The PA and HA Gene-Mediated High Viral Load and Intense Innate Immune Response in the Brain Contribute to the High Pathogenicity of H5N1 Avian Influenza Virus in Mallard Ducks

Abstract

Most highly pathogenic avian influenza A viruses cause only mild clinical signs in ducks, serving as an important natural reservoir of influenza A viruses. However, we isolated two H5N1 viruses that are genetically similar but differ greatly in virulence in ducks. A/Chicken/Jiangsu/k0402/2010 (CK10) is highly pathogenic, whereas A/Goose/Jiangsu/k0403/2010 (GS10) is low pathogenic. To determine the genetic basis for the high virulence of CK10 in ducks, we generated a series of single-gene reassortants between CK10 and GS10 and tested their virulence in ducks. Expression of the CK10 PA or hemagglutinin (HA) gene in the GS10 context resulted in increased virulence and virus replication. Conversely, inclusion of the GS10 PA or HA gene in the CK10 background attenuated the virulence and virus replication. Moreover, the PA gene had a greater contribution. We further determined that residues 101G and 237E in the PA gene contribute to the high virulence of CK10. Mutations at these two positions produced changes in virulence, virus replication, and polymerase activity of CK10 or GS10. Position 237 plays a greater role in determining these phenotypes. Moreover, the K237E mutation in the GS10 PA gene increased PA nuclear accumulation. Mutant GS10 viruses carrying the CK10 HA gene or the PA101G or PA237E mutation induced an enhanced innate immune response. A sustained innate response was detected in the brain rather than in the lung and spleen. Our results suggest that the PA and HA gene-mediated high virus replication and the intense innate immune response in the brain contribute to the high virulence of H5N1 virus in ducks.

Prevailing PA Mutation K356R in Avian Influenza H9N2 Virus Increases Mammalian Replication and Pathogenicity

“…However, Vines et al showed that this single residue has little or no effect on viral replication in ducks (42). Likewise, the PA-I353R mutation in avian H5N1 viruses contributes to viral infection only in mice but not in avian hosts (39,43). Thus, these mutations also seem to be mammal-specific regulators.…”

Endemicity of H9N2 and H5N1 avian influenza viruses in poultry in China poses a serious threat to poultry industry and public health

“…Our team previously isolated a highly pathogenic H5N1 strain that exhibited high virulence in chickens, ducks, mice and guinea-pigs [64] . We have shown that critical amino acids, 101G and 237E in the PA protein, contributed to its high virulence in ducks [120] . Interestingly, the PA gene also accounts for the high virulence of this H5N1 strain in mice and the amino acid 353R acts as the critical molecular marker [51] .…”

“…In its natural host, wild waterfowl, infection with the highly pathogenic avian influenza viruses (HPAIVs) generally, leads to minimal or no disease signs [113][114][115] . However, since 2002, accumulating field observations and experimental studies have determined the increased virulence of the currently circulating H5N1 HPAIVs in both waterfowl and wild birds [116][117][118][119][120][121][122][123][124][125][126][127][128][129][130] . Our team previously isolated a highly pathogenic H5N1 strain that exhibited high virulence in chickens, ducks, mice and guinea-pigs [64] .…”

“…Zhang et al have shown that most of the H9N2 viruses isolated before 2000 fail to be transmitted efficiently to aerosol contact groups of chickens [66] . However, a number of viruses isolated from 2001 onwards not only replicate efficiently in the infected chickens but also transmit [56] Low to high [120,123,136,137] Low to high [56,136] Variability of virulence in migratory waterfowl [138] ; Lethal to quails [139,140] , budgerigars [140] , geese [141] , cats [8,142] , tigers [9,11] and dogs [10] ; Nonlethal to guineapigs [143] , pigs [140,144] and pigeons [121,145] H5N2 Low to high [63] ; High [146,147] Low to high [63] Low to moderate [148] ; Moderate [146] Nonlethal to dogs [149] H5N5 High [62,131] Mild to moderate [131] Moderate [62,131] / H5N6 High [133,134,150] / Low [151] ; Moderate [133] Lethal to cats and swan geese [132] ; Lethal to human [152] H5N8 High [131] ; Low [153] Moderate …”

Crucial role of PA in virus life cycle and host adaptation of influenza A virus

“…Generally, they develop mild or no disease signs upon infection with IAv [6, 104,105]. However, after the long-term evolution, IAv breaks through this virus-host equilibrium and gains the ability to kill their natural host [69][70][71][72][106][107][108][109][110][111][112][113]. Research data from different groups have shown that the PA gene is a key determinant for the lethality of IAv to waterfowl [69][70][71][72].…”