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Two decades after its original publication, a new human genome sequence has just been published. Far from being an incremental improvement, it is at last really complete, covering each chromosome from one end to the other, with full elucidation of repeated sequences and an extremely low error rate. This is a major advance that tremendously increases our knowledge of our genome and will lead to important scientific and clinical developments.

International audience; Genetic recombination is a major force driving the evolution of some species of positive sense RNA viruses. Recombination events occur when at least two viruses simultaneously infect the same cell, thereby giving rise to new genomes comprised of genetic sequences originating from the parental genomes. The main mechanism by which recombination occurs involves the viral polymerase that generates a chimera as it switches templates during viral replication. Various experimental systems have alluded to the existence of recombination events that are independent of viral polymerase activity. The origins and the frequency of such events remain to be elucidated to this day. Furthermore, it is not known whether non-replicative recombination yields products that are different from recombinants generated by the viral polymerase. If this is the case, then non-replicative recombination may play a unique role in the evolution of positive sense RNA viruses. Finally, the sparse data available suggest that non-replicative recombination does not necessarily involve only virus-specific sequences. It is thus possible that the non-replicative recombination observed in virus-focused studies may in fact reveal a more generalized mechanism that is non-specific to virus RNAs.

A systematic search for non-conventional open reading frames in human DNA reveals a large number of small ORFs encoding peptides generally smaller than 100 amino-acids. These ORFs are transcribed and translated into small proteins, which are demonstrated to have functional significance by bulk CRISPR inactivation. Evidence is also found for bicistronic mRNAs including such a small ORF upstream of a canonical coding sequence. These findings add a new facet to our understanding of biological processes.

Current data estimate that mobile genetic elements represent more than one-half of the human genome. The literature is constantly updating data following the evolution of sequencing techniques and of algorithms for genome analyses. This review aims to provide an overview of the topic showing the complexity given by the various designations and classifications found in scientific papers. A particular focus is made on retrotransposons, including Endogenous RetroViruses (ERV), to introduce a second article focusing on their activation and their involvement in physiological functions and/or pathological mechanisms associated with diseases like multiple sclerosis (MS) or amyotrophic lateral sclerosis (ALS).

Complex diseases usually harbour hereditary factors linked with multiple susceptibility genes. The additive effects of genetic and environmental factors are responsible for the pathology. The impact of heredity has been demonstrated through family studies, but also, and mostly, through the study of adopted people and twins. Recently, genome wide association studies (GWAS) allowed the identification of many susceptibility genes for most complex diseases. However, a large part of the heritability is still missing, probably because of insufficient exploration of rare genetic variants and/or epigenetic factors. The ultimate goal of these genetic studies is the definition of an individual risk leading to specific preventive measures (model "predict and prevent"), but this purpose remains very remote for the majority of complex diseases.

Cancers can be considered as gene diseases. A number of mechanisms leading to cancer have been identified through the discovery of structural alterations of genes called 'oncogenes' and 'tumour suppressor genes'. Somatic and germinal mutations are rare but play a determinant role in the emergence of cancer, while common and frequent variations (polymorphisms) play a role in cancer susceptibility and in the effects of anticancer drugs (efficacy and toxicity). After a general overview on the structural and functional organisation of the human genome, we present here some of the techniques aimed at the identification of structural DNA variations. We present afterwards some examples of the role that play polymorphic constitutive variations of the genome in the occurrence of cancer (molecular epidemiology) and the activity of anticancer drugs (pharmacogenetics).

A detection and quantification method specific of genotype G hepatitis B virus (G-HBV) was developed and used to study the epidemiology and evolution of G-HBV co-infection during antiviral therapy.A multiplex real time PCR was developed and validated on A-HBV and G-HBV plasmid mixes. G-HBV infection was sought for on chronic hepatitis B carriers followed in our institution.Two primers, flanking the specific G-HBV 36 nucleotide insertion and two probes, including one specific for the insertion, were validated for a multiplex real time PCR. This new tool was well correlated to commercially available quantification systems within a 2 to 7 log(10) IU/mL range. On A- and G-HBV clonal mixes, G-HBV was quantified with a precision between 5 to 10%. On HBV-HIV infected patients, a 25% G-HBV prevalence was found, while it was below 1% in HBV mono-infected patients. Longitudinal G-HBV quantifications on five treated patients indicate that G-HBV is not selected by antiviral treatment.A specific method for G-HBV quantification within multi-genotype mixes was set up. The important G-HBV prevalence in HIV patients was unexpected. Our data are not in favour of an intrinsic resistance of G-HBV to current antivirals.

C. difficile is a spore-forming anaerobic enteropathogen. This bacillus is responsible for virtually all cases of pseudomembranous colitis and for 15 to 25% of cases of antibiotic-associated diarrhoea. Clostridium difficile associated-infections (CDI) have a wide range of clinical features which vary from mild uncomplicated diarrhoea to severe debilitating disease, paralytic ileus, toxic megacolon, or even perforation and sometimes death. Risk factors for CDI include age65 years, previous hospitalization and recent antibiotic therapy. Main virulence factors for this pathogen are toxins A and B. A third toxin, the binary toxin, has been found in up to 10% of strains from infected patients. For some years, a new hypervirulent strain has emerged. This epidemic strain belongs to PCR-ribotype 027 and is responsible for outbreaks with increased mortality and severity in North America and Europe. The most effective antibiotics for treatment are oral metronidazole and vancomycin. Control of CDI needs to prevent the emergence of CDI by minimizing the number of patients exposed to antimicrobials and to limit cross transmission.

Unlike group A, a few studies have interested other groups of the rotavirus, especially in Tunisia. The role of rotavirus C (RVC) infection is underestimated because of its sporadic nature. The aim of our study was to develop rapid diagnostic procedures of RVC by using an internal positive control of reverse transcription PCR (RT-PCR).The internal positive control (386pb) was designed from the recombinant baculovirus BacVP6C containing the full length cDNA of the Cowden strain gene 5 (1353pb). A fragment of 596pb was amplified by PCR using the BacVP6C DNA ds as template. Then, a central part of 210pb was deleted and the remaining fragment (386pb) was cloned into pGEM-3Zf(+) plasmid between SP6 and T7 RNA polymerase promoters.The obtained recombinant plasmid "pIAM1" was then used for the generation of the internal positive control by in vitro transcription. The sensibility of the RT-PCR was about 3.66×10(5) molecules of RNA/μl.The use of a shorter positive control, as compared to the wild type, allows increased specificity of the RT-PCR reaction, and could be used for efficient diagnostic and surveillance of RVC-caused diseases.

In 1998, the description of Phlebotomus riouxi emphasised the difficulty to differentiate the female from the closely related P. chabaudi, a suspected vector of Leishmania killicki in several foci in Tunisia. In order to be able to distinguish the females of these two species, we started a molecular study based on 37 Algerian and Tunisian specimens. The alignment of the sequences of the cytochrome b of the mitochondrial DNA and their analysis using Neighbor-Joining, maximum likelihood and maximum parsimony shows the individualisation of two species including an intraspecific variability. Following a morphological approach, it is not possible to distinguish the females on the basis of their spermathecae. A new character is proposed: the presence of anterolateral teeth of the pharyngeal armature for P. chabaudi, never observed in P. riouxi. However, a molecular typing is necessary at the present time for a sure identification of the females.

Leber congenital amaurosis (LCA) is a the earliest and most severe form of retinal dystrophy responsible for congenital blindness. LCA has genetic heterogeneity and the study of this disease is elucidating the genetics and molecular interactions involved in the development of the retina. To date, 11 LCA genes have been mapped, ten of which have been identified. The CEP290 gene has been shown to account for Joubert and Senior-Loken syndromes and to be a frequent cause of nonsyndromic LCA. We report here the first Arab patient, born to consanguineous parents, with Leber congenital amaurosis attributable to mutation of the CEP290 gene.

This study aimed at describing the evolution of the epidemiological pattern of hepatitis A in New Caledonia since 1986 and the recent epidemic which occurred in 2005-2006, regarding particularly its demographic and virological aspects and the public health response implemented. The annual or monthly activity records for Hepatitis A sero-diagnostic performed at the Pasteur Institute of New Caledonia were processed in a retrospective analysis (9723 samples tested for the detection of IgM to hepatitis A). Over the 2004-2006 period, a phylogenetic study of representative strains from New Caledonia and other Pacific islands was carried out by the French National Reference Laboratory for hepatitis A (Paul-Brousse hospital, Villejuif, France).The continuous improvement of hygiene that occurred in New Caledonia during the last two decades led to a dramatic drop in the frequency of hepatitis A among patients tested, ranging from an average value of 79 cases (14%) for the 1986-1999 period to 0 case from 2002. However, in 2005, a strong increasing number of confirmed cases was notified, mainly among young people (78% were under the age of 20). In 2006, this epidemic reached the island of Futuna where it involved more than 1% of the total population (56 cases). The phylogenetic study has confirmed the clonality of the virus circulating during this epidemic, not related to other regional strains (Fiji, Vanuatu, New Zealand) nor with a New Caledonian strain from the previous endemic period. This transition situation, with persistence of a high epidemic risk, should encourage the health authorities to implement adapted response strategies, based in particular on systematic case declaration and targeted immunisation programmes.

We report the first case of primary amoebic meningoencephalitis in a 9-year-old boy in Guadeloupe. The outcome was rapidly fatal in 7 days. The patient presumably acquired the infection by swimming and diving in a basin supplied by natural thermal water 1 week before onset of the disease. The possibility of a free-living amoeba infection was suspected both on the negativity of all bacterial and viral initial tests and on the observation of peculiar cells in stained cerebrospinal fluid samples. Although the amoeba was not isolated, Naegleria fowleri could be identified by polymerase chain reaction with specific primers on DNA extracted from frozen cerebrospinal fluid samples. Furthermore, as the internal transcribed spacer (ITS1) region of DNA is variable in length between the different strains of N. fowleri, sequencing of the amplified ITS1 demonstrated that the responsible N. fowleri strain belongs to a common genotype present in the American and European continent.

Ten years after their discovery, anelloviruses combine some characteristics making them particularly intriguing. In support of their extreme genetic diversity and high prevalence in various populations, their natural history is still poorly understood along with their implication in human health. These viruses have been identified in blood and blood-derived products, and are probably remarkable examples of co-existence and co-evolution in their various hosts. This article presents epidemiological and molecular characterizing this new viral family.

Almost 10% of mammalian coding mRNAs contain in their 3' untranslated region a sequence rich in adenine and uridine residues known as AU-rich element (ARE). Many of them encode oncogenes (for instance c-Myc and c-Fos), cell cycle regulators (cyclin D1, A1, B1), cytokines (TNFalpha, IL2) and growth factors (GM-CSF) which are overexpressed in cancer or inflammatory diseases due to increased mRNA stability and/or translation. AREs are recognized by a group of proteins, collectively called AUBPs which display various functions. For instance, HuR/ELAV is mainly known to protect ARE-containing mRNAs from degradation, while AUF1, TTP and KSRP act to destabilize their bound target mRNAs and TIA/TIAR to inhibit their translation. Alterations in ARE sequences or AUBP abundance, cellular localization or activity due to post-translational modifications such as phosphorylation can promote or enhance malignancy or perturb immune homeostasis. Here, c-myc and TNFalpha are chosen as examples to illustrate how altered 3' UTR gene regulation impacts on pathologies.

Microsatellite markers have a great discriminating power. Widely exploited in many disciplines such as forensic science, medical genetics, conservation biology and molecular ecology, they are also used in human population genetics to illuminate our origins. However, strikingly, their fundamental evolutionary mechanisms remain obscure, because of the difficulty of disentangling the complex and numerous factors involved. After a brief summary of their basic characteristics, the concept of life cycle size-dependant is explored. The major mechanisms known to explain the four different phases of their life (conception, birth, growing and senescence/renaissance) are discussed. Emerging questions about their neutrality are also investigated, pointing out a real need to improve our understanding of their mutational dynamics.

Twenty years ago, the production of mice whose genomes have been deliberatly modified revolutionised biology. Indeed, it is now possible to eliminate a gene's expression to various levels in desired locations, and also to broadcast these genetic modifications created in vitro to the progeny. The isolation and culture of embryonic stem cells (ES) and the discovery of the mechanism of homologous recombination between two sequences of DNA in the 80's, have contributed to the development of site-directed mutagenesis. Today, site specific mutagenesis by homologous recombination in embryonic stem cells is a powerful technique and is widely used throughout the world. In parallel, new techniques to invalidate targeted genes are emerging. These genetics tools, which we will introduce, allow for a better understanding of a gene's function both in fundamental and clinical research. It is now possible to create murine models of human genetic diseases including Lesch-Nhyan syndrome, Adenomatous Polyposis and Duchenne muscular dystrophy which we will discuss as examples.

Down syndrome caused by chromosome 21 trisomy is the most common genetic cause of mental retardation in humans. Disruption of the phenotype is thought to be the result of gene-dosage imbalance. Variations in chromosome 21 gene expression in Down syndrome were analyzed in lymphoblastoid cells derived from patients and control individuals. Of the 359 genes and predictions displayed on a specifically designed high-content chromosome 21 microarray, one-third were expressed in lymphoblastoid cells. We performed a mixed-model analysis of variance to find genes that are differentially expressed in Down syndrome independent of sex and interindividual variations. In addition, we identified genes with variations between Down syndrome and control samples that were significantly different from the gene-dosage effect (1.5). Microarray data were validated by quantitative polymerase chain reaction. We found that 29% of the expressed chromosome 21 transcripts are overexpressed in Down syndrome and correspond to either genes or open reading frames. Among these, 22% are increased proportional to the gene-dosage effect, and 7% are amplified. The other 71% of expressed sequences are either compensated (56%, with a large proportion of predicted genes and antisense transcripts) or highly variable among individuals (15%). Thus, most of the chromosome 21 transcripts are compensated for the gene-dosage effect. Overexpressed genes are likely to be involved in the Down syndrome phenotype, in contrast to the compensated genes. Highly variable genes could account for phenotypic variations observed in patients. Finally, we show that alternative transcripts belonging to the same gene are similarly regulated in Down syndrome but sense and antisense transcripts are not.

It has long been known that gene regulation is mostly achieved via protein-nucleic acid interactions. However, the role of RNA factors in gene control has been recently growing given the implication of new RNA-based gene regulation mechanisms such as microRNAs and related short-interfering RNAs gene expression inactivation mechanisms. Recent studies have demonstrated that the involvement of RNA in fundamental gene-control processes is even more extensive. Prokaryotic messenger RNAs carry highly structured domains known as riboswitches within their 5'-untranslated regions. Each riboswitch is able to bind with high specificity their cellular target metabolite, without the involvement of a protein cofactor. Upon metabolite binding, the messenger RNA undergoes structural change that will ultimately lead to the modulation of its genetic expression. Riboswitches can alter gene expression at the level of transcription attenuation or translation initiation, and can up- or down-regulate gene expression by harnessing appropriate changes in the mRNA structure. Here, we provide an overview of the adenine riboswitch, one of the smallest riboswitch and one of the few that activates gene expression upon ligand binding. Several crystal structures have been obtained for the ligand-binding domain of this riboswitch providing us with an unprecedented glimpse about how riboswitches use their ligand to regulate gene expression. Moreover, mechanistic studies have recently shed light on the transcriptional regulation mechanisms of the adenine riboswitch suggesting that riboswitches may rely on the kinetics of ligand binding and the speed of RNA transcription, rather than simple ligand affinity. Riboswitches are particularly interesting because RNA-ligand interactions are potentially very important in the elaboration of antimicrobial agents.

The glycoprotein B (gB) is the major glycoprotein of the envelope of the human cytomegalovirus, it is encoded by UL55 open reading frame, implicated in host cell, entry cell to cell virus transmission and fusion of infected cells and a significant is an important target for immuno-reactions humoral and cellular. A prospective analysis of cytomegalovirus glycoprotein B genotype was conducted on 31 immunodepressed (transplant recipients and AIDS patients). The DNA was extracted directly from the bronchoalveolar liquid (BAL) of these patients. The gB genotypes of CMV was determined by using the polymerase chain reaction, followed by the digestion of two enzymes of restriction HinfI and RsaI. The distribution of the gB genotype of the CMV was: gB1 38,70%; gB2 25,80%; gB3 16,12% and gB4 19,35%. The analysis of the peptide sequence of this region (codons 437-520), indicate the variation was more frequent between codons 448-480.

Bacterial sterilization by the technology of plasma in post-discharge shows a growing interest. The main appeal of this new process resides in its action at dry and low temperature (60 degrees C). This technology would be therefore useful for the complex medical equipment, sensitive to the oxidization, humidity and/or requiring a temperature lower than 60 degrees C. The objective of this survey is to demonstrate the activity of an atomic flux emanating a plasma of pure molecular nitrogen on the bacterial DNA: does the plasma of nitrogen damage the genetic material?The bacteria tested (Bacillus stearothermophilus, Staphylococcus aureus MRSA and Helicobacter pylori) are cultivated on suitable agar, and the bacterial DNA is extracted from every CFU by the technique of the columns (High Pure PCR Template, Roche). Every quantity of DNA extracted is diluted in 1 ml pure water. Then, 50 microl of each of these solutions of DNA are laid down in sterile Nunclon's plates holes, which undergo an advanced emptiness cycle during 60 minutes. The DNA residues will be then introduced during 40 minutes in a plasma sterilization surrounding wall (Plasmalyse), Satelec) where the debit of nitrogen, the pressure and the temperature are adjusted respectively to 1 L x min(-1), 5 Torrs and 60 degrees C. The DNA so ''plasmalysé'', is recovered then by aspiration in 500 mul pure water and processed to undergo an amplification/detection by Real-Time PCR (LightCycler2.0, Roche). The DNA ''plasmalysé'' will be compared to the intact DNA control(1), to the DNA control control(2) having undergone the cycle of emptiness solely, as well as to the DNA control(3) solely heated to 60 degrees C during 40 min.The amplification curves demonstrated that the only advanced emptiness and the only heat don't have any activity on the bacterial DNA. On the other hand, the DNA ''plasmalysé'' shows a deterioration of the amplified sequences.The genomic bacterial DNA, once extracted, is damaged by the gaseous flux of nitrogen plasma. A new sterilization process of the medical material will presumably impose itself in medium-term.

Congenital antithrombin (AT) deficiency is the most thrombotic genetic abnormality of haemostasis. Total quantitative deficits are lethal as early as life intra-uterine. Only homozygous mutations concerning the heparin-binding site are compatible with life. We report here the case of an 18 years old patient with recurrent deep venous thrombosis of the inferior members. Haemostasis exploration shows a decreased AT activity (11%) in the presence of heparin while AT progressive activity and AT antigen are normal. Two other homozygous sisters are identified in this family study. Molecular study of AT gene show Arg47-Cys substitution, already reported in the literature with patients of different geographic origins. Treatment of patients with homozygous AT type HBS deficiency is similar that for patients with heterozygous AT deficiency; a continuous prophylactic anticoagulant treatment is always necessary and AT concentrates infusions are required in all situations needing curative heparin treatment.

Bruton's disease is the most frequently primary X-linked immunodeficiency. Patients are more susceptible to early and recurring infections associated with hypo/agammaglobulinemia and a severe B-cell deficiency. Moreover, 400 mutations were found in the XLA gene which codes the Btk tyrosine kinase and were identified as responsible for Bruton's disease. Genetic study was carried out with one group of patients named NECKER, composed by five XLA patients and two parents whose XLA gene was sequenced by an Italian crew. Results were obtained by PCR of 19 exons and initial/terminal intron's parts, followed by PCR-sequencing with universal primers and sequencing. The results from this study allowed the validation of the sequencing technique by comparing NECKER group data (equivalent results with Italian data). In addition, the mutation multiplicity (described or not, coding/non coding) need an exact analysis that should be given to clinicians through clear and trustful results. In this way, a strategy to analyse untreated results was created based on the mutation type. The genetic analysis could help physicians for uncertain diagnosis in immune defficiencies, allows proposing a genetic advice to the patient's family and the construction of a data base permits a best understanding of this disease.

Non-functional paraganglioma have not clinical or biological characteristics, so that the diagnostic is most of the time delayed and made on the occasion of advanced abdominal tumor or symptomatic metastasis management. Hereditary forms, notably those with SDHB mutation, seem to have a poor prognosis. On the other hand, and on the oposite to sporadic forms, they are the only ones to benefit from genetic testing which make possible, if positive, an earlier diagnostic, before apparition of symptoms, recurrence or metastasis. We report a case of non-functional malignant hereditary paraganglioma diagnosed belatedly and we will consider management problems raised by non-functional forms.

Every 10 seconds, one person in the world dies of tuberculosis (TB). It is estimated that one third of the world's population is latently infected with Mycobacterium tuberculosis. The proportion of multidrug-resistant strains of M. tuberculosis is increasing at an alarming rate in some parts of the world linked in part with the human immunodeficiency virus epidemic. For these reasons, TB remains a major public health problem, both in less-developed countries and in many industrialized countries, with 8-10 million new cases and 2 million deaths yearly in the world. Clinical, radiological and histological signs are not specific for tuberculosis or for other mycobacterial infections and allow only a presumptive diagnosis. In the same way, the tuberculin skin test is useful if the reaction is strong or phlyctenular because this test depends on various factors as previous BCG vaccination, contact or primary infection and host immune responses. The diagnosis of mycobacterial infection is proved only when bacilli are present in biological samples. Nevertheless, only 50% of cases in adults and 30% in infants have a positive bacteriological result. It seems necessary to develop new methods for a rapid and efficient diagnosis to optimize the therapy and the control of the epidemic. Laboratory testing in the mycobacterium field is experiencing more changes today than ever before. Determining what assays will be most useful to the clinician is a challenge, and acceptance of the new technology is under discussion. Progress in future will be linked probably to the progress of the genomic area. However the incidence rate is higher in less-developed countries, it is also important to develop now techniques possible to use in these countries. This review focuses on the current state-of-the-art resources useful for accurate and rapid laboratory diagnosis of mycobacterial infections.

International audience; The oculo-cerebro-renal syndrome of Lowe is a rare X-linked disorder, caused by the inositol biphosphate 5-phosphatase deficiency, localized to the Golgi complex. Several mutations were reported in patient's OCRL gene leading to enzyme deficiency. We report a Moroccan case of OCRL syndrome of Lowe with a neo mutation in exon 10. The patient aged of 19 months was referred to our medical centre because of a psychomotor retardation. He had a medical history of eye abnormalities including cataract and bilateral glaucoma, diagnosed when he was 5 weeks old. Cataract has been treated after chirurgical therapy but ocular hypertonia persisted. Physical examination revealed an axial hypotonia and walking difficulties. Laboratory tests revealed a moderate acidosis (20 mmol/L), a slight decrease of serum phosphate level (24 mg/L) and an increased serum phosphatase activity. Further studies showed mild proteinuria, urinary bicarbonates loosing and generalised hyperaminoaciduria. Based on both clinical and biological data, Lowe syndrome has been suggested. In this context, molecular investigation has been performed using dHPLC/sequencing techniques which allow identifying an original mutation c.776T>C (p.Phe259Ser), localized on the exon 10 of the OCRL gene. The mutation was not found in the probant's mother suggesting a neo mutation. Lowe syndrome is a rare hereditary X-linked disorder resulting from a variety of heterogeneous mutations of OCRL gene. Indeed, numerous mutations have been reported, variations were noted concerning their localization as well as their type. To our knowledge, this is the first report of the neo mutation c.776T>C of OCRL gene and the first published case report of the Lowe syndrome in a Moroccan patient.

The differentiation within the Mycobacterium tuberculosis complex (MTBC) based on phenotypic methods is long and does not give an unambiguous result in every case whereas the advance in genetic knowledge leads to new views. Thus, regions of difference (RD), that seem to characterize the species of the MTBC, have been identified. Amplification methods, targeted on these zones, have been developed then. The study of four regions (RD1, RD5, RD9, RD10) has been done on 64 isolates formerly identified thanks to phenotypic methods. Genotypic results confirm phenotypic identifications except in one case. This strain initially identified as M. tuberculosis and isolated from a Gabonese patient, corresponds, according to genotypic identification, to M. africanum. Since phenotypic characterization of M. africanum is difficult, this method would allow to accurately determine the true prevalence of this specie. Moreover, the study of RD10 doesn't seem to be informative. The amplification of only two RD, RD1 and RD9 carries out the identification of all M. tuberculosis and M. bovis BCG isolates. M. bovis and M. africanum will be then identified thanks to RD5. Thus, this easy and rapid method of identification of the major species of MTBC seems to be appropriated for a routine use.

Transitional cell carcinomas of the upper urinary tract (UUT-TCCs) are rare: they account for approximately 5% of all urothelial carcinomas. 30% of patients with UUT-TCC have a history of bladder TCC, but fewer than 2% of patients with bladder TCC have UUT-TCC. Tumor microsatellite instability (MSI) is an indicator of the clonal expansion of neoplasms; it was first identified in tumors from patients with hereditary non-polyposis colorectal carcinoma (HNPCC). UUT-TCC occurs in 5% of patients with HNPCC. High-frequency microsatellite instability is present in almost 20% of cases of sporadic UUT-TCC. In cases of UUT-TCC with high-frequency MSI, hereditary cancer must be sought, especially if the patient is younger than 60 years or has a personal or family history of an HNPCC-related cancer: such patients should undergo DNA sequencing for the MSH2 gene germline mutation. Invasive UUT-TCC has a poor prognosis. 5-year survival is less than 50% for stage T2-T3 tumors and less than 10% for T4 or N+/M+ tumors. The main prognostic factors are age and tumor stage and grade. High-frequency MSI is a positive prognostic factor, especially in patients younger than 70 years with T2/T3/N0-M0 tumors.

To study the lymphoid clonality on Tunisian B-cell lymphomas cases by polymerase chain reaction (PCR)-based techniques using DNA from paraffin-embedded tissues.Here we conducted a retrospective PCR clonality study on 73 cases of B-cell lymphomas and 12 reactive lymphoid tissues. The quality of DNA extracted was tested by beta-globin PCR. Consensus primers directed at the FRIII-VH and FRII-VH regions of the immunoglobulin heavy chain (IgH) gene were used to detect clonality.The results showed that 52 of 73 (71%) B-cell lymphomas exhibited good quality of amplifiable DNA. Clonality was found in 77% of cases using the set of primers FRIIIa/LJH/VLJH and in 65.5% using the set of primers FRIIa/LJH/VLJH. Lymphomas derived from pregerminal centre showed a high rate detection of clonal IgH gene rearrangement (100%) compared to other group of tumors derived from germinal centre or postgerminal centre (74.5%). None of the polyclonal controls gave a clonal pattern.This is the first large series of PCR clonality study of IgH gene rearrangements on B-cell lymphoma from Tunisia. Our results were similar to other reports in terms of sensitivity and specificity of these techniques and confirm the interest of that PCR for detecting clonal IgH gene rearrangements in lymphoma.

The diagnostic of invasive fungal infection is often difficult because of the low sensitivity of fungal culture from infected tissues. Here we have assessed the ability of a panfungal PCR targeted on the DNA region encoding the RNA genes followed by sequencing of the amplification products to detect and identify fungi from tissue biopsies. This assay allowed us to identify the microorganism responsible for an invasive fungal infection in three of our patients. In two cases, hepatosplenic candidiasis was suspected and Candida albicans DNA was detected from liver biopsies. The third patient was cared for a thymome and developed a manubrium osteitis caused by Scedosporium apiospermum.

The female of Phlebotomus (Anaphlebotomus) fertei Depaquit, LégerRobert, 2002 is described from Madagascar. The spermathecae have separated ducts, a body with an enlarged-sclerified dilatation and a deriviated large bladder with thin walls and long head. P. fertei female has the following characters: long antennal segments, pharyngeal armature with small teeth and cibarium with 15 to 30 denticles, and very long legs. Its remarkable spermatheca is closely related to that of P. berentiensis. The status of P. (Anaphlebotomus) berentiensis (LégerRodhain, 1978) is revised into this paper. The female is redescribed and the male is described for the first time. The latter has an unusual antennal formula (2/III-XII), a cibarium with four teeth and about 30 denticles, no tuft of setae on the coxite, four spines on the style, including a very long proximal one. Males and females were linked to each other based on morphological and molecular characteristics (second internal transcribed spacer of the ribosomal DNA and ND4 gene of the mitochondrial DNA). The inclusion of these taxa into the subgenus Anaphlebotomus Theodor, 1948 is exclusively based on male characters as listed by Theodor (style with four spines, no basal lobe on the coxite, paramere with two or three branches). The very heterogeneous characters of the females do not allow their use for the definition of this subgenus, which is probably not homogeneous.

The hepatitis C virus (HCV) non-structural NS5A protein has been controversially implicated in the resistance of HCV to interferon therapy in clinical studies. In Japan, mutations in the interferon sensitivity-determining region (ISDR) in the NS5A gene were associated with response to interferon therapy in patients infected with genotype 1b. In contrast, studies from Europe did not confirm such association. More recently, it has been suggested that the V3 domain outside the putative ISDR might also have amino acids changes that may be involved in the resistance to IFN. In this study, the relationship between NS5A mutations in ISDR and V3 domains and virological response to therapy were investigated.The NS5A gene was sequenced from 35 HCV genotype 1b infected patients at D0 of a prospective clinical trial of interferon therapy and interferon plus Ribavirin combination therapy.In the ISDR domain, we did not observe any significant differences in amino acids changes between responders (1.7 +/- 1.8, n = 19, range 0-6) and non-responders (1.1 +/- 0.8, n = 14, range: 0-3), (P = 0.483), to therapy before the beginning of treatment. In the V3 domain, we found more mutations in responders (6.5 +/- 1.9, range: 2-11) than in non-responders (4.7 +/- 1.2, range: 3-8) (P = 0.0013), before the beginning of treatment.Our results confirm that, in Europe, the ISDR domain is not predictive for treatment success but suggest that the V3 domain have greater variability in responders than non-responders.

In 1997, a group of hereditary corneal dystrophies was related to mutations in the TGFBI (BIGH3) gene. Within this group, some corneal dystrophies present particular biochemical features in that they are characterized by corneal amyloid deposition. Contrary to clinical and genetic knowledge, the biochemical characteristics of the encoded protein (Big-h3) and the mechanisms of its amyloid conversion remain unclear. We review the current knowledge on the Big-h3 protein and focus on the behavior of the codon 124 region. We discuss this protein's mechanisms of amyloid conversion from our results and previous reports as well as from other types of amyloidosis. These data provide a better understanding of the putative processes leading to the phenotypic variations linked with their respective codon 124 mutation.

Congenital toxoplasmosis results from foetus contamination by Toxoplasma gondii during pregnancy. It is a frequent and severe condition calling for close monitoring of mothers at risk. During the last decades, numerous advances have been made specially in the antenatal diagnosis. The congenital toxoplasmosis diagnosis relies currently on PCR test of amniotic fluid, with a sensitivity of 80%. More recently, real-time quantitative PCR has been developed to improve toxoplasmosis diagnosis. We therefore compared the diagnosis value of quantitative real-time PCR with our conventional PCR-hybridization for the diagnosis of congenital toxoplasmosis.

As a non-isotopic molecular cytogenetic technique, the primed in situ (PRINS) labelling reaction represents a major technological progress achieved in the past decade. It has become a routine technique for the microscopic visualization of specific DNA sequences in cells and nuclei and constitutes a good alternative to the fluorescence in situ hybridization (FISH) procedure. Among the multiple advantages that characterize the PRINS technique, specificity, rapidity, reliability, reproducibility, and cost-effectiveness can be mentioned. PRINS can be in addition associated with other techniques like FISH, indirect immunofluorescence, and nick translation. The most recent developments show the great potential of this technique. Now PRINS can be used to study single-copy genes and, consequently, can be routinely used to investigate deletions associated with microdeletion syndromes. Therefore, the PRINS technique has the potential to become a widely used molecular cytogenetic tool in clinics and research. This short review presents how the PRINS technique contributed to further the understanding of biological phenomena and describes the different possibilities and applications of the PRINS method in several biological and clinical fields (pre-implantation testing, prenatal, constitutional and oncologic genetic diagnosis).