Sunday, July 10, 2016

J. Virol: PA Mutation K356R in Avian H9N2 Increases Mammalian Replication & Pathogenicity.


While it is always possible that a chance reassortment between two influenza viruses could produce a pandemic-ready virus, the limited evidence we have suggests that protopandemic viruses may require years - perhaps even decades - to accrue enough host adaptations before they are ready for prime time.

In 2010's Charting the Host Adaptation of Influenza Viruses, the authors describe the long evolutionary road taken by the 2009 H1N1 pandemic virus:

In the late 1970s, an independent “Eurasian swine” H1N1 lineage resulted from a direct transmission from an avian host to pigs (Pensaert et al. 1981). In the late 1990s, a series of reassortant viruses appeared in pigs in North America that initially combined genetic elements from human H3N2 (PB1, H3, and N2) with classical swine viruses followed by the introduction of genetic elements from avian influenza (PA and PB2) (Zhou et al. 1999). This “triple-reassortant” strain then underwent various reassortments acquiring genetic elements from classical swine (H1) and Eurasian swine (N1 and MP) before undergoing a host shift to humans, resulting in the novel “swine origin” influenza virus (pandemic H1N1 2009).

Among the many host adaptations that pH1N1 acquired prior to jumping to humans, was the PA-K356R mutation.   The authors explained.

The pandemic H1N1 2009 PA and PB2 proteins have high human adaptedness, even relative to the distribution found in the swine triple reassortants. Contributing to this are the PB2 A684S and PA K356R substitutions that have occurred in these two proteins prior to the 2009 pandemic (Tamuri et al. 2009).

PA-K356R - which has also been found in human infections with H7N9 and H10N8 - is considered an important mammalian adaptation, and linked to the successful transition  of an avian virus to human hosts.

Enter the LPAI H9N2 virus, which is ubiquitous in Asian and Middle Eastern poultry, and which has a long history of successfully reassorting with other avian flu viruses (see PNAS: Reassortment Of H1N1 And H9N2 Avian viruses & PNAS: Reassortment Potential Of Avian H9N2).

And indeed we find H9N2's genetic contributions in a variety of novel viruses including H5N1, H7N9, H5N6, and H10N8 and we recently saw A Canine H3N2 Virus With PA Gene From Avian H9N2 - Korea.

To put it charitably, H9N2 gets around.  And over the years we've watched as it has evolved, picking up more and more mammalian adaptations along the way.  

While human infection with H9N2 has only rarely been reported, and is generally mild, the incidence has increased greatly in the past couple of years. Seroprevalence studies in Egypt and China suggest the rate of infection is much higher than previously believed.

Whether as a standalone virus, or an accomplice to some other subtype, H9N2 is gaining street creds as a virus to watch. 

So much so, that in early 2014, The Lancet carried a report entitled Poultry carrying H9N2 act as incubators for novel human avian influenza viruses by Chinese researchers Di Liu a, Weifeng Shi b & George F Gao that recommended the slaughter of all poultry carrying the virus to help prevent the emergence of new novel subtypes.

Which brings us to a new report, this time in the Journal of Virology, that finds H9N2 continues to evolve, and that the PA-K356R mutation has become predominant in the past couple of years.

Prevailing PA mutation K356R in avian influenza H9N2 virus increases mammalian replication and pathogenicity

Guanlong Xua, Xuxiao Zhanga, Weihua Gaoa, Chenxi Wanga, Jinliang Wanga, Honglei Suna, Yipeng Suna, Lu Guob, Rui Zhanga, Kin-Chow Changc, Jinhua Liua and Juan Pua*


Adaptation of viral polymerase complex comprising PB1, PB2 and PA is necessary for efficient influenza A virus replication in new host species. We found that mutation PA-K356R has become predominant since 2014 in avian H9N2 viruses in China as with seasonal human H1N1 viruses.
The same mutation is also found in most human isolates of emergent avian H7N9 and H10N8 viruses whose six internal gene segments are derived from the H9N2 virus.
We further demonstrated the mammalian adaptive functionality of PA-K356R mutation. Avian H9N2 virus with PA-K356R mutation in human A549 cells showed increased nuclear accumulation of PA, and raised viral polymerase activity that resulted in elevated viral transcription and virus output. The same mutant virus in mice also enhanced virus replication and caused lethal infection.
In addition, combined mutations of PA-K356R with PB2-E627K, a well-known mammalian adaptive marker, in H9N2 virus showed further cooperative increase in virus production and severity of infection in vitro and in vivo.
In summary, PA-K356R behaves as a novel mammalian tropism mutation which along with other mutations such as PB2-E627K might render avian H9N2 viruses adapted for human infection.


Mutations of polymerase complex (PB1, PB2 and PA) of influenza A virus are necessary for viral adaptation to new hosts. This study reports on a novel and predominant mammalian adaptive mutation PA-K356R in avian H9N2 viruses and human isolates of emergent H7N9 and H10N8 viruses.
We found that PA-356R in H9N2 virus causes significant increase in virus replication and severity of infection in human cells and mice, and that PA-K356R cooperates with PB2-E627K mutation, a well characterized human adaptive marker, to exacerbate mammalian infection in vitro and in vivo. Therefore, PA-K356R mutation is a significant adaptation in H9N2 viruses and related H7N9 and H10N8 reassortants towards human infectivity. 

The other mutation that gets a lot of mention in this report is PB2-E627K, which we've addressed before, but briefly:

Birds run `hotter’ than mammals, which means avian flu viruses (which typically replicate in the avian gut) must adapt to lower temperatures if they are to succeed in human hosts.
The (E627K) substitution in the (PB2) protein makes an influenza virus better able to replicate at the lower temperatures (roughly 33C) normally found in the upper human respiratory tract (see Eurosurveillance: Genetic Analysis Of Novel H7N9 Virus).

Like the 2009 H1N1 virus, H9N2 has been kicking around for decades, accruing changes over time. Being an LPAI virus - one that causes little direct or immediate harm - it gets far less attention than H5N1, H5N6, or H5N8.

But as the numerous studies cited above illustrate, its genetic fingerprints can be found on many of the recently emerged novel flu viruses of the past two decades. 

Making any improvements in its ability to infect, and replicate, in mammals of more than just academic interest.

No comments: