Update on the Efficacy of Vaccination on Host Immunity to Influenza Virus Infection

Abstract

Background

Influenza is a highly contagious viral disease that infects the epithelial cells of the upper and lower respiratory tract. Among circulating viruses, influenza A and B strains are responsible for the majority of clinically significant disease and seasonal epidemics.

Summary

The severity of influenza infection and the effectiveness of vaccination are largely determined by the magnitude and quality of the host immune response. Both innate and adaptive immune mechanisms contribute to viral clearance, recovery, and the establishment of protective immunity. Although vaccination remains the most effective strategy to reduce influenza-related disease burden, its efficacy varies due to viral evolution, host factors, and immune response variability.

Key Messages

Ongoing surveillance of circulating influenza strains and continued advancements in vaccine design are essential for improving preventive strategies and protecting public health. Further research into novel vaccine approaches and immune mechanisms is critical to enhance vaccine efficacy, address vaccine hesitancy, and ultimately reduce influenza-associated morbidity and mortality.

Introduction

Influenza viruses have a persistent impact on human well-being. Prior to the COVID-19 outbreak, there were approximately 0.2–0.6 million annual deaths due to influenza infections and associated far greater number of hospitalizations. Influenza incidence declined dramatically during COVID-19 coinciding with broad non-pharmaceutical interventions (masking, distancing, school closures, travel restrictions) [1, 2]. However, the levels of influenza infections are returning to those seen before the pandemic particularly those due to the A(H1N1)pdm09, A(H3N2), and B/Victoria strains [3, 4].

Prior to COVID-19, 25–60% of annual influenza cases were associated with infection by influenza B strains of the virus (IBVs) [5, 6]. Seasonal IBV infections tend to result in fewer hospitalizations compared to those caused by influenza A strains (IAVs) such as H3N2, yet they still lead to more hospitalizations than IAV H1N1. Interestingly, the IBV Yamagata strain has not been reported since the pandemic and may have been eradicated by COVID-19 preventive strategies [7].

As a member of the Orthomyxoviridae ribonucleic acid (RNA) viruses, antigenic differences between strains are used to classify influenza viruses, namely, IAV, IBV, and the C (ICV) and D (IDV) strains [8, 9]. IAV infects humans and many animals with IBV and ICV predominantly infecting man while IDV does not infect humans but does infect cattle and pigs [10, 11]. This review will primarily focus on IAV infection, following a brief overview of IBVs.

IBVs, the cause of seasonal epidemics, mainly infect children and older adults triggering severe disease, but since there appears to be no reservoir in animals, it is likely to have limited pandemic potential. Seasonal influenza vaccination generally restricts the IBV infection burden; however, due to the antigenic mismatches, in complete immunity, there occurs some cases. Therefore, elucidating the mechanisms of host immunity toward viral infection is essential for combating future infections. IBV is commonly considered as being a less severe influenza virus due, in part, to the limited clinical and public health studies investigating outcomes in hospitalized patients.

Despite similarities between IAV and IBV, these viruses are molecularly different with respect to proteins, species and cell preference, and the ability to trigger antiviral pathways. Two lineages of IBVs exist, namely, B/Victoria and B/Yamagata, although as noted above, the Yamagata lineage may have been eradicated. Both IBV and IAV lineages are composed of clades and sub-clades. Airway epithelial cells are the major replicative target for both IAV and IBV via viral hemagglutinin (HA) protein-mediated binding to host cell receptors for sialic acid. Virus attachment to the receptor triggers membrane fusion, endosomal localization, and the secretion of viral ribonucleoproteins (vRNP). There are eight vRNPs that are required for nuclear replication [12]. A distinguishing feature of IBV is the lack of the expression of polymerase basic protein (PB)1-F2 and polymerase acidic protein (PA)-X. These are virulence factors which are produced by IAV [13].

After replication of viral RNA and expression of the related proteins, the assembly of viral proteins occurs followed by budding under the control of neuraminidase (NA) produced by the virus. The IAV nonstructural 1 (NS1) protein suppresses recognition of viral RNA by retinoic acid-inducible gene I (RIG-I) and interferes with Toll-like receptor (TLR) signaling, both of which are critical for initiating the type I interferon (IFN-1) response. The IFN-1s IFN-α and IFN-β are rapidly produced upon infection and, in turn, upregulate IFN-stimulated genes which have a negative feedback effect on viral replication [14, 15].

Epidemiological studies indicate a number of subject groups that are highly susceptible to severe, potentially lethal outcomes from influenza infection including subjects with comorbidities, pregnant women, children, and subjects more than 65 years of age [16]. Secondary lung infection following IAV infection can result in lung alveolar damage and reduced alveolar barrier function leading to infiltration of a protein-rich fluid into the airspace. This is the main cause of death from IAV which is a leading cause of death worldwide. Antiviral drugs are used for IAV-induced lung injury but are ineffective once lung injury has commenced, underscoring an urgent need for new therapeutic strategies.

Infection Cycle with Influenza Virus

After virus internalization by the body, they target epithelial cells within the airway which represent the main target for infection [17]. Detection of the acute infection and initiation of early antiviral and inflammatory responses involve these infected epithelial cells together with innate immune cells residing in the airway such as alveolar macrophages and dendritic cells (DCs). The secretion of cytokines and chemokines by resident and circulating immune and inflammatory cells results in the recruitment and activation of macrophages, granulocytes, and natural killer cells and eosinophils to the site of infection [18].

Infective complete IAV virions require both viral and host proteins for assembly [19]. Ten structural and 9 nonstructural viral proteins are encoded by single-stranded viral RNAs and include polymerase basic proteins (PB)1 and 2, PA, nucleoprotein (NP), matrix protein (M1), NS1 and 2, and nuclear export protein (NEP) which are deposited within the lipid membrane. Furthermore, M2, HA, and NA enable antibody detection as they protrude through the membrane envelope [20–23].

The surface glycoprotein HA of IAVs is the main mediator of viral entry. Its main receptor is α-2,6-linked sialic acid, which is found across the whole of the respiratory epithelium. HA may also bind to non-sialylated phosphoglycans [24]. IAV is endocytosed by host cells, and this is followed by acidification of early endosome through M2 ion channels resulting in HA structural changes enabling fusion of the viral envelope with the endosomal membrane [25]. The uncoated virus is or vRNPs are released into the cytoplasm and subsequently undergo nuclear import. Initially, the negative-sense strand RNAs of virus makes a template for production of viral RNA by converting into positive-sense RNAs which generate complementary ribonucleoprotein (cRNP) [26].

Viral mRNAs are generated by transcription of vRNA, translocated into the cytoplasm where they generate viral proteins with the assistance of endoplasmic reticulum-related ribosomes. Some of the translated products reenter the nucleus to assemble vRNP, followed by transfer of these vRNP to the cytoplasm and then to the plasma membrane to be assembled with surface HA and NA, thereby creating new virions [27, 28]. The final step involves the shedding, assembly, and release of mature virus particles [29]. NA mediates the cleavage of HA on newly formed viral particles from the host cell’s sialic acid receptors enabling the release of viral progeny. This, in turn, stimulates the replication, transcription, translation, and release of the virus and its subsequent infection of neighboring cells – their propagation through exhaled droplets [30]. The cycle of IAV infection from cell entry to virion budding and release as new virions is shown in Figure 1.

Fig. 1.

IAV structure, infection cycle, and replication in a host cell. a. Schematic diagram of IAV structure shows that the virus contains eight RNA segments, M1 matrix protein, surrounded by a lipid bilayer expressing proteins like HA, NA, and M2 ion channels. b. Infection cycle starts with the attachment of HA proteins of the virus to the sialylated glycoprotein receptors expressed on the host cells and virus internalization by endocytosis. Viral membrane and host cell membrane are fused due to the low pH in the endosomes, and subsequently, viral compartments including vRNPs are released into the cytoplasm. After internalization of vRNPs into the nucleus, the vRNA(−) converts into a cRNA(+). Viral mRNA(+) is also generated from vRNA(−) by the aid of the host’s transcriptional system which is responsible for the synthesis of viral proteins. The generated vRNPs which are exported from the nucleus to the cytoplasm, together with the other viral proteins, are gathered into a new virion that buds from the host cell. This process is catalyzed by the aid of NA which cleaves the bond between HA and sialylated receptor to release the new virus. IAV, influenza A virus; HA, hemagglutinin; NA, neuraminidase; vRNP: viral ribonucleoprotein; vRNA(−), negative sense viral RNA; cRNA(+), positive sense RNA; NS1: nonstructural protein 1; NS2: nonstructural protein 2; PB1: polymerase basic protein 1; PB2: polymerase basic protein 2.

Immune System Responses Subsequent to Influenza Virus Recognition

Innate Immune Responses

Airway ciliated and secretory epithelial cells utilize several mechanisms to activate primary innate immunity against inhaled pathogens such as IVs. Secretory epithelial cells produce a mucin-rich layer that physically protects the airway by capturing viruses and other pathogens which are then removed through mucociliary clearance. The virus has to pass through this barrier to reach and infect the epithelial cells because the viral attack is prevented by the sialylated decoy receptors of the mucus which protect the epithelial cell. However, the virus is capable of breaking these decoy interactions and penetrating through the mucus layer [31]. Of note, the severest form of influenza infection involves evoking alveolar cell death following viral spread [32].

The primary line of immune defense involves recognition of intracellular virus by pattern recognition receptors such as TLRs and RIG-I. Intracellular TLRs sense intracellular viral antigens and/or viral antigens generated during replication [15].

Endosomal TLRs such as TLR7 and TLR8 are present in numerous immune and nonimmune cells where they recognize single-stranded RNA, but differ in their oligonucleotide responsiveness [33]. In contrast, endosomal TLR3 expressed mainly in alveolar and airway epithelial cells recognizes double-stranded RNA produced during viral replication [31]. Recent studies confirm that influenza virus RNA induces IRF3 and IRF7 following detection by TLR3, TLR7, and TLR8 indicating a robust IFN-1 antiviral defense [15].

The release of damage-associated molecular patterns such as S100A9, high-mobility group box 1, oxidized phospholipids, and calgranulin B/MRP-14 during IAV infection is recognized by TLR4 on host cells, triggering proinflammatory cytokine and IFN release [34, 35]. During IAV infection, activation and upregulated expression of TLR10, along with elevated secretion of proinflammatory cytokines such as IL-6, IL-8, IL-29, and IFN-β, have also been reported [36].

Cytosolic receptors such as RIG-I, expressed in epithelial cells, macrophages, and DCs also recognize IV. IAV-derived genomic material, such as viral RNA with phosphorylated 5′ ends [37], including short viral RNAs produced by IAV polymerase, is recognized by RIG-I in infected cells [38]. The downstream signaling pathways of IFNs, which are triggered by RIG-I, can be antagonized by NS1 of IAV [14]. Despite TLRs initiating protective antiviral immunity upon viral sensing, uncontrolled activation of these receptors may exaggerate the outcome of a viral infection. For instance, immune-mediated fatal influenza virus infection is not seen in the absence of TLR3 in mice [39]. Moreover, inhibition of overactive TLR3-mediated signaling may alleviate disease symptoms [40]. Antagonism of TLR7 has similar effects in reducing the viral-driven inflammation and mortality rates [41]. Interestingly, sialic acid receptor-mediated internalization of IV in lung platelets [42] triggers TLR7 signaling and subsequent neutrophil aggregation, enhancing myocardial infarction risk [43].

Pattern recognition receptor signaling enhances IFN-1 and IFN-3 production and activation of ISGs that mediate antiviral responses. IFN-λ, a type 3 IFN, is generated before that of IFN-1 and IFN-2 (IFN-α and IFN-β) [44] to attenuate the spread of virus through the lungs [39]. Sequential generation of different IFN subtypes attenuates tissue damage resulting from continual presence of IFN-1 [45, 46] although this is not complete as sustained IFN-3 activation prevents the regeneration of damaged lung epithelial cells thereby worsening the severity of the infection [47]. IFN-1 (IFN-α/β) act systemically to induce antiviral defenses, whereas IFN-3 (IFN-λ) primarily protects epithelial surfaces and limits virus spread to the lungs [44]. Overall, raised IFN activation is essential for the generation of effective antiviral responses and may be induced by pro-inflammatory cytokines secreted by resident and infiltrating immune-inflammatory cells.

Adaptive Immune Responses

After detecting IAV in the airways and lungs, activated antigen presentation cells such as DCs migrate to local lymph nodes where antigen presentation to naïve T lymphocytes occurs via major histocompatibility complexes (MHCs) to generate effector CD4⁺ and CD8⁺ T cells [48]. Antigen presentation is required for CD8⁺ T-cell activation and expansion but their levels and polyfunctionality are shaped by IL-1-dependent signaling pathways during viral infections [49]. Th1 cell differentiation triggers cytokine production including that of IL-2, IFN-γ, and TNFα. CD4⁺ T cells are pivotal in generating humoral antiviral responses by assisting the differentiation of antibody-producing B cells [50]. Generated plasma cells produce neutralizing antibodies against exposed viral antigens such as HA, NA, and ion channels (M2 protein). These antibodies assist phagocytosis of virally infected cells through opsonization and antibody-dependent cellular cytotoxicity [51, 52].

Presentation of viral antigens to MHC-1 on DCs also activates naïve CD8⁺ T cells within local lymph nodes [53]. Differentiation of CD8⁺ cells into cytotoxic T cells and their migration into the lungs result in apoptosis of viral-infected cells following membrane pore formation by perforin and granzyme [54]. Figure 2 shows triggering of innate and adaptive immune pathways by IAV.

Fig. 2.

Schematic diagram of the innate and adaptive immune responses triggered by IAV. The mucosal surfaces of the respiratory system’s epithelial barrier contain mucins, as natural primary defense, which have terminal sialic acid residues and bind with HA of IAV and prevent the penetration of the virus to the lower layer. However, viral NA cleaves this bond and releases the trapped virus to attach to the sialylated receptors on the host epithelial cells. After virus endocytosis and envelope fusion, internal antigens like genomic materials are recognized by intracellular TLRs like TLR3 and TLR7 that induce both NF-κB and type I interferon pathways. Viral RNA in the cytosol is also recognized by RIG-I, inducing IFN pathways (suppressed by viral NS1 protein). Membrane PRRs such as TLR4 and TLR10 also recognize DAMPs and some viral antigen and trigger NF-κB signaling. After detection of IAV by the innate immune system, APCs are activated to present viral antigens and prime adaptive immune cells. Activation of APCs triggers the differentiation of naïve CD4⁺ T cells into T helper and TFH cells. TFH cells activate B cells which later generate plasma cells and neutralize antibodies against surface viral antigens including HA, NA, and M2 ion channels. Antigen-specific CD8⁺ T cells differentiate into CTLs which mediate the elimination of infected cells. IAV, influenza A virus; vRNP, viral ribonucleoprotein; NA, neuraminidase; NS1, nonstructural 1; NS2, nonstructural protein 2; PB1, polymerase basic protein 1; PB2, polymerase basic protein 2; HA, hemagglutinin; NA, neuraminidase; RIG-I, retinoic acid-inducible gene I; TLR, Toll-like receptor; MHC-I, major histocompatibility complex-I; MHC-II, major histocompatibility complex-II; CTL, cytotoxic T lymphocytes; APC, antigen presenting cell; TFH, T follicular helper; NF-κB, nuclear factor kappa-light-chain-enhancer of activated B cells; PRRs, pattern recognition receptors; DAMPs, damage-associated molecular patterns.

Immunity Induced by Vaccination

An annual influenza vaccination has been recommended for all individuals aged ≥ 6 months who do not have contraindications since 2010 by the Centers for Disease Control and Prevention (CDC) and also by the Advisory Committee on Immunization Practices (ACIP) [55]. Vaccination gives significant protection against influenza infection, but its efficacy is modified by various factors including patient age, recipient health status, vaccine type, the strain of influenza viruses present within the community, and the similarity between the strain for which the vaccine was generated against and that which is prevalent in the community [56].

Individuals at risk of infection should receive a vaccine appropriate for their age, except for adults having undergone solid organ transplantation, who are receiving immunosuppressive medications, who may be treated with either HD-IIV3 or aIIV3. The timing of vaccination will depend upon the onset, peak activity, and duration of any outbreak which varies each year [57].

The timing of influenza vaccinations should take into account seasonal differences between hemispheres. Vaccination generally begins in September and continues through December in the North, while in the Southern Hemisphere, it usually starts in April or May, before the winter influenza season [55]. However, administration of vaccines can continue any time during the circulation period of the virus. Routine visits to health care providers or hospitalizations provide good opportunities for offering vaccinations [57]. Boosters, except for the elderly, is generally not beneficial and should not be recommended [58–61].

However, giving the influenza vaccine in July or August may be appropriate for individuals who might not have the chance to get vaccinated at the optimal time. The decline in vaccine efficacy varies with no link to age, season, or viral type/subtype. Immunity against influenza A(H3N2) viruses has been shown to wane more rapidly than immunity to influenza A (H1N1) or B viruses [62].

Antibodies Induced after Natural Infection and Vaccination

Viral surface glycoproteins, including HA and NA, induce antibodies naturally, and this exerts a protective role against influenza infection [63]. Indeed, for many years, serum antibodies have been considered protective [62]. Therefore, a lack of adequate antibody protection within the population is a major cause of pandemic emergence. Indeed, influenza viruses undergo changes in some of their features, especially with respect to point mutations in surface glycoproteins (antigenic drift). The globular head region of the virus has substantial plasticity which overcomes the impact of these changes [64, 65]. Obviously, there is a need for annual updates to influenza virus vaccines annually, due to antigenic drift [66]. The similarity between the circulating virus strain and that against which the vaccine was generated is an important factor in vaccine effectiveness [67, 68].

Eight genomic segments are present in IAV and IBV, which encode 11 proteins, that are potential antibody targets. However, antibody responses against these proteins are not uniform, and the final response depends on features of the targeted proteins, including accessibility [69].

As expected, the surface glycoproteins, the trimeric HA (to a higher degree), and tetrameric NA, present on virions and infected cells, are accessible to antibodies. M2 ion channel of IAV is also an accessible target for antibodies, in addition to patches of NP, which are detected on the surface of infected cells [70]. During an influenza infection, cell lysis can expose internal viral proteins such as M1, NP, PB1, PB2, PA, NS1, and NEP. These proteins are not typically secreted into body fluids but can become accessible to the host immune system following cell death, enabling antibody recognition [71].

After inhalation of the influenza viruses, the HA on the virion binds to the mucosal surfaces via the terminal sialic acid residues present on mucins [72], as a natural defense strategy against host cell binding [73]. NA is responsible for the cleavage of this bond and releasing the trapped virus [74], leading to virus penetration into the airway lining fluid providing access to epithelial and immune cells. The influenza virus is endocytosed following binding to the sialylated receptor on host cells. The immune system is capable of recognizing the viral HA and NA during this period and thereby triggering an antibody response [26]. Endosomal acidification enables fusion of endosomal and viral membranes which facilitates the viral genome to be released into the cytoplasm, its nucleus translocation, and the generation of vRNA, cRNA, and mRNA and finally viral protein expression.

In contrast to HA, NA, and M2, other viral proteins such as M1, NP, PB1, PB2, PA, and NEP are transferred into the new virion. New virions do not typically include NSP1, PB1-F2, and PA-X. B lymphocytes usually detect the cell surface fragment of viruses including HA, NA, and M2, triggering an antibody response against these proteins. Moreover, although NP is usually present inside the cells, it can be detected on patches on the cell surface by B cells [75].

While the new budding virion utilizes HA binding to sialic acids on host cell membranes, NA breaks the bond to release the nascent virus. During this step, B lymphocytes easily detect HA, NA, and even extremely low levels of M2; however, the internal proteins shielded by the virus membrane are not detectable for B cells. The viral HA is eventually proteolytically degraded into its component subunits HA1 and HA2 in the respiratory tract to yield the infectious virus. Moreover, B cells can detect almost all the viral proteins expressed by dying cells and cell debris.

HA-specific antibodies are important in modulating the response to infection, and several methods such as the HA inhibition assays and HA neutralization assays have been utilized to evaluate the polyclonal responses to influenza virus infection. Natural infection induces antibodies which mostly target HA, while to a lower extent recognize NA and internal proteins. The results obtained from HA inhibition assays, ELISA, and microneutralization assays have shown that natural infection induces seroconversion in most infected individuals [75–78]. Previous exposures of individuals to the virus affect the extent of antibody responses seen with natural influenza infection against HA. Adults usually exhibit wider antibody responses than children, who have had limited exposure histories. However, children may also exhibit broader responses toward the virus and induce antibody responses to a previous strain or even the current strain [63].

Antibody Production by Vaccination

Current influenza vaccines are mainly categorized into three main groups including inactivated, recombinant, and live-attenuated influenza vaccines, with no preference expressed by the CDC for any of these vaccine platforms. Age considerations suggest that recombinant vaccines should be used in adults (18+ years) [79], while people aged between 2 and 49 years of age and those who are not at high risk may be treated with live-attenuated influenza vaccines [80]. Older individuals (65+ years) are advised to receive adjuvant-containing vaccines at a high dose to generate a greater immune response against infection [81, 82].

There are three primary categories of inactivated influenza vaccines: subunit vaccines containing HA and NA; split-virus vaccines consist of viral proteins lacking a lipid envelope; and purified chemically inactivated whole-virus vaccines [83]. Antigenic drift, driven by HA and NA mutations, significantly affects vaccine efficacy [82] and may contribute to vaccine escape [75].

Antigenic shift induces a novel HA or NA, creating a distinct influenza subtype which may be amplified by genetic reassortment due to mixing with animal-derived strains [84]. Antibodies against HA epitopes used in inactivated influenza vaccines [85] are vulnerable to reduced efficacy because of antigenic drift [77]. Moreover, the ability to generate a clinically useful antibody response in vaccinated individuals will depend upon strain-specific protection induced by many vaccines [86].

Influenza viruses remain a world-wide concern with the risk of seasonal epidemics [87]. In the USA, 38 million subjects were infected during the 2019 influenza season costing USD 10 billion in medical expenses [88, 89]. Most high-risk individuals are equally susceptible to both influenza and COVID-19 infection [87, 90] since they possess skewed immunity that may impact both the evolution of the virus and the pathophysiology of the infection [3, 91].

Host factors also affect vaccine response. High-risk populations including obese individuals do not benefit as well as others from vaccination despite having similar seroconversion rates to that observed in nonobese subjects. In addition, obese subjects have more severe symptoms and more frequent infections [91, 92]. This is counteracted by the greater likelihood of obese adults receiving an influenza vaccine [93]. The poor vaccine-induced innate and adaptive immune responses seen in these patients compromise the duration of any protection conferred [94]. In addition, adjuvant-containing vaccines, which should generate more antibodies than other vaccines, are not as efficient at protecting obese mice from influenza infection [95, 96].

Another important aspect influencing vaccine effectiveness is the attenuating effect of repeated annual vaccination on antibody responses. Studies show that repeated influenza vaccination either attenuates antibody titers or causes immune imprinting according to similarity between strains used to generate vaccines and viruses within the community. This phenomenon suggests that prior immune history can modulate the quality and durability of subsequent vaccine-induced immunity. Understanding this attenuation is critical for designing vaccination schedules not only for influenza but also for other respiratory vaccines such as those for SARS-CoV-2, where similar immune interference mechanisms have been observed [97, 98].

Although activation of CD4⁺ and CD8⁺ T cells protect against severe influenza, the degree of activation must be controlled to prevent immune and inflammatory cell cascades and the induction of immunopathological damage [99]. Current research indicates that while vaccines effectively reduce the severity and duration of influenza symptoms, their effectiveness varies upon the strain that the vaccine is directed against and their presence, individual age and health status, and the overall immune response. Continuous monitoring and updates to vaccine formulations are essential to improve outcomes, especially in high-risk populations.

Overall, the efficacious public health approach to preventing and mitigating against infection by influenza remains vaccination. This emphasizes the importance of annual vaccination in promoting robust immunity against this evolving virus.

Conflict of Interest Statement

The authors report no conflicts interest in relation to this manuscript.

Funding Sources

No funding was obtained for this study.

Author Contributions

E.M. wrote the manuscript. S.A.-V. helped with the figures and revising. N.K.D. and F.H. helped with acquisition of new data and overviewed the manuscript. H.J., M.V., and P.T. as clinicians helped on the clinical relevance of data. I.M.A. was the principal of part of the project related to this paper and revised all parts of manuscript.

Funding Statement

No funding was obtained for this study.

Data Availability Statement

This study is a review; no new data were generated or analyzed, and data sharing is not applicable.