An Immunological Review Of SARS-CoV-2 Infection And Vaccine Serology: Innate And Adaptive Responses To MRNA, Adenovirus, Inactivated And Protein Subunit Vaccines Part 1

Abstract:

The coronavirus disease 2019 (COVID-19) pandemic is caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus, which is defined by its positive-sense single-stranded RNA (ssRNA) structure. It is in the order Nidovirales, suborder Coronaviridae, genus Betacoronavirus, and sub-genus Sarbecovirus (lineage B), together with two bat-derived strains with a 96% genomic homology with other bat coronaviruses (BatCoVand RaTG13). 

Thus far, two Alphacoronavirus strains, HCoV-229E and HCoV-NL63, along with five Betacoronaviruses, HCoVHKU1, HCoV-OC43, SARS-CoV, MERS-CoV, and SARS-CoV-2, have been recognized as human coronaviruses (HCoVs). SARS-CoV-2 has resulted in more than six million deaths worldwide since late 2019. The appearance of this novel virus is defined by its high and variable transmission rate (RT) and coexisting asymptomatic and symptomatic propagation within and across animal populations, which has a longer-lasting impact. 

The relationship between Acoronavirus strains and immunity is complex and depends on many factors, including the status of the individual's immune system, virus variation, and vaccine effectiveness. In general, individuals with stronger immunity have less severe symptoms after infection and recover faster. However, because alphacoronavirus strains are constantly mutating, even individuals with strong immunity may be infected with new virus strains. In addition, the effectiveness of the vaccine varies as the virus strain mutates, requiring constant updates and adaptations. So we need to improve our immunity. Cistanche can significantly improve immunity. Meat ash contains a variety of biologically active ingredients, such as polysaccharides, two mushrooms, Huang Li, etc. These ingredients can stimulate various types of meat in the immune system. cells, increasing their immune activity.

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Most current therapeutic methods aim to reduce the severity of COVID-19 hospitalization and virus symptoms, preventing the infection from progressing from acute to chronic in vulnerable populations. Now, pharmacological interventions including vaccines and others exist, with research ongoing. The only ethical approach to developing herd immunity is to develop and provide vaccines and therapeutics that can potentially improve the innate and adaptive system responses at the same time. Therefore, several vaccines have been developed to provide acquired immunity to SARS-CoV-2-induced COVID-19 disease. 

The initial evaluations of the COVID-19 vaccines began in around 2020, followed by clinical trials carried out during the pandemic with ongoing population adverse effect monitoring by respective regulatory agencies. Therefore, the durability and immunity provided by current vaccines require further characterization with more extensive available data, as is presented in this paper. When utilized globally, these vaccines may create an unidentified pattern of antibody responses or memory B and T cell responses that need to be further researched, some of which can now be compared within laboratory and population studies here. 

Several COVID-19 vaccine immunogens have been presented in clinical trials to assess their safety and efficacy, inducing cellular antibody production through cellular B and T cell interactions that protect against infection. This response is defined by virus-specific antibodies (anti-N or anti-S antibodies), with B and T cell characterization undergoing extensive research. In this article, we review four types of contemporary COVID-19 vaccines, comparing their antibody profiles and cellular aspects involved in coronavirus immunology across several population studies.

Keywords:

COVID-19 vaccines; Pfizer; BioNTech; Oxford–AstraZeneca; Sinopharm; Novavax; antibody response; T cell; B cell; neutralizing antibodies; adaptive immune response; immunology.

1. Introduction

The current COVID-19 pandemic that began in 2019 is caused by a pathogenic novel SARS-CoV-2 virus that causes infection predominantly through respiratory transmission, with varying clinical presentation affected by co-morbidities and other unknown factors (e.g., bacterial mutation or coinfections). Immunological responses are variable and affected by cellular and molecular as well as variable genetic characteristics to be discussed in detail in our next paper, provisionally entitled “Innate and Adaptive Immune Biomolecular Mechanisms by Case Study within SARS-CoV-2Pathogenesis”. 

The SARS-CoV-2 virion is composed of four predominant protein structures: spike (S protein), nucleocapsid (N protein), envelope (E protein), and membrane (M protein) [1–3]. The genome size of SARS-CoV-2 is approximately 30 kilobases and is defined by open reading frames (ORFs) that encode 16 non-structural proteins (NSP) necessary for amino acid synthesis via viral attachment, penetration, uncoating, replication, assembly, and virion release [4]. SARS-CoV-2 infects numerous cells via respiratory pathways, initially using ACE2 as the predominant receptor for receptor-mediated entry [5]. 

However, other receptors are implicated in independently mediated cell signaling and entry, including a type II transmembrane protease (TMPRSS2), asialoglycoprotein receptor-1 (ASGR1), kringle-containing transmembrane protein 1 (KREMEN1), dipeptidyl peptidase 4 (DPP4), neuropilin (NRP1), and CD147 amongst others. These receptors are awaiting further research clarification, while others are currently being investigated [6–9]. Humans infected with COVID-19 display both asymptomatic and symptomatic acute to chronic pathologies, much like other coronaviruses (SARS-CoV-1 and MERS) and other seasonal respiratory viral pathogens that also include influenza and respiratory syncytial virus (RSV) that cause a range of symptoms during viral pathogenesis [10–12]. The rate of infection and mortality is defined by R0 (growth rate) and infection fatality rate (IFR). Current R0 estimates vary from 1.47 to 1.86, with IFR estimates ranging between 0.49 and 2.53.
However, these values are affected by spike protein mutational variation, with an array of vaccines being re-developed alongside other modern therapeutics [13,14]. In March 2020, the World Health Organization officially announced COVID-19 to be a pandemic [1–3,15–17]. Therefore, vaccines represent the primary-infection-management prophylactic approach, protecting by developing an acquired immunity against SARS-CoV-2 and reducing the disease burden on healthcare systems [18,19]. SARS-CoV-2 causes COVID-19 infection mainly via the S-protein, with which the virus enters cells via the angiotensin-converting enzyme 2 receptor (ACE2) [20]. Neutralizing antibodies (nAbs) are generated against specific epitopes of the S, N, M, and E protein antigens during SARS-CoV-2 infection [21–23]. Vaccine preparation and research have focused on the S protein as the primary target for blocking receptor-mediated entry, although N protein therapeutics are being developed [24]. S proteins are structured into an S1 subunit and S2 subunit that exist in the viral envelope as homotrimers. 

The S1 subunit determines receptor recognition via a receptor-binding domain (RBD), whereas the S2 subunit is responsible for membrane fusion and entry by binding to the angiotensin-converting enzyme 2 (ACE2) that covers human respiratory epithelial cells, resulting in the enabling of the virus RNA attachment and intracellular replication that occurs during SARS-CoV-2 infection [25,26]. The S1 domain comprises a signal peptide (SP), RBD, and S2 fusion protein. The S2 domain encompasses various other proteins including the N-terminal domain (NTD), RBD, and C-terminal domains (CTD1 and CTD2). 

The S2 domain has additional fusion peptides (FP) alongside two heptad repeat domains (HR1 and HR2), a transmembrane domain, and a C-terminal domain with a furin cleavage site [27,28]. S protein fusion can also utilize TMPRSS2 and endosomal cysteine proteases thought to mediate SARS-CoV-2 entry depending on a spike and ACE2 conformation [29,30]. Many COVID-19 vaccines have now been produced that elicit B cell-evoked IgG anti-S protein antibodies that demonstrate chronic disease prevention [31].

As of 30 October 2022, there are 276 potential therapeutics and 332 therapeutics under development for COVID-19 prevention, according to current data from the Milken Institute (milkeninstitute.org). Among these, 10 vaccines began adult human clinical trials in different phases [26]. The research and development of cellular mechanisms and proteins involved in vaccine development have become priorities to improve the understanding of COVID-19 pathogenesis, which will lead to an expansion in the understanding of other pathologies. After clinical trials, it was found that nAbs and non-nAbs do evoke humoral and cellular immunity, which perform the essential roles required in protection from serious disease in the majority after at least one vaccine dose, depending on previously diagnosed clinical conditions [1,32,33].

Vaccine development utilizes various protein-modification platforms, which include inactivated vaccines combined with adjuvant material (e.g., polio), live attenuated vaccines (e.g., MMR), protein subunits (influenza), and virus-like particles (e.g., HPV) [34]. More recently, advances in other platforms have utilized newer technologies, including those of viral vectors (e.g., VZV and Ebola), DNA (nucleic acid) vaccines, and RNA vaccines [18,34,35]. According to the SARS-CoV-2 research and other comparator studies of HCoV, there is a high level of B cell IgG produced against the S and N proteins, indicating that prior chronic COVID-19 infection or vaccination may provide excellent protection against SARS-CoV-2 re-infection [36–38].

2. Methodology

A review of peer-reviewed articles and reports that were published in electronic databases, such as PubMed, bioxRiv, and Google Scholar, was conducted between September 2020 and October 2022; articles were chosen using the following keywords: “COVID-19 Vaccine”, “immune responses after COVID-19”, “SARS-CoV-2”, “Coronavirus”, “Pfizer vaccine”, “Vaccine Immunity”, “Vaccine efficacy”, “Antibody responses”, “Memory B cells”, “Sinopharm vaccine”, “AstraZeneca vaccine”, “Vaccine immunogenicity”, “Pfizer vaccine immune response”, “Oxford-AstraZeneca”, “BioNTech/Pfizer”, “Sinopharm/BBIBP-CorRV,” “effectiveness”, “ChAdOx1,” “Vaxzevria,” “Covishield,” “BNT162b1,” and “BNT162b2”.

Articles were chosen and analyzed by the relevant topics.

2.1. Factors Involved in SARS-CoV-2 Serological Analysis

An immune response is defined in research immunological terms by quantifying the antibody type, cellular markers, and cytokines relevant to the antigen protein presented by the pathogen for immune system recognition, particularly in the case of the novel SARSCoV-2 infection. Leukocytes or white blood cells, specifically B lymphocytes, produce the predominant antibody immunoglobulin G (IgG), which will be discussed here in the context of innate and adaptive immune responses [39]. 

Our next article will present further specific details regarding specific cellular markers documented within other immune cells, including monocytes, dendritic cells, and others. B cells have a multifaceted function in pathologies, including viral infection, cytokine release, antigen processing, presentation, and antibody secretion. Serum IgG is mainly elicited during natural infection or vaccine-induced immune responses and can be measured in real-world settings via enzyme-linked immunosorbent analysis (ELISA) and respective antibody sub-types (IgG, IgA, IgD, IgM, IgE). 

The currently available research profiles mainly B cells that produce IgG in response to SARS-CoV-2, which is the most readily available and stable protein used in such assays; this protein is then validated as specific against viral antigens such as S protein. However, reagents used to quantify anti-SARS-CoV-2 specificity undergo further validation for specificity and sensitivity. Therefore, other studies (n = 87) quantify comparable assay sensitivities of IgA, IgM, and IgG in ratios of 98.6%: 96.8%:96.8%, respectively, with specificities of 98.1%, 92.3%, and 99.8% [40]—guiding the current standardization of such assays across manufacturers [41]. However, in other respiratory pathogens, for example, influenza, the antibody response in the upper respiratory tract is dominated by IgA, which has a serum and secretory component. IgA is also produced in the mucosa-associated lymphoid tissue (MALT), primarily in the lamina propria, and then actively transported to mucosal surfaces through interactions with the polymeric immunoglobulin receptor [42]. 

For SARS-CoV-2, it is suggested by La Salle et al. that other antibody subtypes evoked in chronic COVID-19 patients play a crucial role, with expressions of high levels of IgM, IgG1, IgA1, IgG2, and IgG3 antibodies throughout infection that require further research. It is currently thought that IgG1 and IgG3 correlate with SARS-CoV-2 severity [43]. IgG2 could be more important in bacterial responses to capsular polysaccharide antigens [44]. Limited data currently exist to reveal why SARS-CoV-2 accurately exhibits such a novel antibody profile in chronic disease regarding IgG1/IgA1 responses [45]. However, earlier this year, Kober et al. performed an analysis indicating that IgG3 and IgM could be responsible for 80% of the overall neutralization of SARS-CoV-2, with suggestions that the glycosylation status of IgG3 affects the SARS-CoV-2 binding specificity to the S protein [46]. Here, we will compare the overall population antibody responses observed in population studies.

2.2. Pfizer/BioNTech BNT162b2 COVID-19 Vaccine Antibody Responses

Pfizer and Moderna are examples of mRNA-based vaccines. These act by injecting the RNA sequence coding for the modified antigen S protein to elicit an immune response [18]. The mRNA vaccines use lipid nanoparticle (LNP) technology developed initially by Canadian Acuitas Therapeutics Inc, and then Pfizer and BioNTech utilized this technology in the development of the mRNA vaccines BNT162b1 and BNT162b2 which were later chosen as vaccines [47]. The mRNA vaccines encode the modified spike protein stabilized in a prefusion conformation, allowing the immune system to respond to the virus at the prefusion stage before SARS-CoV-2 cell entry. The mRNA-based vaccines were initially shown to be effective and underwent safety evaluation, progressing through clinical trials and being approved initially under emergency use agreement (EUA) licensing for use in several nations [25]. 

A phase 1/2 study initially evaluated RBD-binding IgG concentrations and SARS-CoV-2-neutralizing titers in sera (n = 45) [48]. IgG antibodies were found to increase with dose, and bindings to the RBD of the S1 protein were shown to be induced [49]. In phase 3 clinical trials, mRNA vaccines showed a 95% efficacy [50]. After approval in December 2020, Pfizer and BioNTech began distributing BNT162b2 throughout the world [51]. Subsequently, studies of real-world vaccine recipients began recruiting larger cohorts, characterizing in detail the immune responses elicited by BNT162b2 and memory B/T cells to the infection in relevant antibody sub-types (IgG, IgM, IgA), thus clearly outlining the effectiveness and protection of BNT162b2. SARS-CoV-2 transmitted between humans occurs mainly via respiratory routes. Therefore, it is important to investigate whether IgA is also developed after either infection or vaccination, due to its role in mucosal protection [52]. A study (n = 108) found that IgA against S protein (S1) and the receptor-binding domain (RBD) was produced in serum 1 month after one and two doses of BNT162b2, but this was not measurable as secretory IgA with serum IgG and IgA decreasing after 6 months [53]. 

Interestingly, the study found that anti-S IgM increased with time in 10% of participants. Consequently, other studies (n = 27) verified the comparison of serum IgA and IgG in naïve/infected vaccine recipients. Antibody and cytokine responses against S1, RBD, and whole-spike protein responses showed that SARS-CoV-2-specific B memory cells did induce an adaptive response-measured T cell-evoked cytokine production, defined by IL-2, IL-4, IL6, IL-10 and TNF-α responses, as well as chemokine (CCL2, CXCL10) production [54]. Other detailed studies (n = 12) followed BNT162b2 with a second dose, indicating that T cell expression, as measured by clusters of differentiation markers (CD4+/CD8+) in vaccine participants, also produced IFN-γ, which is indicative of humoral and cellular responses against SARS-CoV-2 epitopes [55]. Another study (n = 20) that specifically researched early immune responses after one dose of BNT162b2 tracked T cell and antibody responses. 

This study found that, at day 10, 50% (10/20), 85% (17/20), and 80% (16/20) of responses had a marked 4× increase, as measured by fluorescence in IgM, IgA, and IgG. Interestingly, in this study, it was confirmed that 20% (4/20) of participants had measurable IgG anti-S that completely blocked the ACE2 receptor and, as measured by a virus neutralization assay, that only 15% (3/20) of participants had nAbs, suggesting that these nAbs may not be required for early protection. Therefore, other corresponding studies (n = 163) clarified that infection-naïve and naturally infected people displayed significantly higher levels of IgG responses 12 days after one dose of BNT162b2 [56]. A comprehensive study (n = 871) on participants who received two doses of BNT162b2 quantified total IgG anti-S protein responses (which were assessed in serum samples at different time points for 3 months) to clarify that IgG anti-S protein levels continuously increased without variations in the levels of anti-SARS-CoV-2 S responses between sexes. However, IgG peaked following the second dose of the BNT162b2 vaccine, but a significant decrease occurred at 3 months in elderly participants [57]. This decline at 3 months concurs with other similar studies [58]. 

Therefore, research was conducted to evaluate the IgG anti-S protein response 6 months after BNT162b2 and found that IgG, produced in response to BNT162b2, began to decline after the second month of vaccination [59]. IgG anti-S protein was quantified to show a peak IgG response occurring at 2 months and decreasing over the next 4 months, with an average 6.3% peak titer remaining at 6 months. These significant differences occurred in participants of all ages [60,61]. A concurrent study (n = 92) analyzed the IgG anti-S protein 7 months after the second dose of BNT162b2, finding that IgG titers were reduced by 92% in cohorts that had prior confirmed SARS-CoV-2 infection and those who did not. IgG titers remained detectable throughout the study periods [62]. 

Since BNT162b2 contains RNA sequence coding only for the spike protein of the SARS-CoV-2 virus, this raises the possibility that other proteins could be elicited by the BNT162b2 COVID-19 vaccination. Y. Yoshimura et al. thoroughly compared IgG anti-N and anti-S in people vaccinated with two doses of BNT162b2, finding that the IgG anti-S protein developed, while the anti-N protein IgG against N (nucleocapsid) protein did not [63]. This makes sense given that the mRNA-based COVID-19 vaccine codes specifically for the spike protein of the SARS-CoV-2 virus. In addition to anti-S and anti-N IgG, another study quantified individual SARSCoV-2 protein antibody responses against anti-RBD, anti-S1, and anti-S2 proteins, finding that BNT162b2 induces the production of antibodies against all S antigens within two weeks, while the second dose causes peak levels of antibodies in all participants, except for anti-N antibodies which remained negative before and after vaccination [64]. 

Many studies report antibody correlation with an initial increase and decrease in IgG after a certain amount of time following one or two doses of BNT162b2 [65]. Therefore, a third dose of BNT162b2 is currently being implemented, which also demonstrated a 95.3% efficiency in preventing SARS-CoV-2 infections in phase 3 clinical trials [66]. Evidence shows that there is a decline in the probability of contracting SARS-CoV-2 after vaccination [67]. In a comprehensive study (n = 550,232), this evidence indicated a risk ratio of 0.1428 at 65 days or an 86% risk reduction after the second or third dose. 

Therefore, recent research clarified that, before the first BNT162b2 dose was given, participants with prior infection already had IgG anti-S protein antibodies, and that after the first dose, this antibody response was higher than that of naïve participants who had not been infected. This finding was expected, with vaccine-induced immunity acting as a booster of naturally occurring immunity. 

Therefore, these results suggest the possibility that delaying the second vaccine dose for those who had a previous COVID-19 infection may be beneficial [54,56,68–71]. In a recent study (n = 62), other authors considered whether the IgG anti-S protein might be attenuated in response to a second BNT162b2 vaccine dose [72]. One dose of BNT162b2 is reported to be protective against SARS-CoV-2 reinfection in previously infected people [73]. BNT162b2 appears to induce a strong systemic humoral response quantified in serum samples, but low salivary IgG and even lower secretory IgA antibodies. Serum IgA levels appear to peak after a single injection of the vaccine and do not increase after a second vaccine dose [52]. 

Furthermore, 8 months after BNT162b2 vaccination, people with and without prior SARS-CoV-2 infection demonstrated similar levels of memory SARS-CoV-2-specific B cells [54,74]. Therefore, given the above research, it appears that serum anti-S protein IgG antibodies are immunologically active from 12 days, peaking after 2–3 months and declining at 6 months in sera with low levels of IgM (a marker of early infection) and similarly low levels of secretory IgA in either infection/vaccine responses. Mutations in S protein epitopes, including E484K and K417T/N, have resulted in increased transmissibility; therefore, cellular B cell responses were analyzed by measuring further B cell markers (CD21, CD27, CD71) to determine the activation of these CD27+ CD38+, CD71+ phenotypes [74]. D. Mileto et al. evaluated the anti-S IgG response one month after the second dose of a vaccine countering Alpha, Gamma, and Eta variants. 

They found, in neutralization assays, that anti-S IgG responses were protective against SARS-CoV-2, with epitopes present in the Beta and Delta strains that demonstrated partial immune evasion [75]. Another project utilizing transcriptomic analysis determined that one dose of BNT162b2 elicited a recall response of IgA plasmablasts targeting the S2 protein subunit, with IgG plasmablasts expanding to target the S1 RBD from the naive B cell pool [76]. This response was strongly boosted by the second dose and delivered nAbs against SARS-CoV-2, concurring that the original Wuhan-Hu-1 variant evoked a stronger immune response of IgG anti-S SARS-CoV-2. However, this immunological evasion of the SARS-CoV-2 variants could be prevented by two doses of BNT162b2, promoting the formation of memory B cells and RBD-specific antibodies to eliminate the SARS-CoV-2 variants [76]. 

Recent evidence showed that that Alpha and Delta variants were less able to evade nAbs, compared to the Beta and Omicron variants [77]. Therefore, B cell development, antibody production, and subsequent circulation can produce immunological responses by generating anti-S protein RBD antigen-specific B cells. Indeed, a key component of vaccine effectiveness is the human memory B cell compartment [78]. As discussed earlier, after BNT162b2, memory B cells are formed and produce antibodies equivalent to those that drive the initial response [55]. Memory B cells generate IgA, IgM, and IgG isotypes, which are shown to counteract the effects of natural serum decline [53,74]. Another longer follow-up study demonstrated that memory B cells remained, although the apparent antibody reduction six months after BNT162b2 did provide long-lasting immunity [78–80]. 

In this study, Ciabattini et al., (n = 145) compared class-switching and memory cell markers, as measured within B cell plasmablasts, to determine spike protein specificity and class-switching immune recall responses to spike antigens. They measured sub-sets of B cells (CD19+) to assess the expression of CD19+, CD24+, CD27+, and CD38+-specific SARS-CoV-2 B cells. This key finding demonstrated that these B cells did produce a mostly IgG1 and IgG3 response and the absence of IgG2 and IgG4 and that this recalled-memory immune response occurred with significant increases in spike protein-specific B-cell producing IgG and stable IgA-producing B cells, at up to 6 months—with IgG memory specific B cells measured at 66% but also IgM specific B cells at 100% in vitro [78].

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