Multiplex Minisequencing of the HBB Gene: A Rapid Strategy to Confirm the Most Frequent ß-Thalassemia Mutations in the Tunisian Population.

The ß hemoglobinopathies [ß-thalassemia (ß-thal) and structural hemoglobin (Hb) variants such as Hb S (HBB: c.20A > T) and Hb E (HBB: c.79G > A)] are among the most common inherited diseases worldwide. In Tunisia, due to the high prevalence of consanguineous marriages, the recurrent risk of this disease is high. The average prevalence of hemoglobinopathies is 4.48%, reaching 12.50% in some focus regions. The molecular investigations on thalassemia contributed to establishing the spectrum of mutations in the Tunisian population. The total number of HBB gene mutations identified was 24. The two most frequent mutations, codon 39 (C > T) (HBB: c.118C > T) and IVS-I-110 (G > A) (HBB: c.93-21G > A) accounted for 70.0% of the total encountered ß-thal cases. These two mutations together with IVS-I-2 (T > G) (HBB: c.92 + 2T > G) and the Hb S variant account for more than 90.0% of all HBB genetic variants in Tunisia. Thus, developing rapid, inexpensive and reliable mutation-specific molecular diagnostic assays targeting our Tunisian populations is our aim to facilitate routine detection of hemoglobinopathies. In this report, we describe the successful application of the multiplex minisequencing assay as an alternative strategy for genetic diagnosis of HBB gene disorders in Tunisia.

Cribriform plate dehiscence with encephalomeningocele revealed by recurrent meningitis: A case report.

INTRODUCTION: Meningitis is a potentially life threatening illness. It requires prompt diagnosis and treatment. Recurrent meningitis needs detailed investigations to identify the underlying cause. OBSERVATION: We report a case of recurrent pneumococcal meningitis in a 9-year-old boy with an underlying congenital skull base abnormality. Brain computed tomography (CT) scan showed no obvious skull base defects. A magnetic resonance imaging (MRI) of the brain revealed a dehiscence of the cribriform plate with encephalomeningocele. The patient underwent an endoscopic repair of the bony defect and had not developed any new infections ever since. CONCLUSION: This case highlights the need to investigate recurrent bacterial meningitis with CT scan and MRI of the brain and skull base. Repair of these congenital skull base defects are mandatory to prevent the recurrence of meningitis.

Genetic investigation of the ubiquitin-protein ligase E3A gene as putative target in Angelman syndrome.

BACKGROUND: Angelman syndrome (AS) is caused by maternal chromosomal deletions, imprinting defects, paternal uniparental disomy involving chromosome 15 and the ubiquitin-protein ligase UBE3A gene mutations. However the genetic basis remains unclear for several patients. AIM: To investigate the involvement of UBE3A gene in AS and identifying new potential genes using exome sequencing. METHODS: We established a cohort study in 50 patients referred to Farhat Hached University Hospital between 2006 and 2021, with a strong suspicion of AS and absence of chromosomal aberrations. The UBE3A gene was screened for mutation detection. Two unrelated patients issued from consanguineous families were subjected to exome analysis. RESULTS: We describe seven UBE3A variants among them 3 none previously described including intronic variants c.2220+14T>C (intron14), c.2507+43T>A (Exon15) and insertion in Exon7: c.30-47_30-46. The exome sequencing revealed 22 potential genes that could be involved in AS-like syndromes that should be investigated further. CONCLUSION: Screening for UBE3A mutations in AS patients has been proven to be useful to confirm the diagnosis. Our exome findings could rise to new potential alternative target genes for genetic counseling.

Array-CGH study of partial trisomy 9p without mental retardation.

Partial trisomy 9p is one of the most common detected autosomal structural anomalies, so the phenotype-genotype correlation of this rearrangement has been well described. Despite variation in size of the 9p duplications, trisomy 9p syndrome is characterized by typical dysmorphic features and a variable but constant psychomotor and mental retardation. Previously reported phenotype genotype correlation studies proposed that the critical region for phenotype is located in 9p22. We report here on a new patient with partial trisomy 9p13.3→9pter in an 8-year-old boy with typical trisomy 9p dysmorphic features but a normal mental development. Cytogenetics investigations showed that our patient karyotype was 47,XY,+ der(22)t(9;22)(p13.q11) inherited by a 3:1 disjunction of a maternal reciprocal translocation t(9;22)(p13.q11). FISH and array CGH analysis were used to better characterize duplicated chromosomal regions and showed a large duplication of chromosome 9p13.3→9pter associated to microduplication in 22q11.1. The size of the duplications in chromosomes 9p and 22q were estimated about 33.9 and 2.67 Mb, respectively. The comparison between this case and those reported in the literature allows us to support that all syndromes show variability and that not all partial trisomies 9p are associated with intellectual disability.

Further report of MEDS syndrome: Clinical and molecular delineation of a new Tunisian case.

Recently, an autosomal recessive disorder including the triad of microcephaly, infantile epileptic encephalopathy, and permanent neonatal diabetes syndrome (MEDS, OMIM#614231) has emerged as a new distinguishing syndrome. Eight cases of whom seven from Arab countries, have been reported in association with biallelic variants in the IER3IP1 gene (Immediate early response-3 interacting protein-1). Here, we describe a Tunisian boy who presented with permanent neonatal diabetes, microcephaly, generalized seizures and hypovirilized external genitalia consisting of a small genitalia and unilateral cryptorchidism. Chromosomal analysis indicated a 46, XY karyotype in all metaphases. Exome sequencing identified a homozygous missense variant (c.62 T > G; p. Val21Gly) in the IER3IP1 gene, that is predicted to alter the protein structure within the hydrophobic/transmembrane. This variant was previously reported in two cases associated with MEDS. This is the first reported case of MEDS in Tunisia. Our report focuses on the IER3IP1 related phenotypic spectrum and assumes abnormal genitalia as part of the syndrome. Consequently, we recommend to perform hormonal testing on this topic to understand the effect of the IER3IP1 variant on the male genital pathway.

Rituximab in subacute sclerosing panencephalitis.

Subacute sclerosing panencephalitis (SSPE) is a progressive, fatal neurological disorder of childhood and early adolescence. It is caused by a persistent measles virus infection of the brain without any available treatment to date. The physiopathology of the disease is largely unknown. Considering the potential role of humoral immunity in the pathogenesis of SSPE, one patient was given compassionate anti-CD20 antibodies. However, disease progression under treatment led to reconsider B cell involvement in this pathology. Nevertheless, we observed that carbamazepine was useful in improving life quality in our patient, and should be considered as a first-line drug. To date, measles vaccination remains the only solution to SSPE.

Genomic Microarray in Intellectual Disability: The Usefulness of Existing Systems in the Interpretation of Copy Number Variation.

Whole genome array technology is an essential tool for the detection of a large number of copy number variants (CNVs) in patients with ID and/or multiple congenital anomalies. However, the clinical significance of some microimbalances is not known. In this article, we succeeded to detect seven new variations of unknown significance (dup12p13.33, dup2p16.3, dupXq13.2, del12q24.33, dup16p13.11, trip4q22.1, and dup9p21.3), one CNV classified as known pathogenic syndrome (del22q13.31-q33), and one CNV classified as potentially pathogenic (del11q24.3). We emphasize the role of comparative genomic hybridization arrays in the investigation of intellectual disability and evaluate the usefulness of existing systems in the interpretation of CNVs.

Microarray Analysis of 8p23.1 Deletion in New Patients with Atypical Phenotypical Traits.

We describe two patients carrying deletions of chromosome 8p23.1 with a commonly critical region identified by means of oligonucleotide array comparative genomic hybridization (array CGH). They didn't present congenital heart defects or behavioral problems. Only one patient presented with intellectual disability and carrying deletion of TNKS gene. We presumed the inclusion of TNKS gene in the mental impairment.

Molecular cytogenetic and phenotypic characterization of ring chromosome 13 in three unrelated patients.

We report on the cytogenetic and molecular investigations of constitutional de-novo ring chromosome 13s in three unrelated patients for better understanding and delineation of the phenotypic variability characterizing this genomic rearrangement. The patient's karyotypes were as follows: 46,XY,r(13)(p11q34) dn for patients 1 and 2 and 46,XY,r(13)(p11q14) dn for patient 3, as a result of the deletion in the telomeric regions of chromosome 13. The patients were, therefore, monosomic for the segment 13q34 → 13qter; in addition, for patient 3, the deletion was larger, encompassing the segment 13q14 → 13qter. Fluorescence in situ hybridization confirmed these rearrangement and array CGH technique showed the loss of at least 2.9 Mb on the short arm and 4.7 Mb on the long arm of the chromosome 13 in patient 2. Ring chromosome 13 (r(13)) is associated with several phenotypic features like intellectual disability, marked short stature, brain and heart defects, microcephaly and genital malformations in males, including undescended testes and hypospadias. However, the hearing loss and speech delay that were found in our three patients have rarely been reported with ring chromosome 13. Although little is known about its etiology, there is interesting evidence for a genetic cause for the ring chromosome 13. We thus performed a genotype-phenotype correlation analysis to ascertain the contribution of ring chromosome 13 to the clinical features of our three cases.

Phenotype and micro-array characterization of duplication 11q22.1-q25 and review of the literature.

Partial duplication of 11q is related to several malformations like growth retardation, intellectual disability, hypoplasia of corpus callosum, short nose, palate defects, cardiac, urinary tract abnormalities and neural tube defects. We have studied the clinical and molecular characteristics of a patient with severe intellectual disabilities, dysmorphic features, congenital inguinal hernia and congenital cerebral malformation which is referred to as cytogenetic exploration. We have used FISH and array CGH analysis for a better understanding of the double chromosomic aberration involving a 7p microdeletion along with a partial duplication of 11q due to adjacent segregation of a paternal reciprocal translocation t(7;11)(p22;q21) revealed after banding analysis. The patient's karyotype formula was: 46,XY,der(7)t(7;11)(p22;q21)pat. FISH study confirmed these rearrangement and array CGH technique showed precisely the loss of at least 140 Kb on chromosome7p22.3pter and 33.4Mb on chromosome11q22.1q25. Dysmorphic features, severe intellectual disability and brain malformations could result from the 11q22.1q25 trisomy. Our study provides an additional case for better understanding and delineating the partial duplication 11q.

Xp22.3 interstitial deletion: a recognizable chromosomal abnormality encompassing VCX3A and STS genes in a patient with X-linked ichthyosis and mental retardation.

X-linked ichthyosis is a genetic disorder affecting the skin and caused by a deficit in the steroid sulfatase enzyme (STS), often associated with a recurrent microdeletion at Xp22.31. Most of the STS deleted patients have X-linked ichthyosis as the only clinical feature and it is believed that patients with more complex disorders including mental retardation could be present as a result of contiguous gene deletion. In fact, VCX3A gene, a member of the VCX (variable charge, X chromosome) gene family, was previously proposed as the candidate gene for X-linked non-specific mental retardation in patients with X-linked ichthyosis. We report on a boy with familial ichthyosis, dysmorphic features and moderate mental retardation with approximately 2 Mb interstitial deletion on Xp22.3 involving VCX3A and STS genes.

Chromosomal microarray analysis of functional Xq27-qter disomy and deletion 3p26.3 in a boy with Prader-Willi like features and hypotonia.

Duplications of the long arm of the X chromosome are rare. The infantile phenotype shares some resemblance with the Prader-Willi syndrome, presenting severe psychomotor retardation, facial dysmorphic features with a broad face, a small mouth and a thin pointed nose, hypotonia, urogenital malformation and proneness to infections. We report a boy with an additional Xq27-qter chromosome segment translocated onto the short arm of chromosome 3. The karyotype was 46,XY,der(3)t(X;3)(q27.3; p26.3)mat. This cryptic unbalanced X-autosome translocation resulted in Xq27-qter functional disomy and a deletion 3p26.3. A detailed analysis of the constitutional chromosomal changes in the patient was performed using array-CGH, FISH and PCR. The aim was to characterize the size and the location of the duplication Xq27-qter (8.18 Mb) and of the deletion 3p26.3 (1.05 Mb), to establish phenotype-genotype correlations and to offer genetic counselling.

Trisomy and tetrasomy 15q11-q13 diagnosed by molecular cytogenetic analysis in two patients with mental retardation.

In this study, we report two patients with the supernumerary marker chromosome (15)s. The first case is an 8.5-year-old girl with an inv dup (15) syndrome, mental retardation and dysmorphic features. The second case is a 13-year-old boy with a ring chromosome 15, who was referred to the Laboratory of Cytogenetic and Biology of Reproduction in Sousse, Tunisia for mental retardation, epilepsy, speech delay, hypotonia and other mild dysmorphic features. R banding showed the presence of a marker chromosome in both cases. Molecular cytogenetic investigation using fluorescence in situ hybridization method allowed us to characterize the markers including the Prader-Willi syndrome locus that contains the small nuclear ribonucleoprotein polypeptide N (SNRPN) gene. Tetrasomy and trisomy for the 15q11-q13 chromosomal region were found in the first and the second patient, respectively. This observation reinforces the hypothesis that additional copies of proximal chromosome 15q11 segment may be causally related to mental retardation and dysmorphic features.

Cytogenetic analysis in a large series of children with non-syndromic mental retardation.

Mental retardation affects 1-3% of the population. To evaluate the implication of chromosomal abnormalities in the etiology of mental retardation, 1420 patients with non-syndromic mental retardation recruited at the department of cytogenetics of Farhat Hached hospital (Sousse, Tunisia) between January 2005 and December 2009, were analyzed using standard cytogenetic techniques. Age ranged between 3 and 18 years with a median of 8 years. Chromosomal abnormalities were detected in 7.8% of patients and an increased prevalence of chromosome anomalies was observed in patients when the mental retardation is associated with a severe degree of intellectual disability, facial dysmorphic features and/or congenital malformations or epilepsy.

Chromosomal microarray analysis in a girl with mental retardation and spina bifida.

Chromosomal imbalances comprise a major cause of mental retardation, particularly in association with congenital malformations and dysmorphic features. Chromosomal analysis using banded karyotyping is limited by the low resolution of this technique, and cryptic chromosomal rearrangements cannot be detected. We describe a 6-year-old girl with mental retardation, mild growth, congenital malformation, and facial anomalies. Chromosomal analysis with karyotyping produced normal results. Because the phenotype suggested chromosomal abnormality, microarray comparative genomic hybridization was used to search for a possible cryptic anomaly. A subtelomeric chromosomal imbalance, consisting of partial trisomy 2q35 and partial monosomy 3p26, was detected and confirmed using fluorescence in situ hybridization. This rearrangement was inherited from an equilibrated maternal t(2;3) reciprocal translocation. Comparative genomic hybridization array in similar situations is useful in detecting cryptic chromosomal rearrangements, identifying genes contained in deleted or duplicated regions, establishing a precise phenotype-genotype correlation, and offering unambiguous genetic counseling.