Catherine Thompson

The potential impact of neuraminidase inhibitor resistant influenza.

Purpose of review Neuraminidase inhibitor resistant influenza virus has recently emerged, and circulated, in untreated persons. Influenza virus evolution is causing antiviral susceptibility to change. We review the latest research in this rapidly moving field. Recent findings Oseltamivir-resistant influenza H1N1 emerged globally, without drug selection pressure, during the 2007–2008 northern hemisphere influenza season. This unexpected event, coupled with reports of reducing susceptibilities of influenza B and H5N1, contradicts our understanding of the properties of neuraminidase inhibitor resistant influenza viruses. Knowledge of the structure of the neuraminidases and impact of mutations on drug binding has now expanded. Surveillance and clinical studies have identified key areas which require focused research such as the incidence of resistance in children and immunocompromised populations and the need for improved methodologies for detecting resistant virus on an individual and population level. Summary Neuraminidase inhibitors, oseltamivir in particular, are the drugs of choice against seasonal influenza, zoonotic H5N1 and are stockpiled as the primary mitigating strategy for pandemic influenza containment and control. Further clinical and animal studies are essential to fully understand the capacity of neuraminidase inhibitor resistant influenza to be tolerated in the virus population, whilst retaining virulence and transmissibility. Vigilance, policy review and development of new anti-influenza drugs are essential.

Effectiveness of seasonal influenza vaccine for adults and children in preventing laboratory-confirmed influenza in primary care in the United Kingdom: 2015/16 end-of-season results

The United Kingdom (UK) is in the third season of introducing universal paediatric influenza vaccination with a quadrivalent live attenuated influenza vaccine (LAIV). The 2015/16 season in the UK was initially dominated by influenza A(H1N1)pdm09 and then influenza of B/Victoria lineage, not contained in that season’s adult trivalent inactivated influenza vaccine (IIV). Overall adjusted end-of-season vaccine effectiveness (VE) was 52.4% (95% confidence interval (CI): 41.0–61.6) against influenza-confirmed primary care consultation, 54.5% (95% CI: 41.6–64.5) against influenza A(H1N1)pdm09 and 54.2% (95% CI: 33.1–68.6) against influenza B. In 2–17 year-olds, adjusted VE for LAIV was 57.6% (95% CI: 25.1 to 76.0) against any influenza, 81.4% (95% CI: 39.6–94.3) against influenza B and 41.5% (95% CI: −8.5 to 68.5) against influenza A(H1N1)pdm09. These estimates demonstrate moderate to good levels of protection, particularly against influenza B in children, but relatively less against influenza A(H1N1)pdm09. Despite lineage mismatch in the trivalent IIV, adults younger than 65 years were still protected against influenza B. These results provide reassurance for the UK to continue its influenza immunisation programme planned for 2016/17.

Accumulation of Human-Adapting Mutations during Circulation of A(H1N1)pdm09 Influenza Virus in Humans in the United Kingdom

ABSTRACT The influenza pandemic that emerged in 2009 provided an unprecedented opportunity to study adaptation of a virus recently acquired from an animal source during human transmission. In the United Kingdom, the novel virus spread in three temporally distinct waves between 2009 and 2011. Phylogenetic analysis of complete viral genomes showed that mutations accumulated over time. Second- and third-wave viruses replicated more rapidly in human airway epithelial (HAE) cells than did the first-wave virus. In infected mice, weight loss varied between viral isolates from the same wave but showed no distinct pattern with wave and did not correlate with viral load in the mouse lungs or severity of disease in the human donor. However, second- and third-wave viruses induced less alpha interferon in the infected mouse lungs. NS1 protein, an interferon antagonist, had accumulated several mutations in second- and third-wave viruses. Recombinant viruses with the third-wave NS gene induced less interferon in human cells, but this alone did not account for increased virus fitness in HAE cells. Mutations in HA and NA genes in third-wave viruses caused increased binding to α-2,6-sialic acid and enhanced infectivity in human mucus. A recombinant virus with these two segments replicated more efficiently in HAE cells. A mutation in PA (N321K) enhanced polymerase activity of third-wave viruses and also provided a replicative advantage in HAE cells. Therefore, multiple mutations allowed incremental changes in viral fitness, which together may have contributed to the apparent increase in severity of A(H1N1)pdm09 influenza virus during successive waves. IMPORTANCE Although most people infected with the 2009 pandemic influenza virus had mild or unapparent symptoms, some suffered severe and devastating disease. The reasons for this variability were unknown, but the numbers of severe cases increased during successive waves of human infection in the United Kingdom. To determine the causes of this variation, we studied genetic changes in virus isolates from individual hospitalized patients. There were no consistent differences between these viruses and those circulating in the community, but we found multiple evolutionary changes that in combination over time increased the viruss ability to infect human cells. These adaptations may explain the remarkable ability of A(H1N1)pdm09 virus to continue to circulate despite widespread immunity and the apparent increase in severity of influenza over successive waves of infection.

A new laboratory-based surveillance system (Respiratory DataMart System) for influenza and other respiratory viruses in England: results and experience from 2009 to 2012.

During the 2009 influenza A(H1N1) pandemic, a new laboratory-based virological sentinel surveillance system, the Respiratory DataMart System (RDMS), was established in a network of 14 Health Protection Agency (now Public Health England (PHE)) and National Health Service (NHS) laboratories in England. Laboratory results (both positive and negative) were systematically collected from all routinely tested clinical respiratory samples for a range of respiratory viruses including influenza, respiratory syncytial virus (RSV), rhinovirus, parainfluenza virus, adenovirus and human metapneumovirus (hMPV). The RDMS also monitored the occurrence of antiviral resistance of influenza viruses. Data from the RDMS for the 2009–2012 period showed that the 2009 pandemic influenza virus caused three waves of activity with different intensities during the pandemic and post pandemic periods. Peaks in influenza A(H1N1)pdm09 positivity (defined as number of positive samples per total number of samples tested) were seen in summer and autumn in 2009, with slightly higher peak positivity observed in the first post-pandemic season in 2010/2011. The influenza A(H1N1)pdm09 virus strain almost completely disappeared in the second postpandemic season in 2011/2012. The RDMS findings are consistent with other existing community-based virological and clinical surveillance systems. With a large sample size, this new system provides a robust supplementary mechanism, through the collection of routinely available laboratory data at minimum extra cost, to monitor influenza as well as other respiratory virus activity. A near real-time, daily reporting mechanism in the RDMS was established during the London 2012 Olympic and Paralympic Games. Furthermore, this system can be quickly adapted and used to monitor future influenza pandemics and other major outbreaks of respiratory infectious disease, including novel pathogens.

End-of-season influenza vaccine effectiveness in adults and children, United Kingdom, 2016/17

Introduction The United Kingdom is in the fourth season of introducing a universal childhood influenza vaccine programme. The 2016/17 season saw early influenza A(H3N2) virus circulation with care home outbreaks and increased excess mortality particularly in those 65 years or older. Virus characterisation data indicated emergence of genetic clusters within the A(H3N2) 3C.2a group which the 2016/17 vaccine strain belonged to. Methods: The test-negative case–control (TNCC) design was used to estimate vaccine effectiveness (VE) against laboratory confirmed influenza in primary care. Results: Adjusted end-of-season vaccine effectiveness (aVE) estimates were 39.8% (95% confidence interval (CI): 23.1 to 52.8) against all influenza and 40.6% (95% CI: 19.0 to 56.3) in 18–64-year-olds, but no significant aVE in ≥ 65-year-olds. aVE was 65.8% (95% CI: 30.3 to 83.2) for 2–17-year-olds receiving quadrivalent live attenuated influenza vaccine. Discussion: The findings continue to provide support for the ongoing roll-out of the paediatric vaccine programme, with a need for ongoing evaluation. The importance of effective interventions to protect the ≥ 65-year-olds remains.

Detection of influenza A(H3N2) virus in children with suspected mumps during winter 2014/15 in England

Influenza A(H3N2) virus was detected in oral fluid from 16/107 children (aged 2 to 12 years) with a clinical diagnosis of mumps, who were sampled between December 2014 and February 2015 in England, during the peak of the 2014/15 influenza season. Sequence analysis of an A(H3N2) virus from a child with suspected mumps showed the virus was similar to other circulating A(H3N2) viruses detected in winter 2014/15, which were antigenically drifted from the A(H3N2) vaccine strain.

Population susceptibility to North American and Eurasian swine influenza viruses in England, at three time points between 2004 and 2011

Age-stratified sera collected in 2004, 2008 and 2010 in England were evaluated for antibody to swine influenza A(H3N2) and A(H1N1) viruses from the United States or Europe as a measure of population susceptibility to the emergence of novel viruses. Children under 11 years of age had little or no measurable antibody to recent swine H3N2 viruses despite their high levels of antibody to recent H3N2 seasonal human strains. Adolescents and young adults (born 1968–1999) had higher antibody levels to swine H3N2 viruses. Antibody levels to swine H3N2 influenza show little correlation with exposure to recent seasonal H3N2 (A/Perth/16/2009) strains, but with antibody to older H3N2 strains represented by A/Wuhan/359/1995. Children had the highest seropositivity to influenza A(H1N1)pdm09 virus, and young adults had the lowest antibody levels to A/Perth/16/2009. No age group showed substantial antibody levels to A/Aragon/RR3218/2008, a European swine H1N1 virus belonging to the Eurasian lineage. After vaccination with contemporary trivalent vaccine we observed evidence of boosted reactivity to swine H3N2 viruses in children and adults, while only a limited boosting effect on antibody levels to A/Aragon/RR3218/2008 was observed in both groups. Overall, our results suggest that different vaccination strategies may be necessary according to age if swine viruses emerge as a significant pandemic threat.

Comparability of neuraminidase inhibition antibody titers measured by enzyme-linked lectin assay (ELLA) for the analysis of influenza vaccine immunogenicity.

Neuraminidase-inhibition (NI) antibody titers can be used to evaluate the immunogenicity of inactivated influenza vaccines and have provided evidence of serologic cross-reactivity between seasonal and pandemic H1N1 viruses. The traditional thiobarbituric acid assay is impractical for large serologic analyses, and therefore many laboratories use an enzyme-linked lectin assay (ELLA) to determine serum NI antibody titers. The comparability of ELLA NI antibody titers when measured in different laboratories was unknown. Here we report a study conducted through the Consortium for the Standardisation of Influenza SeroEpidemiology (CONSISE) to evaluate the variability of the ELLA. NI antibody titers of a set of 12 samples were measured against both N1 and N2 neuraminidase antigens in 3 independent assays by each of 23 laboratories. For a sample repeated in the same assay, ≥96% of N1 and N2 assays had less than a 4-fold difference in titer. Comparison of the titers measured in assays conducted on 3 different days in the same laboratory showed that a four-fold difference in titer was uncommon. Titers of the same sera measured in different laboratories spanned 3 to 6 two-fold dilutions (i.e., 8-64 fold difference in titer), with an average percent geometric coefficient of variation (%GCV) of 112 and 82% against N1 and N2 antigens, respectively. The difference in titer as indicated by fold range and %GCV was improved by normalizing the NI titers to a standard that was included in each assay. This study identified background signal and the amount of antigen in the assay as critical factors that influence titer, providing important information toward development of a consensus ELLA protocol.

Virological self-sampling to monitor influenza antiviral susceptibility in a community cohort

Abstract Objective To perform antiviral susceptibility monitoring of treated individuals in the community during the 2009 influenza A(H1N1) pandemic in England. Patients and methods Between 200 and 400 patients were enrolled daily through the National Pandemic Flu Service (NPFS) and issued with a self-sampling kit. Initially, only persons aged 16 and over were eligible, but from 12 November (week 45), self-sampling was extended to include school-age children (5 years and older). All samples received were screened for influenza A(H1N1)pdm09 as well as seasonal influenza [A(H1N1), A(H3N2) and influenza B] by a combination of RT–PCR and virus isolation methods. Influenza A(H1N1)pdm09 RT–PCR-positive samples were screened for the oseltamivir resistance-inducing H275Y substitution, and a subset of samples also underwent phenotypic antiviral susceptibility testing by enzyme inhibition assay. Results We were able to detect virus by RT–PCR in self-taken samples and recovered infectious virus enabling further virological characterization. The majority of influenza A(H1N1)pdm09 RT–PCR-positive NPFS samples (n = 1273) were taken after oseltamivir treatment had begun. No reduction in phenotypic susceptibility to neuraminidase inhibitors was detected, but five cases with minority quasi-species of oseltamivir-resistant virus (an H275Y amino acid substitution in neuraminidase) were detected. Conclusions Self-sampling is a useful tool for community surveillance, particularly for the follow-up of drug-treated patients. The virological study of self-taken samples from the NPFS provided a unique opportunity to evaluate the emergence of oseltamivir resistance in treated individuals with mild illness in the community, a target population that may not be captured by traditional sentinel surveillance schemes.

Mouse Models of Influenza Infection with Circulating Strains to Test Seasonal Vaccine Efficacy

Influenza virus infection is a significant cause of morbidity and mortality worldwide. The surface antigens of influenza virus change over time blunting both naturally acquired and vaccine induced adaptive immune protection. Viral antigenic drift is a major contributing factor to both the spread and disease burden of influenza. The aim of this study was to develop better infection models using clinically relevant, influenza strains to test vaccine induced protection. CB6F1 mice were infected with a range of influenza viruses and disease, inflammation, cell influx, and viral load were characterized after infection. Infection with circulating H1N1 and representative influenza B viruses induced a dose-dependent disease response; however, a recent seasonal H3N2 virus did not cause any disease in mice, even at high titers. Viral infection led to recoverable virus, detectable both by plaque assay and RNA quantification after infection, and increased upper airway inflammation on day 7 after infection comprised largely of CD8 T cells. Having established seasonal infection models, mice were immunized with seasonal inactivated vaccine and responses were compared to matched and mismatched challenge strains. While the H1N1 subtype strain recommended for vaccine use has remained constant in the seven seasons between 2010 and 2016, the circulating strain of H1N1 influenza (2009 pandemic subtype) has drifted both genetically and antigenically since 2009. To investigate the effect of this observed drift on vaccine induced protection, mice were immunized with antigens from A/California/7/2009 (H1N1) and challenged with H1N1 subtype viruses recovered from 2009, 2010, or 2015. Vaccination with A/California/7/2009 antigens protected against infection with either the 2009 or 2010 strains, but was less effective against the 2015 strain. This observed reduction in protection suggests that mouse models of influenza virus vaccination and infection can be used as an additional tool to predict vaccine efficacy against drift strains.