Two Escape Mechanisms of Influenza A Virus to a Broadly Neutralizing Stalk-Binding Antibody
Fig 8
Reduced viral fitness of the A/Perth/16/2009 resistant viruses.
(A) A multiple sequence alignment of 11,981 HA amino acid sequences from human and zoonotic isolates of the H1–16 subtypes was used to assess the genetic diversity at positions corresponding to H3 HA 387 and 391. The results are shown as pie charts. (B) In vitro fitness. MDCK cells were infected with the WT or 39.29-resistant A/Perth/16/2009 viruses at MOI 0.01. Released viral genome in the supernatant was quantitated daily by qPCR of the viral M1 Matrix gene. Genome copy numbers per 50 μl of supernatant are shown as histograms. The assay was done in triplicate with data presented as Mean +/- SEM. Statistics were calculated between WT and each of the mutant viruses using a multiple t test with the Prism 6.0 software (* P ≤ 0.05, indicating significant difference). (C) In vivo fitness. DBA/2J mice were infected with same dose of WT or 39.29-resistant A/Perth/16/2009 viruses. At 6 hr, 24 hr, 48 hr and 72 hr post-infection, lung homogenates were prepared and viral titers in the homogenates were determined on MDCK cells. Each group at each time point contained 5 mice. Lung titers were presented as Mean +/- SEM. Statistics were calculated between WT and each of the mutant viruses using a multiple t test with the Prism 6.0 software (* P ≤ 0.05, indicating significant difference).