Prenatal diagnosis in known fragile X carriers

Abstract

Prenatal diagnosis for fragile X syndrome was performed in 34 pregnancies of 33 known carriers, on 22 chorionic villus samples (CVS), and 15 amniocentesis samples. Fetal and maternal DNA were analyzed by the EagI/EcoRI Southern blot of Rousseau et al. [1991: N Engl J Med 325:1673-1681], with detection of full mutations ensured by a second loading with brief electrophoresis. As a supplemental assay for full mutations, cytogenetic induction was performed in 20 cases. Positive cytogenetic results were helpful in confirming full mutations in CVS cases where the fetal DNA was intermediate in appearance, between a large premutation and a small full mutation. Of 8 mothers with full mutations, the fetal results were 5 full, 2 normal, and 1 premutation (whose mother was a full/pre compound heterozygote). Of 26 mothers with premutations, the fetal results were 5 full, 13 normal, 7 premutation, and 1 uninterpretable (maternal contamination). Maternal premutations were sized in kb by Southern blot and in CGG repeat number by PCR; the predicted correlation between maternal length and penetrance was seen. Follow-up studies include 3 full mutations and 2 premutations confirmed by DNA analysis at birth. Maternal contamination of CVS samples was encountered in 3 of 22 cases, illustrating the value of EagI in detecting maternal (lyonized) chromosomes.

ACMG Standards and Guidelines for fragile X testing: a revision to the disease-specific supplements to the Standards and Guidelines for Clinical Genetics Laboratories of the American College of Medical Genetics and Genomics

“…In addition, they found that the premutation carriers were well distributed among all the Jewish ethnic groups, in contrast to a previous study. 39 An Italian study of ~2,000 mothers and their newborns in the general population found a premutation carrier frequency of 1 in 109 females and 1 in 225 newborn males (56)(57)(58)(59)(60)(61)(62)(63)(64)(65)(66)(67)(68)(69)(70). 48 A fragile X screen of 10,000 newborn males in Taiwan showed a prevalence of 1 in 1,674.…”

“…However, because it is unwarranted to use methylation status or X-inactivation for phenotypic prediction of a full mutation, the possible hypomethylation of this tissue is no disadvantage, provided that the tissue-specific basis of the hypomethylation is understood. 56,57 It is an acceptable option to omit methylation analysis entirely when testing CVS specimens. In the minor fraction of CVS cases with a result that is ambiguous between a large premutation and a small full mutation by size criteria alone, a follow-up amniocentesis may be required; • The degree of somatic variation in a full mutation "smear" has a wider range of possibilities than is typically seen in blood specimens, from very limited to extraordinarily diffuse; • Mosaicism between trophoblasts and somatic cells is theoretically possible.…”

A simple multiplex FRAXA, FRAXE, and FRAXF PCR assay convenient for wide screening programs

“…As the PCR methylation test is rapid and simple and requires small amounts of the patients DNA sample, we used it to perform prenatal diagnoses on DNA extracted from amniocytes or fetal blood. In chorionic villi, the methylation status is generally not completely established [Maddalena et al, 1994], and the use of chorionic villi samples for methylation analysis is still to be studied.…”

Misconceptions about mosaicism

“…The remainder of the culture was studied with fluorescence in situ hybridization (FISH). Analysis using alpha satellite X probe revealed approxi- Cytogenetic analysis of cultured chorionic villi often reveals tetraploid cells and maternal contamination is seen in 0·03-13 per cent of cultures (Ledbetter et al, 1992;Maddalena and Hicks, 1994). Therefore subsequent use of cultured cells for FISH is inappropriate.…”