Euchromatic 9q13-q21 duplication variants are tandem segmental amplifications of sequence reciprocal to 9q13-q21 deletions.

BACKGROUND: There are four known pericentromeric euchromatic variants of chromosome 9 in the literature that are increasingly being observed in diagnostic cytogenetic laboratories. These variants pose diagnostic and counselling dilemmas, especially in prenatal settings, as distinction of a pathogenic alteration from a euchromatic variant is difficult. The molecular characterisation of three of these four variants has been reported. In this study, the genomic structure of the fourth variant, an additional G-positive band at 9q13-q21, is characterised. METHODS: Two unrelated families with the 9q13-q21 duplication variant, and a third individual with a cytogenetically visible 9q13-q21 deletion, were studied using conventional and molecular cytogenetics techniques, as well as microarrays. The highly repetitive nature of the segmental duplications in the region also necessitated the use of both interphase and metaphase fluorescence in situ hybridisation (FISH). RESULTS: It was determined that the DNA that constitutes this variant was ∼ 15-20 megabases in size and tandemly repeated as 3-4 cassettes of intrachromosomal segmental duplication. The variant appeared constitutively similar in sequence content and organisation between the two unrelated individuals, and it was inherited without apparent change. Sequences found amplified in the two duplication carriers were absent in the carrier of the deletion variant. CONCLUSIONS: The sequences involved in both the 9q13-q21 duplication and deletion appear the same, implying reciprocity and suggesting non-allelic homologous recombination as the underlying mechanism. All four known euchromatic variants of chromosome 9 have now been shown to encompass segmental duplications. Importantly, a set of validated FISH probes was defined for the detection and characterisation of this 9q13-q21 amplification in the context of other chromosome 9 variants, allowing apparently benign variants to be distinguished from pathogenic changes.

Microarray-based CGH detects chromosomal mosaicism not revealed by conventional cytogenetics.

Somatic chromosomal mosaicism is a well-established cause for birth defects, mental retardation, and, in some instances, specific genetic syndromes. We have developed a clinically validated, targeted BAC clone array as a platform for comparative genomic hybridization (aCGH) to enable detection of a wide range of pathologic copy number changes in DNA. It is designed to provide high sensitivity to detect well-characterized submicroscopic micro-deletion and duplication disorders while at the same time minimizing detection of variation of uncertain clinical significance. In the course of studying 2,585 samples submitted to our clinical laboratory, chromosomal mosaicism was detected in 12 patient samples; 10 of these cases were reported to have had a normal blood chromosome analysis. This enhanced ability of aCGH to detect mosaicism missed by routine chromosome analysis may be due to some combination of testing multiple cell lineages and/or failure of cytogenetically abnormal T lymphocytes to respond to mitogens. This suggests that aCGH may detect somatic chromosomal mosaicism that would be missed by conventional cytogenetics.

Metabolic syndrome features and risk of neural tube defects.

BACKGROUND: Maternal obesity and pre-pregnancy diabetes mellitus, features of the metabolic syndrome (MetSyn), are individual risk factors for neural tube defects (NTD). Whether they, in combination with additional features of MetSyn, alter this risk is not known. We evaluated the risk of NTD in association with maternal features of the MetSyn. METHODS: We used a population-based case-control study design in the province of Ontario, Canada. Cases and controls were derived from women who underwent antenatal maternal screening (MSS) at 15 to 20 weeks' gestation. There were 89 maternal cases with, and 434 controls without, an NTD-affected singleton pregnancy. Maternal features of MetSyn were defined by the presence of pre-pregnancy diabetes mellitus, body weight > or = 90th centile among controls, non-white ethnicity and/or serum highly sensitive C-reactive protein (hsCRP) > or = 75th centile of controls. Since hsCRP naturally increases in pregnancy, analyses were performed with, and without, the inclusion of hsCRP in the model. RESULTS: Mean hsCRP concentrations were exceptionally high among study cases and controls (6.1 and 6.4 mg/L, respectively). When hsCRP was excluded from the model, the adjusted odds ratios for NTD were 1.9 (95% confidence interval 1.1-3.4) in the presence 1 feature of MetSyn, and 6.1 (1.1-32.9) in the presence of 2 or more features. When hsCRP was included, the respective risk estimates were attenuated to 1.6 (0.88-2.8) and 3.1 (1.2-8.3). CONCLUSION: We found about 2-fold and 6-fold higher risk for NTD in the presence 1, and 2 or more features, of the metabolic syndrome, respectively. It is not clear whether this risk is altered by the presence of a high serum hsCRP concentration.

RETIRED: Prenatal screening for fetal aneuploidy.

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The implications of a false positive second-trimester serum screen for Down syndrome.

OBJECTIVE: To investigate the genetic and obstetric implications of false positive Down syndrome serum screening results. METHODS: The study population comprised 162774 women underwent triple marker screening in the Ontario Maternal Serum Screening program between October 1995 and September 1998, with outcomes obtained from the Canadian Institute of Health Information. The study compares the incidence of chromosomal abnormalities other than Down syndrome in screen positive women with background incidence from the literature. It also compares the risks of having a fetus with congenital abnormalities or of developing obstetric complications in 11549 screen positive women with their matched controls. RESULTS: A higher incidence of trisomy 13 (12.4 per 10000) was seen in screen positive women; the incidence of other chromosomal abnormalities in screen positive women was not increased relative to the general population. The higher incidence of trisomy 13 may have been biased by the selective uptake of amniocentesis in women who had high risks for Down syndrome or abnormal ultrasound findings. Incidences of fetal congenital abnormality in screen positive and negative women were similar. Women who screened positive for Down syndrome had increased risk of spontaneous fetal loss (odds ratio 1.80; 95% confidence interval 1.54, 2.07) but no other obstetric complications. CONCLUSION: Among women who screened positive for Down syndrome, we found a higher number of spontaneous fetal losses and a possibly higher risk of having a fetus with trisomy 13. We did not find an increased risk for other chromosomal abnormalities, congenital abnormalities, or other adverse obstetric outcomes.

Prospective experience with integrated prenatal screening and first trimester combined screening for trisomy 21 in a large Canadian urban center.

OBJECTIVES: To evaluate the performance of integrated prenatal screening (IPS) and first trimester combined screening (FTS) for trisomy 21 in a large Canadian urban center. METHOD: Prospective data collection on women having FTS at one center from 1 November 2003 to 31 December 2005, or IPS at another from 1 January 2003 to 31 December 2005. A positive screen was defined as adjusted risk for trisomy 21 >or= 1/200 at term or nuchal translucency >or= 3.5 mm. RESULTS: 32 227 and 14 487 women were screened in the IPS and FTS programs, respectively. Detection rates (DRs) and positive rates (PRs) for trisomy 21 were 88.4% (95% CI: 81.6-91.5) and 3.3% (95% CI: 3.1-3.5) for IPS, and 83.9% (95% CI: 74.7-93.0) and 4.0% (95% CI: 3.7-4.3) for FTS. DR adjusted for viability bias was 85.2% for IPS and 78.6% for FTS. Applying both the screens to the 78 134 women who submitted prenatal screens in Ontario in 2005, thereby eliminating the effect of differences in the distribution of maternal age between screens, gave a DR (corrected for viability bias) and PR of 81 and 3.1% for IPS, and 76 and 3.4% for FTS. CONCLUSIONS: Both IPS and FTS perform well and are feasible in a practical clinical setting.

Second trimester prenatal screening for Down syndrome: the associations between the levels of serum markers in successive pregnancies.

OBJECTIVE: To investigate the association of Down syndrome screening results in successive pregnancies, and assess the impacts of including previous screening results in the current risk estimation on screening performance. METHODS: The study was based on 56,951 women who had triple marker screening in two or more singleton pregnancies in the Ontario Maternal Serum Screening (MSS) program between October 1993 and September 2000. The problem of recurrent false positive results was examined by comparing screening results from different pregnancies in the same individuals. Between-pregnancy associations in the levels of serum markers were estimated using correlation analysis. A published method was used to adjust current risk estimation for previous screening results. The effect of this adjustment was assessed by comparing screening performances prior and subsequent to the adjustment. RESULTS: The observed false positive rate (FPR) in subsequent pregnancies was 2.5 times higher than that expected (26.4% vs 10.7%) among women who screened positive in one previous pregnancy, and 3.9 times higher (47.4% vs 12.1%) among women who screened positive in two previous pregnancies. Adjusting for a previous screening result will significantly reduce the recurrent FPR without compromising detection. CONCLUSION: Risk estimation for Down syndrome may be adjusted using the screening result from a previous pregnancy.

Repeated measurement of pregnancy-associated plasma protein-A (PAPP-A) in Down syndrome screening: a validation study.

OBJECTIVES: To confirm that measuring pregnancy-associated plasma protein-A (PAPP-A) in both first- and second-trimester serum samples improves Down syndrome screening. METHODS: We selected paired first- and second-trimester stored serum samples from 34 Down syndrome pregnancies (cases) and 514 unaffected pregnancies (controls) and tested the second-trimester samples for PAPP-A and dimeric inhibin-A (DIA). First-trimester PAPP-A measurements were already available, as were second-trimester measurements of alpha-fetoprotein, unconjugated estriol (uE3), and human chorionic gonadotrophin (hCG). RESULTS: PAPP-A was lower among cases than controls (0.47 MoM) in the first trimester (at an average of 12.5 weeks); in the second trimester, it was not different (0.91 MoM). Using repeated measures of PAPP-A alone, 21 of 34 cases were detected (62%, 95%CI 44% to 78%) with 5% false positives. At an observed 2% false-positive rate, the detection rates (DR) for the quadruple (69%) and serum integrated (69%) tests were lower than for the repeated measures test (75%). Modelled performance at 12 weeks was similar to these observed findings (70, 75, and 82%, respectively). If the first-trimester samples were collected at 10 weeks, however, DR would be higher (70, 81, and 91%, respectively). CONCLUSIONS: Adding a repeated measure of PAPP-A to existing serum markers improves Down syndrome screening to levels that are currently obtainable only by including ultrasound measurement of nuchal translucency (NT). Serum-based screening has the advantages of higher availability and reliability at a lower cost, resulting in a more effective screening strategy. A serum-based repeated measures test has a place in routine Down syndrome screening.

Triploidy identified through second-trimester serum screening.

OBJECTIVES: To describe the maternal serum marker patterns of triploid pregnancies and estimate the second-trimester prevalence of triploidy. METHODS: Forty-two cases of triploidy were identified in six serum screening programmes, five in the United Kingdom, one in Canada. This study describes the serum marker patterns, serum screening results for Down syndrome, trisomy 18 and open neural tube defects, and maternal age of these triploidy cases. The risk cut-off levels were > or = 1 in 250 for Down syndrome, > or =2.5 MoMs alpha-fetoprotein for open neural tube defects and > or =1:100 for trisomy 18 screening. The estimated second-trimester prevalence of triploidy was based on 22 triploidy cases ascertained in 599 934 pregnancies from three routine screening programmes, which attempted complete ascertainment of aneuploidy cases. RESULTS: The observed second-trimester rate of triploidy was 0.37 per 10 000 fetuses. Two different serum marker patterns were seen in triploid pregnancies, distinguished from each other by typically very high or very low levels of total hCG or free beta-hCG. The median maternal ages were respectively 33 years for triploidy with human chorionic gonadotrophin levels < 1.0 MoM, and 26 years for those with hCG levels > or =1.0 MoM. Fifty-seven percent of the pregnancies with a triploid fetus had a risk estimate > or =1:100 for trisomy 18 alone, 10% had an alpha-fetoprotein > or =2.5 MoM, 5% were screen positive for Down syndrome alone, and 19% had an increased risk or positive results for more than one anomaly. CONCLUSION: The simultaneous use of maternal serum tests designed to screen prenatally for Down syndrome, neural tube defects, and an increased risk of trisomy 18 resulted in a screen-positive result for 90% of pregnancies with triploidy.

FISH-mapping of telomeric 14q32 deletions: search for the cause of seizures.

Ring chromosome 14 is a rare cytogenetic disorder. Individuals with r(14) generally have developmental delay and seizures. Other features include hypotonia, microcephaly, mild facial dysmorphism, and retinal pigmentation. Most of these features are also found in patients with linear terminal deletions of chromosome 14, except for seizures and retinal abnormalities. The objective of the study was to determine if deletion of a specific chromosome region is a possible explanation for the occurrence of seizures in patients with ring chromosome 14. Patients diagnosed either with r(14) (six patients) or a deletion of distal 14q (three patients) were analyzed by FISH (fluorescence in situ hybridization) with BAC probes. We observed differences in the size of deletions in the studied group. In two r(14) patients, we did not detect any deletion; the four other patients had deletions of various sizes, ranging from 0.8 Mb to 5 Mb. Two linear deletions were 3.2 Mb and 5.3 Mb in length, respectively; the third case had an interstitial deletion that did not overlap with the others. The deleted regions in ring chromosomes showed overlap with those in the two linear terminal deletions. We conclude that there is unlikely to be a specific deleted locus in 14q32.3 that predisposes r(14) patients to seizures or retinal pigmentation. The cause is probably related to the formation of the ring itself and the effect this may have on local chromatin structure.

The identification of risk of spontaneous fetal loss through second-trimester maternal serum screening.

OBJECTIVE: The purpose of this study was to investigate associations between risk of spontaneous fetal loss and risk estimates for Down syndrome, trisomy 18, and neural tube defects assigned by second-trimester maternal serum screening. STUDY DESIGN: The study involved 264,653 women with available pregnancy outcomes who were screened between 15 and 20 weeks of gestation in the Ontario Maternal Serum Screening Program between October 1995 and September 2000. Pregnancies complicated by fetal chromosomal or structural abnormalities, insulin-dependent diabetes mellitus, and multiple pregnancies were excluded. Spontaneous fetal loss was defined as spontaneous miscarriage and intrauterine fetal demise as classified by the ICD-9 system, but including only those > or = 15 weeks of gestation. Women were grouped according to risk estimates for Down syndrome, trisomy 18, and neural tube defects, respectively. Spontaneous fetal loss rates by each risk group were evaluated after adjusting for losses associated with maternal age and amniocentesis. RESULTS: Fetal loss rates increased in women with risk estimates of > or = 1 in 1110 for trisomy 18 or neural tube defects, and > or = 1 in 130 for Down syndrome. The excessive fetal loss rates for these 3 groups of women were 14.4%, 3.2%, and 1.5% respectively. CONCLUSION: Fetal loss rate markedly increased in women with high-risk estimates for trisomy 18, neural tube defects, and Down syndrome. Risk estimates assigned by triple marker screening may provide an early means of stratifying pregnancies into risk for fetal loss.

Characterization of a novel cation transporter ATPase gene (ATP13A4) interrupted by 3q25-q29 inversion in an individual with language delay.

Specific language impairment (SLI) is defined as failure to acquire normal language skills despite adequate intelligence and environmental stimulation. Although SLI disorders are often heritable, the genetic basis is likely to involve a number of risk factors. This study describes a 7-year-old girl carrying an inherited paracentric inversion of the long arm of chromosome 3 [46XX, inv(3)(q25.32-q29)] having clinically defined expressive and receptive language delay. Fluorescence in situ hybridization (FISH) with locus-specific bacterial artificial chromosome clones (BACs) as probes was used to characterize the inverted chromosome 3. The proximal and distal inversion breakpoint was found to reside between markers D3S3692/D3S1553 and D3S3590/D3S2305, respectively. ATP13A4, a novel gene coding for a cation-transporting P-type ATPase, was found to be disrupted by the distal breakpoint. The ATP13A4 gene was shown to comprise a 3591-bp transcript encompassing 30 exons spanning 152 kb of the genomic DNA. This study discusses the characterization of ATP13A4 and its possible involvement in speech-language disorder.

Accuracy of trisomy 18 screening using the second-trimester triple test.

OBJECTIVE: To assess the accuracy of the calculated risk for trisomy 18 assigned to individual women screened with the second-trimester triple test. METHODS: The study was based on 382598 women screened in the Ontario Maternal Serum Screening Programme between October 1993 and September 2000. Of the women screened, 111 cases of trisomy 18 were identified. Originally, 92874 women were screened using a risk cut-off level method. Estimated risks of trisomy 18 were calculated by applying published population parameters for the remaining women screened using a fixed analyte cut-off method. Women were ranked according to their individual risk for trisomy 18 syndrome in decreasing order and divided into 12 groups. The mean calculated risks of having an affected pregnancy at term for each group were compared with the birth prevalence of the corresponding group after allowing for spontaneous fetal losses. RESULTS: Agreement between the mean calculated risks and the observed prevalence was seen across the entire risk range, although women identified as having high-risk pregnancies had an actual prevalence that was somewhat lower than that estimated by the screen. CONCLUSION: The calculated risk for trisomy 18 syndrome assigned to the individual woman on the basis of the risk cut-off method accurately reflects their risk of having a term trisomy 18 syndrome pregnancy.

Maternal serum marker medians in Aboriginal Canadian women.

OBJECTIVES: The study evaluates the differences between Aboriginal and Caucasian women in the levels of maternal serum markers used in second-trimester Down syndrome screening (alpha-fetoprotein, unconjugated estriol, and total human chorionic gonadotrophin). METHODS: A case-control study compared the levels of serum markers in 401 Aboriginal women and 1565 matched controls selected from 7717 Caucasian women. The cases and controls were screened in a single centre and matched for maternal age, parity, and sample date. Women with multiple pregnancies and pregnancies associated with Down syndrome, open neural tube defects, trisomy 18, and insulin-dependent diabetes mellitus as well as women without weight recorded were excluded from the study. RESULTS: No differences in the levels of maternal serum alpha-fetoprotein and total human chorionic gonadotrophin were observed between the two groups. Maternal serum unconjugated estriol was 12% higher in Aboriginal women. DISCUSSION: Since Aboriginal women make up only a small proportion of women screened, correcting the level of uE3 for this group will have little effect on the overall screening performance. However, if these results are confirmed by further study, individual centres may consider making this correction, so optimal screening performance can be achieved in Aboriginal women.

Double-positive maternal serum screening results for down syndrome and open neural tube defects: An indicator for fetal structural or chromosomal abnormalities and adverse obstetric outcomes.

OBJECTIVE: The study was designed to evaluate whether double positive maternal serum screening results for Down syndrome and open neural tube defects indicate an increased risk of adverse perinatal outcome. STUDY DESIGN: A retrospective case-control study was conducted. In a cohort of 170,394 women who underwent maternal serum triple screening in Ontario, Canada, between October 1995 and September 1998, 189 women received positive screening results for both Down syndrome and neural tube defects. Each case was matched to 5 control subjects who had negative screening results for test center, maternal age, and specimen date. The risks for adverse perinatal outcomes were compared. RESULTS: Women with double-positive screening results had significantly higher risks of having fetuses with structural abnormalities (odds ratio, 14.5) and chromosomal anomalies (odds ratio, 36.3). They also had higher risks of having preeclampsia (odds ratio, 6.7), small-for-gestational age (odds ratio, 9.7), preterm delivery (odds ratio, 5.9), miscarriage, and intrauterine fetal death (odds ratio, 11.8). CONCLUSION: Double-positive maternal serum screening results are associated with fetal structural and chromosomal abnormalities and/or adverse pregnancy outcomes. Close fetal and maternal surveillance are indicated when such pregnancies are identified.