The Influenza virus belong to the family of Orthomyxoviridae which consists of eight segments of negative-sense single-stranded RNA. The virus genome encodes 13 proteins which are: hemagglutinin (HA), neuraminidase (NA), M1 matrix protein (M1), M2 ion channel protein (M2), nuclear protein (NP), non-structural protein (NS1, NS2), and RNA polymerase complex (PB1, PB2, PA). There are five subgroups of the virus: influenza A virus; influenza B virus; influenza C virus; influenza D (Asha & Kumar, 2019). These can also be divided based on their surface glycoproteins such as HA and NA. One example is influenza A viruses which can be divided into 18 HA subtypes and 11 NA subtypes. Influenza A and B can lead to epidemics. Influenza C viruses generally cause a mild upper respiratory infection. Influenza D generally affects cattle rather than humans.
Influenza A
Influenza A is an enveloped, segmented, negative-strand RNA virus, belonging to the Orthomyxoviridae family. Influenza A viruses are divided into subtypes based on the proteins on the surface of the virus: hemagglutinin (HA) and neuraminidase (NA). Although its eight viral gene segments encode as many as 18 proteins there may be more (Shao et al, 2017). In fact, there are 18 different hemagglutinin subtypes and 11 different neuraminidase subtypes (Treanor, 2023). Influenza A virus is a highly infectious respiratory pathogen and is a significant threat to global public health (Shoa et al, 2017). Although identified in 1931, with attempts to develop a vaccine against the virus starting soon after the discovery, researchers still have to produce yearly vaccines. The perennial challenge is therefore to produce a one-off vaccine.
Influenza B
Influenza B viruses are not divided into subtypes, but instead are further classified into two lineages or strains rather than their antigenic differences, which are B/Yamagata and B/Victoria. Although not as concerning as influenza A, influenza B viruses are associated with significant morbidity and mortality in those who are immunocompromised (Zaraket et al, 2021).
Why vaccines are important
Public health measures to control the spread of the disease, including non-pharmaceutical measures such as isolation and mask wearing are helpful, but the use of vaccines is a much more effective method of prevention (WHO, 2021). This is because the pathogenesis of influenza is related both to the extent of viral replication for influenzae A and B and response of the host organism enabling it to survive (Treanor, 2023).
The current vaccination recommended for influenza by the World Health Organisation (WHO) for the 2024-2025 flu season for the northern hemisphere is a trivalent egg based or cell culture or recombinant based vaccine. They also recommend a quadrivalent egg- or cell culture-based or recombinant vaccine (WHO, 2024). Following monitoring of the virus they annually recommend the type of flu that we need to vaccinate against in each vaccine such as the following in the trivalent vaccine:
- A/Victoria/4897/2022 (H1N1) pdm09-like virus.
- A/Thailand/8/2022 (H3N2)-like virus.
- B/Austria/1359417/2021 (B/Victoria lineage)-like virus.
Vaccination itself is characterized by the persistence of antibodies and memory cells immune cells that can rapidly reactivate when exposed to the same pathogen. The subpart of a pathogen that initiates the formation of antibodies is called an antigen. These induce cell-mediated immunity, activating subsets of T lymphocytes and humoral immunity by stimulating B lymphocytes to produce specific antibodies. Vaccines induce antibodies that block the binding of the hemagglutinin (HA) which is the major surface glycoprotein of the influenza virus.
The HA is responsible for binding of the virus to cell surface receptors, supporting the infiltration of the viral genome into the cytoplasm of the host cell through membrane fusion, (Nobusawa, 1997). The binding site for the HA to the host cells is sialic acid which is a shallow pocket at the head of the HA, (Zang et al, 2020; Zhao et al, 2022). The Influenza A and B viruses recognize N-acetylneuraminic acid, whereas influenza C virus recognizes the different N-acetyl-9-O-acetylneuraminic acid as the receptor. By blocking the work of HA the virus is unable to attach to cells and replicate (Zhao et al, 2022). Antibodies which are directed against the globular head domain of the HA are strain specific as the HA is subject to constant antigenic drift driven by herd immunity (Chang & Zaia, 2019; Zang et al, 2020). In other words, if they target this domain, they are strain specific and can quickly lose their efficacy against drifted strains. It is because of this action that virus vaccines must be reformulated annually based on recommendations from the WHO and surveillance data from laboratories in both the Northern and Southern hemispheres.
There can also be genome reassortment when two or more IAV strains infect a single host. This is known as antigenic shift, resulting in novel progeny viruses with a new HA and/or NA that are immunologically new to the human immune system significantly increasing the risk of a pandemic (Carrat et al, 2007). The Influenza A virus (IAV) is therefore effective at evading the host's defence as they have multiple strategies to avoid being detected and eliminated by the host immunity (Chang & Zaia, 2019).
Historically whole pathogen vaccines have been manufactured that consist of entire pathogens that have been killed or weakened to prevent them causing disease. They still elicit an immune response from the host. Although they produce a strong protective immune response, they can also cause a range of adverse reactions. Subunit vaccines have since been produced as they are made from components or antigens of the virus. ‘Inactivated‘ vaccines are also available, providing very short-term and highly specific humoral immunity due to the frequent antigenic variations in the influenza virion (Bardiya & Bae, 2005).
The vaccine mediums
Egg-based vaccine manufacturing is the oldest form of manufacturing for the flu vaccine and is used to make both inactivated and live attenuated vaccines. Although they have been found to be more effective than cell derived, or recombinant vaccines production can be slow and may have an allergic reaction to the egg component in the vaccine (Pérez Rubio & Eiros, 2018).
Cell culture-based inactivated flu vaccines are made using cell cultures. Influenza viruses are grown in cultured mammalian cells to make candidate vaccine viruses (CVVs), which are then provided to a vaccine manufacturer. The CVVs are inoculated into cultured mammalian cells allowing it replicate. This is then collected, purified, and deactivated to produce the flu vaccine.
Recombinant vaccines are made using the virus' genetic instructions for making the antigen, HA. This HA gene is then combined with a baculovirus, to help deliver the genetic instructions to make flu HA antigen into a host cell.
Developments in vaccine production.
Advances in genomics and proteomics have meant that several subunit vaccines have been produced which are better tolerated than inactivated or live-attenuated pathogens but are thought to be less immunogenic and require an adjuvant to achieve protective immune responses (Mbow et al, 2010). This is particularly important for populations such as children, elderly and the immunocompromised. These vaccines include adjuvants which are substances which are formulated as part of a vaccine to boost immune responses and enhance the vaccine's effectiveness (NIH, 2023).
Other approaches are that development of nucleic acid-based vaccines which work by inserting DNA or RNA that encode antigenic proteins into body cells which induces antigen presentation to the immune system triggering an immune response. The benefit of this research has led to the development of DNA and mRNA vaccines which deliver genetic information encoding tumour antigens (TAs) to the host leading to an immune responses against cancer cells that express the Tas (Jahanafrooz et al, 2020).
Certainly, nucleic acid vaccines elicit immune responses akin to live attenuated vaccines. Advantages of these vaccines include stimulation of both cell-mediated and humoral immunity, ease of design, rapid adaptability to changing pathogen strains seen during the COVID pandemic, stability, and protein expression and customizable multiantigen vaccines (Ho et al, 2021; Liu, 2019).
Work continues to improve vaccine immunogenicity and effectiveness (Atmar & Keitel, 2020). This is evidenced in some of the at-risk groups such as the elderly where high-dose and MF59-adjuvanted IIV enhance immunogenicity compared with the standard dose vaccine in person ≥65 years of age (Dunkle et al, 2017). The use of recombinant hemagglutinin has significantly improved immunogenicity compared in persons ≥50 years of age (Keitel et al, 2008).
The hope is that future vaccines may be also able to produce reactive antibodies that recognize epitopes, such as those in the hemagglutinin stem or the ectodomain of the M2 protein (M2e). The hope is that a vaccine with a with a long and broad spectrum of action will be developed. This could be effective as the extracellular domain of the M2 protein (M2e) of influenza A virus is a conservative region, and a possible target for a universal influenza vaccine (Mezhenskaya et al, 2019).
Other areas of research focus on the peptides from the virus nucleoprotein or the viral neuraminidase that would elicit a response with less risk of developing the disease or side effects (Atmar & Keitel, 2020).
Newer vaccines in the future may also use self-amplifying RNA (saRNA) which is derived from alphavirus expression vectors and is superior to non-replicating mRNA and DNA as the saRNA can provide very high expression levels and simultaneously induces strong innate responses (Ballesteros-Briones et al, 2020). Certainly, the use of mRNA has been shown to be more beneficial than subunit, killed and live attenuated virus, as well as DNA-based vaccines (Pardi et al, 2018). Hopefully in the future there may develop a universal influenza vaccine using ‘mRNA as a vector’ (Deviatkin et al, 2020).
GLOSSARY OF TERMS
- Glycosylation – this is the modification protein by the addition of a carbohydrate by an enzymatic reaction. This is used in vaccine production to help to induce an adaptive humoral immune response.
- Immunogenicity – the ability of the body's cells to provoke an immune response.
- Inactivated vaccines – contain whole bacteria or viruses which have been killed or have been altered, so that they cannot replicate.
- Live attenuated vaccines – contain whole bacteria or viruses which have been ‘weakened’ or attenuated so that they elicit a protective immune response but do not cause disease in healthy individuals.
- Recombinant vaccines – are made using organic material such as bacterial or yeast cells.
- Trivalent – Trivalent flu vaccines protect against three different influenza viruses.