Selection of a single amino acid substitution in the hemagglutinin molecule by chicken eggs can render influenza A virus (H3) candidate vaccine ineffective

Abstract

This study investigated whether a single amino acid change in the hemagglutinin (HA) molecule influenced the efficacy of formalin-inactivated influenza A (H3N1) vaccine candidates derived from high-growth reassortants between the standard donor of high-yield genes (A/PR/8/34 [H1N1]) and host cell variants generated from the same clinical isolate (A/Memphis/7/90 [H3N2]) by passage in embryonated chicken eggs. Two clones of the isolate generated by growth in eggs differed from the parent virus (represented by an MDCK cell-grown counterpart) solely by the presence of Lys (instead of Glu) at position 156 or Ile (instead of Ser) at position 186 in the HA1 subunit. The protective efficacy of egg-grown HA Lys-156 and HA Ile-186 reassortant variants was compared with that of the MDCK cell-grown reassortant vaccine. Classically, antibody titers in serum have been used to demonstrate vaccine efficacy. Here, parameters of B-cell responsiveness were monitored, including the kinetics, character, and localization of the primary antibody-forming cell (AFC) response and the development of B-cell memory in lymphoid tissues associated with the priming site (spleen) and responsive to pulmonary challenge with infectious virus (upper and lower respiratory tract lymph nodes). We show that the egg-grown HA Lys-156 variant induced an AFC profile vastly different from that elicited by the other two reassortant vaccines. The vaccine was poorly immunogenic; it induced antibodies that were cross-reactive prior to challenge but which, postchallenge with a lethal dose of the MDCK cell-grown reassortant virus, were targeted primarily to the HA Lys-156 variant, were of the immunoglobulin M isotype, were nonprotective, and were derived from the spleen. In contrast, the egg-grown HA Ile-186 variant was remarkably like the MDCK cell-grown virus in that protective immunoglobulin G antibodies were unaffected by the Ile-186 substitution but poorly recognized HA with Lys-156. Furthermore, memory AFC responsiveness was localized to regional lymphoid tissue in the upper respiratory tract, where challenge HA was found. Thus, it is recommended that in the selection of vaccine candidates, virus populations with the egg-adapted HA Lys-156 substitution be eliminated and that, instead, egg-grown isolates which minimally contain Ile-186 be used as logical alternatives to MDCK cell-grown viruses.

Protection of Mice Against a Lethal Influenza Challenge by Immunization with Yeast‐Derived Recombinant Influenza Neuraminidase

“…The preparation resulted in protection against lethal influenza in animals and in humans (Hocart et al, 1995;Kilbourne et al, 1995). However, the production of an influenza vaccine in eggs requires viral adaptation that can be associated with the generation of genetic-drift variants of hemagglutinin or neuraminidase (Robertson et al, 1994), which may provide suboptimal protection (Kodihalli et al, 1995). Moreover, these systems share many disadvantages when industrial scale-up is required.…”

Antibody-forming cell response to virus challenge in mice immunized with DNA encoding the influenza virus hemagglutinin

“…Enzyme-linked immunospot (ELISPOT) assays. Elements of HA-specific Bcell responsiveness, including antibody secretion, were assessed by a modification of the enzyme-linked immunospot (ELISPOT) assay (8,18,19,21,22,31,34). In brief, 96-well plates (Millipore) were coated overnight with isolated PR8 HA (0.5 g of HA in 100 l of PBS at 4ЊC).…”

“…These results raise the question of how HA DNA vaccination induces humoral immunity against influenza virus, in particular, whether the immune response is comparable to that generated by replicating virus or purified antigen. The immune response against influenza virus has been extensively studied (4,6), and the importance of HA-specific antibodies and cognate help provided by virus-specific helper T cells is well established (10,21,22,26,32,33). Two important aspects of protective immunity against live virus challenge are the presence of preexisting HA-specific antibodies and the rapid generation of antibodyforming cells (AFCs).…”

Gene silencing of β-galactosamide α-2,6-sialyltransferase 1 inhibits human influenza virus infection of airway epithelial cells

“…There were more than 29,000 cases of infections, and 3,000 human deaths attributed to the 2009 global H1N1 influenza pandemic [2]. Because of the high mutation rates of the viral genome, vaccines and drugs initially directed against the virus often become ineffective [3,4]. Therefore, measures are urgently needed to prevent and treat influenza virus infections, especially for high-risk groups and in the event of another pandemic.…”