[From gene to disease; progressive myoclonus epilepsy of Unverricht-Lundborg and mutations in the cystatin B gene]. / Van gen naar ziekte; progressieve myoclonusepilepsie van Unverricht-Lundborg en mutaties in het cystatine-B-gen.

Progressive myoclonus epilepsy type 1 of Unverricht-Lundborg (EPM1) is a rare disorder, associated with mutations in the cystatin B (CSTB) gene. The most prevalent molecular abnormality is an expansion of a dodecamer repeat in the promoter region of the CSTB gene, but point mutations in the CSTB gene have also been found. DNA examination may be useful in discriminating EPM1 from juvenile myoclonic epilepsy, and from other types of progressive myoclonus epilepsy. An early diagnosis is important to optimise treatment and to provide an adequate prognosis and prediction of recurrence.

Product and redox potential analysis of sauerkraut fermentation.

The relationships between the redox potential of the brine, during fermentation of white cabbage into sauerkraut of two early and two late fermentation processes, and the changes in the amount of sugars, organic acids, the redox potential of the brine and of the ascorbic acid redox couple, and pH are described. The trend in the change of the redox potential of the brine is the same for all four fermentation processes studied. In the first phase a sharp decrease in redox potential is followed by an increase in redox potential. In the second phase the redox potential is rather constant. This second phase is followed by another decrease in redox potential, which stabilizes at a minimum value, the third phase. It was observed that sugar fermentation and acid production mainly took place during the first and third phases, probably representing, respectively, the heterogeneous and homogeneous fermentation processes.

Submicroscopic Xpter deletion in a boy with growth and mental retardation caused by a familial t(X;14).

In a 3-year-old boy with short stature, developmental delay, and dry skin, steroid sulphatase deficiency and a submicroscopic terminal deletion of Xp were found. Except for the short stature, no major clinical signs of X-linked recessive chondrodysplasia punctata could be observed. His mother had lowered steroid sulphatase activity compatible with carriership for X-linked ichthyosis and a submicroscopic translocation (X;14)(p22.31;p11.1). This finding combined with a normal amplification of exons 1, 5, and 10 of the STS gene from propositus' DNA suggested a breakpoint upstream of the STS gene. The submicroscopic maternal translocation had important implications for genetic counseling. This case report illustrates that contiguous gene syndrome related to the Xpter region may have an atypical clinical presentation and the usefulness of combined clinical, biochemical, molecular, and fluorescence in situ hybridization analysis.

Prospective prenatal investigations on potential uniparental disomy in cases of confined placental trisomy.

In most reported cases of uniparental disomy (UPD) associated with confined placental mosaicism (CPM), a high level of mosaicism or a full trisomy was found in chorionic villi. At the time that we started our investigations, it was not quite clear whether fetal UPD also existed in the more frequently occurring low levels of mosaicism. During a 4-year period, a follow-up amniocentesis was performed in all cases of mosaic or non-mosaic trisomy detected in chorionic villus (CV) semi-direct preparations and suspected to be confined to the placenta. We performed fluorescent in situ hybridization (FISH) on uncultured amniotic fluid cells to differentiate between generalized mosaicism and CPM. We found 29 cases of CPM and we determined the incidence of UPD in 23 of these cases. Normal biparental chromosome contributions were found in 22 cases. In one case, we detected a maternal heterodisomy for chromosome 16. UPD appeared to be a rare phenomenon in the cases of CPM (type I and/or type III) that we encountered in 3958 consecutively investigated CV samples, and is not the cause of the pregnancy complications found in seven out of 23 cases with CPM.

Rapid antibody test for prenatal diagnosis of fragile X syndrome on amniotic fluid cells: a new appraisal.

Fragile X syndrome is caused by mutations in the FMR1 gene and is one of the most frequent forms of inherited mental retardation in males. Postnatal and prenatal diagnosis of fragile X syndrome is feasible by direct DNA analysis. A new approach to prenatal diagnosis of fragile X syndrome in amniotic fluid cells is described, using a rapid and simple antibody test on uncultured amniotic fluid cells. The test requires 1 ml of amniotic fluid and the results of this antibody test are available on the same day as the amniocentesis.

Analysis of unstable DNA sequence in FMR1 gene in Polish families with fragile X syndrome.

The unstable DNA sequence in the FMR1 gene was analyzed in 85 individuals from Polish families with fragile X syndrome in order to characterize mutations responsible for the disease in Poland. In all affected individuals classified on the basis of clinical features and expression of the fragile site at X(q27.3) a large expansion of the unstable sequence (full mutation) was detected. About 5% (2 of 43) of individuals with full mutation did not express the fragile site. Among normal alleles, ranging in size from 20 to 41 CGG repeats, allele with 29 repeats was the most frequent (37%). Transmission of premutated and fully mutated alleles to the offspring was always associated with size increase. No change in repeat number was found when normal alleles were transmitted.

Positional mapping of loci in the DiGeorge critical region at chromosome 22q11 using a new marker (D22S183).

The majority of patients with DiGeorge syndrome (DGS) and velo-cardio-facial syndrome (VCFS) and a minority of patients with non-syndromic conotruncal heart defects are hemizygous for a region of chromosome 22q11. The chromosomal region that is commonly deleted is larger than 2 Mb. It has not been possible to narrow the smallest region of overlap (SRO) of the deletions to less than ca 500 kb, which suggests that DGS/VCFS might be a contiguous gene syndrome. The saturation cloning of the SRO is being carried out, and one gene (TUPLE1) has been identified. By using a cosmid probe (M51) and fluorescence in situ hybridization, we show here that the anonymous DNA marker locus D22S183 is within the SRO, between TUPLE1 and D22S75 (probe N25). A second locus with weak homology to D22S183, recognized by cosmid M56, lies immediately outside the common SRO of the DGS and VCFS deletions, but inside the SRO of the DGS deletions. D22S183 sequences are strongly conserved in primates and weaker hybridizing signals are found in DNA of other mammalian species; no transcripts are however detected in polyA+ RNA from various adult human organs. Probe M51 allows fast reliable screening for 22q11 deletions using fluorescence in situ hybridization. A deletion was found in 11 out of 12 DGS patients and in 3 out of 7 VCFS patients. Two patients inherited the deletion from a parent with mild (atypical) symptoms.

Loss of mutation at the FMR1 locus through multiple exchanges between maternal X chromosomes.

The mutation observed in the fragile X syndrome, an X-linked inherited disorder causing mental retardation, is almost exclusively an expanded CGG repeat in the first exon of the FMR1 gene. Here we describe a daughter of a female carrier, who inherited the fragile X premutation chromosome based on haplotype analysis using flanking markers. However, the CGG repeat sequence and the intragenic polymorphic marker FMRb showed the normal maternal alleles, while two other intragenic markers, FMRa and FRAXAC2 and other, more distant markers, showed the risk haplotype. Since FMRa and FRAXAC2 are located in between the markers CGG and FMRb, this results in patches of normal and fragile X sequences in the FMR1 gene of the daughter. This observation is very likely due to gene conversion. As this daughter received a normal CGG repeat region, we expect that her risk to have affected offspring is the same as the population risk. The observed phenomenon would therefore represent a back mutation at the FMR1 locus.

Refined localization of TSC1 by combined analysis of 9q34 and 16p13 data in 14 tuberous sclerosis families.

Tuberous sclerosis (TSC) is a heterogeneous trait. Since 1990, linkage studies have yielded putative TSC loci on chromosomes 9, 11, 12 and 16. Our current analysis, performed on 14 Dutch and British families, reveals only evidence for loci on chromosome 9q34 (TSC1) and chromosome 16p13 (TSC2). We have found no indication for a third locus for TSC, linked or unlinked to either of these chromosomal regions. The majority of our families shows linkage to chromosome 9. We have refined the candidate region for TSC1 to a region of approximately 5 cM between ABL and ABO.

Strategy for reliable prenatal detection of normal male carriers of the fragile X syndrome.

Prenatal diagnosis of fragile X syndrome identifying full mutations has been described. Here we report on a case of a prenatal test concerning a normal male carrier of the fragile X syndrome. Southern blot analysis of the fragile X gene resulted in the identification of a premutation in DNA isolated from the chorionic villus sample. Using a polymerase chain reaction (PCR)-based assay the CGG repeat length was determined to be 82 CGG repeat units. Confirmation of this premutation in the chorionic villus sample was based on the cytogenetic analysis of the expression of the fragile site at Xq27 applied on a cordocentesis-derived blood sample.

DNA diagnosis of the fragile X syndrome in a series of 236 mentally retarded subjects and evidence for a reversal of mutation in the FMR-1 gene.

The cloning of the FMR-1 gene and the identification of an expanded CGG repeat in DNA of fragile X patients has made reliable DNA diagnosis feasible. Southern blotting and PCR assays of the CGG repeat in an unselected series of 236 mentally retarded subjects resulted in the identification of 10 new fragile X families. Reevaluation of previously assessed fragile X families resulted in the first observation of the presence of a reversal of mutation in the FMR-1 gene.

Conservation of CGG region in FMR1 gene in mammals.

Only two of the fragile sites found in humans (FRAXA and FRAXE) have been associated with a clinical phenotype. In mentally retarded individuals with cytogenetic expression of FRAXA a CGG repeat in the FMR1 gene is amplified. Fragile sites are found in many animals species. We have analyzed the FRAXA region containing the CGG repeat in several different species by PCR amplification. In most mammals this region could be amplified; the number of copies of the repeat is deduced.

Founder effect in a Belgian-Dutch fragile X population.

For many years, the high prevalence of the fragile X syndrome was thought to be caused by a high mutation frequency. The recent isolation of the FMR1 gene and identification of the most prevalent mutation enable a more precise study of the fragile X mutation. As the vast majority of fragile X patients show amplification of an unstable trinucleotide repeat, DNA studies can now trace back the origin of the fragile X mutation. To date, de novo mutations leading to amplification of the CGG repeat have not yet been detected. Recently, linkage disequilibrium was found in the Australian and US populations between the fragile X mutation and adjacent polymorphic markers, suggesting a founder effect of the fragile X mutation. We present here a molecular study of Belgian and Dutch fragile X families. No de novo mutations could be found in 54 of these families. Moreover, we found significant (P < 0.0001) linkage disequilibrium in 68 unrelated fragile X patients between the fragile X mutation and an adjacent polymorphic microsatellite at DXS548. This suggests that a founder effect of the fragile X mutation also exists in the Belgian and Dutch populations. Both the absence of new mutations and the presence of linkage disequilibrium suggest that a few ancestral mutations are responsible for most of the patients with fragile X syndrome.

Linkage analysis with chromosome 15q11-13 markers shows genomic imprinting in familial Angelman syndrome.

Angelman syndrome (AS) and Prader-Willi syndrome (PWS) have become the classical examples of genomic imprinting in man, as completely different phenotypes are generated by the absence of maternal (AS) or paternal (PWS) contributions to the q11-13 region of chromosome 15 as a result of deletion or uniparental disomy. Apparently, most patients are sporadic cases. The genetic mechanism underlying familial AS has remained enigmatic for a long time. Recently, evidence has been emerging suggesting autosomal dominant inheritance of a detectable or undetectable defect in a gene or genes at 15q11-13, subject to genomic imprinting. The present report describes an unusually large pedigree with segregation of AS through maternal inheritance and apparent asymptomatic transmission through several male ancestors. Deletion and paternal disomy at 15q11-13 were excluded. However, the genetic defect is still located in this region, as we obtained a maximum lod score of 5.40 for linkage to the GABA receptor locus GABRB3 and the anonymous DNA marker D15S10, which have been mapped within or adjacent to the AS critical region at 15q11-13. The size of the pedigree allowed calculation of an odds ratio in favour of genomic imprinting of 9.25 x 10(5). This family illustrates the necessity of extensive pedigree analysis when considering recurrence risks for relatives of AS patients, those without detectable deletion or disomy in particular.

The mutation delta F508 on Dutch cystic fibrosis chromosomes: frequency and relation to patients age at diagnosis.

We tested 190 chromosomes from Dutch cystic fibrosis (CF) patients and carriers for the presence or absence of the major CF mutation delta F508. This mutation was found on 77% of the Dutch CF chromosomes. We observed a significant difference in the distribution of the ages at diagnosis between homozygotes for delta F508 and the other patients. delta F508 homozygotes tend to be identified as patients at neonatal or infantile age. The age at diagnosis of patients with at least one unknown allele, on the other hand, ranged between neonatal and young adult age.

Mirror blank testing by real-time holographic interferometry.

This paper describes an application of real-time holographic interferometry to the testing of unworked mirror blanks. The thermal test of a 70-cm diam, fused silica, eggcrate mirror blank and the mechanical test of a 28-cm mirror blank are included.