The Neuraminidase Stalk Deletion Serves as Major Virulence Determinant of H5N1 Highly Pathogenic Avian Influenza Viruses in Chicken

Abstract

Highly pathogenic avian influenza viruses (HPAIV) cause devastating losses in gallinaceous poultry world-wide and raised concerns of a novel pandemic. HPAIV develop from low-pathogenic precursors by acquisition of a polybasic HA cleavage site (HACS), the prime virulence determinant. Beside that HACS, other adaptive changes accumulate in those precursors prior to transformation into an HPAIV. Here, we aimed to unravel such virulence determinants in addition to the HA gene. Stepwise reduction of HPAIV genes revealed that the HPAIV HA and NA form a minimum set of virulence determinants, sufficient for a lethal phenotype in chicken. Abolishing the NA stalk deletion considerably reduced lethality and prevented transmission. Conversely, the analogous stalk deletion reconstructed in the NA of an LPAIV reassortant carrying only the HPAIV HA resulted in 100% lethality both after primary and contact infection. Remarkably, the unmodified LPAIV NA with its long stalk, when exclusively introduced into the H5N1 HPAIV, still enabled high virulence and efficient transmission. Therefore, irrespective of an NA stalk deletion, minor virulence determinants in addition to the essential polybasic HACS contribute to high virulence, whereas the NA stalk deletion alone may serve as major virulence determinant.

Insertion of Basic Amino Acids in the Hemagglutinin Cleavage Site of H4N2 Avian Influenza Virus (AIV) Reduced Virus Fitness in Chickens which is Restored by Reassortment with Highly Pathogenic H5N1 AIV

“…Importantly, high virulence of the current H4N2 virus was only conferred after reassortment with gene segments from HPAIV H5N1. These findings emphasize the role of other gene segments, in addition to the polybasic CS, in the evolution of HPAIV [10,11,49]. H9N2 with polybasic CS and gene segments from HPAIV H5N1 exhibited a low level HP phenotype (IVPI= 1.23) [26].…”

“…A/quail/California/D113023808/2012(H4N2) was kindly provided by Beate Crossley, the California Animal Health and Food Safety Laboratory System, Department of Medicine and Epidemiology, University of California, Davis. Plasmids containing eight gene segments of HPAIV A/swan/Germany/R65/2006(H5N1) were kindly provided by Jürgen Stech, Institute of Molecular Virology and Cell Biology, Friedrich-Loeffler-Institute (FLI), Greifswald-Insel Riems, Germany [10]. Moreover, plasmids containing eight gene segments of HPAIV A/chicken/Germany/AR1385/2015 (H7N7) were previously cloned [29].…”

“…Despite carrying polybasic CS motifs, some H5 and H7 viruses exhibited low virulence in chickens [5,9]. The virulence of several of these viruses was influenced by increasing the numbers of basic aa in the CS or by additional mutations in the HA or other gene segments [10][11][12]. Interestingly, a few natural non-H5/H7 viruses fulfill the criteria of HPAIV.…”

Avian influenza

“…Stech et al, 2009). This indicates that virulence is influenced by further viral determinants either in the HA (Abdelwhab and Abdel-Moneim, 2015) or beyond (Stech et al, 2015). However, the mechanisms driving the different mutations in the field and the reasons why HA CS changes are restricted to the H5 and H7 subtypes remain obscure.…”

Proteome Response of Chicken Embryo Fibroblast Cells to Recombinant H5N1 Avian Influenza Viruses with Different Neuraminidase Stalk Lengths

“…Especially, a deletion of five amino acids in the viral NA stalk has been observed in the novel reassortant H7N9 viruses in China3. To date, numerous studies on influenza viruses with different NA stalk-motif have been explored and indicated the significant role of NA stalk lengths in virus growth, virulence, and even cross-species transmission456789. However, in terms of host factor, little is known about the molecular mechanism of different pathogenesis of H5N1 viruses with different stalk lengths.…”