title: Standardizing the influenza neuraminidase inhibition assay among United States public health laboratories conducting virological surveillance.
abstract: BACKGROUND
  Monitoring influenza virus susceptibility to neuraminidase (NA) inhibitors (NAIs) is vital for detecting drug-resistant variants, and is primarily assessed using NA inhibition (NI) assays, supplemented by NA sequence analysis. However, differences in NI testing methodologies between surveillance laboratories results in variability of 50% inhibitory concentration (IC50) values, which impacts data sharing, reporting and interpretation. In 2011, the Centers for Disease Control and Prevention (CDC), in collaboration with the Association for Public Health Laboratories (APHL) spearheaded efforts to standardize fluorescence-based NI assay testing in the United States (U.S.), with the goal of achieving consistency of IC50 data.
  
  
  METHODS
  For the standardization process, three participating state public health laboratories (PHLs), designated as National Surveillance Reference Centers for Influenza (NSRC-Is), assessed the NAI susceptibility of the 2011-12 CDC reference virus panel using stepwise procedures, with support from the CDC reference laboratory. Next, the NSRC-Is assessed the NAI susceptibility of season 2011-12 U.S. influenza surveillance isolates (n = 940), with a large subset (n = 742) tested in parallel by CDC. Subsequently, U.S. influenza surveillance isolates (n = 9629) circulating during the next three influenza seasons (2012-15), were independently tested by the three NSRC-Is (n = 7331) and CDC (n = 2298).
  
  
  RESULTS
  The NI assay IC50s generated by respective NSRC-Is using viruses and drugs prepared by CDC were similar to those obtained with viruses and drugs prepared in-house, and were uniform between laboratories. IC50s for U.S. surveillance isolates tested during four consecutive influenza seasons (2011-15) were consistent from season to season, within and between laboratories.
  
  
  CONCLUSIONS
  These results show that the NI assay is robust enough to be standardized, marking the first time IC50 data have been normalized across multiple laboratories, and used for U.S. national NAI susceptibility surveillance.
authors.affiliation: Influenza Division, NCIRD, Centers for Disease Control and Prevention (CDC), Atlanta, GA, USA.
authors.forename: M
authors.initials: M
authors.lastname: Okomo-Adhiambo
authors.affiliation: Influenza Division, NCIRD, Centers for Disease Control and Prevention (CDC), Atlanta, GA, USA.
authors.forename: V P
authors.initials: VP
authors.lastname: Mishin
authors.affiliation: Influenza Division, NCIRD, Centers for Disease Control and Prevention (CDC), Atlanta, GA, USA.
authors.forename: K
authors.initials: K
authors.lastname: Sleeman
authors.affiliation: California Department of Public Health (CDPH), Richmond, CA, USA.
authors.forename: E
authors.initials: E
authors.lastname: Saguar
authors.affiliation: California Department of Public Health (CDPH), Richmond, CA, USA.
authors.forename: H
authors.initials: H
authors.lastname: Guevara
authors.affiliation: Wisconsin State Laboratory of Hygiene (WSLH), Madison, WI, USA.
authors.forename: E
authors.initials: E
authors.lastname: Reisdorf
authors.affiliation: Wisconsin State Laboratory of Hygiene (WSLH), Madison, WI, USA.
authors.forename: R H
authors.initials: RH
authors.lastname: Griesser
authors.affiliation: Unified State Laboratories: Public Health (USLPH), Taylorsville, UT, USA.
authors.forename: K J
authors.initials: KJ
authors.lastname: Spackman
authors.affiliation: Unified State Laboratories: Public Health (USLPH), Taylorsville, UT, USA.
authors.forename: M
authors.initials: M
authors.lastname: Mendenhall
authors.affiliation: Maryland Department of Health and Mental Hygiene (MD DHMH) Laboratories Administration, Baltimore, MD, USA.
authors.forename: M P
authors.initials: MP
authors.lastname: Carlos
authors.affiliation: Maryland Department of Health and Mental Hygiene (MD DHMH) Laboratories Administration, Baltimore, MD, USA.
authors.forename: B
authors.initials: B
authors.lastname: Healey
authors.affiliation: Wadsworth Center, New York State Department of Health (NYSDOH), Albany, NY, USA.
authors.forename: K
authors.initials: K
authors.lastname: St George
authors.affiliation: Wadsworth Center, New York State Department of Health (NYSDOH), Albany, NY, USA.
authors.forename: J
authors.initials: J
authors.lastname: Laplante
authors.affiliation: Association of Public Health Laboratories (APHL), Silver Spring, MD, USA.
authors.forename: T
authors.initials: T
authors.lastname: Aden
authors.affiliation: Association of Public Health Laboratories (APHL), Silver Spring, MD, USA.
authors.forename: S
authors.initials: S
authors.lastname: Chester
authors.affiliation: Influenza Division, NCIRD, Centers for Disease Control and Prevention (CDC), Atlanta, GA, USA.
authors.forename: X
authors.initials: X
authors.lastname: Xu
authors.affiliation: Influenza Division, NCIRD, Centers for Disease Control and Prevention (CDC), Atlanta, GA, USA. Electronic address: lgubareva@cdc.gov.
authors.forename: L V
authors.initials: LV
authors.lastname: Gubareva
mesh_terms: D002487:Centers for Disease Control and Prevention, U.S.; D024882:Drug Resistance, Viral; D057075:Enzyme Assays / Q000592:standards*; D062665:Epidemiological Monitoring; D006801:Humans; D007251:Influenza, Human / Q000188:drug therapy* / Q000201:enzymology* / Q000453:epidemiology; D020128:Inhibitory Concentration 50; D009439:Neuraminidase / Q000037:antagonists & inhibitors*; D014481:United States / Q000453:epidemiology
chemical_list: D009439:Neuraminidase
keywords: Assay standardization; Neuraminidase inhibition; Oseltamivir; Peramivir; Zanamivir