key: cord-0894816-39465ap8 authors: Brehony, Carina; Dunford, Linda; Bennett, Charlene; O’Donnell, Joan; Domegan, Lisa; McNamara, Eleanor; De Gascun, Cillian F. title: Neuraminidase characterisation reveals very low levels of antiviral resistance and the presence of mutations associated with reduced antibody effectiveness in the Irish influenza 2018/2019 season date: 2020-10-01 journal: J Clin Virol DOI: 10.1016/j.jcv.2020.104653 sha: eaa103d0cc9d049abd76ba0870a861d1f04296dd doc_id: 894816 cord_uid: 39465ap8 Neuraminidase inhibitor (NAI) resistance levels globally are currently low. However, as antivirals are increasingly being used, but even in the absence of selective pressure, resistance may increase or emerge. The neuraminidase (NA) genes from influenza viruses from the Irish 2018/2019 season were sequenced: 1/144 (0.7%) A(H1N1)pdm09 sequences harboured a substitution associated with highly-reduced susceptibility to NAIs. The very low NAI resistance we describe supports current Irish NAI use recommendations. However, continued monitoring is essential. NA characterisation also identified substitutions associated with reduced antibody effectiveness, thereby highlighting the potential of NA sequence surveillance as an additional tool for investigating influenza vaccine effectiveness (VE).  Very low antiviral resistance in Irish 2018/2019 influenza season  Supports current antiviral use  Continued molecular neuraminidase surveillance essential for resistance emergence  Surveillance also useful for monitoring vaccine effectiveness  Mutations associated with reduced antibody effectiveness detected AbstractNeuraminidase inhibitor (NAI) resistance levels globally are currently low. However, as antivirals are increasingly being used, but even in the absence of selective pressure, resistance may increase or emerge. The neuraminidase (NA) genes from influenza viruses from the Irish 2018/2019 season were sequenced: 1/144 (0.7%) A(H1N1)pdm09 sequences harboured a substitution associated with highly-reduced susceptibility to NAIs. The very low NAI resistance we describe supports current Irish NAI use recommendations. However, continued monitoring is essential. NA characterisation also identified substitutions associated with reduced antibody effectiveness, thereby highlighting the potential of NA sequence surveillance as an additional tool for investigating influenza vaccine effectiveness (VE). Keywords: Influenza; laboratory surveillance; Ireland; antiviral resistance; molecular epidemiology; vaccine effectiveness In the most recent global analysis of human influenza viruses, the level of NA inhibitor (NAI) resistance was found to be 0.2% (1) . The main amino acid substitutions associated with highly reduced or reduced susceptibility are H275Y and Y155H in influenza A(H1N1)pdm09, and E119V & R292K in influenza A(H3N2) viruses (2) . In spite of current low resistance levels, with antivirals increasingly being recommended for use, there is the prospect of emerging resistance. In Ireland, a national network of sentinel general practitioners, covering 6% of the population, monitors and The primary aim of this work was to investigate levels of NAI resistance in influenza isolates for the 2018/2019 season in Ireland. A secondary aim was to investigate the presence of substitutions that may be associated with reduced antibody effectiveness (4)(5). As a WHO National Influenza Centre, the National Virus Reference Laboratory is designated to carry out virus characterisation and antiviral susceptibility monitoring in Ireland. In this study, influenza A samples from the 2018/2019 season, previously subtyped for the influenza HA gene A(H1N1)pdm09 (n=171) and A(H3N2) (n=55)) were subject to whole genome PCR followed by Sanger sequencing with specific primers for A(H1N1)pdm09 and A(H3N2) NA genes (6) . Resulting sequences were assembled using Gap4 (7) and were aligned, translated, and analysed for the presence of amino acid substitutions associated with antiviral resistance as detailed by the WHO (8) Of the A(H1N1)pdm09 (n=144) samples, one sample (0.7%) harboured an amino acid substitution (Y155H) shown to be associated with highly reduced susceptibility to oseltamivir and zanamivir in Vaccination is the primary public health measure used to prevent, reduce, and interrupt the transmission of influenza infection. However, antivirals are also used for prophylaxis as well as J o u r n a l P r e -p r o o f treatment of influenza, and are of particular importance during influenza A(H3N2) predominant seasons when lower influenza VE in older people may occur (11) . Antivirals reduce the risk of severe complications such as hospitalisation and death, as well as the duration of illness, and are recommended to be used as early as possible in the infection (12) . Current Irish guidelines for the use of antivirals recommend antivirals for the treatment of uncomplicated influenza in at-risk groups i.e. those ≥65 years of age, pregnant women, residential care facilities residents, immunosuppressed individuals, individuals with chronic medical conditions and for treatment of clinically complicated influenza (14) . Targeted use of antivirals for post-exposure prophylaxis is also recommended for those in these at-risk groups. The risk of widespread antiviral use is that resistance will increase. However, resistant strains can still emerge even in the absence of selective pressure from widespread antiviral use. This occurred during the 2007/2008 season when oseltamivir resistant A(H1N1) harbouring the H275Y substitution emerged with an average prevalence in Europe of 20% (13) . Mirroring the bacterial antimicrobial resistance 'One Health' framework that has been adopted to tackle it, a multidisciplinary approach has been put forward as a means to detect, contain and prevent the spread of influenza antiviral resistance (14) . Studies have shown the potential for the amplification of antiviral resistance in influenza in natural bird reservoirs upon their ingestion of incompletely degraded antivirals from the environment (15)(16). Seasonal influenza vaccines provide immunity primarily by inducing antibodies that target the protective epitopes on HA. Thus, VE studies primarily concentrate on HA genetic variation: however many factors influence VE, including genetic and antigenic variability, egg adaptation during manufacturing, host related factors (such as age and underlying medical conditions), previous vaccination, and an individual's first influenza infection and subsequent immune response (17) As a major protein in the influenza membrane surface inducing an independent immune response to that of HA, NA may also play a role in VE. Compared to HA there is a relative lack of characterisation of NA immunogenic epitopes and NA characteristics are not standardised during vaccine production (22) . A recent study demonstrated that a reduction or removal of antibody binding and reduced protection against A(H3N2) strains was due to mutations in the NA gene (4). 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