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Biological and immune responses to current anti-SARS- CoV-2 mRNA vaccines beyond anti-Spike antibody production

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Several vaccine strategies are now available to fight current SARS-CoV-2 pandemic. Those based on the administration of lipid-complexed messenger(m)RNA molecules represent the last frontiers in terms of technology innovation. mRNA molecules coding for the SARS-CoV-2 Spike protein are intramuscularly injected thereby entering cells by virtue of their encapsulation into synthetic lipid nanovesicles. mRNA-targeted cells express the Spike protein on their plasma membrane in a way that it can be sensed by the immune system, which reacts generating anti-Spike antibodies. Although this class of vaccines appears as the most effective against SARS-CoV-2 infection and disease, their safety and efficiency are challenged by several factors included, but not limited to: emersion of viral variants, lack of adequate pharmacokinetics/pharmacodynamics studies, inability to protect oral mucosa from infection, and antibody waning. Emersion of viral variants represents an expected consequence of mass vaccination carried out in a pandemic time using sub-optimal vaccines against an RNA virus. On the other hand, understanding remainder flaws could be of some help in designing next generation anti-SARS-CoV-2 vaccines. In this commentary, issues regarding the fate of injected mRNA, the tissue distribution of the induced antiviral antibodies, and the generation of memory B cells, are discussed. Careful evaluation of both experimental and clinical observations on these key aspects should be taken into account before planning third-dose administrations, vaccinations to non-at risk population, and social restrictions.
Biological and immune responses to current anti-SARS-CoV-2 mRNA vaccines beyond anti-Spike
antibody production
Maurizio Federico
National Center for Global Health, Istituto Superiore di Sanità, Viale Regina Elena, 299,
00161, Rome, Italy
* Correspondence to:
Dr. Maurizio Federico,
National Center for Global Health, Istituto
Superiore di Sanità, Viale Regina Elena, 299,
00161 Rome, Italy
Phone: +39-06-4990-6016
Fax: +39-06-49903210
Abbreviations: ACE-2, angiotensin-converting enzyme 2; BRMC, resident B memory
cells; COVID-19, coronavirus disease 2019; Ig, immunoglobulin; LINE, long interspersed
nuclear element; MBC, memory B cells; mRNA, messenger RNA; RBD, receptor-binding
domain; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2; VoC, variant of
Several vaccine strategies are now available to fight the current SARS-CoV-2 pandemic. Those based
on the administration of lipid-complexed messenger(m)RNA molecules represent the last frontiers in
terms of technology innovation. mRNA molecules coding for the SARS-CoV-2 Spike protein are
intramuscularly injected, thereby entering cells by virtue of their encapsulation into synthetic lipid
nanovesicles. mRNA-targeted cells express the Spike protein on their plasma membrane in a way that it
can be sensed by the immune system, which reacts generating anti-Spike antibodies. Although this class
of vaccines appears as the most effective against SARS-CoV-2 infection and disease, their safety and
efficiency are challenged by several factors included, but not limited to: emergence of viral variants, lack
of adequate pharmacokinetics/pharmacodynamics studies, inability to protect oral mucosa from infection,
and antibody waning. Emergence of viral variants can be a consequence of mass vaccination carried out
in a pandemic time using suboptimal vaccines against an RNA virus. On the other hand, understanding
the remainder flaws could be of some help in designing next generation anti-SARS-CoV-2 vaccines. In
this commentary, issues regarding the fate of injected mRNA, the tissue distribution of the induced
antiviral antibodies, and the generation of memory B cells are discussed. Careful evaluation of both
experimental and clinical observations on these key aspects should be taken into account before planning
booster administration, vaccination to non-at-risk population, and social restrictions.
Keywords: mRNA vaccine; SARS-CoV-2; antibody waning; B memory cells; lung immunity
For many years, the use of RNA for therapeutics and vaccines was disregarded due to the supposed
difficulties to be manipulated. The achievement of several technology improvements contributed to put this
technique in the spotlight for its pharmaceutical use. These advancements included the use of base
analogues, addition of a cap at the 5’ end, optimization of codon usage, and inclusion of untranslated
elements at both 5’ and 3’ ends facilitating ribosome recognizing [1].
Starting from November 2011, the Biological Technology Office of the US Defense Department’s
Defense Advanced Research Project Agency invested large funding in several RNA-based vaccine
programs. CureVac and Moderna companies were the first recipients of such financial supports. In the
following years, biotech and Synthetic Genomics companies were funded by both public and private
agencies, including NIH, Gates Foundation, Johnson & Johnson, Bayer, and Genentech to develop similar
programs [2]. The disclosed investment of about 1 billion of dollars provided within a few years was the
best witness of the enormous interest on the application of RNA vaccine technology to fight both tumors
and infectious diseases.
Both preclinical and clinical data have proven both the feasibility and potentiality of the mRNA-
based vaccine platform. Melanoma, triple-negative breast cancer, prostate cancer, and lung cancer were
among the tumor diseases challenged by experimental mRNA vaccines in clinical trials. On the other hand,
the same technique was evaluated against several viral diseases [2]. The anticipated success of this method
in delivering any mRNA sequence into the cells represented the starting point towards the design of current
mRNA-based anti-SARS-CoV-2 vaccines.
These vaccines are composed by “in vitro” synthesized mRNA molecules coding for full-length
SARS-CoV-2 Spike glycoproteins from the ancestral strain (i.e., Wuhan isolate) in a prefusion
conformation. The stabilization of the prefusion conformation is ensured by two consecutive proline
substitutions at amino acid positions 986 and 987, at the top of the central helix of the S2 subunit. The
mRNA molecules are complexed with lipids in a way that they are allowed to enter cells efficiently. In
such a formulation, the mRNA molecules are expected to pass the plasma membrane of any kind of cell,
thereby becoming available for translation by the cell cytoplasmic machinery. In humans, injections are
usually carried out intramuscularly (i.m.) in the deltoid. Neo-synthesized, full-length Spike proteins are
anticipated to remain anchored to the host cell membrane in trimeric complexes, allowing the immune
system to initiate the mechanisms leading to the development of anti-Spike adaptive immunity. It is
supposed that local inflammation generated by co-injected substances and/or cellular responses to Spike
expression may account for co-stimulation needed for an effective immune response.
Here, issues regarding pharmacokinetics/pharmacodynamics of mRNA vaccines, induced mucosal
immunity, and antibody waning are analyzed. Careful evaluation of the weaknesses of current mRNA
vaccines would be helpful for the generation of improved anti-SARS-CoV-2 immunogens.
Still unresolved aspects on pharmacokinetics and pharmacodynamics of mRNA-based vaccines
Several aspects regarding how the organism affects the fate of mRNA vaccines (pharmacokinetics),
as well as how they influence the host physiology (pharmacodynamics) deserve thorough evaluation.
Concerning the fate of mRNA vaccines upon i.m. injection, their formulation implies that virtually
any kind of cell can internalize the lipid-mRNA complexes. Clearly, i.m. inoculation favors the delivery of
mRNA molecules into muscle cells. However, in view of the strong bioactivity of both mRNA and its
translation products, monitoring possible additional sites of vaccine accumulation upon injection is of
major relevance.
On this subject, results from a very accurate study carried out in cynomolgus macaques were
published a few months before the pandemic outbreak [3]. It was clearly demonstrated that, upon vaccine
injection, both muscle cells and diverse types of immune cells express the protein coded by the mRNA.
The authors reported that, 4 hours after injection, mRNA molecules are internalized by immune cells at
both the site of injection and proximal lymph nodes, in amounts appearing inversely proportional to the
distance from the point of injection. After 28 hours, the levels of vaccine mRNA increased in lymph nodes,
while decreasing in the injection site. Monocytes were found to be the immune cells most efficiently
internalizing vaccine mRNA in both muscle tissues and lymph nodes. The latter ones were also found
abundantly infiltrated by both B lymphocytes and dendritic cells expressing the mRNA vaccine.
A relevant message from this study is that mRNA does not localize at the inoculation site only,
however spreading to proximal lymph nodes as early as 28 hours post injection. Considering the observed
rapidity of diffusion, the authors concluded that the spreading can be consequence of vaccine diffusion
rather than cell migration, even if it is also conceivable that mRNA internalizing immune cells may reach
lymph nodes through lymphatic circulation. When translated to current anti-SARS-CoV-2 mRNA vaccines,
a direct consequence of mRNA internalization in immune cells can be that large amounts of the Spike
protein would be persistently expressed in several districts of the body. The fate of SARS-CoV-2 spike
protein expressed by immune cells is essentially unknown. Hence, it is quite difficult to establish whether
free circulation of Spike-expressing immune cells can be beneficial for the antiviral adaptive immune
response. Conversely, membrane-associated Spike proteins can interact with cells expressing the ACE-2
receptor. This binding can perturb the functions of endothelial cells by inhibiting vital mitochondrial
functions, leading to downstream endothelium pathology [4].
Vaccine mRNA molecules comprise 1-methyl-pseudouridine in place of uridine. The rationale for
this change relied on the observation that, after intravenous (i.v.) injection, decreased innate immune
sensing together with increased mRNA stability and translation efficiency compared to unmodified mRNA
have been observed [5]. Not consistently, more recent work emphasized that 1-methyl-pseudouridine
modification of mRNA had no significant effect on both protein expression “in vivo” and mRNA
immunogenicity compared to unmodified mRNA when it was delivered systemically. In particular, we
observed transient extracellular innate immune responses to modified mRNA included neutrophilia,
myeloid cell activation, and upregulation of four serum cytokines, namely, CCL2, CCL5, CXCR9, and G-
CSF [6].
After cell internalization, vaccine mRNA is expected to be translated by the cellular machinery until
its intracellular degradation. Intriguingly enough, however, data from a recent study highlighted that sub-
genomic parts of SARS-CoV-2 RNA can integrate into DNA of human cells and patient-derived tissues
[7]. Integrated SARS-CoV-2-related DNA sequences mostly originated from the 3’ end of the viral
genome, which encompasses Spike-related sequences. Integrations of retrotranscribed SARS-CoV-2
sequences have been detected mostly in exon regions, and correlated with the activity of the ubiquitous
LINE-1 retrotransposon. Even if these findings have been questioned [8], implementing detailed
genotoxicity studies in vaccinated subjects should be strongly recommended.
Once translated, vaccine-derived SARS-CoV-2 Spike is supposed to be embedded into the host cell
membrane. However, based on the observations made with many other virus species, it is more than likely
that at least part of the neo-synthesized protein sheds and circulates into the body. Free circulating Spike
protein might bind ACE-2-expressing cells, thereby inducing cell activation and damage [4], whose overall
consequences depend on the levels of ACE-2 expression on the targeted tissues. When it occurs in blood
vessels or heart tissues, consequent cell damage and inflammation can lead to vasculitis, pericarditis, and
myocarditis [9-11]
Intracellular expression of vaccine mRNA leads to both vigorous and reproducible antibody
response. In this context, possible effects induced by anti-idiotype antibodies should be taken in
consideration, especially in recovered patients undergoing vaccination. Here, the amino acid sequences
(referred to as idiotopes) in antigen-binding domains of anti-Spike antibodies can be immunogenic as
consequence of the very high antibody levels induced. In this way, antibodies against these sequences (anti-
idiotype antibodies) can be induced, and, as consequence of a molecular mimicry, part of them can bind the
ligand of Spike protein, i.e., ACE-2 [12]. Thereby, physiologic functions of ACE-2 could be disturbed, for
instance by blocking natural ligands or abnormally stimulating the receptor. Also, ACE-2 expressing cells
binding anti-idiotype antibodies may undergo cell lysis through mechanisms mediated by the action of
complement and/or immune cells. Notably, all these undesirable effects have the potential to occur after the
disappearance of the Spike antigen. However, no evidences on the generation of anti-idiotype antibodies in
SARS-CoV-2 vaccinees have been produced so far.
In conclusion, several issues regarding both pharmacokinetics and pharmacodynamics of vaccine
mRNA remain open. Deep investigations on them are mandatory to anticipate and, in this case, find
countermeasures against both mid- and long-term side effects.
Anti-Spike antibodies in oral mucosa
Subjects injected with current anti-SARS-CoV-2 vaccines develop anti-Spike antibodies with
different specificities, including anti-receptor binding domain (RBD), anti-N-terminal domain (NTD), and
anti-S2 antibodies. Most part of neutralizing antibodies binds the RBD.
Among all immunoglobulin (Ig) classes, secretory IgAs are the most effective ones in protecting
epithelial cells of mucosal surfaces from the attack of respiratory viruses [13]. In SARS-CoV-2 infected
patients, the virus-neutralizing potency of IgAs was found superior compared to that of virus-specific IgGs
detectable in both serum and saliva [14]. It was calculated that, upon infection, the levels of anti-RBD IgGs
are about five times higher than those of anti-RBD IgA, however, being seven times less efficient in virus
neutralization assays. Moreover, dimeric anti-RBD IgAs from oral/lung mucosa were found more potent
than the monomeric counterpart detectable in serum. Unfortunately enough, the levels of RBD-specific IgA
decay much more rapidly than IgGs [14].
Immunity to the oral mucosa is mandatory for any vaccine conceived to impede the diffusion of
virus spreading through the oral routes. In consequence, evaluating the levels of virus-neutralizing IgA
induced in the oral mucosa by anti-SARS-CoV-2 vaccines attracted the interest of many scientists with the
intent to predict the vaccine effectiveness in limiting viral spread within humans.
Data published by Planas and colleagues demonstrated absence of neutralization activity in nasal
swabs until 2 weeks after the second injection of mRNA vaccine despite good levels of binding activity
(presumably due to the co-existence of anti-Spike IgGs), and in the presence of high titers of both binding
and neutralizing antibodies in sera [15]. More recently, another paper reported the lack of anti-Spike IgA
until three weeks after the second jab in the saliva of 43 health care workers producing very high levels of
antiSpike serum IgAs upon vaccination [16]. Furthermore, Roltgen and colleagues found minimal amounts
of either anti-Spike -RBD and –N-terminal domain antibody IgAs in both serum and oral mucosa of
vaccinated subjects seven weeks after the second injection [17]. In another couple of studies, a modest
neutralizing capacity has been observed in saliva of mRNA vaccinees [18, 19].
Taken together, these data indicate that, differently from natural infection [20, 21], mRNA-
dependent anti-SARS-CoV-2 vaccination seems unable to induce levels of oral immunity adequate to
protect vaccinees from replicating and transmitting infecting viruses. The obvious consequence, as also
supported by real-world evidences, is that also vaccinees can be infected by SARS-CoV-2. Furthermore,
the very recent demonstrations that the levels of viral replication in the mucosa of vaccinated and
unvaccinated subjects are similar [22, 23] support the idea that vaccine-induced immunity is not able to
block virus transmission.
Anti-Spike antibody waning and B memory cells
Antibody waning is a distinctive feature of the immune response against infection with SARS-CoV-2
as well as many other respiratory viruses. Unfortunately, it was reported that in COVID-19 patients
neutralizing antibodies have the most rapid decay kinetics among the different functional families of anti-
Spike antibodies [24]. Accordingly, results from several groups are consistent with the idea that also
vaccine-induced antibodies have a limited half-life. However, antibody waning would not be a major issue
in case the vaccine would be able to generate a well-established B memory activity prompt to react as soon
as the infecting virus is encountered. Despite a limited number of studies suggesting that mRNA vaccines
can generate anti-Spike memory B cells (MBCs), however, their absolute number and, most importantly,
tissue distribution pose relevant questions.
Studies on SARS-CoV-2-specific MBCs induced by vaccines are sparse and refer uniquely to cells
isolated from peripheral blood. Goel and colleagues measured levels of RBD-specific MBCs (i.e., B cells
having the potential to secrete neutralizing antibodies) 1 week after the second injection. By flow
cytometric analysis, they found a mean of 0.1 % of specific cells over the total of MBCs, which comprised
0.3 % of Spike-specific MBCs for the most part expressing IgG [25]. Another study compared the number
of MBCs specific for RBDs from the ancestral virus isolate with that specific for RBDs of a number of
variants of concern (VoCs) two weeks after the second vaccine injection [26]. After three-day “in vitro”
stimulation of PBMCs with human recombinant IL-2 and R848, a maximum of 0.015% specific over the
total of cells was detected by B cell EliSpot analysis, with reduced percentages in the case of RBDs from
VOCs. Mortari and colleagues found a peak of RBD positive MBCs 7 days after the second injection,
slightly decreasing 62 days thereafter. At this time, flow cytometry analysis estimated the percentages of
RBD-specific MBCs around 0.01% of total CD19+ CD24+ CD27+ CD38- cells, in the presence of 15% of
MBCs showing high affinity to trimeric Spike [16]. Sokal and colleagues reported barely detectable RBD-
specific MBCs within PBMCs of double vaccinated subjects 2 months after boosting [27].
Taken together, these data seem not sufficient to establish whether current mRNA vaccines can
generate an adequate, durable, and reactivatable humoral response after antibody waning. Most
importantly, in view of the strict compartmentalization of the lung immune system, additional studies are
needed to evaluate the immune memory at the level of both high and low respiratory compartments.
In fact, the development of lung immune memory is largely not influenced by events occurring in
both the peripheral circulation and lymphoid organs. In many instances, lymphocytes in the lungs are
maintained independently of the pool of circulating lymphocytes, and their continuous loss through
intraepithelial migration towards the airways is constantly replenished by homeostatic proliferation.
Recently, a seminal study on a murine model susceptible to influenza virus infection demonstrated the
presence of lung-resident memory B cells (BRMCs) as the major memory effectors of humoral antiviral
immunity [28]. BRMCs are phenotypically distinguishable from MBCs from lymphoid tissues by virtue of
high CXCR3 levels and absence of CD62L. Formation of BRMCs requires encounter with antigens in the
lung, where they can differentiate. Antigen-specific BRMCs are maintained in the lung and do not
recirculate and respond to infection/re-infection very rapidly. All these key observations have been very
recently confirmed in mice infected with Streptococcus Pneumoniae [29]. Of major interest, data from the
same paper demonstrated the presence of B memory cells showing the BRMCs phenotype in human lungs.
Consistently, Weisel and colleagues identified tissue-resident BRMCs in the human gut [30].
Blocking virus replication in the oral mucosa represents a key step in the fight against SARS-CoV-2
spread. Optimal levels of both humoral and cellular immunity should be achieved locally at the viral port of
entry, rather than peripherally. Based on the here above summarized observations, it is more than desirable
that new antibody-based anti-SARS-CoV-2 vaccines designed to induce effective and long-lived protection
would be delivered to the respiratory tract. This argument is strengthened also by the evidence that mucosal
anti-SARS-CoV-2 dimeric IgAs are several fold more potent that the serum-derived monomeric
counterpart [14, 31]. In addition, oral delivery of new immunogens/vaccines has the potential to generate
very limited systemic side effects even after repeated administrations.
Taken together, these observations strongly support the idea that orally administrated vaccines would
represent a relevant amelioration compared to current ones, which generate an immunity not strong enough
to impede virus replication in the oral mucosa and transmission [32].
Possible consequences of repeated vaccinations using the same immunogen
Systemic administration of vaccines against respiratory viruses often associates with unsatisfactory
outcomes. For instance, the efficacy of seasonal anti-influenza vaccines rarely overcomes 50% of
The immunological correlates of protection against SARS-CoV-2 infection are still unknown, where
the term “correlate of protection” refers to a laboratory parameter associated with protection from a clinical
disease [33]. A coordinated action of CD4+ T cells, CD8+ T cells, and neutralizing antibodies seems
necessary to control SARS-CoV-2 infection. In this scenario, neutralizing antibodies certainly play a key
role in protecting from infection. However, in the case of current vaccines, which exclusively rely on
humoral immunity, antibody waning and uncertain BMC effectiveness in the lung tissue represent not
easily surmountable limitations. It is not obvious to predict whether additional vaccine inoculations could
improve the quality, intra-tissue distribution, and relative duration of the immune response. It has been
observed that repeated antigen encountering can select for plasmablasts/plasmacells producing antibodies
with increasing affinity, as also demonstrated for SARS-CoV-2 infections [34-36]. However, one should
consider that this phenomenon could occur also for non-neutralizing antibodies, some of which might have
pathogenic effects.
Virus replication in the context of suboptimal antiviral action of vaccine-induced antibodies can lead
to emergence of resistant virus quasispecies. In the SARS-CoV-2 case, this process could have contributed
to the selection of VoCs, whose rapid emergence paralleled mass vaccination. This phenomenon can affect
the anticipated outcomes from additional vaccine cycles. In fact, it is well known that repeated vaccinations
against pathogen evolving mutants like SARS-CoV-2 have the risk to meet with the phenomenon referred
to as “original antigenic sin” [37]. In detail, the humoral immune response elicited against
immunodominant epitopes of the first pathogen generates a sort of “immune imprinting” in a way that the
response against subsequent infection with a mutated form of the pathogen cannot be able to recognize new
emerging immunodominant epitopes. As a result, either reduced or no protection against the new
pathogenic strain can be generated (Fig. 1).
Figure 1. The “original antigenic sin”. When the body first encounters a pathogen, it produces effective
antibodies against its dominant antigen and thus eliminates the pathogen. Selective pressure can originate
pathogens with new dominant antigens, with the original antigens now being recessive. In this case, the
immune system still produces the former antibodies against the old “now recessive antigen”, and develops
antibodies against the new dominant one scarcely. The results are the production of ineffective antibodies
and generation of weak immunity.
Intriguingly enough, very recently it has been reported that a third-dose booster with an Omicron-
based mRNA vaccine on macaques previously treated with two doses of mRNA-1273 vaccine (based on
the ancestral Spike sequence) did not offer advantages in terms of protection against infection with the
Omicron VoC respect to the homologous booster [38]. Consistent with the “antigenic original sin” theory,
it is conceivable that the immune system previously educated by the two mRNA-1273 vaccine doses
reacted to the Omicron-based third dose still producing Abs mainly directed to the ancestral S protein.
Current mRNA-based anti-SARS-CoV-2 vaccines have provided protection. However, antibody
waning, unsatisfactory mucosal immunity, and the “antigenic original sin” mechanisms should be
adequately evaluated at the time of decision to proceed towards additional cycles of vaccination. In fact,
challenging an emerging VoC (as in the case of the widespread Omicron variants) with repeated injections
of vaccines designed for a virtually disappeared virus quasispecies may be not the best strategy, as also
suggested by the strong decrease of vaccine effectiveness calculated after the fourth dose (Tab. 1) [39, 40].
On the other hand, the likelihood of side effect occurrence increases with the number of injections.
For all these reasons, repeated mass vaccination of non-at-risk populations, including infants and
adolescents, asks for much more extended and careful evaluation of both mid- and long-term risks.
No specific grants from funding agencies in the public, commercial, or not-for-profit sectors pertain
to the present manuscript.
Conflicts of Interest: The author declares that he has no conflicts of interest.
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Objectives A clear temporal relationship between myocarditis and pericarditis after COVID-19 vaccination has led to the belief that the vaccine may act as a trigger for these cardiologic complications. The aim of this systematic review is to explore the incidence, clinical presentation, management, and association between them. Methods We conducted a systematic literature search on Cochrane, MEDLINE, and EMBASE as per guidelines of PRISMA (Preferred Reporting Items for Systematic Reviews). A total of 41 case reports and case series describing 97 patients, and 5 original articles describing 15,585,309 participants were selected as part of this review. Results Of the 97 reported cases describing vaccine-associated myocarditis/pericarditis, 67 (69%) patients received Pfizer-BioNTech and 25 (25.7%) received Moderna. The mean onset of symptoms after vaccine administration was 3.8 ± 4.5 days with three-quarters developing symptoms after the second dose. Chest pain (n = 88, 90%) and fever (n = 33, 34%) were the most common presenting complaints. Out of 97, 80 (82.5%) patients recovered while 4 (4.1%) patients expired. The pooled incidence of myocarditis and pericarditis extrapolated from original studies is 0.001% and 0.0004%, respectively. In the original studies, nearly all the cases of myocarditis and pericarditis were mild. Chest pain and fever were the most common presenting symptoms. Conclusion Myocarditis and pericarditis after the COVID-19 vaccine have been reported more in young adult males and are most likely to occur after the second dose of mRNA vaccines. The presentation is mild and the majority of the patients recover either completely or partially.
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Mucosal immunity plays a key role in prevention of SARS-CoV-2 virus spread to the lungs. In this study, we evaluated systemic and mucosal immune signatures in asymptomatic SARS-CoV-2–infected versus symptomatic COVID-19 adults compared with RSV-infected adults. Matched serum and nasal wash pairs were subjected to cytokine/chemokine analyses and comprehensive antibody profiling including epitope repertoire analyses, antibody kinetics to SARS-CoV-2 prefusion spike and spike RBD mutants, and neutralization of SARS-CoV-2 variants of concern. The data suggest independent evolution of antibody responses in the mucosal sites as reflected in differential IgM/IgG/IgA epitope repertoire compared with serum. Antibody affinity against SARS-CoV-2 prefusion spike for both serum and nasal washes was significantly higher in asymptomatic adults compared with symptomatic COVID-19 patients. Last, the cytokine/chemokine responses in the nasal washes were more robust than in serum. These data underscore the importance of evaluating mucosal immune responses for better therapeutics and vaccines against SARS-CoV-2. Copyright © 2021 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. Distributed under a Creative Commons Attribution NonCommercial License 4.0 (CC BY-NC).
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We found no significant difference in cycle threshold values between vaccinated and unvaccinated, asymptomatic and symptomatic groups infected with SARS-CoV-2 Delta. Given the substantial proportion of asymptomatic vaccine breakthrough cases with high viral levels, interventions, including masking and testing, should be considered for all in settings with elevated COVID-19 transmission.
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Background Cases of immune complex vasculitis have been reported following COVID-19 infections; so far none in association with novel mRNA-based COVID-19 vaccination. This case report describes a cutaneous immune complex vasculitis after vaccination with BNT162b2. Case presentation A 76-year old male with liver cirrhosis developed an immune complex vasculitis 12 days after the second injection of BNT162b2. On physical examination, the patient presented with pruritic purpuric macules on hands and feet, flexor and extensor parts of both legs and thighs and lower abdomen, and bloody diarrhoea. Laboratory testing showed elevated inflammatory markers. After short treatment with oral steroids all clinical manifestations and laboratory findings resolved. Conclusions An increasing number of clinical manifestations have been attributed to COVID-19 infection and vaccination. This is the first written report of immune complex vasculitis after vaccination with BNT162b2. We present our case report and a discussion in the light of type three hypersensitivity reaction.
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The first line of defense against SARS-CoV-2 is the upper respiratory tract, yet we know little about the amount, type, and kinetics of mucosal anti-Spike antibodies (Ab) in response to intramuscular (i.m.) COVID-19 vaccination. We analyzed salivary Ab against SARS-CoV-2 Spike following mRNA/mRNA and adenovirus (Ad)/mRNA regimes. While anti-Spike/RBD IgG was detected in the saliva and correlated with the systemic response, anti-Spike/RBD IgA associated with the secretory component (sIgA) was also detected, and did not necessarily correlate with serum Ab. Only modest levels of neutralizing capacity were observed in saliva at 2 weeks post-dose 2, and by 6 months, anti-Spike/RBD IgG were greatly diminished. In contrast, low levels of anti-Spike sIgA persisted up to 6 months after dose 2. Our results show that SARS-CoV-2 vaccination induces an IgG response in the saliva that decays over time and an sIgA response that does not necessarily correlate with systemic immunity. One-Sentence Summary Our study delves into how intra-muscular mRNA/mRNA or mRNA/Ad COVID-19 vaccination regimes confer immunity in the oral cavity with important implications for understanding protection against breakthrough infections in healthy vaccinated people.
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There is interest in understanding the mechanisms that underlie reports that patients infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) remain PCR positive many weeks after initial infection. The recent paper by Zhang et al. (1) suggests a potential explanation of this phenomenon by claiming that SARS-CoV-2 RNA can integrate into the genome of infected human cells. The authors also reanalyze RNA se-quencing (RNA-seq) data and report that SARS-CoV-2− host chimeric reads are present in cells and patient tissues. Given the potential implications of this research on the long-term impacts of COVID-19, we feel that it's necessary to scrutinize the evidence presented. To determine whether SARS-CoV-2 RNA might be retrotranscribed and integrated into the genome, the authors conducted a proof-of-principle experiment where human lung cells (Calu3) and kidney cells overexpress-ing class I transposable elements and wild type (HEK293T-L1/HEK293T) were infected with SARS-CoV-2 and subjected to high-throughput DNA sequencing (1). The very low frequency of identified chimeric events (Table 1) suggests that SARS-CoV-2 integration into the host genome is unlikely. Given that the HEK293T-L1 model increases detection of "rare integration events" and L1 can retrotranspose any poly-adenylated cellular RNA (2) and "insertions" are found preferentially in protein-coding exons, a bias unknown to L1 endonuclease insertions (3), these findings are likely spurious. Additionally, 2 of the identified 61 chime-ric nanopore genomic DNA (gDNA) reads contain human DNA from separate chromosomes (chr1,chr22 and chr18,chrX, respectively), suggesting a portion of chime-ric gDNA nanopore reads have arisen due to infrequent technical artifacts, such as base-calling software not recognizing an open-pore state between distinct molecules. Further to this, the authors reanalyzed published sequencing data and identified SARS-CoV-2 and human host chimeric reads in vitro and in patient RNA-seq (1). They show that the fraction of human-viral chimeric reads derived from negative-sense SARS-CoV-2 RNAs is higher in patients than that observed in vitro. These data are presented as evidence of SARS-CoV-2 integration and transcription. We question the presented data as evidence for this phenomenon for several key reasons. Firstly, chimeric virus−host reads are often reported in RNA-seq (4), including SARS-CoV-2 (5). This is partly due to complementary DNA fusions introduced during the reverse transcription step of library preparation (6). Secondly, larger pools of negative-sense SARS-CoV-2 chimeric reads may be due to differences in RNA extraction, library preparation, and sequencing platform. Given that negative-sense RNA is formed during the replication of SARS-CoV-2 and a template for messenger RNA production, significant variation of negative-sense reads is expected in patient RNA samples. Finally, there is no evidence of coronaviruses ever having integrated into the germ-line of host species, as might be expected if retro-transcription and integration occurs in nature, as systematic screening of >750 animal species failed to identify any coronavirus-derived endogenous viral elements (7).
SARS-CoV-2 Omicron is highly transmissible and has substantial resistance to antibody neutralization following immunization with ancestral spike-matched vaccines. It is unclear whether boosting with Omicron-specific vaccines would enhance immunity and protection. Here, nonhuman primates that received mRNA-1273 at weeks 0 and 4 were boosted at week 41 with mRNA-1273 or mRNA-Omicron. Neutralizing antibody titers against D614G were 4760 and 270 reciprocal ID 50 at week 6 (peak) and week 41 (pre-boost), respectively, and 320 and 110 for Omicron. Two weeks after boost, titers against D614G and Omicron increased to 5360 and 2980, respectively, for mRNA-1273 and 2670 and 1930 for mRNA-Omicron. Following either boost, 70-80% of spike-specific B cells were cross-reactive against both WA1 and Omicron. Significant and equivalent control of virus replication in lower airways was observed following either boost. Therefore, an Omicron boost may not provide greater immunity or protection compared to a boost with the current mRNA-1273 vaccine.
Importance: Vaccination against COVID-19 provides clear public health benefits, but vaccination also carries potential risks. The risks and outcomes of myocarditis after COVID-19 vaccination are unclear. Objective: To describe reports of myocarditis and the reporting rates after mRNA-based COVID-19 vaccination in the US. Design, setting, and participants: Descriptive study of reports of myocarditis to the Vaccine Adverse Event Reporting System (VAERS) that occurred after mRNA-based COVID-19 vaccine administration between December 2020 and August 2021 in 192 405 448 individuals older than 12 years of age in the US; data were processed by VAERS as of September 30, 2021. Exposures: Vaccination with BNT162b2 (Pfizer-BioNTech) or mRNA-1273 (Moderna). Main outcomes and measures: Reports of myocarditis to VAERS were adjudicated and summarized for all age groups. Crude reporting rates were calculated across age and sex strata. Expected rates of myocarditis by age and sex were calculated using 2017-2019 claims data. For persons younger than 30 years of age, medical record reviews and clinician interviews were conducted to describe clinical presentation, diagnostic test results, treatment, and early outcomes. Results: Among 192 405 448 persons receiving a total of 354 100 845 mRNA-based COVID-19 vaccines during the study period, there were 1991 reports of myocarditis to VAERS and 1626 of these reports met the case definition of myocarditis. Of those with myocarditis, the median age was 21 years (IQR, 16-31 years) and the median time to symptom onset was 2 days (IQR, 1-3 days). Males comprised 82% of the myocarditis cases for whom sex was reported. The crude reporting rates for cases of myocarditis within 7 days after COVID-19 vaccination exceeded the expected rates of myocarditis across multiple age and sex strata. The rates of myocarditis were highest after the second vaccination dose in adolescent males aged 12 to 15 years (70.7 per million doses of the BNT162b2 vaccine), in adolescent males aged 16 to 17 years (105.9 per million doses of the BNT162b2 vaccine), and in young men aged 18 to 24 years (52.4 and 56.3 per million doses of the BNT162b2 vaccine and the mRNA-1273 vaccine, respectively). There were 826 cases of myocarditis among those younger than 30 years of age who had detailed clinical information available; of these cases, 792 of 809 (98%) had elevated troponin levels, 569 of 794 (72%) had abnormal electrocardiogram results, and 223 of 312 (72%) had abnormal cardiac magnetic resonance imaging results. Approximately 96% of persons (784/813) were hospitalized and 87% (577/661) of these had resolution of presenting symptoms by hospital discharge. The most common treatment was nonsteroidal anti-inflammatory drugs (589/676; 87%). Conclusions and relevance: Based on passive surveillance reporting in the US, the risk of myocarditis after receiving mRNA-based COVID-19 vaccines was increased across multiple age and sex strata and was highest after the second vaccination dose in adolescent males and young men. This risk should be considered in the context of the benefits of COVID-19 vaccination.