Ian Lipkin, a virus hunter from Columbia University, recently received a blood sample from colleagues at the National Institutes of Health. They came from a man who had received a bone-marrow transplant and had fallen mysteriously ill, with evidence of severely inflamed blood vessels. In analyzing a similar case a few years back, Lipkin had discovered a new polyomavirus, part of a family that can cause disease in people with compromised immune systems. Perhaps this new case would yield another new virus.
It didn't. Instead, when Lipkin's team ran the sample through a system that they had devised to detect human viruses, they found that the man was infected with dengue virus. In hindsight, that made sense—he had recently returned from Vietnam, where dengue is prevalent. But the thing is: The team wasn't looking for dengue virus.
“It wasn't what we anticipated, but we didn't have to make a priori decisions about what we planned to find,” Lipkin says. “When people analyze samples from people who are ill, they have some idea in mind. This is probably an enterovirus, or maybe it's a herpesvirues. They then do a specific assay for that particular agent. They don't usually have the capacity to look broadly.”
The new system, known as VirCapSeq-VERT, barrels past this limitation. Lipkin, together with fellow Columbia professors Thomas Briese and Amit Kapoor, designed it to detect all known human viruses, quickly, efficiently, and sensitively. By searching for thousands, perhaps millions, of viruses at once, it should take a lot of the (educated) guesswork out of viral diagnosis.
In the 120 or so years since viruses were first discovered, our ability to identify them, and diagnose the diseases they cause, has improved enormously. But even the most cutting-edge of techniques have limitations. Sequencing technologies allow scientists to unambiguously decipher the genetic material of viruses in a sample, but they suffer from poor sensitivity—that is, they sometimes miss what they're trying to find. That's because viral genes are often swamped by those of their hosts, so sequencing them is like trying to find a needle in a haystack.
PCR, a method for amplifying DNA, solves that problem by making lots and lots of copies of the needle beforehand. It is exquisitely sensitive but it's also hard to do in bulk, and you need to have some idea of what you're looking for in the first place.
VirCapSeq-VERT combines the best features of these techniques, and throws in a few more for good measure. Think of it as a massive exercise in fishing for viruses. To make the hooks, the team identified and synthesized distinctive stretches of DNA from the genomes of every known group of virus that affects humans and other vertebrates. They ended up with two million of these hooks, each of which was baited to snag a different virus. If you dangle them in a blood sample, yank them out, and then sequence everything that's attached to them, you end up with the full genome of every virus present.
The team tested the system using tissue samples spiked with genes from many infamous viruses, including those responsible for Ebola, dengue, flu, and MERS. They also tried analyzing a nasal swab from a patient and a stool sample from a bat. VirCapSeq-VERT successfully identified all the viruses in these samples, even when they were present at miniscule amounts.
And since the technique offers up the full genomes of whatever virus it detects, it shouldn't throw up any false positives. “If you get a genome, you know what it is. It's unequivocal,” says Lipkin. “It also allows you to find mutations that would circumvent traditional diagnostics, or that might would affect resistance to drugs or vaccines.”
The team can also analyze many patients at once. To do that, they fuse a unique barcode sequence to the viral DNA from each individual sample, before mixing them together and running them through VirCapSeq-VERT. After the system does its thing, the team can check the barcodes to work out which sample each virus came from. “We can do up to 21 at a time, which makes it financially viable,” says Lipkin.
Finally, the system is flexible. It should pull out any virus that's even a modest fit to the baited hooks, which includes mutant strains and, potentially, previously unknown viruses. “We probably won't find anything entirely new, but we'll be able to find anything that's within 60 percent similarity to something represented in our library,” says Lipkin, whose team has discovered over 700 viruses. Already, they have used VirCapSeq-VERT to identify a new virus in tilapia, an economically important fish that is being increasingly farmed in aquaculture.
“The results look very promising,” says Nick Loman from the University of Birmingham, who was not involved in the study. “Going forward, we can combine techniques like this with portable sequencing and have a diagnostic device which provides incredibly rich data for clinicians and epidemiologists. Ultimately what we would like is an entirely unbiased method that captured all pathogens—known and unknown—with exquisite sensitivity.”