MEMORANDUM OF UNDERSTANDING CMS/IOSEA/MOS8/Inf.10.5 ON THE CONSERVATION AND 28 August 2019 MANAGEMENT OF MARINE TURTLES AND THEIR HABITATS OF THE INDIAN Original: English OCEAN AND SOUTH-EAST ASIA 8TH MEETING OF THE SIGNATORY STATES Da Nang, Viet Nam, 21-25 October 2019 Agenda Item 10.5 CRACKING THE CODE: DEVELOPING A TORTOISESHELL DNA EXTRACTION AND SOURCE DETECTION METHOD (Prepared by WWF) ATMOSPH ND ER A IC IC A N D A M E I THIS REPORT C N WORKING I O S L T HAS BEEN A R TOGETHER TO N A T O I I O PRODUCED IN T PROTECT THE N A N COLLABORATION HAWKSBILL U . E S C . R WITH D E TURTLE E M PA M RT O MENT OF C REPORT 2019 CRACKING THE CODE DEVELOPING A TORTOISESHELL DNA EXTRACTION AND SOURCE DETECTION METHOD ACKNOWLEDGEMENTS This report was written by Michael Jensen (Research Contractor to WWF), in collaboration with Erin LaCasella (NOAA) and Peter Dutton (NOAA), with contributions and review by Christine Madden Hof (WWF-Australia). We appreciate the support of Christine Madden Hof, Dr Ian Bell, turtle conservation rangers of the Arnavon Community Marine Park, Dr Richard Hamilton, Dr Jeff Kinch and Rachel Groom with the collection of samples for this study. Southwest Fisheries Science Center/NOAA The SWFSC is the research arm of NOAA’s National Marine Fisheries Service in the Southwest Region. Center scientists conduct marine biological, economic and oceanographic research, observations and monitoring of living marine resources and their environment. As the NOAA-Fisheries’ National Sea Turtle Genetics Lab, the SWFSC Marine Turtle Genetics Program (MTGP) has the lead responsibility for generating, analysing and interpreting genetic data to address the scientific and management needs for the agency. A priority for NOAA- SWFSC is to implement Recovery Plans for sea turtles under the U.S. Endangered Species Act (ESA) which includes engaging with international partners (such as WWF), to further the science of conservation of sea turtles throughout their range and life cycle. WWF WWF is one of the world’s largest and most experienced independent conservation organizations, with over 5 million supporters and a global Network active in more than 100 countries. WWF’s mission is to stop the degradation of the planet’s natural environment and to build a future in which humans live in harmony with nature, by: conserving the world’s biological diversity, ensuring that the use of renewable natural resources is sustainable, and promoting the reduction of pollution and wasteful consumption. WWF-Australia in coordinating Southeast Asia and Pacific (Asia-Pacific) efforts to address unsustainable use and illegal trade in marine turtles and turtle products. With a focus on hawksbill turtles, the program of work is multifaceted and draws on collaborations between a number of organisations. One of the initiative’s objectives is to develop and apply new innovative approaches to effectively track marine turtles and turtle products along the trade chain from source to sale. To catalyse enforcement and aid conservation, traceability of hawksbill turtle stocks targeted in trade will rely on WWF’s collaborative trans-Pacific forensic study characterising hawksbill turtle genetic origins and relationships between nesting and foraging populations. Royal Caribbean Cruises: This project would not have been possible without the generous support of Royal Caribbean Cruises. Since 2018, Royal Caribbean Cruises and WWF-Australia have been working together to better protect the hawksbill turtle from the illegal turtle trade. Suggested citation: Jensen MP, LaCasella EL, Dutton PH & Madden Hof CA. 2019. CRACKING THE CODE: Developing a tortoiseshell DNA extraction and source detection method. Report to WWF-Australia. Cover image: Close-up of a hawksbill turtle being caught for monitoring, in Papua New Guinea. Hawksbill turtles are under threat and affected by illegal wildlife trade. WWF-Australia is working with partners to monitor populations and ensure the protection of hawksbill turtles that use the area as a route between Australia and Papua New Guinea. © Christine Madden Hof. A WWF-Australia production. Published in August 2019 by WWF-Australia (Sydney, Australia). Any reproduction in full or in part must mention the title and credit the above-mentioned publisher as the copyright owner. All rights reserved. CONTENTS BACKGROUND Conservation status of hawksbill turtles 4 The tortoiseshell trade 4 The role of DNA 5 Aims of this study 6 HOW DOES IT WORK? A brief introduction to hawksbill turtle population genetics 7 DNA EXTRACTION METHOD Sample collection 8 Sequence data 9 Nesting data 9 Shell and foraging samples 9 PRELIMINARY RESULTS Expansion of the nesting baseline 11 Validation of DNA quality from tortoiseshell 11 Market product samples 12 DISCUSSION Future challenges 14 Conclusion 16 REFERENCES Conservation Status of Hawksbill Turtles BACKGROUND The Asia-Pacific region is home to six of the seven species of marine turtles including green, hawksbill, loggerhead, flatback, olive ridley, and leatherback. All species have been assessed at risk of extinction in the IUCN’s Red List and are all listed as Vulnerable to Critically Endangered, except for Australia’s data deficient endemic flatback turtle (IUCN Red List 2015). The Tortoiseshell Trade Marine turtles have survived for 120 million years, yet marine turtles in the Asia- Pacific region are facing a range of threats, including harvesting for food, bycatch in commercial fisheries, loss of nesting beaches and habitats, and climate change. Also, the hawksbill turtle is highly prized for its beautiful carapace and for centuries this species has been exploited for its tortoiseshell (bekko) to make jewellery such as earrings, necklaces, bracelets etc. It has been recently estimated that the tortoiseshell trade network, concentrated in Southeast Asia, harvested approximately 9 million hawksbill turtles over 150 years, over six times previous estimates (Miller et al. 2019). The abundance of hawksbill turtles is estimated to be at least 80% lower than historical levels, with many populations still declining (Mortimer and Donnelly 2008). As such, it is listed as Critically Endangered (IUCN Red List, 2015). THE TORTOISESHELL Hawksbills are still subject to a significant illegal trade in many countries of TRADE NETWORK Southeast Asia and Polynesia (Miller et al. 2019), which is now a major threat HAS HARVESTED thought to be driving populations to near extinction. APPROXIMATELY Although interventions by the Convention on International Trade in Endangered 9 MILLION Species (CITES) resulted in a downward trend in the bekko industry, the HAWKSBILL TURTLES tortoiseshell trade is alive and intact (CITES Secretariat, 2019), and is considered OVER 150 YEARS to be having long-lasting detrimental effects on hawksbill turtle populations in the Asia-Pacific region (IOSEA, 2014). Between 1950-1992, almost 500,000 hawksbills were exported from the region for the bekko industry (Mortimer and Donnelly 2008). Between 2000 and 2008, a trade volume of over 9,180 marine turtle ©WWF-TNC JOINT MARINE PROGRAM, BERAU / TURTLE FOUNDATION 1 including the South east Asia, The Coral Triangle, Western Pacific and Australia 4 WWF-Australia · Cracking the Code Report 2019 products, 2,062 whole marine turtle, 6,161 crafted products and over 1 tonne of raw shell (789 scutes and 919 kg) were seized in East Asia (IOSEA, 2014). More recently between January 2015 – August 2018, seizures from Indonesia, Malaysia and Vietnam recorded trade of 84,116 eggs, 1,836 live turtles, 300 dead, 174 stuffed/ taxidermied, 687kg of meat, 936 crafted products and around 1 tonne of raw shell (1,023 scutes and 739kg shell) (CITES Secretariat, 2019). While we know the key nations poaching hawksbill turtles for shell, we also know poachers are encroaching the national waters and countries of the Coral Triangle and Western Pacific (Lam et al., 2011). The ©ROGER LEGUEN / WWF illegal trade is thought to range from small scale harvests where turtles are harvested to support local markets to large scale operations where turtles are poached and then transported (overland, shipped or airlifted) internationally (Miller et al., 2019; Madden Hof, 2018). But because turtles may be poached both on nesting beaches and at feeding grounds it can be difficult to estimate where turtles are being sourced, how many, which nesting population[s] (genetic stock[s]) are being impacted the most. This knowledge is critical for designing targeted interventions for conserving and recovering hawksbill turtle populations. The Role of DNA Because the marine turtle supply chain has changed significantly and tortoiseshell trade has seemingly shifted from open market to a more covert form (CITES Secretariat, 2019; Madden Hof, 2018) we need to use new technologies to track the source population of turtles poached. In addition to physical tagging and satellite transmitters, molecular (DNA) sampling is being used as a more time-efficient and cost-effective tracking tool to determine the geographical origin of individuals. Novel genetic approaches have been a useful tool in wildlife forensic investigations. These techniques have been successfully applied to illegal poaching of elephants, tigers, rhinoceros, birds etc. (Nishant et al. 2018). For example, DNA extracted from ivory has been used to identify the location of the poaching hotspots for African elephants, by statistically matching the genetic signature of confiscated ivory to geographic or population- specific genetic signatures (Ehman et al. 2015).