Extrafollicular memory b cell responses during influenza infection - protective antibody responses to influenza virus infection are generated in germinal center (GC) reactions, which generate…

The National Institute of Allergy and Infectious Diseases awarded The Johns Hopkins University $100,228 on November 27, 2026, under the Allergy and Infectious Diseases Research program (CFDA 93.855) to investigate extrafollicular memory B cell responses during influenza infection.

The project uses single-cell CITE-seq with antigen barcoding to characterize B cells isolated from influenza virus-infected subjects, testing the hypothesis that extrafollicular responses generate broadly reactive antibody pools functionally preserved in the extrafollicular-derived memory B cell compartment. The research aims to establish whether this compartment provides heterosubtypic immunity both locally in the respiratory tract and systemically, addressing a gap in understanding how extrafollicular-derived memory cells contribute to systemic and tissue-specific long-term immunity, particularly regarding their antigenic repertoire and capacity to participate in recall responses.

Performance runs through June 30, 2029, at Johns Hopkins' Baltimore, Maryland location. This is a Project Grant, a standard assistance type under the NIAID program supporting biomedical research and research training in infectious diseases and related areas at universities, research organizations, and individual investigators.

Extrafollicular memory b cell responses during influenza infection - protective antibody responses to influenza virus infection are generated in germinal center (GC) reactions, which generate antigen-specific, affinity-matured, class-switched plasma cells as well as memory b cells (mbcs). GC-derived mbcs will respond vigorously to RE-encounter, mostly through rapid differentiation to plasmablasts in so-called extrafollicular foci, located outside the primary follicle in the interfollicular and medullary cord areas of the respiratory tract draining mediastinal lymph nodes. Both during primary infection and during recall, extrafollicular responses (EFR) produce antibody- secreting plasmablasts rapidly and prior to GC formation. They are increasingly recognized as essential for early control of viral infections. Regulation of EFR is incompletely understood but known to involve TLR signaling of b cells. These innate signals may contribute to the activation of b cell clones with a broad range of affinities for the activating antigen, resulting in a broadly reactive antibody pool, prioritizing breadth of coverage over specificity. Additionally, the EFR also generates mbcs (EF- mbcs). These memory cells are largely unswitched but antigen-experienced and though to contribute mainly to GC responses following a recall infection. However, EF-MBC generation represents an emerging area of research and their contributions to systemic and tissue-specific long-term immunity remain poorly understood, particularly in terms of their antigenic repertoire and capacity to participate in recall responses. This project aims to test our hypothesis that the EFR generates a broadly reactive antibody pool, that is functionally preserved in the EFR derived MBC compartment, providing heterosubtypic immunity both locally in the respiratory tract and systemically. For that aim 1 will use single-cell cite-SEQ with antigen barcoding of b cells isolated from individual influenza virus a//puerto rico 8/34 infected mice to determine the breadth and depths of the virus-specific b cell repertoire of both, plasmablasts and mbcs. Aim 2 will test the functional potential of EF-mbcs in recall responses using adoptive cell transfer of time and IG-allogenic labeled cells and homo and heterologous challenge of recipient mice. Mice will be analyzed by flow cytometry, elisa, elispot and affinity measurements of the antibody pool to define and contrast the capacity of EF-MBC for reactivation to that of GC- derived mbcs in the context of both homologous and heterologous influenza viral challenge. By characterizing both the repertoire and functional capacity of EF-derived memory following influenza infection, this work will advance our understanding of an underexplored arm of humoral immunity. These findings may inform future vaccine strategies aimed at eliciting broad, heterosubtypic protection against diverse and evolving influenza strains.