Sunday, July 30, 2017

Study: Virulence Of HPAI H5N8 Enhanced By 2 Amino Acid Substitutions


The big saving grace with avian HPAI H5N8 - which last winter sparked Europe's largest avian epizootic on record, while moving at the same time into the Middle East and Africa  - is that it hasn't shown any signs (yet) of being able to infect humans.
The virus that showed up in Europe last October, however, was more virulent in wild and migratory birds, and spread faster, farther, and infected more avian  species than we'd ever seen before.
Studies over the winter (see EID Journal: HPAI A(H5Nx) Viruses With Altered H5 Receptor-Binding Specificity) suggested recent evolutionary changes in the virus may have increased its host range, and its ability to spread.

HPAI H5N8, like all flu viruses, continues to evolve via reassortment and/or antigenic drift.  We know the virus underwent a major reassortment during the spring or summer of 2016 - probably in China or Russia (see EID Journal: Reassorted HPAI H5N8 Clade - Germany 2016). 
But smaller changes due to antigenic drift can produce big effects as well, particularly if they occur in a non-avian host
Influenza viruses, as negative-sense single-stranded segmented RNA viruses, are notoriously sloppy replicators. Transcription `accidents' that favor replication in the host tend produce more progeny, and if biologically `fit enough', can drown out the earlier `wild type’ virus in the host. This is called host adaptation.
And while it can (and does) happen in the wild, it can be easily simulated in the laboratory as well, via a classic serial passage study (see chart above).
Researchers inoculate a host with a `wild type’ strain of a virus, let it replicate awhile, then take the virus from the first host and inoculate a second, and then repeat the process five, ten, fifteen times or more. Over time, the virus tends to adapt to the new host (assuming there are no species barriers to prevent it).

Last January, in Sci Rpts: H5N8 - Rapid Acquisition of Virulence Markers After Serial Passage In Mice, we looked at just this sort of experiment that showed H5N8 may not remain benign in mammals forever. The authors wrote:
These results suggest that H5N8 viruses can rapidly acquire virulence markers in mammalian hosts; thus, rapid spread as well as repeated viral introduction into the hosts may significantly increase the risk of human infection and elevate pandemic potential.
This week we've another serial passage study on HPAI H5N8 in mice that identifies two specific amino acid changes that enhance its virulence in mice (and possibly other mammals).

Of the two (PB2 E627K and HA A149V), we've run into this PB2 mutation the most often, including earlier this month in Arch. Of VIrology: Rapid Virulence Shift Of An LPAI H5N2 Virus During A Single Passage In Mice.
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 or mammalian hosts.

The E627K substitution in the PB2 protein (swapping out Glutamic acid (E) for Lysine (K)) makes an avian influenza virus better able to replicate at the lower temperatures (roughly 33C) found in the upper respiratory tract of mammals (see Eurosurveillance: Genetic Analysis Of Novel H7N9 Virus). 

The E627K substitution alone isn't enough adapt an avian virus to a mammalian host, and the virus will need help from other amino acid changes to succeed, but it is an important start.

This report (alas, the full study is behind a pay wall) comes from Infection, Genetics and Evolution.

Research paper
Virulence of an H5N8 highly pathogenic avian influenza is enhanced by the amino acid substitutions PB2 E627K and HA A149V
Haibo Wua, Xiuming Penga, Rufeng Lub, Lihua Xuc, Fumin Liua, Linfang Chenga, Xiangyun Lua, Hangping Yaoa, Nanping Wua, rights and content


  • The virulence of mouse-adapted H5N8 avian influenza virus was increased and HA(A149V) and PB2(E627K) mutations were detected.•
  • Substitutions led to enhanced viral virulence, expanded tissue tropism, and increased replication kinetics in cells.•
  • Continued surveillance in poultry for amino acid changes is required.

A novel reassortant H5N8 highly pathogenic avian influenza (HPAI) virus was recently identified in Asia, Europe, and North America. The H5N8 HPAI virus has raised serious concerns regarding the potential risk for human infection.
However, the molecular changes responsible for allowing mammalian infection in H5N8 HPAI viruses are not clear. The objective of this study was to identify amino acid substitutions that are potentially associated with the adaptation of H5N8 HPAI viruses to mammals.
In this study, an avian-origin H5N8 virus was adapted to mice through serial lung-to-lung passage. The virulence of mouse-adapted virus was increased and adaptive mutations, HA (A149V) and PB2 (E627K), were detected after the ninth passage in each series of mice.
Reverse genetics were used to generate reassortants of the wild type and mouse-adapted viruses. Substitutions in the HA (A149V) and PB2 (E627K) proteins led to enhanced viral virulence in mice, the viruses displayed expanded tissue tropism, and increased replication kinetics in mammalian cells.
Continued surveillance in poultry for amino acid changes that might indicate H5N8 HPAI viruses pose a threat to human health is required.
(Continue . . . )

While we've no reports of human infection with HPAI H5N8, we have seen some scattered reports of H5N8 infection in dogs exposed during poultry outbreaks  (see H5N8 Antibodies Detected In South Korean Dogs (Again)), proving that natural mammalian infection is possible, even if the infection is mild or asymptomatic.

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