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Increasing the Uniformity of Genome Fragment Coverage for High-Throughput Sequencing of Influenza A Virus

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Abstract

The high variability of the influenza A virus poses a significant threat to public health, therefore monitoring viral strains and studying their genetic properties are important tasks. One part of this monitoring includes sequencing of influenza A viruses of any subtype and analysis of their whole genomes, which is especially important in cases of interspecies adaptation and reassortment. High-throughput sequencing techno-logies have significantly extended the capabilities of influenza virus epidemiological surveillance. The preparation stages for next generation sequencing (NGS) of influenza A virus include whole genome amplification using one-step RT-PCR, the results of which vary greatly depending on the sample type and quality, that, in turn, affects the coverage of virus fragments and the sequencing results in general. In this work, we propose to supplement the aforementioned technique of whole genome amplification of influenza A virus with sequential suppression PCRs to obtain an even coverage of viral segments of different lengths, which allows sequencing of samples with lower read coverage without decreasing the sequencing quality.

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REFERENCES

  1. Poon L.L., Song T., Rosenfeld R., Lin X., Rogers M.B., Zhou B., Sebra R., Halpin R.A., Guan Y., Twaddle A., DePasse J.V., Stockwell T.B., Wentworth D.E., Holmes E.C., Greenbaum B., et al. 2016. Quantifying influenza virus diversity and transmission in humans. Nat. Genet. 48, 195–200.

    Article  CAS  Google Scholar 

  2. Desselberger U., Racaniello V.R., Zazra J.J., Palese P. 1980. 3′-Terminal and 5′-terminal sequences of influenza-A, influenza-B and influenza-C virus-RNA segments are highly conserved and show partial inverted complementarity. Gene. 8, 315–328.

    Article  CAS  Google Scholar 

  3. Hoffmann E., Stech J., Guan Y., Webster R.G., Perez D.R. 2001. Universal primer set for the full-length amplification of all influenza A viruses. Arch. Virol. 146, 2275–2289.

    Article  CAS  Google Scholar 

  4. Meinel D.M., Heinzinger S., Eberle U., Ackermann N., Schonberger K., Sing A. 2018. Whole genome sequencing identifies influenza A H3N2 transmission and offers superior resolution to classical typing methods. Infection. 46, 69–76.

    Article  CAS  Google Scholar 

  5. Zhou B., Donnelly M.E., Scholes D.T., St George K., Hatta M., Kawaoka Y., Wentworth D.E. 2009. Single-reaction genomic amplification accelerates sequencing and vaccine production for classical and Swine origin human influenza a viruses. J. Virol. 83, 10309–10313.

    Article  CAS  Google Scholar 

  6. Inoue E., Wang X., Osawa Y., Okazaki K. 2010. Full genomic amplification and subtyping of influenza A virus using a single set of universal primers. Microbiol. Immunol. 54, 129–134.

    Article  CAS  Google Scholar 

  7. McGinnis J., Laplante J., Shudt M., George K.S. 2016. Next generation sequencing for whole genome analysis and surveillance of influenza A viruses. J. Clin. Virol. 79, 44–50.

    Article  CAS  Google Scholar 

  8. Lee H.K., Lee C.K., Tang J.W., Loh T.P., Koay E.S. 2016. Contamination-controlled high-throughput whole genome sequencing for influenza A viruses using the MiSeq sequencer. Sci Rep. 6, 33318.

    Article  CAS  Google Scholar 

  9. Shagin D.A., Lukyanov K.A., Vagner L.L., Matz M.V. 1999. Regulation of average length of complex PCR product. Nucleic Acids Res. 27, e23.

    Article  CAS  Google Scholar 

  10. Bankevich A., Nurk S., Antipov D., Gurevich A.A., Dvorkin M., Kulikov A.S., Lesin V.M., Nikolenko S.I., Pham S., Prjibelski A.D., Pyshkin A.V., Sirotkin A.V., Vyahhi N., Tesler G., Alekseyev M.A., Pevzner P.A. 2012. SPAdes: A new genome assembly algorithm and its applications to single-cell sequencing. J. Comput. Biol. 19, 455–477.

    Article  CAS  Google Scholar 

  11. Hunt M., Gall A., Ong S.H., Brener J., Ferns B., Goulder P., Nastouli E., Keane J.A., Kellam P., Otto T.D. 2015. IVA: Accurate de novo assembly of RNA virus genomes. Bioinformatics. 31, 2374–2376.

    Article  CAS  Google Scholar 

  12. Chevreux B., Pfisterer T., Drescher B., Driesel A.J., Muller W.E., Wetter T., Suhai S. 2004. Using the miraEST assembler for reliable and automated mRNA transcript assembly and SNP detection in sequenced ESTs. Genome Res. 14, 1147–1159.

    Article  CAS  Google Scholar 

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The writing of this work did not require special funding.

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Authors and Affiliations

  1. Central Research Institute of Epidemiology, of the Federal Service for Surveillance on Consumer Rights Protection and Human Wellbeing, 111123, Moscow, Russia

    Y. V. Mikhaylova, A. A. Shelenkov, Y. G. Yanushevich & D. A. Shagin

  2. Pirogov Russian National Research Medical University, 117997, Moscow, Russia

    D. A. Shagin

Authors
  1. Y. V. Mikhaylova
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  2. A. A. Shelenkov
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  3. Y. G. Yanushevich
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  4. D. A. Shagin
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Corresponding author

Correspondence to Y. V. Mikhaylova.

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All procedures performed in this work are in accordance with the ethical standards of the institutional committee on research ethics and the 1964 Declaration of Helsinki and its subsequent amendments or comparable standards of ethics.

Conflict of interests. The authors declare they have no conflict of interest.

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Mikhaylova, Y.V., Shelenkov, A.A., Yanushevich, Y.G. et al. Increasing the Uniformity of Genome Fragment Coverage for High-Throughput Sequencing of Influenza A Virus. Mol Biol 54, 851–856 (2020). https://doi.org/10.1134/S0026893320060084

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  • DOI: https://doi.org/10.1134/S0026893320060084

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