Reversion of neurovirulent mutations, recombination and high intra-host diversity in vaccine-derived poliovirus excreted by patients with primary immune deficiency.

Patients with Primary immunodeficiency (PIDs) may be infected by Polioviruses (PVs), especially when vaccinated with live Oral Polio Vaccine before diagnosis. They may establish long-term shedding of divergent strains and may act as reservoirs of PV transmission. This study delved into the effect of the genetic evolution of complete PV genomes, from MHC class II-deficient patients, on the excretion duration and clinical outcomes. Stool samples from three PID patients underwent analysis for PV detection through inoculation on cell culture and real-time PCR, followed by VP1 partial sequencing and full genome sequencing using the Illumina technology. Our findings revealed a low number of mutations for one patient who cleared the virus, while two exhibited a high intra-host diversity favoring the establishment of severe outcomes. Neurovirulence-reverse mutations were detected in two patients, possibly leading to paralysis development. Furthermore, a recombination event, between type 3 Vaccine-Derived Poliovirus and Sabin-like1 (VDPV3/SL1), occurred in one patient. Our findings have suggested an association between intra-host diversity, recombination, prolonged excretion of the virus, and emergence of highly pathogenic strains. Further studies on intra-host diversity are crucial for a better understanding of the virus evolution as well as for the success of the Global Polio Eradication Initiative.

Clinical, immunological, molecular and therapeutic findings in monogenic immune dysregulation diseases: Middle East and North Africa registry.

Monogenic immune dysregulation diseases (MIDD) are caused by defective immunotolerance. This study was designed to increase knowledge on the prevalence and spectrum of MIDDs, genetic patterns, and outcomes in Middle East and North Africa (MENA). MIDD patients from 11 MENA countries (Iran, Turkey, Kuwait, Oman, Algeria, Egypt, United Arab Emirates, Tunisia, Jordan, Qatar, and Azerbaijan) were retrospectively evaluated. 343 MIDD patients (58% males and 42% female) at a median (IQR) age of 101 (42-192) months were enrolled. The most common defective genes were LRBA (23.9%), LYST (8.2%), and RAB27A (7.9%). The most prevalent initial and overall manifestations were infections (32.2% and 75.1%), autoimmunity (18.6% and 41%), and organomegaly (13.3% and 53.8%), respectively. Treatments included immunoglobulin replacement therapy (53%), hematopoietic stem cell transplantation (HSCT) (14.3%), immunosuppressives (36.7%), and surgery (3.5%). Twenty-nine (59.2%) patients survived HSCT. Along with infectious complications, autoimmunity and organomegaly may be the initial or predominant manifestations of MIDD.

A Novel Point-of-Care Rapid Diagnostic Test for Screening Individuals for Antibody Deficiencies.

PURPOSE: No rapid diagnostic test exists to screen individuals for primary antibody deficiencies (PAD) at or near the point of care. In settings at risk for polio where live oral polio vaccine is utilized, undiagnosed PAD patients and cases with delayed diagnosis constitute a potential reservoir for neurovirulent polioviruses, undermining polio eradication. This research aimed to develop a rapid screening test suited for use in resource-limited settings to identify individuals with low immunoglobulin G (IgG) levels, enabling early diagnosis and appropriate treatment. METHODS: Three prototype tests distinguishing low and normal IgG levels were evaluated with a blinded panel of serum/plasma specimens from 32 healthy controls and 86 primary immunodeficiency-confirmed patients with agammaglobulinemia, common variable immunodeficiency, and hyper-IgM syndrome, including 57 not receiving IgG therapy. Prototype tests were compared to laboratory reference and clinical case definition. RESULTS: The leading prototype correctly identified 32 of 32 healthy controls. Among primary antibody deficiency patients not receiving IgG treatment, 17 of 19 agammaglobulinemia, 7 of 24 common variable immunodeficiency, and 5 of 14 hyper-IgM were correctly identified by the prototype, with 67% agreement with the reference assay. CONCLUSION: The Rapid IgG Screen (RIgGS) test can differentiate between low IgG levels associated with agammaglobulinemia and normal IgG antibody levels. Differentiating CVID and hyper IgM was challenging due to the wide range in IgG levels and influence of high IgM. This test can facilitate the identification of patients with primary antibody deficiencies and support polio surveillance initiatives.

Profound differences in IgE and IgG recognition of micro-arrayed allergens in hyper-IgE syndromes.

BACKGROUND: The specificities of IgE and IgG for allergen molecules in patients with inborn errors of immunity (IEI) have not been investigated in detail. OBJECTIVE: To study IgE and IgG antibody specificities in patients with defined hyper-IgE syndromes (HIES) using a comprehensive panel of allergen molecules. METHODS: We used chips containing micro-arrayed allergen molecules to analyze allergen-specific IgE and IgG levels in sera from two groups of HIES patients: Autosomal recessive mutations in phosphoglucomutase-3 (PGM3); Autosomal dominant negative mutations of STAT3 (STAT3); and age-matched subjects with allergic sensitizations. Assays with rat basophil leukemia cells transfected with human FcεRI were performed to study the biological relevance of IgE sensitizations. RESULTS: Median total IgE levels were significantly lower in the sensitized control group (212.9 kU/L) as compared to PGM3 (5042 kU/L) and STAT3 patients (2561 kU/L). However, PGM3 patients had significantly higher allergen-specific IgE levels and were sensitized to a larger number of allergen molecules as compared to STAT3 patients. Biological relevance of IgE sensitization was confirmed for PGM3 patients by basophil activation testing. PGM3 patients showed significantly lower cumulative allergen-specific IgG responses in particular to milk and egg allergens as compared to STAT3 patients and sensitized controls whereas total IgG levels were comparable to STAT3 patients and significantly higher than in controls. CONCLUSION: The analysis with multiple micro-arrayed allergen molecules reveals profound differences of allergen-specific IgE and IgG recognition in PGM3 and STAT3 patients which may be useful for classification of IEI and clinical characterization of patients.

Genetic Approaches for Definitive Diagnosis of Agammaglobulinemia in Consanguineous Families.

Autosomal recessive agammaglobulinemia (ARA) is a primary immunodeficiency characterized by absent peripheral B cells, severe hypogammaglobulinemia, and absent BTK gene mutations. In ARA, mutations occur in genes encoding the pre-B cell receptor (pre-BCR) or downstream signaling proteins. In this work, we used candidate gene and whole-exome sequencing to investigate the molecular basis of ARA in 6 patients from 4 consanguineous North-African families. Sanger sequencing of candidate genes encoding the pre-BCR components (ΙGΗΜ, CD79A, CD79B, IGLL1, and VPREB1) was initially performed and determined the genetic defect in five patients. Two novel mutations in IGHM (p.Val378Alafs*1 and p.Ile184Serfs*21) were identified in three patients from two unrelated kindred and a novel nonsense mutation was identified in CD79A (p.Trp66*) in two siblings from a third kindred. Whole-exome sequencing (WES) was performed on the sixth patient who harbored a homozygous stop mutation at position 407 in the RAG2 gene (p.Glu407*). We concluded that conventional gene sequencing, especially when multiple genes are involved in the defect as is the case in ARA, is costly and time-consuming, resulting in delayed diagnosis that contributes to increased morbidity and mortality. In addition, it fails to identify the involvement of novel and unsuspected gene defects when the phenotype of the patients is atypical. WES has the potential to provide a rapid and more accurate genetic diagnosis in ARA, which is crucial for the treatment of the patients.

A Founder Effect of c.257 + 2T > C Mutation in NCF2 Gene Underlies Severe Chronic Granulomatous Disease in Eleven Patients.

Chronic granulomatous disease (CGD) is the prototypic functional neutrophil disorder caused by genetic defects in one of the five genes encoding the superoxide-generating nicotinamide adenine dinucleotide phosphate (NADPH)-oxidase subunits of phagocytes. Mutations causing the most prevalent form of CGD in western populations are located in the X-linked-CYBB gene. The four remaining autosomal recessive (AR) forms collectively account for one-third of CGD cases. We investigated the clinical and molecular features of eleven patients with CGD from 6 consanguineous families, originating from contiguous regions in the west of Tunisia. The patients' clinical phenotype is characterized by a high incidence of mycobacterial infections. Five out of the eleven patients died despite treatment arguing in favor of a severe clinical form of CGD. These findings correlated with the absence of functional p67phox protein as well as the absence of residual reactive oxygen intermediates (ROI) production. Genetic analysis showed the presence, in all patients, of a unique mutation (c.257 + 2T > C) in NCF2 gene predicted to affect RNA splicing. Segregating analysis using nine polymorphic markers overlapping the NCF2 gene revealed a common haplotype spanning 4.1 Mb. The founder event responsible for this mutation was estimated to have arisen approximately 175 years ago. These findings will facilitate the implementation of preventive approaches through genetic counseling in affected consanguineous families.

Report of the Tunisian Registry of Primary Immunodeficiencies: 25-Years of Experience (1988-2012).

PURPOSE: Primary immunodeficiencies (PIDs) are a large group of diseases characterized by susceptibility to not only recurrent infections but also autoimmune diseases and malignancies. The aim of this study was to describe and analyze the distribution, clinical features and eventual outcome of PID among Tunisian patients. METHODS: We reviewed the record of 710 patients diagnosed with Primary Immunodeficiency Diseases (PIDs) from the registry of the Tunisian Referral Centre for PIDs over a 25-year period. RESULTS: The male-to-female ratio was 1.4. The median age at the onset of symptoms was 6 months and at the time of diagnosis 2 years. The estimated prevalence was 4.3 per 100,000 populations. The consanguinity rate was found in 58.2 % of families. According to the International Union of Immunological Societies classification, spectrums of PIDs were as follows: combined T-cell and B-cell immunodeficiency disorders account for the most common category (28.6 %), followed by congenital defects of phagocyte (25.4 %), other well-defined immunodeficiency syndromes (22.7 %), predominant antibody deficiency diseases (17.7 %), diseases of immune dysregulation (4.8 %), defect of innate immunity (0.4 %) and complement deficiencies (0.4 %). Recurrent infections, particularly lower airway infections (62.3 %), presented the most common manifestation of PID patients. The overall mortality rate was 34.5 %, mainly observed with combined immunodeficiencies. CONCLUSION: The distribution of PIDs was different from that reported in Western countries, with a particularly high proportion of Combined Immunodeficiencies and phagocyte defects in number and/or function. More is needed to improve PID diagnosis and treatment in our country.

A 1,100-year-old founder effect mutation in IL12B gene is responsible for Mendelian susceptibility to mycobacterial disease in Tunisian patients.

Mendelian susceptibility to mycobacterial disease (MSMD) is a rare disorder predisposing apparently healthy individuals to infections caused by weakly virulent mycobacteria such as bacille Calmette-Guerin (BCG), environmental mycobacteria, and poorly virulent Salmonella strains. IL-12p40 deficiency is the first reported human disease due to a cytokine gene defect and is one of the deficiencies that cause MSMD. Nine mutant alleles only have been identified in the IL12B gene, and three of them are recurrent mutations due to a founder effect in specific populations. IL-12p40 deficiency has been identified especially in countries where consanguinity is high and where BCG vaccination at birth is universal. We investigated, in such settings, the clinical, cellular, and molecular features of six IL-12p40-deficient Tunisian patients having the same mutation in IL12B gene (c.298_305del). We found that this mutation is inherited as a common founder mutation arousing ~1,100 years ago. This finding facilitates the development of a preventive approach by genetic counseling and prenatal diagnosis especially in affected families.