International laboratory comparison of influenza microneutralization assays for A(H1N1)pdm09, A(H3N2), and A(H5N1) influenza viruses by CONSISE

The microneutralization assay is commonly used to detect antibodies to influenza virus, and multiple protocols are used worldwide. These protocols differ in the incubation time of the assay as well as in the order of specific steps, and even within protocols there are often further adjustments in individual laboratories. The impact these protocol variations have on influenza serology data is unclear. Thus, a laboratory comparison of the 2-day enzyme-linked immunosorbent assay (ELISA) and 3-day hemagglutination (HA) microneutralization (MN) protocols, using A(H1N1)pdm09, A(H3N2), and A(H5N1) viruses, was performed by the CONSISE Laboratory Working Group. Individual laboratories performed both assay protocols, on multiple occasions, using different serum panels. Thirteen laboratories from around the world participated. Within each laboratory, serum sample titers for the different assay protocols were compared between assays to determine the sensitivity of each assay and were compared between replicates to assess the reproducibility of each protocol for each laboratory. There was good correlation of the results obtained using the two assay protocols in most laboratories, indicating that these assays may be interchangeable for detecting antibodies to the influenza A viruses included in this study. Importantly, participating laboratories have aligned their methodologies to the CONSISE consensus 2-day ELISA and 3-day HAMNassay protocols to enable better correlation of these assays in the future

H1N1 viral proteome peptide microarray predicts individuals at risk for H1N1 infection and segregates infection versus pandemrix(®) -vaccination

A high content peptide microarray containing the entire Influenza A virus (A/California/08/2009(H1N1)) proteome and hemagglutinin proteins from 12 other influenza A subtypes, including the hemagglutinin from the (A/South Carolina/1/1918(H1N1)) strain, was used to gauge serum IgG epitope signatures prior to and after pandemrix vaccination/ or H1N1 infection in a Swedish cohort during the pandemic influenza season 2009. A very narrow pattern of pandemic flu specific IgG epitope recognition was observed in the serum from individuals who later contracted H1N1 infection. Moreover the pandemic influenza infection generated IgG reactivity to two adjacent epitopes of the neuraminidase protein. The differential serum IgG recognition was focused on hemagglutinin 1 (H1) and restricted to classical antigenic sites (Cb) in both the vaccinated controls and individuals with flu infections. We further identified a novel epitope VEPGDKITFEATGNL on the Ca antigenic site (251-265) of the Pandemic flu hemagglutinin, which was exclusively recognized in serum from individuals with previous vaccinations and never in serum from individuals with H1N1 infection (confirmed by RNA PCR analysis from nasal swabs). This epitope was mapped to the receptor-binding domain of the influenza hemagglutinin (HA) and could serve as a correlate of immune protection in the context of pandemic flu. The study shows that unbiased epitope mapping using peptide microarray technology leads to the identification of biologically and clinically relevant target structures, it also shows that H1N1 infection induced a different footprint of IgG epitope recognition patterns as compared to the pandemic H1N1 vaccine. This article is protected by copyright. All rights reserved

Memory-like innate response to booster vaccination with MF-59 adjuvanted influenza vaccine in children

Abstract The pediatric population receives the majority of vaccines globally, yet there is a paucity of studies on the transcriptional response induced by immunization in this special population. In this study, we performed a systems-level analysis of immune responses to the trivalent inactivated influenza vaccine adjuvanted with MF-59 in children (15–24 months old) and in young, healthy adults. We analyzed transcriptional responses elicited by vaccination in peripheral blood, as well as cellular and antibody responses following primary and booster vaccinations. Our analysis revealed that primary vaccination induced a persistent transcriptional signature of innate immunity; booster vaccination induced a transcriptional signature of an enhanced memory-like innate response, which was consistent with enhanced activation of myeloid cells assessed by flow cytometry. Furthermore, we identified a transcriptional signature of type 1 interferon response post-booster vaccination and at baseline that was correlated with the local reactogenicity to vaccination and defined an early signature that correlated with the hemagglutinin antibody titers. These results highlight an adaptive behavior of the innate immune system in evoking a memory-like response to secondary vaccination and define molecular correlates of reactogenicity and immunogenicity in infants