Bird flu can be transmitted between mammals — and possible humans — needing only four mutations to do so, a new study published this week in the journal Nature suggests. But the mutant virus is not deadly, and the work could show virologists how to combat others like it.
The research, by Yoshihiro Kawaoka, professor of virology at the University of Wisconsin-Madison, is one of two studies that already made headlines when bits of the results were made public. That's because the studies outline how to make a more easily transmissible — and deadly — version of H5N1, or avian flu. The other paper, by Ron Fouchier, of the Erasmus Medical Center in the Netherlands, also described a method of breeding a virulent and transmissible flu in ferrets.
Debate flared as to whether either study should be published at all, because of fears that a terrorist group or hostile government could use it to make biological weapons. This kind of research is classified as "dual use," much like that involving nuclear material. In the end, the National Science Advisory Board on Biosecurity (NSABB), which advises the government on publicly financed research, decided to allow the publication of Kawaoka's work. In the Netherlands, Fouchier was granted an export license, allowing him to submit the work to the journal Science. Fouchier, however, had previously said he would go ahead with publication regardless of a license. [ Inside the H5N1 Debate ]
What's in H5N1?
Full details of the experiments hadn't been made public until now. On seeing the actual paper, Vincent Racaniello, a professor of microbiology and immunology at Columbia University, called it "a significant piece of work."
"After waiting to read it for so long, it was like eating again after fasting," Racaniello wrote in an email to LiveScience.
Kawaoka was able to create a hybrid flu virus that is transmitted via respiratory droplets, meaning it can spread through the fluid a person (or in this case, a ferret) sneezes out or coughs up. [ 10 Deadly Diseases That Hopped Across Species ]
Two key factors made the virus transmissable. One involved a protein called haemagglutinin, or HA, which is what gives H5N1 its name; the number 5 says what type of haemagglutinin it is. The other involved combining the genes from the H5N1 virus with that of another called H1N1, or swine flu, which caused a pandemic in 2009 in Asia.
The HA protein only connects to a molecule called alpha 2,3 linked sialic acid. Birds carry that receptor in their respiratory and digestive tracts. Humans and other mammals also have it, but it is deeper in the lungs and harder for the virus to reach. Flu viruses like H1N1 that infect humans link to a receptor called alpha 2,6 linked sialic acid, which resides in mammals' respiratory systems.
So the first thing was to change the HA protein into something that would bind to the molecules in a mammalian respiratory tract. To do that, Kawaoka and his team introduced random changes in the "head" of the HA molecule, and looked for ones that would bind to alpha 2,6.
Once they found those, they created a hybrid H5N1/H1N1 flu virus, using genes from H1N1. The result was an H5N1 (strictly speaking, it is an H5 HA/ H1N1) with genes that coded for that altered HA protein.
The next step was infecting ferrets, which acted as a stand-in for mammals, including humans. More changes happened in the viral gene as it reproduced in the ferrets. That was expected, as viral RNA often gets altered as it infects a host's cells.
Four little mutations
It turned out the new H5N1 did not transmit well between ferrets, that is, until four key changes had occurred in the HA protein. Three of those changes allowed the virus to live in the ferret's bodies, but it was the fourth — an amino acid called T318I — that kept the HA protein stable so that the virus would bind to the ferret's cells long enough to reproduce and live in the droplets that get spread whenever a ferret sneezes.
While that may sound scary, the virus had other surprises. For instance, the H5N1 hybrid wasn't very deadly ; Kawaoka noted in his paper that none of the ferrets died. The researchers also tested an H5N1 vaccine against the virus and found that it was effective, as was Tamiflu.
Racaniello noted that Kawaoka's discovery shows the importance of the stability of HA in transmitting the virus between animals. The results also revealed which genes are important in making an airborne virus, and that the mutations that made the virus bind to animal cells actually made HA less stable until another amino acid, far from the site where the binding happens, changed also.
With that knowledge, scientists now know what to look for. "This is a mechanism we can work on," Racaniello said. "How universal is it [among flu viruses] that stabilizing the HA is needed for transmission? Do we see stabilizing mutations arising in nature?"
While Racaniello supports the publication of the research, not everyone was so enthusiastic. Michael Osterholm, director of the Center for Infectious Disease Research and Policy, which studies bioterrorism threats, has said in the past that H5N1 is more dangerous than people realize, though he voted for publication of Kawaoka's paper when it came before the NSABB. In Fouchier's case, he voted against. [ Blockbuster Bioterrorism? 11 Infectious Movies ]
In an April 12 letter to Dr. Amy Patterson, the associate director for Science Policy at the National Institutes of Health, Osterholm said that even if H5N1 was not spread in human populations, a terrorist group might try to infect pigs, devastating local livestock industries because people would fear infection from eating them (the viruses aren't transmitted that way in any case).
Also, a virus that escaped to reproduce in pigs might still pose a danger to humans. "I can't think of a worse scenario than having H5N1 virus circulating widely in swine with a critical reassortment likely to occur and human transmission not far off," Osterholm wrote in the letter. Reassortment refers to the natural changes in viral RNA that occur when it is "cooking" in a population.
Kawaoka did not return calls or emails for this story, nor was Osterholm willing to speak.
Fouchier's paper has been submitted; it may appear in the next several weeks. When it does it may reveal more about the inner workings of bird flu.
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