Viruses: Genetic Evolution of Avian Influenza A (H9N2) Viruses in Uganda & Evidence of Mammalian Adaptation

Range Of Endemic H9N2 Viruses
From 2019's A Global Perspective on H9N2 Avian Influenza Virus by
T. (Thomas) P. Peacock ,Joe James ,Joshua E. Sealy and Munir Iqbal

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Despite being an LPAI (low path avian influenza) virus - and therefore not a `reportable' disease -  and usually (but not always) producing relatively mild symptoms in humans, H9N2 is viewed by many as an important and influential player in the avian flu world.

• H9N2 has become widespread - even ubiquitous - among poultry across Europe and the Middle East (see 2019 map above), and has more recently extended its range into Africa.

• Serological studies suggest human infection may be far more common than standard surveillance (see FluTrackers List) would have us believe (see J. Infect & Public Health: High Seroprevalence Of Avian Influenza H9 Among Poultry Professionals In Pakistan)

• H9N2 reassorts easily with other viruses, and its internal genes are often found inside many HPAI viruses (including H5N1, H5N6, H7N9, and most recently zoonotic H3N8) -  (see The Lancet's Poultry carrying H9N2 act as incubators for novel human avian influenza viruses)

• H9N2 viruses continue to accrue evolutionary changes, which include mammalian adaptations, and increased binding to human receptor cells (see PLoS Path: Genetics, Receptor Binding, and Transmissibility Of Avian H9N2).

So, while H9N2 may not be at the very top of our pandemic threats list, it is still regarded as having at least some pandemic potential (see CDC IRAT SCORE), and several candidate vaccines have been developed. 

Control of H9N2 has been difficult, as the virus continues to mutate, and many countries continue to use outdated and ineffectual vaccines (see J. Virus Erad.: Ineffective Control Of LPAI H9N2 By Inactivated Poultry Vaccines - China), some of which may be driving its evolution.

Much of what we know about the evolution and threat from avian H9N2 comes from Chinese research and surveillance (see HK CHP Reports 6 More Human H9N2 Cases On the Mainland (3 - 2021, 3 - 2022), where the virus has been monitored for decades. 

Far less is known about how this highly promiscuous H9 virus is evolving in the Middle East and sub-Saharan Africa.  Today, however, we have a comprehensive review of the evolution of H9N2 viruses isolated from poultry in Uganda, one that shows worrisome signs of mammalian adaptation and drug resistance.

I've only reproduced the Abstract, and a snippet from the Conclusions section of the report, so you'll want to follow the link to read it in its entirety.  I'll have a brief postscript when you return. 

Genetic Evolution of Avian Influenza A (H9N2) Viruses Isolated from Domestic Poultry in Uganda Reveals Evidence of Mammalian Host Adaptation, Increased Virulence and Reduced Sensitivity to Baloxavir

by Gladys Atim 1,Titus Tugume 1,Qouilazoni A. Ukuli 1,Bernard Erima 1, Andrew Mubiru 1,Hannah Kibuuka 1,Edison Mworozi 1,Pamela McKenzie 2,Jasmine C. M. Turner 2,David Walker 2,Trushar Jeevan 2,Robert G. Webster 2,Jeremy Jones 2,Richard J. Webby 2,Mariette F. Ducatez 3,Fred Wabwire-Mangen 1,4 and Denis K. Byarugaba 1,5,*

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Abstract

A(H9N2) avian influenza A viruses were first detected in Uganda in 2017 and have since established themselves in live bird markets. The aim of this study was to establish the subsequent genetic evolution of H9N2 viruses in Uganda.

Cloacal samples collected from live bird market stalls in Kampala from 2017 to 2019 were screened by RT-PCR for influenza A virus and H9N2 viruses were isolated in embryonated eggs. One hundred and fifty H9N2 isolates were subjected to whole genome sequencing on the Illumina MiSeq platform. The sequence data analysis and comparison with contemporary isolates revealed that the virus was first introduced into Uganda in 2014 from ancestors in the Middle East. 

There has since been an increase in nucleotide substitutions and reassortments among the viruses within and between live bird markets, leading to variations in phylogeny of the different segments, although overall diversity remained low. The isolates had several mutations such as HA-Q226L and NS-I106M that enable mammalian host adaptation, NP-M105V, PB1-D3V, and M1-T215A known for increased virulence/pathogenicity and replication, and PA-E199D, NS-P42S, and M2-S31N that promote drug resistance.

The PA-E199D substitution in particular confers resistance to the endonuclease inhibitor Baloxavir acid, which is one of the new anti-influenza drugs. Higher EC50 was observed in isolates with a double F105L+E199D substitution that may suggest a possible synergistic effect. 

These H9N2 viruses have established an endemic situation in live bird markets in Uganda because of poor biosecurity practices and therefore pose a zoonotic threat. Regular surveillance is necessary to further generate the needed evidence for effective control strategies and to minimize the threats.

(SNIP)

Conclusions 

Our findings indicate that G1‐lineage H9N2 viruses were introduced into Uganda from the Middle East around 2014. The viruses subsequently became established, quickly reaching a prevalence of 49%. Through comparative whole genome analysis, the viruses were found to have evolved extensively through reassortment events within and between the LBMs in segment‐specific manners.

There were also multiple molecular markers identified that have been associated with cross species infection; these include HA Q226L and those that enhance pathogenicity and reduce antiviral drug sensitivity. The documented zoonotic capacity of G1‐lineage H9N2 viruses highlights the need for regular and exhaustive surveillance of all influenza viruses in circulation within the country and subsequent risk‐assessment to provide a complete picture of prevalence, distribution, and risk 

          (Continue . . . )

Three years ago, in EID Journal: Two H9N2 Studies Of Note, we looked at two reports which suggest that H9N2 continues to evolve away from current (pre-pandemic and poultry) vaccines and is potentially on a path towards better adaptation to human hosts.

Three years ago, the CDC added a new lineage (H9N2 Y280 lineage [A/Anhui-Lujiang/13/2018]) to their short list of novel flu viruses with at least some pandemic potential, although it would not be expected to have anywhere near the impact as an H5 or H7 avian flu.

But H9N2's biggest threat likely comes from its ability to easily reassort with other (avian, human, swine, canine, etc.) influenza viruses, which is believed can increase their ability to infect mammals, including humans. 

 

While probably not a pandemic powerhouse on its own, H9N2's internal genes have shown up as the backbone of some of the most concerning avian viruses of the past 2 decades (PNAS: Evolution Of H9N2 And It’s Effect On The Genesis Of H7N9).

Making it one very much worth keeping our eye on.