Temporal Alterations in CD8+ T Cells During the Progression from Stage 1 to Stage 3 Type 1 Diabetes.

CD8+ T cells are perceived to play a major role in the pathogenesis of type 1 diabetes (T1D). In this study, we characterized the function and phenotype of circulating CD8+ memory T cells in samples from individuals at different stages of T1D progression using flow cytometry and single-cell multiomics. We observed two distinct CD8+ T-cell signatures during progression of T1D within the highly differentiated CD27-CD8+ memory T cell subset. A proinflammatory signature, with an increased frequency of IFN-γ+TNF-α+ CD27-CD8+ memory T cells, was observed in children with newly diagnosed T1D (stage 3) and correlated with the level of dysglycemia at diagnosis. In contrast, a co-inhibitory signature, with an increased frequency of KLRG1+TIGIT+ CD27-CD8+ memory T cells, was observed in islet autoantibody-positive children who later progressed to T1D (stage 1). No alterations within CD27-CD8+ memory T cells were observed in adults with established T1D or in children during the initial seroconversion to islet autoantibody positivity. Single-cell multiomics analyses suggested that CD27-CD8+ T cells expressing the IFNG+TNF+ proinflammatory signature may be distinct from those expressing the KLRG1+TIGIT+ co-inhibitory signature at the single-cell level. Collectively, our findings suggest that distinct blood CD8+ T-cell signatures could be employed as potential biomarkers of T1D progression.

The effect of type 1 diabetes protection and susceptibility associated HLA class II genotypes on DNA methylation in immune cells.

The HLA region, especially HLA class I and II genes, which encode molecules for antigen presentation to T cells, plays a major role in the predisposition to autoimmune disorders. To clarify the mechanisms behind this association, we examined genome-wide DNA methylation by microarrays to cover over 850,000 CpG sites in the CD4+ T cells and CD19+ B cells of healthy subjects homozygous either for DRB1*15-DQA1*01-DQB1*06:02 (DR2-DQ6, n = 14), associated with a strongly decreased T1D risk, DRB1*03-DQA1*05-DQB1*02 (DR3-DQ2, n = 19), or DRB1*04:01-DQA1*03-DQB1*03:02 (DR4-DQ8, n = 17), associated with a moderately increased T1D risk. In total, we discovered 14 differentially methylated CpG probes, of which 10 were located in the HLA region and six in the HLA-DRB1 locus. The main differences were between the protective genotype DR2-DQ6 and the risk genotypes DR3-DQ2 and DR4-DQ8, where the DR2-DQ6 group was hypomethylated compared to the other groups in all but four of the differentially methylated probes. The differences between the risk genotypes DR3-DQ2 and DR4-DQ8 were small. Our results indicate that HLA variants have few systemic effects on methylation and that their effect on autoimmunity is conveyed directly by HLA molecules, possibly by differences in expression levels or function.

Household Transmission and Clinical Features of SARS-CoV-2-Positive and -Negative Respiratory Tract Infections.

BACKGROUND: Comparative data on the transmission of respiratory infections positive and negative for SARS-CoV-2 in households with children are limited. METHODS: In June-August 2020, we recruited 700 participants (175 households, 376 children, 324 adults) to be prospectively followed for all respiratory tract infections. Follow-up lasted from recruitment till April 2022. Daily symptoms were monitored by weekly electronic questionnaires. SARS-CoV-2 PCR testing from nasopharyngeal specimens was performed for symptomatic participants and twice (one-week interval) for the household members of positive participants. Clinical features and secondary attack rates (SARs), based on the onset of symptoms, were compared between SARS-CoV-2-positive and -negative respiratory infections. RESULTS: Most (90%) SARS-CoV-2 infections occurred from January to April 2022 when Omicron BA.1 and BA.2 were the dominant variants. SARS-CoV-2-positive infections were transmitted more often than SARS-CoV-2-negative infections (SAR, 41% vs 24%; P < .001). SARS-CoV-2 transmission was similar for child and adult index cases (SAR, 40% vs 43%; P = .47), but the transmission of SARS-CoV-2-negative infections was higher for child index cases (SAR, 27% vs 18%; P < .001). CONCLUSIONS: Our findings demonstrate that SARS-CoV-2 Omicron viruses spread more effectively within households compared to other respiratory infections.

Distinct cellular immune responses in children en route to type 1 diabetes with different first-appearing autoantibodies.

Previous studies have revealed heterogeneity in the progression to clinical type 1 diabetes in children who develop islet-specific antibodies either to insulin (IAA) or glutamic acid decarboxylase (GADA) as the first autoantibodies. Here, we test the hypothesis that children who later develop clinical disease have different early immune responses, depending on the type of the first autoantibody to appear (GADA-first or IAA-first). We use mass cytometry for deep immune profiling of peripheral blood mononuclear cell samples longitudinally collected from children who later progressed to clinical disease (IAA-first, GADA-first, ≥2 autoantibodies first groups) and matched for age, sex, and HLA controls who did not, as part of the Type 1 Diabetes Prediction and Prevention study. We identify differences in immune cell composition of children who later develop disease depending on the type of autoantibodies that appear first. Notably, we observe an increase in CD161 expression in natural killer cells of children with ≥2 autoantibodies and validate this in an independent cohort. The results highlight the importance of endotype-specific analyses and are likely to contribute to our understanding of pathogenic mechanisms underlying type 1 diabetes development.

Neutralizing antibodies after the third COVID-19 vaccination in healthcare workers with or without breakthrough infection.

BACKGROUND: Vaccinations against the SARS-CoV-2 are still crucial in combating the ongoing pandemic that has caused more than 700 million infections and claimed almost 7 million lives in the past four years. Omicron (B.1.1.529) variants have incurred mutations that challenge the protection against infection and severe disease by the current vaccines, potentially compromising vaccination efforts. METHODS: We analyzed serum samples taken up to 9 months post third dose from 432 healthcare workers. Enzyme-linked immunosorbent assays (ELISA) and microneutralization tests (MNT) were used to assess the prevalence of vaccine-induced neutralizing antibodies against various SARS-CoV-2 Omicron variants. RESULTS: In this serological analysis we show that SARS-CoV-2 vaccine combinations of BNT162b2, mRNA-1273, and ChAdOx1 mount SARS-CoV-2 binding and neutralizing antibodies with similar kinetics, but with differing neutralization capabilities. The most recent Omicron variants, BQ.1.1 and XBB.1.5, show a significant increase in the ability to escape vaccine and infection-induced antibody responses. Breakthrough infections in thrice vaccinated adults were seen in over 50% of the vaccinees, resulting in a stronger antibody response than without infection. CONCLUSIONS: Different three-dose vaccine combinations seem to induce considerable levels of neutralizing antibodies against most SARS-CoV-2 variants. However, the ability of the newer variants BQ1.1 and XBB 1.5 to escape vaccine-induced neutralizing antibody responses underlines the importance of updating vaccines as new variants emerge.

During the COVID-19 pandemic, mass vaccination efforts against SARS-CoV-2 infection have provided effective protection against the virus and helped reduce the severity of symptoms in infected individuals. However, it is not well established whether the existing vaccines can provide the same protection against new and emerging SARS-CoV-2 variants that develop over time as the virus evolves. In this study, we tested combinations of three-dose COVID-19 vaccines given in random order to protect against all SARS-CoV-2 variants in circulation including the newest being Omicron variants. We demonstrate that more than half of the population who received the three-dose vaccine combinations were infected with SARS-CoV-2 Omicron variants after receiving the last vaccine dose. These findings indicate the need to develop new vaccine candidates against emerging SARS-CoV-2 variants.

Increased Frequency of the HLA-DRB1*04:04-DQA1*03-DQB1*03:02 Haplotype Among HLA-DQB1*06:02-Positive Children With Type 1 Diabetes.

HLA-DR/DQ haplotypes largely define genetic susceptibility to type 1 diabetes (T1D). The DQB1*06:02-positive haplotype (DR15-DQ602) common in individuals of European ancestry is very rare among children with T1D. Among 4,490 children with T1D in the Finnish Pediatric Diabetes Register, 57 (1.3%) case patients with DQB1*06:02 were identified, in comparison with 26.1% of affected family-based association control participants. There were no differences between DQB1*06:02-positive and -negative children with T1D regarding sex, age, islet autoantibody distribution, or autoantibody levels, but significant differences were seen in the frequency of second class II HLA haplotypes. The most prevalent haplotype present with DQB1*06:02 was DRB1*04:04-DQA1*03-DQB1*03:02, which was found in 27 (47.4%) of 57 children, compared with only 797 (18.0%) of 4,433 among DQB1*06:02-negative case patients (P < 0.001 by χ2 test). The other common risk-associated haplotypes, DRB1*04:01-DQA1*03-DQB1*03:02 and (DR3)-DQA1*05-DQB1*02, were less prevalent in DQB1*06:02-positive versus DQB1*06:02-negative children (P < 0.001). HLA-B allele frequencies did not differ by DQB1*06:02 haplotype between children with T1D and control participants or by DRB1*04:04-DQA1*03-DQB1*03:02 haplotype between DQB1*06:02-positive and -negative children with T1D. The increased frequency of the DRB1*04:04 allele among DQB1*06:02-positive case patients may indicate a preferential ability of the DR404 molecule to present islet antigen epitopes despite competition by DQ602.

CXADR polymorphism rs6517774 modifies islet autoimmunity characteristics and exhibits sex disparity.

Enteroviral infections have been linked to the development of islet autoimmunity (IA) and type 1 diabetes (T1D), and the coxsackie and adenovirus receptor (CXADR) is one of the ligands used by adenoviruses and enteroviruses for cell internalization. Two CXADR single nucleotide polymorphisms (SNPs), rs6517774 and rs2824404, were previously associated with an increased susceptibility to IA in the international TEDDY study (The Environmental Determinants of Diabetes in the Young). This study aimed to replicate the results by genotyping 2886 children enrolled in the Finnish Diabetes Prediction and Prevention study (DIPP). In our preliminary analysis of the SNPs' allelic distributions, we could not find any association with IA susceptibility. However, a stratified analysis revealed a sex disparity, since the allelic distribution of rs6517774 was different when comparing autoantibody positive females with males; a difference not seen in healthy subjects. By using HLA risk groups and sex as covariates, a Cox regression survival analysis found that the rs6517774 (A/G) SNP was associated with a lower age at seroconversion in females (Female*rs6517774-AA; HR = 1.53, p = 0.002), while introducing a protective effect in males. Accordingly, we propose that rs6517774 alters IA characteristics by modifying the age at seroconversion in a sex-dependent manner. In light of this observation, rs6517774 now joins a limited set on SNPs found to introduce sex-dependent risk effects on the age at IA initiation.

DNA methylation differences within INS, PTPN22 and IL2RA promoters in lymphocyte subsets in children with type 1 diabetes and controls.

We elucidated the effect of four known T1D-susceptibility associated single nucleotide polymorphism (SNP) markers in three genes (rs12722495 and rs2104286 in IL2RA, rs689 in INS and rs2476601 in PTPN22) on CpG site methylation of their proximal promoters in different lymphocyte subsets using pyrosequencing. The study cohort comprised 25 children with newly diagnosed T1D and 25 matched healthy controls. The rs689 SNP was associated with methylation at four CpG sites in INS promoter: -234, -206, -102 and -69. At all four CpG sites, the susceptibility genotype AA was associated with a higher methylation level compared to the other genotypes. We also found an association between rs12722495 and methylation at CpG sites -373 and -356 in IL2RA promoter in B cells, where the risk genotype AA was associated with lower methylation level compared to the AG genotype. The other SNPs analyzed did not demonstrate significant associations with CpG site methylation in the examined genes. Additionally, we compared the methylation between children with T1D and controls, and found statistically significant methylation differences at CpG -135 in INS in CD8+ T cells (p = 0.034), where T1D patients had a slightly higher methylation compared to controls (87.3 ± 7.2 vs. 78.8 ± 8.9). At the other CpG sites analyzed, the methylation was similar. Our results not only confirm the association between INS methylation and rs689 discovered in earlier studies but also report this association in sorted immune cells. We also report an association between rs12722495 and IL2RA promoter methylation in B cells. These results suggest that at least part of the genetic effect of rs689 and rs12722495 on T1D pathogenesis may be conveyed by DNA methylation.

Circulating metabolic signatures of rapid and slow progression to type 1 diabetes in islet autoantibody-positive children.

Aims/hypothesis: Appearance of multiple islet cell autoantibodies in early life is indicative of future progression to overt type 1 diabetes, however, at varying rates. Here, we aimed to study whether distinct metabolic patterns could be identified in rapid progressors (RP, disease manifestation within 18 months after the initial seroconversion to autoantibody positivity) vs. slow progressors (SP, disease manifestation at 60 months or later from the appearance of the first autoantibody). Methods: Longitudinal samples were collected from RP (n=25) and SP (n=41) groups at the ages of 3, 6, 12, 18, 24, or ≥ 36 months. We performed a comprehensive metabolomics study, analyzing both polar metabolites and lipids. The sample series included a total of 239 samples for lipidomics and 213 for polar metabolites. Results: We observed that metabolites mediated by gut microbiome, such as those involved in tryptophan metabolism, were the main discriminators between RP and SP. The study identified specific circulating molecules and pathways, including amino acid (threonine), sugar derivatives (hexose), and quinic acid that may define rapid vs. slow progression to type 1 diabetes. However, the circulating lipidome did not appear to play a major role in differentiating between RP and SP. Conclusion/interpretation: Our study suggests that a distinct metabolic profile is linked with the type 1 diabetes progression. The identification of specific metabolites and pathways that differentiate RP from SP may have implications for early intervention strategies to delay the development of type 1 diabetes.

T cell immunity following COVID-19 vaccination in adult patients with primary antibody deficiency - a 22-month follow-up.

Primary antibody deficiencies, such as common variable immunodeficiency (CVID), are heterogenous disease entities consisting of primary hypogammaglobulinemia and impaired antibody responses to vaccination and natural infection. CVID is the most common primary immunodeficiency in adults, presenting with recurrent bacterial infections, enteropathy, autoimmune disorders, interstitial lung diseases and increased risk of malignancies. Patients with CVID are recommended to be vaccinated against SARS-CoV-2, but there are relatively few studies investigating humoral and cellular responses to immunization. We studied the dynamics of humoral and cell-mediated immunity responses up to 22 months in 28 patients with primary immunodeficiency and three patients with secondary immunodeficiency receiving ChAdOx1, BNT162b2 and mRNA-1273 COVID-19 vaccines. Despite inadequate humoral response to immunization, we demonstrate a robust T cell activation likely protecting from severe COVID-19.

Coronavirus spike protein-specific antibodies indicate frequent infections and reinfections in infancy and among BNT162b2-vaccinated healthcare workers.

The prevalence of seasonal human coronavirus (HCoV) infections in early childhood and adults has not been well analyzed in longitudinal serological studies. Here we analyzed the changes in HCoV (229E, HKU1, NL63, OC43, MERS, and SARS-CoV-2) spike-specific antibody levels in follow-up serum specimens of 140 children at the age of 1, 2, and 3 years, and of 113 healthcare workers vaccinated for Covid-19 with BNT162b2-vaccine. IgG antibody levels against six recombinant HCoV spike subunit 1 (S1) proteins were measured by enzyme immunoassay. We show that by the age of three years the cumulative seropositivity for seasonal HCoVs increased to 38-81% depending on virus type. BNT162b2 vaccinations increased anti-SARS-CoV-2 S1 antibodies, but no increase in seasonal coronavirus antibodies associated with vaccinations. In healthcare workers (HCWs), during a 1-year follow-up, diagnostic antibody rises were seen in 5, 4 and 14% of the cases against 229E, NL63 and OC43 viruses, respectively, correlating well with the circulating HCoVs. In 6% of the HCWs, a diagnostic antibody rise was seen against S1 of HKU1, however, these rises coincided with anti-OC43 S1 antibody rises. Rabbit and guinea pig immune sera against HCoV S1 proteins indicated immunological cross-reactivity within alpha-CoV (229E and NL63) and beta-CoV (HKU1 and OC43) genera.

Childhood manifestations of 22q11.2 deletion syndrome: A Finnish nationwide register-based cohort study.

AIM: The aim of the study was to describe the clinical manifestations of 22q11.2 deletion syndrome patients in the Finnish paediatric population. METHODS: Nationwide registry data including all diagnoses and procedures of every public hospital in Finland between 2004 and 2018 along with mortality and cancer registry data were retrieved. Patients born during the study period and with an ICD-10 code of D82.1 or Q87.06 were included as having 22q11.2 deletion syndrome. A control group was formed with patients born during the study period and with benign cardiac murmur diagnosed under the age of 1 year. RESULTS: We identified 100 pediatric patients with 22q11.2 deletion syndrome (54% males, median age at diagnosis <1 year, median follow-up 9 years). Cumulative mortality was 7.1%. Among patients with 22q11.2 deletion syndrome, 73.8% had congenital heart defects, 21.8% had cleft palate, 13.6% had hypocalcaemia, and 7.2% had immunodeficiencies. Furthermore, 29.6% were diagnosed with autoimmune diseases, 92.9% had infections, and 93.2% had neuropsychiatric and developmental issues during follow-up. Malignancy was found in 2.1% of the patients. CONCLUSION: The 22q11.2 deletion syndrome is associated with increased mortality and substantial multimorbidity in children. A structured multidisciplinary approach is necessary for managing patients with 22q11.2 deletion syndrome.

HLA-DQ-conferred risk for type 1 diabetes does not alter neutralizing antibody response to a widely used enterovirus vaccine, the poliovirus vaccine.

This study investigated whether children with HLA-DQ-conferred risk for type 1 diabetes (T1D) have an altered immune response to the widely-used enterovirus vaccine, namely poliovirus vaccine, and whether initiation of autoimmunity to pancreatic islets modulates this response. Neutralizing antibodies induced by the inactivated poliovirus vaccine against poliovirus type 1 (Salk) were analysed as a marker of protective immunity at the age of 18 months in a prospective birth cohort. No differences were observed in antibody titers between children with and without genetic risk for T1D (odds ratio [OR] = 0.90 [0.83, 1.06], p = 0.30). In the presence of the genetic risk, no difference was observed between children with and without islet autoimmunity (OR = 1.00 [0.78, 1.28], p = 1.00). This did not change when only children with the autoimmunity before 18 months of age were included in the analyses (OR = 1.00 [0.85, 1.18], p = 1.00). No effect was observed when groups were stratified based on autoantigen specificity of the first-appearing autoantibody (IAA or GADA). The children in each comparison group were matched for sex, calendar year and month of birth, and municipality. Accordingly, we found no indication that children who are at risk to develop islet autoimmunity would have a compromised humoral immune response which could have increased their susceptibility for enterovirus infections. In addition, the proper immune response supports the idea of testing novel enterovirus vaccines for the prevention of T1D among these individuals.

SARS-CoV-2 and type 1 diabetes in children in Finland: an observational study.

BACKGROUND: Some epidemiological studies have suggested an increase in incidence of type 1 diabetes during the COVID-19 pandemic, however the mechanism(s) behind such an increase have yet to be identified. In this study we aimed to evaluate the possible role of the SARS-CoV-2 virus in the reported increase in the rate of type 1 diabetes. METHODS: In this observational cohort study using data from the Finnish Pediatric Diabetes Register (FPDR), we assessed the incidence of type 1 diabetes (number of children with newly diagnosed type 1 diabetes per 100 000 person-years during the pandemic and the reference period) during the first 18 months of the COVID-19 pandemic in children in Finland younger than 15 years old compared with a reference period which included three corresponding pre-pandemic periods also obtained from the FPDR. Children with confirmed monogenic diabetes were excluded. We also compared the phenotype and HLA genotype of the disease between these two cohorts, and analysed the proportion of newly diagnosed people with type 1 diabetes testing positive for SARS-CoV-2 antibodies. FINDINGS: 785 children and adolescents in Finland were diagnosed with type 1 diabetes from March 1, 2020, to Aug 31, 2021. In the reference period, which comprised three similar 18-month terms (from March 1, 2014, to Aug 31, 2015; March 1, 2016, to Aug 31, 2017; and March 1, 2018, to Aug 31, 2019) 2096 children and adolescents were diagnosed. The incidence of type 1 diabetes was 61·0 per 100 000 person-years (95% CI 56·8-65·4) among children younger than 15 years old during the pandemic, which was significantly higher than during the reference period (52·3 per 100 000 person-years; 50·1-54·6). The incidence rate ratio adjusted for age and sex for the COVID-19 pandemic was 1·16 (1·06-1·25; p=0·0006) when compared with the reference period. The children diagnosed during the COVID-19 pandemic had more often diabetic ketoacidosis (p<0·001), had a higher HbA1c (p<0·001), and tested more frequently positive for glutamic acid debarboxylase antibodies at diagnosis (p<0·001) than those diagnosed before the pandemic. There were no significant differences in the distribution of HLA genotypes between the two periods. Only five of those diagnosed during the pandemic (0·9%) of 583 tested positive for infection-induced SARS-CoV-2 antibodies. INTERPRETATION: Children and adolescents diagnosed with type 1 diabetes during the pandemic had a more severe disease at diagnosis. The observed increase in type 1 diabetes incidence during the first 18 months could be a consequence of lockdown and physical distancing rather than a direct effect of SARS-CoV-2 infection. FUNDING: Helsinki University Hospital Research Funds, EU Horizon 2020 (Versatile emerging infectious disease observatory project), Academy of Finland, Sigrid Jusélius Foundation, Jane & Aatos Erkko Foundation, and Medicinska understödsföreningen Liv och Hälsa. TRANSLATIONS: For the Finnish and Swedish translations of the abstract see Supplementary Materials section.

Persistent T cell-mediated immune responses against Omicron variants after the third COVID-19 mRNA vaccine dose.

Introduction: The prime-boost COVID-19 mRNA vaccination strategy has proven to be effective against severe COVID-19 disease and death. However, concerns have been raised due to decreasing neutralizing antibody levels after COVID-19 vaccination and due to the emergence of new immuno-evasive SARS-CoV-2 variants that may require additional booster vaccinations. Methods: In this study, we analyzed the humoral and cell-mediated immune responses against the Omicron BA.1 and BA.2 subvariants in Finnish healthcare workers (HCWs) vaccinated with three doses of COVID-19 mRNA vaccines. We used enzyme immunoassay and microneutralization test to analyze the levels of SARS-CoV-2 specific IgG antibodies in the sera of the vaccinees and the in vitro neutralization capacity of the sera. Activation induced marker assay together with flow cytometry and extracellular cytokine analysis was used to determine responses in SARS-CoV-2 spike protein stimulated PBMCs. Results: Here we show that within the HCWs, the third mRNA vaccine dose recalls both humoral and T cell-mediated immune responses and induces high levels of neutralizing antibodies against Omicron BA.1 and BA.2 variants. Three weeks after the third vaccine dose, SARS-CoV-2 wild type spike protein-specific CD4+ and CD8+ T cells are observed in 82% and 71% of HCWs, respectively, and the T cells cross-recognize both Omicron BA.1 and BA.2 spike peptides. Although the levels of neutralizing antibodies against Omicron BA.1 and BA.2 decline 2.5 to 3.8-fold three months after the third dose, memory CD4+ T cell responses are maintained for at least eight months post the second dose and three months post the third vaccine dose. Discussion: We show that after the administration of the third mRNA vaccine dose the levels of both humoral and cell-mediated immune responses are effectively activated, and the levels of the spike-specific antibodies are further elevated compared to the levels after the second vaccine dose. Even though at three months after the third vaccine dose antibody levels in sera decrease at a similar rate as after the second vaccine dose, the levels of spike-specific CD4+ and CD8+ T cells remain relatively stable. Additionally, the T cells retain efficiency in cross-recognizing spike protein peptide pools derived from Omicron BA.1 and BA.2 subvariants. Altogether our results suggest durable cellmediated immunity and protection against SARS-CoV-2.

Finnish children carrying the high-risk HLA genotype have a 45-fold increased risk of type 1 diabetes compared to peers with neutral or protective genotypes.

The association between HLA genotypes and type 1 diabetes is well known. We set out to examine incidence rates and ratios of type 1 diabetes depending on the risk afflicted by HLA genotype. Children with the high-risk genotype have a 45-fold disease risk compared to peers with neutral or protective genotypes.

Strong Neutralizing Antibody Responses to SARS-CoV-2 Variants Following a Single Vaccine Dose in Subjects With Previous SARS-CoV-2 Infection.

Background: Previous severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection primes the immune system; thus individuals who have recovered from infection have enhanced immune responses to subsequent vaccination (hybrid immunity). However, it remains unclear how well hybrid immunity induced by severe or mild infection can cross-neutralize emerging variants. We aimed to compare the strength and breadth of antibody responses in vaccinated recovered and uninfected subjects. Methods: We measured spike-specific immunoglobulin (Ig)G and neutralizing antibodies (NAbs) from vaccinated subjects including 320 with hybrid immunity and 20 without previous infection. From 29 subjects with a previous severe or mild infection, we also measured NAb responses against Alpha (B.1.1.7), Beta (B.1.351), Delta (B.1.617.2), and Omicron (B.1.1.529/BA.1) variants following vaccination. Results: A single vaccine dose induced 2-fold higher anti-spike IgG concentrations and up to 4-fold higher neutralizing potency of antibodies in subjects with a previous infection compared with vaccinated subjects without a previous infection. Hybrid immunity was more enhanced after a severe than a mild infection, with sequentially decreasing NAb titers against Alpha, Beta, Delta, and Omicron variants. We found similar IgG concentrations in subjects with a previous infection after 1 or 2 vaccine doses. Conclusions: Hybrid immunity induced strong IgG responses, particularly after severe infection. However, the NAb titers were low against heterologous variants, especially against Omicron.

Non-HLA Gene Polymorphisms in the Pathogenesis of Type 1 Diabetes: Phase and Endotype Specific Effects.

The non-HLA loci conferring susceptibility to type 1 diabetes determine approximately half of the genetic disease risk, and several of them have been shown to affect immune-cell or pancreatic ß-cell functions. A number of these loci have shown associations with the appearance of autoantibodies or with progression from seroconversion to clinical type 1 diabetes. In the current study, we have re-analyzed 21 of our loci with prior association evidence using an expanded DIPP follow-up cohort of 976 autoantibody positive cases and 1,910 matched controls. Survival analysis using Cox regression was applied for time periods from birth to seroconversion and from seroconversion to type 1 diabetes. The appearance of autoantibodies was also analyzed in endotypes, which are defined by the first appearing autoantibody, either IAA or GADA. Analyzing the time period from birth to seroconversion, we were able to replicate our previous association findings at PTPN22, INS, and NRP1. Novel findings included associations with ERBB3, UBASH3A, PTPN2, and FUT2. In the time period from seroconversion to clinical type 1 diabetes, prior associations with PTPN2, CD226, and PTPN22 were replicated, and a novel association with STAT4 was observed. Analyzing the appearance of autoantibodies in endotypes, the PTPN22 association was specific for IAA-first. In the progression phase, STAT4 was specific for IAA-first and ERBB3 to GADA-first. In conclusion, our results further the knowledge of the function of non-HLA risk polymorphisms in detailing endotype specificity and timing of disease development.

Detection of enterovirus RNA in peripheral blood mononuclear cells correlates with the presence of the predisposing allele of the type 1 diabetes risk gene IFIH1 and with disease stage.

AIMS/HYPOTHESIS: Enteroviral infection has been implicated consistently as a key environmental factor correlating with the appearance of autoimmunity and/or the presence of overt type 1 diabetes, in which pancreatic insulin-producing beta cells are destroyed by an autoimmune response. Genetic predisposition through variation in the type 1 diabetes risk gene IFIH1 (interferon induced with helicase C domain 1), which encodes the viral pattern-recognition receptor melanoma differentiation-associated protein 5 (MDA5), supports a potential link between enterovirus infection and type 1 diabetes. METHODS: We used molecular techniques to detect enterovirus RNA in peripheral blood samples (in separated cellular compartments or plasma) from two cohorts comprising 79 children or 72 adults that include individuals with and without type 1 diabetes who had multiple autoantibodies. We also used immunohistochemistry to detect the enteroviral protein VP1 in the pancreatic islets of post-mortem donors (n=43) with type 1 diabetes. RESULTS: We observed enhanced detection sensitivity when sampling the cellular compartment compared with the non-cellular compartment of peripheral blood (OR 21.69; 95% CI 3.64, 229.20; p<0.0001). In addition, we show that children with autoimmunity are more likely to test positive for enterovirus RNA than those without autoimmunity (OR 11.60; 95% CI 1.89, 126.90; p=0.0065). Furthermore, we found that individuals carrying the predisposing allele (946Thr) of the common variant in IFIH1 (rs1990760, Thr946Ala) are more likely to test positive for enterovirus in peripheral blood (OR 3.07; 95% CI 1.02, 8.58; p=0.045). In contrast, using immunohistochemistry, there was no correlation between the common variant in IFIH1 and detection of enteroviral VP1 protein in the pancreatic islets of donors with type 1 diabetes. CONCLUSIONS/INTERPRETATION: Our data indicate that, in peripheral blood, antigen-presenting cells are the predominant source of enterovirus infection, and that infection is correlated with disease stage and genetic predisposition, thereby supporting a role for enterovirus infection prior to disease onset.

Long-Lasting T Cell Responses in BNT162b2 COVID-19 mRNA Vaccinees and COVID-19 Convalescent Patients.

The emergence of novel variants of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has made it more difficult to prevent the virus from spreading despite available vaccines. Reports of breakthrough infections and decreased capacity of antibodies to neutralize variants raise the question whether current vaccines can still protect against COVID-19 disease. We studied the dynamics and persistence of T cell responses using activation induced marker (AIM) assay and Th1 type cytokine production in peripheral blood mononuclear cells obtained from BNT162b2 COVID-19 mRNA vaccinated health care workers and COVID-19 patients. We demonstrate that equally high T cell responses following vaccination and infection persist at least for 6 months against Alpha, Beta, Gamma, and Delta variants despite the decline in antibody levels.