Mutational landscape of severe combined immunodeficiency patients from Turkey.

Severe combined immunodeficiency (SCID) has a diverse genetic aetiology, where a clinical phenotype, caused by single and/or multiple gene variants, can give rise to multiple presentations. The advent of next-generation sequencing (NGS) has recently enabled rapid identification of the molecular aetiology of SCID, which is crucial for prognosis and treatment strategies. We sought to identify the genetic aetiology of various phenotypes of SCIDs and assessed both clinical and immunologic characteristics associated with gene variants. An amplicon-based targeted NGS panel, which contained 18 most common SCID-related genes, was contumely made to screen the patients (n = 38) with typical SCID, atypical SCID or OMENN syndrome. Allelic segregations were confirmed for the detected gene variants within the families. In total, 24 disease-causing variants (17 known and 7 novel) were identified in 23 patients in 9 different SCID genes: RAG1 (n = 5), RAG2 (n = 2), ADA (n = 3), DCLRE1C (n = 2), NHEJ1 (n = 2), CD3E (n = 2), IL2RG (n = 3), JAK3 (n = 4) and IL7R (n = 1). The overall success rate of our custom-made NGS panel was 60% (39.3% for NK+ SCID and 100% for NK- SCID). Incidence of autosomal-recessive inherited genes is more frequently found in our cohort than the previously reported populations probably due to the high consanguineous marriages in Turkey. In conclusion, the custom-made sequencing panel was able to identify and confirm the previously known and novel disease-causing variants with high accuracy.

Type I IFN-related NETosis in ataxia telangiectasia and Artemis deficiency.

BACKGROUND: Pathological inflammatory syndromes of unknown etiology are commonly observed in ataxia telangiectasia (AT) and Artemis deficiency. Similar inflammatory manifestations also exist in patients with STING-associated vasculopathy in infancy (SAVI). OBJECTIVE: We sought to test the hypothesis that the inflammation-associated manifestations observed in patients with AT and Artemis deficiency stem from increased type I IFN signature leading to neutrophil-mediated pathological damage. METHODS: Cytokine/protein signatures were determined by ELISA, cytometric bead array, or quantitative PCR. Stat1 phosphorylation levels were determined by flow cytometry. DNA species accumulating in the cytosol of patients' cells were quantified microscopically and flow cytometrically. Propensity of isolated polymorhonuclear granulocytes to form neutrophil extracellular traps (NETs) was determined using fluorescence microscopy and picogreen assay. Neutrophil reactive oxygen species levels and mitochondrial stress were assayed using fluorogenic probes, microscopy, and flow cytometry. RESULTS: Type I and III IFN signatures were elevated in plasma and peripheral blood cells of patients with AT, Artemis deficiency, and SAVI. Chronic IFN production stemmed from the accumulation of DNA in the cytoplasm of AT and Artemis-deficient cells. Neutrophils isolated from patients spontaneously produced NETs and displayed indicators of oxidative and mitochondrial stress, supportive of their NETotic tendencies. A similar phenomenon was also observed in neutrophils from healthy controls exposed to patient plasma samples or exogeneous IFN-α. CONCLUSIONS: Type I IFN-mediated neutrophil activation and NET formation may contribute to inflammatory manifestations observed in patients with AT, Artemis deficiency, and SAVI. Thus, neutrophils represent a promising target to manage inflammatory syndromes in diseases with active type I IFN signature.

DCLRE1C (ARTEMIS) mutations causing phenotypes ranging from atypical severe combined immunodeficiency to mere antibody deficiency.

Null mutations in genes involved in V(D)J recombination cause a block in B- and T-cell development, clinically presenting as severe combined immunodeficiency (SCID). Hypomorphic mutations in the non-homologous end-joining gene DCLRE1C (encoding ARTEMIS) have been described to cause atypical SCID, Omenn syndrome, Hyper IgM syndrome and inflammatory bowel disease-all with severely impaired T-cell immunity. By whole-exome sequencing, we investigated the molecular defect in a consanguineous family with three children clinically diagnosed with antibody deficiency. We identified perfectly segregating homozygous variants in DCLRE1C in three index patients with recurrent respiratory tract infections, very low B-cell numbers and serum IgA levels. In patients, decreased colony survival after irradiation, impaired proliferative response and reduced counts of naïve T cells were observed in addition to a restricted T-cell receptor repertoire, increased palindromic nucleotides in the complementarity determining regions 3 and long stretches of microhomology at switch junctions. Defective V(D)J recombination was complemented by wild-type ARTEMIS protein in vitro. Subsequently, homozygous or compound heterozygous DCLRE1C mutations were identified in nine patients from the same geographic region. We demonstrate that DCLRE1C mutations can cause a phenotype presenting as only antibody deficiency. This novel association broadens the clinical spectrum associated with ARTEMIS mutations. Clinicians should consider the possibility that an immunodeficiency with a clinically mild initial presentation could be a combined immunodeficiency, so as to provide appropriate care for affected patients.

Hematopoietic stem cell transplantation from unrelated donors in children with DOCK8 deficiency.

DIDS is a unique form of combined immune deficiency characterized by an unusual susceptibility to cutaneous viral infections, severe allergies with eosinophilia and elevated immunoglobulin E titers, autoimmunity, and cancer. HSCT is considered the standard of care for this deadly disease. We have retrospectively analyzed the outcome of allogeneic HSCT from unrelated donors in patients with DIDS. Data from four patients, with five transplants, are presented. All patients received transplants from unrelated donors' BM, except for one patient who received a cord blood transplant. The conditioning regimens were based on myeloablative protocols for BM derived transplants; a NM regimen was pursued for the patient who received a cord blood transplant, which resulted in graft rejection. Although recurrent pneumonia and skin infections resolved immediately after transplantation, all patients subsequently developed human herpesvirus infection, including cutaneous herpetic lesions, cytomegalovirus reactivation, and zona zoster, which could be attributed to the use of ATG. Despite the presence of serious morbidities prior to transplantation, all patients recovered successfully. DIDS can be successfully treated with allogeneic HSCT from unrelated donors following a myeloablative conditioning regimen, with a reasonable safety profile.

How effective are the 6 European Society of Immunodeficiency warning signs for primary immunodeficiency disease?

BACKGROUND: The European Society of Immunodeficiency (ESID) developed 6 warning signs to promote the awareness of adult primary immunodeficiency disease (PID). OBJECTIVE: To screen adult patients for the presence of PID using these 6 warning signs to determine the effectiveness of this protocol. METHODS: Questions related to the ESID warning signs for adult PID were added to the standard outpatient clinic file system and asked of 3,510 patients who were admitted to our clinic for any reason. Patients with signs and/or suspicion of PID based on their medical history underwent immunologic investigation. RESULTS: In total, 24 patients were diagnosed as having a PID. The most common reason that patients with PID were admitted was frequent infection (n=18 [75%]), and the most common PID subgroup was common variable immunodeficiency (n=12 [50%]). Twenty patients with PID had at least one positive finding according to the ESID warning signs. Two patients with gastrointestinal concerns and 2 with dermatologic symptoms were also diagnosed as having a PID, although they did not have any of the ESID warning signs. CONCLUSION: The ESID warning signs do not specify the need for symptoms to diagnose a PIDs and do not include a comprehensive list of all signs and symptoms of PIDs. As a result, more than infection-centric questions are needed to identify adult patients with immunodeficiencies.

Leigh Syndrome due to MT-ATP6 Variants: A Case Presentation and the Review of the Literature.

Introduction: Leigh syndrome is a rare mitochondrial disorder characterized by subacute necrotizing encephalomyelopathy, resulting from defects in mitochondrial respiratory enzymes or pyruvate dehydrogenase complex. Symptoms can manifest in infancy, childhood, or adulthood. We present a case of a 7-month-old girl initially misdiagnosed with septic shock but was later found to have Leigh encephalomyelopathy due to MT-ATP6 deficiency. Case Presentation: A 7-month-old girl was admitted with fever, drowsiness, and wheezing, initially diagnosed with septic shock. She had a history of parental consanguinity and hypotonia. Physical examination revealed unconsciousness, miotic pupils, and respiratory distress. Initial laboratory tests showed significant metabolic acidosis and elevated lactate, creatine kinase, and ammonia levels. The patient was treated for sepsis and shock, but her condition worsened with elevated lactate and liver transaminases, eventually leading to hypertrophic cardiomyopathy and multiorgan failure. Her basic metabolic scans showed extremely low citrulline levels, whole-exome sequencing analysis did not show any pathologic change in nuclear genome, and mitochondrial genome analysis revealed an MT-ATP homoplasmic variant. She passed away on the 22nd day of hospitalization. Discussion/Conclusion: While mitochondrial disorders are broadly acknowledged for their phenotypic diversity, it is essential to note that specific disorders, such as Leigh syndrome, display distinctive presentations with varying degrees of severity. Factors such as the percentage of homoplasmy contribute to the variability in manifestations. Notably, MT-ATP6-associated Leigh syndrome is predominantly characterized by an early onset, typically occurring before the age of 2 years. Low citrulline levels have been observed in approximately 90% of patients with MT-ATP6-related disorders, distinguishing them from other mitochondrial disorders. The exact mechanisms underlying this specific metabolic alteration are not fully understood, but it could be linked to disruptions in the mitochondrial energy production process. The mitochondria are essential for various metabolic pathways, including the urea cycle, where citrulline is involved. The association between low citrulline levels and MT-ATP6-related disorders raises the possibility of using citrulline as a potential biomarker for disease identification. MT-ATP6 defects should be kept in mind in cases with mitochondrial disease and low plasma citrulline levels.

Primary antibody deficiencies in Turkey: molecular and clinical aspects.

Primary antibody deficiencies (PAD) are the most common subtype of primary immunodeficiencies, characterized by increased susceptibility to infections and autoimmunity, allergy, or malignancy predisposition. PAD syndromes comprise of immune system genes highlighted the key role of B cell activation, proliferation, migration, somatic hypermutation, or isotype switching have a wide spectrum from agammaglobulinemia to selective Ig deficiency. In this study, we describe the molecular and the clinical aspects of fifty-two PAD patients. The most common symptoms of our cohort were upper and lower respiratory infections, bronchiectasis, diarrhea, and recurrent fever. Almost all patients (98%) had at least one of the symptoms like autoimmunity, lymphoproliferation, allergy, or gastrointestinal disease. A custom-made next-generation sequencing (NGS) panel, which contains 24 genes, was designed to identify well-known disease-causing variants in our cohort. We identified eight variants (15.4%) among 52 PAD patients. The variants mapped to BTK (n = 4), CD40L (n = 1), ICOS (n = 1), IGHM (n = 1), and TCF3 (n = 1) genes. Three novel variants were described in the BTK (p.G414W), ICOS (p.G60*), and IGHM (p.S19*) genes. We performed Sanger sequencing to validate pathogenic variants and check for allelic segregation in the family. Targeted NGS panel sequencing can be beneficial as a suitable diagnostic modality for diagnosing well-known monogenic PAD diseases (only 2-10% of PADs); however, screening only the coding regions of the genome may not be adequately powered to solve the pathogenesis of PAD in all cases. Deciphering the regulatory regions of the genome and better understanding the epigenetic modifications will elucidate the molecular basis of complex PADs.

The frequency of vitamin B12, iron, and folic acid deficiency in the neonatal period and infancy, and the relationship with maternal levels.

AIM: The most important function of vitamin B12 is to accomplish DNA synthesis, which is necessary for cell division and proliferation. Deficiency of vitamin B12 causes megaloblastic anemia, retardation of growth, and delay in neuromotor maturation. Newborns whose mothers have vitamin B12 deficiency are born with low vitamin B12 storages, and are at risk in terms of vitamin B12 deficiency symptoms during infancy. The aim of our study was to investigate the frequency of anemia and deficiency of vitamin B12, folic acid, and iron in pregnant women living in our region, in their newborn babies, and during the infancy period of these babies. Another aim of our study was to investigate the correlation between the levels of these vitamins in newborns and in their mothers. MATERIAL AND METHODS: In our study, 250 pregnant women at 38-42 gestational weeks, who were admitted for delivery to Gynecology and Obstetrics Clinic and their babies with a birth weight over 2500 g were included in the study. RESULTS: We determined that 24.8% of the pregnant women had anemia, 28% had low ferritin levels, 90.4% had vitamin B 12 deficiency, and 22.4% had folic acid deficiency. Some 3.2% of the newborns had anemia, 2.8% had low ferritin levels, and 72.4% had vitamin B12 deficiency. Among the infants who presented for a follow-up visit at 6 months of age, 22.3% had anemia, 14.9% had low ferritin levels, 40.4% had vitamin B12 deficiency, and 1.06% had folic acid deficiency. In addition, we found that the levels of vitamin B12 and folic acid in newborns were related to the levels of vitamin B12 and folic acid in their mothers. CONCLUSION: Development of low vitamin B12 stores in newborns and the development of vitamin B12 deficiency during infancy, which may result in irreversible complications including neurologic complications, can be prevented by preventing vitamin B12 deficiency during pregnancy.

International Efforts to Save Healthcare Personnel during COVID-19.

Coronavirus disease 2019 (COVID-19) pandemic is a global challenge. Several governments of the world have decided to take drastic actions in order to combat the spread of the disease, including the closing of air, maritime and land borders, as an extreme measure of isolation of each country/region. However, such measures had not prevented the disease from spreading globally; as COVID-19 has already spread in almost all countries. This virus's main victims are the healthcare personnel (HCP), who are physically and psychologically affected. The HCP serves as the first line of defense against this pandemic, what if we faced a significant loss in their number? And what if our HCP was going through a deep dark depression? The condition would be terrifying not only for now but also in the future. This raises the need for an intensified International collaboration, that mainly supports the HCP. We are throwing by challenging moments, and it is clear that social distancing, cooperation, hygiene awareness and abide by the recommendation and help of all governments, as well as obtaining the support of international organizations could be an excellent tool for preventing an increase in the number of cases, principally in countries and regions were COVID-19 is in the early stage of the epidemic. However, this is not the final solution for the current pandemic. An intensified global program, which mainly supports the HCP, then considers the other aspects of the COVID19 pandemic might bring this pandemic to a peaceful end.

Inherited IFNAR1 deficiency in otherwise healthy patients with adverse reaction to measles and yellow fever live vaccines.

Vaccination against measles, mumps, and rubella (MMR) and yellow fever (YF) with live attenuated viruses can rarely cause life-threatening disease. Severe illness by MMR vaccines can be caused by inborn errors of type I and/or III interferon (IFN) immunity (mutations in IFNAR2, STAT1, or STAT2). Adverse reactions to the YF vaccine have remained unexplained. We report two otherwise healthy patients, a 9-yr-old boy in Iran with severe measles vaccine disease at 1 yr and a 14-yr-old girl in Brazil with viscerotropic disease caused by the YF vaccine at 12 yr. The Iranian patient is homozygous and the Brazilian patient compound heterozygous for loss-of-function IFNAR1 variations. Patient-derived fibroblasts are susceptible to viruses, including the YF and measles virus vaccine strains, in the absence or presence of exogenous type I IFN. The patients' fibroblast phenotypes are rescued with WT IFNAR1 Autosomal recessive, complete IFNAR1 deficiency can result in life-threatening complications of vaccination with live attenuated measles and YF viruses in previously healthy individuals.

Ocular Findings in Children With 22q11.2 Deletion Syndrome.

PURPOSE: To identify the ocular features of children diagnosed as having 22q11.2 deletion syndrome in a Turkish population, which is the most common microdeletion syndrome with a wide range of facial and ocular abnormalities. METHODS: Sixteen children aged between 4 months and 18 years with a microdeletion in chromosome 22q11.2 underwent a detailed ophthalmological examination including uncorrected and best corrected visual acuity testing, stereoscopic vision examination, biomicroscopic and indirect fundus examination, and ocular motility testing. RESULTS: All patients had at least one ocular abnormality. The major abnormalities were eyelid abnormalities (eye hooding, narrow palpebral fissure, telecanthus, hypertelorism, sparse and thin eyebrows and eyelashes, blepharitis, and distichiasis), posterior embryotoxon, and tortuous retinal vessels in at least half of the patients. Other ophthalmological disorders were refractive errors, iris remnants, and strabismus. CONCLUSIONS: The chromosome 22q11.2 deletion syndrome is associated with a wide range of ocular disorders, which necessitates a comprehensive eye examination for appropriate treatment and follow-up. Ocular findings sometimes can provide a clue to the diagnosis of 22q11.2 deletion. [J Pediatr Ophthalmol Strabismus. 2016;53(4):218-222].