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Purpose: CTLA4 deficiency is an inborn error of immunity (IEI) due to heterozygosity for germline loss-of-function variants of the CTLA4 gene located on chromosome 2q33.2. CTLA4 deficiency underlies pleiotropic immune and lymphoproliferation-mediated features with incomplete penetrance. It has been identified in hundreds of patients but copy number variants (CNVs) have been reported in only 12 kindreds, including nine which displayed large 2q33.1-2q33.2 deletions encompassing CTLA4., Methods: We conducted a nationwide study in France to identify patients with 2q33 deletions encompassing CTLA4. We investigated the clinical and immunological phenotypes and genotypes of these patients., Results: We identified 12 patients across six unrelated kindreds with clinical immunodeficiency. Neurological features were recorded in three patients, including one with syndromic neurodevelopmental disorder. Single-nucleotide polymorphism (SNP) or comparative genomic hybridization (CGH) array analysis, and targeted high-throughput sequencing revealed five different heterozygous 2q33 deletions of 26 kilobases to 7.12 megabases in size and encompassing one to 41 genes. We identified a contiguous gene syndrome (CGS) due to associated KLF7 deficiency in a kindred with a neurodevelopmental phenotype., Conclusion: Deletions within the 2q33 region encompassing CTLA4 are rare and not extensively explored, and are probably underdiagnosed in cytogenetic practice. A literature review identified 14 different CGS loci including at least one gene responsible for an IEI. The deletions involved in IEIs should be systematically delimited, to facilitate screening for CGS., Competing Interests: Declarations. Conflicts of Interest: The authors have no conflict of interest to declare. Ethics Approval: Informed consent for participation in this study was obtained in accordance with local regulations, with approval from the institutional review board (IRB). Consent to Participate: Written informed consent to participate was obtained from the patients or their parents. Consent for Publication: Consent to publish this report was obtained from the patients or their parents. All the authors approved the final version of the manuscript., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Purpose: Inborn errors of IFN-γ immunity underlie Mendelian susceptibility to mycobacterial disease (MSMD). Twenty-two genes with products involved in the production of, or response to, IFN-γ and variants of which underlie MSMD have been identified. However, pathogenic variants of IFNG encoding a defective IFN-γ have been described in only two siblings, who both underwent hematopoietic stem cell transplantation (HCST)., Methods: We characterized a new patient with MSMD by genetic, immunological, and clinical means. Therapeutic decisions were taken on the basis of these findings., Results: The patient was born to consanguineous Turkish parents and developed bacillus Calmette-Guérin (BCG) disease following vaccination at birth. Whole-exome sequencing revealed a homozygous private IFNG variant (c.224 T > C, p.F75S). Upon overexpression in recipient cells or constitutive expression in the patient's cells, the mutant IFN-γ was produced within the cells but was not correctly folded or secreted. The patient was treated for 6 months with two or three antimycobacterial drugs only and then for 30 months with subcutaneous recombinant IFN-γ1b plus two antimycobacterial drugs. Treatment with IFN-γ1b finally normalized all biological parameters. The patient presented no recurrence of mycobacterial disease or other related infectious diseases. The treatment was well tolerated, without the production of detectable autoantibodies against IFN-γ., Conclusion: We describe a patient with a new form of autosomal recessive IFN-γ deficiency, with intracellular, but not extracellular IFN-γ. IFN-γ1b treatment appears to have been beneficial in this patient, with no recurrence of mycobacterial infection over a period of more than 30 months. This targeted treatment provides an alternative to HCST in patients with complete IFN-γ deficiency or at least an option to better control mycobacterial infection prior to HCST., (© 2024. The Author(s).)

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., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Patients with autosomal dominant (AD) hyper-IgE syndrome (HIES) suffer from a constellation of manifestations including recurrent bacterial and fungal infections, severe atopy, and skeletal abnormalities. This condition is typically caused by monoallelic dominant-negative (DN) STAT3 variants. In 2020, we described 12 patients from eight kindreds with DN IL6ST variants resulting in a new form of AD HIES. These variants encoded truncated GP130 receptors, with intact extracellular and transmembrane domains, but lacking the intracellular recycling motif and the four STAT3-binding residues, resulting in an inability to recycle and activate STAT3. We report here two new DN variants of IL6ST in three unrelated families with HIES-AD. The biochemical and clinical impacts of these variants are different from those of the previously reported variants. The p.(Ser731Valfs*8) variant, identified in seven patients from two families, lacks the recycling motif and all the STAT3-binding residues, but its levels on the cell surface are only slightly increased and it underlies mild biological phenotypes with variable clinical expressivity. The p.(Arg768*) variant, identified in a single patient, lacks the recycling motif and the three most distal STAT3-binding residues. This variant accumulates at the cell surface and underlies severe biological and clinical phenotypes. The p.(Ser731Valfs*8) variant shows that a DN GP130 expressed at near normal levels on the cell surface can underlie heterogeneous clinical presentations, ranging from mild to severe. The p.(Arg768*) variant demonstrates that a truncated GP130 protein retaining one STAT3-binding residue can underlie severe HIES., (© 2023. The Author(s).)

Background: Cryptococcosis is a potentially life-threatening fungal disease caused by encapsulated yeasts of the genus Cryptococcus, mostly C. neoformans or C. gattii. Cryptococcal meningitis is the most frequent clinical manifestation in humans. Neutralizing autoantibodies (auto-Abs) against granulocyte-macrophage colony-stimulating factor (GM-CSF) have recently been discovered in otherwise healthy adult patients with cryptococcal meningitis, mostly caused by C. gattii. We hypothesized that three Colombian patients with cryptococcal meningitis caused by C. neoformans in two of them would carry high plasma levels of neutralizing auto-Abs against GM-CSF., Methods: We reviewed medical and laboratory records, performed immunological evaluations, and tested for anti-cytokine auto-Abs three previously healthy HIV-negative adults with disseminated cryptococcosis., Results: Peripheral blood leukocyte subset levels and serum immunoglobulin concentrations were within the normal ranges. We detected high levels of neutralizing auto-Abs against GM-CSF in the plasma of all three patients., Conclusions: We report three Colombian patients with disseminated cryptococcosis associated with neutralizing auto-Abs against GM-CSF. Further studies should evaluate the genetic contribution to anti-GM-CSF autoantibody production and the role of the GM-CSF signaling pathway in the immune response to Cryptococcus spp., (© 2023. The Author(s).)

Purpose: The first molecular evidence of a monogenic predisposition to mycobacteria came from the study of Mendelian susceptibility to mycobacterial disease (MSMD). We aimed to study this Mendelian susceptibility to mycobacterial diseases in Moroccan kindreds through clinical, immunological, and genetic analysis., Methods: Patients presented with clinical features of MSMD were recruited into this study. We used whole blood samples from patients and age-matched healthy controls. To measure IL-12 and IFN-γ production, samples were activated by BCG plus recombinant human IFN-γ or recombinant human IL-12. Immunological assessments and genetic analysis were also done for patients and their relatives., Results: Our study involved 22 cases from 15 unrelated Moroccan kindreds. The average age at diagnosis is 4 years. Fourteen patients (64%) were born to consanguineous parents. All patients were vaccinated with the BCG vaccine, and twelve of them (55%) developed locoregional or disseminated BCG infections. The other symptomatic patients had severe tuberculosis and/or recurrent salmonellosis. Genetic mutations were identified on the following genes: IL12RB1 in 8 patients, STAT1 in 7 patients; SPPL2A, IFNGR1, and TYK2 in two patients each; and TBX21 in one patient, with different modes of inheritance. All identified mutations/variants altered production or response to IFN-γ or both., Conclusion: Severe forms of tuberculosis and complications of BCG vaccination may imply a genetic predisposition present in the Moroccan population. In the presence of these infections, systematic genetic studies became necessary. BCG vaccination is contraindicated in MSMD patients and should be delayed in newborn siblings until the exclusion of a genetic predisposition to mycobacteria., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Purpose: Hypogammaglobulinemia in a context of lymphoma is usually considered as secondary and prior lymphoma remains an exclusion criterion for a common variable immunodeficiency (CVID) diagnosis. We hypothesized that lymphoma could be the revealing symptom of an underlying primary immunodeficiency (PID), challenging the distinction between primary and secondary hypogammaglobulinemia., Methods: Within a French cohort of adult patients with hypogammaglobulinemia, patients who developed a lymphoma either during follow-up or before the diagnosis of hypogammaglobulinemia were identified. These two chronology groups were then compared. For patients without previous genetic diagnosis, a targeted next-generation sequencing of 300 PID-associated genes was performed., Results: A total of forty-seven patients had developed 54 distinct lymphomas: non-Hodgkin B cell lymphoma (67%), Hodgkin lymphoma (26%), and T cell lymphoma (7%). In 25 patients, lymphoma developed prior to the diagnosis of hypogammaglobulinemia. In this group of patients, Hodgkin lymphoma was overrepresented compared to the group of patients in whom lymphoma occurred during follow-up (48% versus 9%), whereas MALT lymphoma was absent (0 versus 32%). Despite the histopathological differences, both groups presented with similar characteristics in terms of age at hypogammaglobulinemia diagnosis, consanguinity rate, or severe T cell defect. Overall, genetic analyses identified a molecular diagnosis in 10/47 patients (21%), distributed in both groups and without peculiar gene recurrence. Most of these patients presented with a late onset combined immunodeficiency (LOCID) phenotype., Conclusion: Prior or concomitant lymphoma should not be used as an exclusion criteria for CVID diagnosis, and these patients should be investigated accordingly., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Mendelian susceptibility to mycobacterial disease (MSMD) is a rare genetic disorder characterized by impaired immunity against intracellular pathogens, such as mycobacteria, attenuated Mycobacterium bovis-Bacillus Calmette-Guérin (BCG) vaccine strains, and environmental mycobacteria in otherwise healthy individuals. Retrospective study reviewed the clinical, immunological, and genetic characteristics of patients with MSMD in Mexico. Overall, 22 patients diagnosed with MSMD from 2006 to 2021 were enrolled: 14 males (64%) and eight females. After BCG vaccination, 12 patients (70%) developed BCG infection. Furthermore, 6 (22%) patients developed bacterial infections mainly caused by Salmonella, as what is described next in the text is fungal infections, particularly Histoplasma. Seven patients died of disseminated BCG disease. Thirteen different pathogenic variants were identified in IL12RB1 (n = 13), IFNGR1 (n = 3), and IFNGR2 (n = 1) genes. Interleukin-12Rβ1 deficiency is the leading cause of MSMD in our cohort. Morbidity and mortality were primarily due to BCG infection., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Background: Autosomal recessive (AR) PKCδ deficiency is a rare inborn error of immunity (IEI) characterized by autoimmunity and susceptibility to bacterial, fungal, and viral infections. PKCδ is involved in the intracellular production of reactive oxidative species (ROS)., Material and Methods: We studied a 5-year old girl presenting with a history of Burkholderia cepacia infection. She had no history of autoimmunity, lymphocyte counts were normal, and no auto-antibodies were detected in her plasma. We performed a targeted panel analysis of 407 immunity-related genes and immunological investigations of the underlying genetic condition in this patient., Results: Consistent with a history suggestive of chronic granulomatous disease (CGD), oxidative burst impairment was observed in the patient's circulating phagocytes in a dihydrorhodamine 123 (DHR) assay. However, targeted genetic panel analysis identified no candidate variants of known CGD-causing genes. Two heterozygous candidate variants were detected in PRKCD: c.285C > A (p.C95*) and c.376G > T (p.D126Y). The missense variant was also predicted to cause abnormal splicing, as it is located at the splice donor site of exon 5. TOPO-TA cloning confirmed that exon 5 was completely skipped, resulting in a truncated protein. No PKCδ protein was detected in the patient's neutrophils and monocyte-derived macrophages. The monocyte-derived macrophages of the patient produced abnormally low levels of ROS, as shown in an Amplex Red assay., Conclusion: PKCδ deficiency should be considered in young patients with CGD-like clinical manifestations and abnormal DHR assay results, even in the absence of clinical and biological manifestations of autoimmunity., (© 2022. The Author(s).)

Severe combined immunodeficiency, SCID, is a pediatric emergency that represents the most critical group of inborn errors of immunity (IEI). Affected infants present with early onset life-threatening infections due to absent or non-functional T cells. Without early diagnosis and curative treatment, most die in early infancy. As most affected infants appear healthy at birth, newborn screening (NBS) is essential to identify and treat patients before the onset of symptoms. Here, we report 47 Brazilian patients investigated between 2009 and 2020 for SCID due to either a positive family history and/or clinical impression and low TRECs. Based on clinical presentation, laboratory finding, and genetic information, 24 patients were diagnosed as typical SCID, 14 as leaky SCID, and 6 as Omenn syndrome; 2 patients had non-SCID IEI, and 1 remained undefined. Disease onset median age was 2 months, but at the time of diagnosis and treatment, median ages were 6.5 and 11.5 months, respectively, revealing considerable delay which affected negatively treatment success. While overall survival was 51.1%, only 66.7% (30/45) lived long enough to undergo hematopoietic stem-cell transplantation, which was successful in 70% of cases. Forty-three of 47 (91.5%) patients underwent genetic testing, with a 65.1% success rate. Even though our patients did not come from the NBS programs, the diagnosis of SCID improved in Brazil during the pilot programs, likely due to improved medical education. However, we estimate that at least 80% of SCID cases are still missed. NBS-SCID started to be universally implemented in the city of São Paulo in May 2021, and it is our hope that other cities will follow, leading to early diagnosis and higher survival of SCID patients in Brazil., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Background: Autosomal recessive (AR) complete IRF8 deficiency is a rare severe inborn error of immunity underlying an absence of blood myeloid mononuclear cells, intracerebral calcifications, and multiple infections. Only three unrelated patients have been reported., Materials and Methods: We studied an Argentinian child with multiple infectious diseases and severe pulmonary alveolar proteinosis (PAP). We performed whole-exome sequencing (WES) and characterized his condition by genetic, immunological, and clinical means., Results: The patient was born and lived in Argentina. He had a history of viral pulmonary diseases, disseminated disease due to bacillus Calmette-Guérin (BCG), PAP, and cerebral calcifications. He died at the age of 10 months from refractory PAP. WES identified two compound heterozygous variants in IRF8: c.55del and p.R111*. In an overexpression system, the p.R111* cDNA was loss-of-expression, whereas the c.55del cDNA yielded a protein with a slightly lower molecular weight than the wild-type protein. The mutagenesis of methionine residues downstream from c.55del revealed a re-initiation of translation. However, both variants were loss-of-function in a luciferase assay, suggesting that the patient had AR complete IRF8 deficiency. The patient had no blood monocytes or dendritic cells, associated with neutrophilia, and normal counts of NK and other lymphoid cell subsets., Conclusion: We describe the fourth patient with AR complete IRF8 deficiency. This diagnosis should be considered in children with PAP, which is probably due to the defective development or function of alveolar macrophages., (© 2022. The Author(s).)

Purpose: Germline heterozygous mutations of GATA2 underlie a variety of hematological and clinical phenotypes. The genetic, immunological, and clinical features of GATA2-deficient patients with mycobacterial diseases in the familial context remain largely unknown., Methods: We enrolled 15 GATA2 index cases referred for mycobacterial disease. We describe their genetic and clinical features including their relatives., Results: We identified 12 heterozygous GATA2 mutations, two of which had not been reported. Eight of these mutations were loss-of-function, and four were hypomorphic. None was dominant-negative in vitro, and the GATA2 locus was found to be subject to purifying selection, strongly suggesting a mechanism of haploinsufficiency. Three relatives of index cases had mycobacterial disease and were also heterozygous, resulting in 18 patients in total. Mycobacterial infection was the first clinical manifestation in 11 patients, at a mean age of 22.5 years (range: 12 to 42 years). Most patients also suffered from other infections, monocytopenia, or myelodysplasia. Strikingly, the clinical penetrance was incomplete (32.9% by age 40 years), as 16 heterozygous relatives aged between 6 and 78 years, including 4 older than 60 years, were completely asymptomatic., Conclusion: Clinical penetrance for mycobacterial disease was found to be similar to other GATA2 deficiency-related manifestations. These observations suggest that other mechanisms contribute to the phenotypic expression of GATA2 deficiency. A diagnosis of autosomal dominant GATA2 deficiency should be considered in patients with mycobacterial infections and/or other GATA2 deficiency-related phenotypes at any age in life. Moreover, all direct relatives should be genotyped at the GATA2 locus.

Purpose: Nocardiosis is a life-threatening infectious disease. We aimed at describing nocardiosis in patients with primary immunodeficiency diseases (PID)., Methods: This international retrospective cohort included patients with PID and nocardiosis diagnosed and/or published from Jan 1, 2000, to Dec 31, 2016. To identify nocardiosis cases, we analyzed PID databases from the French National Reference Center for PID (Paris, France) and the National Institute of Health (NIH, United States of America) and we performed a literature review on PubMed., Results: Forty-nine cases of nocardiosis associated with PID were included: median age at diagnosis of nocardiosis was 19 (0-56) years and most cases were observed among chronic granulomatous disease (CGD) patients (87.8%). Median time from symptoms to diagnosis of Nocardia infection was 20 (2-257) days. Most frequent clinical nocardiosis presentation was pneumonia (86.7%). Twelve-month mortality rate was 4.2%, and 11.9% of patients experienced a possible recurrence of infection. Nocardiosis more frequently led to the diagnosis of PID among non-CGD patients than in CGD patients. Non-CGD patients experienced more cerebral nocardiosis and more disseminated infections, but mortality and recurrence rates were similar. Highest incidences of nocardiosis among PID cohorts were observed among CGD patients (0.0057 and 0.0044 cases/patient-year in the USA and in France, respectively), followed by IL-12p40 deficiency., Conclusions: Among 49 cases of nocardiosis associated with PID, most patients had CGD and lung involvement. Both mortality and recurrence rates were low.

Mendelian susceptibility to mycobacterial diseases (MSMD) is a rare congenital condition characterized by a selective predisposition to infections caused by weakly virulent mycobacteria and other types of intra-macrophagic pathogens. The 16 genes associated with MSMD display a considerable level of allelic heterogeneity, accounting for 31 distinct disorders with variable clinical presentations and prognosis. Most of MSMD deficiencies are isolated, referred to as selective susceptibility to mycobacterial diseases. However, other deficiencies are syndromic MSMD, defined by the combination of the mycobacterial infection with another, equally common, infectious, specific phenotypes. Herein, we described a series of 32 Iranian MSMD cases identified with seven distinct types of molecular defects, all of which are involved in the interferon gamma (IFNγ) immunity, including interleukin IL-12 receptor-β1 (IL-12Rβ1) deficiency (fifteen cases), IL-12p40 deficiency (ten cases), and IL-23R deficiency (three cases), as well as IFNγ receptor 1 (IFNγR1) deficiency, IFNγ receptor 2 (IFNγR2) deficiency, interferon-stimulated gene 15 (ISG15) deficiency, and tyrosine kinase 2 (TYK2) deficiency each in one case. Since the first report of two MSMD patients in our center, we identified 30 other affected patients with similar clinical manifestations. As the number of reported Iranian cases with MSMD diagnosis has increased in recent years and according to the national vaccination protocol, all Iranian newborns receive BCG vaccination at birth, early diagnosis, and therapeutic intervention which are required for a better outcome and also prevention of similar birth defects. Therefore, we investigated the clinical and molecular features of these 32 patients. The current report also defined novel classes of pathological mutations, further expanding our knowledge of the MSMD molecular basis and associated clinical manifestations.

Down syndrome (DS) is characterized by the occurrence of three copies of human chromosome 21 (HSA21). HSA21 contains a cluster of four interferon receptor (IFN-R) genes: IFNAR1, IFNAR2, IFNGR2, and IL10RB. DS patients often develop mucocutaneous infections and autoimmune diseases, mimicking patients with heterozygous gain-of-function (GOF) STAT1 mutations, which enhance cellular responses to three types of interferon (IFN). A gene dosage effect at these four loci may contribute to the infectious and autoimmune manifestations observed in individuals with DS. We report high levels of IFN-αR1, IFN-αR2, and IFN-γR2 expression on the surface of monocytes and EBV-transformed-B (EBV-B) cells from studying 45 DS patients. Total and phosphorylated STAT1 (STAT1 and pSTAT1) levels were constitutively high in unstimulated and IFN-α- and IFN-γ-stimulated monocytes from DS patients but lower than those in patients with GOF STAT1 mutations. Following stimulation with IFN-α or -γ, but not with IL-6 or IL-21, pSTAT1 and IFN-γ activation factor (GAF) DNA-binding activities were significantly higher in the EBV-B cells of DS patients than in controls. These responses resemble the dysregulated responses observed in patients with STAT1 GOF mutations. Concentrations of plasma type I IFNs were high in 12% of the DS patients tested (1.8% in the healthy controls). Levels of type I IFNs, IFN-Rs, and STAT1 were similar in DS patients with and without recurrent skin infections. We performed a genome-wide transcriptomic analysis based on principal component analysis and interferon modules on circulating monocytes. We found that DS monocytes had levels of both IFN-α- and IFN-γ-inducible ISGs intermediate to those of monocytes from healthy controls and from patients with GOF STAT1 mutations. Unlike patients with GOF STAT1 mutations, patients with DS had normal circulating Th17 counts and a high proportion of terminally differentiated CD8 + T cells with low levels of STAT1 expression. We conclude a mild interferonopathy in Down syndrome leads to an incomplete penetrance at both cellular and clinical level, which is not correlate with recurrent skin bacterial or fungal infections. The constitutive upregulation of type I and type II IFN-R, at least in monocytes of DS patients, may contribute to the autoimmune diseases observed in these individuals.

Purpose: Chronic granulomatous disease (CGD) is a primary immunodeficiency characterized by an inability of phagocytes to produce reactive oxygen species, impairing their killing of various bacteria and fungi. We summarize here the 93 cases of CGD diagnosed in Mexico from 2011 to 2019., Methods: Thirteen Mexican hospitals participated in this study. We describe the genetic, immunological, and clinical features of the 93 CGD patients from 78 unrelated kindreds., Results: Eighty-two of the patients (88%) were male. All patients developed bacterial infections and 30% suffered from some kind of fungal infection. Fifty-four BCG-vaccinated patients (58%) presented infectious complications of BCG vaccine. Tuberculosis occurred in 29%. Granulomas were found in 56% of the patients. Autoimmune and inflammatory diseases were present in 15% of patients. A biological diagnosis of CGD was made in 89/93 patients, on the basis of NBT assay (n = 6), DHR (n = 27), and NBT plus DHR (n = 56). The deficiency was complete in all patients. The median age of biological diagnosis was 17 months (range, 0-186 months). A genetic diagnosis was made in 83/93 patients (when material was available), corresponding to CYBB (n = 64), NCF1 (n = 7), NCF2 (n = 7), and CYBA (n = 5) mutations., Conclusions: The clinical manifestations in these Mexican CGD patients were similar to those in patients elsewhere. This cohort is the largest in Latin America. Mycobacterial infections are an important cause of morbidity in Mexico, as in other countries in which tuberculosis is endemic and infants are vaccinated with BCG. X-linked CGD accounted for most of the cases in Mexico, as in other Latin American countries. However, a significant number of CYBA and NCF2 mutations were identified, expanding the spectrum of known causal mutations.

Purpose: Caspase-associated recruitment domain-9 (CARD9) deficiency is an inborn error of immunity that typically predisposes otherwise healthy patients to single fungal infections and the occurrence of multiple invasive fungal infections is rare. It has been described as the first known condition that predisposes to extrapulmonary Aspergillus infection with preserved lungs. We present a patient that expands the clinical variability of CARD9 deficiency., Materials and Methods: Genetic analysis was performed by Sanger sequencing. Neutrophils and mononuclear phagocyte response to fungal stimulation were evaluated through luminol-enhanced chemiluminescence and whole blood production of the proinflammatory mediator interleukin (IL)-6, respectively., Results: We report a 56-year-old Argentinean woman, whose invasive Exophiala spinifera infection at the age of 32 years was unexplained and reported in year 2004. At the age of 49 years, she presented with chronic pulmonary disease due to Aspergillus nomius. After partial improvement following treatment with caspofungin and posaconazole, right pulmonary bilobectomy was performed. Despite administration of multiple courses of antifungals, sustained clinical remission could not be achieved. We recently found that the patient's blood showed an impaired production of IL-6 when stimulated with zymosan. We also found that she is homozygous for a previously reported CARD9 loss-of-function mutation (Q289*)., Conclusions: This is the first report of a patient with inherited CARD9 deficiency and chronic invasive pulmonary aspergillosis (IPA) due to A. nomius. Inherited CARD9 deficiency should be considered in otherwise healthy children and adults with one or more invasive fungal diseases.

Blau syndrome (BS) is an auto-inflammatory granulomatous disease that possibly involves abnormal response to interferon gamma (IFNγ) due to exaggerated nucleotide-binding oligomerization domain containing 2 (NOD2) activity. Mendelian susceptibility to mycobacterial diseases (MSMD) is an infectious granulomatous disease that is caused by impaired production of or response to IFNγ. We report a mother and daughter who are both heterozygous for NOD2 c.2264C˃T variant and dominant-negative IFNGR1 818del4 mutation. The 17-year-old patient displayed an altered form of BS and milder form of MSMD, whereas the 44-year-old mother was completely asymptomatic. This experiment of nature supports the notion that IFNγ is an important driver of at least some BS manifestations and that elucidation of its involvement in the disease immunopathogenesis may identify novel therapeutic targets.

Live-attenuated vaccines (LAVs) can protect humans against 12 viral and three bacterial diseases. By definition, any clinical infection caused by a LAV that is sufficiently severe to require medical intervention attests to an inherited or acquired immunodeficiency that must be diagnosed or identified. Self-healing infections can also result from milder forms of immunodeficiency. We review here the inherited forms of immunodeficiency underlying severe infections of LAVs. Inborn errors of immunity (IEIs) underlying bacille Calmette-Guérin (BCG), oral poliovirus (OPV), vaccine measles virus (vMeV), and oral rotavirus vaccine (ORV) disease have been described from 1951, 1963, 1966, and 2009 onward, respectively. For each of these four LAVs, the underlying IEIs show immunological homogeneity despite genetic heterogeneity. Specifically, BCG disease is due to inborn errors of IFN-γ immunity, OPV disease to inborn errors of B cell immunity, vMeV disease to inborn errors of IFN-α/β and IFN-λ immunity, and ORV disease to adaptive immunity. Severe reactions to the other 11 LAVs have been described yet remain "idiopathic," in the absence of known underlying inherited or acquired immunodeficiencies, and are warranted to be the focus of research efforts. The study of IEIs underlying life-threatening LAV infections is clinically important for the affected patients and their families, as well as immunologically, for the study of the molecular and cellular basis of host defense against both attenuated and parental pathogens.

DOCK2 is a guanine-nucleotide-exchange factor for Rac proteins. Activated Rac serves various cellular functions including the reorganization of the actin cytoskeleton in lymphocytes and neutrophils and production of reactive oxygen species in neutrophils. Since 2015, six unrelated patients with combined immunodeficiency and early-onset severe viral infections caused by bi-allelic loss-of-function mutations in DOCK2 have been described. Until now, the function of phagocytes, specifically neutrophils, has not been assessed in human DOCK2 deficiency. Here, we describe a new kindred with four affected siblings harboring a homozygous splice-site mutation (c.2704-2 A > C) in DOCK2. The mutation results in alternative splicing and a complete loss of DOCK2 protein expression. The patients presented with leaky severe combined immunodeficiency or Omenn syndrome. The novel mutation affects EBV-B cell migration and results in NK cell dysfunction similar to previous observations. Moreover, both cytoskeletal rearrangement and reactive oxygen species production are partially impaired in DOCK2-deficient neutrophils.

Purpose: Mendelian susceptibility to mycobacterial disease (MSMD) is a rare primary immunodeficiency, triggered by non-tuberculous mycobacteria or Bacillus Calmette-Guérin (BCG) vaccines and characterized by severe diseases. All known genetic etiologies are inborn errors of IFN-γ-mediated immunity. Here, we report the molecular, cellular, and clinical features of patients from 15 Iranian families with disseminated disease without vaccination (2 patients) or following live BCG vaccination (14 patients)., Methods: We used whole blood samples from 16 patients and 12 age-matched healthy controls. To measure IL-12 and IFN-γ, samples were activated by BCG plus recombinant human IFN-γ or recombinant human IL-12. Immunological assessments and genetic analysis were also done for the patients., Results: Eight patients affected as a result of parental first-cousin marriages. Seven patients originated from multiplex kindred with positive history of death because of tuberculosis or finding the MSMD-related gene mutations. Two patients died due to mycobacterial disease at the ages of 8 months and 3.7 years. The remaining patients were alive at the last follow-up and were aged between 2 and 13 years. Patients suffered from infections including chronic mucocutaneous candidiasis (n = 10), salmonellosis (n = 2), and Leishmania (responsible for visceral form) (n = 2). Thirteen patients presented with autosomal recessive (AR) IL-12Rβ1 deficiency, meaning their cells produced low levels of IFN-γ. Bi-allelic IL12RB1 mutations were detected in nine of patients. Three patients with AR IL-12p40 deficiency (bi-allelic IL12B mutations) produced low levels of both IL-12 and IFN-γ. Overall, we found five mutations in the IL12RB1 gene and three mutations in the IL12B gene. Except one mutation in exon 5 (c.510C>A) of IL12B, all others were previously reported to be loss-of-function mutations., Conclusions: We found low levels of IFN-γ production and failure to respond to IL12 in 13 Iranian MSMD patients. Due to complicated clinical manifestations in affected children, early cellular and molecular diagnostics is crucial in susceptible patients.

Purpose: Inborn errors of IFN-γ-mediated immunity underlie Mendelian Susceptibility to Mycobacterial Disease (MSMD), which is characterized by an increased susceptibility to severe and recurrent infections caused by weakly virulent mycobacteria, such as Bacillus Calmette-Guérin (BCG) vaccines and environmental, nontuberculous mycobacteria (NTM)., Methods: In this study, we investigated four patients from four unrelated consanguineous families from Isfahan, Iran, with disseminated BCG disease. We evaluated the patients' whole blood cell response to IL-12 and IFN-γ, IL-12Rβ1 expression on T cell blasts, and sequenced candidate genes., Results: We report four patients from Isfahan, Iran, ranging from 3 months to 26 years old, with impaired IL-12 signaling. All patients suffered from BCG disease. One of them presented mycobacterial osteomyelitis. By Sanger sequencing, we identified three different types of homozygous mutations in IL12RB1. Expression of IL-12Rβ1 was completely abolished in the four patients with IL12RB1 mutations., Conclusions: IL-12Rβ1 deficiency was found in the four MSMD Iranian families tested. It is the first report of an Iranian case with S321* mutant IL-12Rβ1 protein. Mycobacterial osteomyelitis is another type of location of BCG infection in an IL-12Rβ1-deficient patient, notified for the first time in this study.

Purpose: CARD9 deficiency is an inborn error of immunity that predisposes otherwise healthy humans to mucocutaneous and invasive fungal infections, mostly caused by Candida, but also by dermatophytes, Aspergillus, and other fungi. Phaeohyphomycosis are an emerging group of fungal infections caused by dematiaceous fungi (phaeohyphomycetes) and are being increasingly identified in patients with CARD9 deficiency. The Corynespora genus belongs to phaeohyphomycetes and only one adult patient with CARD9 deficiency has been reported to suffer from invasive disease caused by C. cassiicola. We identified a Colombian child with an early-onset, deep, and destructive mucocutaneous infection due to C. cassiicola and we searched for mutations in CARD9., Methods: We reviewed the medical records and immunological findings in the patient. Microbiologic tests and biopsies were performed. Whole-exome sequencing (WES) was made and Sanger sequencing was used to confirm the CARD9 mutations in the patient and her family. Finally, CARD9 protein expression was evaluated in peripheral blood mononuclear cells (PBMC) by western blotting., Results: The patient was affected by a large, indurated, foul-smelling, and verrucous ulcerated lesion on the left side of the face with extensive necrosis and crusting, due to a C. cassiicola infectious disease. WES led to the identification of compound heterozygous mutations in the patient consisting of the previously reported p.Q289* nonsense (c.865C > T, exon 6) mutation, and a novel deletion (c.23_29del; p.Asp8Alafs10*) leading to a frameshift and a premature stop codon in exon 2. CARD9 protein expression was absent in peripheral blood mononuclear cells from the patient., Conclusion: We describe here compound heterozygous loss-of-expression mutations in CARD9 leading to severe deep and destructive mucocutaneous phaeohyphomycosis due to C. cassiicola in a Colombian child.

Purpose: Mendelian suceptibility to mycobacterial disease (MSMD) is a rare primary immunodeficiency predisposing to severe disease caused by mycobacteria and other intracellular pathogens. Delay in diagnosis can have an impact on the patient's prognosis., Methods: We evaluated the IFN-γ circuit by studying IFN-γ production after mycobacterial challenge as well as IL-12Rβ1 expression and STAT4 phosphorylation in response to IL-12p70 stimulation in whole blood of a 6-year-old Peruvian girl with disseminated recurrent mycobacterial infection diagnosed as multidrug-resistant tuberculosis. Genetic studies with Sanger sequencing were used to identify the causative mutation. Microbiological studies based on PCR reactions were used to diagnose the specific mycobacterial species., Results: We identified a homozygous mutation in the IL12RB1 gene (p. Arg211*) causing abolished expression of IL-12Rβ1 and IL-12 response. MSMD diagnosis led to a microbiological reevaluation of the patient, revealing a BCG vaccine-related infection instead of tuberculosis. Treatment was then adjusted, with good response., Conclusions: We report the first Peruvian patient with IL-12Rβ1 deficiency. Specific mycobacterial species diagnosis within Mycobacterium tuberculosis complex is still challenging in countries with limited access to PCR-based microbiological diagnostic techniques. Awareness of MSMD warning signs and accurate microbiological diagnosis of mycobacterial infections are of the utmost importance for optimal diagnosis and management of affected patients.

Purpose: Inborn errors of IFN-γ immunity underlie Mendelian susceptibility to mycobacterial disease (MSMD). Autosomal recessive complete IL-12Rβ1 deficiency is the most frequent genetic etiology of MSMD. Only two of the 84 known mutations are copy number variations (CNVs), identified in two of the 213 IL-12Rβ1-deficient patients and two of the 164 kindreds reported. These two CNVs are large deletions found in the heterozygous or homozygous state. We searched for novel families with IL-12Rβ1 deficiency due to CNVs., Methods: We studied six MSMD patients from five unrelated kindreds displaying adverse reactions to BCG vaccination. Three of the patients also presented systemic salmonellosis, two had mucocutaneous candidiasis, and one had disseminated histoplasmosis. We searched for CNVs and other variations by IL12RB1-targeted next-generation sequencing (NGS)., Results: We identified six new IL-12Rβ1-deficient patients with a complete loss of IL-12Rβ1 expression on phytohemagglutinin-activated T cells and/or EBV-transformed B cells. The cells of these patients did not respond to IL-12 and IL-23. Five different CNVs encompassing IL12RB1 (four deletions and one duplication) were identified in these patients by NGS coverage analysis, either in the homozygous state (n = 1) or in trans (n = 4) with a single-nucleotide variation (n = 3) or a small indel (n = 1). Seven of the nine mutations are novel. Interestingly, four of the five CNVs were predicted to be driven by nearby Alu elements, as well as the two previously reported large deletions. The IL12RB1 locus is actually enriched in Alu elements (44.7%), when compared with the rest of the genome (10.5%)., Conclusion: The IL12RB1 locus is Alu-enriched and therefore prone to rearrangements at various positions. CNVs should be considered in the genetic diagnosis of IL-12Rβ1 deficiency.

The pathogenesis of life-threatening influenza A virus (IAV) disease remains elusive, as infection is benign in most individuals. We studied two relatives who died from influenza. We Sanger sequenced GATA2 and evaluated the mutation by gene transfer, measured serum cytokine levels, and analyzed circulating T- and B-cells. Both patients (father and son, P1 and P2) died in 2011 of H1N1pdm IAV infection at the ages of 54 and 31 years, respectively. They had not suffered from severe or moderately severe infections in the last 17 (P1) and 15 years (P2). A daughter of P1 had died at 20 years from infectious complications. Low B-cell, NK- cell, and monocyte numbers and myelodysplastic syndrome led to sequence GATA2. Patients were heterozygous for a novel, hypomorphic, R396L mutation leading to haplo-insufficiency. B- and T-cell rearrangement in peripheral blood from P1 during the influenza episode showed expansion of one major clone. No T-cell receptor excision circles were detected in P1 and P3 since they were 35 and 18 years, respectively. Both patients presented an exuberant, interferon (IFN)-γ-mediated hypercytokinemia during H1N1pdm infection. No data about patients with viremia was available. Two previously reported adult GATA2-deficient patients died from severe H1N1 IAV infection; GATA2 deficiency may predispose to life-threatening influenza in adulthood. However, a role of other genetic variants involved in immune responses cannot be ruled out. Patients with GATA2 deficiency can reach young adulthood without severe infections, including influenza, despite long-lasting complete B-cell and natural killer (NK) cell deficiency, as well as profoundly diminished T-cell thymic output.

Purpose: Mendelian susceptibility to mycobacterial disease (MSMD) is a rare primary immunodeficiency predisposing congenitally affected individuals to diseases caused by weakly virulent mycobacteria, such as Bacillus Calmette-Guérin (BCG) vaccine strains and environmental mycobacteria. IL-12p40 deficiency is a genetic etiology of MSMD resulting in impaired IL-12- and IL-23-dependent IFN-γ immunity. Most of the reported patients with IL-12p40 deficiency originate from Saudi Arabia (30 of 52) and carry the recurrent IL12B mutation c.315insA (27 of 30)., Methods: Whole-exome sequencing was performed on three patients from two unrelated kindreds from Saudi Arabia with disseminated disease caused by a BCG vaccine substrain., Results: Genetic analysis revealed a homozygous mutation, p.W60X, in exon 3 of the IL12B gene, resulting in complete IL12p40 deficiency. This mutation is recurrent due to a new founder effect., Conclusions: This report provides evidence for a second founder effect for recurrent mutations of IL12B in Saudi Arabia.

Purpose: Mendelian susceptibility to mycobacterial disease is a rare clinical condition characterized by a predisposition to infectious diseases caused by poorly virulent mycobacteria. Other infections such as salmonellosis and candidiasis are also reported. The purpose of this article is to describe a young boy affected with various infectious diseases caused by Mycobacterium tuberculosis complex, Salmonella sp, Klebsiella pneumonie, Citrobacter sp., and Candida sp, complicated with severe enteropathy and transient hypogammaglobulinemia., Methods: We reviewed medical records and performed flow cytometry staining for lymphocyte populations, lymphocyte proliferation in response to PHA, and intracellular IFN-γ production in T cell PHA blasts in the patient and a healthy control. Sanger sequencing was used to confirm the genetic variants in the patient and relatives., Results: Genetic analysis revealed a bi-allelic mutation in IL12RB1 (C291Y) resulting in complete IL-12Rβ1 deficiency. Functional analysis demonstrated the lack of intracellular production of IFN-γ in CD3+ T lymphocytes from the patient in response to rhIL-12p70., Conclusions: To our knowledge, this is the third patient with MSMD due to IL-12Rβ1 deficiency complicated with enteropathy and hypogammaglobulinemia and the first case of this disease to be described in Colombia.

Introduction: Chronic granulomatous disease (CGD) is an inherited mutational defect in any of the NADPH oxidase complex, CYBB (gp91-phox), NCF1 (p47-phox), CYBA (p22-phox), NCF2 (p67-phox), or NCF4 (p40-phox) leading to inability of phagocytes to perform effective respiratory burst and thus diminished killing of bacteria and fungi. The identification of defective proteins aids in establishing a diagnosis prior to genetic analysis, which is rather labor-intensive, expensive, and time-consuming., Aim: The present study aims at assessing the NADPH proteins by performing the intracellular staining with specific monoclonal antibodies and their assessment on flow cytometry. The use of flow cytometry is less laborious and faster to perform than western blot. It also confirms the diagnosis of CGD and detects the affected components allowing proper management of patients., Materials and Methods: Twenty-eight patients from 25 different kindred, clinically suspected as CGD were recruited in Egypt. Dihydrorhodamine test was performed to confirm the diagnosis of the patients. Intracellular staining of NADPH components using specific monoclonal antibodies was performed followed by flow cytometric analysis., Results: The present study revealed that the most common defective protein in our cohort is p22-phox, found in 13 patients (46.4 % of cases) followed by p47-phox in 8 patients (28.6 %), gp91-phox in 5 patients (17.9 %), and finally p67-phox in 2 patients (7.1 %)., Conclusion: In countries with limited resources and yet large number of CGD patients, the analysis of the defective proteins by flow cytometry is an optimum solution for confirming the diagnosis and is a step for targeted sequencing in families seeking prenatal diagnosis.

Mendelian susceptibility to mycobacterial disease (MSMD) is a rare condition characterized by clinical disease caused by weakly virulent mycobacteria. All genes mutated in MSMD patients are involved in IFN-γ immunity. Autosomal partial dominant (PD) interferon-γ receptor 1 (IFN-γR1) deficiency is the most frequent abnormality affecting the group of MSMD patients leading to impaired response of IFN-γ. We describe here a patient from India with disseminated infection due to Mycobacterium avium intracellulare (MAC) including multifocal osteomyelitis and BCG disease. A heterozygous mutation in exon 6 of IFNGR1 gene was identified, conferring an autosomal PD IFN-γR1 deficiency. Patient had recurrence of mycobacterial disease during antibiotic therapy for which subcutaneous IFN-γ was added as a modality of treatment for resistant MAC infection.

Interferon-γ receptor 2 (IFN-γR2) deficiency is a rare primary immunodeficiency characterized by predisposition to infections with weakly virulent mycobacteria, such as environmental mycobacteria and BCG vaccines. We describe here two children with IFN-γR2 deficiency, from unrelated, consanguineous kindreds of Arab and Israeli descent. The first patient was a boy who died at the age of 4.5 years, from recurrent, disseminated disease caused by Mycobacterium simiae. His IFN-γR2 defect was autosomal recessive and complete. The second patient was a girl with multiple disseminated mycobacterial infections, including infection with M. simiae. She died at the age of 5 years, a short time after the transplantation of umbilical cord blood cells from an unrelated donor. Her IFN-γR2 defect was autosomal recessive and partial. Autosomal recessive IFN-γR2 deficiency is life-threatening, even in its partial form, and genetic diagnosis and familial counseling are therefore particularly important for this condition. These two cases are the first of IFN-γR2 deficiency associated with M. simiae infection to be described.