1887

Journal of General Virology header logo

Volume 99, Issue 3

An I436N substitution confers resistance of influenza A(H1N1)pdm09 viruses to multiple neuraminidase inhibitors without affecting viral fitness Free

Abstract

The resistance of influenza viruses to neuraminidase (NA) inhibitors (NAIs; i.e. oseltamivir, zanamivir, peramivir and laninamivir) can be associated with several NA substitutions, with differing effects on viral fitness. To identify novel molecular markers conferring multi-NAI resistance, the NA gene of oseltamivir-resistant (H275Y, N1 numbering) 2009 pandemic influenza [A(H1N1)pdm09] virus was enriched with random mutations. This randomly mutated viral library was propagated in Madin–Darby canine kidney (MDCK) cells under zanamivir pressure and gave rise to additional changes within NA, including an I436N substitution located outside the NA enzyme active site. We generated four recombinant A(H1N1)pdm09 viruses containing either wild-type NA or NA with single (I436N or H275Y) or double (H275Y-I436N) substitutions. The double H275Y-I436N mutation significantly reduced inhibition by oseltamivir and peramivir and reduced inhibition by zanamivir and laninamivir. I436N alone reduced inhibition by all NAIs, suggesting that it is a multi-NAI resistance marker. I436N did not affect viral fitness or in a murine model; however, H275Y and I436N together had a negative impact on viral fitness. Further, I436N alone did not have an appreciable impact on viral replication in the upper respiratory tract or transmissibility in ferrets. However, the rg-H275Y-I436N double mutant transmitted less efficiently than either single mutant via the direct contact and respiratory droplet routes in ferrets. Overall, these results highlight the usefulness of a random mutagenesis approach for identifying potential molecular markers of resistance and the importance of I436N NA substitution in A(H1N1)pdm09 virus as a marker for multi-NAI resistance.

  • Received:
  • Accepted:
  • Published Online:
Keyword(s): I436N , Influenza , multi-NAI resistance , neuraminidase and random mutagenesis
© 2018 The Authors
Loading

Article metrics loading...

/content/journal/jgv/10.1099/jgv.0.001029
2018-03-01
2024-04-18
Loading full text...

Full text loading...

/deliver/fulltext/jgv/99/3/292.html?itemId=/content/journal/jgv/10.1099/jgv.0.001029&mimeType=html&fmt=ahah

References

  1. Hancock K, Veguilla V, Lu X, Zhong W, Butler EN et al. Cross-reactive antibody responses to the 2009 pandemic H1N1 influenza virus. N Engl J Med 2009; 361:1945–1952 [View Article][PubMed]
     [Google Scholar]
  2. Centers for Disease Control and Prevention (CDC) Update: influenza activity - United States, September 28, 2008-January 31, 2009. MMWR Morb Mortal Wkly Rep 2009; 58:115–119[PubMed]
     [Google Scholar]
  3. Hurt AC, Hardie K, Wilson NJ, Deng YM, Osbourn M et al. Characteristics of a widespread community cluster of H275Y oseltamivir-resistant A(H1N1)pdm09 influenza in Australia. J Infect Dis 2012; 206:148–157 [View Article][PubMed]
     [Google Scholar]
  4. Centers for Disease Control and Prevention (CDC) Oseltamivir-resistant 2009 pandemic influenza A (H1N1) virus infection in two summer campers receiving prophylaxis-North Carolina, 2009. MMWR Morb Mortal Wkly Rep 2009; 58:969–972[PubMed]
     [Google Scholar]
  5. Le QM, Wertheim HF, Tran ND, van Doorn HR, Nguyen TH et al. A community cluster of oseltamivir-resistant cases of 2009 H1N1 influenza. N Engl J Med 2010; 362:86–87 [View Article][PubMed]
     [Google Scholar]
  6. Hurt AC, Besselaar TG, Daniels RS, Ermetal B, Fry A et al. Global update on the susceptibility of human influenza viruses to neuraminidase inhibitors, 2014-2015. Antiviral Res 2016; 132:178–185 [View Article][PubMed]
     [Google Scholar]
  7. Meijer A, Rebelo-de-Andrade H, Correia V, Besselaar T, Drager-Dayal R et al. Global update on the susceptibility of human influenza viruses to neuraminidase inhibitors, 2012-2013. Antiviral Res 2014; 110:31–41 [View Article][PubMed]
     [Google Scholar]
  8. Takashita E, Meijer A, Lackenby A, Gubareva L, Rebelo-de-Andrade H et al. Global update on the susceptibility of human influenza viruses to neuraminidase inhibitors, 2013-2014. Antiviral Res 2015; 117:27–38 [View Article][PubMed]
     [Google Scholar]
  9. Ferraris O, Lina B. Mutations of neuraminidase implicated in neuraminidase inhibitors resistance. J Clin Virol 2008; 41:13–19 [View Article][PubMed]
     [Google Scholar]
  10. Samson M, Pizzorno A, Abed Y, Boivin G. Influenza virus resistance to neuraminidase inhibitors. Antiviral Res 2013; 98:174–185 [View Article][PubMed]
     [Google Scholar]
  11. Ives JA, Carr JA, Mendel DB, Tai CY, Lambkin R et al. The H274Y mutation in the influenza A/H1N1 neuraminidase active site following oseltamivir phosphate treatment leave virus severely compromised both in vitro and in vivo. Antiviral Res 2002; 55:307–317 [View Article][PubMed]
     [Google Scholar]
  12. de Jong MD, Tran TT, Truong HK, Vo MH, Smith GJ et al. Oseltamivir resistance during treatment of influenza A (H5N1) infection. N Engl J Med 2005; 353:2667–2672 [View Article][PubMed]
     [Google Scholar]
  13. Okomo-Adhiambo M, Demmler-Harrison GJ, Deyde VM, Sheu TG, Xu X et al. Detection of E119V and E119I mutations in influenza A (H3N2) viruses isolated from an immunocompromised patient: challenges in diagnosis of oseltamivir resistance. Antimicrob Agents Chemother 2010; 54:1834–1841 [View Article][PubMed]
     [Google Scholar]
  14. Piralla A, Gozalo-Margüello M, Fiorina L, Rovida F, Muzzi A et al. Different drug-resistant influenza A(H3N2) variants in two immunocompromised patients treated with oseltamivir during the 2011-2012 influenza season in Italy. J Clin Virol 2013; 58:132–137 [View Article][PubMed]
     [Google Scholar]
  15. Babady NE, Laplante JM, Tang YW, St George K. Detection of a transient R292K mutation in influenza A/H3N2 viruses shed for several weeks by an immunocompromised patient. J Clin Microbiol 2015; 53:1415–1418 [View Article][PubMed]
     [Google Scholar]
  16. Nguyen HT, Trujillo AA, Sheu TG, Levine M, Mishin VP et al. Analysis of influenza viruses from patients clinically suspected of infection with an oseltamivir resistant virus during the 2009 pandemic in the United States. Antiviral Res 2012; 93:381–386 [View Article][PubMed]
     [Google Scholar]
  17. Little K, Leang SK, Butler J, Baas C, Harrower B et al. Zanamivir-resistant influenza viruses with Q136K or Q136R neuraminidase residue mutations can arise during MDCK cell culture creating challenges for antiviral susceptibility monitoring. Euro Surveill 2015; 20: [View Article][PubMed]
     [Google Scholar]
  18. Tu V, Abed Y, Barbeau X, Carbonneau J, Fage C et al. The I427T neuraminidase (NA) substitution, located outside the NA active site of an influenza A(H1N1)pdm09 variant with reduced susceptibility to NA inhibitors, alters NA properties and impairs viral fitness. Antiviral Res 2017; 137:6–13 [View Article][PubMed]
     [Google Scholar]
  19. Song MS, Marathe BM, Kumar G, Wong SS, Rubrum A et al. Unique determinants of neuraminidase inhibitor resistance among N3, N7, and N9 avian influenza viruses. J Virol 2015; 89:10891–10900 [View Article][PubMed]
     [Google Scholar]
  20. Kim CU, Lew W, Williams MA, Liu H, Zhang L et al. Influenza neuraminidase inhibitors possessing a novel hydrophobic interaction in the enzyme active site: design, synthesis, and structural analysis of carbocyclic sialic acid analogues with potent anti-influenza activity. J Am Chem Soc 1997; 119:681–690 [View Article][PubMed]
     [Google Scholar]
  21. Tamura D, Debiasi RL, Okomo-Adhiambo M, Mishin VP, Campbell AP et al. Emergence of multidrug-resistant influenza A(H1N1)pdm09 virus variants in an immunocompromised child treated with oseltamivir and zanamivir. J Infect Dis 2015; 212:1209–1213 [View Article][PubMed]
     [Google Scholar]
  22. van der Vries E, Stelma FF, Boucher CA. Emergence of a multidrug-resistant pandemic influenza A (H1N1) virus. N Engl J Med 2010; 363:1381–1382 [View Article][PubMed]
     [Google Scholar]
  23. L'Huillier AG, Abed Y, Petty TJ, Cordey S, Thomas Y et al. E119D neuraminidase mutation conferring pan-resistance to neuraminidase inhibitors in an A(H1N1)pdm09 isolate from a stem-cell transplant recipient. J Infect Dis 2015; 212:1726–1734 [View Article][PubMed]
     [Google Scholar]
  24. Baek YH, Song MS, Lee EY, Kim YI, Kim EH et al. Profiling and characterization of influenza virus N1 strains potentially resistant to multiple neuraminidase inhibitors. J Virol 2015; 89:287–299 [View Article][PubMed]
     [Google Scholar]
  25. Hurt AC, Chotpitayasunondh T, Cox NJ, Daniels R, Fry AM et al. Antiviral resistance during the 2009 influenza A H1N1 pandemic: public health, laboratory, and clinical perspectives. Lancet Infect Dis 2012; 12:240–248 [View Article][PubMed]
     [Google Scholar]
  26. Belser JA, Gustin KM, Maines TR, Blau DM, Zaki SR et al. Pathogenesis and transmission of triple-reassortant swine H1N1 influenza viruses isolated before the 2009 H1N1 pandemic. J Virol 2011; 85:1563–1572 [View Article][PubMed]
     [Google Scholar]
  27. Baranovich T, Webster RG, Govorkova EA. Fitness of neuraminidase inhibitor-resistant influenza A viruses. Curr Opin Virol 2011; 1:574–581 [View Article][PubMed]
     [Google Scholar]
  28. Choi WS, Jeong JH, Kwon JJ, Ahn SJ, Lloren KKS et al. Screening for neuraminidase inhibitor resistance markers among avian influenza viruses of the N4, N5, N6, and N8 neuraminidase subtypes. J Virol 2018; 92:e01580-17 [View Article][PubMed]
     [Google Scholar]
  29. Colman PM, Varghese JN, Laver WG. Structure of the catalytic and antigenic sites in influenza virus neuraminidase. Nature 1983; 303:41–44 [View Article][PubMed]
     [Google Scholar]
  30. Imperiali B, O'Connor SE. Effect of N-linked glycosylation on glycopeptide and glycoprotein structure. Curr Opin Chem Biol 1999; 3:643–649 [View Article][PubMed]
     [Google Scholar]
  31. Hariono M, Abdullah N, Damodaran KV, Kamarulzaman EE, Mohamed N et al. Potential new H1N1 neuraminidase inhibitors from ferulic acid and vanillin: molecular modelling, synthesis and in vitro assay. Sci Rep 2016; 6:38692 [View Article][PubMed]
     [Google Scholar]
  32. Russell RJ, Haire LF, Stevens DJ, Collins PJ, Lin YP et al. The structure of H5N1 avian influenza neuraminidase suggests new opportunities for drug design. Nature 2006; 443:45–49 [View Article][PubMed]
     [Google Scholar]
  33. Nguyen HT, Fry AM, Gubareva LV. Neuraminidase inhibitor resistance in influenza viruses and laboratory testing methods. Antivir Ther 2012; 17:159–173 [View Article][PubMed]
     [Google Scholar]
  34. Song MS, Kumar G, Shadrick WR, Zhou W, Jeevan T et al. Identification and characterization of influenza variants resistant to a viral endonuclease inhibitor. Proc Natl Acad Sci USA 2016; 113:3669–3674 [View Article][PubMed]
     [Google Scholar]
  35. Marathe BM, Lévêque V, Klumpp K, Webster RG, Govorkova EA. Determination of neuraminidase kinetic constants using whole influenza virus preparations and correction for spectroscopic interference by a fluorogenic substrate. PLoS One 2013; 8:e71401 [View Article][PubMed]
     [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/jgv.0.001029
Loading
An I436N substitution confers resistance of influenza A(H1N1)pdm09 viruses to multiple neuraminidase inhibitors without affecting viral fitness
J. Gen. Virol. 99, 292 (2018); https://doi.org/10.1099/jgv.0.001029
/content/journal/jgv/10.1099/jgv.0.001029
/content/journal/jgv/10.1099/jgv.0.001029
Loading

Data & Media loading...

Supplements

Supplementary File 1

PDF

Most cited Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error