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Background In Tunisia a first SARS-CoV-2 confirmed case was reported in March 03, 2020. Since then, an increase of cases number was observed from either imported or local cases. The aim of this preliminary study was to better understand the molecular epidemiology and genetic variability of SARS-CoV-2 viruses circulating in Tunisia and worldwide. Methods Whole genome sequencing was performed using NGS approach on six SARS. CoV-2 highly positive samples detected during the early phase of the outbreak. Results Full genomes sequences of six Tunisian SARS-CoV-2 strains were obtained from imported and locally transmission cases during the COVID-19 outbreak. Reported sequences were non-identical with 0.1% nucleotide divergence rate and clustered into 6 different clades with worldwide sequences. SNPs results favor the distribution of the reported Tunisian sequences into 3 major genotypes. These SNP mutations are critical for diagnosis and vaccine development. Conclusions These results indicate multiple introductions of the virus in Tunisia and add new genomic data on SARS-CoV-2 at the international level.

International audience; We sought to determine the relative value of conventional molecular methods and whole-genome sequencing (WGS) for subtyping Salmonella enterica serovar Enteritidis recovered from 2000 to 2015 in Tunisia and to investigate the genetic diversity of this serotype. A total of 175 Salmonella Enteritidis isolates were recovered from human, animal, and foodborne outbreak samples. Pulsed-field gel electrophoresis (PFGE), multiple locus variable-number tandem repeat analysis (MLVA), and wholegenome sequencing were performed. Eight pulsotypes were detected for all isolates with PFGE (DI = 0.518). Forty-five Salmonella Enteritidis isolates were selected for the MLVA and WGS techniques. Eighteen MLVA profiles were identified and classified into two major clusters (DI = 0.889). Core genome multilocus typing (cgMLST) analysis revealed 16 profiles (DI = 0.785). Whole-genome analysis indicated 660 single-nucleotide polymorphism (SNP) divergences dividing these isolates into 43 haplotypes (DI = 0.997). The phylogenetic tree supported the classification of Salmonella Enteritidis isolates into two distinct lineages subdivided into five clades and seven subclades. Pairwise SNP differences between the isolates ranged between 302 and 350. We observed about 311 SNP differences between the two foodborne outbreaks, while only less or equal to 4 SNP differences within each outbreak. SNP-based WGS typing showed an excellent discriminatory power comparing with the conventional methods such as PFGE and MLVA. Besides, we demonstrate the added value of WGS as a complementary subtyping method to discriminate outbreak from non-outbreak isolates belonging to common subtypes. It is important to continue the survey of Salmonella Enteritidis lineages in Tunisia using WGS.

Alpha-Mannosidosis (AM) is an ultra-rare storage disorder caused by a deficiency of lysosomal alpha-mannosidase encoded by the MAN2B1 gene. Clinical presentation of AM includes mental retardation, recurrent infections, hearing loss, dysmorphic features, and motor dysfunctions. AM has never been reported in Tunisia. We report here the clinical and genetic study of six patients from two Tunisian families with AM. The AM diagnosis was confirmed by an enzymatic activity assay. Genetic investigation was conducted by Sanger sequencing of the mutational hotspots for the first family and by ES analysis for the second one. In the first family, a frameshift duplication p.(Ser802GlnfsTer129) was identified in the MAN2B1 gene. For the second family, ES analysis led to the identification of a missense mutation p.(Arg229Trp) in the MAN2B1 gene in four affected family members. The p.(Ser802GlnfsTer129) mutation induces a premature termination codon which may trigger RNA degradation by the NMD system. The decrease in the levels of MAN2B1 synthesis could explain the severe phenotype observed in the index case. According to the literature, the p.(Arg229Trp) missense variant does not have an impact on MAN2B1 maturation and transportation, which correlates with a moderate clinical sub-type. To explain the intra-familial variability of cognitive impairment, exome analysis allowed the identification of two likely pathogenic variants in GHR and SLC19A3 genes potentially associated to cognitive decline. The present study raises awareness about underdiagnosis of AM in the region that deprives patients from accessing adequate care. Indeed, early diagnosis is critical in order to prevent disease progression and to propose enzyme replacement therapy.

International audience; Purpose. Pertussis remains a public health concern in most countries. Our study aimed to prospectively explore the epidemiology of pertussis in the Tunis area of Tunisia between 2007 and 2016, and to characterize the virulence-associated genes of the collected Bordetella pertussis isolates.Methodology. Infants and children hospitalized at the Children’s Hospital of Tunis, Tunisia, between 2007 and 2016 for suspicion of pertussis were enrolled in the study. Culture and real-time PCR (qPCR) assays targeting IS481, IS1001, recA, H-IS1001 and ptxP were used to confirm the pertussis diagnosis. Phenotypic and genotypic characterization of recovered isolates was performed.Results/Key findings. A total of 1844 children were included in the study. Overall, 306 children (16.6 %) with Bordetella infection were confirmed by qPCR. Among them, 265 (86.6 %) were confirmed as having B. pertussis (IS481+, ptxP+, H-IS1001−), 18 (5.9 %) as having Bordetella parapertussis (IS481−, IS1001+) and 11 (3.6 %) as having Bordetella spp. (IS481+, ptxP−, H-IS1001−). No Bordetella holmesii (IS481+, IS1001−, H-IS1001+) was identified. The estimated pertussis incidence in the Tunis area was 134/100 000 in children aged less than 5 years. Two epidemic peaks were observed in 2009 and 2014. Ten B. pertussis isolates were cultured and characterized. Deficiency in pertactin expression was not observed, and genotyping of the isolates revealed a predominant allelic profile: ptxP3-ptxA1-prn2-fim2-1-fim3-2.Conclusion. This study demonstrated that pertussis is still present as a cyclical disease in Tunisia, despite high primo-vaccination coverage with a pertussis whole-cell vaccine. The predominant genotype of Tunisian B. pertussis isolates is similar to isolates circulating in countries using the acellular vaccine.

International audience; Septoria tritici blotch caused by the fungal pathogen Mycosphaerella graminicola (anamorph: Zymoseptoria tritici) is one of the most frequently occurring diseases on both bread and durum wheat crops worldwide. One hundred and sixty four durum wheat-adapted isolates of this fungus were sampled during the 2012 growing season from five distinct geographical locations of Tunisia (Bizerte, Béja, Kef, Jendouba and Siliana) in order to examine the status of strobilurin resistance of M. graminicola in this country. Resistance was assessed by screening the G143A substitution (Cytochrome b) which confers resistance to this class of fungicides. We used a PCR-based mismatch mutation assay allowing the amplification of either G143 (sensitive) or A143 (resistant) allele. All isolates were found to contain the sensitive wild-type G143 allele and therefore to be sensitive. Our study confirms recent reports on M. graminicola in Tunisia and shows that the Tunisian population of the fungus remains fully sensitive to strobilurins. A durability-oriented management of strobilurin applications in Tunisia is thereby recommended to prevent the development and widespread of the corresponding resistance such as in Europe, where pathogen populations are nowadays fully resistant to strobilurins.

International audience; Polyphenols, flavonoids and condensed tannins contents, as well as the antioxidant activity of the ethanolic and aqueous extracts obtained from aerial parts of 10 wild Tunisian plants, have been determined. Extracts showed appreciable levels of polyphenols and flavonoids, which reached 215.16 mg GAE g-1 DW in Lavandula stoechas ethanolic extract, and 49.12 mg RE g-1 DW in Thapsia garganica aqueous extract. The majority of tested extracts exhibited low total condensed tannins content, except for Rhus tripartitum and Periploca laevigata. The antioxidant activity tests showed great activity, especially for R. tripartitum and Lavandula multifida (IC50 = 5.16 and 5.1 μg mL-1, respectively). Canonical Correspondence Analysis revealed clear groupings of species according to the solvent used.

The genomic sequence data generated in this work were submitted to the European Nucleotide Archive and are available from the International Nucleotide Sequence Database Collaboration (NCBI/ENA/DDBJ) databases under project accession number PRJEB27412 and run data accession numbers ERS2572942 to ERS2572951.; International audience; The recent increase in pertussis cases observed in some countries may have several causes, including the evolution of Bordetella pertussis populations towards escape of vaccine-induced immunity. Most genomic studies of B. pertussis isolates performed so far are from countries that use acellular vaccines. The objective was to analyse genomic sequences of isolates collected during the 2014 whooping cough epidemic in Tunisia, a country where whole-cell vaccines are used. Ten Tunisian isolates and four vaccine strains were sequenced and compared to 169 isolates from countries where acellular vaccines are used. Phylogenetic analysis showed that Tunisian isolates are diverse, demonstrating a multi-strain 2014 epidemic peak, and are intermixed with those circulating in other world regions, showing inter-country transmission. Consistently, Tunisian isolates have antigen variant composition observed in other world regions. No pertactin-deficient strain was observed. The Tunisian B. pertussis population appears to be largely connected with populations from other countries.

Background Lynch syndrome (LS) is a highly penetrant inherited cancer predisposition syndrome, characterized by autosomal dominant inheritance and germline mutations in DNA mismatch repair genes. Despite several genetic variations that have been identified in various populations, the penetrance is highly variable and the reasons for this have not been fully elucidated. This study investigates whether, besides pathogenic mutations, environment and low penetrance genetic risk factors may result in phenotype modification in a Tunisian LS family. Patients and methods A Tunisian family with strong colorectal cancer (CRC) history that fulfill the Amsterdam I criteria for the diagnosis of Lynch syndrome was proposed for oncogenetic counseling. The index case was a man, diagnosed at the age of 33 years with CRC. He has a monozygotic twin diagnosed at the age of 35 years with crohn disease. Forty-seven years-old was the onset age of his paternal uncle withCRC. An immunohistochemical (IHC) labeling for the four proteins (MLH1, MSH2, MSH6 and PMS2) of the MisMatchRepair (MMR) system was performed for the index case. A targeted sequencing of MSH2, MLH1 and a panel of 85 DNA repair genes was performed for the index case and for his unaffected father. Results The IHC results showed a loss of MSH2 but not MLH1, MSH6 and PMS2 proteins expression. Genomic DNA screening, by targeted DNA repair genes sequencing, revealed an MSH2 pathogenic mutation (c.1552C>T; p.Q518X), confirmed by Sanger sequencing. This mutation was suspected to be a causal mutation associated to the loss of MSH2 expression and it was found in first and second degree relatives. The index case has smoking and alcohol consumption habits. Moreover, he harbors extensive genetic variations in other DNA-repair genes not shared with his unaffected father. Conclusion In our investigated Tunisian family, we confirmed the LS by IHC, molecular and in silico investigations. We identified a novel pathogenic mutation described for the first time in Tunisia. These results come enriching the previously reported pathogenic mutations in LS families. Our study brings new arguments to the interpretation of MMR expression pattern and highlights new risk modifiers genes eventually implicated in CRC. Twins discordance reported in this work underscore that disease penetrance could be influenced by both genetic background and environmental factors. Electronic supplementary material The online version of this article (10.1186/s12967-019-1961-9) contains supplementary material, which is available to authorized users.

International audience; Genome data, with underlying new knowledge, are accumulating at exponential rate thanks to ever-improving sequencing technologies and the parallel development of dedicated efficient Bioinformatics methods and tools. Advanced Education in Bioinformatics and Genome Analyses is to a large extent not accessible to students in developing countries where endeavors to set up Bioinformatics courses concern most often only basic levels. Here, we report a pioneering pilot experience concerning the design and implementation, from scratch, of a three-months advanced and extensive course in Bioinformatics and Genome Analyses in the Institut Pasteur de Tunis. Most significantly the outcome of the course was upgrading the participants’ skills in Bioinformatics and Genome Analyses to recognized international standards. Here we detail the different steps involved in the implementation of this course as well as the topics covered in the program. The description of this pilot experience might be helpful for the implementation of other similar educational projects, notably in developing countries, aiming to go beyond basics and providing young researchers with high-level skills.

International audience; A variety of viruses can cause acute flaccid paralysis (AFP). However, the causative agent, sometimes, remains undetermined. Metagenomics helps in identifying viruses not diagnosed by conventional methods. Stool samples from AFP (n = 104) and non-AFP (n = 114) cases that tested enterovirus-negative by WHO standard methods were investigated. A metagenomics approach, first used on five pools of four samples each, revealed the presence of adenovirus sequences. Amplification in A549 cells and full-genome sequencing were used for complete virus identification and for designing a PCR assay to screen individual related samples. Metagenomic analysis showed that adenovirus sequences that were closely to the A31 and A61 genotypes were the most abundant. Two out of the corresponding 20 individual samples were found positive by PCR, and isolates were obtained in cell culture. Phylogenetic analysis based on complete genome sequences showed that the viruses belong to HAdV-A31 genotype (98-100% nucleotide sequence identity). PCR analysis of stool samples from all AFP and non-AFP cases revealed that a larger proportion of the positive samples were from AFP cases (17.3%) than from non-AFP cases (2.4%). These results open the way to studies aiming to test a possible role of HAdV-A31 in the pathogenesis of AFP.

International audience; Toxoplasmosis is an important zoonosis caused by an obligate intracellular parasitic protozoan, Toxoplasma gondii. The disease is distributed worldwide and can affect all warm-blooded vertebrates, including humans. The present review aimed to collect, compile and summarize the data on the prevalence of T. gondii infection in humans and animals in the five North African countries (Morocco, Algeria, Tunisia, Libya and Egypt). Published data from national and international databases were used. Distribution patterns and risk factors for T. gondii infection are discussed, focusing on biotic and abiotic factors. This review is a comprehensive epidemiological analysis of T. gondii infection in North Africa and will therefore be a useful tool for researchers. It can also be used to propose or enhance appropriate national toxoplasmosis control programs.; La toxoplasmose est une zoonose importante causée par un protozoaire parasite intracellulaire obligatoire, Toxoplasma gondii. La maladie est répandue dans le monde entier, chez tous les vertébrés à sang chaud, y compris les humains. La présente étude visait à collecter, compiler et résumer les données sur la prévalence de l’infection par T. gondii chez l’homme et les animaux dans les cinq pays d’Afrique du Nord (Maroc, Algérie, Tunisie, Libye et Égypte). Les données publiées dans des bases de données nationales et internationales ont été utilisées. Les schémas de distribution et les facteurs de risque d’infection par T. gondii sont discutés, en se concentrant sur les facteurs biotiques et abiotiques. Cette synthèse est une analyse épidémiologique complète de l’infection par T. gondii en Afrique du Nord et sera donc un outil intéressant pour les chercheurs. Elle peut également être utilisée pour proposer ou renforcer des programmes nationaux appropriés de contrôle de la toxoplasmose.

The aim of the current study is to assess the prevalence ofCampylobacterinfection in broiler chickens, raised in intensive production conditions, and to evaluate the antimicrobial susceptibility of recoveredCampylobacterisolates. A total of 590 cloacal swab samples were taken from 13 broiler chicken flocks in the North East of Tunisia. All samples were tested for the presence of thermophilicCampylobacterby culture and PCR, targeting themapAandceuEgenes, respectively. Susceptibility to antimicrobial drugs was tested against 8 antibiotics. Prevalence ofCampylobacterinfection, relationship with geographic origins and seasons, antimicrobial resistance rates and patterns were analyzed. Total prevalence ofCampylobacterinfection in broiler flocks was in the range of 22.4%, with a predominance ofC. jejuni(68.9%), followed byC. coli(31.1%). Positive association was highlighted between the infection level and the season (P< 0.001), but no link was emphasized considering the geographic origin. Antimicrobial susceptibility testing revealed very high resistance rates detected against macrolide, tetracycline, quinolones, and chloramphenicol, ranging from 88.6% to 100%. Lower resistance prevalence was noticed forβ-lactams (47% and 61.4%) and gentamicin (12.9%). 17 R-type patterns were observed, and a common pattern was found in 30.3% of isolates. This study provides updates and novel data on the prevalence and the AMR of broiler campylobacters in Tunisia, revealing the occurrence of high resistance to several antibiotics and emphasizing the requirement of better surveillance and careful regulation of antimicrobials use.

Background Breast cancer is the most common cancer in women worldwide. Around 50% of breast cancer familial risk has been so far explained by known susceptibility alleles with variable levels of risk and prevalence. The vast majority of these breast cancer associated variations reported to date are from populations of European ancestry. In spite of its heterogeneity and genetic wealth, North-African populations have not been studied by the HapMap and the 1000Genomes projects. Thus, very little is known about the genetic architecture of these populations. Methods This study aimed to investigate a subset of common breast cancer loci in the general Tunisian population and to compare their genetic composition to those of other ethnic groups. We undertook a genome-wide haplotype study by genotyping 135 Tunisian subjects using the Affymetrix 6.0-Array. We compared Tunisian allele frequencies and linkage disequilibrium patterns to those of HapMap populations and we performed a comprehensive assessment of the functional effects of several selected variants. Results Haplotype analyses showed that at risk haplotypes on 2p24, 4q21, 6q25, 9q31, 10q26, 11p15, 11q13 and 14q32 loci are considerably frequent in the Tunisian population (> 20%). Allele frequency comparison showed that the frequency of rs13329835 is significantly different between Tunisian and all other HapMap populations. LD-blocks and Principle Component Analysis revealed that the genetic characteristics of breast cancer variants in the Tunisian, and so probably the North-African populations, are more similar to those of Europeans than Africans. Using eQTl analysis, we characterized rs9911630 as the most strongly expression-associated SNP that seems to affect the expression levels of BRCA1 and two long non coding RNAs (NBR2 and LINC008854). Additional in-silico analysis also suggested a potential functional significance of this variant. Conclusions We illustrated the utility of combining haplotype analysis in diverse ethnic groups with functional analysis to explore breast cancer genetic architecture in Tunisia. Results presented in this study provide the first report on a large number of common breast cancer genetic polymorphisms in the Tunisian population which may establish a baseline database to guide future association studies in North Africa. Electronic supplementary material The online version of this article (10.1186/s12885-018-5133-8) contains supplementary material, which is available to authorized users.

International audience; 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.

International audience; Background: Variants in the fat mass and obesity-associated (FTO) gene are associated with obesity and type 2 diabetes mellitus.Aim of the study: This study aims to assess the association of the rs9939609 variant and haplotypes in FTO gene with metabolic syndrome (MetS) components in a Tunisian population sample.Methods: A total of 685 Tunisian subjects were genotyped for the rs9939609T>A using TaqMan allelic discrimination assay. Two variants rs1421085T>C and rs8057044A>G already genotyped in a previous study were used to test haplotype association of the FTO gene.Results: Genotype distribution of the variant rs9939609 was different between MetS and controls (P = 0.017). Individuals carrying TA genotypes had a significantly increased risk independently of body mass index or age (P = 0.009). The variant rs9939609 was also associated with impaired fasting glucose (IFG) (P = 0.002). Among the eight haplotypes in the population, the haplotype GCA was significantly associated with a higher risk of developing the MetS, higher systolic blood pressure, and higher levels of fasting glucose and triglycerides (TGs) in the total sample and females, separately. Separate analysis by gender revealed a protective haplotype TGT among women (P = 0.023).Conclusions:FTO haplotypes have a strong influence on blood pressures and TG and IFG levels. These findings provide evidence that FTO gene may play a critical role in leading to MetS in Tunisian population.

International audience; AIMS: Maturity Onset Diabetes of the Young (MODY) is a monogenic form of diabetes with autosomal dominant inheritance pattern. The diagnosis of MODY and its subtypes is based on genetic testing. Our aim was investigating MODY by means of next-generation sequencing in the Tunisian population.METHODS: We performed a targeted sequencing of 27 genes known to cause monogenic diabetes in 11 phenotypically suspected Tunisian patients. We retained genetic variants passing filters of frequency in public databases as well as their probable effects on protein structures and functions evaluated by bioinformatics prediction tools.RESULTS: Five heterozygous variants were found in four patients. They include two mutations in HNF1A and GCK that are the causative genes of the two most prevalent MODY subtypes described in the literature. Other possible mutations, including novel frameshift and splice-site variants were identified in ABCC8 gene.CONCLUSIONS: Our study is the first to investigate the clinical application of targeted next-generation sequencing for the diagnosis of MODY in Africa. The combination of this approach with a filtering/prioritization strategy made a step towards the identification of MODY mutations in the Tunisian population.

Background Hard ticks are hematophagous ectoparasites characterized by their long-term feeding. The saliva that they secrete during their blood meal is their crucial weapon against host-defense systems including hemostasis, inflammation and immunity. The anti-hemostatic, anti-inflammatory and immune-modulatory activities carried out by tick saliva molecules warrant their pharmacological investigation. The Hyalomma dromedarii Koch, 1844 tick is a common parasite of camels and probably the best adapted to deserts of all hard ticks. Like other hard ticks, the salivary glands of this tick may provide a rich source of many compounds whose biological activities interact directly with host system pathways. Female H. dromedarii ticks feed longer than males, thereby taking in more blood. To investigate the differences in feeding behavior as reflected in salivary compounds, we performed de novo assembly and annotation of H. dromedarii sialotranscriptome paying particular attention to variations in gender gene expression. Results The quality-filtered Illumina sequencing reads deriving from a cDNA library of salivary glands led to the assembly of 15,342 transcripts. We deduced that the secreted proteins included: metalloproteases, glycine-rich proteins, mucins, anticoagulants of the mandanin family and lipocalins, among others. Expression analysis revealed differences in the expression of transcripts between male and female H. dromedarii that might explain the blood-feeding strategies employed by both genders. Conclusions The annotated sialome of H. dromedarii helps understand the interaction of tick-host molecules during blood-feeding and can lead to the discovery of new pharmacologically active proteins of ticks of the genus Hyalomma. Electronic supplementary material The online version of this article (10.1186/s13071-018-2874-9) contains supplementary material, which is available to authorized users.

International audience; During the time periods of June 2015 and from July to August 2016, sandflies were collected among seven collection sites of the three leishmaniasis endemic villages of Sidi Bouzid, Tunisia. A total of 690 sandflies were captured and identified (380 males and 310 females). Four species belonging to genus Phlebotomus (Ph.) and two species belonging to genus Sergentomyia were identified. Leishmania DNA was detected in four out of 310 females (one Ph. sergenti and three Ph. papatasi). The overall sensitivity of the Prepronociceptin gene detection reached 76%. The concurrent presence of Ph. papatasi and Ph. sergenti vectors, the analysis of blood-meals, together with the detection of L. major in Ph. papatasi, confirms the ultimate conditions for the transmission of the disease in center Tunisia. These results expand the known epidemiological area of distrubtion of leishmaniasis and its vectors in this part of Tunisia, highlighting the need for ongoing entomological and parasitological surveillance.

International audience; An entomological investigation performed in 2013 covering different bio-geographical areas varying from humid in the north to the arid in the center showed that sand flies of the subgenus Larroussius including Phlebotomus perniciosus, Phlebotomus perfiliewi, and Phlebotomus longicuspis are abundant and widely distributed in Tunisia A total of 3992 collected and pooled with up to 30 specimens per pool based on sex, trapping location and collection date were tested for the presence of phleboviruses by nested reverse transcriptase polymerase chain reaction and sequencing Of a total of 135 pools, 23 were positive, yielding and minimum infection rate of 0.6%. Phylogenetic analysis performed using partial amino acid sequence in the polymerase gene showed that all these phleboviruses were grouped in one cluster clearly distinct from but closely related to Massilia virus and Granada virus This putative novel virus, tentatively called Saddaguia virus (SADV), is widely distributed in Tunisia Together with Toscana, Punique, and Utique viruses, SADV is the fourth recognized phlebovirus to be transmitted by sand flies in Tunisia The medical and public health interest of SADV remains to be investigated.

International audience; Recurrent miscarriage (RM) is defined as three or more consecutive pregnancy losses before 24 weeks of gestation. Parental chromosomal abnormalities represent an important etiology of RM. The aim of the present study was to identify the distribution of chromosome abnormalities among Tunisian couples with RM referred to the Department of Cytogenetic at the Pasteur Institute of Tunis (Tunisia) during the last five years. Standard cytogenetic analysis was carried out in a total of 163 couples presenting with two or more spontaneous abortions. Karyotypes were analyzed by R-banding. We identified 14 chromosomal abnormalities including autosomal reciprocal translocation, Robertsonian translocation, inversion, mosaic aneuploidy and heteromorphysm. The overall prevalence of chromosomal abnormalities was 8.5% in our cohort. This finding underlies the importance of cytogenetic investigations in the routine management of RM.

Background In North African populations, G2019S mutation in LRRK2 gene, encoding for the leucine-rich repeat kinase 2, is the most prevalent mutation linked to familial and sporadic Parkinson’s disease (PD). Early detection of G2019S by fast genetic testing is very important to guide PD’s diagnosis and support patients and their family caregivers for better management of their life according to disease’s evolution. Methods In our study, a genetic PD’s diagnosis tool was developed for large scale genotyping using Kompetitive Allele Specific PCR (KASP) technology. We investigated G2019S’s frequency in 250 Tunisian PD patients and 218 controls. Results We found that 33.6% of patients and 1.3% of controls were carriers. Demographic characteristics of patients with G2019S had no differences compared with non-carrier patients. Thereby, we could emphasize the implication of G2019S in PD without any distinctive demographic factors in the studied cohort. Sixty patients out of 250 were genotyped using Taqman assay and Sanger sequencing. The genotyping results were found to be concordant with KASP assay. Conclusions The G2019S mutation frequency in our cohort was similar to that reported in previous studies. Comparing to Taqman assay and Sanger sequencing, KASP was shown to be a reliable, time and cost effective genotyping assay for routine G2019S screening in genetic testing laboratories.

Summary Background Ampicillin, the first semi-synthetic penicillin active against Enterobacteriaceae, was released onto the market in 1961. The first outbreaks of disease caused by ampicillin-resistant strains of Salmonella enterica serotype Typhimurium were identified in the UK in 1962 and 1964. We aimed to date the emergence of this resistance in historical isolates of S enterica serotype Typhimurium. Methods In this retrospective, whole-genome sequencing study, we analysed 288 S enterica serotype Typhimurium isolates collected between 1911 and 1969 from 31 countries on four continents and from various sources including human beings, animals, feed, and food. All isolates were tested for antimicrobial drug susceptibility with the disc diffusion method, and isolates shown to be resistant to ampicillin underwent resistance-transfer experiments. To provide insights into population structure and mechanisms of ampicillin resistance, we did whole-genome sequencing on a subset of 225 isolates, selected to maximise source, spatiotemporal, and genetic diversity. Findings 11 (4%) of 288 isolates were resistant to ampicillin because of acquisition of various β lactamase genes, including bla TEM-1 , carried by various plasmids, including the virulence plasmid of S enterica serotype Typhimurium. These 11 isolates were from three phylogenomic groups. One isolate producing TEM-1 β lactamase was isolated in France in 1959 and two isolates producing TEM-1 β lactamase were isolated in Tunisia in 1960, before ampicillin went on sale. The vectors for ampicillin resistance were different from those reported in the strains responsible for the outbreaks in the UK in the 1960s. Interpretation The association between antibiotic use and selection of resistance determinants is not as direct as often presumed. Our results suggest that the non-clinical use of narrow-spectrum penicillins (eg, benzylpenicillin) might have favoured the diffusion of plasmids carrying the bla TEM-1 gene in S enterica serotype Typhimurium in the late 1950s. Funding Institut Pasteur, Sante publique France, the French Government's Investissement d'Avenir programme, the Fondation Le Roch-Les Mousquetaires.

International audience; In Tunisia, chronic cutaneous leishmaniasis due to Leishmania tropica is an important health problem. Its spreading has not been fully elucidated. Information on sandfly vectors, as well as their associated Leishmania species, is of paramount importance since vector dispersion is one of the major factors responsible for pathogen dissemination. Ninety-seven unfed females belonging to the genera Sergentomyia and Phlebotomus were collected between June and August 2015 using sticky paper traps. Polymerase chain reaction-restriction fragment length polymorphism analysis of the internal transcribed spacer 1and sequencing were used for Leishmania detection and identification. In total, 650 sandflies were captured and identified (380 males and 270 females). Ninety-seven unfed females were tested for the presence of Leishmania parasite DNA. Six Phlebotomus sergenti were found positive for L. tropica. This novel finding enhances the understanding of the cycle extension of L. tropica outside its original focus of Tataouine.

International audience; Autoimmune lymphoproliferative syndrome (ALPS) is a prototypic disorder of impaired apoptosis characterized by autoimmune features and lymphoproliferation. Heterozygous germline or somatic FAS mutations associated with preserved protein expression have been described. Very rare cases of homozygous germline FAS mutations causing severe autosomal recessive form of ALPS with a complete defect of Fas expression have been reported. We report two unrelated patients from highly inbred North African population showing a severe ALPS phenotype and an undetectable Fas surface expression. Two novel homozygous mutations have been identified underlying rare splicing defects mechanisms. The first mutation breaks a branch point sequence and the second alters a regulatory exonic splicing site. These splicing defects induce the skipping of exon 6 encoding the transmembrane domain of CD95. Our findings highlight the requirement of tight regulation of FAS exon 6 splicing for balanced alternative splicing and illustrate the importance of such studies in highly consanguineous populations.

In the last few years, different surveillances have been published in Africa, especially in northern countries, regarding antimicrobial resistance among husbandry animals. Information is still scarce, but the available data show a worrying picture. Although the highest resistance rates have been described against tetracycline, penicillins and sulphonamides, prevalence of plasmid-mediated quinolone resistance genes and extended spectrum β-lactamase (ESBL) are being increasingly reported. Among ESBLs, the CTX-M-1 group was dominant in most African surveys. Within this group, CTX-M-15 was the main variant both in animals and humans, except in Tunisia where CTX-M-1 was more frequently detected among Escherichia coli from poultry. Certain blaCTX-M-15-harbouring clones (ST131/B2 or ST405/D) are mainly identified in humans, but they have also been reported in livestock species from Tanzania, Nigeria or Tunisia. Moreover, several reports suggest an inter-host circulation of specific plasmids (e.g. blaCTX-M-1-carrying IncI1/ST3 in Tunisia, IncY- and Inc-untypeable replicons co-harbouring qnrS1 and blaCTX-M-15 in Tanzania and the worldwide distributed blaCTX-M-15-carrying IncF-type plasmids). International trade of poultry meat seems to have contributed to the spread of other ESBL variants, such as CTX-M-14, and clones. Furthermore, first descriptions of OXA-48- and OXA-181-producing E. coli have been recently documented in cattle from Egypt, and the emergent plasmid-mediated colistin resistance mcr-1 gene has been also identified in chickens from Algeria, Tunisia and South Africa. These data reflect the urgent need of a larger regulation in the use of veterinary drugs and the implementation of surveillance programmes in order to decelerate the advance of antimicrobial resistance in this continent.

International audience; Toxoplasmosis is an important zoonosis caused by an obligate intracellular parasitic protozoan, Toxoplasma gondii. The disease is distributed worldwide and can affect all warm-blooded vertebrates, including humans. The present review aimed to collect, compile and summarize the data on the prevalence of T. gondii infection in humans and animals in the five North African countries (Morocco, Algeria, Tunisia, Libya and Egypt). Published data from national and international databases were used. Distribution patterns and risk factors for T. gondii infection are discussed, focusing on biotic and abiotic factors. This review is a comprehensive epidemiological analysis of T. gondii infection in North Africa and will therefore be a useful tool for researchers. It can also be used to propose or enhance appropriate national toxoplasmosis control programs.; La toxoplasmose est une zoonose importante causée par un protozoaire parasite intracellulaire obligatoire, Toxoplasma gondii. La maladie est répandue dans le monde entier, chez tous les vertébrés à sang chaud, y compris les humains. La présente étude visait à collecter, compiler et résumer les données sur la prévalence de l’infection par T. gondii chez l’homme et les animaux dans les cinq pays d’Afrique du Nord (Maroc, Algérie, Tunisie, Libye et Égypte). Les données publiées dans des bases de données nationales et internationales ont été utilisées. Les schémas de distribution et les facteurs de risque d’infection par T. gondii sont discutés, en se concentrant sur les facteurs biotiques et abiotiques. Cette synthèse est une analyse épidémiologique complète de l’infection par T. gondii en Afrique du Nord et sera donc un outil intéressant pour les chercheurs. Elle peut également être utilisée pour proposer ou renforcer des programmes nationaux appropriés de contrôle de la toxoplasmose.

Background Skin cancers (SC) are complex diseases that develop from complex combinations of genetic and environmental risk factors. One of the most severe and rare genetic diseases predisposing to SC is the Xeroderma pigmentosum (XP) syndrome. Objectives First, to identify the genetic etiology of XP and to better classify affected patients. Second, to provide early molecular diagnosis for pre-symptomatic patient and finally to offer genetic counseling for related individuals. Methods Whole Exome Sequencing (WES) and Run Of Homozygosity (ROH) were performed for two patients belonging to two different multiplex consanguineous families. The identified mutations were confirmed by Sanger sequencing and researched in ten Tunisian families including a total of 25 affected individuals previously suspected as having XP group V (XP-V) form. All patients had mild dermatological manifestations, absence of neurological abnormalities and late onset of skin tumors. Results Screening for functional variations showed the presence of the ERCC2 p.Arg683Gln in XP14KA-2 patient and a novel mutation, DDB2 p. (Lys381Argfs*2), in XP51-MAH-1 patient. Sanger sequencing and familial segregation showed that the ERCC2 mutation is present at a homozygous state in 10 affected patients belonging to 3 families. The second mutation in DDB2, is present at a homozygous state in 5 affected cases belonging to the same family. These two mutations are absent in the remaining 10 affected patients. The ERCC2 c.2048G > A mutation is present in a medium ROH region (class B) suggesting that it mostly arises from ancient relatedness within individuals. However, the c.1138delG DDB2 mutation is present in a large ROH region (class C) suggesting that it arises from recent relatedness. Conclusion To our knowledge, this is the first study that identifies XP-D and XP-E complementation groups in Tunisia. These two groups are very rare and under-diagnosed in the world and were not reported in North Africa.

International audience; الخلاصة: تعتبر النيسرية السحائية - أحد الأسباب الرئيسية لالتهاب السحايا الجرثومي وغيره من الأمراض المُعدية الخطيرة - مسؤولة عن حوالي ثلث حالات التهاب السحايا الجرثومي في مستشفى الأطفال بمدينة تونس. وقد حُدِّد توزع المجموعات المصلية، والحساسية للمضادات الحيوية، والمميزات المستضدية والجزيئية، للنيسرية السحائية المعزولة من مرضى تتراوح أعمارهم بين 3 أيام و 13 عاماً في الفترة ما بين فبراير/شباط 1998 ويونيو/حزيران 2013. لقد تم عزل 107 سلالات غازية للنيسيرية السحائية. وشوهد انخفاض في الحساسية للبنسلين G لدى %55.7 من المعزولات، مع تدني مستوى المقاومة في جميع الحالات؛ كما أظهرت %28.4 مستوى متدنياً من المقاومة للأموكسيسيلين. وكانت السلالات المعزولة للمجموعة المصلية B الأكثرَ شيوعاً (%80.4)، تليها المجموعة المصلية C (%12.2) و A (%5.6). وكان لمعزولات المجموعة المصلية A نفس الصيغة المستضدية (A:4:P1.9)، ونفس النواحي المتغيرة VR1 و VR2 و VR3، وتنتمي إلى نفس المجمع النسيلي (CC5). وكانت معزولات المجموعتين المصليتين B و C أكثر تغايراً وذات صيغ مستضدية متعددة. وكان المجمع النسيلي الأكثر شيوعاً في هذه السلالات CC35. وقد استأثرت المجموعة المصلية B بنسبة مئوية كبيرة من معزولاتنا، مع وجود تنوع ملحوظ.; Neisseria meningitidis, a leading cause of bacterial meningitis and other serious infections, is responsible for approximately one-third of cases of bacterial meningitis in the Children's Hospital of Tunis. The serogroup distribution, antibiotic susceptibility and antigenic and molecular characteristics of N. meningitidis isolates were determined in patients aged 3 days-13 years between February 1998 and June 2013. In all 107 invasive strains of N. meningitidis were isolated. Reduced susceptibility to penicillin G was seen in 55.7% of isolates, with a low level of resistance in all cases; 28.4% showed a low level of resistance to amoxicillin. Serogroup B isolates were the most frequent (80.4%), followed by serogroups C (12.2%) and A (5.6%). Isolates of serogroup A had the same antigenic formula (A:4:P1.9), the same variable regions VR1, VR2 and VR3, and belonged to the same clonal complex (CC5). Isolates of serogroups B and C were more heterogeneous with several antigenic formulae. The most frequent clonal complex in these isolates was CC35. Serogroup B accounted for a large percentage of our isolates with marked diversity.; La bactérie Neisseria meningitidis, l'une des causes principales de méningite bactérienne et d'autres infections graves, est responsable d'environ un tiers des cas de méningite bactérienne à l'Hôpital d'enfants de Tunis. La distribution par sérogroupe, la sensibilité aux antibiotiques et les propriétés antigéniques et moléculaires des isolats de N. meningitidis ont été déterminées chez des patients âgés de 3 jours à 13 ans entre février 1998 et juin 2013. En tout, 107 souches invasives de N. meningitidis ont été isolées. Une sensibilité réduite à la pénicilline G a été observée dans 55,7 % des isolats, avec un bas niveau de résistance pour tous les cas ; et 28,4 % ont montré un bas niveau de résistance à l'amoxicilline. Les isolats du sérogroupe B étaient les plus fréquents (80,4 %), suivis par les sérogroupes C (12,2 %) et A (5,6 %). Les isolats du sérogroupe A avaient la même formule antigénique (A:4:P1.9), les mêmes régions variables VR1, VR2 et VR3, et appartenaient au même complexe clonal (CC5). Les isolats des sérogroupes B et C étaient davantage hétérogènes, avec plusieurs formules antigéniques. Le complexe clonal le plus fréquent de ces isolats était CC35. Le sérogroupe B comptait pour un pourcentage élevé des isolats avec une diversité marquée.

International audience; HLA-G is highly expressed in cancer. Also, it is associated to its progression. Here, we explored the relationship between two HLA-G polymorphisms with breast cancer (BC) and tried to make a correlation with sHLA-G levels. We genotyped 104 patients with BC and 83 controls (CTRL) for HLA-G 14-bp insertion/deletion (Ins/Del) and HLA-G +3142 C > G polymorphisms. The mutations were identified with PCR and PCR-RFLP. The sHLA-G dosage was performed on plasma samples by a specific ELISA. A significant association with BC was found concerning the G allele in the +3142 C > G polymorphism (p = 0.0004). The G/G genotype is the protective genotype (1 % in BC patients vs. 13.1 % in CTRL, OR 0.065, 95 % CI 0.008-0.523). No statistically significant differences were observed for the 14-bp Ins/Del polymorphism between BC patients and controls frequencies. The protection by G/G genotype of +3142 C > G polymorphism is maintained in young patients (< 50 years, p = 0.0006) and in early-diagnosed BC patients (< 50 years, p = 0.0033). In addition, an association was found between the haplotypes inferred by both HLA-G polymorphisms and BC susceptibility. Indeed, the (DelG) haplotype is found as the protective haplotype against BC (OR 0.269, 95 % CI 0.081-0.895, p = 0.023). The ELISA dosage of sHLA-G revealed increased levels in BC compared to CTRL (p < 0.0001). We demonstrated also that sHLA-G is closely associated with advanced stages of BC without significance. sHLA-G is increased in TNM IV and SBR III subgroups. It is also enhanced in patients with a tumor size over 20 mm and in triple-negative patients. Taken together, our findings demonstrate, for the first time, the association of HLA-G +3142 C > G polymorphism with BC susceptibility in Tunisian population. Our results revealed also a potential implication of sHLA-G in advanced stages of BC.

International audience; To investigate the possible role of the hospital environment in the dissemination of extended-spectrum-beta-lactamase (ESBL)-producing Escherichia coli isolates, 300 samples were taken during 2013 from abiotic surfaces (n = 250), healthcare worker hands (n = 27), and hands of patients (n = 23) in a Tunisian Hospital. ESBL-producing E. coli isolates were recovered in 3.7% of analyzed samples (4% abiotic surfaces; 4.3% hands of patients; 0% in healthcare worker hands), and one isolate/sample was further studied. The characterization of beta-lactamase genes, as well as the genetic environment of bla CTX-M gene, was performed by PCR and sequencing. The ESBL genes found were as follows: bla CTX-M-15 (eight isolates), bla CTX-M-15 +bla SHV-12 (two isolates), and bla SHV-12 (one isolate). The bla TEM-1b gene was detected in seven ESBL-positive isolates. The orf477 was found downstream of bla CTX-M-15 gene in 10 strains, whereas the ISEcp1 sequence was identified upstream of this gene in two isolates. The analysis of class 1 integrons by PCR and sequencing revealed five positive isolates with the following gene cassette arrangements: dfrA1-aadA1 (two isolates), aadA1 (two isolates), and aadA2 (one isolate). The virulence-encoding genes aer, eae, bfp, and hly were detected by PCR in six, four, four, and three isolates, respectively. The following sequence types and associated phylogroups were detected among ESBL-producing strains: ST167-phylogroup-A (six isolates) and ST131-phylogroup-B2 (two isolates). In conclusion, the hospital environment could be a reservoir of multiresistant bacteria, including ESBL-positive E. coli isolates, which could be acquired by the patient population, and strict control measures should be established to minimize this problem.

International audience; Spore-forming bacteria, principally Bacillus species, are important contaminants of milk. Because of their high heat resistance, Bacillus species spores are capable of surviving the heat treatment process of milk and lead to spoilage of the final product. To determine the factors influencing the contamination of milk, spore-forming bacteria occurrence throughout the UHT milk production line during winter, spring, and summer was studied. The obtained results confirm that the total viable rate decreases rapidly throughout the production line of UHT milk showing the efficiency of thermal treatments used. However, the persistent high rate of spore-forming bacteria indicates their high heat resistance , especially in spring and summer. In addition, a significant variation of the quality of raw milk according to the location of the collecting centers was revealed. The molecular identification showed a high degree of diversity of heat-resistant Bacillus species, which are isolated from different milk samples. The distribution of Bacillus species in raw milk, stored milk, bactofuged milk, pasteurized milk, and UHT milk were 28, 10, 16, 13, and 33%, respectively. Six Bacillus spp. including Bacillus licheniformis (52.38%), Bacillus pumilus (9.52%), Bacillus sp. (4.76%), Bacillus sporothermodurans (4.76%), Terribacillus aidingensis (4.76%), and Paenibacillus sp. (4.76%) were identified in different milk samples.

International audience; The presence of extended-spectrum beta-lactamase and plasmid-mediated AmpC beta-lactamase producing Enterobacteriaceae (ESBL-Eb and pAmpC-Eb, respectively) was analyzed in 57 wastewater and 57 surface-water samples in Tunisia. Twenty-four of the 57 wastewater samples (42.1%) and one of the 57 surface-water samples (1.7%, a river that received effluents of a wastewater-treatment-plant) contained ESBL-Eb or pAmpC-Eb; one ESBL/pAmpC-Eb per positive sample was further characterized. Beta-lactamase genes detected were as follows: blaCTX-M-1 (10 Escherichia coli),blaCTX-M-15 (eight E. coli, one Klebsiella pneumoniae, one Citrobacter freundii), blaCTX-M-14 (one E. coli) and blaCMY-2 (four E. coli). The blaTEM-1, blaOXA-1 or blaSHV-1 genes were also found in 72% of these isolates. The ISEcp1, orf477 or IS903 sequences were found upstream or downstream of blaCTX-M genes. Class 1 integrons were present in 16 of the 25 ESBL-Eb/pAmpC-Eb strains (64%), and contained five different gene-cassette arrays. Most of the strains (76%) showed a multiresistant phenotype and qnr genes were identified in four strains. Molecular typing of ESBL/CMY-2-producing E. coli isolates showed 23 different PFGE-patterns and 15 different sequence-types (ST10, ST46, ST48, ST58, ST69, ST101, ST117, ST131, ST141, ST288, ST359, ST399, ST405, ST617, and the new ST4530); these strains were ascribed to phylogroups A (11 isolates), B1 (3 isolates), D (6 isolates) and B2 (3 isolates). From one to five plasmids were detected in each strain (size from 30 kb to > 240 kb) and ESBL or pAmpC genes were transferred by conjugation in 69.5% of the E. coli strains. In conclusion, ESBL-Eb and pAmpC-Eb strains are frequently detected in wastewater samples and they might be a source for dissemination in other environments with repercussion in public health.

International audience; X-linked agammagobulinemia (XLA) is a primary immunodeficiency caused by Bruton's tyrosine kinase (BTK) gene defect. XLA patients have absent or reduced number of peripheral B cells and a profound deficiency in all immunoglobulin isotypes. This multicenter study reports the clinical, immunological and molecular features of Bruton's disease in 40 North African male patients. Fifty male out of 63 (male and female) patients diagnosed with serum agammaglobulinemia and non detectable to less than 2 % peripheral B cells were enrolled. The search for BTK gene mutations was performed for all of them by genomic DNA amplification and Sanger sequencing. We identified 33 different mutations in the BTK gene in 40 patients including 12 missense mutations, 6 nonsense mutations, 6 splice-site mutations, 5 frameshift, 2 large deletions, one complex mutation and one in-frame deletion. Seventeen of these mutations are novel. This large series shows a lower frequency of XLA among male patients from North Africa with agammaglobulinemia and absent to low B cells compared with other international studies (63.5 % vs 85 %). No strong evidence for genotype-phenotype correlation was observed. This study adds to other reports from highly consanguineous North African populations, showing lower frequency of X-linked forms as compared to AR forms of the same primary immunodeficiency. Furthermore, a large number of novel BTK mutations were identified and could further help identify carriers for genetic counseling.

International audience; Objectives. Although a relationship between obesity and metabolic consequences with thyroid function has been reported, the underlying pathogenesis is not completely known. In the current study, we evaluated the thyroid function in obese and/or diabetic patients compared to healthy normal weight peers, exploring the possible association between components of metabolic syndrome and thyroid function parameters. Methods. We recruited 108 subjects (56 male and 52 female). In all subjects, thyroid stimulating hormone (TSH), free thyroxine (FT4), fasting plasma levels of insulin and glucose, homeostasis model assessment for insulin resistance, and obesity parameters were assessed. Results. We found that circulating levels of TSH and FT4 were significantly increased in overweight and obese subjects. However, the data do not reveal any change of these hormones in diabetics. Multivariate linear regression analysis showed that TSH was directly associated with both obesity and insulin resistance parameters (p < 0.05). FT4 was negatively associated only with obesity parameters (p < 0.05). Conclusions. Our data strongly support that the changes of thyroid hormones may be influenced by adiposity and its metabolic consequences, such as insulin resistance. This relationship can be explained by a cross talk between adipose tissue release and thyroid function. Nevertheless, metformin treatment seems to affect thyroid function in diabetic patients by maintaining plasma thyrotropin levels to subnormal levels.

A novel thermophilic, anaerobic, Gram-stain-positive, terminal-spore-forming bacterium was isolated from an upflow anaerobic filter treating abattoir wastewaters in Tunisia. This strain, designated LIND6LT2T, grew at 40–60 °C (optimum 50–55 °C) and at pH 6.0–8.5 (optimum pH 7.0–7.5). It did not require NaCl for growth, but tolerated it up to 2 %. Sulfate, thiosulfate, elemental sulfur, sulfite, nitrate and nitrite were not used as electron acceptors. Growth of LIND6LT2T was inhibited by sulfite (2 mM). Strain LIND6LT2T used cellobiose, glucose, mannose, maltose, mannitol, sucrose and xylose as electron donors. The main fermentation products from glucose metabolism were acetate, formate, butyrate and isobutyrate. The predominant cellular fatty acids were C16 : 0 (68.4 %) and C14 : 0 (8.3 %). The G+C content of the genomic DNA was 35.2 mol%. On the basis of its phylogenetic and physiological properties, a new genus and species, Defluviitalea saccharophila gen. nov., sp. nov., are proposed to accommodate strain LIND6LT2T, placed in Defluviitaleaceae fam. nov. within the phylum Firmicutes, class Clostridia, order Clostridiales. Strain LIND6LT2T ( = DSM 22681T = JCM 16312T) is the type strain of Defluviitalea saccharophila, which itself is the type species of Defluviitalea.

International audience; Several outbreaks of human West Nile virus (WNV) infections were reported in Tunisia during the last two decades. Serological studies on humans as well as on equine showed intensive circulation of WNV in Tunisia. However, no virus screening of mosquitoes for WNV has been performed in Tunisia. In the present study, we collected mosquito samples from Central Tunisia to be examined for the presence of flaviviruses. A total of 102 Culex pipiens mosquitoes were collected in September 2014 from Central Tunisia. Mosquitoes were pooled according to the collection site, date and sex with a maximum of 5 specimens per pool and tested for the presence of flaviviruses by conventional reverse transcription heminested PCR and by a specific West Nile virus real time reverse transcription PCR. Of a total of 21 pools tested, 7 were positive for WNV and no other flavivirus could be evidenced in mosquito pools. In addition, WNV was isolated on Vero cells. Phylogenetic analysis showed that recent Tunisian WNV strains belong to lineage 1 WNV and are closely related to the Tunisian strain 1997 (PAH 001). This is the first detection and isolation of WNV from mosquitoes in Tunisia. Some areas of Tunisia are at high risk for human WNV infections. WNV is likely to cause future sporadic and foreseeable outbreaks. Therefore, it is of major epidemiological importance to set up an entomological surveillance as an early alert system. Timely detection of WNV should prompt vector control to prevent future outbreaks. In addition, education of people to protect themselves from mosquito bites is of major epidemiological importance as preventive measure against WNV infection.

International audience; The -158 (C→T) nucleotide change, known as Xmn I polymorphism, occurs in (G)γ-globin gene promoter, and results in elevated fetal hemoglobin (HbF). We found this mutation in cis of a β(0)-thalassemia splicing mutation. Despite the complete absence of adult HbA, the phenotype was only moderately severe with no detectable alteration of α-globin gene expression. Interestingly, the β-globin locus haplotype has not been described to bear the (G)γ promoter mutation. Using a gene-specific real-time RT-PCR approach, we found a dramatic increase of both (G)γ and (A)γ mRNA accumulated in the reticulocytes, suggesting that the (G)γ-promoter mutation, alone or in association with another genetic modification, alters in concert the transcription of both (G)γ and (A)γ. This observation is discussed in light of recent regulatory model for β-globin locus.

International audience; Cutaneous leishmaniasis (CL), a public health problem in Tunisia, is associated to three species: Leishmania (L.) infantum, L. major and L. killicki. Accurate and sensitive procedures for the diagnostic of Leishmania infection and for species identification are required to enable adequate treatment and appropriate control measures. Several PCR-methods are applied for the diagnosis and the identification of Leishmania parasites such as PCR-restriction fragment length polymorphism (PCR-RFLP), DNA sequencing, hybridization probes and real-time PCR (RT-PCR). In this study, PCR-RFLP and RT-PCR were performed on skin scrapings from 27 patients with confirmed CL by microscopic examination, in order to compare their usefulness and efficiency for identification of Leishmania species in routine diagnostic laboratories. Identification of Leishmania species was successfully achieved in 96.3% and 81.5% respectively. Agreement between using internal transcribed spacer 1 (ITS1)-PCR-RFLP and kDNA-RT-PCR assays was 70% (19/27). Characterization problems using RT-PCR were mainly due to the difficulties in analyzing the melting temperatures. ITS1-PCR-RFLP and kDNA-RT-PCR presented an interesting alternative to conventional methods for the identification of Leishmania parasites from clinical samples. Both PCR assays can be used in a routine diagnostic, however, further prospective studies including largest sampling, are required to determine their performances in a routine use.

Many infectious diseases were described in North Africa in 18th-19th centuries by European travellers. Most of them were allegedly imported by new migrant populations coming from sub-Saharan, European or Middle East countries. Plague outbreaks have been described since the Black Death as diseases of the Mediterranean harbours. Charles Nicolle and his collaborators at the Pasteur Institute were witnesses to the extinction of plague and typhus fever in Tunisia. Both could be considered as endemo-epidemic diseases propagated by ancient nomad communities for centuries. Typhus was exported to other countries; plague was imported by Mediterranean travellers but also hid in unknown wild-animal reservoirs. The role of the bite of a rat's flea was not confirmed and the pneumonic form might have prevailed in the medieval North African cities. Association between plague, typhus, flu and other causes of immune deficiencies could explain the high morbidity and mortality caused by plague in the past. The authors comment the local history of plague at the light of the evolutionary laws of infectious disease proposed by Charles Nicolle in 1930.; De nombreuses maladies infectieuses ont été décrites en Afrique du Nord aux XVIIIe et XIXe siècles par des voyageurs européens. La plupart d’entre elles, soupçonnait-on, étaient importées par les nouvelles populations migrantes venues des pays sub-sahariens, d’Europe ou d’Asie. Au début du XXe siècle, Charles Nicolle et ses collaborateurs ont assisté à l’extinction de la peste et du typhus exanthématique au nord de l’Afrique. Il ressort de leurs observations de nouvelles hypothèses sur l’origine, l’évolution et les modalités de transmission locale de la peste: celle d’ancêtres nomades réservoirs et propagateurs de maladies très anciennes contrastant avec des maladies nouvelles importées par les vagues de migrations successives, celle de la prédominance des formes pulmonaires dans les épidémies urbaines, celle de l’association de la peste avec la grippe, le typhus et autres causes de déficits immunitaires collectifs expliquant la morbidité et la mortalité spécifique exceptionnelle observées certaines années dans certaines villes. Les idées qu’on peut dire évolutionnistes sur le destin des maladies infectieuses, proposées par Charles Nicolle en 1930, ont été à la fois l’aboutissement et la trame des recherches pasteuriennes en Afrique du Nord.

A total of 271 stool specimens were collected from children (diarrheagenic, n = 115 and control, n = 54) and adults (diarrheagenic, n = 73 and control, n = 29) from Tunis, Tunisia, and processed to detect bacterial enteropathogens, parasites, and viruses. Diarrheagenic Escherichia coli (DEC) were identified by their virulence genes (polymerase chain reaction) and adherence patterns (tissue culture assays). The most frequently isolated enteric pathogens from diarrheagenic children were enterotoxigenic E. coli (ETEC, 32.3%), enteroaggregative E. coli (EAEC, 11.3%), enteroinvasive E. coli (EIEC, (11.3%), adenovirus (10.4%), enterohemorrhagic E. coli (EHEC, 10.4%), and Salmonella spp. (9.5%). For children in the control group, ETEC (37%), EAEC (15%), EHEC (11.1%), and typical enteropathogenic E. coli (EPEC, 11.1%) were the most common enteric pathogens. In adults in the diarrheagenic group, Salmonella spp. (34.2%), ETEC (12.3%), adenovirus (7%), and Shigella spp. (4%) were the most common enteric pathogens. In adults in the control group, ETEC (31%) was the most common enteric pathogen. Multiple pathogens were recovered from 22% of the diarrheagenic children and 7% of the diarrheagenic adults. Escherichia coli strains showed high resistance rates to tetracycline, streptomycin, and β-lactams. The most frequent combinations were ETEC-rotavirus and ETEC-adenovirus. Pulsed-field gel electrophoresis for DEC indicated a large number of DEC clones (five major clones) persistent in the community reservoir for a considerable period of time that caused diarrhea in the population. This suggests the confluence of small epidemics by clonally related DEC strains circulating in this region.

International audience; Hypereosinophilic syndromes (HES) are a heterogenous group of rare disorders characterized by sustained and otherwise unexplained overproduction of eosinophils with organ involvement and consecutive dysfunction. Detection of the FIP1L1-PDGFRA fusion gene or the corresponding cryptic 4q12 deletion in HES supports the diagnosis of chronic eosinophilic leukemia (CEL) and provides a molecular explanation for the pathogenesis of this disorder. We screened seven Tunisian patients fulfilling the WHO criteria of HES for the presence of the FIP1L1-PDGFRA fusion gene using nested reverse transcription polymerase chain reaction on peripheral blood samples. Four of the seven patients were positive for this fusion gene. Sequence analysis revealed a substantial heterogeneity of the fusion transcripts due to the involvement of several FIP1L1 exons. All patients were male. The median age at diagnosis was 24 years (range, 18-50); one patient had a history of hypereosinophilia of more than 10 years. Two patients had clinically important and symptomatic eosinophilic endomyocardial disease with thrombotic events. Splenomegaly was constant in FIP1L1-PDGFRA positive CEL but not in the other HES patients (only 1/3).; Les hyperéosinophilies essentielles sont caractérisées par une hyperéosinophilie supérieure à 1500 éléments/mm 3 persistante au-delà de sixmois, après exclusion d ’ une éosinophilie réactionnelle. Différentes altérations viscérales peuvent s’y associer leur pronostic est souvent redou-table et jusqu’à une période récente, il n’existait pas de traitement spécifique ou réellement efficace. En 2003 un gène de fusion :FIP1L1-PDGFR alpha a été retrouvée de façon récurrente dans ces syndromes, ce gène de fusion est généré à la suite d’une délétion interstitielle en 4q12 qui est cryptique à l’ analyse cytogénétique standard, la protéine chimérique résultante a une activité tyrosine kinase constitutive qui répond au traitement par l’ imatinib utilisé par ailleurs pour le traitement de la leucémie myéloïde chronique. Nous rapportons quatre cas d’hyperéosinophilie associéesà ce remaniement génétique

International audience; Background: Tunisia is classified as an area of middle endemic for hepatitis B virus (HBV) infection, however little is known about hepatitis Delta virus (HDV) infection. Objectives: This study aimed to address the prevalence of HDV infection, to identify possible risks factors, and to analyze the genetic diversity of HDV strains that are spreading in Tunisia. Study design: A retrospective large-scale study including 1615 HBsAg positive patients, native of the North East coast of Tunisia, recruited from Gastroenterology departments, was conducted. Demographic, epidemiological, ethnical, clinical and biological data were recorded. HBV and HDV serological analyses and DNA and RNA viral load quantification were performed. Genotyping of HBV and HDV strains was performed using nucleotide sequencing followed by phylogenetic analyses. Results: The study population included 819(50.7%) men and 796(49.3%) women; aged 12-90 years (mean age 41 + 13 years). A very low prevalence of HDV infection, 2% was observed. No risk factor, except a history of hospitalization for surgery was found. All HDV strains belonged to genotype 1, with a wide distribution within the HDV-1 group. They all share the African amino acid marker, a serine at position 202 of the large Delta protein. HBV genotypes were distributed as follows: HBV/D1 (56.8%), HBV/D7 (40.9%), and HBV/A2 (2.3%). Conclusion: Tunisia is a low endemic region for HDV infection, due to an efficient policy of HBV infection control. HDV-1 is the sole genotype found, with a high diversity within this group. Further studies are ongoing in order to better characterize and manage the HBV/HDV-infected patients according to the genetic variability of the viral strains. (C) 2015 Elsevier B.V. All rights reserved.

International audience; Pistacia lentiscus L. is known in some Tunisian forest area by its fixed oil used in traditional medicine as an antiseptic product. This investigation is the first to study the antimicrobial activity of P.lentiscus edible oil and its phenolic extract. Oil was extracted from fruits harvested from six provenances located in Tunisia. The antimicrobial activity was tested using disc diffusion assay and the broth dilution method. Kbouch and Sidi Zid oils were most efficient (p

International audience; OBJECTIVE: To assess the value of multifocal chorioretinitis and of clinical manifestations and biologic parameters in the diagnosis of West Nile virus (WNV) infection.PATIENTS AND METHODS: We conducted a prospective, controlled case series study during an outbreak of WNV infection between August 15 and October 24, 2003, of 64 consecutive patients who presented with clinical manifestations consistent with WNV disease. In each patient, standardized clinical and biologic data were collected. An ophthalmologic examination searching particularly for multifocal chorioretinitis was performed.RESULTS: Of 64 patients who presented primarily with meningitis and/or encephalitis, 36 had IgM antibodies against WNV. The WNV-infected patients tended to be older (median age of 54 years vs 46 years in WNV infection and control groups, respectively) and more frequently had diabetes (30% vs 7% in WNV infection and control groups, respectively; P = .03). Multifocal chorioretinitis was found in 75% of WNV-infected patients but in no patient in the control group (P = .001). Blood glucose and amylase levels were higher in WNV-infected patients, whereas serum sodium levels were lower. The cerebrospinal fluid leukocyte count and protein levels were significantly higher in WNV meningitis or encephalitis. Overall, multifocal chorioretinitis had 100% specificity and 73% sensitivity (88% when only patients with meningitis or encephalitis were analyzed) for the diagnosis of WNV. Multivariate analysis disclosed multifocal chorioretinitis as the only predictor of WNV infection (odds ratio, 62; 95% confidence interval, 6-700; P = .001).CONCLUSION: Multifocal chorioretinitis appears to be a specific marker of WNV infection, particularly in patients who present with meningoencephalitis. An ophthalmologic examination should be part of the routine evaluation of such patients.

International audience; Six temporary wetlands in the region of Sejenane (Mogods, NW Tunisia) were studied in order to characterize the aquatic flora and fauna and to quantify their spatio-temporal variability. Samplings of aquatic fauna, phytosociological relevés, and measurements of the physicochemical parameters of water were taken during four different field visits carried out during the four seasons of the year (November 2009-July 2010). Despite the strong anthropic pressures on them, these temporary wetlands are home to rich and diversified biodiversity, including rare and endangered species. Spatial and temporal variations affect fauna and flora differently, as temporal variability influences the fauna rather more than the plants, which are relatively more dependent on spatial factors. These results demonstrate the interest of small water bodies for maintaining biodiversity at the regional level, and thus underscore the conservation issues of Mediterranean temporary wetlands that are declining on an ongoing basis currently.

International audience; Echinococcus granulosus infection is a preventable zoonosis of human and veterinary public health importance in Tunisia. We aimed to comprehensively quantify human and animal echinococcosis losses in Tunisia. Itemized cost menus were developed for the health of both domestic animals and humans, and for productivity monetary losses. The incidence and prevalence of the disease in all species were obtained from national and personal reports. The domestic animal and human costs of echinococcosis in Tunisia were estimated using age-stratified rates and losses, productivity losses (including those not formally employed) and Monte Carlo sampling to represent the uncertainty inherent in some epidemiological and economic values. Echinococcosis in Tunisia causes significant direct and indirect losses in both humans and animals of approximately US dollars 10-19 million annually. These estimates are based on numerous methodological improvements over previous studies and are of considerable consequence relative to Tunisia's US dollars 21.2 billion gross domestic product. A cost-benefit analysis of control programmes using the methodological advances presented here and regional comparison to other endemic diseases is warranted. These may provide information to assist policy decision-makers in prioritizing the allocation of scarce resources.

International audience; BackgroundCord blood is an important source of stem cells. However, nearly 90% of public cord blood banks have declared that they are struggling to maintain their financial sustainability and avoid bankruptcy. The objective of this study is to evaluate how characteristics of cord blood units influence their utilization, then use this information to model the economic viability and therapeutic value of different banking strategies.MethodsRetrospective analysis of cord blood data registered between January 1st, 2009 and December 31st, 2011 in Bone Marrow Donor Worldwide. Data were collected from four public banks in France, Germany and the USA. Samples were eligible for inclusion in the analysis if data on cord blood and maternal HLA typing and biological characteristics after processing were available (total nucleated and CD34+ cell counts). 9,396 banked cord blood units were analyzed, of which 5,815 were Caucasian in origin. A multivariate logistic regression model assessed the influence of three parameters on the CBU utilization rate: ethnic background, total nucleated and CD34+ cell counts. From this model, we elaborated a Utilization Score reflecting the probability of transplantation for each cord blood unit. We stratified three Utilization Score thresholds representing four different banking strategies, from the least selective (scenario A) to the most selective (scenario D). We measured the cost-effectiveness ratio for each strategy by comparing performance in terms of number of transplanted cord blood units and level of financial deficit.ResultsWhen comparing inputs and outputs over three years, Scenario A represented the most extreme case as it delivered the highest therapeutic value for patients (284 CBUs transplanted) along with the highest financial deficit (USD 5.89 million). We found that scenario C resulted in 219 CBUs transplanted with a limited deficit (USD 0.98 million) that charities and public health could realistically finance over the long term. We also found that using a pre-freezing level of 18 x 108 TNC would be the most cost-effective strategy for a public bank.ConclusionOur study shows that a swift transition from strategy A to C can play a vital role in preventing public cord blood banks worldwide from collapsing.

Summary Purpose To assess vision loss, identify affected anatomical sites, determine etiologies and potentially avoidable causes in students attending two institutions for the visually disabled in Tunis city. Methods A visit for a complete ophthalmological examination was performed. All students attending these schools were recruited in our study. The World Health Organisation Programme for the Prevention of Blindness (WHO/PBL) examination record for children was used. Data was analysed by the SPSS version 17 statistical software. Results A total of 172 students were recruited with mean age of 11.9 ± 3.3 years (between 6 and 18 years). One hundred and thirty-seven (79.6%) were under 16 years. The sex-ratio was 1.17. Ninety students (52.3%) had low vision and eighty-two (47.7%) were blind. We reported retina (29%), whole globe (29%), globe appears normal (11%) and optic nerve (9.8%) as the common sites of ocular abnormalities. Retinal dystrophy (22.7%) and congenital glaucoma (22.7%) were the most reported ocular diseases. The main etiologies were hereditary (54.1%) and unknown (30.8%). Consanguinity was reported in 108 students (62.8%), and fifty-five students (32%) had a positive family history. Overall, 50.5% (87/172) of ocular diseases were potentially treatable or preventable. Conclusion Retinal dystrophy and congenital glaucoma were the most common eye diseases. Heredity was the main etiology, and consanguinity was high. To decrease their incidence, awareness of the family members of the risks of consanguinous marriage and appropriate therapy for congenital glaucoma/cataract may significantly improve the child's visual prognosis.

International audience; One-hundred-nine samples of 18 different farms (49 of food-vegetables, 41 of soil and 19 of irrigation water) and 45 vegetable food samples of 13 markets were collected in Tunisia. These samples were inoculated in MacConkey agar plates supplemented with cefotaxime (2 μg/ml). ESBL-producing Enterobacteriaceae (ESBL-Eb) were detected in 10 of the 109 farm samples (vegetables, 8.2%; soil, 7.3%; water, 15.8%), and in 4 of 45 vegetables of markets (8.9%), recovering 15 ESBL-Eb. Isolates and ESBL genes detected were: Escherichia coli (n = 8: 5 blaCTX-M-1, 2 blaCTX-M-15 and one blaCTX-M-14), Citrobacter freundii (n = 4: 3 blaCTX-M-15 and one blaSHV-12), Enterobacter hormaechei (n = 2: 2 blaCTX-M-15) and Klebsiella pneumoniae (n = 1, blaCTX-M-15). The ISEcp1 sequence was found upstream of blaCTX-M genes in 13 of 14 strains (in three cases truncated by IS5), and orf477 or IS903 downstream. Class 1 integrons were detected in five strains and contained two gene cassette arrangements (dfrA17-aadA5 and aadA1). Most isolates tested showed a multiresistant phenotype. All blaCTX-M-15-positive strains carried the aac(6′)-1b-cr gene, that affects to amikacin–tobramycin–kanamycin–ciprofloxacin. Five ESBL-Eb strains carried genes of the qnr family. The 8 ESBL-positive E. coli isolates were typed as: ST58/B1 (n = 3) and ST117/D, ST131/B2, ST10/A, ST23/A, and the new ST3496/D (one strain, each). From 1–2 plasmids were detected in all ESBL-positive E. coli isolates (63–179 kb). The ESBL genes were transferred by conjugation in 4 blaCTX-M-1-positive E. coli strains, and transconjugants acquired a 97 kb IncI1 plasmid. ESBL-Eb isolates are frequently disseminated in vegetable farms and potentially could be transmitted to humans through the food chain.

International audience; Physiological and molecular characteristics of natural populations of Culex pipiens Linnaeus, 1758 (Diptera: Culicidae) were investigated to elucidate how this species is potentially involved in the transmission of West Nile virus in Tunisia. A total of 215 Cx. pipiens females from 11 breeding habitats were analysed in the laboratory to estimate autogeny and stenogamy rates. They were tested individually for the locus CQ11 to distinguish between the two Cx. pipiens forms, pipiens and molestus. All tested Cx. pipiens populations were stenogamous. Females from underground breeding sites were all autogeneous, whereas females from above-ground habitats were mostly anautogeneous. Of all the females tested, 59.7% were identified as pipiens, 22.4% as molestus, and 17.9% as hybrid pipiens/molestus. Furthermore, both Cx. pipiens forms and their hybrids were found to co-occur in sympatry in all sites. The results of this study represent the first evidence that both Cx. pipiens forms and their hybrids are present in Tunisia. Because hybrids able to act as bridge vectors are present in all studied habitats, Tunisia can be considered to have a high degree of receptivity for the establishment of West Nile virus zoonotic cycles.