Whole genome sequencing of type A influenza virus by next-generation sequencing

Kim Heui Man, Lee Namjoo, Kim Mi-Seon, Chung Yoon-Seok, Kang Chun
Division of Viral Diseases, Center for Laboratory Control of Infectious Diseases, KCDC

Background: Since influenza viruses are mutated frequently, the sequencing of their hemagglutinin (HA) antigen helps researchers to compare newly discovered clinical isolates with existing vaccine strains to characterize their receptor affinity and pathogenicity. Similarly, sequencing of the neuraminidase (NA) antigen is important for estimating anti-influenza drug resistance.
Present condition: Traditional genetic sequencing techniques, such as the Sanger method, have has several limitations. First, sequencing of a single gene segment of a virus sample involves several laboratory steps that can cause contamination. Second, several polymerase chain reactions (PCRs) need to be performed successively due to the long length of some genes, such as the polymerase genes. Finally, in case of influenza A, frequent mutations make it necessary to redesign the primer sequences because of mismatches with the template sequence. Therefore, next-generation sequencing is employed to prevent contamination of samples and to allow whole genome sequencing regardless of mutations.
Future Perspective: In this study, we performed whole-genome sequencing of influenza isolates obtained from respiratory specimens of Korean subjects and vaccine strain by multi-segment reverse tranion - PCR and NGS analyses. The continual use of NGS could produce a large amount of information for analysis of genetic characteristics of influenza strains as well as an estimation of the performance of commercial vaccine stocks. It could also be used to construct an effective diagnostic system and assess disease risks to humans by analysis of the mutations of recently evolved influenza A strains.

Keywords: Whole genome sequencing, Influenza A virus, Next generation sequencing, Pathogenicity, Mutation