Background Patients with inborn errors of immunity (IEI) have high risks of severe complications after SARS-CoV-2 infection. Although vaccination is effective in preventing severe COVID-19, boosters are required as the protection provided wanes over time. A greater number of boosters may be required in IEI patients compared to healthy individuals due to their immunodeficient state, but no such data are available. Objective To investigate and compare the immunogenicity between 4 doses of COVID-19 vaccination for patients with IEI and 3 doses for healthy individuals. Methods In the current study (NCT04800133), either BNT162b2 or CoronaVac was administered as the fourth dose. Healthy individuals who received three doses were included for comparison. Humoral and cellular immunogenicity against wild-type (WT) and JN.1 SARS-CoV-2 was assessed between IEI patients and healthy individuals. Results IEI patients had lower humoral and cellular immunogenicity than healthy individuals after three doses of COVID-19 vaccination. After the fourth dose, IEI patients achieved immunogenicity similar to that of healthy individuals who received three doses. A fourth dose of BNT162b2 significantly enhanced humoral and cellular immunogenicity against WT SARS-CoV-2 and humoral responses against JN.1 SARS-CoV-2. In contrast, CoronaVac had minimal effect on humoral responses to WT and JN.1 SARS-CoV-2. Six patients experienced breakthrough infections, which did not result in hospitalisation or death. Conclusion A fourth dose of BNT162b2 was immunogenic and safe for most IEI patients. The fourth dose of vaccination is recommended for IEI patients to improve protection against COVID-19, particularly prior to travel to areas with high endemic transmission.

Variant calling in segmental duplications is challenging for short-read sequencing because of ambiguous read origins. We present SDrecall, a method for sensitive variant detection in these regions. Upon constructing a network of homologous sequences, SDrecall realigns reads to each segmental duplication from its homologous counterparts. Realignments are phased and assembled into haplotypes via graph-based algorithms, followed by integer linear programming to retain the two most plausible haplotypes. Tested against long-read benchmarks, SDrecall achieved 95% sensitivity, while maintaining manageable false positives for short variants. SDrecall thus offers significant value for molecular diagnosis in terms of causal mutation detection within homologous regions.

Background: Patients with inborn errors of immunity (IEI) have high risks of severe complications after severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Although vaccination is effective in preventing severe coronavirus disease 2019 (COVID-19), boosters are required as the protection provided wanes over time. A greater number of boosters may be required in IEI patients compared to healthy individuals because of their immunodeficient state, but no such data are available. Objective: We investigated and compared the immunogenicity between 4 doses of COVID-19 vaccination for patients with IEI and 3 doses for healthy individuals. Methods: In the current study (NCT04800133), either BNT162b2 or CoronaVac was administered as dose 4. Healthy individuals who received 3 doses were included for comparison. Humoral and cellular immunogenicity against wild-type and JN.1 SARS-CoV-2 was assessed between IEI patients and healthy individuals. Results: IEI patients had lower humoral and cellular immunogenicity than healthy individuals after 3 doses of COVID-19 vaccination. After dose 4, IEI patients obtained immunogenicity similar to that of healthy individuals who received 3 doses. A fourth dose of BNT162b2 significantly enhanced humoral and cellular immunogenicity against wild-type SARS-CoV-2 and humoral responses against JN.1 SARS-CoV-2. In contrast, CoronaVac had minimal effect on humoral responses to wild-type and JN.1 SARS-CoV-2. Six patients experienced breakthrough infections, which did not result in hospitalization or death. Conclusion: A fourth dose of BNT162b2 was immunogenic and safe for most IEI patients. The fourth dose of vaccination is recommended for IEI patients to improve protection against COVID-19, particularly before travel to areas with high endemic transmission.

Background:The transcription factors signal transducer and activator of transcription 1 and 3 (STAT1 and STAT3) play essential roles in immune and non-immune cell function. The clinical characterization of patients carrying germline gain or loss-of-function (GOF or LOF) mutations in these genes has significantly improved our understanding of their physiological and pathological roles. Although patients with STAT3 LOF, STAT3 GOF, and STAT1 GOF mutations are classified into distinct inborn errors of immunity (IEI) categories, namely Hyper-IgE Syndrome, Regulatory T cell defects, and predisposition to Mucocutaneous Candidiasis, respectively, there is notable clinical overlap among these disorders. Case summary:We describe a 17-year-old girl with recurrent lung infection leading to bronchiectasis, chronic onychomycosis, recurrent vulvovaginal candidiasis, and oral thrush. Additional findings included short stature, delayed puberty, and retained primary teeth. Laboratory results revealed eosinophilia and elevated IgE serum levels, with a NIH HIES score of 53. A rare heterozygous deletion within the 3'UTR of the STAT3 gene (c.*351_*353del) was identified through a candidate gene approach. Although the variant is in a non-coding region, increased STAT3 phosphorylation and elevated suppressor of cytokine signaling 3 (SOCS3) expressions suggested a potential GOF effect. In silico analysis further predicted that the deletion disrupts microRNA (miRNA) binding sites and RNA binding proteins (RBP), potentially impairing post-transcriptional regulation and contributing to STAT3 overexpression. Given the complexity of the phenotype and the atypical location of the STAT3 variation, whole-exome sequencing (WES) was performed, revealing a heterozygous missense mutation in the STAT1 DNA-binding domain (c.1053G>T, p.L351F), previously reported in autosomal dominant chronic mucocutaneous candidiasis (AD-CMC). Functional assays on lymphocytes confirmed an increased STAT1 phosphorylation compared to both STAT1 LOF patient and healthy controls. Conclusion:This case highlights the diagnostic complexity of overlapping IEI phenotypes and the value of combining targeted and WES strategies. This dual molecular diagnosis, comprising a regulatory variant in STAT3 and a pathogenic coding mutation in STAT1, emphasizes the need to include non-coding regions in genetic analyses. It also underscores the value of using techniques that offer a broader genomic view and capture all coding exons, enabling a more comprehensive correlation with the clinical and immunological phenotype.

BackgroundCD19 is an essential component of a membrane protein complex on B cells, which also includes complement receptor 2 (CD21), CD81, and CD225. It amplifies B cell receptor (BCR) signaling by recruiting regulatory molecules and facilitating the phosphorylation of key kinases. Mutations in the CD19 gene disrupt the integrity of this complex and impair BCR signaling, ultimately leading to antibody deficiency.Purposewe report here a novel mutation in the CD19 gene in two patients from consanguineous Algerian kindred.MethodsWe conducted a comprehensive analysis of the clinical, genetic, and immunological characteristics of two siblings with CD19 deficiency.ResultsBoth siblings began experiencing upper and lower respiratory tract infections in early childhood. Over time, the older sibling developed recurrent fungal and viral skin infections, as well as episodes of pyelonephritis. Whole exome sequencing identified a novel homozygous mutation in the CD19 gene, leading to an out-of-frame translation predicted to trigger nonsense-mediated decay and result in absent gene expression. Flow cytometry revealed a complete absence of CD19 and reduced CD21 expression on CD20+ B cells in both siblings, while CD81 expression remained normal. Despite normal total peripheral B cell counts, the older patient exhibited reduced memory B cells. Additionally, both patients displayed circulating autoantibodies and an increased frequency of circulating follicular helper T cells.ConclusionThese findings highlight the critical role of CD19 not only in the initial activation of B lymphocytes by T-dependent antigens, but also in the maturation and/or selection of activated B cells within the memory compartment.

The calcium release-activated calcium modulator-2A (CRACR2A) protein plays a critical role in regulating store-operated calcium entry (SOCE) through CRAC channels, a process essential for numerous cellular functions, particularly T cell activation. Herein, we report the first Tunisian patient harboring a homozygous CRACR2A mutation. The patient is a 45-year-old female, born to consanguineous parents, who presented with lymph node and urinary tuberculosis, as well as recurrent fungal infections, including oral, genital, and urinary candidiasis, raising suspicion of an inborn error of immunity (IEI). HIV serology was negative. Immunophenotypic analysis revealed profound CD4+ and CD8+ T cell lymphopenia associated with a markedly decreased percentage of naïve CD4 T lymphocytes (CD4+ naïve T cells: 17.3%, normal: 33–66%). Lymphocyte proliferation to phytohemagglutinin (PHA) was impaired, and functional studies of IL-12/IFN-γ and TH17 pathways studies were normal, excluding other primary immunodeficiencies (PIDs). Whole-exome sequencing identified a homozygous missense variant in CRACR2A (c.1058G>A, p.Gly353Glu), predicted as disease-causing by MutationTaster. This variation was confirmed by Sanger sequencing. To the best of our knowledge, only one case of CRACR2A deficiency has been reported in the literature. The previously reported Asian patient presented with chronic diarrhea and recurrent pulmonary infections since adolescence, associated with severe hypogammaglobulinemia, progressive CD4 T cell lymphopenia, and a reduced proliferative response to mitogens, consistent with a late-onset combined immunodeficiency (LOCID). CRACR2A deficiency is presumed to result in functional impairment of T lymphocytes. Further immunological investigations are needed to better characterize the impact of the novel variant described herein. In conclusion, we describe the first Tunisian patient with a homozygous CRACR2A mutation, making her the second reported case worldwide. This finding broadens the clinical and immunological spectrum of CRACR2A-associated immunodeficiency and contributes to expanding the catalogue of genetic variants underlying this rare inborn error of immunity.

Signal transducer and activator of transcription 1 (STAT1) gene mutations have broad clinical phenotypes, classified by the inheritance pattern and functional state. Individuals with autosomal dominant STAT1 deficiency are more susceptible to intracellular bacteria, the hallmark of which is Mendelian susceptibility to mycobacterial diseases (MSMDs) that are associated with increased risks of invasive disease by weakly virulent mycobacteria. We report a novel de novo heterozygous missense mutation in exon 23 of the STAT1 gene (NM_007315.4):c.2129C>T(p.Ser710Phe) (S710F), located in the transactivation domain (TAD) for two Chinese siblings, whereby the index patient presented with multifocal osteomyelitis after Bacillus Calmette-Guerin (BCG) vaccine, while the younger sibling was spared the infection, as BCG vaccination was withheld at birth. STAT1 loss-of-function was confirmed by the gamma-activated sequence reporter assay, representing the first loss-of-function mutation in the TAD of the STAT1 gene. Both parents did not have the same mutation, and this finding is suggestive of gonadal mosaicism.

Background Some individuals may not retain adequate immunity against measles and rubella years after 2 doses of measles, mumps, and rubella (MMR) vaccination due to vaccine failure. This study aimed to investigate the rates of vaccine failure and seroconversion by administering an MMR booster to young adults.Methods We first assessed measles and rubella antibody levels using the Luminex multiplex assay, Vitek Immunodiagnostic Assay System (VIDAS) immunoglobulin G assay, and plaque reduction neutralization test among individuals aged 18-30 years who had received 2 doses of MMR vaccine. Participants with low measles and/or rubella antibody levels as confirmed by VIDAS received an MMR booster. Antibody levels were measured at 1 month postbooster.Results Among 791 participants, the measles and rubella seroprevalence rates were 94.7% (95% confidence interval [CI], 92.9%-96.0%) and 97.3% (95% CI, 96.0%-98.3%), respectively. Lower seroprevalence rates were observed among older participants. One hundred thirteen participants who received an MMR booster acquired higher measles and rubella antibody levels at 1 month postbooster compared to baseline.Conclusions Although measles and rubella vaccine failures were observed among 5.3% and 2.7% of young adults, respectively, an MMR booster triggered a significant antibody response. Young adults in Hong Kong had failure rates of 5.3% and 2.7% to measles and rubella vaccines, respectively. One month after an MMR booster, antibody levels against measles and rubella increased significantly, even for participants who had vaccine failures.

This report describes two brothers from India and a Chinese patient with somatic reversion of an inherited deleterious mutation in the WAS gene. Both the Indian siblings had inherited a single nucleotide deletion causing a frameshift mutation (c.1190del, p.Pro397Argfs*48) (variant 1: marked in blue) from the mother. Another variant (variant 2: marked in red), a 12-nucleotide deletion at position 1188-1199 (c.1188_1199del, p.P401_P404del) was also found, which resulted in restoration of the frame and subsequent rescue of the protein sequence. DNA sequencing from buccal mucosal cells revealed only the inherited variant (variant 1), while no reversion mutation was identified in the mucosal cells. Similarly, the Chinese patient was found to have a novel germline 14-base duplication (ACGAAAATGCTTGG) c.120_132 + 1dup (variant 1). This resulted in abolishment of the original splice junction coupled with the creation of a new junction 14 bases 3 ' and a frameshift mutation with predicted protein truncation p. Thr45Aspfs*. DNA from the patient's PBMC showed co-existence of wild-type and mutated sequences, but only the mutant was present in the buccal cells. Genomic and mRNA analysis of the isolated CD3+ T lymphocytes, CD3- mononuclear cells, and EBV-transformed B lymphocytes indicated that the reverant variant (germline variant was restored to wild-type sequence) were selectively found in CD3+ T lymphocytes.image

Sharma et al. define a new primary atopic disorder caused by heterozygous gain-of-function variants in STAT6. This results in severe, early-onset allergies, and is seen in 16 patients from 10 families. Anti–IL-4Rα antibody and JAK inhibitor treatment were highly effective.

Background Autosomal dominant signal transducer and activator of transcription 1 (STAT1) deficiency, part of the Mendelian susceptibility to mycobacterial disease (MSMD) group, frequently causes disseminated Bacillus Calmette-Guérin (BCG) infections, but has not been reported from Sub-Saharan Africa (SSA) where routine birth BCG vaccination is practiced. Case presentation Two half-siblings presented five years apart, with multifocal osteomyelitis as the dominant feature of disseminated BCG, which was successfully treated with antimycobacterial therapy. Whole exome sequencing demonstrated a novel heterozygous substitution in the splice site between intron 13 and exon 14 of the STAT1 gene, NM_007315: c.1128-1G>A, in the proband and his mother and was later confirmed in his half-brother. Conclusions Children with BCG vaccine complications in SSA should be referred for further investigation and particular consideration of MSMD.

Purpose Inborn errors of immunity (IEI) are typically monogenic. Data from the Indian subcontinent are relatively scarce. This paper evaluates IEI diagnosed in Sri Lanka. Methods Data of patients diagnosed with IEI from 2010 to 2022 at the Department of Immunology, Medical Research Institute, Colombo, Sri Lanka, were retrospectively analyzed. Results Two hundred and six patients were diagnosed with IEI, with a prevalence of 0.94 per 100,000. The onset of disease was below 12 years in 84.9%, whereas in 10.9%, it was after 18 years. The male: female ratio was 1.78:1. Consanguinity was identified in 26.6%. IEI were found in all but one (bone marrow failure) of the 10 IUIS categories. Predominantly antibody deficiencies were the most common category among the nine identified (30.1%), followed by combined immune deficiencies with syndromic features (21.3%), immunodeficiencies affecting cellular and humoral immunity (19.9%), congenital defects of phagocyte number or function (13.1%), and defects in intrinsic and innate immunity (8.2%). Severe combined immune deficiency (SCID) was the commonest disease (14.6%), followed by chronic granulomatous disease (CGD) (10.6%) and X linked agammaglobulinemia (8.7%). Of the patients with a known outcome ( n = 184), 51 died (27.7%). Mortality rates were high in SCID (83.3%), Omenn syndrome (OS) (100%), and CGD (31.8%) patients. Conclusion IEI in Sri Lanka are diagnosed mainly in childhood. The low diagnosis rates suggest a need for educating clinicians regarding IEI in adulthood. The high mortality rates associated with some IEI indicate the need of transplant services in the country.

Cryopyrin-associated periodic syndrome (CAPS) comprises a group of disorders characterized by recurrent bouts of systemic inflammation related to overactivation of inflammasome. So far, neither large cases of the correlation between genotype and phenotype nor treatment strategies have been clearly stated in China. Here, we studied the clinical and genetic characteristics and their correlation from 30 CAPS patients in China. We identified the pathogenesis for novel mutations by activating NLRP3 inflammasome for peripheral cells with ATP plus LPS, compared characteristics with other case series, and analyzed treatment outcomes of these patients. The patients harbored 19 substitutions in NLRP3, and 8 of them were novel mutations. Among these novel mutations, percentages of severe musculoskeletal, ophthalmologic, and neurological symptoms were higher compared with other case serials. The correlation of phenotypes and their variants seemed different in our cases, such as T350M, S333G/I/R, and F311V (somatic mosaicism). Ten patients received Canakinumab treatment, which proved effective at alleviating musculoskeletal, neurological, auditory, visual manifestations, fever, and rash for 10–20 months follow-up. Patients treated with prednisolone or prednisolone plus thalidomide or methotrexate, tocilizumab, TNF inhibiting agents, and sirolimus achieved only partial remission. Importantly, we firstly identified somatic mosaicism mutation of F311V, which was severe. Our study extended the spectrum of genotype and phenotype and characteristics of their correlations and provided detailed responses to different treatment strategies. These data provide guidance for future diagnosis and management for CAPS.

Hyper IgE syndromes (HIES) is a heterogeneous group of Inborn Errors of Immunity characterized by eczema, recurrent skin and lung infections associated with eosinophilia and elevated IgE levels. Autosomal dominant HIES caused by loss of function mutations in Signal transducer and activator of transcription 3 (STAT3) gene is the prototype of these disorders. Over the past two decades, advent in genetic testing allowed the identification of ten other etiologies of HIES. Although Dedicator of Cytokinesis 8 (DOCK8) deficiency is no more classified among HIES etiologies but as a combined immunodeficiency, this disease, characterized by severe viral infections, food allergies, autoimmunity, and increased risk of malignancies, shares some clinical features with STAT3 deficiency. The present study highlights the diagnostic challenge in eleven patients with the clinical phenotype of HIES in a resource-limited region. Candidate gene strategy supported by clinical features, laboratory findings and functional investigations allowed the identification of two heterozygous STAT3 mutations in five patients, and a bi-allelic DOCK8 mutation in one patient. Whole Exome Sequencing allowed to unmask atypical presentations of DOCK8 deficiency in two patients presenting with clinical features reminiscent of STAT3 deficiency. Our study underlies the importance of the differential diagnosis between STAT3 and DOCK8 deficiencies in order to improve diagnostic criteria and to propose appropriate therapeutic approaches. In addition, our findings emphasize the role of NGS in detecting mutations that induce overlapping phenotypes.

X-linked lymphoproliferative disease (XLP1) is an inborn error of immunity (IEI) with severe immune dysregulation caused by a mutation in the SH2D1A gene resulting in the absence or dysfunction of signaling lymphocytic activation molecule (SLAM)-associated protein (SAP). The severe acute respiratory syndrome (SARS) caused by SARS-coronavirus (CoV), a highly pathogenic CoV, has been shown to only cause mild diseases in Asian children. We report on a 5-year-old Nepalese boy with agammaglobulinemia and probable SARS who died of diffuse alveolar damage 22 days after admission amid the SARS outbreak. The index patient and his younger brother were genetically confirmed to have XLP1. In the current coronavirus disease 2019 (COVID-19) pandemic, most children also had mild disease only. Children with severe COVID-19 would warrant investigations for underlying IEI, particularly along the pathways leading to immune dysregulation.

Our study (NCT04800133) aimed to determine the safety and immunogenicity in patients with IEIs receiving a 3-dose primary series of mRNA vaccine BNT162b2 (age 12+) or inactivated whole-virion vaccine CoronaVac (age 3+) in Hong Kong, including Omicron BA.1 neutralization, in a nonrandomized manner. Intradermal vaccination was also studied. Thirty-nine patients were vaccinated, including 16 with homologous intramuscular 0.3ml BNT162b2 and 17 with homologous intramuscular 0.5ml CoronaVac. Two patients received 3 doses of intradermal 0.5ml CoronaVac, and 4 patients received 2 doses of intramuscular BNT162b2 and the third dose with intradermal BNT162b2. No safety concerns were identified. Inadequate S-RBD IgG and surrogate virus neutralization responses were found after 2 doses in patients with humoral immunodeficiencies and especially so against BA.1. Dose 3 of either vaccine increased S-RBD IgG response. T cell responses against SARS-CoV-2 antigens were detected in vaccinated IEI patients by intracellular cytokine staining on flow cytometry. Intradermal third dose vaccine led to high antibody response in 4 patients. The primary vaccination series of BNT162b2 and CoronaVac in adults and children with IEIs should include 3 doses for optimal immunogenicity.

Transmembrane 29 (Tmem29) gene with unknown function is a gene located on the X chromosome of the mouse genome. The gene showed differential expression in the Vannucci neonatal hypoxic-ischemic mouse brain model. We found the gene expresses with different molecular forms, including a group of long non-coding RNA forming a family of transcripts. It was predominantly expressed in the testes, brain, and kidney of mouse. In vitro identification and functional characterization were carried out in Neuro2a cells. Using fluorescence microscopy, Tmem29 protein was found to be constitutively expressed in mouse cell lines of different origins. Oxygen glucose deprivation (OGD) induced apoptotic cell death in Neuro2a cells and was confirmed by activations of caspase 3. Tmem29 protein was found to be associated with cell death especially at the time points of caspase 3 activations. A similar response was obtained in glucose deprivation (GD) cultures suggesting Tmem29 response to a common mechanism induced by OGD and GD. Downregulation of Tmem29 was induced by OGD and GD, further validating its response to hypoxia-ischemia (HI) insults. Our findings contributed to further understanding of molecular events after hypoxic-ischemic insults and opens new avenues for developing protective and therapeutic strategies for hypoxic-ischemic encephalopathy or even pathological programmed cell death.

To address inborn errors of immunity (IEI) which were underdiagnosed in resource-limited regions, our centre developed and offered free genetic testing for the most common IEI by Sanger sequencing (SS) since 2001. With the establishment of The Asian Primary Immunodeficiency (APID) Network in 2009, the awareness and definitive diagnosis of IEI were further improved with collaboration among centres caring for IEI patients from East and Southeast Asia. We also started to use whole exome sequencing (WES) for undiagnosed cases and further extended our collaboration with centres from South Asia and Africa. With the increased use of Next Generation Sequencing (NGS), we have shifted our diagnostic practice from SS to WES. However, SS was still one of the key diagnostic tools for IEI for the past two decades. Our centre has performed 2,024 IEI SS genetic tests, with in-house protocol designed specifically for 84 genes, in 1,376 patients with 744 identified to have disease-causing mutations (54.1%). The high diagnostic rate after just one round of targeted gene SS for each of the 5 common IEI (X-linked agammaglobulinemia (XLA) 77.4%, Wiskott–Aldrich syndrome (WAS) 69.2%, X-linked chronic granulomatous disease (XCGD) 59.5%, X-linked severe combined immunodeficiency (XSCID) 51.1%, and X-linked hyper-IgM syndrome (HIGM1) 58.1%) demonstrated targeted gene SS should remain the first-tier genetic test for the 5 common X-linked IEI.