Contribution of SARS-CoV-2 infection preceding COVID-19 mRNA vaccination to generation of cellular and humoral immune responses in children

Primary COVID-19 vaccination for children, 5-17 years of age, was offered in the Netherlands at a time when a substantial part of this population had already experienced a SARS-CoV-2 infection. While vaccination has been shown effective, underlying immune responses have not been extensively studied. We studied immune responsiveness to one and/or two doses of primary BNT162b2 mRNA vaccination and compared the humoral and cellular immune response in children with and without a preceding infection. Antibodies targeting the original SARS-CoV-2 Spike or Omicron Spike were measured by multiplex immunoassay. B-cell and T-cell responses were investigated using enzyme-linked immunosorbent spot (ELISpot) assays. The activation of CD4+ and CD8+ T cells was studied by flowcytometry. Primary vaccination induced both a humoral and cellular adaptive response in naive children. These responses were stronger in those with a history of infection prior to vaccination. A second vaccine dose did not further boost antibody levels in those who previously experienced an infection. Infection-induced responsiveness prior to vaccination was mainly detected in CD8+ T cells, while vaccine-induced T-cell responses were mostly by CD4+ T cells. Thus, SARS-CoV-2 infection prior to vaccination enhances adaptive cellular and humoral immune responses to primary COVID-19 vaccination in children. As most children are now expected to contract infection before the age of five, the impact of infection-induced immunity in children is of high relevance. Therefore, considering natural infection as a priming immunogen that enhances subsequent vaccine-responsiveness may help decision-making on the number and timing of vaccine doses

The adaptive immune system in early life: The shift makes it count.

Respiratory infectious diseases encountered early in life may result in life-threatening disease in neonates, which is primarily explained by the relatively naive neonatal immune system. Whereas vaccines are not readily available for all infectious diseases, vaccinations have greatly reduced childhood mortality. However, repeated vaccinations are required to reach protective immunity in infants and not all vaccinations are effective at young age. Moreover, protective adaptive immunity elicited by vaccination wanes more rapidly at young age compared to adulthood. The infant adaptive immune system has previously been considered immature but this paradigm has changed during the past years. Recent evidence shows that the early life adaptive immune system is equipped with a strong innate-like effector function to eliminate acute pathogenic threats. These strong innate-like effector capacities are in turn kept in check by a tolerogenic counterpart of the adaptive system that may have evolved to maintain balance and to reduce collateral damage. In this review, we provide insight into these aspects of the early life’s adaptive immune system by addressing recent literature. Moreover, we speculate that this shift from innate-like and tolerogenic adaptive immune features towards formation of immune memory may underlie different efficacy of infant vaccination in these different phases of immune development. Therefore, presence of innate-like and tolerogenic features of the adaptive immune system may be used as a biomarker to improve vaccination strategies against respiratory and other infections in early life

Is fatigue in Marfan syndrome related to orthostatic intolerance?

Patients with Marfan syndrome have a tall stature, which could be associated with low orthostatic tolerance. Fatigue, a common complaint of these patients, is also related to orthostatic intolerance. Treatment with beta-blockers, to prevent aortic complications, could be a reinforcing factor of both. This study aimed to investigate (1) the relationship between symptoms of orthostatic tolerance and in patients with Marfan syndrome, and (2) whether termination of beta-blocker therapy improves orthostatic tolerance. Symptoms of fatigue and orthostatic complaints were assessed in 49 subjects using questionnaires (MFI-20 and Autonomic Symptoms Profile). Marfan patients have a high level of fatigue and orthostatic complaints when compared to the general population. Fatigue and orthostatic tolerance are significantly correlated. Orthostatic tolerance was assessed in 9 additional subjects by an active-standing test and head-up tilt for 5 minutes, and 24 hours blood pressure monitoring, once during beta-blocker therapy and once after ceasing beta-blockers for 2 weeks. During hemodynamic testing Marfan patients frequently showed Initial Orthostatic Hypotension and an abnormally high initial heart rate response. Ceasing beta-blockers did not affect the initial blood pressure response. Patients with Marfan syndrome are fatigued and have low orthostatic tolerance, which are significantly correlated. Patients could be educated in physical counterpressure maneuvers to increase orthostatic toleranc

SARS-CoV-2 Omicron BA.4/BA.5 Mutations in Spike Leading to T Cell Escape in Recently Vaccinated Individuals.

SARS-CoV-2 Omicron (B.1.1.529) lineages rapidly became dominant in various countries reflecting its enhanced transmissibility and ability to escape neutralizing antibodies. Although T cells induced by ancestral SARS-CoV-2-based vaccines also recognize Omicron variants, we showed in our previous study that there was a marked loss of T cell cross-reactivity to spike epitopes harboring Omicron BA.1 mutations. The emerging BA.4/BA.5 subvariants carry other spike mutations than the BA.1 variant. The present study aims to investigate the impact of BA.4/BA.5 spike mutations on T cell cross-reactivity at the epitope level. Here, we focused on universal T-helper epitopes predicted to be presented by multiple common HLA class II molecules for broad population coverage. Fifteen universal T-helper epitopes of ancestral spike, which contain mutations in the Omicron BA.4/BA.5 variants, were identified utilizing a bioinformatic tool. T cells isolated from 10 subjects, who were recently vaccinated with mRNA-based BNT162b2, were tested for functional cross-reactivity between epitopes of ancestral SARS-CoV-2 spike and the Omicron BA.4/BA.5 spike counterparts. Reduced T cell cross-reactivity in one or more vaccinees was observed against 87% of the tested 15 non-conserved CD4+ T cell epitopes. These results should be considered for vaccine boosting strategies to protect against Omicron BA.4/BA.5 and future SARS-CoV-2 variants

SARS-CoV-2 Omicron BA.4/BA.5 Mutations in Spike Leading to T Cell Escape in Recently Vaccinated Individuals

SARS-CoV-2 Omicron (B.1.1.529) lineages rapidly became dominant in various countries reflecting its enhanced transmissibility and ability to escape neutralizing antibodies. Although T cells induced by ancestral SARS-CoV-2-based vaccines also recognize Omicron variants, we showed in our previous study that there was a marked loss of T cell cross-reactivity to spike epitopes harboring Omicron BA.1 mutations. The emerging BA.4/BA.5 subvariants carry other spike mutations than the BA.1 variant. The present study aims to investigate the impact of BA.4/BA.5 spike mutations on T cell cross-reactivity at the epitope level. Here, we focused on universal T-helper epitopes predicted to be presented by multiple common HLA class II molecules for broad population coverage. Fifteen universal T-helper epitopes of ancestral spike, which contain mutations in the Omicron BA.4/BA.5 variants, were identified utilizing a bioinformatic tool. T cells isolated from 10 subjects, who were recently vaccinated with mRNA-based BNT162b2, were tested for functional cross-reactivity between epitopes of ancestral SARS-CoV-2 spike and the Omicron BA.4/BA.5 spike counterparts. Reduced T cell cross-reactivity in one or more vaccinees was observed against 87% of the tested 15 non-conserved CD4+ T cell epitopes. These results should be considered for vaccine boosting strategies to protect against Omicron BA.4/BA.5 and future SARS-CoV-2 variants

Daily, weekly, monthly, and seasonal patterns in the occurrence of vasovagal syncope in an older population

AIMS: The aim of this study was to assess the frequency of vasovagal episodes over the day, week, month, and seasons. METHODS AND RESULTS: This study was part of the multi-centre International Study on Syncope of Uncertain Etiology-2 (ISSUE-2), which included patients, aged 30 years or older, with severe neurally mediated syncope between June 2002 and July 2004. The Implantable Loop Recorder (ILR) was used to document the syncope-related ECG periods. For this study patients with recorded syncopal episodes after ILR-implantation was selected. At least one episode was documented in 106 patients. A higher number of episodes were documented during the morning than during other periods of the day (P < 0.01). There was no difference between various days of the week, episodes per month, or between seasons. There was no difference between age and gender groups, although elderly patients seemed to be responsible for the peak in the morning. CONCLUSION: A circadian pattern in the frequency of vasovagal episodes exists, with a peak in the morning. This is in accordance with reports of diurnal variations in blood pressure and heart rate. No difference was observed in syncope distribution between days of the week, months, or season

Omicron BA.1 Mutations in SARS-CoV-2 Spike Lead to Reduced T-Cell Response in Vaccinated and Convalescent Individuals.

Omicron BA.1 variant can readily infect people with vaccine-induced or naturally acquired SARS-CoV-2 immunity facilitated by escape from neutralizing antibodies. In contrast, T-cell reactivity against the Omicron BA.1 variant seems relatively well preserved. Here, we studied the preexisting T cells elicited by either vaccination with the mRNA-based BNT162b2 vaccine or by natural infection with ancestral SARS-CoV-2 for their cross-reactive potential to 20 selected CD4+ T-cell epitopes of spike-protein-harboring Omicron BA.1 mutations. Although the overall memory CD4+ T-cell responses primed by the ancestral spike protein was still preserved generally, we show here that there is also a clear loss of memory CD4+ T-cell cross-reactivity to immunodominant epitopes across the spike protein due to Omicron BA.1 mutations. Complete or partial loss of preexisting T-cell responsiveness was observed against 60% of 20 nonconserved CD4+ T-cell epitopes predicted to be presented by a broad set of common HLA class II alleles. Monitoring such mutations in circulating strains helps predict which virus variants may escape previously induced cellular immunity and could be of concern

Treg-cell marker frequency and density are increased on live BCG-activated CD8⁺ vs. CD4⁺ T cells.

<p>A: BCG induces Treg-cell marker expression on CD4⁺ and CD8⁺ T cells; after live BCG stimulation the percentage of total CD8⁺ T cells expressing CD25, Foxp3, CD39, LAG-3 or CCL4 is significantly higher compared to CD4⁺ T cells, depicted here as frequency of CD8⁺ or CD4⁺ population. Differences in Treg marker expression between heatkilled BCG–activated CD8⁺ vs. CD4⁺ T cells were not significant, except for expression of CCL4; CCL4 expression was also significantly higher on CD8⁺ T cells compared to CD4⁺ T cells in samples not stimulated with BCG (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0094192#pone.0094192.s001" target="_blank">Fig. S1</a>) (<i>*p</i> < 0.05, Wilcoxon signed-ranks test). B: Mean fluorescence intensities (MFIs) of CD25 and CD39 are increased on live BCG-activated CD8⁺ T cells as compared to CD4⁺ T cells. Gating was performed as demonstrated in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0094192#pone-0094192-g001" target="_blank">figure 1A</a>. To assess differences in intrinsic intensity of expression on CD4⁺ and CD8⁺ T cells, respectively, MFIs of positive Treg marker populations in samples not stimulated with BCG were compared; this was similar on CD4⁺ and CD8⁺ T cells for MFIs of CD25, Foxp3 and CD39. Data are representative of seven <i>in vitro</i> PPD-responders six days after heatkilled or live BCG stimulation (*<i>p</i> < 0.05; Wilcoxon signed-ranks test).</p

Co-expression of multiple Treg-cell markers enriches for CD8⁺, and not CD4⁺ T cells.

<p>A. The percentage of total CD8⁺ T cells co-expressing CD39, LAG-3, CCL4, CD25 and/or Foxp3 in different combinations is significantly increased, compared to CD4⁺ T cells. Demonstrated is a combined analysis using Boolean gating of cells from ten donors six days after live BCG infection. Gating was performed as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0094192#pone-0094192-g001" target="_blank">figure 1A</a>. Boxes: 25th to 75th percentiles; line at median; whiskers: minimum to maximum (*<i>p</i> < 0.05, **<i>p</i> < 0.01; Wilcoxon signed-ranks test). B. Combining Treg markers enriches for CD8⁺ T cells as opposed to CD4⁺ T cells. Boolean gating was performed on CD3⁺ T cells of ten donors; CD8 vs. CD4 gating is demonstrated (top) and the CD8⁺ proportion of these gated populations is demonstrated (bottom) for a selection of CD3⁺ Boolean gates. The CD8⁺ proportion increased significantly using a combination of Treg markers as compared to the complete CD3⁺ population. Boxes: minimum to maximum, line at median (Wilcoxon signed-ranks test).</p

Heatkilled vs. live BCG-activated expression of Treg-cell markers on CD4⁺ and CD8⁺ T cells.

<p>A: Gating strategy: cells were gated on single cells, live lymphocytes, CD3⁺ and CD4⁺CD8⁻ vs. CD4⁻CD8⁺. Demonstrated is the synchronized gating on the positive population of interest for CD4⁺CD8⁻ and CD8⁺CD4⁻ T cells; here the CD25-positive population. B: Heatkilled and live BCG activate CD25⁺Foxp3⁺ and LAG-3⁺CD39⁺ T cells. Expression of regulatory T cell markers on CD4⁺ and CD8⁺ T cells of <i>in vitro</i> PPD responders was analysed by flowcytometry six days after heatkilled or live BCG stimulation. For each donor gating was compared to samples not stimulated with BCG (demonstrated in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0094192#pone.0094192.s001" target="_blank">Fig. S1</a>). Data are representative of seven responders.</p