In 2009, fish in Israel began dying in droves. And not just any fish, but the St. Peter’s fish, tilapia in the Sea of Galilee—the fish famed in the Bible for feeding the multitudes and paying the temple tax for St. Peter.
As head of the fish disease laboratory for Israel’s Ministry of Agriculture, Avi Eldar was called in to investigate die-offs that were also occurring in Israel’s freshwater fish farms. “It started in the northern part of the Jordan Valley and then a year later, it arrived all over Israel,” he says.
“Generally, fish are not very symptomatic animals,” Eldar explains, but the fish had some skin erosions, swelling, eye problems, and diseased brain tissue. It was clear that something was killing off up to 70% of the fish farmers’ stocks. Also, in 2009, the amount of wild tilapia caught by fisherman on the Sea of Galilee (also known as Kinneret Lake) plummeted. “This fishery is very important for its historical meaning, its religious meaning, and its ecological meaning,” says Eldar, who is based at The Kimron Veterinary Institute in Bet Dagan. [image: Dead fish that succumbed to Tilapia Lake Virus at a fish farm in Israel near the Mediterranean coast. Photo courtesy of Avi Eldar.]
Eldar could not culture any bacteria from the diseased fish. But he could infect fish cells by culturing them with brain tissue from diseased fish, so he suspected a virus. He got in touch with an old friend from graduate school, molecular virologist Eran Bacharach at Tel Aviv University.
The two friends isolated some candidate viral genes from infected cells and “got a glimpse that it was an RNA virus with no homology to other known viruses in the sequence databases,” says Bacharach.
With few leads to build upon, in 2011 the two spoke with a researcher visiting from Columbia University who happened to be a colleague of Ian Lipkin, the renowned virus hunter. Lipkin wanted a crack at characterizing this mystery virus, which only appeared to affect tilapia species. “For us, it was like winning Bingo,” says Eldar.
Around the same time, Hugh Ferguson, a marine pathologist based at St. George’s University in Grenada also sent Lipkin some diseased tilapia samples from nearby fish farms in Ecuador. These tilapia had also been part of massive die-offs but with different symptoms and with the disease lodging in their livers.
Lipkin’s team, led by Nischay Mishra, pursued a negative selection strategy—extracting all of the non-ribosomal RNA from the diseased fish samples, then sequencing everything, and subtracting out sequences that matched up to known fish or other contaminating genes. Whatever was leftover should be the viral culprit.
They found 10, short RNA gene segments, which they report in mBio [add link] this week. One of the segments had what Bacharach calls “a very gentle signature” that matches a subunit of the influenza C RNA polymerase. But the other nine gene segments were a complete enigma.
To be sure they all belonged to the mystery virus, which had been named the Tilapia Lake Virus (TiLV), the team had to resort to some ‘old school’ methods.
“We started using all sorts of classical methods [to characterize this virus],” says Lipkin, who directs Columbia’s Center for Infection and Immunity in New York. That included doing Northern blots to see that each of the 10 segments was present in diseased tissues. This was complemented with mass spectroscopy to identify the 10 viral proteins predicted by the 10 gene segments.
These data and others led the team to conclude that TiLV was a segmented RNA virus closely related to the Orthomyxoviridae family, which includes the influenza viruses. When the team ran parallel experiments in the Ecuadorian fish samples, they found nearly the exact same viral gene sequences.
“It’s almost identical,” says Bacharach, which suggests that, “it is a modern, emerging virus that evolved recently and probably spread by transport.”
But he also has another hypothesis about how the virus might spread globally. 500 million birds fly over Israel twice a year on their migrations between Europe and Africa. Many of those birds stop to rest in Kinneret Lake, he says, and could be viral vectors.
Regardless of how the virus moves around, this finding now gives researchers and fish farmers a way to diagnose TiLV, track the virus’s movement, and better contain it to stop the spread of disease among fish farming operations. It also paves the way to develop a vaccine against TiLV, which could protect this $7.5 billion industry. Such protection would have important trickle-down effects as well, including protecting an important source of protein in the world’s food supply and preserving employment in developing countries in Asia and South America that export some 225,000 tons of tilapia to the US alone each year.
Eldar says the discovery may have come too late for two tilapia species that have practically vanished from Kinneret Lake—the blue tilapia Oreochromis aureus and Tristamella simonis. But luckily, it appears that St. Peter’s fish, Sarotherodon galilaeus, has made a partial recovery. Eldar says, “For our colleagues worldwide, this will be a key preventive tool to stop the transfer of infected stock and stop the disease from spreading.”
-- Kendall Powell