Determining the genome-wide kinship coefficient seems unhelpful in distinguishing consanguineous couples with a high versus low risk for adverse reproductive outcome.

BACKGROUND: Offspring of consanguineous couples are at increased risk of congenital disorders. The risk increases as parents are more closely related. Individuals that have the same degree of relatedness according to their pedigree, show variable genomic kinship coefficients. To investigate whether we can differentiate between couples with high- and low risk for offspring with congenital disorders, we have compared the genomic kinship coefficient of consanguineous parents with a child affected with an autosomal recessive disorder with that of consanguineous parents with only healthy children, corrected for the degree of pedigree relatedness. METHODS: 151 consanguineous couples (73 cases and 78 controls) from 10 different ethnic backgrounds were genotyped on the Affymetrix platform and passed quality control checks. After pruning SNPs in linkage disequilibrium, 57,358 SNPs remained. Kinship coefficients were calculated using three different toolsets: PLINK, King and IBDelphi, yielding five different estimates (IBDelphi, PLINK (all), PLINK (by population), King robust (all) and King homo (by population)). We performed a one-sided Mann Whitney test to investigate whether the median relative difference regarding observed and expected kinship coefficients is bigger for cases than for controls. Furthermore, we fitted a mixed effects linear model to correct for a possible population effect. RESULTS: Although the estimated degrees of genomic relatedness with the different toolsets show substantial variability, correlation measures between the different estimators demonstrated moderate to strong correlations. Controls have higher point estimates for genomic kinship coefficients. The one-sided Mann Whitney test did not show any evidence for a higher median relative difference for cases compared to controls. Neither did the regression analysis exhibit a positive association between case-control status and genomic kinship coefficient. CONCLUSIONS: In this case-control setting, in which we compared consanguineous couples corrected for degree of pedigree relatedness, a higher degree of genomic relatedness was not significantly associated with a higher likelihood of having an affected child. Further translational research should focus on which parts of the genome and which pathogenic mutations couples are sharing. Looking at relatedness coefficients by determining genome-wide SNPs does not seem to be an effective measure for prospective risk assessment in consanguineous parents.

Neonatal screening for medium-chain acyl-CoA dehydrogenase (MCAD) deficiency in The Netherlands: the importance of enzyme analysis to ascertain true MCAD deficiency.

The outcome was determined of population-wide neonatal screening for medium-chain acyl-CoA dehydrogenase (MCAD) deficiency using tandem mass spectrometry (MS/MS) in The Netherlands, between October 2003 and September 2005. Prospective population-wide neonatal screening for MCAD deficiency was performed in the northern part of The Netherlands. In newborns with blood octanoylcarnitine (C(8:0)) concentrations > or =0.3 micromol/L, clinical and laboratory follow-up was initiated, including MCAD enzymatic measurements which played a decisive role. In a 2-year period, 66 216 newborns were investigated for MCAD deficiency and follow-up was initiated in 28 newborns. True-positives (n = 14) were identified based upon MCAD enzyme activity <50%, measured with hexanoyl-CoA as substrate. The observed prevalence of MCAD deficiency was 1/6600 (95% CI: 1/4100-1/17 400). In addition to an elevated C(8:0) concentration, a C(8:0)/C(10:0) molar ratio >5.0 turned out to differentiate between false-positives and true-positives. Measurement of MCAD activity using phenylpropionyl-CoA as a substrate further discriminated between newborns with MCAD deficiency and so-called mild MCAD deficiency. To summarize, neonatal screening for MCAD deficiency in the northern part of The Netherlands resulted in the predicted number of affected newborns. Measurement of MCAD activity in leukocytes or lymphocytes using phenylpropionyl-CoA as a substrate can be regarded as the gold standard to diagnose MCAD deficiency upon initial positive screening test results.

[Rapid prenatal diagnosis of chromosomal abnormalities; limitations and possibilities]. / Snelle prenatale diagnostiek van chromosomale afwijkingen; beperkingen en mogelijkheden.

In the past, prenatal diagnosis of chromosomal abnormalities has been achieved using a microscope to make a visual assessment of the chromosomes in foetal cells which had been obtained by invasive procedures. The results were usually not available until 10-14 days later. Now molecular techniques have become available which provide a result within 24-48 hours. The first generation of these techniques combines speed with a focus on a limited number of frequent and important chromosomal abnormalities. Examples include fluorescence in-situ hybridisation (FISH), quantitative polymerase chain reaction (PCR) and multiple ligation-dependent probe amplification (MLPA). One drawback, therefore, is that other clinically significant chromosomal abnormalities will not be detected. More recently other new techniques have been making their appearance, such as array-comparative genomic hybridisation (CGH), which will allow the detection of clinically significant submicroscopic aberrations.

Possibilities and barriers in the implementation of a preconceptional screening programme for cystic fibrosis carriers: a focus group study.

OBJECTIVE: This qualitative study aimed to explore possibilities and barriers in the implementation of a nationwide preconceptional cystic fibrosis (CF) carrier screening programme. METHODS: Sessions were held with two focus groups of CF patients and CF relatives, one focus group of people from the target population (couples planning a pregnancy), and two focus groups of potential providers (general practitioners (GPs) and municipal health service workers). RESULTS: Important barriers in the implementation of a preconceptional CF carrier screening programme included the problem of reaching the target population, the heavy workload of GPs, the limited public knowledge about CF in general, and the absence of a preconceptional consultation setting. In general, there was a positive attitude among the participants towards CF carrier screening. CONCLUSION: This study revealed some important barriers in the implementation of CF carrier screening programmes. More research is needed to specify and quantify the importance of the various barriers. Eventually, different intervention strategies should be included in an implementation plan to overcome the most important barriers in the organization and execution of screening.

Preconception cystic fibrosis carrier couple screening: impact, understanding, and satisfaction.

The impact, understanding of test-results, and satisfaction among participating couples in a preconception cystic fibrosis (CF) carrier screening project were assessed 6 months after testing. Questionnaire data were obtained from 17/18 identified carriers, 15 partners of carriers with negative test results, and 794 (73%) other participants. None of the carriers changed their reproductive plans because of their test results. Eight participants were worried about their results, including four carriers. Those who attended a general practitioner (GP) consultation for pretest education were less worried than those who attended an educational session. Seven carriers felt less healthy. Predictors of a correct understanding of test results (correct in 62% of participants) were: positive test results, high level of knowledge of CF, high level of education, attending an educational session, and previously heard of CF. All participants who reported that they were worried, all carriers, and 95% of the other participants said that they would make the same decision to be tested again. Although couples who were educated during a GP consultation were less worried, the results of the study suggest that understanding is more correct in couples attending an educational session. The results further suggest that since satisfaction with the screening was high, worries and feeling less healthy due to the test results are probably not a great burden.

Survival analysis in familial ovarian cancer, a case control study.

OBJECTIVE: familial ovarian cancer patients have been found to differ from sporadic cases, clinically as well as in the molecular make-up of the tumour. Here, a case control study is performed to analyse potential differences in survival. STUDY DESIGN: 31 families with a strong history of ovarian and/or breast cancer presenting to a family cancer clinic 44 ovarian cancer patients were included. Each patient was matched for age and stage with controls from a cancer registry. Survival rates and the effect of several prognostic factors were analysed. RESULTS: median survival in the study group differed significantly from controls. A survival benefit for familial cases was maintained up to 5 years after diagnosis. Long-term survival was equally poor in both groups. CONCLUSION: the difference in survival between familial ovarian cancer cases and matched controls may reflect differences in biological behaviour. This may have important implications for the management and prevention of familial ovarian cancer.

Polymorphisms in the tumor necrosis factor and lymphotoxin-alpha gene region and preeclampsia.

OBJECTIVE: To investigate potential association or linkage among nine polymorphisms in the genes encoding tumor necrosis factor (TNF) alpha or lymphotoxin (LT) alpha and preeclampsia. METHODS: Four di-allelic polymorphisms and five microsatellite markers in the genes encoding TNF-alpha (TNF) and LTalpha (LTA) and their haplotypes were studied in 150 Dutch families. These families contained sib-pairs of women affected with preeclampsia; eclampsia; the hemolysis, elevated liver enzymes, low platelets (HELLP) syndrome (strict criteria); or pregnancy-induced hypertension (mild criteria). Frequencies were compared with 98 healthy controls. Nonparametric affected sib-pair analyses for allele sharing among siblings were carried out for all nine markers. Each sibship was composed of an affected index woman and one or more affected sisters. RESULTS: Although we found a striking association with the TNF-I haplotype in 30 index women with (pre-)eclampsia or HELLP syndrome compared with controls (odds ratio [OR] 3.8; 95% confidence interval [CI] 1.6, 8.9), this association was not found in their 30 sisters meeting similar disease criteria. Analyses in all 150 families showed a similar TNF-I association in 122 index women meeting the strict criteria compared with controls (OR 1.9; 95% CI 1.1, 3.3), but, again, not in their 91 sisters meeting similar disease criteria. This association was stronger in a subgroup of 75 index women with preeclampsia only (OR 2.3; 95% CI 1.2, 4.2). No excess allele sharing for any marker was seen between the siblings. CONCLUSION: The nine polymorphisms studied in the TNF-LTA region did not show evidence for association or linkage with familial preeclampsia.

Mutations in the gene for methylenetetrahydrofolate reductase, homocysteine levels, and vitamin status in women with a history of preeclampsia.

OBJECTIVE: This study was undertaken to assess frequencies of the methylenetetrahydrofolate reductase gene mutations cytosine-to-thymine substitution at base 677 (C677T) and adenine-to-cytosine substitution at base 1298 (A1298C) and their interactions with homocysteine and vitamin levels among Dutch women with preeclampsia. STUDY DESIGN: Mutations were studied in the following 5 groups: 47 consecutive women with preeclampsia, 49 women with preeclampsia and with hyperhomocysteinemia, 36 women with preeclampsia but without hyperhomocysteinemia, 127 women with familial preeclampsia (typed for C677T mutations only), and 120 control subjects. Plasma levels of homocysteine, folate, and vitamin B12 were measured. RESULTS: Although 10.6% of the consecutive women with preeclampsia had strictly defined hyperhomocysteinemia (values >97.5th percentile), neither mutation was found in excess relative to the control group. Women with preeclampsia who had mild hyperhomocysteinemia (values >75th percentile) had a significant excess of the TT genotype (homozygosity for C677T mutation) relative to the women with preeclampsia who did not have hyperhomocysteinemia (odds ratio, 8.2; 95% confidence interval, 1.8-39). They also had significantly lower vitamin levels. CONCLUSION: Hyperhomocysteinemia in women with preeclampsia was associated with mutations in the gene for methylenetetrahydrofolate reductase, but the high frequency of hyperhomocysteinemia itself cannot be explained by these mutations alone.

Attitudes towards reproductive issues and carrier testing among adult patients and parents of children with cystic fibrosis (CF).

Assessing the reproductive choices of parents of children with cystic fibrosis (CF) is important in getting a greater insight into the possible needs for counselling of carrier couples in the reproductive decision-making process. Also, parents' reproductive attitudes might indicate critical issues for discussion with regard to introducing general population screening programmes. Data were available from two groups of subjects: 287 adult patients with CF (mean age 27.5 years) and 288 parents of a child with CF (mean age of child 7.7 years) who participated in a Dutch national survey. Attitudes towards reproductive issues and carrier screening and parents' reproductive behaviour were analysed in terms of responses to questionnaires. To avoid having another child with CF, most parents decided against further pregnancies or used other reproductive options. Prenatal diagnosis was used by 72% of parents, whereas 76% of parents planning more children intended using this option. Intention to use prenatal diagnosis was associated with the strength of religious conviction, and was not associated with perceived severity of CF, health of the child, or future therapeutic possibilities. Respondents demonstrated difficulties in deciding to abort for CF. A majority of parents and adult patients supported carrier couple identification within CF families. The results suggest that those most closely involved with CF will accept the reproductive choices of carrier couples identified in population screening programmes.

A genome-wide scan for preeclampsia in the Netherlands.

Preeclampsia, hallmarked by de novo hypertension and proteinuria in pregnancy, has a familial tendency. Recently, a large Icelandic genome-wide scan provided evidence for a maternal susceptibility locus for preeclampsia on chromosome 2p13 which was confirmed by a genome scan from Australia and New Zealand (NZ). The current study reports on a genome-wide scan of Dutch affected sib-pair families. In total 67 Dutch affected sib-pair families, comprising at least two siblings with proteinuric preeclampsia, eclampsia or HELLP-syndrome, were typed for 293 polymorphic markers throughout the genome and linkage analysis was performed. The highest allele sharing lod score of 1.99 was seen on chromosome 12q at 109.5 cM. Two peaks overlapped in the same regions between the Dutch and Icelandic genome-wide scan at chromosome 3p and chromosome 15q. No overlap was seen on 2p. Re-analysis in 38 families without HELLP-syndrome (preeclampsia families) and 34 families with at least one sibling with HELLP syndrome (HELLP families), revealed two peaks with suggestive evidence for linkage in the non-HELLP families on chromosome 10q (lod score 2.38, D10S1432, 93.9 cM) and 22q (lod score 2.41, D22S685, 32.4 cM). The peak on 12q appeared to be associated with HELLP syndrome; it increased to a lod score of 2.1 in the HELLP families and almost disappeared in the preeclampsia families. A nominal peak on chromosome 11 in the preeclampsia families showed overlap with the second highest peak in the Australian/NZ study. Results from our Dutch genome-wide scan indicate that HELLP syndrome might have a different genetic background than preeclampsia.

Efficacy of cascade testing for fragile X syndrome.

OBJECTIVE: Fragile X syndrome is the most common cause of mental retardation from a single gene defect, transmitted in an X-linked semidominant fashion. Cloning of the gene responsible for fragile X syndrome has made it possible to identify carriers who are at risk of giving birth to a child with fragile X syndrome. One of the proposed strategies for identifying carriers is cascade testing, in which relatives of a patient with fragile X syndrome (the index case) are tested. Because the effectiveness of this type of testing is unknown, the objective of this study was to develop a simulation model for studying the consequences of cascade testing for fragile X syndrome. METHODS: With this model, 100,000 five-generation pedigrees were simulated to assess the maximum number of carriers that would be detected for three scenarios: (a) only first degree relatives of the index case are tested; (b) relatives up to the third degree are tested; (c) relatives up to the fifth degree are tested. RESULTS: In the start-up phase of the testing programme, 18% of couples who will have a fragile X syndrome child are detected. After this phase the (stabilised) cascade testing programme detects 7% of undetected couples who would have a fragile X syndrome child if only first degree relatives were tested, 12% if first to third degree relatives were tested, and 15% if first to fifth degree relatives were tested. To detect 90% of all premutation and full mutation carriers at least eight consecutive generations need to be tested. CONCLUSIONS: The results of our analysis show that cascade testing is not very effective in detecting carriers.

Molecular analysis of SALL1 mutations in Townes-Brocks syndrome.

Townes-Brocks syndrome (TBS) is an autosomal dominantly inherited malformation syndrome characterized by anal, renal, limb, and ear anomalies. Recently, we showed that mutations in the putative zinc finger transcription factor gene SALL1 cause TBS. To determine the spectrum of SALL1 mutations and to investigate the genotype-phenotype correlations in TBS, we examined 23 additional families with TBS or similar phenotypes for SALL1 mutations. In 9 of these families mutations were identified. None of the mutations has previously been described. Two of these mutations are nonsense mutations, one of which occurred in three unrelated families. Five of the mutations are short deletions. All of the mutations are located 5' of the first double zinc finger (DZF) encoding region and are therefore predicted to result in putative prematurely terminated proteins lacking all DZF domains. This suggests that only SALL1 mutations that remove the DZF domains result in TBS. We also present evidence that in rare cases SALL1 mutations can lead to phenotypes similar to Goldenhar syndrome. However, phenotypic differences in TBS do not seem to depend on the site of mutation.

Preconceptional screening of couples for carriers of cystic fibrosis: a prospective evaluation of effects, costs and savings for different mutation detection methods.

OBJECTIVE: The aim of this study was to perform a prospective evaluation of the effects, costs and savings of a preconceptional screening programme of couples for carriers of cystic fibrosis (CF). METHODS: A decision model for both single-entry two-step (SETS) and double-entry two-step (DETS) couple screening was constructed. Two mutation detection methods were considered: allele-specific oligonucleotide (ASO) hybridisation screening of 32 mutations, with a sensitivity of 90%, and denaturing gradient gel electrophoresis (DGGE), with a sensitivity of 98%. In our model, the following combinations were used: (1) ASO for both steps; (2) DGGE for both steps, and (3) ASO for the first step and DGGE for the second step. The model is demonstrated using figures from the Netherlands, where there is a carrier frequency of 1:30. We estimated the value of different choices and probabilities in a decision tree and determined the costs of screening for CF and the costs of the illness itself. RESULTS: We found that with most of the combinations of mutation detection methods, SETS couple screening could offer positive net savings in the Netherlands. The ASO/DGGE combination resulted in the highest net savings. DETS couple screening for all combinations, including testing with DGGE in both steps, did not show a positive cost-savings balance at all, not even with an uptake rate of 100%. The maximum number of carrier couples identified when screening 100,000 couples with the ASO/DGGE combination was 98 (SETS). This could result in about 25 fewer children born with CF each year in the Netherlands, under the following assumptions: (1) each couple has two children and 10% of couples are unable to have children; (2) of detected carrier couples, 15% decide not to have children and 85% make use of prenatal diagnosis; (3) of those fetuses diagnosed with CF, 80% are aborted, and (4) prenatal diagnosis carries a 0.75% risk of iatrogenic abortion. CONCLUSIONS: The results of this evaluation show that there are no financial objections to the preconceptional screening of couples in the Netherlands when the above-mentioned assumptions apply; thus, further evaluation can concentrate on the balance of the non-economic consequences of screening for participants and for society.