subsequent efforts in integrative alpha taxonomy. Result included the description of a new species of Longnose skate (Dipturus argentinensis) and the recognition of the Brazilian cusk eel (Genypterus brasiliensis) as a valid species, which Neilsen, Cohen, Markle and Robins regarded as an unpatterned form of the pink-cusk eel (Genypterus blacodes) in their FAO catalog of Ophiidiform species. Additionally, the pattern of diversification implied by the molecular data among eight Bathyraja species implies the presence of a phylogenetic constraint acting on the physiological plasticity of these species with respect to their differing habitat preferences.

Subject: Fish

Second International Barcode of Life Conference 18-20 September 2007 88 Conference Abstracts Poster Abstracts

DETECTION OF MARINE BIOINVADERS IN A MEDITERRANEAN INDUSTRIAL PORT BY DNA BARCODING.

Catherine Vogler, DIRK ERPENBECK, Gert Wörheide,

Dept. of Geobiology, University of Göttingen

The introduction of non-indigenous species (NIS) into the Mediterranean Sea has been ongoing since centuries. As a hub of steadily increasing commercial shipping, and encircled by major ports, it is very susceptible to the introduction of ship-borne organisms. Ballast water discharges and hull fouling are recognised as major vectors of NIS, and consequently commercial ports are often the point of inoculation. Moreover, the extent to which invertebrate NIS have invaded the Mediterranean is not entirely known. To minimise these species' invading potential and their spread into neighbouring costal areas, discovering them as early as possible in their progression is essential, but often hindered through their difficult identification and the problem posed by morphologically hard to distinguish cryptic invaders. In this study we target the port of Marseille (France), to test the applicability of DNA assisted identification techniques (DNA barcoding and taxonomy) coupled with morphological methods, to detect potential marine invaders. These techniques will subsequently be taken further by applying them not only to invertebrate communities in ports, but also to ballast water, characterising its biotic contents by using similar molecular techniques, greatly facilitating the identification of otherwise taxonomically hard to determine larval stages.

Subject: Marine

Second International Barcode of Life Conference 18-20 September 2007 89 Conference Abstracts Poster Abstracts

CLEANING UP THE MESS - APPLIED DNA TAXONOMY OF SPONGES WITH THE SPONGE BARCODING DATABASE (SBD, WWW.SPONGEBARCODING.ORG)

DIRK ERPENBECK1,2, John N.A. Hooper2, Steve C. de Cook3, Judith Pöppe1, Gert Wörheide1

1Dept of Geobiology, University of Göttingen, Germany 2Biodiversity Program, Queensland Museum, South Brisbane, Queensland, Australia 3Auckland University of Technology, Auckland, New Zealand

Sponges are among the most important marine producers of secondary metabolites with demonstrated antibiotic, anticancer or otherwise pharmaceutically interesting properties. Additionally they are among the first branching Metazoa and pivotal for the study of animal evolution. Unfortunately, they simultaneously belong to the most difficult to identify metazoan taxa. This is due to their primitive nature of morphological characters and their frequent lack of complexity. Sponge morphological systematics is almost entirely focussed on mineral skeleton features. Those sponges without such mineral skeleton elements provide problems even to the taxonomy expert. Slight misinterpretation of morphological features can lead to classification of a keratose sponge sample to a different order or family and therefore extremely influence the results of biochemical or evolutionary studies negatively. To aid in circumventing such problems, the Sponge Barcoding Project (www.spongebarcoding.org) and the Sponge Barcoding Database have recently been established with the aim to facilitate sponge species identification using DNA barcoding techniques. Here, we present first results of a successful application of sponge barcoding, ranging from the re-classification of higher keratose sponge taxa to unravelling cryptic species in a species complex of keratose sponge genera of particular interest for pharmaceutical and biochemical purposes.

Subject: Marine

Second International Barcode of Life Conference 18-20 September 2007 90 Conference Abstracts Poster Abstracts

THE SPONGE BARCODING PROJECT (SBP) AND THE SPONGE BARCODING DATABASE (SBD) - AIDING IN THE IDENTIFICATION AND DESCRIPTION OF PORIFERAN TAXA (WWW.SPONGEBARCODING.ORG)

Gert Wörheide1,DIRK ERPENBECK1,2

1Dept of Geobiology, University of Göttingen, Germany 2Biodiversity Program, Queensland Museum, South Brisbane, Queensland, Australia

Sponges are among the most ancestral metazoans and notoriously difficult to identify, even by taxonomic experts due to their depauperate suite of complex morphologial characters. However, as a group they are highly diverse, ecologically important and of significant commercial importance to the pharmaceutical and biomaterials industry as producers of highly potent secondary metabolites. Therefore, means of unambiguous identification are urgently needed. Sponge DNA- barcodes will provide a set of indispensable tools to aid taxonomists and ecologists in the identification of sponge species, and will enhance the discovery of drug-producing species.

The Sponge Barcoding Project (SBP) is the first worldwide barcoding project on any diploblast taxon, and covers the complete taxonomic range of Porifera. The Sponge Barcoding Website (www.spongebarcoding.org) has been set up to serve the project's aims and provide a centralized platform for data exchange. Its core component is the Sponge Barcoding Database (SBD), which is developed with the aim to function as the primary access point for DNA signature sequences together with providing information on conventional morphological taxonomic characters to aid species discovery, description and characterization. The unique combination of sponge-specific conventional taxonomic information and DNA signature sequences is the distinguishing feature, in which the SBD differs from other database systems, such as Genbank or the Barcode of Life Data Systems, with which the SBD is cross-linked.

Subject: Marine

Second International Barcode of Life Conference 18-20 September 2007 91 Conference Abstracts Poster Abstracts

DNA BARCODING OF PESTS AND INVASIVE SPECIES: APHIDS, ADELGIDS AND SCALES

ROBIN FLOYD1, Leland Humble2, Robert Foottit3, Eric Maw3, Paul D.N. Hebert1

1Biodiversity Institute of Ontario, University of Guelph 2Canadian Forest Service, Natural Resources Canada, University of British Columbia 3Agriculture and Agri-Food Canada, Ottawa

DNA barcoding promises to enable rapid species identification using a standard methodology by non-taxonomic specialists. One of the areas in which such a technology could potentially have the greatest benefit to society is in the monitoring of harmful species, such as agricultural and forestry pests, where there is often a pressing need for rapid and reliable species diagnoses—often from eggs or larval life-stages which are difficult or impossible to identify by morphology. The insect order Hemiptera (true bugs and their allies) contains many species of significant pest status for gardeners, agriculture and forestry, including the aphids (superfamily Aphidoidea), adelgids (superfamily Adelgoidea) and scale insects (superfamily Coccoidea). We are in the process of generating barcode sequences from aphids, adelgids and scales collected from across North America, and have generated sequences from over 2000 specimens, covering nearly 600 species. This presentation will summarise progress on these, and some smaller projects also relating to North American pest species (Lygus plant bugs; bark beetles). This data will facilitate species diagnosis in the future, as well as providing valuable insight into biogeographic and population genetic patterns in these economically important animal groups.

Subject: Insects

BARCODING DINOFLAGELLATES USING COI AND ITS

JILLIAN FERRELL and Margaret Beaton

Mount Allison University, Canada

Dinoflagellates are a morphologically diverse group of protists encompassing more than 2000 extant species. These microscopic organisms represent an important group from both biological and economic perspectives, yet their taxonomic classification remains challenging. The use of barcoding to delineate species of this group could represent a powerful taxonomic tool and aid in clarifying evolutionary relationships. To test the efficacy of COI as a suitable barcode for unambiguous species identification in dinoflagellates we focused on the genus Prorocentrum of which some members are known to be toxic. Sequence comparisons among 76 strains, representing 13 species, typically revealed that conspecific divergence ranged from 0-1% (0-7 substitutions) with a mean of 0.56±0.05%. At the congeneric level the divergence was on average much higher (3.59±0.04%), and although as many as 64 substitutions were found, certain taxa only differed by a single nucleotide. Preliminary results suggest that COI may provide adequate resolution at the genus level, but due to its low resolving power for a few closely related species, a second genetic marker will likely be required. This study tested the utility of the ITS region to complement the COI data and revealed substantial divergence allowing the unambiguous discrimination of all tested species.

Subject: Protists

Second International Barcode of Life Conference 18-20 September 2007 92 Conference Abstracts Poster Abstracts

MOLECULAR PROBES: A TOOL TO IDENTIFY MARINE INVERTEBRATE LARVAE

CHETAN A. GAONKAR, Arga Chandrashekar Anil

National Institute of Oceanography, Dona Paula, Goa- India.

Species specific information on marine invertebrate larvae is vital for understanding their ecology. In the recent years translocation of organisms through ship’s ballast water has gained importance in marine bio-invasion. Identification of organisms within the ballast tank and the discharge ballast is important in ballast water management efforts.

In view of this, efforts are underway to develop molecular probes for the larvae of barnacle (Balanus amphitrite) and bivalves (Mytilopsis sallei, Perna veridis and Crassosstrea graphoides). Adult specimens were collected in sufficient quantity for DNA extraction and sequencing. Zooplankton samples were also collected from Mandovi and Zuari estuaries of Goa (West Coast of India) from 8 different stations, over a period of 2 years on a bi-monthly basis. These samples were analysed microscopically to group levels. Efforts are underway to design species specific probes for comparison.

Subject: Marine

DNA BARCODE PHYLOGENY IDENTIFIES DUAL ORIGINS OF CLEPTOPARASITIC BEHAVIOUR IN THE BEE SUBGENUS DIALICTUS (HALICTIDAE: LASIOGLOSSUM)

JASON GIBBS

York University, Toronto, Ontario, Canada

DNA barcodes are an effective tool for integrative approaches to bee taxonomy and have identified numerous cryptic species. Although, primarily used for species identification the utility of DNA barcodes for phylogenetics should be explored. Although more robust phylogenies are desirable, the availability of DNA barcode sequences allows for rapid testing of standing evolutionary hypotheses and can aid in the formulation of new ones. The bee subgenus Dialictus (Halictidae: Lasioglossum) displays an incredible array of social systems and is an ideal group for studying the evolution of social behaviour. Solitary, communal, primitively eusocial and parasitic Dialictus species have been identified. Parasitic Dialictus are social parasites of other Dialictus species. A phylogenetic analysis of North American Dialictus based on DNA barcode sequences shows for the first time that parasitic Dialictus are polyphyletic. Each parasitic lineage of Dialictus appears to have evolved according to the loose form of Emery's rule. The utility of DNA barcodes for phylogenetics and the evolution of social parasitism in Dialictus are discussed

Subject: Insects

Second International Barcode of Life Conference 18-20 September 2007 93 Conference Abstracts Poster Abstracts

IDENTIFICATION OF TRADITIONAL CHINESE MEDICINE RADIX ASTRAGALI BY DNA BARCODING

HAIYAN GUO, Weiwei Wang, Qishi Sun, Jun Yu

Beijing Genomic Institute, Chinese Academy of Sciences, China

DNA barcoding has been proved to be more accurate and efficient for identification of animal and plant species based on standard region of COI gene. It has never been applied to identification of traditional Chinese medicine. In this study, we propose to evaluate authentication of traditional Chinese medicine using DNA barcoding through phylogenetic study on Radix Astragali in China and distinguishing it from adulterants with suitable biomarkers. Since approximate 90% of Chinese medicine for clinical therapy is botanical materials and their adulterants are usually found under the same genus and species, we plan to screen out effective biomarkers from five candidate genes including cox1, matK, rbcl, trnL-F, and ITS. We extracted total DNA from 54 samples covered 11 families including 17 Radix Astragali with both leaves and dried roots. We observed average K2P distance of 0.039, 0.005 and 0.002 separately within families, genus and species based on of cox1 gene, while ITS and matK were more effective markers and ensured well-separated phylogenetic trees. Moreover, two significant deletions were found in 5’ region of matK gene between Radix Astragali and adulterants which made it a promising marker for qualification of Chinese medicine.

Subject: Plants

GENETIC RELATIONSHIP AMONG FOUR SUBSPECIES OF MASU SALMON COMPLEX

JIN-CHYWAN GWO and Te-Hua Hsu

Department of Aquaculture, Taiwan National Ocean University, Keelung 20224, TAIWAN

Of the seven species of Pacific salmon (genus Oncorhynchus), four masu salmon (Oncorhynchus masou) complex are native to Far East Asia. Three subspecies occur in Japan: masu (Oncorhynchus masou masou), Amago (O. masou ishikawae) and Biwa (Oncorhynchus masou subsp.) salmon. The Formosan landlocked salmon (Taiwan salmon), Oncorhynchus masou formosanus (Jordan and Oshima 1919), is the fourth subspecies of this masu salmon complex, and located in the southern geographical limit of masu salmon complex. Historically, the taxonomy and nomenclature of masu salmon complex has been greatly debated among researchers, partly due to the similarities in morphological and ecological traits shared by all four subspecies. The scientific name and systematic position of the Formosan landlocked salmon especially has been an exceptional subject of controversy - currently its scientific name has been changed four times. Here, a DNA fingerprinting method, amplified fragment length polymorphisms (AFLP) was applied to clarify the genetic relationship of these four subspecies of masu salmon complex. After examining near thousand of polymorphisms distributed across the genome, we propose that masu, Amago, Biwa, and the Formosan landlocked salmon (Taiwan salmon) should be grouped as four distinct subspecies.

Subject: Fish

Second International Barcode of Life Conference 18-20 September 2007 94 Conference Abstracts Poster Abstracts

IBARCODE.ORG - ANALYTICAL AND VISUAL TOOLS FOR DNA BARCODING

MEHRDAD HAJIBABAEI1, Greg Singer2

1Canadian Centre for DNA Barcoding, Biodiversity Institute of Ontario, University of Guelph, Guelph, Ontario, Canada 2Human Cancer Genetics Program, The Ohio State University, Columbus, OH, USA iBarcode.org is a web-based application server that provides visualization and analysis tools for barcoding data. The services are free and do not require registration. Below, we outline the first such tools that we have developed.

HapMap: This data visualization module provides a graphical view of the nucleotide character variation in a barcode data set. It allows the user to quickly pinpoint nucleotide positions within the barcode sequence that account for barcode variation in a set of species.

Haplotype Variation Analysis: This tool identifies unique haplotypes for each species in a set of barcode sequences and provides statistical information on haplotype frequency and nucleotide variation in a table format.

Between- vs. Within-species Sequence Variation: For each species with 3 or more individuals, this tool plots maximum Within Species Divergence against minimum Between Species Divergence.

Tree Collapse: This tool uses bootstrap values as a benchmark for visualizing statistical support of a given barcode-based tree. This is done by collapsing all the branches that are supported by bootstrap values smaller than a user-selected cut-off value.

Tree Tip colorization: This tool takes a Newick tree and plots it, colorizing the branches leading to individuals of each species, allowing one to visually inspect the robustness of species- assignments based on the barcode sequences.

Although this original suite of tools has been developed based on our own analytical needs, we encourage feedback from the DNA barcoding community, and in particular we would like suggestions for new tools. In the future iBarcode.org will have an API that will allow users to plug their own analysis modules into the system.

Subject: Methods

Second International Barcode of Life Conference 18-20 September 2007 95 Conference Abstracts Poster Abstracts

UNMASKING HIDDEN BUTTERFLY DIVERSITY WITH DNA BARCODES: PUTATIVE SPECIES OF NYMPHALIDAE

John J Wilson1, MEHRDAD HAJIBABAEI1, Daniel H Janzen2, Winnie Hallwachs2, Paul DN Hebert1

1Canadian Centre for DNA Barcoding, Biodiversity Institute of Ontario & Department of Integrative Biology, University of Guelph, Guelph, Ontario, Canada 2Department of Biology, University of Pennsylvania. Philadelphia, PA 19104, USA

Butterflies are among the most conspicuous invertebrates in terrestrial ecosystems and represent a benchmark group for the assembly of an "encyclopedia of life". We are completing a taxonomic inventory of the Lepidoptera of the Area de Conservacion Guanacaste, a reserve in Costa Rica with an estimated 9,600 species of non-leaf-mining lepidopterans. The Nymphalidae make an abundant and attractive component of this diversity, comprising the majority of the Costa Rican butterfly fauna. DNA barcode sequence (COI mtDNA) patterns have flagged numerous records of potential species unknown to science. Distinctive barcode clusters, often supported by ecological or morphological characters, can be used as the basis of "primary" species hypotheses. In this study we test these hypotheses through genealogical concordance with other molecular sequences. If these clusters represent phylogenetic species and not tokogenetic structure, then this should be corroborated by diagnostic characters in sequences from the nuclear genome.

Subject: Insects

Second International Barcode of Life Conference 18-20 September 2007 96 Conference Abstracts Poster Abstracts

HIGH THROUGHPUT BARCODING OF FUNGAL SPECIES: A FEASIBILITY ANALYSIS

Isabelle Meusnier1, Pedro Antunes1, Magali Solé2, Daniel Royse3, Alison Fischer1, Tiarella Hanna1 and MEHRDAD HAJIBABAEI1

1 Canadian Centre for DNA Barcoding, Biodiversity Institute of Ontario, University of Guelph, Guelph, Ontario, N1G 2W1 Canada 2 Helmholtz Centre for Environmental Research, Lepzig, Germany 3 Department of Plant Pathology, Pennsylvania State University, University Park, PS, USA

A 648bp segment of mitochondrial cytochrome c oxidase 1 (CO1) has been extensively used as a DNA barcode for animal species identification and discovery. However, the applicability of this gene as a barcode for the fungal kingdom has not been fully evaluated. Factors such as the presence of nuclear pseudogenes and intronic regions in the mitochondrial genes of fungi can potentially complicate the use of CO1 as a DNA barcode in this kingdom. Here we test the feasibility of a high throughput barcoding system for fungi by using COI and two ribosomal loci, LSU and ITS. We designed 2 sets of primers based on available CO1 sequences from Ascomycota, Basidiomycota, and Zygomycota. One set was aimed at a short fragment (110bp), in the 5’ region of COI barcode, thereby avoiding probable intronic sequences. The other primer set targets the entire CO1 barcoding region. We tested these sets of primers on Basidio and Ascomycetes collected in 2 natural communities of Northern Ontario (Thunder Bay) and Manitoba (Churchill) with 282 and 182 specimens, respectively, and on 38 Agaricus, 30 Hyphomycetes and 8 Glomeromycota specimens from fungal collections. CO1 results were compared with ITS and LSU sequences. Our results suggest that short COI fragments are much more robustly recovered from fungi. However, perhaps due to the presence of introns and limitations associated with primer design, the effectiveness of long primers varied across groups. These preliminary results indicate that CO1 may be used in the future for species identification across the kingdom fungi. Nevertheless, barcode recovery can be enhanced by designing specific primers for each phylum and by using RNA as template in RT-PCR system to avoid intronic sequences within barcode region.

Subject: Fungi

Second International Barcode of Life Conference 18-20 September 2007 97 Conference Abstracts Poster Abstracts

BARCODING EPHEMEROPTERA, PLECOPTERA, AND TRICHOPTERA OF NORTH AMERICA

Xin Zhou1, MEHRDAD HAJIBABAEI1, Tim Pascoe2, Donald Baird3 and Paul Hebert1

1 Canadian Centre for DNA Barcoding, Biodiversity Institute of Ontario, University of Guelph, Guelph, Ontario, Canada N1G 2W1. 2 Environment Canada, Canada Centre for Inland Waters, 867 Lakeshore Road, Burlington, Ontario, Canada L7R 4A6 3 Canadian Rivers Institute, Department of Biology, 10 Bailey Drive, PO Box 45111, University of New Brunswick, Fredericton, New Brunswick, Canada E3B 6E1

The three insect orders Ephemeroptera (mayflies), Plecoptera (stoneflies), and Trichoptera (caddisflies), or “EPT”, are aquatic macroinvertebrates commonly encountered in various freshwater habitats. Because of their great diversity and abundance, they are widely used as bioindicators of water quality. However, the difficulties in species-level identification, especially of immature stages, have hindered such activity. The DNA barcoding initiative provides an alternative route in species identification with a number of advantages that complement the conventional morphological approach. For example, DNA barcoding can improve resolution on taxonomic identification by discriminating cryptic species and enabling confident association of larval and adult stages.

We conducted a pilot study to test the feasibility of high throughput barcoding in aquatic macroinvertebrates. Our initial results suggest that specimens collected by conventional methods in many bioinventory and biomonitoring programs may serve as good sources of DNA for barcoding. COI barcodes were also collected from more than 1600 EPT samples collected from Churchill, Manitoba, Canada, from 2004 to 2006, and from the Great Smoky Mountains National Park, USA in 2007. These specimens cover a variety of species and life stages (male and female adults, pupae, and larvae or nymphs). COI has shown its great potential in recognizing EPT species and associating life stages.

Subject: insects

Second International Barcode of Life Conference 18-20 September 2007 98 Conference Abstracts Poster Abstracts

A TWO-PRONGED MOLECULAR APPROACH

Jason Cooper1 , Natalia V Ivanova2, Steve King1, Greg Sykes1, Karin Cottrill1, Pranvera Ikonomi1, ROBERT HANNER2

1ATCC (American Type Culture Collection), 10801, University Blvd. Manassas, Va. 20110 2Biodiversity Institute of Ontario, University of Guelph, Guelph, Ontario, N1G 2W1

Species identification and detection of inter-species cross contamination in cell culture is crucial for scientific research. Traditional isoenzymological identification tools suffer from a limited taxonomic scope and are not easily replicated among laboratories. Here we describe a two-pronged molecular approach taking advantage of nucleotide sequence variation in the mitochondrial cytochrome c oxidase subunit I (COI) gene. First, we designed species-specific primers to rapidly identify the most common model species in culture. The primer sets are designed to produce size- specific amplicons for each species, resulting in a multiplex PCR-based screening assay that can be used to detect cross-contamination of cultures involving these species. Second, for less commons species, we evaluated a sequence-based approach for identification targeting the barcode region of COI. Identifications were made using the Barcode of Life Data System (BOLD), which contains reference sequences derived from morphological voucher specimens. Taxonomic authentication is achieved for cultures that yield an appropriate match. We accurately identified 67 cell cultures representing 44 distinct animal species with this method. Similarly, cultures of known provenance can be barcoded and added to BOLD, serving as standard reference materials (e.g. as controls in forensic analysis) and thereby enhancing their marketability. We recommend this two-pronged approach, now implemented at ATCC, to rapidly and cost-effectively reduce the problems of misidentification and inter-species cross-contamination in cell culture.

Subject: Methods

Second International Barcode of Life Conference 18-20 September 2007 99 Conference Abstracts Poster Abstracts

BARCODING DEEP SEA FISHES FROM THE NORTHERN MID ATLANTIC

Ingvar Byrkjedal1, Endre Willassen1, and ROBERT HANNER2

1Dept Natural History, Bergen Museum, University of Bergen, P.O. Box 7800, N-5020 Bergen, 2Biodiversity Institute of Ontario & Department of Integrative Biology, University of Guelph, Guelph, ON N1G 2W1, Canada

Under the auspices of Census of Marine Life (CoML), the project MAR-ECO conducted a trawling cruise in summer 2004 with the R/V G.O.Sars, supplemented by the longliner M/S Loran, to study marine life along the northern Mid Atlantic Ridge. Pelagic sampling and bottom trawling were undertaken down to about 3600 m depth. In excess of 60,000 individual fish were collected, representing about 300 species identified from morphology by international specialists. About 3,000 voucher lots were deposited in the collections of the Bergen Museum in Norway. A selection of tissue samples from this collection, including 319 specimens representing 127 species, was successfully sequenced in collaboration with the Canadian Centre for DNA Barcoding (CCDB) to date. The species are generally well segregated into distinct K2P clusters, even in species rich groups like the Myctophiformes (27 spp) and Stomiiformes (19 spp). Test searches with the BOLD identification engine suggest that North Atlantic species are distinct from other congeners in the East and West Pacific that have been barcoded by other research teams. The sequencing results did, however, reveal taxonomic problems in some families, including the Alepocephalidae (Osmeriformes), now under taxonomic revision. The resulting COI sequences and trace files were submitted to GenBank and received annotation with the reserved keyword BARCODE. They represent a significant contribution to the international Fish Barcode of Life (FISH-BOL) initiative.

Subject: Fish

Second International Barcode of Life Conference 18-20 September 2007 100 Conference Abstracts Poster Abstracts

DIFFERENTIATING BETWEEN ERROR AND INFORMATION IN DNA SEQUENCE DATA

Taika von Königslöw1, Dan Ashlock2, PAUL HEBERT1,

1Department of Integrative Biology, University of Guelph, Ontario, Canada 2Department of Mathematics and Statistics, University of Guelph, Ontario, Canada

The Barcode of Life Data System is a publicly accessible online database of cytochrome c oxidase subunit I (COI) mitochondrial DNA sequences that are collected for a global identification system for animal life. The technique of identifying a specimen to species level by matching its COI sequence to similar or exact COI sequences in the database is the central theme in DNA barcoding. It is sometimes the case in DNA barcoding that species that have recently diverged are differentiated by a very small number of differences, sometimes as few as 1-2 base pairs out of the approximately 650 found in their DNA barcode. Unfortunately this small fraction of differences can fall within the range of sequencing reaction error. We propose to address this problem by building a profile hidden Markov model. This model would estimate the probability of variation at a site given the sequence diversity of the related sequences from which the model was built. This information would complement the error probability estimates assigned to the raw sequence data (Phred scores) which are unbiased by sequence history but cannot always correct for systematic error or random error introduced in areas of single-strand coverage.

Subject: Methods

BARCODING BATS: PROGRESS ON GLOBAL AND REGIONAL CAMPAIGNS

Elizabeth L. Clare, and PAUL HEBERT

Department of Integrative Biology, University of Guelph, Ontario, Canada

Bats are the second largest order of mammals but with fewer than 1300 described species, the campaign to collect DNA barcodes for all bats has made significant progress providing insights into the patterns we expect to see among mammals. The dataset encompass >11, 000 barcode sequences from >450 species representing diverse geographic locations. The initiative has been carried out by chiropteran taxonomists from several institutions worldwide, and coordinated at the Biodiversity Institute of Ontario in Canada. Here, I will outline the results of barcode collection emphasizing the Neotropical regions of Central America and northern South America. I will provide details on the resolution of alpha-level taxonomy and phylogeographic patterns of genetic diversity including recent work on spatial sequence analysis and I will examine the results of a detailed investigation of DNA barcode variation in 87 species of bats from Guyana. Ninety-three percent of these species showed low intraspecific variation (mean = 0.60%), and clear sequence divergence from their congeners (mean = 7.80%). The remaining 7% show intraspecific lineages with up to 14% COI sequence divergence between them. The present study strongly supports the effectiveness of DNA barcoding as an identification tool for bat species.

Subject: Mammals

Second International Barcode of Life Conference 18-20 September 2007 101 Conference Abstracts Poster Abstracts

BARCODING BIRDS: PROGRESS TOWARDS A GLOBAL PERSPECTIVE

Kevin C.R. Kerr1, DarÌo A. Lijtmaer2, Pablo L. Tubaro2, Sharon Birks3, Mikhail Kalyakin4, Carla J. Dove5, and PAUL HEBERT1.

1University of Guelph 2Museo Argentino de Ciencias Naturales 3The Burke Museum 4Zoological Museum of Moscow University 5Smithsonian Institution

Avian taxonomy is arguably the most robust of any class of animals; hence, birds offer a unique test for DNA barcoding. A subsample of North American birds was used as an early indicator of the efficacy of barcode-based species delimitation. Promising results aroused interests as well as critical commentaries. An international effort ñ the All Birds Barcoding Initiative ñ was established shortly thereafter to co-ordinate efforts to barcode the worldís avifauna. We have expanded the sampling of North American birds creating comprehensive coverage of breeding species. Furthermore, we have generated DNA barcodes for birds of Argentina through collaboration with the Museo Argentino de Ciencias Naturales, as well as birds of the eastern Palearctic through collaboration with the Burke Museum and the Zoological Museum of Moscow University. While data collection is ongoing, current progress is propitious. Broadly ranging species are represented from mulitple continents. Initial trends in mean intraspecific and interspecific genetic distances vary little between biogeographical realms. While a few sister species with overlapping barcode sequences have emerged, they are mostly limited to North America. Only one sequence is shared between species from different continents. Deep genetic splits within species are often predictable and tend to be limited to shy, nondescript species. These findings address several of the rudimentary criticisms of barcoding.

Subject: Birds

Second International Barcode of Life Conference 18-20 September 2007 102 Conference Abstracts Poster Abstracts

IDENTIFICATION OF IMPORTANT CRABS OF TAIWAN USING MITOCHONDRIAL COI GENE SEQUENCE AND PCR-RFLP ANALYSIS

SHENG-TAI HSIAO, Hong-Yen Liang, Jwu-Lan Lai

Marine Fisheries Division, Fisheries Research Institute, Taiwan

Species identification for fishery resources management, trading and conserving of aquatic species are very important, as a primary step, is being able to identify taxa correctly and unambiguously. Nucleotide sequences of mitochondrial cytochrome oxidase subunit I (COI) gene were compared among economically important crabs species and recognition sites of diagnostically useful restriction enzymes were examined. In this study, we also compared nucleotide divergence of full sequence of COI (1534 bp) and the partial fragment of COI (recommended by Barcode of life, 657 bp). The mean sequence divergence among genera was about 20% based on Folmer region. However, the mean sequence divergence among genera was only about 14% based on the full length of COI. The transitions of the partial COI data set were found more saturated than the full COI data set, but they were retained for analysis since they contain the majority of the phylogenetic information. The PCR-RFLP analysis using 111 individuals confirmed that 15 species could be correctly identified by cleavage patterns generated by the endonucleases TaiI and TaqI. Restriction enzyme, TaiI, allowed discrimination of ten species. PCR-RFLP is a powerful diagnostic tool for rapid and reliable identification of economically important crab species.

Subject: Marine

Second International Barcode of Life Conference 18-20 September 2007 103 Conference Abstracts Poster Abstracts

COMPARATIVE PERFORMANCE OF THE COI AND cyt b GENES IN DNA BARCODING OF FISHES

KUI-CHING HSU, Kwang-Tsao Shao

Research Center for Biodiversity, Academia Sinica

Many worldwide species can be separated into several populations or evolutionary significant units. Therefore, we need a DNA barcoding dataset to identify species, populations, or even to reveal some phylogenetic signals. Hence, a preliminary test was undertaken to compare with the two datasets, cytochrome b (cyt b) and cytochrome oxidase I (COI). The differences of COI sequences within species were close to zero, so it was not be able to be used for identifying populations. According to the base compositions within Cheiloactylidae, we found the transition /transversion ratios were 3.6 (1st =3.6, 2nd=4.3, 3rd = 2.1) and 2.9 (1st = 2.9, 2nd= 5.3, 3rd = 0.3) in average for cyt b and COI, respectively. These saturation constraints of COI would influence the relative evolution rate. Within a species or genus, the phylogenetic tree, restructured based on the cyt b sequences revealed, turned out well-resulted and had more phylogenetic information. The phylogenetic analysis within family or order, based on both the cyt b and COI gene, would take many genetic constraints, e.g. homoplasy or low rates of evolution. According to these results, we consider that the cyt b sequences can perform as good as or even better than COI sequence sets in assigning sequences to populations or species as well as supplying phylogenetic signals.

Subject: Fish

MULTIGENES MATCH MASS EXTRAORDINARY SEXUAL DIMORPHISM AND MALE POLYMORPHISM OF TWELVE FIG WASPS FROM SMALL FIG SYCONIA

Jin-Hua Xiao and DA-WEI HUANG

Institute of Zoology, Chinese Academy of Sciences, Beijing 100101

Fig and fig wasps has attracted much interest as a good model in the study of evolution. The extreme sexual dimorphism and extraordinary male polymorphism of fig wasps made the species classification especially male identification difficult and false-prone. The assistance of molecular techniques in the species identification process in this fancy species assemblage is requisite. In the present study, we take the 13 species 8 genus fig wasps associated with Ficus benjamina (Ficus, Moraceae) as the object, analyzed the four fragments of EF1-alpha;, COI, cyt b, and ITS2 for their utilization in phylogenetic studies and species identification. Our results showed that, ITS2 was the best marker used in recognition of species identity of dimorphic sexes and polymorphic males, and COI could also act well in these jobs except some little flaws or dubiousness. On the contrary, cyt b and EF1-alpha; provided ambiguous phylogenetic information in the relationship we expected. We thus discussed the indispensable utilization of ITS2 in the process of DNA taxonomy, especially for sexual dimorphic and male polymorphic groups, or in arthropod groups frequently infected by symbionts, in which mitochondrial COI sequences alone are deficient and even misleading.

Subject: Insects

Second International Barcode of Life Conference 18-20 September 2007 104 Conference Abstracts Poster Abstracts

METHODS FOR STREAMLINING HIGH THROUGHPUT DNA BARCODING

NATALIA V. IVANOVA, Chris M. Grainger, Alex V. Borisenko*

Canadian Centre for DNA Barcoding, Biodiversity Institute of Ontario, University of Guelph, 579 Gordon Street, Guelph, ON, Canada, N1G 2W1

The ability to generate DNA barcodes in a high throughput facility requires vigorous optimization of data flow and analytical protocols. In 2006-2007 the Canadian Centre for DNA Barcoding integrated automation for many steps of the laboratory analytical chain. In many cases conversion to more expensive, but reliable protocols or reagents (e.g., Platinum® Taq for PCR, E-gel® for PCR check) allowed us to cut down overall analytical costs, due to increased efficiency and reduced labor costs. Furthermore, integration of automated sequencing cleanup from Agencourt enabled reducing of BigDye® volume due to increased sensitivity. At the same time we employed various 'home-made' approaches (e.g., Glass Fiber DNA extraction protocol, pre-made frozen PCR and cycle sequencing plates) to reduce the cost of consumables and ensure operation efficiency. For increased data management requirements, an electronic lab book facilitates easy data conversion with BOLD, sequencer and liquid handling stations, while minimizing human error. Efficient operation depends on a well designed workflow. Automation aids in all these areas by reducing errors for the tedious and labor intensive pipetting and cherry-picking tasks. Although all protocols described here were optimized for a high throughput automated environment, most of them could be employed in a lower throughput barcoding lab.

Subject: Methods

BARCODING DARWIN’S MEADOW: A 21ST CENTURY BOTANICAL INVENTORY AT HISTORIC DOWN HOUSE

KAREN JAMES, Mark Spencer, Fred Rumsey, Mark Carine, Rob Huxley, Harald Schneider and Johannes Vogel

Department of Botany, The Natural History Museum, London

In 1855, Charles Darwin surveyed Great Pucklands meadow near his home at Down House. ? In an age when specimen rarity was prized above all, Darwin’s vision was radical: to identify all plant species in a small, unremarkable plot. As it did for Darwin, Great Pucklands still serves as an ideal training ground for the development of plant identification skills. In 2005-7, we re-surveyed Darwin’s meadow, aiming to 1) develop a seamless approach to collection, management and storage of morphological and molecular collections for the 21st century including high throughput DNA barcoding, and 2) measure performance of the proposed combinatorial plant barcodes (including rpoC, rpoB, matK, trnH-psbA and rbcL). We collected ~200 specimens of ~160 flowering plant species with 5-10 fold sampling density for ~5 species. We will report on amplification and sequencing success with relation to sampling method (Whatman FTA vs. silica), and discuss prospects for taxonomic identification of plants using either single or up to five loci combined. Future aims include barcode-based identification of tester and real unknowns from the meadow including mixed environmental samples. Findings and procedures will be applied to a wider suite of case studies covering the taxonomic depth and breadth of the British flora.

Subject: Plants

Second International Barcode of Life Conference 18-20 September 2007 105 Conference Abstracts Poster Abstracts

THE JAPANESE BARCODE OF LIFE INITIATIVE (JBOLI)

UTSUGI JINBO, Hiraku Yoshitake & Motomi Ito

Japanese Barcode of Life Initiative, University of Tokyo

There is increasing interest in DNA barcoding in Japan. However, it has never been promoted sufficiently to date due to the lack of a domestic stronghold. We are working on the establishment of the Japanese Barcode of Life Initiative (JBOLI; http://www.jboli.org/) which has the following goals: 1) to disseminate information on the concept, protocols, and latest issues of DNA barcoding, 2) to provide a web-based system which acts as an information and communication center for projects concerning DNA barcoding, 3) to coordinate activities among CBOL, DDBJ, institutions such as museums and herbariums, and researchers, 4) to provide a data repository and management system to store and analyze data produced by researchers, and 5) to provide an identification system for public use in Japan. Although JBOLI is a working group consisting of researchers, it is associated with GBIF, Union of Japanese Society for Systematic Biology and The National Bioresource Project. The infrastructure of JBOLI is supported by the GBIF Japan Node. In this presentation, we report on our current progress and future directions.

Subject: Initiatives

Second International Barcode of Life Conference 18-20 September 2007 106 Conference Abstracts Poster Abstracts

APPLICATION OF DNA BARCODING TO INVASIVE POPULATIONS OF LEAFMINERS, LIRIOMYZA SPP. (DIPTERA: AGROMYZIDAE) IN THE PHILIPPINES

S. J. Scheffer1, M. L. Lewis1, RAVI C. JOSHI2

1Systematic Entomology Laboratory, PSI, ARS, USDA, Bldg. 005, Rm. 137, BARC-W, 10300 Baltimore Ave., Beltsville, MD 20705, USA

2Chief Science Research Specialist, Office of the Executive Director, Philippine Rice Research Institute, Maligaya, Science City of Muñoz, Nueva Ecija 3119, Philippines

Outbreak populations of invasive leafminers, Liriomyza spp. comprise of morphologically cryptic species complexes that are known economic plant pests globally. They require rapid identification to design management strategies and prevent their further spread. Traditional identification of species based on morphology is difficult even for taxonomic specialists. Since DNA barcoding offers a powerful means for rapid identification, and it can be performed on any life stages, we therefore, applied this approach to invasive leafminers in the Philippines for reliable identification. The main objectives were to: (1) determine the identity of the pest species present, and (2) to evaluate the utility of DNA barcoding in an economically important agricultural pest group in which considerable complexity exists in mitochondrial structure.

We sequenced a 527-bp portion of mitochondrial cytochrome oxidase I (COI) from 258 leafminers collected from 26 plant host species (primarily crops), from a total of 10 provinces in two islands of the Philippines. Neighbor-joining and parsimony analysis were used to compare COI sequences from the Philippines to an extensive database of 307 COI sequences previously obtained from samples of the invasive leafminers Liriomyza huidobrensis (Blanchard), Liriomyza trifolii (Burgess), and Liriomyza sativae (Blanchard) from locations worldwide.

We conclude that although a DNA barcoding approach can provide rapid species identifications, in certain instances it is likely to either overestimate or underestimate the number of species present, depending on which sequence were used as reference sequences. In general, DNA barcoding holds tremendous promise as a means of generating large amounts of systematic data. But, DNA barcoding is most valuable when combined with taxonomic data from other sources, such as insect morphology or even behavior.

Subject: Insects

Second International Barcode of Life Conference 18-20 September 2007 107 Conference Abstracts Poster Abstracts

MOLECULAR IDENTIFICATION OF THE PERINEREIS SPECIES (NEREIDIDAE: POLYCHAETA) FROM INDO-WEST PACIFIC SHORES

Tae Seo Park1, SEUNG HYUN KANG2, Hyeyoung Koo3, and Won Kim2

1National Institute of Biological Resources, Environmental Research Complex, Incheon, 404-708, Korea 2School of Biological Sciences, Seoul National University, Seoul 151-747, Korea 3Department of Biological Science, Sangji University, Wonju 220-702, Korea

Perinereis polychaete worms are an important part of the invertebrate community of intertidal flats. Their feeding mode is known as surface deposit feeder, and the sediment reworking rate by their feeding activity maybe high, which is very important to the natural purification of the excess organic matter in the coastal ecosystem. Despite its important role in coastal ecosystem, some of Perinereis species were frequently mis-identified or mis-recorded. In this study, we evaluate sequence divergence in the mitochondrial cytochrome c oxidase I (COI) gene as a tool for resolving differences among Perinereis species from Indo-West Pacific shores. Sequence divergences within species were generally less than 1%, whereas divergences between species were greater than 8% in all cases. The present results suggest that DNA barcoding based on the COI gene sequences is a useful method for identifying species of the Perinereis polychaetes.

Subject: Invertebrates

DNA BARCODING ENDEMIC FRESHWATER FINFISH OF INDIA

SMITA KAPOOR, W. S. Lakra

National Bureau of Fish Genetic Resources, India

Freshwater finfish species richness and level of endemism were analyzed using published information and database. Variation at the mitochondrial cytochrome c oxidase subunit I gene was examined in 50 species including important carps, catfishes and murrels from India for the first time. Most species had unique sequences indicating that DNA barcoding is a powerful technology for species identification and biodiversity conservation.

Subject: Fish

Second International Barcode of Life Conference 18-20 September 2007 108 Conference Abstracts Poster Abstracts

EXPERIENCE OF PARTICIPATION IN BARCODING OF THE FAR EASTERN FISH SPECIES OF RUSSIA ON CYTOCHROME OXIDASE 1 GENE SEQUENCE DATA

YURY KARTAVTSEV1, S.N. Sharina1, A.Y. Chichvarkhin2, A.S. Sokolovsky1, A.A. Balanov1, K.A. Vinnikov3, V.N. Ivankov3

1A.V. Zhirmunsky Institute of Marine Biology of Far Eastern Branch of Russian Academy of Sciences, Vladivostok 690041, Russia 2Institute of Biology and Soil Science, of Far Eastern Branch of Russian Academy of Sciences, Vladivostok 690022, Russia 3Far Eastern State University, Vladivostok 690095, Russia

Sequence of nucleotides at cytochrome oxidase 1 (COI) gene was determined for 200 specimens in a frame of Fish-BOL in Russia. Flatfishes of the order Pleuronectiformes were in the focus; two genes, cytochrome b (Cyt-b) and COI were used. Based on GenBank and own data 67 and 21 sequences at Cyt-b and COI were analyzed. Nucleotide diversity measured in p-distances was estimated at four phylogenetic levels: (1) intraspecies, (2) intragenus, (3) intrafamily, and (4) intraorder. Scores of mean p-distances revealed the pattern of increased values, (1) 0.46±0.19%, (2) 11.74±2.26%, (3) 17.51±3.13%, and (4) 25.60±0.42% (mean ± SE) for Cyt-b and (1) 0.09±0.06%, (2) 10.97±1.87%, (3) 11.98±0.63%, and (4) 20.09±0.17% for COI, correspondingly. These data support the earlier findings and may show that geographic mode of speciation and later independent phyletic evolution prevailing in nature. Four tree building approaches, NJ, BA, ML, and MP give similar topologies of branches, proving the monophyly of 6 investigated flatfish families. Well genetically distinct are the majority of genera and species of flatfish. Main conclusion is made that the species barcoding on per-individual basis is efficient for both, COI and Cyt-b genes. Main problems of BOL in Russia are discussed.

Subject: Fish

Second International Barcode of Life Conference 18-20 September 2007 109 Conference Abstracts Poster Abstracts

TAXONOMY AND NATURAL HISTORY OF BEES OF SRI LANKA

W. A. I. P. KARUNARATNE and J. P. Edirisinghe

Department of Zoology, University of Peradeniya, Sri Lanka

Bees (Hymenoptera: Apoidea) are the most important pollinators of flowering plants resulting in fruit and seed production. Bees are unique among all other pollinators due to the presence of special pollen carrying hairs on their body. The very first record of taxonomic work on bees of Sri Lanka dates back to 1897 and was later followed by the Smithsonian Surveys in the late 1970’s. A recently conducted field study in 29 collection sites distributed among 13 districts of Sri Lanka led to the identification of 148 species of bees in 38 genera and 4 families.

These included 15 previously unrecorded species and 5 genera and a species new to Science. Based on the location of the 29 collection sites, the Low Country Dry Zone, middle peneplain, and the agricultural habitats harbored the highest number of species. Floral hosts consisted of 167 species in 115 genera and 44 families. Weeds were the most preferred hosts (129 spp.) followed by crops and trees. Flowers of 6 naturalized plant species attracted an unusual number (> 20 species) of bees. According to pollen relationships, 131 bee species are generalists and 7 species are pollen specialists. Twelve species of bees are buzz pollinators of crops whose pollen is concealed in anthers. Several stem nesting (16 spp.), ground nesting (13 spp.), and hive building (4 spp.) bees were recorded. Certain species were recorded only from their nests.

The lack of taxonomic revisions in Asia for majority of bee groups led certain groups to be identified only up to the generic level. Subfamily Nomiinae was studied in detail and was found that there are many cryptic species of Nomiinae and also Meliponini that need especial attention. DNA barcoding combined with morphological and natural history data of bees will provide the correct identity of this important group of pollinators in the country.

Subject: Insects

Second International Barcode of Life Conference 18-20 September 2007 110 Conference Abstracts Poster Abstracts

DNA BARCODE OF THE KOREAN MARINE INVERTEBRATES

CHOONG-GON KIM, Dageum Jeong, Go Eun Kim, Ki-Woong Nam, Heung-Sik Park, Se-Jin Pae, and Youn-Ho Lee

Korea Ocean Research and Development Institute, P.O. Box 29, Ansan 425-600, Korea

In the present study, we aimed the taxonomic revision of Korean marine invertebrates such as echinoderms, molluscs, annelida and sponges using the DNA barcoding system. We analyzed the invertebrates DNA barcode using the mitochondrial DNA sequences of cytochorome oxidase subunit I (COI) gene. After extraction of genomic DNA from the samples using QIAGEN kit, COI sequences were amplified by COI universal primer: COI universal primers (F: 5'- ggt caa caa atc ata aag ata ttg g - 3', R: 5'- taa act tca ggg tga cca aaa aat ca -3') recommended by Barcode of Life. The amplified DNA fragments are sequenced and analyzed by molecular phylogenetic method.

We analyzed 36 species of the echinoderms, 30 of annelida, 10 of sponges and 9 of mollusca species. The Echinoderms including sea urchin, sea stars and sea cucumbers are a major group of marine invertebrates that can be found in almost every marine habitat. The Annelida (Polychaetas) are extremely variable in both morphology and lifestyle. Sponges constitute a group of marine invertebrates whose its morphological classification is very difficult because of a few morphological characters and a high degree of similarity. Some mollusca (mussel) species are has been controversy the identity of introduced species caused by uncertainty of morphological distinction. These marine invertebrates are all of the dominating benthic groups in terms of biomass and species diversity, yet studies on molecular biodiversity and genetic structure are still scarce.

The COI barcode results showed a significant differentiation between each species using this study except the sponge species. Mitochondrial DNA COI barcoding result shows homogeneity across sponge species with scant variability and no geographical resolution. We also report the phylogenetic relationships of the invertebrates used in this study.

Subject: Marine

Second International Barcode of Life Conference 18-20 September 2007 111 Conference Abstracts Poster Abstracts

DEVELOPMENT OF PLANT DNA BARCODING MARKERS FROM THE VARIABLE NONCODING REGIONS OF CHLOROPLAST GENOME

Hae-Lim Lee, Dong-Keun Yi, Jeong-Seong Kim, and KI-JOONG KIM

School of Life Sciences and Biotechnology, Korea University, Seoul 136-701, Korea

Complete chloroplast DNA sequences were generated from seven species of Panax, one species of Aralia and a species of Eleuthrococcus (Araliaceae). We reconstruct the species phylogeny using both coding and noncoding regions of chloroplast DNA. Based on the comparative sequence analyses of fully sequenced chloroplast genomes, we selected the seven spacer regions including rps15/ycf1, ndhG/ndhI, trnH/psbA, psbK/pabI, atpF/atpH, petA/psbJ and atpH/atpI spacers as the promising barcoding markers for flowering plants. The regions show the high sequence divergence, the relatively low length variation, and no spacer interruption. Among them, ndhG/ndhI, psbK/pabI, atpF/atpH, and atpH/atpI regions are transcribed as a single cistron. The spacer regions, rps15/ycf1, ndhG/ndhI, are located on the SSC and other five regions are located on LSC. We also apply the DNA barcoding markers to 10 other genera of land plants including Syringa(Oleaceae), Taraxacum(Asteraceae), Quercus(Fagaceae), Carex(Cyperaceae), Pinus(Pinaceae), Dryopteris (ferns), etc. Furthermore, we extended the evaluation of the markers in terms of the universality, variability, and alignment to the 100 selected genera from flowering plants, gymnosperms, fern and their allies, and mosses. Two noncoding markers (atpF/atpH and psbK/psbI) were identified as extremely reliable DNA barcoding markers for a whole plant kingdom. Therefore, we proposed the three marker barcoding system (a coding region of matK and two noncoding region of atpF/atpH and psbK/psbI) for plant kingdom. The proposed three maker barcodes can be identified species from a notorious apomictic Taraxacum complex. The three barcode system showed the identification power among all tested multi-species from various groups of plant genera. We believe that the currently available bioinformatics algorithms do not discourage the usages of length variable regions for the DNA barcode markers.

Subject: Plants

Second International Barcode of Life Conference 18-20 September 2007 112 Conference Abstracts Poster Abstracts

DNA BARCODES AND SIBLING SPECIES COMPLEXES OF ANOPHELINE MOSQUITOES IN INDIA (DIPTERA: CULICIDAE)

N. PRADEEP KUMAR, A.R. Rajavel and P. Jambulingam

Vector Control Research Centre (Indian Council of Medical Research), Indira Nagar, Puducherry –605006, India

DNA Barcodes based on mitochondrial cytochrome c oxidase-1 gene sequences has emerged as an important tool for identification of species among diverse group of animals ranging from Class Pisces to Class Insecta. Also its utility for identification of cryptic species has been validated among Lepidopterans and Dipterans. Mosquitoes belonging to the Genus Anopheles (Family: Culicidae) are notorious owing to their role as vectors of malaria in different regions of the Globe. This genus includes many sibling species complexes. Among 56 species of Anopheline mosquitoes reported in India at least 6 species are reported to be sibling species complexes. We have initiated studies on the taxonomy of these complexes based on DNA Bar code approach and the initial results are presented here.

Anopheles culicifacies complex is the most important vector of malaria in India. This complex constitutes 5 sibling species based on chromosomal polymorphisms. However, PCR methodologies based on ribosomal DNA (rDNA) polymorphism could identify only two species among this complex. We constructed DNA Barcodes for 6 specimens of this species complex, identified by PCR methodology to be species A and B. DNA Bar code approach also identified them, as belonging to two species, the genetic distances (Kimura 2 parameter) between these species being 5.21%. They grouped into distinct clades in phylogenetic tree constructed for Anopheles mosquitoes. An. fluviatilis complex is another important vector species of malaria in India. It constitutes 3 sibling species. PCR methodologies had been reported to identify these 3 sibling species complexes based on rDNA polymorphism. 20 individuals of these mosquitoes identified to be either belonging to Species S or T was subjected to DNA Barcode taxonomical approach. The analysis indicated the mean genetic distances of these two sibling species members to be only 0.08%, and these two sibling species did not separate into two distinct clades in the phylogenetic tree constructed, denoting them to be not two distinct species, by DNA barcode approach.

An. minimus complex, another important malaria vector comprise of 3 sibling species; however only species A have been reported from India. We identified 10 specimens of this species to be species A and compared it with the lone COI sequence available in the GenBank for species C in the complex. The DNA Barcode approach could distinguish these sibling species members of the complex, the genetic distance between the members being 2.94%. A few studies reported that An. minimus C is synonymous with An. fluviatilis S based on the sequences of D3 region of rDNA. However, analysis of DNA Barcodes with the available COI sequence of An. minimus C denoted them to be two distinct species. The genetic distance estimated between these two species was estimated to be 4.75%.

An. subpictus is another species of Genus Anopheles in which sibling species complexes have been reported. We analyzed 13 specimens of this species collected from different regions of the country identified morphologically to be species A and B. DNA Barcode approach could distinguish the 2 sibling species, their genetic distances being 12.45%. Further studies are ongoing on other sibling species complexes and members of Anophelines.

Subject: Insects

Second International Barcode of Life Conference 18-20 September 2007 113 Conference Abstracts Poster Abstracts

THE UNION OF BARCODING AND TAXONOMY: A CASE STUDY IN THE COMMERCIALLY VALUABLE BLUE SWIMMING CRAB, PORTUNUS PELAGICUS SPECIES COMPLEX

JOELLE C.Y. LAI1, Peter F. Davie2, and Peter K.L. Ng1

1National University of Singapore 2Queensland Museum, Bisbane, Australia

Whilst DNA barcoding has been shown to uncover hidden pockets of biodiversity in different animal groups, species discovery via barcoding should nevertheless be carried out in tandem with careful morphological analysis, knowledge of biogeographic information and judicious checking of historical literature to maximise its full value to the scientific community as well as in practical situations. In our case, analyses of COI sequences and morphology of the marine swimming crab Portunus pelagicus showed that it is in fact a complex of at least four species. However, while unique Molecular Operational Taxonomic Units (MOTUs) may be morphologically indistinguishable, we also have what would be considered as a single MOTU comprise of two species with pronounced morphological differences between them. Using a complement of various methods, we have clarified the systematics of this commercially important group of species group. In the process, we have maintained nomenclatural stability as far as possible yet assign pre-existing names to each rediscovered species.

Subject: Marine

MOLECULAR PHYLOGENY SUGGESTS POLYPHYLY OF BOTH CYCLOPHORINAE AND AYCAEINAE (MOLLUSCA: CYCLOPHORIDAE)

YEN-CHEN LEE1,2, Wen-Long Wu2, Kuang-Yang Lue1

1Department of life science, National Taiwan Normal University 2Research Center for Biodiversity, Academia Sinica

Cyclophoridae is the dominant group of operculated territorial snail in East Asia. The group consist 4 subfamilies and about 300 species currently arranged in 34 genera, occupied varies habitats, and with great morphological diversity. The molecular phylogenetic relationship of the group was never discussed before. In order to uncover the relationship between Cyclophorinae and Aycaeinae, we used 27 species of 8 genera of Cyclophorinae and Aycaeinae. Then use maximum parsimony, maximum likelihood and neighbor joining analyses on two genes dataset comprising part of COI and 16S rRNA separately. The phylogenetic relationship based on mtDNA sequences suggests that neither the subfamily Cyclophorinae, nor Aycaeinae are monophyletic. In so-called Aycaeinae, both COI and 16S rRNA indicate Chamalycaeus is more basic group than Dioryx. Dioryx is more closely related to Cyclophorinae genera Pterocyclos and Cyclotus in 16S rRNA phylogenetic relationship.

Subject: Invertebrates

Second International Barcode of Life Conference 18-20 September 2007 114 Conference Abstracts Poster Abstracts

BARCODES OF MYCORRHIZAL FUNGI IN SUBTROPICAL FOREST ECOSYSTEMS: APPLICATIONS IN SPECIES IDENTIFICATION AND BIODIVERSITY ESTIMATION.

YU LIANG, Qiong Ding, Ying Chen, Kequan Pei, Keping Ma

Key Laboratory of Vegetation and Environmental Change, Institute of Botany, the Chinese Academy of Sciences

Mycorrhiza is a mutualistic symbiosis between plant and fungus localized in a root or root-like structure in which energy moves primarily from plant to fungus and inorganic resources move from fungus to plant. There are over 6,000 mycorrhizal fungal species in the world and species diversity of mycorrhizal fungi is high in subtropical forests of China. Comparing with animals and plants, morphological identification of mycorrhizal fungal species is quite difficult due to the following reasons: (1) the up-ground parts of mycorrhizal fungi, i.e. the sporocarps or fruitbodies, are short- lived (usually rot in two days after emergence) and seasonal and yearly variances in sporocarp compositions make it difficult to collect the fruitbody needed; (2) identification is infeasible based on morphological information of underground structures of mycorrhizal fungi; (3) because many mycorrhizal species do not form sporocarps or their sporocarps cannot be easily found, the species compositions of up ground and under ground mycorrhizal communites are usually not matched.

In an on-going project, we try to use DNA barcodes, cytochrome oxidase I (COI) gene and the nuclear ribosomal internal transcribed spacer (ITS) regions, in species identification of mycorrhizal fungi and species diversity estimation of mycorrhizal communities. DNA has been extract from over 2,000 mycorrhizal samples, i.e. infected roots and sporocarps, and over 150 ITS sequences have been obtained. Sequences of mycorrhizal fungi are compared and we are trying to find species-specific oligos to measure the species composition and diversity in the ecosystem using a microcoding strategy. We hope this project be helpful in species identification of mycorrhizal fungi and diversity evaluation of mycorrhizal communities in natural ecosystems.

Subject: Fungi

Second International Barcode of Life Conference 18-20 September 2007 115 Conference Abstracts Poster Abstracts

EVOLUTIONARY MITOGENOMICS OF THE HEXACORALLIAN ANTHOZOANS (CNIDARIA; ANTHOZOA): GENE ORDER, INTRON EVOLUTION, AND PHYLOGENY

MEIFANG LIN, Chienhsun Chen,Yao-Yang Chuang, Yu-Tseng Wu, Chaolun Allen Chen

Research Center for Biodiversity, Academia Sinica

Colonial scleractinians are the main reef-building organisms of the planet, and sustain amongst the most species rich marine ecosystems. Among the 1,314 currently know valid scleractinian species, 656 are zooxanthellate and 669 are azooxanthellate. If shallow-water corals are fairly well studied, the less accessible, deep-water fauna is still poorly know and new azooxanthellate general are still being described. However, in 2006, Medina et al indicated that the scleractinian skeleton was likely lost in the ancestry of Corallimorpharia and supported the “naked corals” hypothesis. But the poorly know azooxanthellate Scleractinia still need more research. Evolutionary genomics of mitochondrial genomes studies the gene arrangement with phylogenetic inferences of complete length of protein-coding genes or nucleotide sequences. In this study, the complete mitochondrial genomes of a fungiacyathid, two caryophylliids and, one oculinid will be characterized. The results will be utilized to compare those anthozoans mt genomes available in the GenBank, and gene arrangement, group I intron evolution, and phylogenetic implications will be analyses to elucidate the evolution of scleractinian.

Subject: Marine

Second International Barcode of Life Conference 18-20 September 2007 116 Conference Abstracts Poster Abstracts

EVALUATION OF SPECIES DIVERSITY OF THE SOUTH AMERICAN SUCKERMOUTH ARMORED CATFISHES (SILURIFORMES: LORICARIIDAE) IN THE AQUARIUM MARKETS USING DNA BARCODE TECHNIQUES

SI-MIN LIN1, Wei-Ling Kao1, Chun-Yi Ku2, and Shen-Horn Yen3

1Department of Life Science, Chinese Culture University 2Wiser Publishing Company 3Department of Biological Sciences, National Sun Yat-Sen University

The Loricariidae (the South American suckermouth armored catfishes) have more than 600 species in over 40 genera in the Neotropical region and harbour the highest specific diversity among all the catfish families. These catfishes are well-known for their popularity in tropical fish trade, and thus some species of the subfamily Hypostominae have been incidentally invaded into various areas of Asia through impropriate fish release and caused serious ecological problems for freshwater ecosystem. In contrast with the rapid Introduction of these catfishes to tropical fish trade and the significant increase of new “fishes” recognized by fish traders, only very few of the loricariid fishes circulated in the fish trade can taxonomically be justified due to the historical difficulties in confirming type locality of each “species” as well as the highly variable colour pattern of “a species” recognized by traders.

In the present study, we use mitochondrial DNA sequence data to evaluate the validity of the “L- numbers”, a commercial naming system developed for coding the loricariid fishes that have not been scientifically described based on external morphology, and to test the species membership of the species which are already recognized based on morphological characters. We selected two genera, Baryancistrus and Panaque, both common in tropical fish trade and with complex colour pattern variations among species, as our target taxa. The result shows that the barcode data and morphological information match well, and thus each of the L-numbers involving these two genera can be recognized as good species. This indicates that the diversity of loricariids may have been under-estimated, and more future studies should be carried out to evaluate the taxonomic identities of the species with ecological impacts and evolutionary significances.

Subject: Fish

Second International Barcode of Life Conference 18-20 September 2007 117 Conference Abstracts Poster Abstracts

IDENTIFICATION OF MULTIPLE MARKERS FOR PHYLOGENETIC AND DNA BARCODING STUDIES FOR MEDICINAL PLANTS

Loucia Chan, Stanley Chi Chung Chik, Allan SY Lau, and CHARLIE CHANG LIU.

L6-27, Molecular Chinese Medicine Laboratory, LKS Faculty of Medicine, 21 Sassoon Road, The University of Hong Kong, Hong Kong SAR, P.R.China

There are an estimated 12000 medicinal plants used in the traditional Chinese medicine system and they are of tremendous economic and medical significance in China and other countries. Our long term goal is to compile multiple markers to build a phylogenetic foundation and to generate a barcode system for species discovery and identification of these medicinal plants. In collaboration with the Institute of Medicinal Plant Development in P.R.China, we collected a sample set containing ~360 medicinal plants described in Chinese Pharmacopeia. Four marker genes, ITS, 18S, rbcL, atpB have been chosen. First, the plant DNA was extracted. Second, degenerate primers were used to amplify sequences of the marker regions. And last, the amplified fragments were subjected to DNA sequencing. In our preliminary study, we have generated ~220 phylogenetic marker/barcode sequences from 52 samples. Thirty-one of them have sequences generated for all four markers. The DNA sequences of the four markers were aligned using Clustal W software and subjected to parsimony analysis individually or in concatenation using PAUP software. Our analysis has demonstrated the possibility of using short DNA sequences as phylogenetic markers and DNA barcodes, and the advantage of using multiple markers over the use of single marker.

Subject: Plants

FINGERPRINT: VISUAL DEPICTION OF VARIATION IN MULTIPLE SEQUENCE ALIGNMENTS

MELANIE LOU and G.B. Golding

McMaster University, Hamilton, Ontario, Canada

We have created a web server, Fingerprint, which provides a simple, intuitive web interface to programs that provide a visual representation of genetic variation in BARCODE sequences. It can represent identity, variability, charge, hydrophobicity, solvent accessibility, and structure along with other visualizations based on composition, heterogeneity, dN/dS, and nucleotide diversity.

For illustrative purposes, Fingerprint was applied to 9195 lepidopteran sequences. We found that lepidopteran families were very similar in residue composition. However, the heterozygostity, variability and nucleotide diversity fingerprints revealed distinct patterns of variation between families. Within each family, the locations of the sites exhibiting diversity were similar; however, the extent of diversity differed between corresponding sites.

In summary, Fingerprint is effective for identifying sequence variation and for preparing high resolution, intuitive graphics for presentation. The identification of variation permits one to see the distribution of a particular feature and to make inferences about the structure, function, and evolution of the sequences in question.

Fingerprint is available at http://evol.mcmaster.ca/fingerprint/.

Subject: Methods

Second International Barcode of Life Conference 18-20 September 2007 118 Conference Abstracts Poster Abstracts

DEVELOPMENT OF A BIOCHIP FOR RAPID DISCRIMINATION OF THE SPECIES OF FOUR TEPHRITID FRUIT FLIES (DIPTERA: TEPHRITIDAE)

Chen, Y.-H. W.-B. Yeh, M.-M. Yang, M.-H. Kuo and K.-H. LU

Department of Entomology, National Chung Hsing University, Taichung, Taiwan

Traditionally, morphological characters are regularly adopted to discriminate the species of insect pests that are intercepted in quarantine inspection. However, it is not easy to identify the species from larvae or even eggs of most tephritid fruit flies, which often occur in imported or exported fruits, based only on their morphology. In order to overcome this difficulty, an attempt to develop a biochip for species identification was carried out in this study. The cytochrome oxidase I (COI) gene of four tephritid fruit flies, including Bactrocera dorsalis, Ceratitis capitata, Anastrepha suspensa and Rhagoletis pomonella, were selected for this work. We first obtained the COI fragments from the genomic DNA of each species with PCR amplification using a pair of universal primers. After DNA sequencing and alignment, several potential species-specific oligonucleotide probes (size between 20~30 mers) were designed and synthesized according to the most variable regions of each species. After a series of screening and tests, a biochip prototype was successfully developed; and this biochip possesses the criteria of high sensitivity and species specification for rapid discriminating at least these four species of tephritid fruit flies.

Subject: Insects

INFORMATION VISUALIZATION AND FUSION FOR BARCODES OF LIFE IN ENVIRONMENT AND SOCIETY

Paul Yu Chen, Tai-Been Chen, Chun-Jui Chen, Guan-Cheng Li, HENRY HORNG-SHING LU

Institute of Statistics, National Chiao Tung University

Visualizing information is a key step to fuse information from different sources related to the Barcode of Life. In order to provide a user friendly interface for such purpose, we have developed a web application allowing information visualization and fusion at http://140.113.114.100/BOL/main.htm. For example, the unfortunate mislabeling of toxic puffer fish as monkfish (http://www.ens-newswire.com/ens/may2007/2007-05-28-09.asp) could be detected and avoided by the use of Barcode of Life and related information. In cases like this the information visualization can integrate the information of Barcode of Life and biologic features with news in the environment and society. This example is demonstrated in our web pages through searching results from mitochondrial sequences, taxonomy trees, images and related information from Consortium for the Barcode of Life (http://barcoding.si.edu/), FishBase (http://www.fishbase.org/), NCBI (http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi), FISH-BOL (http://www.fishbol.org/) and related web pages as well as subsequent databases.

Users will be able to operate the interface with ease allowing them to search for news articles or web resources, identify specimens, and even link to some of the most useful and valuable sources related to the BOL. As the progress of Barcode of Life advances, the information visualization and fusion system can be incorporated to make use of the majority of information available to serve all concerned users in international communities for both individual and professional needs.

Subject: Methods

Second International Barcode of Life Conference 18-20 September 2007 119 Conference Abstracts Poster Abstracts

ORDER OUT OF CHAOS: DNA BARCODING IN THE LAURACEAE

SANTIAGO MADRIÑAN

Laboratorio de Botanica y Sistematica, Universidad de los Andes, Bogota, Colombia

The Lauraceae are a major component of tropical floras. Worldwide they comprise 52 genera and ca. 2,750 species. Lauraceae are notorious among botanists who consider identification of genera and species extremely difficult. One striking characteristic of Lauraceae is that species almost identical in leaf characters can differ profoundly in floral or fruiting morphology, both within and between genera. Another serious problem in identifying Lauraceae is that vegetative characters often vary considerably within species, populations, and even individuals. Different collections of the same species have been described in different genera. Recent studies of molecular variation within Lauraceae coupled with the efforts in establishing a DNA barcode for land plants may prove a solution to what Alwyn Gentry once referred to as the taxonomic chaos that prevails in the Lauraceae. As part of a multi institutional project geared towards establishing a DNA barcode for land plants, We sequenced tentative barcode chloroplast DNA regions (matK, rpoB, rpoC1 and trnH- psbA) for selected Lauraceae specimens. Ease of sequence alignment and nucleotide variation were scored for each region. Identification matches using BLAST with available GenBank sequences were tried for each specimen with high success rate.

Subject: Plants

Second International Barcode of Life Conference 18-20 September 2007 120 Conference Abstracts Poster Abstracts

DNA BARCODING IN FORENSIC ENTOMOLOGY

TADEUSZ MALEWSKI, Agnieszka Draber-Monko, Wieslaw Bogdanowicz

Museum and Institute of Zoology, Polish Academy of Sciences, Warsaw, Poland

A carrion-fly maggot is the most common type of insect evidence collected during investigation of dead bodies. Determining the species composition of such sample is an important first step in a forensic entomological analysis. Although species-diagnostic anatomical characters have been reported for the eggs and early larvae of some taxa, an anatomical approach does not provide 100% identification of immature stages of many species. Mitochondrial DNA can be used in species identification of all life stages of a carrion fly. Insects of the greatest value to forensic entomology are blowflies (family Calliphoridae) - they are usually the first insects to colonize a body after death, often within hours. Out of 66 species of blowflies occurring in Poland, twelve of them are of forensic importance. The majority of these species have already been sequenced, and this information is available for use by police and other investigative agencies. Based on results we obtained it will be possible to design the sequence-specific probes. Moreover, beyond the applications to forensic entomology, an understanding of the COI sequences of these flies may be also treated as a valuable addition to the DNA barcoding initiative.

Subject: Insects

DNA BARCODING REVEALS SUBSTANTIAL CRYPTIC DIVERSITY AMONG MACROALGAE IN CANADA'S THREE OCEANS

DAN MCDEVIT and Gary W. Saunders.

Centre for Environmental & Molecular Algal Research, Biology, University of New Brunswick, NB,Canada.

This presentation will highlight successes to date in our DNA barcoding effort to generate a complete floristic account of the marine macroalgae in Canada. With ca. 2300 barcodes completed at the time of writing this abstract, we have uncovered ca. 70 overlooked (cryptic) species or records in the Canadian flora including six unique to the Churchill region in the low Arctic. In addition to altering radically our perspectives on the actual numbers of species in Canadian waters, we are uncovering a bewildering array of tales that are: altering our perspectives on algal distribution at both ecological and geographical scales; challenging widely accepted morphological species concepts; rewriting phylogeographical hypotheses; resolving the dynamic nature of speciation and various hybrid populations; and uncovering key examples of niche exclusion with a corresponding restriction in phenotypic plasticity. We will present an overview of some of the highlights of our work with the aim of sharing the enthusiasm that has gripped our lab in what is truly the golden age of molecular-assisted alpha taxonomy.

Subject: Protists

Second International Barcode of Life Conference 18-20 September 2007 121 Conference Abstracts Poster Abstracts

DELINEATION OF SPECIES BOUNDARIES IN PAVONA SPP.

K. RUBY MOOTHIEN PILLAY1, Takashi Asahida2, C. Allen Chen3, Hiroaki Terashima2, Keiichi Nomura4, Hitoshi Ida2

1Mauritius Oceanography Institute, Mauritius 2School of Fisheries Sciences, Kitasato University, Japan 3Research Centre for Biodiversity, Academia Sinica, Taiwan 4Kushimoto Marine Park Centre, Kushimoto, Japan

Pavona cactus and P. decussata are sympatric on the reef flats of Mauritius and are usually intricately associated. Using the nuclear ribosomal DNA internal transcribed spacers (rDNA ITS-1 and ITS-2) and the mitochondrial cytochrome b (mtDNA Cyt.b) gene, the species boundaries between both species were examined. The phylogenetic analyses of ITS sequence types separated out P. cactus and P. decussata into two distinct clades. Moreover, the significant difference detected by the Nested Clade Analyses indicated that both P. cactus and P. decussata in Mauritius constitute statistically distinguishable lineages. A set of ribosomal ITS DNA sequences of both species from the high latitude reef of Kushimoto (Japan) further confirmed these species to be distinct evolutionary entities. The complementary use of the Cyt.b, characterized by a very slow rate of evolution in corals, contributed largely in determining the relationship between these closely related taxa. The conservative change at the amino acid level confirmed P. cactus and P. decussata to be distinct species. Additional Cyt.b sequence data from a third species, P. danai, similarly showed a conservative change at the amino acid level from P. cactus and P. decussata. The genetic markers used in this study proved useful for species delineation.

Subject: Marine

WOOD DECAY FUNGAL DIVERSITY IN BANGLADESH

M.A.U. MRIDHA1; S. Dewan2; M.S.R. Rahman2, and M.K. Bhuiyan2

1Department of Botany 2Institute of Forestry and Environmental Sciences, University of Chittagong, Chittagong-4331, Bangladesh.

The wood decay fungi (macro fungi) have a great importance in the viewpoint of economic and ecosystem management. Samples of wood decay fungi were collected from different selected locations of Comilla, Chittagong and Chittagong Hill Tracts. Forty four different types of macro fungi belonging to 16 genera were encountered from Bangladesh during the present study. They were identified by following the standard keys and confirmation was made from the Field Museum, Chicago University, USA. The identified genera were Amauroderma, Daedalea, Fomitopsis, Fomes, , Favolus Ganoderma, Hexagonia , Inonotus, Irpex, Lenzites, Phellinus, Polyporus, Pycnoporus,,Pleurotus, Trametes, Schizophyllum,. The diversity of macrofungi is host independent Strong host specificity was not found among identified fungi growing on different host tree species. The implications of diversity of fungi in Bangladesh are discussed.

Subject: Fungi

Second International Barcode of Life Conference 18-20 September 2007 122 Conference Abstracts Poster Abstracts

CONSTRUCTION OF DNA BARCODE OF PTERIDOPHYTES BY USING rbcL GENE AND ITS FLANKING REGION

MOSTAFA K. PASHA, and M. I. H. Reza

Department of Botany, University of Chittagong, Chittagong-4331, Bangladesh.

Chloroplast synthesizes RUBISCO, has two subunits, LSU and SSU. The LSU is translated from the rbcL gene located in cpDNA. The exact position and the nucleotide sequence are detected in many plants. The gene has no intron, about 1425-1431 bases coding region, as revealed from the amino acid sequences and also from DNA sequence analysis. The rbcL gene can be separated intact when EcoRI and BamHI are used. The fragment contains the gene with 3' and 5' flanking regions having about 1700bp. This fragment when treated further with EcoRI and BglII a small fragment yielded 472bp length of the gene. This size fragment is suitable for both cloning and DNA sequencing. The nucleotide sequences have found high conserved regions and some silent mutated regions; yielded up to 16% variations which are sufficient to establish species identity and to construct DNA Barcode for every species to be identified. With this view in mind a project proposal is proposed to construct DNA Barcode of the Pteridophytes of Bangladesh (we have about 250 species identified and the herbarium specimens are deposited in the Chittagong University Herbarium located at Botany Department). This project will be extended for flowering plants of Bangladesh in future.

Subject: Plants

EXPLORING CORAL REEF BIODIVERSITY USING DNA BARCODING

LAETITIA PLAISANCE, Nancy Knowlton

Center for Marine Biodiversity and Conservation (CMBC), Scripps Institution of Oceanography, University of California San Diego

Coral reefs are the most diverse marine ecosystems on the planet and among the most threatened. Unfortunately we lack the taxonomic expertise and the time needed to characterize this diversity, as well as the extent of biodiversity loss related to human impacts, using traditional methods. However, the revolution in molecular genetics dramatically changes the potential for reef scientists to make progress in this area.

In this study, we investigated the diversity of crustaceans inhabiting coral reefs using DNA barcoding. In order to achieve a semi quantitative sampling, crustaceans were collected from similar sized dead heads of Pocillopora. Sampling was performed in Moorea, French Polynesia (8 heads) and in four localities of the Line Islands across a gradient of human disturbances: Kingman Reef (4 heads); Palmyra Atoll (3 heads); Fanning Island (2 heads); Christmas Island (5 heads); all heads were from 10m depth. Crustaceans were removed from the heads (517 specimens) and a DNA sample was collected in order to sequence partial cytochrome oxidase subunit I (403 sequences). For this very restricted habitat, that is a dead head of Pocillopora at a single depth, we estimated that the species richness for crustaceans could be as high as 150 in the Line Islands and 77 in Moorea.

Subject: Marine

Second International Barcode of Life Conference 18-20 September 2007 123 Conference Abstracts Poster Abstracts

MOLECULAR ANALYSIS OF THE GENUS PHYSALIA (CNIDARIA: SIPHONOPHORA) IN NEW ZEALAND

DAVID R. PONTIN and R.H. Cruickshank

Lincoln University Lincoln, Canterbury, New Zealand

Physalia physalis (Portuguese Man O War) and P. utriculus (bluebottle) have both been described as present in the Pacific Ocean, however neither has been confirmed in New Zealand. The species are morphologically similar, with the main features used to differentiate the species being the size of the specimen and number of main tentacles, which creates the possibility of misidentification. Moreover, the ability to identify cnidarians through cytochrome oxidase (COI) is uncertain as Hebert et al. (2003) suggested that COI evolves much slower in Cnidaria than other phyla. Twenty six specimens were collected from five locations from around New Zealand and sequenced for both COI and the internal transcribed spacers (ITS) of the nuclear ribosomal gene cluster. Further COI sequences were sourced from Hawaii (6 specimens) and for P. physalis. Four distinct clades were identified with 7.7%-12.7% divergence for COI and 1.8% divergence for ITS1 (New Zealand clades only). Based on these results it would appear that New Zealand has two distinct species of Physalia, neither of which are P. physalis. Furthermore, COI is an appropriate gene for species identification of Physalia and may be applicable to other genera.

Subject: Marine

DNA BARCODES AND MOLECULAR ALPHA-TAXONOMY OF A POORLY EXPLOREDBUT HYPERDIVERSE GROUP OF MARINE GASTROPODS : THE TURRINAE (GASTROPODA).

NICOLAS PUILLANDRE, Sarah Samadi, Marie-Catherine Boisselier, Philippe Bouchet.

“Systématique, Adaptation et Evolution”, UMR 7138 CNRS-IRD-UMPC-ENS-MNHN, Muséum National d'Histoire Naturelle (USM 603), Département Systématique et Evolution, Case postale 26, 57 Rue Cuvier, F-75231 PARIS CEDEX 05 France.

Conoidea, which includes subfamily Turrinae, is, a hyperdiverse group of marine gastropods, with about 10000 living species defined mostly by shell characters. The great and sometimes continuous variability of these morphological characters renders the delimitation of species complicated. Molecular taxonomy has received increasing attention (Vogler 2006) to better define species boundaries in poorly known groups, where traditionnal characters are difficult to assess. Recent cruises in West-Pacific permitted to sample more than 500 specimens of Turrinae. Species were not a priori defined in order to accurately estimate intra and interspecific variabilities. Sequencing was performed for two genes : the COI mitochondrial gene used for the barcode project and a portion of the 28S nuclear gene. Species boundaries were defined using phylogenetic reconstructions associated to different methods method, as for example the one described in Pons et al. 2006. Several tens of clusters were thus delimited, mostly including several specimens. Geographic and morphologic informations were then used to discuss alpha-taxonomy of the group. If some clusters were easily recognisable on the basis of morphological characters, some included specimens with apparently identical shells, while several shell morphologies were observed in others. Indeed, molecular taxonomy appears to be a fast and accurate way to define clusters of individuals within Turrinae.

Subject: Marine

Second International Barcode of Life Conference 18-20 September 2007 124 Conference Abstracts Poster Abstracts

THE MOOREA BIOCODE PROJECT: ENABLING A MODEL ECOSYSTEM APPROACH TO CONSERVATION SCIENCE

GEORGE RODERICK1, Neil Davies2, John Deck1, Matteo Garbelotto1, Rosemary Gillespie1, Joyce Gross1, Christopher Meyer1, Jean-Yves Meyer3, Brent Mishler1, Craig Moritz1, Serge Planes4, Gustav Paulay5, Claude Payri6

1University of California, Berkeley, USA 2UC Berkeley Gump South Pacific Research Station, French Polynesia 3Délégation à la Recherche, French Polynesia 4CRIOBE (UMS CNRS EPHE), Moorea, French Polynesia 5University of Florida, Gainesville, USA 6IRD, Noumea, New Caledonia

The Moorea Biocode Project (MBP) is an integrated approach to biodiversity science that bridges the gap between ecology, genomics, and systematics. Strongly focused on field research, the MBP promotes biocomplexity science based on model ecosystems. The goal is to put in place the conditions necessary to achieve radical advances in the way we monitor and manage ecosystems and conserve biodiversity. One objective of the MBP is to generate a vouchered, All Taxon Barcode Inventory (ATBI) for the tropical island of Moorea, French Polynesia. A complementary objective is to build the identification tools needed to access this inventory. A fundamental principle of the MBP is to ground the development of informatics in real-world experience. Thus, computer scientists and informatics experts are working directly in conjunction with field, laboratory, and museum researchers, to ensure that solutions are user-driven and practical. In a pilot study (Dec 2005-June 2007) we catalogued approximately 30% of the known diversity for the Moorea ecosystem, including both marine and terrestrial species. Particular targets for the pilot study included fish, marine invertebrates, terrestrial arthropods, terrestrial herps, and ferns. Here, we describe the details of the project, collaborations, successes, pitfalls, and prospects for the future.

Subject: Methods

Second International Barcode of Life Conference 18-20 September 2007 125 Conference Abstracts Poster Abstracts

THE BARCODING OF ANTARCTIC FISHES OF TERRE ADÉLIE

Olivier Rey1, Céline Bonillo2, Agnès Dettaï1, Catherine Ozouf-Costaz1, SARAH SAMADI1, and Guillaume Lecointre1

1 “Systématique, Adaptation et Evolution”, UMR 7138 CNRS-IRD-UMPC-ENS-MNHN, Muséum National d'Histoire Naturelle (USM 603), Département Systématique et Evolution, Case postale 26, 57 Rue Cuvier, F-75231 PARIS CEDEX 05 France. 2Service de systématique moléculaire (CNRS, IFR101), Muséum National d'Histoire Naturelle, Département Systématique et Evolution, CP 26, 57 Rue Cuvier, F-75231 Paris Cedex 05, France.

Forty years of fish sampling in the Antarctic sector Terre Adélie and other areas of the Southern Ocean have provided an important collection of Antarctic marine organisms stored in the Muséum National d’Histoire Naturelle in Paris, France. This collection was gathered thanks to the ICOTA program (1996-2006) directed by Philippe Koubbi and Catherine Ozouf-Costaz and recurrent sampling efforts in this area and other sectors make the fish barcoding initiative relevant for creating a reference collection of specimens and sequences. In Terre Adélie several interesting issues concern species delineation among coastal Antarctic fishes. Sequences from several markers including COI, cytochrome b and the mitochondrial Control Region, to which we have added nuclear markers like the Rhodopsin gene will be coupled with voucher specimens stored in formalin, pictures and chromosome structures of the same individuals. Among results, the sequences used for barcoding clearly separate Trematomus hansoni from Terre Adélie from those of the same species in South Georgia. Gymnodraco acuticeps from Terre Adélie is not distinguisable from Gymnodraco victori neither from the sequenced genes nor from the cytogenetic analysis. Moreover specimens with intermediate morphological features have been recently recorded (and barcoded).

Subject: Fish

Second International Barcode of Life Conference 18-20 September 2007 126 Conference Abstracts Poster Abstracts

A BARCODE INVENTORY OF DEEP-SEA FISHES FROM THE SANTO 2006 EXPEDITION.

SARAH SAMADI1, Julien Brisset1, Damien Hisinger1, Nicolas Puillandre1, Céline Bonillo§, Agnés Dettaï1,Guillaume Lecointre1

1 “Systématique, Adaptation et Evolution”, UMR 7138 CNRS-IRD-UMPC-ENS-MNHN, Muséum National d'Histoire Naturelle (USM 603), Département Systématique et Evolution, Case postale 26, 57 Rue Cuvier, F-75231 PARIS CEDEX 05 France. ([email protected]) 2Service de systématique moléculaire (CNRS, IFR101), Muséum National d'Histoire Naturelle, Département Systématique et Evolution, CP 26, 57 Rue Cuvier, F-75231 Paris Cedex 05, France.

During the Santo 2006 expedition deep-sea fishes were sampled using small gears as a dredge and a four meters wide beam-trawl. Around 120 stations were explored in the range of 100 to 1 000 meter depth. More than 800 fish specimens were sampled representing at least 150 species. Small fishes (<5 cm lenght) were directly preserved in 80% ethanol. For other specimens a standardized piece of tissue was preserved in ethanol and the specimen stored in formalin. For each specimen, standardized picture, geographic position and depth of the station were recorded. Specimens were first identified at least to the family level. A database containing all these data was constituted. All specimens were then sequenced for the COI gene and the Rodopsin nuclear gene. Additional COI sequences were obtained from a set of 150 reference species used in phylogenetic analyses of acanthomorph teleosteans (Dettaï and Lecointre, 2005) in order to determine the systematic position of the fishes inventoried in Santo.

Subject: Marine

TESTING BARCODE MARKERS IN ASCOMYCOTA: INTRON DISTRIBUTION ALONG MITOCHONDRIAL GENES.

MONICA SANTAMARIA1, Saverio Vicario1, Claudio Scazzocchio2,3, Cecilia Saccone1,4

1Istituto di Tecnologie Biomediche, Sede di Bari, CNR, 70126 Bari, Italy 2Institut de Gènètique et Microbiologie, UMR 8621 CNRS, Universitè Paris-Sud (XI) 3Institut Universitaire de France, B‚timent 409, Centre d'Orsay, 91405, Orsay Cedex, France 4Dipartimento di Biochimica e Biologia Molecolare, Università di Bari, 70126 Bari, Italy

A rapid, standardized, and cost-effective identification system is needed for Fungi, owing to the frequent lack of distinctive morphological characters and the preponderance of microscopic species. The present work would contribute to the development of effective species molecular markers in Ascomycota, a phylum with a large economic impact both as pests and as beneficial organisms being involved in the quality and safety of agri-food products. Currently, the molecular identification of species in Fungi is based primarily on nuclear DNA, but the potential use of mitochondrial markers has also been considered, along with the possible complications in the PCR- based genetic divergence surveys due to the presence of mobile introns in almost all the mitochondrial genes of these organisms. Our results confirm this trend in Ascomycota: only few sequences seem to be free from introns and, among them, ND6 gene could really be the best barcode candidate.

Subject: Fungi

Second International Barcode of Life Conference 18-20 September 2007 127 Conference Abstracts Poster Abstracts

DERIVING AND USING BARCODE BASED DIAGNOSTICS TO FACILITATE INFORMATION RETRIEVAL

INDRA NEIL SARKAR

MBLWHOI Library, Marine Biological Laboratory, Woods Hole, MA USA

One of the great promises of the Barcode of Life initiative is the integration and linking of data types from multiple, often disparate, sources (including biogeography, images, and literature). As a first step towards this goal, barcode-based diagnostics may facilitate the ability to identify relevant information from existing resources (e.g., GBIF). This poster will first demonstrate how such diagnostics can be derived from publicly available “barcode annotated” sequences (sensu NCBI) using automated techniques, such as the Characteristic Attribute Organization System (CAOS). Next, metrics will be shown for how CAOS-derived diagnostics can be used to assess the classification space of taxonomic hierarchies, through both descriptive statistics and graphically. Finally, this poster will describe how CAOS-based diagnostics can be used as a classifier of new barcode data, within the context of an information retrieval search tool.

Subject: Methods

APPLYING DNA BARCODE ON IDENTIFICATION OF LARVAL FISH CAN IMPROVE COMMUNITY ANALYSIS

Hui-Ling Ko1, Li-Lian Liu1, Hsin-Ming Yeh2, Kui-Ching Hsu3, Ching-Yi Chen3, and KWANG- TSAO SHAO3

1Institute of Marine Biology, National Sun Yat-Sen University 2Coastal and Offshore Resources Research Center, Fisheries Research Institute, COA 3Research Center for Biodiversity, Academia Sinica

Biological communities often show tempo-spatial variations along environmental gradients. However, ichthyoplankton assemblages usually do not show any tempo-spatial distribution along environmental factors. This phenomenon may be due to the species identification problem or the irregularities of ichthyoplankton assemblages themselves. In this preliminary research, we collected 1,549 specimens of fish larvae in the postflexion stage by light trap in the Kenting coral reef area during 2005-2007. Fifteen types of damselfish larvae and five types of butterflyfish larvae were identified based on morphology. Then, we used the COI sequences as DNA barcodes to identify the morphological types of damselfishes and butterflyfishes to species level. Fifteen morphological types of damselfishes (Pomacentridae) could be identified to 11 species and three genera. Five morphological types of butterflyfishes (Chaetodontidae) could be assigned to four species and one genus. After adopting COI identification to the lower taxonomic levels, the ordination analysis clearly revealed a significant connection to the seasonal variation of the larval fishes. In other words, our result showed that applying the DNA barcode technique to identify larval fishes to lower taxonomic categories can indeed further assist the study of temporal or spatial variation of the community structure of ichthyoplankton. Additionally, DNA barcoding can help us to compile the diagnostic keys for fish larvae and publish illustrations or guide books for larval fish identification.

Subject: Fish

Second International Barcode of Life Conference 18-20 September 2007 128 Conference Abstracts Poster Abstracts

CONSTRUCTION OF A DNA DATABASE FOR BARCODING CHINESE MEDICINAL MATERIAL

PANG-CHUI SHAW1, Ka-Lok Wong1 and Paul Pui-Hay But1,2

1Institute of Chinese Medicine, Department of Biochemistry, The Chinese University of Hong Kong, Shatin, N.T., Hong Kong, China 2Institute of Chinese Medicine, Department of Biology, The Chinese University of Hong Kong, Shatin, N.T., Hong Kong, China

The Pharmacopoeia of the People's Republic of China (2005 edition) covers the information of 668 plant species and 104 animal and fungal species. Previously, we have adopted a battery of DNA techniques for the authentication of Chinese medicinal material. At present, we have gathered the concerned DNA sequences available in the GenBank and determined the DNA sequences of additional medicinal plants and animals. The sequences are being complied for generating the first TCM DNA database in the world. Our experience shows that isolation of intact DNA from medicinal material is a limiting factor, owing to the storage and processing of the material. The criteria of a useful DNA barcode for medicinal material are: (1) high interspecific divergence; (2) low intraspecific divergence; (3) short PCR product and (4) available universal primers for DNA amplification. According to these criteria, the differentiating power of five DNA sequences, including the ITS, 5S rRNA spacer, trnL-trnF, psbA-trnH and rpl36-rps8, from four medicinal Gentiana species (Longdan in Chinese) and their adulterants are being tested.

Subject: Plants

DNA BARCODING THE HAWAIIAM RED ALGAL FLORA

SHERWOOD, ALISON1, Kurihara, Akira1, Conklin, Kimberly1, Sauvage, Thomas1, Wang, Norman2, Presting, Gernot2

1Botany Department, University of Hawaii, Honolulu, Hawaii, USA 96822 2Molecular Biosciences and Bioengineering, University of Hawaii, Honolulu, Hawaii, USA 96822

The Hawaiian marine red algal flora has been the subject of intense morphological examination for a number of decades, and currently approximately 370 species are recognized, with a rate of endemism of almost 20%. Several freshwater red algae are also known from the streams in the archipelago. We are currently starting the second year of a three-year effort to DNA barcode the marine and freshwater red algae of Hawaii, as part of a biodiversity survey of the flora. Collections for DNA barcoding originate from many sources: the majority are obtained via seasonal sampling efforts throughout the main Hawaiian islands, but others are included through access to deep-water surveys of the main Hawaiian islands, annual NOAA surveys of the main and Northwestern Hawaiian islands, archived material at Bishop Museum, and the Census of Marine Life collections from the 2006 CReefs expedition to French Frigate Shoals. All collections are vouchered (as herbarium sheets, formalin vouchers, microscope slides, dried vouchers, or in a few cases, photographs) and DNA extracts are archived for most samples. Approximately 250 consensus COI sequences have been obtained thus far, representing 13 orders. We will illustrate progress to date in barcoding this diverse flora.

Subject: Protists

Second International Barcode of Life Conference 18-20 September 2007 129 Conference Abstracts Poster Abstracts

DNA BARCODING TO DISTINGUISH SPECIES OF BUTTERFLIES FROM WESTERN GHATS, INDIA

Bipinchandra Salunke1, Sandeep Amberkar1, Dhiraj Dhotre1, Rakesh Chandode1, Rahul Choudhary1, Avinash Khandagale1, Milind S Patole1, Hemant V Ghate2, and YOGESH S SHOUCHE1

1National Centre for Cell Science, Pune University Campus, Ganeshkhind, Pune411007 INDIA 2Modern College of Arts & Science, Shivajinagar, Pune 411 005 INDIA

The Western Ghats mountain range in peninsular India, one of the Gondwana breakup landmasses, is among the most important global hotspots of biodiversity. It is a unique centre for diversity due to extraordinary endemism and special evolutionary relationships with other biogeographical regions. There are nearly 1400 species of butterflies in India of which about 400 are present in Western Ghats. There is very little information about how many species in India are really endemic. In addition, there is considerable regional, seasonal variation within species. In an attempt to resolve these issues, we have used cytochrome oxidase subunit I DNA barcodes to effectively discriminate amongst species in four butterfly families Pieridae, Papilionidae, Nymphalidae and Lycaenidae from Western Ghats. We found that all the morphologically distinct species that were sampled so far have distinctive cytochrome oxidase barcode. Our results suggest that the butterflies from the Western Ghats are poorly represented in molecular databases such as Genbank and BOLD. Thus our study adds new sequences to the existing vast data available for butterflies. This will help defining phylogeny among oriental as well as world butterflies. These results also show that DNA barcoding can significantly aid species identification of butterflies from India.

Subject: Insects

DNA BARCORDING OF TAIWANESE FISH SPECIES

HORNG-TZER SHY1, Kui-Ching Hsu1, Po-Feng Lee2, Kwang-Tsao Shao1

1Research Center for Biodiversity, Academia Sinica, Taiwan 2Institute of Ecology and Evolutionary Biology College of Life Science, National Taiwan University, Taiwan

Four hundred and seven species of fish (108 families, 33 orders), mostly marine fish from Taiwan, were sequenced (barcoded) for a 600 bp region of the mitochondrial cytochrome oxidase subunit I gene (COI). Most species were represented by multiple specimens. The GC content of the 382 species of the teleosts was higher than the 25 species sharks and rays (46.6% versus 40.2%), largely due to a higher GC content of codon position 3 in the former (42.2% versus 30.1%). Average Kimura two parameter (K2P) distances within-species, genus, family, and order were 0.5%, 11.1%, 22.3% and 26.8%, respectively. Although species compositions have some disparities, the K2P distances data that we obtained were approximately the same as that from the Australian fish species (Ward et al. 2005). All species could be identified by their COI sequence, and the differences between congeneric species were, on average, eleven times higher than the differences within species. Our results also suggest that COI sequencing can be used to identify fish species effectively.

Subject: Fish

Second International Barcode of Life Conference 18-20 September 2007 130 Conference Abstracts Poster Abstracts

THE DNA BARCODING OF CRYPTOSPORIDIUM-BANGLADESH PERSPECTIVE

A.Z. SIDDIKI1 and R. Haque2

1Dept of Pathology and Parasitology, Chittagong Veterinary and Animal Sciences University, Pahartali, Chittagong- 4202, Bangladesh 2Centre for Health and Population Research, International Centre for Diarrhoeal Disease Research, Bangladesh, Dhaka, Bangladesh.

The protistan phylum Apicomplexa comprises a large, heterogenous, assemblage of obligate intracellular parasites classified as coccidian, gregarines, haemogregarines, piroplasms and malarial organisms. Coccidia are exceptionally common protist parasites of both vertebrates and invertebrates. Cryptosporidium is a member of the coccidia which is responsible for widespread zoonotic infection in 152 host species including man. So far, molecular tools have been developed to identify Cryptosporidium at the species/genotype and subtype levels. The techniques are now increasingly used in the characterization of the transmission of Cryptosporidium spp. More importantly the recent development of subtyping tools has led to better understanding of the population genetics and transmission of Cryptosporidium in humans as well as animals. However, the total number of species has not been confirmed yet throughout the world. There are two well known species of Cryptosporidium, namely C. parvum and C. hominis and they have more complicated population structure. We have a collection of over hundred Cryptosporidial DNA samples derived from human patients in Bangladesh (at Laboratory Sciences Division, ICDDRB). Alongside identifying the types and subtypes, the DNA barcoding of those samples using available molecular tools will lead to the identification of geographic and temporal differences in the transmission of C. parvum and C. hominis. The project also will facilitate development of further experimental methodologies for DNA barcoding of other parasitic protozoa.

Subject: Protists

GoogleGene: A VISUAL, WEB-BASED TOOL FOR LINKING DNA BARCODE SEQUENCES TO SPECIES INFORMATION.

SINGER G.A.C.1, Hajibabaei M.2, Nikbakht, H.1, and Hickey, D.A.1

1Concordia University, Department of Biology, 7141 Sherbrooke Street West Montreal, Quebec H4B 1R6 Canada 2Canadian Centre for DNA Barcoding, Biodiversity Institute of Ontario, University of Guelph, 579 Gordon Street, Guelph, ON, Canada, N1G 2W1

We have developed a rapid method, based on the Google search algorithm, to compare an unknown DNA barcode sequence to the DNA barcode database. Rather than simply retrieving matching barcode sequences, however, our method also retrieves information on species images, taxonomy, phylogeny and scientific articles. The taxonomic and phylogenetic information is obtained from independent sources, and is not inferred from the barcode sequences themselves. We believe that this tool will allow a broad community of potential users, including both biologists and non- biologists to fully appreciate the power and utility of DNA barcoding.

Subject: Methods

Second International Barcode of Life Conference 18-20 September 2007 131 Conference Abstracts Poster Abstracts

A GENERALIZED ROC ANALYSIS FOR COMPARING THE EFFECTIVENESS OF VARIOUS LOCI AND DISTANCES FOR PLANT BARCODING

JOHN L. SPOUGE1, D. Erickson', A. Resch1, W.J. Kress2

1National Center for Biotechnology Information 2National Museum of Natural History, 'Smithsonian Institution

The use of DNA barcoding for the identification of the world's 10 million species requires the effective summarization of large amounts of DNA sequence data. This presentation summarizes and then compares the performance of various candidate plant barcode loci (e.g., matK, rbcL, and trnH-psbA) and metrics (e.g., sequence alignment and Kimura distance). First, we assembled a database consisting of GenBank sequences of the candidate plant barcode loci. Second, we calculated nearest neighbors in the sequence database under the various metrics. Third, by extracting each sequence from the database in turn and using its nearest neighbor's distance to classify it, we estimated the probability of a correct answer to the question: does another member of the extracted sequence's taxon remain in the database? We also estimated the probability of correct classification. Our comparison has three merits most analyses lack: (1) it can produce a single figure of merit with an intuitive interpretation; (2) it suppresses distant sequences, which have little influence on classification; and (3) the (largely irrelevant) numerical value of the distance is ignored in favor of ranking criteria.

Subject: Plants

Second International Barcode of Life Conference 18-20 September 2007 132 Conference Abstracts Poster Abstracts

DNA BARCODING NEW ZEALAND MARINE FISHES: PROOF OF PRINCIPLE AND APPLICATIONS

Peter J. Smith1 and DIRK STEINKE2

1National Institute of Water & Atmospheric Research Ltd., Private Bag 14 901 Wellington, New Zealand 2Canadian Centre for DNA Barcoding, Biodiversity Institute of Ontario, University of Guelph, 579 Gordon Street, Guelph, Ontario, N1G 2W1 Canada

New Zealand has the fourth largest Exclusive Economic Zone with over 1200 recorded fish species, of which 380 species and 1100 specimens have been barcoded. A further 38 species and 106 specimens of fishes have been barcoded from the Ross Sea area managed by New Zealand. Average within-species and genus K2P distances were 0.3% and 9.3%, respectively. All species could be differentiated by their COI sequence, although single individuals of 16 species had haplotypes characteristic of a congener. Combined COI and cytochrome b analyses of deepwater (Bathyraja) skates in the Southern Ocean and the Ross Sea have revealed a new species of skate Bathyraja n. sp. cf eatonii that differs from Bathyraja eatonii from the Kerguelen Plateau (the type locality).

Barcoding is being routinely applied to specimen and product identification for fisheries managers and for species verification of record specimens. The southern bluefin tuna Thunnus maccoyii and Pacific bluefin tuna T. orientalis for example can be difficult to discriminate based on external characters, but COI sequences derived from finclip samples differ by 7 base positions making barcoding the preferred identification method. In addition to routine fillet identification, barcoding has been applied to the identification of smoked fish products from a range of fishes. It allowed identification to the species level, even among closely related species in the same genus.

Subject: Fish

ACCELERATED BIODIVERSITY SURVEYS THROUGH DNA BARCODING

DIRK STEINKE, and Paul D.N. Hebert

Canadian Centre for DNA Bardcoding, University of Guelph

Small invertebrates dominate the animal kingdom. Even in well-studied regions, taxonomic resolution is often only possible to a family level, preventing fine-grained assessments of species distributions. Our research seeks to resolve this taxonomic impediment. We are in the process of assembling a DNA barcode library for all animal species at a site in the Canadian arctic - Churchill, Manitoba. This location was chosen because species diversity is relatively low and past taxonomic work has been intense. Despite this fact, members of certain species-rich families of Diptera and Hymenoptera remain effectively unidentifiable. We have developed protocols to quantify species diversity in these groups through the use of rule sets derived from the analysis of lineages with well-established taxonomic systems. Early results, based on the sequencing of 6200 specimens from some 1000 species across varied animal phyla, confirm that congeneric species regularly possess substantial COI sequence divergence whereas conspecifics show little variation. As result, species diversity can be accurately estimated in all groups. Although past studies have demonstrated the effectiveness of DNA barcoding in the identification of known species, this study shows that barcoding can also enable rapid estimates of species diversity in whole ecosystems.

Subject: Methods

Second International Barcode of Life Conference 18-20 September 2007 133 Conference Abstracts Poster Abstracts

THE CENSUS OF ANTARCTIC MARINE LIFE (CAML) BARCODING INITIATIVE

Rachel Grant1 and DIRK STEINKE2

1 CAML Barcoding Data Manager c/o British Antarctic Survey. High Cross, Madingley Road CAMBRIDGE CB3 0ET, United Kingdom 2 Campaign Coordinator, Marine Barcode of Life Biodiversity Institute of Ontario, University of Guelph 579 Gordon Street Guelph, Ontario N1G 2W, Canada

CAML aims to provide and co-ordinate molecular barcode data for all marine Antarctic species, from plankton to top predators. Where possible we will use the standard barcode regions, CO1, but considering other genes where this is difficult to sequence or unable to resolve species-level differences. Certain groups will be prioritized over the course of the project, such as fish where there is a lot of genetic work underway, and rare or difficult to work with groups such as gelatinous zooplankton. An image and voucher reference will be linked to the species and its sequence where possible, along with appropriate metadata. CAML hopes to collect specimens from ten different bio-geographic areas of the Antarctic to allow sequence comparisons. The products of this coordination are anticipated to be available in 2010 and will include a metadata system detailing what data are held in where they are stored, interactive maps linked to sequence information which can be downloaded for bio-geographic comparisons and public-access to Antarctic sequence data via user-friendly web based systems such as BOLD and OBIS.

Subject: Marine

DNAPRINTS - BIONFORMATICS SOFTWARE FOR VNTR ANALYSIS/VISUALISATION

ROBERT STONES and Neil Parkinson

Central Science Laboratory, UK

DNAPrints is a standalone bioinformatics software package developed at the Central Science Laboratory (CSL is a UK government research agency) for the rapid analysis and visualisation of variable number tandem repeats (VNTR). The software incorporates a tandem repeat analysis algorithm with options to change a number of running parameters. The software provides a user interface, which makes analysis and interpretation of potential biomarkers across whole genomic datasets. There is a facility to screen out non-ACGT characters that may lie in whole genomic shotgun sequenced contigs. There is the option to analyse thousands of contigs downloaded from a public genomic sequencing project.

The software has the facility to visualize identified tandem repeats on whole chromosomes within a scalable graphic in the user interface.† The Clustal alignment algorithm has been incorporated to align and compare tandem repeats identified between related organisms/bacterial strains.† DNA Tandem repeats of interest can be exported along with the flanking regions surrounding the repeats. The software has a PCR primer design algorithm for the user to design primers.

The development and provision of bioinformatics software packages to be accessible to laboratory- based scientists; will aid in the rapid identification of potential biomarkers for species identification.

Subject: Methods

Second International Barcode of Life Conference 18-20 September 2007 134 Conference Abstracts Poster Abstracts

DNA BARCODES OF THE GENUS SCIARA (INSECTA, DIPTERA, SCIARIDAE): APPLICATION TO THE STUDY OF ARMYWORMS

MITSUAKI SUTOU1, Toshihide Kato1, and Motomi Ito2

1The University of Tokyo 2The Union of Japanese Societies for Systematic Biology

Since the 19th century, the spectacular migratory habits of sciarid larvae in forests have been documented worldwide; vast numbers of larvae form moving columns, which sometimes reach several meters long. They have been called armyworms. Morphological identification of sciarid larvae is usually difficult, so the cause and function of this interesting habit remain unknown. Although two species of the genus Sciara, S. militaris and S. hemerobioides, have been identified as armyworms in Europe, there are other potential armyworm species in this genus or related genera. Sutou et al. (submitted) conducted a case study addressing the usefulness of DNA barcodes in identifying sciarid larvae.

The genus Sciara contains 19 Palearctic species according to the recent taxonomic studies (Menzel & Mohrig, 2000; Sutou et al., 2004). Of these, we show the immature morphology and DNA barcodes of the following six species with photographs of armyworms recently discovered in Japan: S. hemerobioides, S. helvola, S. humeralis, S. thoracia, S. melanostyla, and S. kitakamiensis. We will determine the DNA barcodes of most of the Palearctic species of this genus in the near future. Using the barcodes as a molecular identification tool, we will conduct an ecological and evolutionary study of armyworms.

Subject: Insects

Second International Barcode of Life Conference 18-20 September 2007 135 Conference Abstracts Poster Abstracts

MOLECULAR PHYLOGENY OF INDO-AUSTRALIAN GLYPHODES AND ITS ALLIED GENERA (LEPIDOPTERA: CRAMBIDAE)

SUTRISNO, HARI1, Azuma, N.2, Higashi, S.2

1Museum Zoologicum Bogoriense, Research Center for Biology –LIPIJl. Raya Bogor-Jakarta Km 46, Cibinong, Bogor, Indonesia 2Graduate School of Environmental Earth Science, Hokkaido University, Sapporo 060-0810, Japan

The phylogeny of Indo-Australian Glyphodes (15 species), Talanga (three species), Agrioglypta (four species) and two outgroup species Ostrinia nubilalis Hubner, 1825 and Metallarcha aurodiscalis (Hampson, 1918) was inferred from nucleotide sequence variation in the mitochondrial COI (686 bp) and COII (687) genes and the nuclear EF-1 alpha (973). While the 3 nd codon position in COI and EF-1 alpha were not saturated with substitution, those in COII were saturated when sequences were compared between genera. MP analysis based on pooled data of COI, COII and EF-1 alpha resulted in two MP trees. The strict consensus tree showed that Glyphodes, Talanga and Agrioglypta were all not monophyletic. Another analysis based on pooled data of all molecular and morphological data resulted in a single tree, in which monophyly of the genus Glyphodes was barely supported (bootstrap value). Glyphodes species were divided into the same three clades as in an earlier published, morphology based tree, one of these clades (they so- called group 3) being supported by a bootstrap value of 100%. In all analysis the genus Agrioglypta was showen to be paraphyletic, since A. eurytusalis (Walker, 1859) fell within the genus Talanga.

Subject: Insects

TARGETS AND CHALLENGES FOR THE FISH BARCODE OF LIFE INITIATIVE (FISH-BOL) TO BARCODE ALL AFRICAN FISH

ERNST R. SWARTZ1, Monica Mwale1, Robert Hanner2, Denis Tweddle1, Roger Bills1, Luis Moreira da Costa1, Elaine Heemstra1, Phil C. Heemstra1 and Paul H. Skelton1

1South African Institute for Aquatic Biodiversity, Private Bag 1018 Grahamstown, 1640, South Africa 2Canadian Centre for DNA Barcoding, Biodiversity Institute of Ontario, University of Guelph, Guelph, Canada

Ichthyologists have organized a Regional Working Group within the FISH-BOL initiative to barcode the diverse African freshwater and marine ichthyofauna. A species checklist is required to organise sampling activities and to gauge progress. FishBase was adopted by FISH-BOL for this. We compared this list with other regional species lists and through consultation with taxonomists developed a more comprehensive checklist of African fishes, focusing on southern Africa and the western Indian Ocean. This checklist that includes more species, is used to develop a strategy for addressing the many challenges faced in barcoding African fishes. The most serious challenge is to collect, curate and analyse species not represented in biomaterial collections. Despite limited funding, infrastructure and few taxonomic experts, the FISH-BOL project in Africa has already barcoded 570 specimens of 328 species (295 marine and 33 freshwater) through collaborative efforts with existing research programmes, demonstrating the ability to differentiate more than 90% of the species with the COI barcode. The collaborative network and these results are promising and suggest that barcoding the fishes of Africa is feasible. Funding, training field workers, establishing links with regional co-ordination centres, and building capacity in curation at regional centres will raise the profile of this initiative.

Subject: Fish

Second International Barcode of Life Conference 18-20 September 2007 136 Conference Abstracts Poster Abstracts

EVOLUTIONARY HISTORY OF THE GENUS SUS IN CYTOCHROME B SEQUENCES

Stefano Mona1, Ettore Randi 2, Marcella Attimonelli3, MILA TOMMASEO-PONZETTA4

1Department of Genetics and Microbiology, University of Bari, Italy 2National Institute for the Study of Wildlife Fauna, Ozzano Emilia (Bo), Italy 3Department of Biochemistry and Molecular Biology, University of Bari, Italy 4Department of Zoology, University of Bari, Italy

The systematic status of species belonging to the genus Sus is still a matter of debate. According to a recent review of its taxonomy, there are 8 living species (Sus scrofa, Sus salvanius, Sus barbatus, Sus verrucosus, Sus celebensis, Sus philippensis, Sus cebifrons and Sus bucculentus) and approximately 22 subspecies (Groves and Grubb, 1993; Groves et al., 1997; Groves, 1997). Among these species, three are critically endangered (Sus salvanius, Sus cebifrons, Sus verrucosus) and one is virtually extinct (Sus bucculentus). Moreover, data are lacking for the various subspecies. To determine the genetic variability and the phylogenetic relationship within the genus Sus, we performed phylogenetic and barcoding analyses of the available species using 172 cytochrome b sequences, as a possible alternative to the use of COI gene region. Bayesian phylogenetic analyses, under different models, showed high level of polyphyly within Sus; at the same time, barcoding analyses were unsuccessful in assigning sequences to the correct species. The results obtained highlighted a recent adaptive radiation of the genus which determined the difficulty of discriminating the various species using mtDNA. However, the morphological diversity among Sus species suggests that natural selection has shaped nuclear (but not mitochondrial) DNA evolution within the various species.

Subject: Mammals

Second International Barcode of Life Conference 18-20 September 2007 137 Conference Abstracts Poster Abstracts

WHAT DNA BARCODES TELL US ABOUT HIDDEN BIODIVERSITY: A STORY OF INTERTIDAL ACORN BARNACLES

LING MING TSANG1, Benny K.K. Chan 2,3, Tsz Huen Wu1, Ka Hou Chu1

1Department of Biology, The Chinese University of Hong Kong 2Research Centre for Biodiversity, Academia Sinica 3The Swire Institute of Marine Science, The University of Hong Kong

Identification and delimitation of species are fundamental to biological sciences. However, phenotypic convergence and plasticity hamper the characterization of species, especially in marine invertebrates. DNA barcode provides a new short-cut for exploring the hidden biodiversity. In this study, we evaluated the applicability of COI barcode system in delimiting species and discovering new species using two common speciose barnacle genera, Tetraclita and Chthamalus, from the Indo-West Pacific as models. Both genera include a wide array of undescribed lineages within nominated species, representing putative cryptic species. Comparison of morphological and ecological characteristics of these genetically diverged groups has revealed that some of the sibling species are morphologically indistinguishable. In contrast, morphologically diverged individuals may be genetically homogenous. Moreover, because some sibling species with sympatric distribution share similar microhabitat, species identification in the field could be extremely difficult. Our results indicate that species cannot be defined by morphological features alone and the DNA barcode system provides a first step in examining species boundaries and discovering cryptic species. Together with multiple lines of information from ecology and morphology, we can explore the hidden biodiversity and lay the foundation for further studies on the ecology and evolution of this important intertidal fauna.

Subject: Marine

DNA-BASED RECOGNITION OF PAPILIONIODEA FROM TAIWAN

WEI-CHI TSO, Wen-Bin Yeh

Department of Entomology, National Chung Hsing University, Taichung 40227, Taiwan

A short fragment of cytochrome oxidase I (COI) is used to establish the nucleotide sequences of Papilionoidea in Taiwan. Five hundred and fifty-one specimens in 179 species of 115 genera and four families, i.e., Papilionidae, Nymphalidae, Pieridae, and Lycaenidae have been investigated. In general, the proportional nucleotide divergence of COI increased with taxonomic distance, with a low level of divergence, less than 1.6%, was observed within species. Nucleotide divergence among species of a given genus was about 2% to 11%, that among genera in the same family was 7% to 17%, and that among families was 13% to 20%. Furthermore, sequences of 89 individuals in 34 papiliolid butterflies from Taiwan, Hong Kong and mainland China have been determined and compared. Subspecies of Byasa alcinous, Papilio demoleus, P. helenus, P. nephelus, and Pazala eurous from different geographic regions were deep mitochondrial divergences should probably be split into separate species. Undifferentiated COI data also suggest that some subspecies of Papilio bianor, P. demoleus, P. memnon, P. nephelus, P. paris, P. polytes, and P. protenor may therefore not to be completely isolated from each other.

Subject: Insects

Second International Barcode of Life Conference 18-20 September 2007 138 Conference Abstracts Poster Abstracts

DNA BARCODING OF THE FLORA OF THE KRUGER NATIONAL PARK (SOUTH AFRICA)

MICHELLE VAN DER BANK1, Renaud Lahaye1, Olivier Maurin1, Sylvie Duthoi1, and Vincent Savolainen2

1Molecular Systematic Laboratory, Department of Botany and Plant Biotechnology, University of Johannesburg, APK Campus, PO BOX 524, Auckland park, 2006, South Africa 2 Jodrell Laboratory, Royal Botanic Gardens, Kew, Richmond, Surrey, TW9 3DS, United Kingdom

This is the first application of the technique of DNA barcoding to a large and renowned area such as the Kruger National Park (KNP). The KNP is one of the largest protected areas in the world with a floral biodiversity estimated at approximately 2000 species. Various habitats and ecological regions exist within the boundary of this Park, with at least 16 macro ecozones recognised, in all covering nearly 2 million hectares. It also now form part of the Great Limpopo Transfrontier Park, which links the KNP with Gonarezhou National Park in Zimbabwe and the Limpopo National Park in Mozambique. UNESCO has designated this new Park as a World Heritage Site. Surprisingly, no large inventory has recently been done in the KNP. We are currently performing a large-scale collection and DNA barcoding of all plant species of the KNP.

Results based on nine plastid DNA regions (coding and non-coding) were evaluated, across a phylogenetically diverse set of taxa which include 20 families, 26 genera and 39 species (in triplicate). Our results suggest that a combination of genes is recommended as a barcode for the flora of the KNP. An overview of the management of samples and data basing of information will also be discussed. This study will improve the knowledge of plant biodiversity of the KNP as well as conservation management by providing exact localities of native, alien, rare and endangered plants. Lastly this study will also reduce or limit plant bio-prospecting in the KNP since Herbarium specimens and DNA samples will be made available to the scientific community.

Subject: Plants

Second International Barcode of Life Conference 18-20 September 2007 139 Conference Abstracts Poster Abstracts

BARCODE DATA SUBMISSION AND RETRIEVAL FROM THE EMBL NUCLEOTIDE DATABASE

BOB VAUGHAN, Guy Cochrane, Rolf Apweiler, Ewan Birney

EMBL-EBI

The EMBL nucleotide database is the European partner in the International Nucleotide Sequence Database Consortium (INDSC), sharing data with Genbank and DDBJ. We provide a variety of data submission methods to the biological community, suitable for both large and small scale datasets, and a wide selection of data retrieval options. Some of these submission and retrieval tools may be of particular interest to the CBOL community. Our web-based submission system, Webin, is adapted to allow quick and efficient submission of small datasets (where the submitter completes the submission themselves), and larger datasets, where a template can be created from an initial submission, and then filled in either by direct entry into the web forms, or by upload of a formatted fasta file. We offer a variety of data retrieval systems, allowing the collection of sets of sequences on the basis of journal citations, geographic location or other more complex criteria via SRS (sequence retrieval system).Again, some of these may be of particular interest to those working with BARCODE data.

Subject: Methods

EXPLORING MORE PHYLOGENETIC EXPLICIT PROTOCOLS FOR SPECIES ASSIGNMENT TO BE IMPLEMENTED IN A BARCODE SERVER

VICARIO, SAVERIO1, Monica Santamaria1, Antonio Turi6,9, Giacinto Donvito4, Maria Mirto6,7,8, Gaetano Scioscia4, Graziano Papada’5, Vicenzo Quinto5, Giovanni Aloisio6,7,8, and Cecilia Saccone1,2

1Istituto di Tecnologie Biomediche, Sede di Bari, CNR, 70126 Bari, Italy 2Dipartimento di Biochimica e Biologia Molecolare, Universit‡ di Bari, 70126 Bari, Italy 3IBM Italy Innovation Center, 70125 Bari, Italy 4INFN, Bari, Italy 5Exhicon ICT, 70125 Bari, Italy 6SPACI Consortium, Lecce, Italy 7ISUFI University of Salento, Lecce, Italy 8CACT of NNL/CNR-INFM, Lecce, Italy 9Dipartimento di Informatica, Universit‡ di Bari, 70126 Bari, Italy

Two aspects of the barcode initiative are the keys for its success: the development of large reference barcode libraries and the setting up of a fast, but accurate procedure to assign new barcode haplotypes to known groups of them. The current assignment of a barcode is based on a very fast procedure that, although very appealing for its implementation ease, is likely to fail when confronted with groups of species having a moderate population structure. We propose a Bayesian phylogenetic inference with a more realistic evolutionary model in which the values of the parameter and topology are pre-estimated on subsets of the reference library. A userís query is directed to the correct subsection by a fast algorithm (BLAST or SVM word count). Then, a fast topological search is implemented to find the best location of the user sequence within the fixed phylogeny of the libraries subsection. This implementation allows having realistic node supports which could be used to identify the species group and assess the adequacy of the assignment.

Subject: Methods

Second International Barcode of Life Conference 18-20 September 2007 140 Conference Abstracts Poster Abstracts

TESTING THE EFFICIENCY OF MT BARCODE MARKERS ACROSS LARGE STRETCHES OF TAXONOMIC DIVERSITY

VICARIO, SAVERIO1, Monica Santamaria1, Giacinto Donvito4, Maria Mirto6,7,8, Gaetano Scioscia3, Graziano Papada’4, Vincenzo Quinto4, Giovanni Aloisio6,7,8, and Saccone Cecilia1,2

The choice of the barcode marker COI stems from a mix of theoretical consideration and historical contingency. To base this choice on a more solid ground, we compare the performance of all the 13 coding mitochondrial genes of Vertebrate from a large group of data set built in an automated way from EMBL. Each data set is composed of at least 5 individuals of a nominal species and at least another co-generic individual. The performance of each gene was tested using two different statistics: first, the comparison between intraspecific and interspecifc divergence, where a bootstrap approach was implemented to assess a confidence interval and then the nodal support for the monophyly of the nominal species. Both statistics derive from a fully-fledged Bayesian phylogenetic inference using as a model of evolution GTR plus gamma distribution for site variation. Using the estimation of site likelihood obtained from those analyses, we were also able to identify the stretch of 600bp maximizing the ratio of likelihood between the topology that presents the monophyly of the nominal species and the one that does not show it.

Subject: Methods

Second International Barcode of Life Conference 18-20 September 2007 141 Conference Abstracts Poster Abstracts

DNA BARCODING REVEALS INCONGRUENCE BETWEEN MOLECULAR CHARACTERS AND NOMINAL SPECIES IN THE CERATITIS FASCIVENTRIS, C. ANONAE, C. ROSA COMPLEX (DIPTERA: TEPHRITIDAE)

MASSIMILIANO VIRGILIO1, Norman Barr2, Marc De Meyer1, Thierry Backeljau3

1Royal Museum for Central Africa, Leuvensesteenweg 13, 3080, Tervuren, Belgium 2PDDML-APHIS-USDA, Moore Air Base, 22675 N. Moorefield Rd., Edinburg, Texas 78541, U.S.A 3Royal Belgian Institute of Natural Sciences, Vautierstraat 29, 1000, Brussels, Belgium

Ceratitis fasciventris, C. anonae and C. rosa (hereafter referred to as FAR) form a complex of polyphagous pests widespread in Africa. Adult male specimens can be readily differentiated by secondary sexual characters but females are almost identical morphologically. The use of DNA barcoding for the molecular characterisation of these species was previously tested by means of mitochondrial (COI, 16S, ND6) and nuclear markers (Period) but none of them allowed to recover the 3 species as monophyletic taxa. In this study we investigate the use of ITS1 as an alternative tool for the molecular diagnosis of these nominal species. Inter- and intra-individual ITS1 polymorphism was therefore analysed in 51 FAR specimens. The alignment of 95 FAR sequences resulted in 35 haplotypes and showed comparable levels of sequence divergence among and within species and within individuals. Shared haplotypes were observed between C. fasciventris and C. rosa and shared indel patterns were observed between C. anonae and C. fasciventris. Taken altogether ITS1 did not allow to recover the three species as monophyletic taxa. Overall, these results suggest that a) the phylogenetic relationships among the three morphospecies are more complex than previously hypothesised and b) the taxonomic status of the three nominal species needs to be further clarified before considering DNA barcoding as an operational tool in their diagnosis.

Subject: Insects

APPLICATION OF DNA BARCODING FOR ENDANGERED GRASSLANDS IN ITALY

MASSIMO VISCHI, Elisa Tomat, Erica Tomat, Alessandro Peressotti, Michele Morgante

Dipartimento di Scienze Agrarie e Ambientali - Università di Udine, Via delle Scienze 208, 33110 Udine, Italy

Intensive agriculture and land-use change during the last century have resulted in the loss of many species-rich grasslands in the Friuli Venezia Giulia region (NE Italy), as it happened throughout NW Europe. These semi-natural permanent grasslands that thrive on shallow calcareous soils support a significant number of flora and fauna species specifically associated with highly stressing habitat conditions (low nutrient content and drought due to the coarse texture of the soil) and very low disturbance (no tilling for even hundreds of years). About 100 different plant species can coexist in such grasslands: many of these are rare or endemic ones and for this reason they have long been recognised as a primary source of biodiversity. Hay cutting for animal nutrition was the main reason for their maintenance in the past, while today, to have a wide acceptance for their conservation, different objectives are to be found. Some DNA barcoding sequences, which at present are proposed for land plants, were tested for their usefulness for managing, preserving and protecting these endangered ecosystems. For these purposes we are considering a metagenomic approach and new sequencing technologies.

Subject: Plants

Second International Barcode of Life Conference 18-20 September 2007 142 Conference Abstracts Poster Abstracts

DNA BARCODING OF MESO-AMERICAN ORCHIDS FOR BIODIVERSITY RESEARCH AND CONSERVATION

JORGE WARNER, Diego Bogarin, Franco Pupulin, Guillaume Gigot, Martyn Powell, and Vincent Savolainen

Lankester Botanical Garden, University of Costa Rica

Costa Rica is one of the richest tropical countries in terms of biodiversity, and especially for orchids with over 1300 species within just 50000 km2. Epiphytic orchids are particularly important as indicators of ecosystem health in tropical areas. Since 2005, the Lankester Botanical Garden (University of Costa Rica) and the Royal Botanic Gardens Kew in the UK have launched an ambitious DNA barcoding project funded by the Darwin Initiative for the Survival of Species (UK). Its aim is to DNA barcode the entire orchid flora of Costa Rica for biodiversity research and conservation. After 22 fieldtrips, we have collected multiple accessions of over 400 orchid species. Testing the DNA regions of the Plant Working Group and others, about 500 trial DNA barcodes have been undertaken for over 50 species, identifying matK as a suitable DNA barcode for this flora. Subsequently a matrix with several matK accessions per species has been assembled for over 200 species. The benefit and caveats of various markers, as well as the practical application of DNA barcoding for trade surveillance of a hyper-diverse flora will be presented. The project will also be discussed within the context of the targets of the Convention on Biological Diversity.

Subject: Plants

A PHYLOGENETIC ANALYSIS OF BRINE SHRIMP IN CHINA USING DNA BARCODING

WEIWEI WANG1,2*, Qibin Luo1,3*, Haiyan Guo1,5*, Songnian Hu1, Peter Bossier3, Gilbert Van Stappen3, Naihong Xin4, Patrick Sorgeloos3, Qishi Sun5, Jun Yu1,3

1Key Laboratory of Genome Sciences and Information, Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing 101300, China 2Graduate University of Chinese Academy of Sciences, Beijing 100094, China 3James D. Watson Institute of Genome Sciences, Zhejiang University, Hangzhou 310008, China 4Laboratory of Aquaculture & Artemia Reference Center, Ghent University, Rozier 44, B-9000 Gent, Belgium 5Salt Research Institute, Yingkou Road 831, 300450 Tanggu, Tianjin, PR China 6School of Traditional Chinese Materia Medica, Shenyang Pharmaceutical University, Shenyang 110016, China

DNA barcoding is a powerful approach to distinguish animal or plant species. In this study, a 648- bp segment of the mitochondrial COI gene was chosen as a standard barcode region to establish polygenetic relationships among Artemia species from major habitats around the world and to further focus on the biodiversity of Artemia species in China, especially in the Tibetan Plateau. Samples from five major salt lakes of the Tibet Plateau located at altitudes of over 4,000 meters, showed clear differences from other populations in China. The variations include two consistent amino acid changes, 153A/V and 183L/F, in COI between high and the low altitude species in China. An Indel in the COI sequence was identified in cyst and adult samples unique to the Co Qen population, demonstrating the need for additional investigations of the mitochondrial genome in Tibetan Artemia populations.

Subject: Marine

Second International Barcode of Life Conference 18-20 September 2007 143 Conference Abstracts Poster Abstracts

DETECTING NEW AND CRYPTIC SPECIES OF CAMAENIDS LAND SNAILS FROM TAIWAN WITH EVIDENCES VIA TRADITIONAL AND MOLECULAR METHODS, AND THE PHYLOGENETIC ANALYSIS WITH DIFFERENT MOLECULAR MARKERS

SHU-PING WU1; Yuh-Wen Chiu2; Hui-Ming Huang1; Yao-Song Lin1 and Chung-Chi Hwang3

1Institute of Ecology and Evolutionary Biology, National Taiwan University. 2Faculty of Biomedical Science and Environmental Biology, Kaohsiung Medical University 3Department of Bioresources, Dayeh University

Traditional taxonomy methods and molecular survey were used to probe into the taxonomic and phylogenetic issues of an endemic arboreal Satsuma snail species complex from Taiwan, the Satsuma albida species complex. In this study, seventeen species were distinguished; fourteen of them were identified and described as new species. Two subspecies of S. albida, S. a. insignis and S. a. mollicula were promoted to independent species based on the characteristic comparison of the genital system and the molecular phylogenetic relationships. Consequently, taxonomy should coincide with the traditional and molecular evidences regardless of different species definition either BSC or PSC.

An explicit phylogenetic relationship of all camaenids land snails from Taiwan were shown, which was based on the molecular phylogeny reflecting their evolutionary history. A total of 52 species were identified, though only 30 are known species on this island. The remnants are undescribed or new species, the biodiversity on the Taiwan Island regarding this family is far higher than known. The universal primers of mtDNA COI gene provided 655 base pairs and can conspicuously discriminate taxa to species level; nevertheless the result reflected by the phylogenetic tree are insufficient, specifically the low bootstraps values at the roots of the phylogram. Elongate marker length or combine sequences from different markers, such as nuclear genes ITS or 28S gene sequences can improve the resolution and bootstraps values at the roots, hence we can construct a well-structured phylogenetic tree.

Subject: Marine

Second International Barcode of Life Conference 18-20 September 2007 144 Conference Abstracts Poster Abstracts

REVISION OF THE Heterocarpus gibbosus Bate, 1888 GROUP (CRUSTACEA: DECAPODA: PANDALIDAE) BY INFERRING TO PARTIAL MITOCHONDRIAL DNA SEQUENCE ANALYSIS

CHIEN-HUI YANG1, Ka-Hou Chu2, and Tin-Yam Chan1

1Institute of Marine Biology, National Taiwan Ocean University, Keelung, Taiwan 2Department of Biology, The Chinese University of Hong Kong, Shatin, Hong Kong

Shrimps of the genus Heterocarpus are common by catches of commercial trawling and trapping from depths of more than 200 meters in tropical and subtropical seas. Heterocarpus gibbosus Bate, 1888 is a widely distributed species in the Indo-West Pacific but with high variations in the development of the rostral crest; from nearly leveled with the carapace to exceptionally high. Recently, a distinct species H. chani Li, 2006 has been separated from H. gibbosus in the Philippines. Other Heterocarpus species closely related to H. gibbosus are H. lepidus De Man, 1917 and H. tricarinatus Alcock and Anderson, 1894 with latter species containing two subspecies. Partial sequences of the mitochondrial 16S rRNA and COI genes were used to examine if the morphological discrepancies observed in the different forms of H. gibbosus are genetically distinct. The molecular data is also used to interpret the phylogenetic relationship of the different taxa in the H. gibbosus group. The COI data set clearly showed that three genetically distinct forms are present in the material currently identified as H. gibbosus but the 16S data set does not. A combined anaylsis of COI and 16S data showed 2-4.6% sequence divergence in the three forms of H. gibbosus and only 0.4-0.8% sequence divergence between the two subspecies of H. tricarinatus. It is concluded that the currently recognized species in the H. gibbosus group are all valid and with two new species in the material previously identified as H. gibbosus. The true H. gibbosus has a moderately high rostral crest while the very low and very high rostral crest form each representing a new species. On the other hand, the two subspecies in H. tricarinatus are likely invalid.

Subject: Marine

Second International Barcode of Life Conference 18-20 September 2007 145 Conference Abstracts Poster Abstracts

IDENTIFICATION OF CETACEAN MEAT IN THE MARKETS OF TAIWAN BASED ON MTDNA MARKERS

CHIOU-JU YAO1,2 Lien-Siang Chou2 Yi-Ju Yang3 Kwang-Tsao Shao1

1Research Center for Biodiversity, Academia Sinica, Taipei, Taiwan, ROC 2Institute of Ecology and Evolutionary Biology, National Taiwan University, Taipei, Taiwan, ROC 3Graduate Institute of Ecology and Environmental Education, National Hualien University of Education, Hualien, Taiwan, ROC

We report the results of molecular identification of cetacean meat in Taiwan markets from 1996 to 2006. As all whales and dolphins are protected under the Wild Animal Conservation Law since 1990, all cetaceans can not be sold or consumed in Taiwan. However, more than fifty illegal trades had been ferreted out by conservation police in the past decade. In order to identify the species of cetacean meat, we have examined thirty-five samples by comparing them with our mtDNA database of twenty-seven cetacean species nearby Taiwanese waters. Either control region or 12SrRNA gene of mtDNA was used for species identification by using parsimony, maximum likelihood, and neighbor-joining methods. Among the examined cases, thirteen spotted dolphins (Stenella attenuata), five Rissos dolphins (Grampus grieseus), four rough-toothed dolphins (Steno bredanensis), two Indo-Pacific bottlenose dolphins (Tursiops aduncus), and two pigmy sperm whales (Kogia breviceps) were identified. The remaining nine samples could only be diagnosed as Dephinidae due to poor supporting values on species category. In addition to species identification, our results suggest that the genera Stenella and Tursiops are polyphyletic, which is also revealed by previous studies based on complete mtDNA cytochrome b gene. In our ongoing project, we are developing an additional database of complete mtDNA COI genes and other nuclear DNA markers in order to compare and improve the resolution of the species identification on cetacean meat or products in the markets of Taiwan.

Subject: Marine

Second International Barcode of Life Conference 18-20 September 2007 146 Conference Abstracts Poster Abstracts

APPLYING DNA BARCODES TO IDENTIFY MEDICINAL PLANTS

Shilin Chen1, HUI YAO1, Jingyuan Song1, Xiwen Li1, Chang Liu2

1Institute of Medicinal Plant Development, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100094, PR China 2Molecular Chinese Medicine Laboratory, LKS Faculty of Medicine, The University of Hong Kong, PR China

DNA barcode as a new technique is a powerful tool for authentication of many animals and plants. However, no one has estimated how well DNA barcode is applied to identify medicinal plants. Medicinal plants and their derived products are not only widely used in Asia, but also popularly used in Europe, Africa and several countries in Americans. In this paper, we make estimation with four molecular tags generally sequenced in plant phylogenetic investigations, the whole chloroplast trnL (UAA) intron, the plastid trnH-psbA intergenic spacer, plastid rbcL and rDNA ITS. Initially, three genus including Panax, Fritillaria and Rheum of medicinal plants are used as materials. These plants are Panax ginseng, P. quinquefolium, P. notoginseng, Fritillaria cirrhosa, F. unibracteata, F. delavayi, F. pallidiflora, F. thunbergii, F. ussuriensis, Rheum palmatum, R. tanguticum and R. officinale. Our results predict available role of the four molecular tags as potential DNA barcodes in the identification of both medicinal plants and their prepared pieces or powders. Furthermore, Taxonomy based on DNA barcode will enhance phylogenetic studies on medicinal plants, and promote development of Pharmaphylogeny and Chemotaxonomy. In conclusion, DNA barcode has a good prospect of application in authentication of medicinal plants.

Subject: Plants

Second International Barcode of Life Conference 18-20 September 2007 147 Conference Abstracts Poster Abstracts

DNA BARCODE DISCOVERS TWO CRYPTIC SPECIES AND TWO GEOGRAPHIC RADIATIONS IN ZAPRIONUS INDIANUS, A RECENT COSMOPOLITAN DROSOPHILID

AMIR YASSIN1,2, Pierre Capy2,3, Lilian Madi-Ravazzi4, David Ogereau2 and Jean R. David2

1Department of Genetics, University of Alexandria, Egypt 2Laboratoire Evolution, Génomes et Spéciation (LEGS), CNRS, Gif-sur-Yvette, France 3Université Paris-Sud XI, Orsay, France 4Depto. Biologia, Unesp, Sao Jose do Rio Preto, 15054-000, SP, Brazil

We studied the distribution of mitochondrial DNA (mtDNA) haplotypes for two genes (COI and COII) among 23 geographical populations of Zaprionus indianus. MtDNA revealed the presence of two well-supported phylads. One phylad included 3 African populations, reinforcing the African- origin of the species. Within the second phylad, a distinct geographical pattern was discovered: Atlantic populations (from the Americas and Madeira) were closer to the ancestral African populations than to Palearctic ones (from Madagascar, Middle-East and India). Discriminant function analysis using 13 morphometrical characters was also able to discriminate between the two molecular phylads (93.34 ± 1.67%), although scanning electron microscopy of male genitalia showed no significant differences. Crossing experiments revealed the presence of reproductive barrier between populations from each phylad, and further between populations within the first phylad. Hence, a bona species status was assigned to two new cryptic species: Z. africanus and Z. gabonicus. The two new species are forest dwellers, which explains their restricted endemic distribution, in contrast to their relative cosmopolitan Z. indianus, known to be a human- commensal. Our results reconfirm the great utility of mtDNA at both inter- and intra-specific levels within the frame of an integrated taxonomical project.

Subject: Insects

Second International Barcode of Life Conference 18-20 September 2007 148 Conference Abstracts Poster Abstracts

MICROPLATE-BASED METHOD FOR DNA BARCODING

WEN-BIN YEH, Wei-Chi Tso

Department of Entomology, National Chung Hsing University, Taichung 40227, Taiwan

In this study, we used an efficient and labor-saving method for barcoding assay. Ninety-six samples can be handled at the same time by using microplate and multichannal pipette for DNA extraction, PCR amplification and purification of PCR products by ExoI/SAP clean-up (Exonuclease I/ Shrimp alkaline phosphatase), and the cleaned-up DNA samples in the microplate are ready for sequencing. In comparison with the traditional procedures by which we spent ca. a year intermittently collecting samples and doing sequence analyses of cytochrome oxidase I (COI) from 68 nymphalid and 24 pierid butterflies; the microplate-based high throughput method allowed us to obtain the sequences in less than ten days once a batch of 96 samples are ready for the analyses. Preliminary work shows that this 96-well microplate-based system makes DNA sequencing faster and cheaper for tissues or specimens in the cryobanks of genetic materials. In 244 individuals of 110 species from Nymaphalidae and Pielidae, we found the proportional nucleotide divergence of COI increased with taxonomic distance, while a low level of divergence, less than 0.8%, was observed within species. Nucleotide divergence among species of a given genus was ca. 2% to 11% while that among genera of the same family was 8% to 17%; and divergence between Nymphalidae and Pielidae families was 13% to 19%.

Subject: Methods

Second International Barcode of Life Conference 18-20 September 2007 149 Conference Abstracts Poster Abstracts

DNA BARCODE ANALYSIS FOR KOREAN BIRDS

HYE SOOK YOO1, Jae-Yong Eah1, Jong Soo Kim2, Young-Jun Kim3, Mi-Sook Min3, Woon Kee Paek4, Hang Lee3, Chang-Bae Kim1

1Biological Resource Center/Korean BioInformation Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon 305-333, Korea 2Hankuk Academy of Foreign Studies, Yongin 449-854, Korea 3Conservation Genome Resource Bank for Korean Wildlife (CGRB), College of Veterinary Medicine, and BK21 Program for Veterinary Science, Seoul National University, Seoul 151-742, Korea 4Natural History Team, National Science Museum, Daejeon 305-705, Korea

The DNA barcoding, inventory of DNA sequences come from a standardized genome region, provides bio-barcode for identifying and discovering bird species. Several recent studies suggest that sequence diversity in a 648bp region of the mitochondrial gene for cytochrome c oxidase I (COI) might serve as a DNA barcode for identifying North American bird species. The present study tested the effectiveness of a COI barcode in discriminating Korean bird species with more comprehensive specimens. We determined 5' terminus of COI barcode for 266 specimens of 105 species of Korean birds and found that species identification was unambiguous; the genetic differences between closely related species were, on average, 25 times higher than the differences within species. We also identified two potential cryptic species, calling for further investigation using more samples. The finding of large COI sequence differences between species confirms the effectiveness of COI barcodes for the identifying of Korean bird species. To bring greater reliability to the identification of species, increased intra- and interspecies sampling including southeast Asian regions, as well as supplementation of the mitochondrial barcodes with nuclear ones is strongly needed. With these data we are developing the microarray chip for identification of bird species responsible for bird striking and suspected as vector of Avian influenza.

Subject: Birds

Second International Barcode of Life Conference 18-20 September 2007 150 Conference Abstracts Poster Abstracts

ALL-CERAM-JPN: A BARCODING PROJECT ON THE LONGICORN BEETLES OF JAPAN

HIRAKU YOSHITAKE1, Nobuo Ohbayashi2, Utsugi Jinbo1, Motomi Ito1, Tatsuya Niisato3 and Hiroshi Makihara4

1The Japanese Barcode of Life Initiative, GBIF Japan Portal; 2Ehime University 3Bioindicator Co., Ltd. 4Forestry & Forest Products Research Institute, Incorporated Administrative Agency

The longicorn beetles belonging to the families Cerambycidae and Disteniidae (Insecta: Coleoptera) are one of the most well-studied groups in Japan, mainly due to their importance in forestry and agriculture. However, many Japanese taxa are still in need of systematic revision at higher level, because phylogenetic relationships among Japanese species have never been studied satisfactorily until now. In addition, some Japanese species are very difficult to identify due to the presence of species that are morphologically similar. Recently, under the leadership of the Japanese Barcode of Life Initiative and Ehime University, we started a project to make a DNA barcode inventory of 946 species-group taxa of longicorn beetles known to occur in Japan. The project was initiated in order to facilitate further studies and species identification based on molecular data. The first step in development of the project is the construction of a comprehensive collection of 99.5% ethanol preserved specimens in cooperation with many professional and amateur researchers. In this presentation, we report on our current progress and the future direction of All-Ceram-JPN.

Subject: Marine

CLASSIFICATION AND CLUSTERING FOR DNA BARCODE DATA

CHING-RAY YU

Department of Statistics and Biostatistics, Rutgers University.

DNA barcode data involved rich information in different biological species. Study showed that sequence divergences at COI (648 base pairs) regularly enable the discrimination of closely allied species in all animal phyla except the Cnidaria. Statistical methods, Clustering and Classification, can be applied to classify the species.

In this talk, we convert the nucleotides {A, T, C, G} into numerical numbers {XA, XT, XC, XG}. We put location and scale constrains on the X’s, and then we estimate X’s optimizing some criteria. We produced two alternative algorithms for estimating the optimal weights. Once the data is in a numeric form the ABC algorithm or any other standard algorithm could be applied for classification and clustering. We use database Birds of North America – Phase II by K.C.R. Kerr et al. to evaluate the performance of our methods.

Subject: Methods

Second International Barcode of Life Conference 18-20 September 2007 151 Conference Abstracts Poster Abstracts

GIBELLULOPSIS, A SUITABLE GENUS FOR VERTICILLIUM NIGRESCENS, AND MUSICILLIUM, A NEW GENUS FOR V. THEOBROMAE

RASOUL ZARE1, W. Gams2, M. Starink-Willemse2 & R.C. Summerbell3

1 Department of Botany, Iranian Research Institute of Plant Protection, P.O. Box 1454 Tehran 19395, Iran 2 Centraalbureau voor Schimmelcultures, P.O. Box 85167, 3508 AD Utrecht, Netherlands 3 27 Hillcrest Park, Toronto Ont. M4X 1E8, Canada

Verticillium nigrescens is not congeneric with Verticillium s. str., as shown in cladograms based on LSU and ITS sequences. Both genera and a few other taxa form a separate, new family, the Plectosphaerellaceae, in which Plectosphaerella cucumerina is the only known teleomorph taxon. The species is conspecific with Gibellulopsis piscis, which provides a suitable generic name. The available isolates are not completely homogeneous as shown in tef1 analysis. This frequent saprotrophic species is neotypified with an isolate from a Dutch soil. It has hyaline, rather short conidiophores arising from vegetative hyphae generally in terminal position, with one or two scant whorls of aculeate phialides. Brown, intercalary or terminal chlamydospores give the colony a more or less dark grey aspect. In the same family, the causal agent of cigar-end rot of bananas, Verticillium theobromae, is also generically distinct and described in the new genus Musicillium.

Subject: Fungi

INITIATIVE DNA BARCODING RESEARCH IN INDONESIA

M SYAMSUL ARIFIN ZEIN, Sri Sulandari, Dwi Astuti, and Hari Sutrisno

Research Center for Biology, the Indonesian Institute of Sciences (LIPI), Indonesia

Research Center for Biology, the Indonesian Institute of Sciences has conducted ex-situ conservation in the form of tissue and blood collection (as material DNA) on Indonesian fauna since 2002. Total number collections are still 20% of number animals available in Indonesia: 1941 bird, 2100 mammals, 741 herpetofauna, and 50 lepidoptera samples. With availability high valuable of the DNA material at the institute and as competence institute for taxonomy field, therefore Research Center for Biology, the Indonesian Institute of Sciences can be pioneer to initiative for doing research on fauna DNA barcoding in Indonesia. As mentioned from literature review: DNA Barcoding is a technique for species identification on fauna by using DNA sequence standard. Barcoding is a useful systematic tool especially in connection with classical methods. It represents a valuable complement to conventional practice when applied to problems beyond the reach of morphological studies, such as the determination of larval stages. DNA barcoding studies detached from the fundament of integrative systematic research, however, are problematical. Cytochrome c Oxidase I (CO I) gene as ‘DNA barcode ‘ will be used in barcoding research.

No DNA barcoding research has been conducted in Indonesia, except a study conducted by Sutrisno et al. (2006) using COI that resolves the phylogenetic relation of glyphodes moths (Lepidoptera). Therefore, a DNA barcoding campaign is the first step that we will do it urgently in Indonesia. We want to do initiative work on DNA barcoding in 2008. Hopefully, the result will solve problems in taxonomy studies. In order to obtain advance results, we will try to make a collaborative research on DNA barcoding with international institute.

Subject: Initiatives

Second International Barcode of Life Conference 18-20 September 2007 152 Conference Abstracts Poster Abstracts

BARCODING EPHEMEROPTERA, PLECOPTERA, AND TRICHOPTERA OF NORTH AMERICA

XIN ZHOU1, Mehrdad Hajibabaei1, Tim Pascoe2, Donald Baird3, and Paul Hebert1

1Canadian Centre for DNA Barcoding, Biodiversity Institute of Ontario, University of Guelph, Guelph, Ontario, Canada N1G 2W1 2Environment Canada, Canada Centre for Inland Waters, 867 Lakeshore Road, Burlington, Ontario, Canada L7R 4A6 3Canadian Rivers Institute, Department of Biology, 10 Bailey Drive, PO Box 45111, University of New Brunswick, Fredericton, New Brunswick, Canada E3B 6E1

The three insect orders Ephemeroptera (mayflies), Plecoptera (stoneflies), and Trichoptera (caddisflies), or EPT, are aquatic macroinvertebrates commonly encountered in various freshwater habitats. Because of their great diversity and abundance, they are widely used as bioindicators of water quality. However, the difficulties in species-level identification, especially of immature stages, have hindered such activity. The DNA barcoding initiative provides an alternative route in species identification with a number of advantages that complement the conventional morphological approach. For example, DNA barcoding can improve resolution on taxonomic identification by discriminating cryptic species and enabling confident association of larval and adult stages. We conducted a pilot study to test the feasibility of high throughput barcoding in aquatic macroinvertebrates. Our initial results suggest that specimens collected by conventional methods in many bioinventory and biomonitoring programs may serve as good sources of DNA for barcoding. COI barcodes were also collected from more than 1600 EPT samples collected from Churchill, Manitoba, Canada, from 2004 to 2006, and from the Great Smoky Mountains National Park, USA in 2007. These specimens cover a variety of species and life stages (male and female adults, pupae, and larvae or nymphs). COI has shown its great potential in recognizing EPT species and associating life stages.

Subject: Insects

Second International Barcode of Life Conference 18-20 September 2007 153 Conference Abstracts Poster Abstracts

DNA DAMAGE IN PRESERVED SPECIMENS AND TISSUE SAMPLES: A MOLECULAR ASSESSMENT

JUERGEN ZIMMERMANN1, Elizabeth Cantin1, Mehrdad Hajibabaei2, David Blackburn3, James Hanken3, Thomas C. Evans, Jr.1

1New England Biolabs Inc. 240 County Rd. Ipswich, MA, 01938 USA 2Canadian Centre for DNA Barcoding Biodiversity Institute of Ontario University of Guelph Guelph, Ontario, N1G 2W1 Canada 3Harvard University Museum of Comparative Zoology Herpetology Department 26 Oxford Street Cambridge, MA 02138 USA

The extraction of genetic information from preserved tissue samples and museum specimens is of great interest in many fields of research, including the ongoing barcoding efforts, forensic studies, scat sample analysis, and cancer research utilizing formalin-fixed paraffin-embedded tissue. Efforts to obtain genetic information from these sources are often hampered by the poor quality of extractable DNA, which leads to poor or no PCR yields.Numerous studies in the past have highlighted techniques for improved DNA extraction from such samples or focused on the effect of damaging agents - such as light, oxygen, formaldehyde - on free nucleotides.Here, we present ongoing work to characterize lesions in DNA samples extracted from preserved specimens. The extracted DNA is digested with a combination of DNase I, Snake Venom Phosphodiesterase and Antarctic Phosphatase to single nucleosides and then analyzed by HPLC-ESI-TOF-MS.We present data for moth specimens that were stored pinned, and for frog tissue samples, preserved either in ethanol or formaldehyde; or fixed in formaldehyde and then preserved in ethanol. We observe changes in the nucleoside content of these samples over time, characterize the fragmentation state of the DNA and aim to identify some more abundant nucleoside lesions.

Subject: Methods

Second International Barcode of Life Conference 18-20 September 2007 154 Conference Abstracts

Blohm, D · 24 Blohm, D. · 82 A BOAKYE, DANIEL A. · 76 Bogarin, Diego · 143 Bogdanowicz, Wieslaw · 121 Adetunji, O.A. · 71 Boisselier, Marie-Catherine · 21, 124 Aladele, S.E · 71 Bonillo, Céline · 126, 127 Aloisio, Giovanni · 140, 141 Borisenko, Alex V. · 105 Amberkar, Sandeep · 130 Bossier, Peter · 143 AMEKA, GABRIEL · 68 Botla, S · 24 ANDERSEN, ROBERT A. · 37 Bouchet, Philippe · 124 Anil, Arga Chandrashekar · 93 Brisset, Julien · 127 Antoniou, A · 24 BRIX, SASKIA · 77 Antunes, Pedro · 97 Brown, Charles A. · 76 Apweiler, Rolf · 140 BUCKLIN, ANN · 17, 77 Araújo, C Castro · 68 Burns, John M. · 33 Arca, M. · 72 But, Paul Pui-Hay · 129 ARITUA, VALENTE · 69 Byrkjedal, Ingvar · 100 Armstrong, Karen · 74 ARMSTRONG, KAREN · 70 Armstrong, Karen F. · 74 Arvanitidis, C. · 82 C Asahida, Takashi · 122 Ashlock, Dan · 101 CAMERON, KENNETH M. · 78 Astuti, Dwi · 152 Campbell, A.J. · 21 Atkinson, J. H. · 69 Campo Falgueras, D · 24 ATOYEBI, O. JOHN · 71 Cantin, Elizabeth · 154 Attimonelli, Marcella · 137 Capy, Pierre · 148 AUSTERLITZ FREDERIC · 72 Cariani, A. · 82 AUSTERLITZ, FREDERIC · 25 Carine, Mark · 105 Avinash Khandagale · 130 Cecilia, Saccone · 141 Azuma, N · 136 Chan, Benny K.K. · 138 Chan, Loucia · 118 Chan, Tin-Yam · 145 B Chandode, Rakesh · 130 CHANG, CHIH-HAN · 79 Chao, Hsin-Yun · 42 Backeljau, Thierry · 142 Chase, Mike · 68 BAGLEY, MARK · 16 Chen, C. Allen · 122 BAILLY, NICOLAS · 73 Chen, Chaolun Allen · 84, 116 Baird, Donald · 98, 153 Chen, Chienhsun · 116 Baker, Andrew · 45 Chen, Ching-Yi · 128 Bakker Freek T. · 56 Chen, Chun-Jui · 119 BAKKER, FREEK T. · 56 CHEN, I-SHIUNG · 80 Balanov, A.A. · 109 Chen, Jiun-Hong · 79 BALL, SHELLEY · 74 Chen, Paul Yu · 119 BALL, SHELLEY L. · 73, 74 Chen, Shilin · 147 Banerjee, M · 78 Chen, Tai-Been · 119 BARCENAS, ROLANDO T. · 75 CHEN, WEN · 81 Barr, Norman · 142 Chen, Y.-H · 119 Barrett, Spencer CH · 60 Chen, Ying · 115 BARTELS, PAUL · 53 Chen, Ying-Rong · 42 Beaton, Margaret · 92 Chichvarkhin, A.Y. · 109 BERMINGHAM, ELDREDGE · 28 Chik, Stanley Chi Chung · 118 Bertolazzi, Paola · 26 CHITIPOTHU, SRUJANA · 82 Bhuiyan, M.K. · 122 Chiu, Yuh-Wen · 144 Bills, Roger · 136 CHO, SOOWON · 83 Bipinchandra, Salunke · 130 Chou, Lien-Siang · 146 Birks, Sharon · 102 Choudhary, Rahul · 130 Birney, Ewan · 140 CHRISTIDIS, LES · 32 Blackburn, David · 154 Chu, Ka Hou · 138 Bleakley, Kevin · 25 CHU, KA HOU · 85

Second International Barcode of Life Conference 18-20 September 2007 155 Conference Abstracts Chu, Ka-Hou · 145 Chuang, Yao-Yang · 116 F CHUANG, YAOYANG · 84 Chung, Jinwook · 23 FAITH, DANIEL P. · 45 Claereboudt, Michel · 84 FELICI, GIOVANNI · 26 Clare, Elizabeth L. · 101 Fernandez-Manjarres J.F. · 72 Cochrane, Guy · 140 Ferrand, N · 68 Conklin, Kimberly · 129 FERRELL, JILLIAN · 92 COOK, LYN · 59 Fischer, Alison · 97 Cooper, Jason · 99 FISHER, BRIAN L. · 41 Copley, Nancy J. · 77 FLOYD, ROBIN · 92 Cottrill, Karin · 99 Flynn, Alan · 70 COWAN, ROBYN · 57 Foottit, Robert · 92 CRAYN, DARREN · 85 Frascaria-Lacoste N. · 72 Cruickshank, R.H. · 124 Fukami, Hironobu · 84 Currie, Doug · 86 CYWINSKA, ALINA · 86 G

D G.A.C, SINGER · 131 Gams, W. · 152 Dai, Chang-Feng · 84 GAONKAR, CHETAN A. · 93 Dapkey, Tanya · 33 Garbelotto, Matteo · 125 David, Jean R. · 148 Garcia Vazquez, E · 24 David, Olivier · 25 Gaudeul M. · 72 Davie, Peter F. · 114 Ghate, Hemant V · 130 Davies, Neil · 125 GIBBS, JASON · 93 DAYRAT, BENOÎT · 29 Gigot, Guillaume · 143 DE BARRO, PAUL · 34 Gillespie, Rosemary · 125 de Cook, Steve C. · 90 Golding, G.B. · 118 De Meyer, Marc · 142 GRAHAM, SEAN W · 60 DE MEYER, MARC · 13 Grainger, Chris M. · 105 de Souza, Dziedzom K. · 76 Grant, Rachel · 134 Deck, John · 125 Gross, Joyce · 125 Dettaï, Agnés · 127 Guo Haiyan · 143 Dettaï, Agnès · 126 GUO, HAIYAN · 94 DEWAARD, JEREMY R. · 87 Gusev, Alexander · 27 Dewan, S. · 122 GWO, JIN-CHYWAN · 94 Dhotre, Dhiraj · 130 DIAZ DE ASTARLOA, JUAN M. · 88 Ding, Qiong · 115 H DONOGHUE, MICHAEL J. · 43 Donvito, Giacinto · 140, 141 Hajibabaei, M. · 131 Dove, Carla J. · 102 Hajibabaei, Mehrdad · 33, 153, 154 Draber-Monko, Agnieszka · 121 HAJIBABAEI, MEHRDAD · 55, 95, 96, 97, 98 Driskell, Amy · 52 Hallwachs, Winnie · 33, 96 Dungan, Roger · 74 Hanken, James · 154 Duthoi, Sylvie · 139 Hanna, Tiarella · 97 Hanner, Robert · 136 HANNER, ROBERT · 15, 19, 99, 100 E Haque, R. · 131 Hauschild, J · 24 Eah, Jae-Yong · 150 Hawkins, Julie A. · 75 Edirisinghe, J. P. · 110 Haxaire, Jean · 30 EDWARDS, JIM · 51 Hebert, Paul · 33, 98, 153 Erickson, D. · 132 HEBERT, PAUL · 18, 101, 102 Erickson, David L. · 58 Hebert, Paul D. N. · 30, 96 ERPENBECK, DIRK · 89, 90, 91 Hebert, Paul D.N. · 22, 92, 133 Evans, Alison · 74 Heemstra, Elaine · 136 Evans, Jr. Thomas C. · 154 Heemstra, Phil C. · 136 EVANS, TOM · 22 Henderson, Rosa · 74 Hervet, C · 24

Second International Barcode of Life Conference 18-20 September 2007 156 Conference Abstracts Hickey, D.A. · 131 KARTAVTSEV, YURY · 109 Higashi, S. · 136 KARUNARATNE, W. A. I. P. · 110 Hinsinger D.D. · 72 Karuneratne, Lalitha · 70 Hisinger, Damien · 127 Kato, Toshihide · 135 Hjörleifsdottir, S. · 24 Kelly, David · 74 Ho, Hsuan-Yi · 42 Kentros, Sotirios · 27 Hong, Yong · 79 Kerr, Kevin C.R. · 102 Hooper, John N.A. · 90 Kim, Chang-Bae · 150 Hreggvidsson, G.O · 24 KIM, CHOONG-GON · 111 HSIAO, SHENG-TAI · 103 Kim, Choongon · 23 Hsu, Kui-Ching · 128, 130 Kim, Go Eun · 111 HSU, KUI-CHING · 104 Kim, Goeun · 23 Hsu, Te-Hua · 94 Kim, Jeong-Seong · 112 Hu, Songnian · 143 Kim, Jong Soo · 150 HUANG, DA-WEI · 104 KIM, KI-JOONG · 112 Huang, Hui-Ming · 144 Kim, Won · 108 Humble, Leland · 92 KIM, WON · 9 Humble, Leland M. · 87 Kim, Young-Jun · 150 Hunt, Jeff · 52 King, Steve · 99 Hunter, Fiona. F. · 86 Kitching, Ian J. · 30 Husband, Brian C · 60 Klaere, Steffen · 45 Huxley, Rob · 105 Knowlton, Nancy · 84, 123 Hwang, Chung-Chi · 144 Ko, Hui-Ling · 128 Hwang, Seongyong · 23 Kochzius, M. · 82 KOCHZIUS, MARC · 24 Königslöw, Taika von · 101 I Koo, Hyeyoung · 108 KRESS, W. JOHN · 58 Kress, W.J. · 132 Ida, Hitoshi · 122 Ku, Chun-Yi · 117 Ikonomi, Pranvera · 99 KUKSA, PAVEL P. · 26 Ito, Motomi · 106, 135, 151 KUMAR, N. PRADEEP · 113 Ivankov, V.N. · 109 Kuo, M.-H. · 119 Ivanova, Natalia V · 99 Kurihara, Akira · 129 IVANOVA, NATALIA V. · 105 IVANOVA, NATALIA V. · 22 L J Lahaye, Renaud · 139 LAI, JOELLE C.Y. · 114 Jaeger, J. · 82 Lai, Jwu-Lan · 103 Jambulingam, P. · 113 Lakra, W. S. · 108 JAMES, KAREN · 105 LAKRA, WAZIR S. · 40 James, Samuel W. · 79 LAMBERT, DAVID · 54 Janzen, Daniel H · 96 Lambourdiëre, Josie · 21 JANZEN, DANIEL H. · 33 Landi, M · 24 Jaspers, Marlene V. · 73 Landi, M. · 82 Jennings, Robert M. · 77 LANE, RICHARD · 46 Jeong, Dageum · 23, 111 Laredo, Catherine · 25 Ji, Eun Lim · 83 Lau, Allan SY · 118 Jinbo, Utsugi · 151 Lecointre, Guillaume · 126, 127 JINBO, UTSUGI · 106 Lee, Hae-Lim · 112 JOSHI, RAVI C. · 107 Lee, Hang · 150 Lee, Po-Feng · 130 LEE, YEN-CHEN · 114 K Lee, Youn-Ho · 111 LEE, YOUN-HO · 23 Kalyakin, Mikhail · 102 Lévesque, C. André · 81 KANG, SEUNG HYUN · 108 Lewis, M. L. · 107 Kao, Wei-Ling · 117 Li, Chi Pang · 85 KAPOOR, SMITA · 108 Li, Guan-Cheng · 119 Kappel, K · 24 Li, Xiwen · 147

Second International Barcode of Life Conference 18-20 September 2007 157 Conference Abstracts Liang, Hong-Yen · 103 LIANG, YU · 115 N Lijtmaer, DarÌo A. · 102 Lilley J. C. · 69 Nabulya, G · 69 LIN, MEIFANG · 116 Nakachwa, R · 69 LIN, SI-MIN · 42, 117 Nam, Ki-Woong · 111 Lin, Yao-Song · 144 Namaganda, J · 69 Lindsay, James · 27 NATARAJAN, CHANDRASEKHAR · 78 LINTON, YVONNE-MARIE · 12 Neetha N.V. · 78 LITTLE, DAMON P. · 27 Newmaster, Steve · 60 Liu, Chang · 147 NG, PETER · 35 LIU, CHARLIE CHANG · 118 Ng, Peter K.L. · 114 Liu, Li-Lian · 128 Nigro, Lisa M. · 77 LOU, MELANIE · 118 Niisato, Tatsuya · 151 LU, HENRY HORNG-SHING · 119 Nikbakht, H. · 131 LU, K.-H. · 119 Nishida, Mutsumi · 44 Lue, Kuang-Yang · 42, 114 Noelte, M. · 82 Luo, Qibin · 143 Nölte, M. · 24 Nomura, Keiichi · 122 Norrbom, Allen · 13 M

M. Starink-Willemse, M. · 152 O M. T. M. · 82 Ma, Keping · 115 Obura, David · 84 Macpherson, Enrique · 21 Odofin, W.T · 71 Madi-Ravazzi, Lilian · 148 Ogereau, David · 148 Magoulas, A · 24 Ohbayashi, Nobuo · 151 Magoulas, A. · 82 Olteanu, Madalina · 25 Makihara, Hiroshi · 151 Onyia, C. · 71 MALEWSKI, TADEUSZ · 121 Ormos, Andrea · 52 Mandoiu, Ion · 27 Ortman, Brian D. · 77 Marteinsson, V · 24 Ouma, Johnson · 38 MASIGA, DANIEL · 38 Oyedapo, O.O. · 71 Maurin, Olivier · 139 OYIEKE, HELIDA · 10 Maw, Eric · 92 Ozouf-Costaz, Catherine · 126 MCDEVIT, DAN · 121 McPheron, Bruce · 13 Meusnier, Isabelle · 97 P MEYER, AXEL · 36 Meyer, Christopher · 125 Pae, Sejin · 23 MEYER, CHRISTOPHER · 20 Pae, Se-Jin · 111 Meyer, Jean-Yves · 125 Paek, Woon Kee · 150 Michel Veuille · 25 Papada, Graziano · 140, 141 Milind S Patole · 130 Park, Heungsik · 23 Millar, Craig · 54 Park, Heung-Sik · 111 MILLER, SCOTT E. · 11 Park, Hyungjin · 83 Min, Mi-Sook · 150 Park, Ik Ju · 83 Mirto, Maria · 140, 141 Park, Tae Seo · 108 Mishler, Brent · 125 Parkinson, Neil · 134 MITCHELL, ANDREW · 21 PASANIUC, BOGDAN · 27 MIYA, MASAKI · 44 Pascoe, Tim · 98, 153 Mona, Stefano · 137 PASHA, MOSTAFA K. · 123 Moreira da Costa, Luis · 136 Paulay, Gustav · 125 Morgante, Michele · 142 Pavlovic, Vladimir · 26 Moritz, Craig · 125 Payri, Claude · 125 MRIDHA, M.A.U. · 122 Pei, Kequan · 115 Muir, Carol · 70 Peressotti, Alessandro · 142 Mwale, Monica · 136 PILLAY, K. RUBY MOOTHIEN · 122 PLAISANCE, LAETITIA · 123 Planes, S · 24 Planes, Serge · 125

Second International Barcode of Life Conference 18-20 September 2007 158 Conference Abstracts POLASZEK, ANDREW · 50 SHAW, PANG-CHUI · 129 PONTIN, DAVID R. · 124 Shen, Huei-Ping · 79 Pöppe, Judith · 90 SHERWOOD, ALISON · 129 Powell, Martyn · 143 Shih, Jar-Hong · 80 Presting, Gernot · 129 SHOUCHE, YOGESH S · 130 Puillandre, Nicolas · 21, 127 SHY, HORNG-TZER · 130 PUILLANDRE, NICOLAS · 124 SIDDALL, MARK E. · 31 Pupulin, Franco · 143 SIDDIKI, A.Z. · 131 Silkenbeumer, N · 24 Singer, Greg · 95 Q Skelton, Paul H. · 136 Smith, Alex · 33, 41 Smith, Peter J. · 133 Quinto, Vicenzo · 140 Sokolovsky, A.S. · 109 Quinto, Vincenzo · 141 Solé, Magali · 97 Solomon, B. · 71 Song, Jingyuan · 147 R Sorgeloos, Patrick · 143 Spencer, Mark · 105 Rahman, M.S.R. · 122 SPOUGE, JOHN L. · 132 Rajavel, A.R. · 113 STEINKE, DIRK · 134 Raman, A. · 21 STEINKE, DIRK · 133 Randi, Ettore · 137 STOECKLE, MARK · 14 Resch, A. · 132 Sulandari, Sri · 152 Rey, Olivier · 126 Summerbell, R.C. · 152 Reza, M. I. H. · 123 Sun, Qishi · 94, 143 Rigg, K.A. · 21 SUTOU, MITSUAKI · 135 RODERICK, GEORGE · 125 Sutrisno, Hari · 152 Rodriguez, Josephine · 33 SUTRISNO, HARI · 136 RONAGHI, MOSTAFA · 24 SWARTZ, ERNST R. · 136 ROSSMAN, AMY · 61 Sykes, Greg · 99 ROUGERIE, RODOLPHE · 30 Royse, Daniel · 97 RUBIM, ALMEIDA-DA-SILVA · 68 T Rumsey, Fred · 105 Terashima, Hiroaki · 122 Tinti, F · 24 S Tinti, F. · 82 Tomat, Elisa · 142 Saccone, Cecilia · 127, 140 Tomat, Erica · 142 Samadi, Sarah · 124 TOMMASEO-PONZETTA, MILA · 137 SAMADI, SARAH · 127 TSANG, LING MING · 138 SAMADI, SARAH · 21, 126 Tseng, Ching-Chi · 84 Santamaria, Monica · 140, 141 Tso, Wei-Chi · 149 SANTAMARIA, MONICA · 127 TSO, WEI-CHI · 138 SARKAR, INDRA NEIL · 128 TUBARO, PABLO · 39 Saunders, Gary W. · 121 Tubaro, Pablo L. · 102 Sauvage, Thomas · 129 Turan, C · 24 SAVERIO, VICARIO · 140, 141 Turi, Antonio · 140 Savolainen, Vincent · 139, 143 Tushemereirwe, W · 69 Scazzocchio, Claudio · 127 Tweddle, Denis · 136 Schaeffer, Brigitte · 25 Scheffer, S. J. · 107 Schmidt, Katrin · 73 U Schneider, Harald · 105 Scioscia, Gaetano · 140, 141 Utsugi, Jinbo · 151 Seidel, C · 24 Seifert, Keith A. · 81 Sequeira, F · 68 Shao, Kwang-Tsao · 80, 104, 130, 146 V SHAO, KWANG-TSAO · 128 Sharina, S.N. · 109 Vaglia, Thierry · 30

Second International Barcode of Life Conference 18-20 September 2007 159 Conference Abstracts Vaidyan, L.K. · 78 VAN DER BANK, MICHELLE · 139 X Van Stappen, Gilbert · 143 VAUGHAN, BOB · 140 Xiao, Jin-Hua · 104 Venugopal, M.N. · 82 Xin, Naihong · 143 Vicario, Saverio · 127 Xu, Minli · 85 Vinnikov, K.A. · 109 VIRGILIO, MASSIMILIANO · 142 VISCHI, MASSIMO · 142 Y Vogel, Johannes · 105 Vogler, Catherine · 89 YANG, CHIEN-HUI · 145 Voice, David · 70 Yang, M.-M. · 119 Yang, Yi-Ju · 146 YAO, HIOU-JU · 146 W YAO, HUI · 147 YASSIN, AMIR · 148 Wang, Norman · 129 Yeh, Hsin-Ming · 128 Wang, Weiwei · 94 Yeh, W.-B. · 119 WANG, WEIWEI · 143 Yeh, Wen-Bin · 138 WARNER, JORGE · 143 YEH, WEN-BIN · 149 Weber, H · 24 Yen, Shen-Horn · 117 Weber, H. · 82 Yi, Dong-Keun · 112 WEIGT, LEE · 52 YOO, HYE SOOK · 150 Weir, T.A. · 21 Yoon, Hyunkye · 23 Whitehead, Michael · 85 Yoshitake, Hiraku · 106 Wiebe, Peter H. · 77 YOSHITAKE, HIRAKU · 151 Willassen, Endre · 100 YU, CHING-RAY · 151 Wilson, John J · 96 Yu, Jun · 94, 143 Wilson, Michael D. · 76 Wong, Ka-Lok · 129 Wood, Monty · 33, 74 Z Wörheide, Gert · 89, 90, 91 Wratten, Steve D. · 73 ZARE, RASOUL · 152 WU, SHU-PING · 144 ZEIN, M. SYAMSUL ARIFIN · 152 Wu, Tsz Huen · 138 ZHANG, YA-PING · 8 Wu, Wen-Long · 114 Zhou, Xin · 98 Wu, Yu-Tseng · 116 ZHOU, XIN · 153 ZIMMERMANN, JUERGEN · 154

Second International Barcode of Life Conference 18-20 September 2007 160 Conference Abstracts