Case Studies in NIPT: NIPT for detection of subchromosomal deletions and duplications

March 5, 2015

Case Studies in NIPT pairs fascinating real case examples involving use of NIPT with commentaries from experts on practical clinical implications of the technology. This case was provided by Sequenom and peer reviewed by series co-chairs Joe Leigh Simpson, MD, and Ronald J. Wapner, MD.

Case Studies in NIPT pairs fascinating real case examples involving use of NIPT with commentaries from experts on practical clinical implications of the technology. This case was provided by Sequenom and peer reviewed by series co-chairs Joe Leigh Simpson, MD, and Ronald J. Wapner, MD.


By Julie Jesiolowski1, Thomas Monroe1, Courtney Fitch3, Jenna Wardrop2, Theresa Boomer2, and Juan-Sebastian Saldivar2

1 Sequenom Laboratories, Morrisville, NC; 2 Sequenom Laboratories, San Diego, CA; 3 Lehigh Valley Maternal Fetal Medicine, Allentown, PA


Introduction: Noninvasive prenatal testing (NIPT) has rapidly changed the prenatal landscape for pregnant women at increased risk of fetal aneuploidy. This technology provides high sensitivity and specificity for Trisomy 21, 18, and 13. Overrepresentation of a chromosome can be detected by an increased Z-score in comparison with a normal euploid genome. Here we report a case involving a partial chromosome 13 duplication that assisted in identifying a maternal balanced translocation and revealed the limitations of suboptimal karyotype resolution.

Methods: Maternal plasma samples were subjected to DNA extraction and library preparation followed by massively parallel sequencing as described by Palomaki et al. Sequencing data were analyzed using a novel algorithm to detect trisomies and other subchromosomal events as described by Chen et al.

Results: A 38-year-old G8P3043 presented for NIPT due to advanced maternal age and a previous pregnancy history of Trisomy 13, confirmed by low-resolution karyotype on peripheral blood. Ultrasound at 12 weeks, 6 days was suspicious for micrognathia. NIPT studies were ordered and results were positive for Trisomy 13. Sequencing data were reviewed based on the clinical history and revealed a 24.3Mb duplication of 13q31.2 and an apparent 27.85Mb deletion of 4q32.2. A fetal demise was noted on a 15 week, 0 day ultrasound. Products of conception (POC) studies were performed at 450–500 band resolution resulting in a normal female karyotype 46,XX, discordant from the NIPT results. Follow-up maternal studies at high-resolution karyotype analysis detected a balanced translocation: 46,XX,t(4;13)(q32;q31). After informing the tissue analysis laboratory of the translocation events, discordance between NIPT and POC studies was attributed to low karyotype resolution.

Conclusion: This case demonstrates the power of NIPT sequencing technology and optimized bioinformatics. Clinicians should be conscious that standard karyotyping does not have sufficient resolution to detect subchromosomal events detected by NIPT. Accurate clinical information provided to the laboratory may aid in additional interpretation. In these cases, microarray studies or high-resolution karyotype should be used to confirm a suspected abnormality.


NEXT: Commentary from Ronald J. Wapner, MD >>




By Ronald J. Wapner, MD

Dr. Wapner is Professor of Maternal Fetal Medicine at Columbia University, New York, and co-chair of this series. He reports performing contracted research for Ariosa, Illumina, Natera, the NIH, and Sequenom. All funds go to the university.

This case demonstrates the ability of NIPT performed by massively parallel sequencing to identify a relatively large subchromosomal deletion and duplication, which raises interesting issues. First, when a fetal structural anomaly is suspected, testing by chorionic villus sampling or amniocentesis with analysis by chromosomal microarray rather than NIPT is the recommended procedure to evaluate the underlying genomic cause. In this case, it was fortunate that the chromosomal abnormalities were sufficiently large to be identified by NIPT. Many clinically relevant pathogenic microdeletions or duplications are too small to be identified by routine NIPT screening or do not involve the chromosomes that NIPT is intended to screen.

Second, it is unusual for a karyotype to miss such a large rearrangement. In general, prenatal samples at the 450–500 band level should be sufficient to identify the chromosomal alterations seen in this case, both of which were over 20 million base pairs. In general, karyotypes identify alteration above 7–10 million base pairs, but admittedly, the threshold may be greater than this when POC are analyzed, especially following a fetal demise. In this case, the fact that the rearranged segments were similar in size and banding patterns further contributed to the error.

This example demonstrates that karyotypes are not without limitations and also illustrates the advantages of molecular-based techniques, which have a resolution of less than 1 million base pairs and do not routinely require tissue culture.

It is likely that with time, NIPT will be able to routinely identify subchromosomal abnormalities significantly smaller than those illustrated in this case. Currently, this is only reliably done by chromosome microarray analysis performed on fetal tissue collected by diagnostic testing. Work is needed to determine the threshold of NIPT for these lesions and to investigate whether the current NIPT approaches can reliably identify subchromosomal alterations for all chromosomes without a significant increase in the false-positive rate. The authors of this report are to be congratulated for their diligence in determining the abnormality.

As our knowledge of sequencing technology for NIPT increases and the cost of sequencing decreases, allowing higher-resolution analysis, it is certain that more such examples will become evident.