Low proportion of whole exon deletions causing phenylketonuria in Denmark and Germany.

Phenylketonuria (PKU) is an autosomal recessive genetic disorder caused by mutations of the gene encoding phenylalanine hydroxylase (PAH). More than 500 different PAH mutations have been identified and about 90% of these are single base mutations. Although the identification rate of the PAH mutations is generally very high, some variants remain unidentified. A fraction of these mutations are the result of genomic deletions or duplications, which are not recognized with standard PCR-based methods. Here we present the results of exon deletion or duplication analysis in a total of 34 families, in which two mutations had not been identified using conventional diagnostic screening techniques. Using multiplex ligation-dependent probe amplification (MLPA), we found a deletion covering exon 1 and exon 2 (c.1-?_168+?del) in one patient, a deletion of exon 3 (c.169-?_352+?del) in four patients, and a deletion of exon 5 (c.442-?_509+?del) in two patients. A deletion was thus detected in about 20% (7/34) of the families tested. Out of a combined cohort of 570 independent PKU patients from Denmark and Germany, exon deletions were identified in a total of four patients. The estimated allelic frequency of exon deletions in PKU patients in these two populations is therefore below 0.5%.

Is zygosity or chorionicity the main determinant of fetal outcome in twin pregnancies?

OBJECTIVE: The purpose of this study was to examine whether fetal outcome in twin pregnancies is dependent on zygosity or chorionicity. STUDY DESIGN: This was a prospective observational study comprised of women with twin pregnancies who attended the fetal medicine unit at St Michael's Hospital, Bristol, Ireland, during the years 1998 to 2000 and who were delivered in hospitals in south west England. After delivery, zygosity was determined with umbilical cord blood with the use of microsatellite markers that were amplified by polymerase chain reaction. Placentae were examined histologically for chorionic type. The perinatal outcomes of 3 groups of monozygotic monochorionic, monozygotic dichorionic, and dizygotic pregnancies were compared with the use of the Mann-Whitney U test and the Fisher's exact test. RESULTS: All 92 dizygotic and 15 monozygotic dichorionic pregnancies resulted in live births. In 7 of the 39 cases in the monozygotic monochorionic group, either both twins were not live born or delivery occurred <24 weeks of gestation. The gestational age at delivery and birth weight were significantly lower, and there were a greater number of cases with birth weight discordancy of >25% in the monochorionic pregnancies compared with the other 2 groups (P < .05). There were no significant differences in any of the study parameters between the monozygotic dichorionic and dizygotic groups. CONCLUSION: Fetal outcome in twin pregnancies is related to chorionicity rather than zygosity.

Comparative multiplex dosage analysis detects whole exon deletions at the phenylalanine hydroxylase locus.

We have developed quantitative comparative multiplex dosage analysis to detect altered copy number of regions of the phenylalanine hydroxylase gene. Out of 41 alleles (4% of 1,010 PKU chromosomes) on which a mutation had not been characterized previously, this technique has highlighted two novel mutations: deletions of exon 5 and of exon 6 on a total of eight alleles. Restriction-enzyme digestion of genomic DNA and hybridization to an amplified segment of the phenylalanine hydroxylase (PAH) cDNA probe PAH247 established the size of the deletion in five individuals to be between 700 and 900 bases. We also report somatic mosaicism in the parent of an affected child previously shown to have a deletion spanning exons 5 and 6. Finally, we report a putative duplication of a region encompassing exon 6 in an affected individual.

Genetic diversity within the R408W phenylketonuria mutation lineages in Europe.

The R408W phenylketonuria mutation in Europe has arisen by recurrent mutation in the human phenylalanine hydroxylase (PAH) locus and is associated with two major PAH haplotypes. R408W-2.3 exhibits a west-to-east cline of relative frequency reaching its maximum in the Balto-Slavic region, while R408W-1.8 exhibits an east-to-west cline peaking in Connacht, the most westerly province of Ireland. Spatial autocorrelation analysis has demonstrated that the R408W-2.3 cline, like that of R408W-1.8, is consistent with a pattern likely to have been established by human dispersal. Genetic diversity within wild-type and R408W chromosomes in Europe was assessed through variable number tandem repeat (VNTR) nucleotide sequence variation and tetranucleotide short tandem repeat (STR) allelic associations. Wild-type VNTR-8 chromosomes exhibited two major cassette sequence organizations: (a1)5-b3-b2-c1 and (a1)5-b5-b2-c1. R408W-1.8 was predominantly associated with (a1)5-B5-B2-C1. Both wild-type vntr-3 and r408w-2.3 chromosomes exhibited a single invariant cassette sequence organization, a2-b2-c1. STR allele distributions associated with the cassette variants were consistent with greater diversity in the wild-type VNTR-8 lineage and were suggestive of different levels of diversity between R408W-1.8 and R408W-2.3. The finding of greater genetic diversity within the wild-type VNTR-8 lineage compared to VNTR-3 suggests that VNTR-8 may be older within the European population. However, in the absence of a more extensive STR data-set, no such conclusions are possible for the respective R408W mutant lineages.