Diana Curtis

Specific diagnosis of medium-chain acyl-CoA dehydrogenase (MCAD) deficiency in dried blood spots by a polymerase chain reaction (PCR) assay detecting a point-mutation (G985) in the MCAD gene

The discovery of a point-mutation, adenine-to-guanine, at position 985 in the gene coding for MCAD (G985), gave the basis for an easy and specific polymerase chain reaction test. We tested the specificity of such a PCR based assay and detected correctly G985 and A985 in sequence verified cDNA clones. We showed that the G985 mutation is present in genomic DNA from 48 of 50 patients with confirmed MCAD deficiency, originating from various European countries, Australia and the USA. On the basis of this high frequency of the G985 mutation among patients, we improved and optimized the assay with respect to reliability and convenience for routine diagnostic and screening purposes. As little as 2 microliters blood from filter-paper blood-spots (Guthrie spots) is sufficient for the test.

Frequency of the G985 MCAD mutation in the general population

1 Holton JB, Allen JT, Green CA, Partington S, Gilbert RE, Berry PJ. Inherited metabolic diseases in the sudden infant death syndrome. Arch Dis Child 1991;66:1315-7. 2 Blakemore AIF, Singleton H, Pollitt RJ, et al. Frequency of the G985 MCAD mutation in the general population. Lancet 1991 ;337:298-9. 3 Matsubara Y, Narisawa K, Tada K, et al. Prevalence of K329E mutation in mediumchain acyl-CoA dehydrogenase gene determined from Guthrie cards. Lancet 1991 ;338:552-3.

Medium-Chain Acyl-CoA Dehydrogenase (MCAD) Deficiency: The Prevalent Mutation G985 (K304E) Is Subject to a Strong Founder Effect from Northwestern Europe

Medium-chain acyl CoA dehydrogenase (MCAD) deficiency is a potentially fatal inherited defect of fatty acid beta-oxidation. Approximately 90% of the disease-causing alleles in diagnosed patients are due to a single base mutation, an A (adenine) to G (guanine) transition at position 985 of MCAD cDNA (G985). In a limited number of cases it was found that this mutation was always associated with a particular haplotype, defined by three intragenic restriction fragment length polymorphisms, indicating a founder effect [Kølvraa et al.; Hum Genet 1991; 87: 425-429]. In addition, recent studies of American patients and their ancestors suggested the existence of a founder from northern Europe [Yokota et al.; Am J Hum Genet 1991; 49: 1280-1291]. In the present study we document (1) that the G985 heterozygous frequency in the Caucasian population of North Carolina in the USA is 1/84, which is 5- to 10-fold higher than in non-Caucasian Americans; (2) that there exists a 100% association of the G985 mutation in 17 families with MCAD-deficient patients to a certain haplotype, defined by the restriction endonucleases BanII, PstI and TaqI; (3) that MCAD deficiency due to the G985 mutation is more frequent in the Netherlands, Ireland, England, Belgium and Denmark than in other western European countries, and (4) that the frequency distribution of G985 mutation carriers is 1/68-1/101 in newborns in the United Kingdom and Denmark, and 1/333 in Italy. These results support the notion of a founder effect in northwestern Europe.

Acrocentric associations in mongol populations

SummaryAssociation patterns of acrocentric chromosomes in 2100 metaphases from sample populations of 30 chromosomally normal individuals, 20 mongols and 20 parents of mongol children have been studied. In this study the data are concerned with the overall numbers of acrocentric chromosomes present in associations and not with the numbers of associations or with the ways in which they may be arranged. The individual pairs of acrocentric chromosomes were identified by Leishman staining following trypsin digestion.Acrocentric chromosomes showed a random participation in associations both in normal and in mongol populations, where the additional 21 chromosome significantly alters expected frequencies. In the parents of mongols populations, however, significant deviations from the model of random participation in associations were documented.ZusammenfassungSatellitenassoziationsmuster akrozentrischer Chromosomen wurden in 2100 Metaphasen aus Stichproben von 30 chromosomal normalen Individuen, 20 Fällen mit Down-Syndrom und 20 Eltern mongoloider Kinder untersucht. In dieser Studie wird die Gesamtzahl akrozentrischer Chromosomen, die an Assoziationen beteiligt sind, untersucht; nicht dagegen die Zahl der Assoziationen oder die genaue Art, in der sie vorkommen. Die einzelnen Paare assoziierter Chromosomen wurden durch Leishman-Färbung mit nachfolgender Trypsinisierung identifiziert.Sowohl bei Normalen als auch bei Patienten mit Down-Syndrom waren die akrozentrischen Chromosomen zufällig an den Assoziationen beteiligt; dabei sind die veränderten Erwartungswerte beim Down-Syndrom berücksichtigt. Bei den Eltern der Patienten dagegen fanden sich signifikante Abweichungen von der zufälligen Beteiligung der Chromosomen.

Hearing Impairment and Pigmentary Disturbancea

Hearing impairment is a variable manifestation of several heritable conditions in which pigmentation of the skin or eyes is abnormal. Some of these disorders are well recognized although uncommon, while others are virtually private syndromes. Practical issues concerning the major conditions of this type are reviewed in this article on a basis of a survey of 4452 profoundly deaf children attending special schools in Southern Africa, together with investigations in affected families. The Waardenburg syndrome (WS), which is the most common deafness-depigmentation disorder, was present in 121 (2.7%) of the 4452 deaf scholars. Further studies in 7 multigeneration affected families confirmed phenotypic variability and indicated a need for internationally agreed diagnostic criteria. In 4 Cape Town families of mixed ancestry the WS-I gene was linked to the 2q37 locus, but in another large kindred no linkage could be demonstrated. Nonallelic heterogeneity is possible. There is uncertainty concerning possible interrelationship between WS and piebaldism. The phenotypic consistency of a South African family in which 7 persons in 3 generations had gross piebaldism in the absence of disturbance of hearing or involvement of the eyes and periorbital structures is suggestive that this disorder and WS are separate entities. Molecular investigations indicate that the gene for piebaldism in this kindred is not situated at the WS-I locus 2q37. Deafness and hyperpigmentation are present in neurofibromatosis type II (acoustic neuromata) and the multiple lentigines syndrome, while retinal pigmentation is a feature of the Usher syndrome. This latter entity is apparently much less common in Southern Africa than in other parts of the world.

The most common mutation causing medium-chain acyl-CoA dehydrogenase deficiency is strongly associated with a particular haplotype in the region of the gene

SummaryRFLP haplotypes in the region containing the medium-chain acyl-CoA dehydrogenase (MCAD) gene on chromosome 1 have been determined in patients with MCAD deficiency. The RFLPs were detected after digestion of patient DNA with the enzymes BanII, PstI and TaqI and with an MCAD cDNA-clone as a probe. Of 32 disease-causing alleles studied, 31 possesed the previously publised A→G point-mutation at position 985 of the cDNA. This mutation has been shown to result in inactivity of the MCAD enzyme. In at least 30 of the 31 alleles carrying this G985 mutation a specific RFLP haplotype was present. In contrast, the same haplotype was present in only 23% of normal alleles (P≤3.4×10-18). These findings are consistent with the existence of a pronounced founder effect, possibly combined with biological and/or sampling selection.

Grebe chondrodysplasia and brachydactyly in a family

A family is reported in which various skeletal abnormalities have been segregating over three generations. The Great‐grandfather (II) of the consultand had features consistent with Grebe chondrodysplasia. The other members of the family have brachydactyly, radiologically characterised by short first metacarpals and short middle phalanges of the index and little fingers. The possibility of association of familial brachydactyly and Grebe chondrodysplasia is discussed. An attempt has been made to deal with the genetic counselling problem in this particular family.

X-linked Ehlers-Danlos syndrome type V; the next generation

Two English families with the X‐linked form (Type V) of the Ehlers‐Danlos syndrome (EDS) who were investigated almost 20 years ago have been re‐studied. In one family, the potentially heterozygous sister of 3 affected brothers had born two sons, of whom one has EDS. The 3 brothers had all procreated, producing a total of 2 sons and 3 daughters, all of whom are clinically normal. These pedigree data provide further evidence to support the syndromic identity and X‐linked mode of inheritance of this form of the EDS.

Piebaldism: an autonomous autosomal dominant entity

Piebaldism is a disorder in which the major clinical features are patchy hypopigmentation of the skin and a white forelock. The manifestations of piebaldism overlap with those of other genodermatoses, in particular the Waardenburg syndrome, and it is uncertain whether piebaldism is a distinct entity. We have documented a family in which seven affected members in three generations have gross piebaldism without any additional stigmata. The intrafamilial phenotypic consistency is suggestive that this autosomal dominant disorder has independent syndromic status. Linkage studies using conventional gene markers failed to identity the locus of the faulty gene.

Chromosome banding: specification of structural features of dyes giving rise to G-banding

SummaryMetaphase chromosomes were stained in a routine G-banding procedure with 39 basic dyes of varied structures substituted for the Giemsa stain. Staining outcomes were categorized as: iverstained, differentially stained, trivially or unstained. Certain structural features of the dyes were described numerically, namely, largest conjugated fragment (LCF), conjugated bond number (CBN) and cationic weight. The staining, outcomes were compared to these numerical structural parameters, and structure-staining correlations sought. Dyes with large conjugated systems (and high LCF values) were seen to be overstained; dyes with low LCF values were often non-staining. At intermediate LCF values, the more hydrophobic dyes (with high Hansch π values) stained differentially; the more hydrophilic dyes failed to stain. Expressed numerically, 89% of the dyes with the following characteristics stained differentially: 30≥LCF≥10; Hansch π>−5.0. It was concluded that contributions to dye-chromosome affinity included coulombic forces and van der Waals attractions and that the selectivity of G-banding was largely due to hydrophobic bonding. Induction of bands could be due to the loss of hydrophilic, histones, amplifying underlying variations in the hydrophobic-hydrophilic character of the chromosome structure. Relatively hydrophobic sites include AT-rich DNA and disulphide-rich proteins.The effects on Romanowsky G-banding of chemically modifying chromosomes were in keeping with this model. Overstaining resulted from formation of either hydrophobic or conjugated derivatives or both, whereas trivial or non-staining arose from the formation of hydrophilic derivatives. Intriguingly, the efficacy of the dyes used for Q-banding also correlated positively with their hydrophobic character.