Threat: Illegal Wildlife Trade
Threat: Illegal Wildlife Trade
Contributor: James Askew
Academic & Industry Experts: Dirke Steinke (International Barcode of Life), Demian Chapman (Florida International University), Diego Cardeñosa (Stony Brook University), Sarah Foster (Project Seahorse), Mark McAnallen & Heidi Norton (Biomeme), Matthew Markus (Pembient), Rebecca Ng (Paul G. Allen Philanthropies), Heidi Norton (Biomeme), Josh Perfetto (ChaiBio), Luke Warwick (Wildlife Conservation Society), and Nathan Walworth (University of Southern California)
Pictured Above: Tens of million of sharks are captured illegally each year; their fins hacked off by fishermen who, in many cases, throw them back in the ocean to die.
Wildlife trade is one of the biggest drivers of biodiversity loss and leads to the direct death of millions of individual animals across tens of thousands of species worldwide (Challender et al., 2015). Biodiversity loss degrades ecological integrity, from food chain dynamics and ecosystem functions to mutualistic relationships. The harm inflicted by the wildlife trade is multiplied since species with an oversized ecological influence are often targeted: apex predators, keystone species, pollinators, dispersers, browsers, and ecosystem engineers (McKlennan et al., 2016; Ripple et al., 2016). The World Economic Forum’s Risks’ Report identified biodiversity loss and ecosystem collapse as one of the major drivers of global risk that may lead to the spread of infectious diseases, food crises, water crises, and man-made environmental disasters (Gascon, 2015).
Legal wildlife trade, including fisheries and timber, is worth an estimated $300 billion globally (TRAFFIC, 2018). Comparably, illegal wildlife trade is a USD $20 billion industry (UNODC, 2016; Global Financial Integrity, 2017). The primary marine organisms (non-fisheries) illegally traded include large-bodied, high-value species traded for food (i.e. sharks, rays, sturgeon, and whales) or for the entertainment industry (i.e. whales and dolphins), and smaller-bodied species traded for food (i.e. european eel) traditional Chinese medicine (i.e. seahorses) or for ornamental purposes (i.e. coral reef products, aquarium fish, and shells).
The scale of killing for the trade in oceanic species is massive.
Between 63 and 270 million sharks and untold numbers of rays are killed each year, primarily for their fins, with a commercial value of USD $540 million to $1.2 billion.
Despite the International Whaling Commission (IWC) enacting a moratorium on commercial whaling, more than 2,000 whales and 90,000 dolphins are killed annually for meat or for fishing bait (IWC; Fisher & Reeves, 2008), and this number is set to rise with Japan’s recent decision to restart commercial whaling in 2019. The other major threat to cetaceans is marine parks, which contain more than 3,000 individuals, including more than 2,000 dolphins, 200 beluga whales, 60 orcas, and 30 porpoises taken from the wild (Lott & Williamson, 2017).
All 27 species of sturgeon are listed by the International Union for the Conservation of Nature (IUCN), and 16 are Critically Endangered, due to the trade and consumption of caviar (i.e. sturgeon eggs). Without intervention against illegal and/or unsustainable trade, sturgeon will go extinct (IUCN Sturgeon Specialist Group, 2018).
Every year approximately 37 million seahorses (genus Hippocampus) are caught in the world’s non-selective fishing gears, and most find their way into international trade for use as traditional medicine (Lawson et al., 2017), selling in Hong Kong for as much as $1,000-$1,200 USD per kilogram of dried individuals (S. Foster, personal communication).
Corals and reef fish are popular in the United States, European Union, and Japanese aquarium trades, with 30 million fish and 1.5 million coral colonies traded per year. In the United States alone, more than 400,000 pieces of coral are traded annually (Rhyne et al., 2014).
Even with the moratoriums and trade limits imposed by CITES (the Convention on International Trade in Endangered Species of Wild Fauna and Flora), enforcement and monitoring are the greatest challenge for both the legal and illegal trade of all marine species. Legal products are often indistinguishable from illegal counterparts once they have entered the supply chain and current methods are limited due to financial and technical considerations and capacity gaps. However, due to recent advances in the field, this Ocean Genomics Horizon Scan identified a number of potential opportunities to employ genomic tools for monitoring and reducing the illegal and/or unsustainable trade of marine wildlife.
PROGRESS TO DATE
Molecular biology, particularly the advances in polymerase chain reaction (PCR) methods, made its first contribution to the detection of illegally-caught marine species when two scientists set up a small sequencing laboratory in a hotel room to identify “fish” being sold in the Tokyo fish market (Baker and Palumbi 1994; Baker et al., 1996; Palumbi & Cipriano, 1998). Dr. C. Scott Baker and Dr. Steve Palumbi identified 28 species of cetaceans among the marine life for sale, including several protected species such as humpback, western gray, fin, Bryde’s, and small-form Bryde’s whales. This work provides the foundation for many current genomic interventions.
In 2010, Baker and colleagues established that whale sashimi sold in Los Angeles and Seoul was sourced from Japanese “scientific” whaling, by comparing mitochondrial sequences and microsatellites of the whale used for sashimi with validated reference genomes curated by DNA Surveillance, a web-based database for cetaceans (Ross et al., 2003). Following this study, molecular registries have been completed for Norwegian minke whales (Glover et al., 2012) and Japanese whales (data not publicly available). Baker and colleagues (2007) also combined these techniques with classic mark-recapture methods to estimate the number of whales entering the market. While these efforts required technical skills and the transportation of samples to the United States and New Zealand for analyses, new technologies and assays are now available to identify whale and dolphin meat in the field.
Building upon these PCR-based methods, Diego Cardeñosa at Stony Brook University and Dr. Demian Chapman at Florida International University recently developed a rapid-tool for detecting CITES-listed sharks, funded by Paul G. Allen Philanthropies (Cardeñosa et al., 2018). The real-time PCR test for 9 CITES-listed shark species is rapid (approximately four hours), reliable (all 9 species are regularly identified from field samples), and cost-effective. After the initial purchase of the portable Chai Bio Open qPCR unit for $4,300, the per-sample cost of running the test is USD $0.94 in reagents. This tool is being championed at CITES meetings.
One of the most prevalent data gaps in fisheries management, including sharks and rays, is the lack of traceability of products. However, Genetic Stock Identification (GSI) methods could be used to assess the stock composition of a fishery or market that could have multiple sources, which would play an essential role in assessing population-specific exploitation levels. In one example, Chapman et al. (2010), reconstructed the natal source population of origin of 62 scalloped hammerhead shark fins sampled from the Hong Kong shark fin market using GSI methods to demonstrate mitochondrial DNA regions exhibited regionally distinct haplotypes.
Because of caviar’s commercial value, moderate progress has been made on sturgeon genomics. Genidaqs, a Sacramento, California-based company, is sequencing the whole genome of white sturgeon (Scott Blankenship, personal communication). In the European Union, genomic techniques of tracking sturgeon have been prioritized as critical to maintaining sustainable trade practices. The SturSNiP program, led by Dr. Rob Ogden at the Tools and Resources for Applied Conservation and Enforcement (TRACE) Network, represents the first major step in the development of a comprehensive suite of new DNA markers for the forensic identification of caviar products traded within the European Union. SturSNiP aims to provide a standardized identification system for fish parts and derivatives and for supporting sustainable aquaculture practices. Researchers within the SturSNiP project are working to discover SNP markers in several sturgeon species: Russian (A. gueldenstaedtii), Persian (A. persicus), Siberian (A. baerii) and Adriatic (A. naccarii). The SNP discovery method was enriched for markers that are polymorphic among species and candidate SNPs were tested to confirm their ability to authenticate pedigree.
Two researchers from the SturSNiP consortium, Dr. Elisa Boscari at University of Padova and Dr. Milos Havelka at University of Hokkaido developed primers and simple PCR-gel and electrophoresis-based tools that can identify species of sturgeon and hybrids from their eggs. These researchers showed that many (though not all) sturgeon products can be identified to the species level by analysis of the mitochondrial cytochrome b gene (Boscari et al., 2014; Havelka, et al., 2017). Dr. Boscari also investigated genetic bases for sex-determination of sturgeon, which can be found on AnaccariiBase, while Chen and colleagues (2017) at the China Academy of Sciences recently completed exploratory CRISPR work that could enable future genetic manipulation of sturgeon. These studies demonstrated the potential to apply genomic techniques for selective breeding and even genetic engineering of farmed sturgeon, which could increase yields of farmed sturgeon and reduce the pressure from the trade on wild populations. Although promising, much more research is necessary before it can be applied in the field.
Seahorses are notably understudied, with fewer than thirty scientists working on the genus around the world (Sarah Foster, personal communication). Currently there are only two whole genomes published: the tigertail seahorse (Quiang Lin et al., 2017) and lined seahorse (Lin et al., 2016). Some efforts have been made to evaluate genetic structure and breeding studies based on microsatellite markers (Mobley et al., 2011 review). Limited studies have utilized DNA barcoding and PCR methods to identify traded species in California (Sanders et al., 2008) and Taiwan (Hou et al., 2018), but these techniques have not been operationalized for a conservation use case (i.e. enforcement) as with the shark fin tool.
Genomic techniques needed to monitor the coral reef trade are still rudimentary. However, a database of mitochondrial DNA genotypes across its geographic range, including data from dried corals, was used to characterize the origin of red coral, Corallium rubrum. Steinke et al (2009) developed genetic assays to identify ornamental reef fish, and genomes have been sequenced for popular reef species: Blacktail butterflyfish (Batista et al., 2018), orange clownfish (Marcionetti et al., 2018), and pygmy angelfish (Fernandez-Silva et al., 2017).
The scale of the wildlife trade and its impacts on ecosystem integrity compound the importance of monitoring and regulation through the rapid identification of species and products at all stages of the wildlife trade supply chain. Until now, these methods have proven too time-consuming, expensive, and technically complex for implementation in the field. Genomics holds tremendous promise of providing portable, cost-effective, and accurate tools that can support monitoring and interdiction of wildlife trade. These tools would enable enforcement agents to act against shipments, illuminate trends in the illegal wildlife trade, and provide evidence for CITES regulation. With proper evidence and legislation, CITES will have the ability to implement international trade sanctions.
The promising mechanism for creating change is through the development and distribution of tools that can enable the rapid identification of illegal or regulated species by enforcement staff at ports and borders. Other potential opportunities include the use of population genetics to monitor the ecology of commonly traded marine wildlife. Longer-term, through the development of either recombinant-based or genetically-engineered tradeable animals or products, it may be possible to reduce or eliminate the demand for wild populations.
To be effective, genomics monitoring and enforcement methods must be portable and accessible to those who need them. The methods also need to be easily deployed in the field without requirements for a fully equipped lab and technical training. The tools must be affordable enough for regular use and capable of running a reasonable number of samples concurrently. Effective detection must identify multiple illegal samples in a shipment. For example, if one sample contains a single illegal fin, this is often considered insufficient to drive prosecution or confiscation, depending on country.
The current tools for species identification include: 1) a sequencing approach to identify DNA-barcodes for a given species 2) RT-PCR with species-specific primers, assays, and tools, developed from DNA barcodes. The highest priorities for immediate genomics work include developing primers and assays for the most vulnerable ocean species in wildlife trade, including whales, dolphins, seahorses, sturgeon, corals, and reef fish.
Where assay development is advanced, there is an opportunity to jump-start the conservation application of these tools through advance market commitments with the vendors that would supply discounted equipment and consumables, and developing training programs with leading researchers. NGOs in Peru have already purchased real-time PCR units, and Florida International University has developed a training plan that is being adopted by Peru and Hong Kong. Final implementation must make the tool available to the most critical nations for CITES regulation (Diego Cardeñosa, personal communication).
In order to successfully transfer and implement these technologies on a much larger scale, including routine inspections, collaborative testing initiatives must be developed involving the various stakeholders (e.g. governments, non-governmental organizations, industry, academic institutions, funders, etc.) to ensure the necessary investments in capacity and financing to enable these efforts. There is a widespread perception that genomic techniques are cost prohibitive for routine screening of products. Accordingly, engagement between nations successfully using these approaches, such as the United States or Hong Kong, and others that need to use them would be a substantial step forward to seeing broader uptake. With successful uptake, a PCR-based approach provides a model that has the potential to transform the monitoring and interdiction of the wildlife trade.
Currently on the market are three commercial products (Chai Bio Open qPCR, Biomeme, and ConservationXLabs Scanner advertising low-to-moderate-cost, portable units, and rapid RT-PCR analyses of DNA barcodes. A fourth company, Thermofisher Scientific, have a range of more expensive units, one of which is currently utilized in Hong Kong. These products are operational for species for which assays and primers have been developed, or for species whose variation is well characterized, and limited assay development is required. Each product is capable of running different numbers of samples in a run and each company offers different costs and services in terms of assay development, need for sample preparation, portability, web interfaces, and training requirements prior to use.
Longer read sequencing approaches would enable a deeper understanding and insight into genetic variation than the tools listed above. This would lower the risk of species misidentifications and provide insights into the extent of geographical variation in DNA sequences. Such insight could be critical for species with only limited samples available for study. Similar to the rest of the field, promising innovations are occurring in this technology. In particular, the Oxford Nanonpore: MinION, is a portable, field-ready device for real-time DNA and RNA sequencing. Each consumable flow cell can generate 10–30 Gb of DNA sequence data across ultra-long read lengths. While this is tremendous power in a miniaturized unit, only one sample can be run at a time, potentially limiting its utility for inspections. Furthermore, the length of the reads is limited by the volume of DNA extracted from a sample, which is in turn limited by lab facilities and labor, again limiting its utility at point of inspection.
RISKS & CHALLENGES
As we explore the opportunities, it is also important to consider the associated risks and challenges. It is unclear whether the policy and enforcement mechanisms in place, especially in the developing world, are stringent enough to achieve tangible results from detecting illegal or regulated species during the trade. Still, enforcement agents, such as the U.S. Fish and Wildlife Service, have used genomics-based tools to identify traded products at airports (i.e. Fields et al., 2015).
The implementation of these tools carries risk as well. Several of these genomics tools require significant expertise (i.e. MinION) or have relatively high start-up costs (i.e. Biomeme), making them undesirable for governments or NGOs. These issues can be mitigated with comprehensive and repeated training courses and long-term sustainable funding. However, cost reductions are ongoing, and innovations in design of rt-PCR units are improving the user-friendliness, compactness, and reliability of the units.
The most significant challenge is a general lack of data; identifying species relies on having a comprehensive database of DNA barcode sequences and variation from all possible target species. For example, seahorses are relatively understudied and difficult to encounter, meaning there is a lack of samples within the Barcode of Life Database (BOLD) library that can be used for barcoding (Dirk Steinke, personal communication). However, this also provides opportunity for researchers to identify DNA barcodes that can become part of the International Barcode of Life database and to develop assays for identifying species. Further, these efforts can be used to develop methods and datasets for eDNA and population genomics to fill in significant gaps in our knowledge of distributions and abundances within the trade.
A range of funders provide support for leading researchers in genomics and the wildlife trade:
Diego Cardeñosa and Dr. Demian Chapman’s work on the rapid shark-fin tool was funded by Vulcan / Paul Allen Philanthropies. Dr. Chapman was a Pew Foundation Marine Fellow, and Pew Charitable Trusts funded some of the earlier work, which enabled the tool’s development. Paul Allen Philanthropies has a history of funding similar projects tackling the wildlife trade including Dr. Sam Wasser’s work determining the geographic origin of poached African elephant ivory (Wasser et al., 2015), which could provide a model for similar work with marine species.
Dr. C. Scott Baker at Oregon State University is a Pew Marine Fellow and has previously been funded by National Geographic and the U.S. government. Dr. Steve Palumbi at Stanford University is funded by various sources, including Chan Zuckerberg’s Biohub.
The E.U. and the European Commission fund significant sturgeon genomics work through the SturSNiP program, a collaboration led by TRACE Network with the Russian and Iranian Fisheries Research Institutes, and Edinburgh and Padova Universities. Research outputs include the work of Dr. Elisa Boscari at University of Padova and Dr. Milos Havelka (now at University of Hokkaido).
Amanda Vincent and Dr. Sarah Foster at Project Seahorse are the world leaders in studying seahorses and the seahorse trade. Very few researchers study seahorses, and therefore there are significant knowledge gaps, including the use of genomic techniques. They are funded by a variety of sources, including their major donor Guylian Belgian Chocolates.
Dr. Dirk Steinke and Dr. Paul Hebert at International Barcode of Life, iBOL project developed assays for barcoding commonly traded ornamental reef fish. Further, iBOL maintains the library of DNA barcodes (available on the BOLD database). They are largely funded by the Canadian government, but lean on additional international infrastructure and multilateral / bilateral funding. A similar project for African terrestrial species, Barcode of Wildlife, is funded by Google.
The Gordon and Betty Moore Foundation is funding the development of the ConservationXLabs Barcode Scanner, and the USFWS’s Combating Wildlife Trafficking Program provides approximately $2m in grants per year.
The leading companies in this space include BioMeme, Oxford Nanopore, Thermofisher Scientific, and ChaiBio. Each company has proprietary technology that contribute to product selection for particular use cases. This competitive environment should continue to foster innovations in technology and design.
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