Photograph by Tim Laman, Nat Geo Image Collection
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Blue tangs are popular saltwater aquarium fish that until recently couldn’t be bred in captivity. It’s believed many of them are caught using cyanide to stun them.

Photograph by Tim Laman, Nat Geo Image Collection

‘Holy Grail' Test for Illegal Cyanide-Caught Aquarium Fish May Be Fatally Flawed

A widely celebrated test believed to be able to determine if tropical marine aquarium fish were caught illegally using cyanide may be based on problematic data, a new study says.

Komang swerves to miss a pothole and then to avoid an oncoming bemo, the ubiquitous minibuses providing public transportation throughout Indonesia. He turns off the main road onto a sandy track. Large leaves slap against the truck’s rusted sides, as bags of reef fish slosh in the back. Ahead, through an insect-splattered windshield, chickens and children scuttle in a cacophony of squawks and laughter until the lushness gives way to a cobble beach with a fishing village huddled against dark hills in the distance. Komang pulls up near a small building. A man leans against an overturned dory surrounded by the detritus of his livelihood—nets, a boat beyond repair, a rusted engine block.

He leads Komang to a concrete pool filled with seawater. A fish darts into the open, gills gaping. Komang nods. There’s a brief negotiation, then the man nets and bags the fish and hands it over.

Komang returns to the truck, placing the fish in the back with the others. He starts to slide in behind the wheel but stops, as if he forgot something. Komang removes a plastic bag containing several small tablets from his pocket and hands it to the fisherman.

See What It Takes to Bring Exotic Fish From the Sea to Aquarium Tanks

High demand has exotic fish exporters capturing the vast majority of the fish from their natural habitats. Learn more.

The scene repeats that day at fishing villages along the northwest coast of Bali. Komang is a middleman. He buys fish from fishermen and drives them back over the island to Denpasar, where he sells them to exporters at a profit. For those concerned about illegal, unreported, and unregulated fishing, the middleman system in Indonesia is a roadblock to sustainability. It often removes traceability from the supply chain, as provenance is lost by the time the fish reach the exporter.

There’s also a more insidious concern: Those tablets Komang handed out are potassium cyanide. Combined with seawater in a squirt bottle, they’re used to paralyze fish, making them easier to catch.

It is estimated that during the past half century more than 2.2 million pounds of cyanide were illegally used on Philippine coral reefs to exploit fish for the aquarium and live food trades. The Philippines is the largest supplier of live fish imported into the U.S. for aquariums. In addition to sometimes killing target fish for the pet trade, cyanide kills other fish, coral, and invertebrates such as sea stars, nudibranchs, snails, and crabs in a part of the world where more than 95 percent of reefs are considered “at risk.”

A lack of supply chain transparency, combined with institutionalized illegal activity and weak law enforcement, make it nearly impossible to identify and prosecute those engaged in the trade of cyanide-caught fish.

Anecdotally the situation has improved in the Philippines, where the Bureau of Fisheries and Aquatic Resources has stepped up efforts to curb the practice. But in Indonesia, government involvement has been lackluster. More than 80 percent of marine aquarium fish bound for U.S. aquariums originate in these two countries. An unknown number are taken with cyanide, but, given the damage it causes, scientists say it doesn’t matter whether it’s 90 percent—a figure often repeated in the media, by NGOs, and among anti-trade activist groups—or significantly lower, as the best available data suggest.

We don’t know because no reliable cyanide detection test exists that can easily be used by authorities or resource managers. Law enforcement, environmental NGOs, and aquarium trade importers, wholesales, and retailers have all talked of an effective, non-invasive, and easy-to-implement cyanide test as a sort of Holy Grail.

And in 2012 researchers in Portugal, led by Ricardo Calado of Universidade de Aveiro in Aveiro, seemed to have discovered it—using something called high performance liquid chromatography to detect thiocyanate, a byproduct of the cyanide that is excreted by fish into the water of their holding bag.

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Here in Tubbataha Reefs Natural Park, in the Philippines, the protected reefs remain vibrant. But overfishing and destructive fishing practices, such as cyanide fishing, have led to 95 percent of Southeast Asia’s reefs being labeled “at risk.”

This test was touted as “easier, safer, and cheaper to use than current techniques,” and it didn’t require killing fish as other methods did. It was considered so promising that the Center for Biological Diversity, a nonprofit conservation group, and others petitioned the U.S. Fish and Wildlife Service and other government agencies to crack down on cyanide fishing by using Calado’s method on live fish imports.

“Nobody was more excited than me when the 2012 paper was published,” says Andrew Rhyne, a scientist at Roger Williams University, in Bristol, Rhode Island. As someone who had focused his career on marine aquarium trade issues, he knew how significant an effective test would be. In 2014 Rhyne, along with Roger Williams chemist Nancy Breen, started what they thought would be a simple experiment to replicate the methodology described in the 2012 paper. What they uncovered, however, is now being characterized as a whistleblower report that will likely result in a call to retract the 2012 paper.

Rhyne, Breen, and colleagues found that the amount of cyanide necessary to produce the levels of thiocyanate reported in the 2012 paper would have to be so high that it would kill almost any animal on Earth. Their findings were published on May 30, 2018, in the journal PLoS ONE.

‘We Couldn’t Figure Out What We Were Doing Wrong’

For years Rhyne and Breen had worked to try to replicate the findings at Calado’s lab. “We thought it would be easy,” Breen says. “We just thought it’s an experiment. Bing!—we’ll prove that it works, and that’ll be great.” But it didn’t work. And then it didn’t work again. And again.

The researchers at Roger Williams, like their Portuguese counterparts, placed fish in a bath that contained cyanide for 60 seconds, during which time the fish became paralyzed. The fish were then rinsed and allowed to recover, and the holding water was subsequently tested for thiocyanate. Calado’s lab detected, on average, about 5.7 micrograms per liter of excreted thiocyanate in the water over the course of four weeks. Breen repeatedly detected none, despite it being clear the fish were taking up the cyanide.

“We beat ourselves up for four years. It’s so simple,” Breen says, “but we couldn’t figure out what we were doing wrong.”

On a long drive to Pennsylvania to pick her daughter up from college, Breen fixated on a new question: Was it physiologically possible for a fish to take up enough cyanide in the test period to excrete the levels of thiocyanate reported in the original paper?

It turns out the answer is no. Without upending what’s known about fish physiology and toxicology, it simply isn’t possible for a fish to absorb enough cyanide to lead to the results at Calado’s lab. In their paper Breen and her co-authors walk readers through the math on the 2012 data and conclude that a dose of 53.6 milligrams per kilogram of cyanide would be necessary to yield the reported data. “It seems improbable that these doses could be achieved from the concentrations of cyanide in the bath that the fish were exposed to,” they write. They point out that the dose required is well above the acute lethal dosage of cyanide in most animal species.

The new paper also upends research presented by fish conservationist Rene Umberger, of the Hawaii-based nonprofit For the Fishes, and Craig Downs of Haereticus Environmental Laboratory, a nonprofit research lab based in Virginia, who presented, but have not published, data claiming to have used the 2012 methodology to detect cyanide exposure in aquarium fish imported into the U.S. This “gray literature,” as research that has not been formally published or peer reviewed is called, is cited by Center for Biological Diversity’s petition and more than 50 times in worldwide media.

Miyoko Sakashita, oceans director and senior counsel for the Center for Biological Diversity, says the organization relied on Umberger’s expertise when preparing both the petition and a 2016 cyanide report called Poisoned Waters. “She’s done some of the most important work on this issue of anyone I know,” Sakashita says. When presented with the conclusions of the new paper, Sakashita emphasized the importance of scientific integrity. “I absolutely support science-based decision-making, and I realize that the best available science is a constantly evolving thing.”

Umberger alleged bias and poor lab work yielded the new paper’s conclusions, which she says, will be proven incorrect. Downs did not respond to repeated requests for comment.

The U.S. government rejected the Center for Biological Diversity’s petition largely because the cyanide detection test developed by Calado’s lab had yet to be validated. In order to prosecute, says a spokeswoman for the National Oceanic and Atmospheric Administration, which the Center for Biological Diversity also petitioned, a judge would need to be assured of the reliability of the test, and without scientific validation, that likely wouldn’t be possible. According to Laury Parramore, a spokeswoman for the Fish and Wildlife Service, the agency hasn’t yet prosecuted anyone for importing fish illegally caught with cyanide because of the lack of a viable test.

Reset on Cyanide Detection Research

When he saw a draft of the new paper by Breen and her colleagues, “I wasn’t very happy,” says Calado, who was also a peer reviewer for it. “They tried to replicate our method, and they did not succeed, but the fact is they actually haven’t used our methods, and they haven’t even used the same fish species.” Breen and Rhyne used the common clownfish, whereas Calado used the Clark’s anemonefish, a closely related species.

“This is not black or white,” Calado says. “They tested one species and then generalized to all the fish. That’s pretty much like you test penguins for flying, and you say, ‘Look! Birds can’t fly because we tested penguins hundreds of times, and they can’t fly.’”

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Blue-green chromis swim among the coral in Maumere Bay, Indonesia. They are among the most popular marine fish imported to the U.S. for aquariums, but it remains unclear what proportion are caught using cyanide.

Rhyne says in response, "It’s not about which species. The data presented on that paper are not possible.”

Calado says he will soon be publishing a paper that establishes the common clownfish as an outlier. Breen and Rhyne are also working on a new paper they say will validate their claim that the amount of cyanide necessary to produce thiocyanate at the reported levels far exceeds the lethal dose, even in this clownfish species.

“We have to reset the framework, and I think that’s really what our paper does,” Rhyne says. “It corrects the record, which I think is really important for the process of science, but it also pushes the reset button and says, ‘If you’re working on this topic, this is no longer a valid method. We demonstrated that clearly.”

Nicole Herz was a master’s student with the Leibniz Center for Tropical Marine Ecology, in Bremen, Germany, when she tried and failed to replicate Calado’s method. Sebastian Ferse, also of the Leibniz Center, was the editor of the original 2012 paper, and he was also a co-author of Herz's paper. He, like many, thought it would be relatively easy to replicate the work, but after seeing Herz's work and now reading Rhyne's paper, he has become increasingly skeptical.

In her paper published by SPC Live Reef Fish Information Bulletin in June 2016, Herz writes, “In recent years there has been reduced interest from the media and science [to detect the presence of cyanide in reef fishes] despite a flourishing trade.” In other words, many in the field supposed that the Calado lab’s easy, safe and cheap cyanide test was on the brink of being ready and lost interest in pursuing other cyanide test methodologies.

“We need to acknowledge we certainly don’t have the panacea here that we thought we had with that 2012 paper,” Rhyne says. “We hope in reporting our findings that it will provide the stimulus needed to look for alternate methods to detect cyanide-caught fish that will lead to the development of certification schemes for cyanide-free supply chains.”

Ret Talbot is a freelance writer covering fisheries at the intersection of science and sustainability. Follow him at the Good Catch Blog and on Facebook and Twitter, where he frequently uses the hashtag #DataMatter.

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