Sonia Abdelhak

The experience of a Tunisian referral centre in prenatal diagnosis of Xeroderma pigmentosum.

Aims: Xeroderma pigmentosum (XP, OMIM 278700-278780) is one of the most severe genodermatoses and is relatively frequent in Tunisia. In the absence of any therapy and to better manage the disease, we aimed to develop a molecular tool for DNA-based prenatal diagnosis. Methods: Six consanguineous Tunisian XP families (4 XP-A and 2 XP-C) have benefited from a prenatal diagnosis. Screening for mutations was performed by direct sequencing, while maternal-foetal contamination was checked by genotyping. Results: Among the 7 prenatal diagnoses, 4 foetuses were heterozygous for the screened mutation. Exclusion of contamination by maternal cells was checked. Mutations were detected at a homozygous state in the remaining cases, and the parents decided to terminate pregnancy. Conclusion: Our study illustrates the implementation of prenatal diagnosis for better health support of XP in Tunisia.

High frequency of exon 15 deletion in the FANCA gene in Tunisian patients affected with Fanconi anemia disease: implication for diagnosis

Tunisian population is characterized by its heterogeneous ethnic background and high rate of consanguinity. In consequence, there is an increase in the frequency of recessive genetic disorders including Fanconi anemia (FA). The aim of this study was to confirm the existence of a founder haplotype among FA Tunisian patients and to identify the associated mutation in order to develop a simple tool for FA diagnosis. Seventy‐four unrelated families with a total of 95 FA patients were investigated. All available family members were genotyped with four microsatellite markers flanking FANCA gene. Haplotype analysis and homozygosity mapping assigned 83 patients belonging to 62 families to the FA‐A group. A common haplotype was shared by 42 patients from 26 families at a homozygous state while five patients from five families were heterozygous. Among them, 85% were from southern Tunisia suggesting a founder effect. Using multiplex ligation‐dependent probe amplification (MLPA) technique, we have also demonstrated that this haplotype is associated with a total deletion of exon 15 in FANCA gene. Identification of a founder mutation allowed genetic counseling in relatives of these families, better bone marrow graft donor selection and prenatal diagnosis. This mutation should be investigated in priority for patients originating from North Africa and Middle East.

Autism in Phenylketonuria Patients From Clinical Presentation to Molecular Defects

Autism has been reported in untreated patients with phenylketonuria. The authors aimed to explore autism in 15 Tunisian and 4 Algerian phenylketonuria patients, and report their clinical, biochemical and molecular peculiarities. The Childhood Autism Rating Scale and the Autism Diagnostic Interview–Revised were used for the diagnosis of autism. Five exons of phenylalanine hydroxylase gene (7, 6, 10, 11, and 5) were amplified by polymerase chain reaction and directly sequenced. Among these patients, 15 were suffering from autism at the time of evaluation. Six mutations were identified: p.E280K, p.G352Vfs, IVS10nt11, p.I224T, p.R261Q, and p.R252W. There was no correlation between autism and mutations affecting the phenylalanine hydroxylase gene, but the age of diet onset was the determining factor in autistic symptoms’ evolution.

Clinical, genealogical and molecular investigation of the xeroderma pigmentosum type C complementation group in Tunisia

DEAR EDITOR, Xeroderma pigmentosum (XP) is an ultraviolet sensitivity syndrome characterized by skin hyperpigmentation, premature photoageing and early onset of skin cancers. XP is a rare disease. In the U.S.A. and Europe the estimated incidence is 1 per 1 million inhabitants, while it is much higher in Japan (1 in 22 000) and both North Africa and the Middle East (estimated 1 in 50 000). In Tunisia the frequency is about 1 in 10 000. There are seven complementation groups with defects in the nucleotide excision repair pathway (XP-A to XP-G). Patients with XP-C develop predominantly skin damage and early malignancies without neurological abnormalities. Nevertheless, several cases with neurological disorders have been reported. Here we report on the clinical, genealogical and molecular investigation of 44 families including a total of 64 patients with XP-C who were followed at three university hospitals (Charles Nicolle, Habib Thameur and La Rabta hospitals) during the period from 2006 to 2013 (Table 1). Patients were suspected to have XP-C on the basis of their clinical features (age at onset of erythema, age at appearance of skin cancers and absence of neurological abnormalities). Written informed consent was obtained from all participants or their legal guardians. All families were interviewed using a specific questionnaire to collect information about genealogical data, familial history and associated diseases. We performed molecular and histological analysis at Institut Pasteur de Tunis. The study was evaluated and approved by the institutional ethical committee and conducted according to the Declaration of Helsinki principles. Blood samples were obtained from all available family members and DNA was extracted using a salting-out procedure or Qiagen FlexiGene DNA kit (51206; Qiagen, Venlo, the Netherlands). Genomic DNA was amplified using XPC gene primers specific to exons 4–16. Polymerase chain reaction products were sequenced in an ABI 3130 Genetic Analyzer (Applied Biosystems, Foster City, CA, U.S.A.). Mutations were annotated using NG 011763 1 according to the Human Genome Variation Society version 2.0 (Mutalyzer 2.0.beta-26; https://www.mutalyzer.nl/). In the case of skin tumours, histopathological analyses were performed on biopsies using routine haematoxylin and eosin staining to define the histological type of the tumours. Acquired data were statistically analysed using SPSS 20.0 for Windows (IBM, Armonk, NY, U.S.A.). Descriptive statistical methods (frequency, mean, median, SD) were performed with their corresponding graphs. The association between protection levels and skin cancer development was calculated using a Pearson correlation test. A P-value ≤ 0 05 was considered statistically significant. The observed number of nonmelanoma skin cancers (NMSCs) was compared with the expected number from regional cancer registries for the 1995–2006 period after adjustment for age and sex. The exact Fisher test was used to predict the relative risk factor odds ratio (Table 2). Genealogical pedigrees were drawn using the R package Kinship2 (http://www.r-project.org/). Mutational analysis showed that 60 patients (94%) belonging to 43 families not known to be related shared the founder mutation XPC c.1643_1644delTG (p.Val548Alafs*25). Among them, 15 patients shared the same haplotype. This founder mutation c.1643_1644delTG has been identified in patients from various regions around the world, with higher frequencies in the Mediterranean region. It has been estimated that the common ancestor mutation occurred 1250 years ago. In addition, three unrelated patients (5%) had the second founder mutation c.850G>T and one patient (2%) had the c.779+1G>A mutation (Fig. 1). It is noteworthy that all of the patients were homozygous for the deleterious mutations, and no compound heterozygous mutation was identified. The mutations were confirmed in the parents. In addition, all available related individuals were screened. Among the latter, 24 were healthy heterozygous carriers and 18 were homozygous for the wild-type allele (Fig. 1). Presymptomatic diagnosis was performed for two patients under the age of 2 years (XP66 2MED and XP30 2MO). Four families (XP13KA, XP26B, XP46MED and XP76MED) asked for prenatal diagnosis (PND). The XP46MED family benefited from two consecutive PNDs. The results showed that one fetus was homozygous for the wild-type allele, three fetuses were heterozygous and one was affected homozygous for the screened mutation in the proband. The parents decided to terminate the pregnancy when the fetus was homozygous for the deleterious allele. Analysis of the geographical distribution of XPC mutations showed that southeast Tunisia and the coasts have the highest

H syndrome: Clinical, histological and genetic investigation in Tunisian patients

H syndrome is a rare autosomal recessive disorder with characteristic dermatological findings consisting of hyperpigmentation and hypertrichosis patches mainly located on the inner thighs and multisystemic involvement including hepatosplenomegaly, hearing loss, heart abnormalities and hypogonadism. The aim of this study was to conduct a clinical and genetic investigation in five unrelated Tunisian patients with suspected H syndrome. Hence, genetic analysis of the SLC29A3 gene was performed for four patients with a clinical diagnosis of H syndrome. We identified a novel frame‐shift mutation in the SLC29A3 gene in a female patient with a severe clinical presentation. Furthermore, we report two mutations previously described, the p.R363Q mutation in a male patient and the p.P324L mutation in two patients of different age and sex. This paper extends the mutation spectrum of H syndrome by reporting a novel frame‐shift mutation, the p.S15Pfs*86 in exon 2 of SLC29A3 gene and emphasizes the relevance of genetic testing for its considerable implications in early diagnosis and clinical management.