ABSTRACT
In this case report we describe a 67-year-old male, admitted to the ICU with pneumonia who unexpectedly developed a fatal coma due to hyperammonaemia. At postmortem the diagnosis late-onset ornithine transcarbamylase deficiency was made. The non-specific clinical presentation, the rapid deterioration and incidentally the fatal outcome all underline the importance of recognition and knowledge of this genetic disorder. Several measures to treat and prevent potentially fatal episodes of hyperammonaemia are available, if only the disorder is recognised in time. In retrospect, several clues to the diagnosis were available in this fatal case, such as voluntary protein avoidance, as well as several male family members who died at a young age of an unknown cause. After his death, two daughters were discovered to be carriers of an OTC gene mutation, as well as his infant grandson. We emphasise the importance of obtaining ammonia levels in all patients with unexplained coma, seizures or cerebral oedema, irrespective of their age, especially in patients in the ICU or in an otherwise catabolic state.
Subject(s)
Delayed Diagnosis , Hyperammonemia/genetics , Late Onset Disorders/genetics , Ornithine Carbamoyltransferase Deficiency Disease/complications , Aged , Coma/genetics , Fatal Outcome , Humans , Male , Ornithine Carbamoyltransferase Deficiency Disease/diagnosisABSTRACT
We report on a child who presented clinical manifestations of both neurofibromatosis type 1 (NF1) and cherubism. With genetic testing, we found a mutation in the NF-1 gene, confirming the neurocutaneous disorder. Histology when correlated with radiological evaluation of a mandibular biopsy was consistent with cherubism. This is the first report in the literature of a child with proven neurofibromatosis type 1 and cherubism without extragnathic lesions. This emphasises that cherubism is a clinical phenotype that can be associated with a number of germline mutations involving SH3BP2, PTPN11 and NF1.
Subject(s)
Cherubism/complications , Genes, Neurofibromatosis 1 , Neurofibromatosis 1/complications , Adolescent , Base Sequence , Cherubism/diagnostic imaging , Cherubism/pathology , Child , Humans , Male , Mutation , Neurofibromatosis 1/diagnosis , Neurofibromatosis 1/genetics , Phenotype , RadiographyABSTRACT
During routine ultrasound screening at 12 weeks 5 days of gestation, a nuchal translucency of 7 mm, an omphalocele, and fetal hydrops were found and prompted chorionic villus sampling at 13 weeks 2 days. Chromosome analysis showed an unbalanced karyotype with an abnormal chromosome 14. The mother was a carrier of a translocation karyotype 46,XX,t(13;14) (q34;q32.2). In the fetus this gave rise to a partial trisomy 13q and partial monosomy 14q (fetal karyotype: 46,XX,der[14]t[13;14][q34;q32.2]). By Array-CGH on DNA extracted from a postmortem skin culture, a duplication of approximately 1.7 Mbp of the distal part of chromosome 13q34 and a deletion of approximately 6.0 Mbp of the distal part of chromosome 14q32.2 was demonstrated. Postmortem findings after termination of pregnancy at 14 weeks 6 days included, among others, a severe hypoplasia of the median part of the maxilla, no recognizable nose, a broad median palatoschisis, nonlobulated lungs, a horseshoe kidney with multicystic dysplasia, and decreased development of cortical cellularity in the thymus. These clinical manifestations and autopsy findings of the fetus are compared with those of previously published cases and the possible involvement in this pathology of the YY1 and JAG2 transcription factors and the BCL11b and SIVA-1 regulators of thymic development is discussed.
Subject(s)
Abnormalities, Multiple/genetics , Chromosomes, Human, Pair 14 , Face/abnormalities , Gene Deletion , Thymus Gland/abnormalities , Abortion, Eugenic , Adult , Chorionic Villi Sampling , Female , Gestational Age , Humans , In Situ Hybridization, Fluorescence , Intercellular Signaling Peptides and Proteins , Jagged-2 Protein , Male , Membrane Proteins/genetics , Membrane Proteins/metabolism , Nuchal Translucency Measurement , Nucleic Acid Hybridization/methods , Pregnancy , Translocation, Genetic , Trisomy , Ultrasonography, Prenatal , YY1 Transcription Factor/genetics , YY1 Transcription Factor/metabolismABSTRACT
OBJECTIVE: This study aimed to identify a marker chromosome and characterize the short arm of a derivative chromosome 5 in a foetus with the following karyotype: mos 47,XX,del(5)(p?),+i(5)(p10)[50]/48,XX,del(5)(p?),+i(5)(p10),+mar[25]. METHOD: Amniocentesis was performed in the 26th week of pregnancy because of ultrasound abnormalities (polyhydramnion and decreased amount of gastric filling). All classic banding techniques were performed. FISH and microdissection combined with reverse painting were used to reveal the exact origin of the marker and any extra material on the deleted chromosome 5p. The parents decided to continue the pregnancy and we compared the clinical features of the child born in week 34 with data from the literature on trisomy 5p. The possible contribution of trisomy of the centromeric region of chromosome 8 and trisomy 8p23.3-->8pter to this clinical picture was evaluated. RESULTS: GTG banding showed one normal and two aberrant chromosomes 5 [del(5)(p?) and i(5)(p10)] in all the cells examined. Furthermore, a supernumerary marker chromosome was present in approximately 30% of the cells. The marker was CBG positive and positive with the pancentromere probe, but dystamicinA/DAPI negative. It did not contain NOR-positive satellites. FISH proved this marker to be derived from the centromeric region of chromosome 8. MicroFISH disclosed the aberrant chromosome 5 as der(5)t(5;8)(p10;p23.3). The parent's karyotypes were normal. The baby showed the characteristic features of trisomy 5p syndrome. She died at the age of 15 days after cardiorespiratory arrest. CONCLUSION: The karyotype was interpreted as mos 47,XX,add(5)(p10).rev ish der(5)t(5;8)(p10;p23.3),+i(5)(p10) (WCP5+,D5S23+)[50]/48,XX,add(5)(p10).rev ish der(5)t(5;8)(p10;p23.3),+i(5)(p10)(WCP5+,D5S23+),+mar.ish 8(p10q10)(D8Z2+,WCP8-)[25]. Therefore, the baby had complete trisomy 5p, with trisomy of the distal part of 8p and of the centromeric region of chromosome 8. The clinical significance of de novo marker chromosomes is a major problem in prenatal counselling. Molecular cytogenetic tools such as FISH and microFISH are indispensable for characterizing markers and determining the breakpoints more precisely in deleted chromosomes.
Subject(s)
Genetic Counseling , Prenatal Diagnosis , Trisomy/diagnosis , Trisomy/genetics , Adult , Amniocentesis , Chromosomes, Human, Pair 5 , Diagnosis, Differential , Esophageal Atresia/diagnostic imaging , Esophageal Atresia/embryology , Fatal Outcome , Female , Humans , In Situ Hybridization, Fluorescence , Infant, Newborn , Polyhydramnios/diagnostic imaging , Pregnancy , Pregnancy Trimester, Third , Ultrasonography, PrenatalABSTRACT
The genetics of A. niger has been developed since 1980. An overview is presented of the advances in developing methods and collecting data. Important tools have been a) the application of essentially different methods to isolate mutants, b) the adaptation to A. niger ofA. nidulans methodology for analysis of the parasexual cycle, c) the choice of marker genes, and in some cases the artificial introduction of such genes, to select homozygous segregants arising from mitotic recombination. With the use of parasexual recombination, a genetic linkage map of A. niger has been established. In total, 110 nuclear and 1 cytoplasmic (mitochondrial) markers are available. The application of A. niger genetics in applied research is illustrated by examples.
Subject(s)
Aspergillus niger/genetics , Chromosome Mapping , Genetic Linkage , Genetic Markers , Karyotyping , Mutation , Selection, GeneticABSTRACT
Somatic cells of males with azoospermia or oligozoospermia (sperm density < 20 million sperm cells/ml) were found to contain increased percentages of chromosomal abnormalities. Subfertile males with a normal somatic karyogram were found to have increased rates of aneuploidy in sperm. This creates risks for the offspring after fertilization with intracytoplasmatic sperm injection (ICSI). Certain gene mutations on the Y chromosome cause severe oligo- or azoospermia and will, in case of successful reproduction with ICSI, be transmitted to male offspring in 100% of the cases. The same holds true, irrespective of sex, of mutations responsible for cystic fibrosis. In non-random groups of ICSI pregnancies, higher proportions of de novo sex-chromosomal abnormalities have been found than expected. In addition, there are increased proportions of paternally inherited structural autosomal anomalies. Extrapolation of the findings is not yet possible, however.
Subject(s)
Chromosome Aberrations , Chromosome Disorders , Fertilization in Vitro/methods , Oligospermia/etiology , Humans , Male , MicroinjectionsABSTRACT
We report on a case of generalized mosaicism for trisomy 22. At chorionic villus sampling (CVS) in the 37th week of pregnancy, a 47,XX,+22 karyotype was detected in all cells. The indication for CVS was severe unexplained symmetrical intrauterine growth retardation (IUGR) and a ventricular septal defect (VSD) was noted. In cultured cells from amniotic fluid taken simultaneously, only two out of ten clones were trisomic. At term, a growth-retarded girl with mild dysmorphic features was born. Lymphocytes showed a normal 46,XX[50] karyotype; both chromosomes 22 were maternal in origin (maternal uniparental disomy). Investigation of the placenta post-delivery using fluorescence in situ hybridization showed a low presence of trisomy 22 cells in only one out of 14 biopsies. In cultured fibroblasts of skin tissue, a mosaic 47,XX,+22[7]/46,XX[25] was observed. Clinical follow-up is given up to 19 months.
Subject(s)
Chromosomes, Human, Pair 22/genetics , Fetal Growth Retardation/genetics , Microsatellite Repeats/genetics , Mosaicism/genetics , Trisomy/genetics , Abnormalities, Multiple/genetics , Adult , Alleles , Biopsy , Chorionic Villi Sampling , Female , Fetal Growth Retardation/complications , Fetal Growth Retardation/physiopathology , Follow-Up Studies , Heart Septal Defects, Ventricular/complications , Heart Septal Defects, Ventricular/embryology , Heart Septal Defects, Ventricular/genetics , Humans , Infant, Newborn , Karyotyping , Parents , Placenta/pathology , Pregnancy , Pregnancy Outcome , Pregnancy Trimester, Third , Prenatal Diagnosis/methods , Trisomy/diagnosisABSTRACT
Single-strand conformational analysis was used to screen for genetic defects in all thirteen exons of the phenylalanine hydroxylase gene (PAH) in phenylketonuria and hyperphenylalaninemia patients in the Netherlands. Exons that showed a bandshift were sequenced directly. In this way, we were able to identify 93% of the PAH mutations in a panel of 34 patients. Twenty-one different mutations were found: 4 of these gene aberrations are novel.
Subject(s)
Mutation , Phenylalanine Hydroxylase/genetics , Phenylketonurias/enzymology , Polymorphism, Single-Stranded Conformational , Amino Acid Metabolism, Inborn Errors/blood , Amino Acid Metabolism, Inborn Errors/enzymology , Amino Acid Metabolism, Inborn Errors/genetics , DNA Mutational Analysis , Exons/genetics , Genetic Heterogeneity , Genotype , Humans , Netherlands , Phenylalanine/blood , Phenylketonurias/blood , Phenylketonurias/geneticsSubject(s)
Prenatal Diagnosis , Turner Syndrome/epidemiology , Diagnostic Errors , Female , Humans , Pregnancy , Prevalence , Turner Syndrome/diagnosisABSTRACT
A genetic transformation system for the aflatoxin-producing fungus Aspergillus parasiticus using two autonomously replicating plasmids from A. nidulans (ARp1 and pDHG25) is reported. Transformation frequencies using the plasmid pDHG25 were from 5 x 10(2) to 2.5 x 10(4) transformants per 10(6) viable protoplasts and microgram DNA. The stability of the plasmids in the transformants was also studied. This transformation system offers a new opportunity to clone genes related to aflatoxin production using appropriate aflatoxin-defective mutants.
Subject(s)
Aspergillus nidulans/genetics , Aspergillus/genetics , DNA Replication , Plasmids , Transformation, Genetic , DNA, Fungal/analysis , Mitosis , Plasmids/genetics , ProtoplastsABSTRACT
We report on a patient with sensorineural deafness, onycho- and osteodystrophy and mental retardation (DOOR syndrome) and review the literature. It appears that abnormal dermatoglyphics are a frequent feature of the DOOR syndrome, as all patients with DOOR syndrome in whom dermatoglyphic investigations were done, had multiple arches on their fingertips.
Subject(s)
Bone Diseases/diagnostic imaging , Deafness , Intellectual Disability/diagnosis , Nail Diseases/diagnosis , Child, Preschool , Dermatoglyphics , Fingers/abnormalities , Humans , Male , Radiography , SyndromeABSTRACT
This paper provides a genetic map of Aspergillus niger. At present 84 markers have been assigned to eight linkage groups. The chromosomal location of 60 markers is presented in this paper. The allocation of markers is based on recombination due to mitotic crossing over. Various methods for selection and analysis of homozygous recombinants were applied, using colour, auxotrophic and resistance markers. In addition, transformants carrying the heterologous Aspergillus nidulans gene coding for acetamidase (amdS) were used for mitotic mapping of markers in several linkage groups. In most of the transformants the amdS insert appeared to be centromere-distal to all known genetic markers, thus extending the genetic map. The linear order of the markers in the eight linkage groups has been determined. On the basis of these and earlier experiments tentative genetic maps for the eight linkage groups are presented. Genetic markers were found on both arms of the chromosomes, except for chromosomes II and IV. The genetic distance between markers and the centromere varies from about 10(-4) (LG I, II, V) up to more than 10(-2) (LG III, VI, VIII). The total frequency of mitotic recombination per genome in this fungus has been estimated to be at least 1.2 x 10(-1).
Subject(s)
Aspergillus niger/genetics , Chromosomes, Fungal , Genetic Linkage , Amidohydrolases/genetics , Aspergillus niger/enzymology , Aspergillus niger/isolation & purification , Chromosome Mapping , Crossing Over, Genetic , Genetic Markers , MitosisABSTRACT
Aspergillus niger mutants defective in arginine or proline biosynthesis have been isolated and 12 genetic loci were identified. Mutation was induced by low doses UV, and mutants were isolated after filtration enrichment. The mutants were classified according to their phenotype in growth tests and were further characterized in complementation tests. The arginine auxotrophic mutants represent nine complementation groups. Three additional complementation groups were found for mutants that could grow on proline (two of them on arginine too). Linkage group analysis was done in somatic diploids obtained from a mutant and a master strain with genetic markers on six chromosomes. The arg genes belong to six different linkage groups and the pro genes to two. One arg-mutant could be complemented by transformation with the A. nidulans argB+ gene, and this A. niger gene thus appeared to be homologous to the A. nidulans argB. We isolated an A. niger strain with the argB gene tightly linked with the nicA1 marker. This strain is very suitable as acceptor for transformation with an argB-plasmid, because transformants with inserts on the homologous site can be recognized and analyzed genetically using the nicA1 marker gene.
Subject(s)
Arginine/genetics , Aspergillus niger/genetics , Mutation/genetics , Proline/genetics , Aspergillus niger/growth & development , Genetic Complementation Test , Genetic Linkage , Genetic Markers/genetics , Transformation, Genetic/geneticsABSTRACT
This study was prompted by the observation that an Aspergillus niger transformant with a multicopy bphA (benzoate-4-hydroxylase gene) insert did not grow on benzoate, whereas a transformant with only one extra copy could grow. Therefore, an extensive survey has been made for other genes involved in the conversion of benzoate into 4-hydroxy-benzoate. A transformant with two copies of the bphA gene was used in part of the mutation experiments in order to avoid the isolation of many bphA mutants. Filtration enrichment was used to isolate mutants defective in the conversion of benzoate. The Bph mutants that have been isolated belong to six complementation groups. Mutants with a defected structural gene (bphA) were again predominantly found but, in addition, five other groups of mutants that could not grow on benzoate were isolated. Genetic analysis of the mutants showed that the six genes were localized in different parts of the genome. This was used as an additional proof that some mutants involved different genes. Diploids with seven copies of the bphA gene and heterozygous for one of the other bph genes were constructed. No indication has been obtained that any one of the mutant classes is responsible for the growth-limiting factor in bphA multicopy transformants. This study shows that the p-hydroxylation of benzoate is very complex, although the metabolic pathway is straight forward.
Subject(s)
Aspergillus niger/genetics , Fungal Proteins/genetics , Genes, Fungal , Mixed Function Oxygenases/genetics , Aspergillus niger/enzymology , Benzoate 4-Monooxygenase , Benzoates/metabolism , Benzoic Acid , Genetic Complementation Test , Genetic Linkage , Recombination, GeneticABSTRACT
An electrophoretic karyotype of Aspergillus niger was obtained using contour-clamped homogeneous electric field (CHEF) gel electrophoresis. Chromosome-sized DNA was separated into four bands. Seven of the eight linkage groups could be correlated with specific chromosomal bands. For this purpose DNA preparations from seven transformant strains of A. niger each carrying the heterologous amdS gene of Aspergillus nidulans on a different chromosome were analysed. Some of the assignments were confirmed with linkage group-specific A. niger probes. The estimated sizes of the A. niger chromosome range from 3.5 to 6.6 Mb, based on gel migration relative to the chromosomes of Schizosaccharomyces pombe strains, Saccharomyces cerevisiae and A. nidulans. The total genome size of A. niger significantly exceeds that of A. nidulans and is estimated to be about 35.5-38.5 Mb. Electrophoretic karyotyping was used to allocate non-mutant rRNA genes and to estimate the number of plasmids integrated in a high copy number transformant.
Subject(s)
Aspergillus niger/genetics , DNA, Fungal/genetics , Genes, Fungal , Chromosome Mapping , Chromosomes, Fungal , DNA, Fungal/isolation & purification , Electrophoresis/methods , Genetic Linkage , KaryotypingABSTRACT
The gene coding for benzoate-para-hydroxylase (bphA) of Aspergillus niger was cloned using differential hybridisation techniques and complementation of mutants deficient in this enzyme activity. The nucleotide sequence of the gene was determined, the presence of two introns was shown and the transcription start and termination sites were determined. The structure of the mRNA upstream from the long open reading frame (ORF) is unusual. It contains two small, overlapping ORFs whose function is unknown. Comparison of the deduced amino acid sequence of the protein with the sequences present in the databanks, indicated a significant similarity of BPH to the superfamily of cytochrome P450 enzymes. Further analysis revealed that this protein is a member of a new P450 gene family designated P450LIII. The gene is designated CYP53. To increase the BPH activity of A. niger, multiple copies of the bphA gene were introduced into the genome of a recipient strain by transformation. Although increased intracellular levels of the BPH protein could be detected, the BPH enzyme activity was decreased, suggesting titration of another essential component.
Subject(s)
Aspergillus niger/genetics , Cytochrome P-450 Enzyme System/genetics , Multigene Family , Oxygenases/genetics , Amino Acid Sequence , Aspergillus niger/enzymology , Base Sequence , Blotting, Western , Cloning, Molecular , Gene Expression Regulation, Fungal , Genes, Fungal , Genetic Complementation Test , Introns , Molecular Sequence Data , Mutation , Oxygenases/metabolism , Restriction Mapping , Sequence Homology, Nucleic Acid , Transcription, GeneticABSTRACT
The Aspergillus nidulans gene coding for acetamidase (amdS) was introduced into A. niger by transformation. Twelve Amd+ transformants were analysed genetically. The amdS inserts were located in seven different linkage groups. In each transformant the plasmid was integrated in only a single chromosome. Our (non-transformed) A. niger strains do not grow on acetamide and are more resistant to fluoroacetamide than the transformants. Diploids hemizygous for the amdS insert have the Amd+ phenotype. We exploited the opportunity for two-way selection in A. niger: transformants can be isolated based on the Amd+ phenotype, whereas counter-selection can be performed using resistance to fluoroacetamide. On this basis we studied the phenotypic stability of the heterologous amdS gene in A. niger transformants as well as in diploids. Furthermore, we mapped the plasmid insert of transformant AT1 to the right arm of chromosome VI between pabA1 and cnxA1, providing evidence for a single transformational insert. The results also show that the amdS transformants of A. niger can be used to localize non-selectable recessive markers and that the method meets the prerequisites for efficient mitotic mapping. We suggest the use of amdS transformants for mitotic gene mapping in other fungi.
Subject(s)
Amidohydrolases/genetics , Aspergillus niger/genetics , Chromosome Mapping , Transformation, Genetic , Amidohydrolases/metabolism , Aspergillus niger/enzymology , Chromosomes, Fungal , Diploidy , Drug Resistance, Microbial , Fluoroacetates/pharmacology , Genes, Fungal , Genetic Linkage , Genotype , Haploidy , Mitosis , Phenotype , Transformation, BacterialABSTRACT
This paper describes the use of chlorate resistant mutants in genetic analysis of Aspergillus niger. The isolated mutants could be divided into three phenotypic classes on the basis of nitrogen utilization. These were designated nia, nir and cnx as for Aspergillus nidulans. All mutations were recessive to their wild-type allele in heterokaryons as well as in heterozygous diploids. The mutations belong to nine different complementation groups. In addition a complex overlapping complementation group was found. Evidence for the existence of eight linkage groups was obtained. Two linked chlorate resistance mutations and two tryptophan auxotrophic markers, which were unlinked to any of the known markers, form linkage group VIII. We used the chlorate resistance mutations as genetic markers for the improvement of the mitotic linkage map of A. niger. We determined the linear order of three markers in linkage group VI as well as the position of the centromere by means of direct selection of homozygous cnxA1 recombinants. In heterozygous diploid cultures diploid chlorate resistant segregants appeared among conidiospores with a frequency of 3.9 x 10(-5) (cnxG13 in linkage group I) to 2.1 x 10(-2) (cnxD6 in linkage group III). The mean frequency of haploid chlorate resistant segregants was 1.3 x 10(-3). The niaD1 and niaD2 mutations were also complemented by transformation with the A. niger niaD+ gene cloned by Unkles et al. (1989). Mitotic stability of ten Nia+ transformants was determined. Two distinct stability classes were found, showing revertant frequencies of 5.0 x 10(-3) and 2.0 x 10(-5) respectively.
Subject(s)
Aspergillus niger/genetics , Chlorates/pharmacology , Genetic Linkage , Mitosis , Mutation , Aspergillus niger/drug effects , Aspergillus niger/growth & development , Drug Resistance, Microbial/genetics , Gene Frequency , Genetic Complementation Test , Genetic Markers/analysis , Recombination, Genetic , Transformation, GeneticABSTRACT
This paper describes a procedure which allows the quantitative selection of auxotrophs of the fungus Aspergillus niger by enzymatic killing of immobilized germinating prototrophic conidiospores. We have applied this procedure to linkage analysis on the basis of mitotic cross-over in this fungus. Starting with a heterozygous diploid strain, we could select auxotrophic homozygous diploid recombinants quantitatively. We estimated the frequency of crossing-over after correction for clonal distribution of recombinants, and localized four auxotrophic markers as well as the centromere on chromosome V of this fungus. The Novozym enrichment procedure proved to be useful in genetic analysis and for the construction of recombinant genotypes in the case of closely linked auxotrophic markers. The determination of gene order and the estimation of distances on the basis of benomyl-induced recombinant haploid segregants may lead to incorrect conclusions. Genetic analysis on the basis of homozygous recombinants, however, can provide reliable estimates of map distances.
Subject(s)
Aspergillus niger/genetics , Chromosomes, Fungal , Crossing Over, Genetic/genetics , Genetic Linkage/genetics , Mitosis , Aspergillus niger/ultrastructure , Chromosome Mapping , Culture Media , Diploidy , Genotype , HaploidyABSTRACT
Mutants of Aspergillus niger requiring adenine and one mutant requiring cytosine were isolated after low-dose mutagenesis and enrichment. In addition we had mutants of two genes involved in the pyrimidine biosynthesis isolated as 5-fluoro-orotic acid-resistant mutants. The fifteen adenine-less mutants could be placed in seven complementation groups. From each group a representative mutant was analyzed in order to determine the linkage group by analysis of the mutants in a heterozygous diploid carrying markers in six linkage groups. AdeF could not be assigned to any one of these linkage groups and proved to be linked to nicB, oliC and cnxC, none of which could be placed in a linkage group. Thus, conclusive evidence was obtained for a seventh linkage group. As pyrA was used as selection marker for transformation, we constructed a pyrA strain with a linked marker which can be used in the genetic analysis of transformations.