Analysis of Anti-Influenza Virus Neuraminidase Antibodies in Children, Adults, and the Elderly by ELISA and Enzyme Inhibition: Evidence for Original Antigenic Sin

Abstract

Antibody responses to influenza virus hemagglutinin provide protection against infection and are well studied. Less is known about the human antibody responses to the second surface glycoprotein, neuraminidase. Here, we assessed human antibody reactivity to a panel of N1, N2, and influenza B virus neuraminidases in different age groups, including children, adults, and the elderly. Using enzyme-linked immunosorbent assays (ELISA), we determined the breadth, magnitude, and isotype distribution of neuraminidase antibody responses to historic, current, and avian strains, as well as to recent isolates to which these individuals have not been exposed. It appears that antibody levels against N1 neuraminidases were lower than those against N2 or B neuraminidases. The anti-neuraminidase antibody levels increased with age and were, in general, highest against strains that circulated during the childhood of the tested individuals, providing evidence for “original antigenic sin.” Titers measured by ELISA correlated well with titers measured by the neuraminidase inhibition assays. However, in the case of the 2009 pandemic H1N1 virus, we found evidence of interference from antibodies binding to the conserved stalk domain of the hemagglutinin. In conclusion, we found that antibodies against the neuraminidase differ in magnitude and breadth between subtypes and age groups in the human population. (This study has been registered at ClinicalTrials.gov under registration no. NCT00336453, NCT00539981, and NCT00395174.)

Meeting report and review: Immunological assays and correlates of protection for next‐generation influenza vaccines

“…In addition, serum neuraminidase inhibition (NI) antibody titers have been identified as independent correlates of protection in field studies by Monto et al and Couch et al Furthermore, this is supported by recent data from an H1N1 human challenge model . Of note, anti‐HA stalk antibodies can interfere with H6NX‐based NI assays which have to be taken into account in their interpretation . Anti‐NA enzyme‐linked immunosorbent assays (ELISAs), which do not suffer from this shortcoming, have recently been used to show a negative correlation between anti‐NA titers and virus shedding in humans in a cohort study .…”

“…21 Of note, anti-HA stalk antibodies can interfere with H6NX-based NI assays which have to be taken into account in their interpretation. 22,23 Anti-NA enzyme-linked immunosorbent assays (ELISAs), which do not suffer from this shortcoming, have recently been used to show a negative correlation between anti-NA titers and virus shedding in humans in a cohort study. 24 In the same cohort, anti-HA stalk antibodies have been shown to be an independent correlate of protection against both infection as well as symptomatic disease.…”

Use of lentiviral pseudotypes as an alternative to reassortant or Triton X‐100‐treated wild‐type Influenza viruses in the neuraminidase inhibition enzyme‐linked lectin assay

“…A previous comparison between NI titers using H11N1 and N1 only PVs in ELLA highlighted differences in titers . Whether this was due to PV stability, HA serum inhibition, or interference by antibodies that bind HA stem is unclear and should be further investigated.…”

“…Further studies are needed to examine the reason for higher titers being measured using H6N1 as the antigen. In fact, it can either depend on different antigen accessibility of antibodies or different sensitivity of the assay . Although a 4‐fold increase is usually considered seroconversion when pre‐ and post‐vaccination sera are evaluated according to international harmonization guidelines, these assay still need to be validated to draw more robust conclusions.…”

Kinetics, Longevity, and Cross-Reactivity of Antineuraminidase Antibody after Natural Infection with Influenza A Viruses

“…The antihemagglutinin (anti-HA) antibodies play a dominant role in protection against HA-matched influenza virus, while anti-NA antibodies may also be protective in the case of mismatched HA (10)(11)(12)(13)(14)(15)(16). HA antibodies block viral attachment to the target cell surface, and if the virus enters into the endolysosome, these antibodies also block viral uncoating and release of the nucleoprotein into cytoplasm.…”