Mary Hoi Yin Tang

Non-invasive prenatal assessment of Trisomy 21 by multiplexed maternal plasma DNA sequencing: large scale validity study

Objectives To validate the clinical efficacy and practical feasibility of massively parallel maternal plasma DNA sequencing to screen for fetal trisomy 21 among high risk pregnancies clinically indicated for amniocentesis or chorionic villus sampling. Design Diagnostic accuracy validated against full karyotyping, using prospectively collected or archived maternal plasma samples. Setting Prenatal diagnostic units in Hong Kong, United Kingdom, and the Netherlands. Participants 753 pregnant women at high risk for fetal trisomy 21 who underwent definitive diagnosis by full karyotyping, of whom 86 had a fetus with trisomy 21. Intervention Multiplexed massively parallel sequencing of DNA molecules in maternal plasma according to two protocols with different levels of sample throughput: 2-plex and 8-plex sequencing. Main outcome measures Proportion of DNA molecules that originated from chromosome 21. A trisomy 21 fetus was diagnosed when the z score for the proportion of chromosome 21 DNA molecules was >3. Diagnostic sensitivity, specificity, positive predictive value, and negative predictive value were calculated for trisomy 21 detection. Results Results were available from 753 pregnancies with the 8-plex sequencing protocol and from 314 pregnancies with the 2-plex protocol. The performance of the 2-plex protocol was superior to that of the 8-plex protocol. With the 2-plex protocol, trisomy 21 fetuses were detected at 100% sensitivity and 97.9% specificity, which resulted in a positive predictive value of 96.6% and negative predictive value of 100%. The 8-plex protocol detected 79.1% of the trisomy 21 fetuses and 98.9% specificity, giving a positive predictive value of 91.9% and negative predictive value of 96.9%. Conclusion Multiplexed maternal plasma DNA sequencing analysis could be used to rule out fetal trisomy 21 among high risk pregnancies. If referrals for amniocentesis or chorionic villus sampling were based on the sequencing test results, about 98% of the invasive diagnostic procedures could be avoided.

Noninvasive prenatal diagnosis of fetal trisomy 18 and trisomy 13 by maternal plasma DNA sequencing.

Massively parallel sequencing of DNA molecules in the plasma of pregnant women has been shown to allow accurate and noninvasive prenatal detection of fetal trisomy 21. However, whether the sequencing approach is as accurate for the noninvasive prenatal diagnosis of trisomy 13 and 18 is unclear due to the lack of data from a large sample set. We studied 392 pregnancies, among which 25 involved a trisomy 13 fetus and 37 involved a trisomy 18 fetus, by massively parallel sequencing. By using our previously reported standard z-score approach, we demonstrated that this approach could identify 36.0% and 73.0% of trisomy 13 and 18 at specificities of 92.4% and 97.2%, respectively. We aimed to improve the detection of trisomy 13 and 18 by using a non-repeat-masked reference human genome instead of a repeat-masked one to increase the number of aligned sequence reads for each sample. We then applied a bioinformatics approach to correct GC content bias in the sequencing data. With these measures, we detected all (25 out of 25) trisomy 13 fetuses at a specificity of 98.9% (261 out of 264 non-trisomy 13 cases), and 91.9% (34 out of 37) of the trisomy 18 fetuses at 98.0% specificity (247 out of 252 non-trisomy 18 cases). These data indicate that with appropriate bioinformatics analysis, noninvasive prenatal diagnosis of trisomy 13 and trisomy 18 by maternal plasma DNA sequencing is achievable.

Early ultrasound prediction of pregnancies affected by homozygous α-thalassaemia-1

Homozygous α‐thalassaemia‐1 is conventionally diagnosed by invasive testing on all at‐risk pregnancies. We evaluated the diagnostic efficacy of non‐invasive abdominal ultrasonographic cardiothoracic ratio measurement in 62 pregnancies at 13–14 weeks and 75 pregnancies at 17–18 weeks. This performed better than placental thickness measurement. Using a cardiothoracic ratio cut‐off level of ≥0·5, 75 per cent of affected pregnancies were detected at 13–14 weeks and all cases were detected at 17–18 weeks. False‐positive rates were 7 and 8 per cent, respectively. There was no false‐positive diagnosis if the cardiothoracic ratio was ≥0·53. With this approach, invasive procedures can be selectively performed and fewer pregnancies will be lost unnecessarily. The reduction in medical expenses is likely to be substantial.

Prenatal ultrasonographic prediction of homozygous type 1 α-thalassemia at 12 to 13 weeks of gestation☆☆☆★

We determined the feasibility of prenatal prediction of type 1 homozygous alpha-thalassemia at 12 to 13 weeks of gestation by either abdominal or vaginal (or both) ultrasonographic examination to measure the fetal cardiothoracic ratio in 135 at-risk pregnancies. Forty-three fetuses were affected by homozygous type 1 alpha-thalassemia. The mean cardiothoracic ratio was significantly larger than that of the unaffected fetuses (0.54 vs 0.45, P <.0005), with no overlap between the 2 groups. A cardiothoracic ratio cutoff point of >/=0.5 was 100% sensitive and specific for disease.

Noninvasive Prenatal Molecular Karyotyping from Maternal Plasma

Fetal DNA is present in the plasma of pregnant women. Massively parallel sequencing of maternal plasma DNA has been used to detect fetal trisomies 21, 18, 13 and selected sex chromosomal aneuploidies noninvasively. Case reports describing the detection of fetal microdeletions from maternal plasma using massively parallel sequencing have been reported. However, these previous reports were either polymorphism-dependent or used statistical analyses which were confined to one or a small number of selected parts of the genome. In this report, we reported a procedure for performing noninvasive prenatal karyotyping at 3 Mb resolution across the whole genome through the massively parallel sequencing of maternal plasma DNA. This method has been used to analyze the plasma obtained from 6 cases. In three cases, fetal microdeletions have been detected successfully from maternal plasma. In two cases, fetal microduplications have been detected successfully from maternal plasma. In the remaining case, the plasma DNA sequencing result was consistent with the pregnant mother being a carrier of a microduplication. Simulation analyses were performed for determining the number of plasma DNA molecules that would need to be sequenced and aligned for enhancing the diagnostic resolution of noninvasive prenatal karyotyping to 2 Mb and 1 Mb. In conclusion, noninvasive prenatal molecular karyotyping from maternal plasma by massively parallel sequencing is feasible and would enhance the diagnostic spectrum of noninvasive prenatal testing.

Comparison of nuchal and detailed morphology ultrasound examinations in early pregnancy for fetal structural abnormality screening: a randomized controlled trial

To compare the effectiveness of a nuchal scan at 10 to 14 + 6 weeks and a detailed morphology scan at 12 to 14 + 6 weeks in screening for fetal structural abnormalities.

Can amnio-polymerase chain reaction alone replace conventional cytogenetic study for women with positive biochemical screening for fetal Down syndrome?

OBJECTIVE To determine whether amnio–polymerase chain reaction (amnio-PCR) can replace conventional cytogenetic study for confirming the karyotype of fetuses in women with positive biochemical screening for fetal Down syndrome. METHOD To check the accuracy of this technique in our laboratory, we first compared the amnio-PCR results with those of conventional cytogenetic study in 235 patients referred from June 1999 to December 2001 for prenatal diagnosis in a referral center in Hong Kong. We then reviewed the results of 1526 amniotic fluid cultures performed for positive fetal Down syndrome screening between January 1997 and December 2001 and classified them as detectable or not detectable by amnio-PCR, using the assumption that we had replaced conventional cytogenetic study with amnio-PCR. RESULTS The 235 amnio-PCR results were all informative, without a false-positive or false-negative result. Of the 1526 cases with positive fetal Down syndrome screening and no ultrasound abnormalities, only two cases of sex chromosome abnormalities and two cases of marker chromosomes would have been missed if conventional cytogenetic study had been replaced by amnio-PCR. CONCLUSION Amnio-PCR can be an alternative to conventional cytogenetic study for women with positive biochemical screening for fetal Down syndrome and no demonstrable fetal structural abnormality.

Prevalence and specificity of clinically significant red cell alloantibodies in Chinese women during pregnancy – a review of cases from 1997 to 2001

Summary.  Guidelines for the prevention and management of red cell alloantibodies during pregnancy, related to anti‐D in particular, are well established in Caucasian populations. However, because of the racial difference of the blood group distribution, applicability to Chinese is unknown as a result of insufficient data on the prevalence and their outcome. In a retrospective review of 28 303 (21 327 Chinese) antenatal attendances from 1997 to 2001, 213 (0·79%) women were found to have a total of 230 irregular antibodies. About 137 (0·64%) were ethnic Chinese, and a total of 160 irregular antibodies were identified in their blood samples. About 58 of these Chinese women (0·27%) were found to have 66 clinically significant antibodies. There was only one case of anti‐D detected in an Rh(D)‐negative subject. Our study shows the overall prevalence of clinically significant antibodies in Chinese women, which was not different from that of the Western population. However, the specificities of the antibodies differ with the commonest antibodies encountered; these being anti‐Mi (57·6%), anti‐E (19·7%), anti‐S (10·6%) and anti‐c (7·6%). Neonatal jaundice was observed in 37 babies and 10 of them required phototherapy. The findings support the previous recommendation that routine antenatal antibody screening for Chinese women may not be worthwhile except in Rh(D)‐negative subjects or those with an antecedent history of haemolytic disease of the newborn (HDN). The relative high incidence of anti‐Mi in the present study and the local population, in general, may warrant a large‐scale prospective study of pregnancy outcome in these subjects, especially in the light of the previous case reports of HDN because of anti‐Mi.

Three‐dimensional extended imaging: a new display modality for three‐dimensional ultrasound examination

To describe Three‐Dimensional eXtended Imaging (3DXI)™ as a new display modality for three‐dimensional (3D) ultrasound examination of the fetus.

Nuchal translucency in pregnancies conceived after assisted reproduction technology

Levels of maternal serum markers of fetal Down syndrome in pregnancies conceived after assisted reproduction are different from those of normal spontaneous pregnancies. The present study examined the effects of conventional in‐vitro fertilization (IVF), intracytoplasmic sperm injection (ICSI) and embryo cryopreservation on nuchal translucency (NT) thickness.