Synergistic PA and HA mutations confer mouse adaptation of a contemporary A/H3N2 influenza virus

Abstract

The mouse is the most widely used animal model for influenza virus research. However, the susceptibility of mice to seasonal influenza virus depends on the strain of mouse and on the strain of the influenza virus. Seasonal A/H3N2 influenza viruses do not replicate well in mice and therefore they need to be adapted to this animal model. In this study, we generated a mouse-adapted A/H3N2 virus (A/Switzerland/9715293/2013 [MA-H3N2]) by serial passaging in mouse lungs that exhibited greater virulence compared to the wild-type virus (P0-H3N2). Seven mutations were found in the genome of MA-H3N2: PA(K615E), NP(G384R), NA(G320E) and HA(N122D, N144E, N246K, and A304T). Using reverse genetics, two synergistically acting genes were found as determinants of the pathogenicity in mice. First, the HA substitutions were shown to enhanced viral replication in vitro and, second, the PA-K615E substitution increased polymerase activity, although did not alter virus replication in vitro or in mice. Notably, single mutations had only limited effects on virulence in vitro. In conclusion, a co-contribution of HA and PA mutations resulted in a lethal mouse model of seasonal A/H3N2 virus. Such adapted virus is an excellent tool for evaluation of novel drugs or vaccines and for study of influenza pathogenesis.

Key Role of the Influenza A Virus PA Gene Segment in the Emergence of Pandemic Viruses

“…The crystal structure of H17N10 bat IAV polymerase in complex with an RNA pol II mimetic peptide shows that residue 547 (corresponds to residue 552 in human and avian IAVs) is located within the second binding pocket and very close to RNA pol II CTD residues [30]. While it has yet to be experimentally verified, it will be interesting if interactions with the RNA pol II C-terminal domain affect host specificity of the vRdRp [77][78][79].…”

Effects of Different Drug Combinations in Immunodeficient Mice Infected with an Influenza A/H3N2 Virus

“…Madin–Darby canine kidney cells overexpressing the α2,6 sialic acid receptor (ST6-GalI-MDCK cells) were kindly provided by Y. Kawaoka from the University of Wisconsin, Madison, WI [ 30 ]. The mouse-adapted A/Switzerland/9715293/2013 (H3N2) IV was generated in our laboratory [ 29 ]. Virus stocks were prepared, titrated in ST6-GalI-MDCK cells and stored at −80 °C.…”

“…First, we confirmed that the lymphoproliferative response and number of total T, B and neutrophil cells was reduced. Then, we used such an IS mouse model and we evaluated whether the combination of OS and polymerase inhibitors (FA or BXM) would improve therapeutic efficacy and reduce the emergence of resistance mutations compared with single therapy following lethal infection with a contemporary mouse-adapted influenza A/H3N2 virus [ 29 ].…”

Animal Models Utilized for the Development of Influenza Virus Vaccines

“…Some viruses are easily adapted to mice, however other viruses may take several passages before they are fully adapted [ 122 , 123 , 124 , 125 ]. Mouse adaptation can cause mutations in many viral genes, including the polymerase machinery (PB1, PB2 and PA) as well as the glycoproteins HA and NA [ 126 , 127 , 128 ]. However, there has been little evidence of antigenic drift between the parental and mouse adapted viruses.…”

“…Typically, wild type mouse strains such as BALB/c or C57BL6 mice are used for mouse adaptation. However, more susceptible mice, such as the DBA/2 strain or pharmacologically induced immune-suppressed, have been used to adapt viruses that are unable to infect wild type mice [ 126 ].…”