title: Comparative mutational analyses of influenza A viruses.
abstract: The error-prone RNA-dependent RNA polymerase (RdRP) and external selective pressures are the driving forces for RNA viral diversity. When confounded by selective pressures, it is difficult to assess if influenza A viruses (IAV) that have a wide host range possess comparable or distinct spontaneous mutational frequency in their RdRPs. We used in-depth bioinformatics analyses to assess the spontaneous mutational frequencies of two RdRPs derived from human seasonal (A/Wuhan/359/95; Wuhan) and H5N1 (A/Vietnam/1203/04; VN1203) viruses using the mini-genome system with a common firefly luciferase reporter serving as the template. High-fidelity reverse transcriptase was applied to generate high-quality mutational spectra which allowed us to assess and compare the mutational frequencies and mutable motifs along a target sequence of the two RdRPs of two different subtypes. We observed correlated mutational spectra (τ correlation P < 0.0001), comparable mutational frequencies (H3N2:5.8 ± 0.9; H5N1:6.0 ± 0.5), and discovered a highly mutable motif "(A)AAG" for both Wuhan and VN1203 RdRPs. Results were then confirmed with two recombinant A/Puerto Rico/8/34 (PR8) viruses that possess RdRP derived from Wuhan or VN1203 (RG-PR8×Wuhan(PB2, PB1, PA, NP) and RG-PR8×VN1203(PB2, PB1, PA, NP)). Applying novel bioinformatics analysis on influenza mutational spectra, we provide a platform for a comprehensive analysis of the spontaneous mutation spectra for an RNA virus.
authors.affiliation: School of Public Health, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong.
authors.forename: Peter Pak-Hang
authors.initials: PP
authors.lastname: Cheung
authors.affiliation: National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, Maryland 20894-6075, USA.
authors.forename: Igor B
authors.initials: IB
authors.lastname: Rogozin
authors.affiliation: School of Public Health, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong.
authors.forename: Ka-Tim
authors.initials: KT
authors.lastname: Choy
authors.affiliation: School of Public Health, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong.
authors.forename: Hoi Yee
authors.initials: HY
authors.lastname: Ng
authors.affiliation: School of Public Health, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong hyen@hku.hk malik@hkucc.hku.hk.
authors.forename: Joseph Sriyal Malik
authors.initials: JS
authors.lastname: Peiris
authors.affiliation: School of Public Health, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong hyen@hku.hk malik@hkucc.hku.hk.
authors.forename: Hui-Ling
authors.initials: HL
authors.lastname: Yen
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D004252:DNA Mutational Analysis; D005260:Female; D005814:Genes, Viral; D057809:HEK293 Cells; D006801:Humans; D053124:Influenza A Virus, H5N1 Subtype / Q000201:enzymology / Q000235:genetics*; D008168:Lung / Q000821:virology; D008810:Mice, Inbred C57BL; D008957:Models, Genetic; D059645:Mutation Rate; D012324:RNA-Dependent RNA Polymerase / Q000502:physiology; D014764:Viral Proteins / Q000502:physiology
chemical_list: D014764:Viral Proteins; D012324:RNA-Dependent RNA Polymerase
keywords: H5N1; bioinformatics; influenza A virus; mutational spectra; polymerase fidelity