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Comparative Study
. 2019 Mar 1;63(sp1):138-144.
doi: 10.1637/11886-042718-Reg.1.

Are Microneutralization and Hemagglutination Inhibition Assays Comparable? Serological Results from Influenza Experimentally Infected Mallard Ducks

Karen M Segovia  1 , Monique S França  1 , Charlie S Bahnson  2 , Neus Latorre-Margalef  2   3 , David E Stallknecht  4
Affiliations
Comparative Study

Are Microneutralization and Hemagglutination Inhibition Assays Comparable? Serological Results from Influenza Experimentally Infected Mallard Ducks

Karen M Segovia et al. Avian Dis. .

Abstract
in English, Spanish

The hemagglutination inhibition (HI) assay is commonly used to assess the humoral immune response against influenza A viruses (IAV). However, the microneutralization (MN) assay has been reported to have higher sensitivity when testing sera from humans and other species. Our objective was to determine the agreement between MN and HI assays and compare the proportion of positive samples detected by both methods in sera of mallards primary infected with the A/mallard/MN/Sg-000169/ 2007 (H3N8) virus and subsequently inoculated with homosubtypic or heterosubtypic IAV. Overall, we found poor to fair agreement (prevalence-adjusted bias-adjusted kappa [PABAK], 0.03-0.35) between MN and HI assays in serum samples collected 2 weeks after H3N8 inoculation; the observed agreement increased to moderate or substantial in samples collected 4 to 5 weeks postinoculation (WPI) (PABAK, 0.52-0.75). The MN assay detected a higher proportion of positive samples compared with HI assays in serum samples collected 2 WPI (P = 0.01). This difference was not observed in samples collected 4 WPI. Also, a boosting effect in MN and HI titers was observed when birds were subsequently inoculated with IAV within the same H3 clade. This effect was not observed when birds were challenged with viruses that belong to a different HA clade. In summary, the agreement between assays varies depending on the postinfection sample collection time point and the similarity between the antigens used for the assays. Additionally, subsequent exposure of ducks to homosubtypic or heterosubtypic strains might affect the observed agreement.

¿Los ensayos de microneutralización e inhibición de la hemaglutinación son comparables? Resultados serológicos de patos de collar infectados experimentalmente con influenza. La prueba de inhibición de la hemaglutinación se usa rutinariamente para evaluar la respuesta inmune humoral contra los virus de influenza aviar, sin embargo, se ha reportado que la prueba de microneutralización tiene una mayor sensibilidad cuando se evalúan muestras de suero de humanos u otras especies. Este estudio tuvo como objetivo determinar la concordancia entre las pruebas de microneutralización e inhibición de la hemaglutinación en suero de patos de collar que fueron desafiados con el virus A/ mallard/MN/Sg-000169/2007(H3N8) y re-inoculados con virus de influenza aviar homosubtípicos o heterosubtípicos. Además, se comparó la proporción de muestras positivas detectadas por ambos métodos. En general, se observó un nivel de concordancia pobre a razonable (PABAK = 0.03 - 0.35) entre las pruebas de microneutralización e inhibición de la hemaglutinación en muestras de suero recolectadas dos semanas post-inoculación del virus H3N8. La concordancia se incrementó a moderada o sustancial en muestras recolectadas cuatro o cinco semanas después de la inoculación (PABAK = 0.52 - 0.75). Una mayor proporción de muestras recolectadas a las dos semanas después de la inoculación fueron positivas por microneutralización en comparación con inhibición de la hemaglutinación (P = 0.01), estas diferencias no fueron observadas con las muestras recolectadas a las cuatro semanas después de la inoculación. Adicionalmente, se observó un incremento en los títulos de anticuerpos cuando las aves fueron re-inoculadas con virus de influenza aviar pertenecientes al mismo clado H3 de la hemaglutinina. Este efecto no fue observado en los patos re-inoculados con virus de influenza aviar pertenecientes a un clado distinto. En resumen, la concordancia entre los ensayos varía según el momento de recolección de la muestra y la similitud entre los antígenos utilizados para los ensayos. Además, la re-inoculación de patos con una cepa homosubtípica or heterosubtípica podría afectar el nivel de concordancia observada.

Keywords: hemagglutination inhibition; influenza A virus; mallards; microneutralization; serology.

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Figures

Figure 1.
Figure 1.. Dynamics of serological response by MN and HI assays.
The graph shows variation of geometric mean antibody titers across time based on HI and MN assays in different experimental groups. Arrows indicate the time points of infections and circles, diamonds, and squares the time-points at which samples were tested by MN and HI assays.
See this image and copyright information in PMC

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References

    1. Alexander DJ A review of avian influenza in different bird species. Vet Microbiol 74:3–13. 2000. - PubMed
    1. Bucukovski J, Latorre-Margalef N, Stallknecht DE, and Miller BL A multiplex label-free approach to avian influenza surveillance and serology. PloS one 10:e0134484 2015. - PMC - PubMed
    1. Gross P, and Davis A Neutralization test in influenza: use in individuals without hemagglutination inhibition antibody. J Clin Microbiol 10:382–384. 1979. - PMC - PubMed
    1. Ito T, Suzuki Y, Mitnaul L, Vines A, Kida H, and Kawaoka Y Receptor specificity of influenza A viruses correlates with the agglutination of erythrocytes from different animal species. Virology 227:493–499. 1997. - PubMed
    1. Kida H, Yanagawa R, and Matsuoka Y Duck influenza lacking evidence of disease signs and immune response. Infect Immun 30:547–553. 1980. - PMC - PubMed

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