A novel, fast-acting eDNA test can help airport customs officials identify illegally trafficked European eels, which as juveniles cannot be visually distinguished from legally-traded species.

Although international treaties have historically provided a framework for imposing restrictions when nations violate agreements, enforcement remains a challenge in part because many trafficked specimens go unnoticed.

Where enforcement proves difficult, technology such as this fast-acting eDNA test can improve monitoring of illegally traded flora and fauna.

Diego Cardeñosa wasn’t expecting any work calls on a Saturday, and certainly not one from the Hong Kong Customs and Excise Department. The urgency of the matter, however, became quickly apparent.

“Diego, there’s two smugglers here, we need your help. Can you come to the airport…now?”

Cardeñosa, a PhD candidate in marine biology at Stony Brook University in New York, United States, grabbed a set of test tubes from his refrigerator, the suitcase next to the TV, and hopped on the next train to the airport. Once there, the customs officials laid out why they called: two men arriving from Portugal had carried with them bags of baby eels–tens of thousands of them in total. They needed to determine if these were legal shipments of American eels, or illegal transport of the endangered European Eel, Anguilla anguilla. Cardeñosa’s DNA test kit was just what they needed.

“So I arrived to the airport, we tested the eels, and the protocol told us it was European eels,” Cardeñosa told Mongabay. “They were able to detain the smugglers, and we actually made the first prosecution of an international smuggler trying to bring European eels into Hong Kong.”

Later genetic sequencing performed on the shipment of eels confirmed this result, thereby validating this test for eel identification, developed by Cardeñosa and collaborators at the Florida International University, as a rapid and efficient tool for customs official worldwide.

The kit makes use of a process, called polymerase chain reaction, or PCR, which unzips DNA strands to identify genetic code. It can determine the presence of the illegally-traded European eel in a matter of minutes. As outlined in a recent paper in Conservation Science and Practice, the portable test kit can detect A. anguilla DNA from water in a transport bag without actually sampling eel tissue. Analyzing environmental DNA, or eDNA, significantly improves the abilities of custom officials to determine the legality of an eel shipment: eels of different species are impossible to visually distinguish as juveniles, the life stage at which most are transported legally and illegally.

“[Previously], if you got caught, there was no evidence on-site that you are bringing in European eels,” Cardeñosa said. “So there’s nothing you can be prosecuted for. They take the eels, you take the next plane out, and then nobody will be able to put you in jail.”

“But now with this tool…we can extract DNA from the water [shipped with the eels], and it will tell you if there is a European eel swimming or not.”

The novel method of conducting PCR on the fly enables customs officials to quickly distinguish illegally-traded European eels from legal species. Video by Diego Cardeñosa, Stony Brook University.

The long journey to your sushi plate

Most European eels are trafficked for human consumption, primarily as unagi sushi, for Asian markets such as in Japan.

“In April 2018, Europol estimated that 100 tonnes, which equates to about 300 million glass eels, are trafficked annually from Europe to Asia,” Florian Stein, Director of Scientific Operations at the Sustainable Eel Group, told Mongabay. “Almost everyone believed this was unreal and way overestimated…[but] we realized that there is a gap of 100 to 200 tonnes that has to be supplied by unknown sources.”

Such volumes of trafficking can seriously impact the eel’s delicate demographics, as noted by global eel expert Matthew Gollock, researcher at the Zoological Society of London and chair of the Anguillid Eel Specialist Group for the International Union for the Conservation of Nature (IUCN).

“The European eel life-cycle is a truly peculiar phenomenon,” Gollock told Mongabay. “We believe they breed in the Sargasso Sea before juveniles migrate thousands of kilometers to continental waters, from Norway to North Africa. The return journey, years later, is made by maturing ‘silver eels’ that are adapted to the oceanic migration required before spawning can take place.”

This distinguishing characteristic of A. anguilla makes the species both fascinating and vulnerable to hunting pressure. The crisscrossing geography of the species’ life cycle means it cannot be bred in captivity, so all consumed eels were at some point harvested from the wild.

Gollock suggested it was hard to predict a bright future for A. anguilla.

“Unsustainable fisheries and trade, be they legal or illegal, have to be considered in the context of other potential threats to the species, such as barriers to migration, habitat loss, pollution and climate change,” he said. “We know the species is in a critical state, but focusing conservation efforts on one threat would be a very risky strategy.”

Big teeth that bite: enforcing CITES regulations

Wildlife trafficking for the pet trade, human consumption, or medicinal purposes, most of which are scientifically unsubstantiated, has been a major driver of population declines for many threatened species. The Convention on International Trade of Endangered Species of Wild Fauna and Flora (CITES), which regulates legal trade in wildlife, has provided a framework for punishing nations for violating trade agreements through member-imposed sanctions. It has led to a number of historic successes in stemming the flow of illegal trade, such as the international crusade that forced Thailand to reduce its ivory trade in 2015.

“CITES is probably the [global wildlife conservation] convention holder with the biggest teeth…they do have the tools to bite really hard it they want to.” Cardeñosa said. “Whether they bite really hard or not is a different story.”

When sanctions are imposed, nations tend to quickly act to reduce trafficking. Yet motivating sanction campaigns requires confident estimates of illegal wildlife trade: difficult to obtain if member nations fabricate trafficking data, or improperly issue permits, as many countries do. Hong Kong, the gateway to China—the biggest consumer of trafficked animals—is no exception, and it is of particular concern for wildlife trafficking. Even when officials seek to comply, identifying some illegally traded species, such as A. anguilla eels, is no small ask.

Eels are not the only wildlife that slip through the hands of custom officials. Several species of shark are threatened with extinction largely from overharvesting for shark fin soup. Often just the fin is traded, which can prove difficult to attribute to a particular species, so many smugglers can claim their shipment is legal. Stricter shark fin legislation continues to gain momentum, with mostly recently a bill in the works in the United States. But as with the shortcomings of CITES sanctions, policy does not necessarily improve enforcement, and may actually undermine conservation efforts by oversimplifying the problems.

Facilitating identification of traded animal products, on the other hand, is perhaps a more targeted fix. The eel DNA test kit was derived from a protocol initially developed for shark DNA to improve shark fin trade enforcement, and for both sharks and eels analyzing a sample costs about $1. Modifying the genetic amplification and identification method from shark DNA to eel DNA took only a few months. Contingent on available research funding, these methods might be scalable to many species.

“You could potentially develop this protocol for any species, and it should be just as cheap,” Cardeñosa said. “For the next few years, we’re looking at going into more countries. We’re prioritizing which countries trade a lot of sharks…and they need actual help to bring the technologies, the funding, and the expertise.”

When governance and enforcement fail to connect, research can fill the gap

Closing the gap between written policy and active enforcement can prove daunting, but research efforts may make enforcement of wildlife trafficking laws cheaper, easier, and/or more accurate.

Aware of the role they can play in turning policy into action, Cardeñosa and the wildlife forensics team designed the novel rapid eDNA method to be easy to implement by non-scientists, so that customs officials can put it to use themselves.

“You don’t need training in genetics, you don’t need to know what the machine is doing,” Cardeñosa said. “The only thing you need to know is how to mix the reagents, put in the DNA, and hit run on the computer.”

To encourage implementation of the DNA kit, Cardeñosa and his collaborators are in discussions with regulators in Indonesia, Malaysia, and Taiwan to implement their protocols for tracking illegal shark fin and meat trade. And the Superintendencia Nacional de Aduanas y de Administración Tributaria (SUNAT), Peru’s customs agency, may begin enforcement using this rapid DNA test later this year.

Cardeñosa, who recently outlined an international program for eDNA-informed trade monitoring in an academic journal, said he believes that contributing his peer-reviewed research to the table can enhance wildlife trafficking enforcement.

“If you have the data, you better get it out and publish it,” he said, adding that some conservation groups have read his team’s papers to inform their campaigns. “So if I get the facts out, we can all push in the same direction.”

Citation

Cardenosa, D., Gollock, M. J., Chapman, D. D. (2019). Development and application of a novel real-time polymerase chain reaction assay to detect illegal trade of the European eel (Anguilla anguilla). Conservation Science and Practice, 1:e39.

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