The viral polymerase mediates adaptation of an avian influenza virus to a mammalian host

Abstract

Mammalian influenza viruses are descendants of avian strains that crossed the species barrier and underwent further adaptation. Since 1997 in southeast Asia, H5N1 highly pathogenic avian influenza viruses have been causing severe, even fatal disease in humans. Although no lineages of this subtype have been established until now, such repeated events may initiate a new pandemic. As a model of species transmission, we used the highly pathogenic avian influenza virus SC35 (H7N7), which is low-pathogenic for mice, and its lethal mouse-adapted descendant SC35M. Specific mutations in SC35M polymerase considerably increase its activity in mammalian cells, correlating with high virulence in mice. Some of these mutations are prevalent in chicken and mammalian isolates, especially in the highly pathogenic H5N1 viruses from southeast Asia. These activity-enhancing mutations of the viral polymerase complex demonstrate convergent evolution in nature and, therefore, may be a prerequisite for adaptation to a new host paving the way for new pandemic viruses.

Fig. 1.

Growth in avian and mammalian cells. Growth curves of SC35 (•) and SC35M (▪) in chicken embryo fibroblasts (CEF) (A), monkey kidney cells (Vero) (B), mouse lung adenoma cells (LA-4) (C), and human epithelial lung cells (A549) (D). Cells were inoculated at a multiplicity of infection of 10^-4. The plaque titers were determined at appropriate time points on MDCK cells. Each curve is the average of two independent experiments.

Fig. 2.

Enhanced polymerase activity of SC35M.(A and B) Polymerase activity of ribonucleoprotein complexes reconstituted from PB1, PB2, PA, and NP plasmids of SC35, SC35M, and their combinations (A) and of their single-point mutants (B). (C) Correlation of polymerase activity of recombinant virus with virulence. The mouse LD₅₀ of viruses was plotted (inverted axis) against their polymerase activities. The polymerase activities were determined by luciferase reporter assay in four independent experiments.

Fig. 3.

Influenza strains convergent with SC35M polymerase mutations. Shown are phylogenetic trees from PB2 (A), PA (B), and NP (C) genes of strains convergent with SC35M polymerase mutations (black) and representative strains without polymerase mutations (gray). Consensus trees are from neighborhood-joining analyses from bootstrap samples. Numbers at nodes are bootstrap values.