Non-Invasive Prenatal Testing (NIPT) has rapidly become an integral part of clinical practice, providing a highly accurate and non-invasive method for detecting common fetal chromosomal abnormalities, such as trisomy 21 (Down syndrome), trisomy 18, and trisomy 13. Traditionally, NIPT focused on these common chromosomal aneuploidies and sex chromosome abnormalities (SCAs). However, technological advancements have broadened the scope of NIPT to a Genome-Wide (GW) approach, which now includes the detection of rare autosomal aneuploidies (RAAs) and microdeletions/duplications. These can be smaller than 7Mb in size, encompassing syndromes like DiGeorge, Prader-Willi/Angelman, Cri du Chat, and 1p36 deletion syndrome.
A key factor in interpreting NIPT results is the confined placental mosaicism (CPM), a condition affecting about 1-2% of pregnancies. This occurs when chromosomal abnormalities are present in the placenta but not in the fetus, leading to discordant test results. In these cases, NIPT may show a false-positive result that does not reflect the fetus’s actual chromosomal status. False-positive results can also be caused by maternal chromosomal abnormalities or tumors, as well as in cases of vanishing twins in twin pregnancies.
Due to the potential for these discordant results, major clinical guidelines from organizations like the American College of Medical Genetics and Genomics (ACMG) and the International Society for Prenatal Diagnosis (ISPD) have recommended caution in the routine clinical use of genome-wide NIPT for rare aneuploidies and structural variations. Nevertheless, studies continue to explore the clinical utility of GW-NIPT in providing a more comprehensive view of fetal health without increasing the number of false positives.
Study overview and methods
This study was conducted to assess the clinical utility of GW-NIPT in a large cohort of 71,883 pregnancies, including both singleton and twin pregnancies, from November 2019 to December 2021. The average maternal age was 38, with a median gestational age of 12 weeks, and the tests were primarily performed for advanced maternal age or patient preference.
The Veriseq NIPT v2 kit (Illumina, Inc.) was used for testing, and the entire analysis process was carried out at Eurofins Genoma Laboratories. The results were analyzed using NIPTFlow, a proprietary algorithm, to detect chromosomal aneuploidies and copy number variations (CNVs), including rare autosomal aneuploidies and microdeletions.
The sensitivity and specificity of the GW-NIPT were calculated using the statistical software MedCalc, and follow-up diagnostic tests were conducted when necessary.
Results
Out of the 71,883 tests conducted, 70,869 results were obtained on the first attempt, while 1,268 required a second analysis. Ultimately, 254 cases (0.35%) did not yield any result. Among the successfully analyzed cases, 1,011 (1.4%) were positive for some chromosomal abnormality. Follow-up diagnostic testing confirmed 781 of these positive cases.
Detection of common trisomies and sex chromosome aneuploidies
Among the 579 positive cases for common trisomies, 440 were classified as trisomy 21, with 437 confirmed through prenatal diagnosis. There were three false-positive and two false-negative results for trisomy 21. In cases of trisomy 18, 93 out of 94 positive cases were confirmed, with only one false positive and no false negatives.
The results for SCAs showed a high degree of accuracy, with 156 out of 173 positive cases confirmed by invasive testing. There were 17 false positives and one false negative. The overall sensitivity and specificity for SCAs were 99.36% and 99.97%, respectively.
Genome-Wide analysis for rare autosomal aneuploidies (RAAs)
The genome-wide screening for rare autosomal aneuploidies was performed on 65% of the cohort (46,724 patients). Out of 69 positive cases, 33 were confirmed through prenatal diagnosis, including several cases of mosaicism. The most frequently detected rare trisomies involved chromosomes 7, 15, and 22.
In twin pregnancies, GW-NIPT successfully distinguished between cases where only one fetus was affected, even in cases where the other twin had been reabsorbed.
Detection of microdeletions
GW-NIPT identified 36 cases of microdeletions in 28,753 patients, with 20 of these confirmed by invasive testing. Microdeletions included 5 confirmed cases of DiGeorge syndrome and other rare deletions. There was only one false-negative result for microdeletions, and the sensitivity for microdeletions was 83.33%, with a specificity of 99.99%.
Discussion
This study demonstrated that the GW-NIPT approach used by Eurofins Genoma Laboratories achieved high clinical sensitivity and specificity, comparable to or exceeding previous studies. For common trisomies, the positive predictive value (PPV) was 97.9%, with particularly high PPVs for trisomy 21 (99.3%) and trisomy 18 (98.9%).
The study also highlighted the challenges posed by confined placental mosaicism, which remains a leading cause of false-positive results, particularly for trisomy 13 and SCAs. Despite this, the high accuracy of NIPT for detecting common and rare aneuploidies, coupled with genetic counseling, supports the clinical utility of GW-NIPT in pregnancy management.
Conclusion
Accurate NIPT analysis requires a careful combination of biological factors, bioinformatic algorithms, and expert interpretation. The use of an algorithm alone is not enough; clinical context, patient history, and ultrasound findings must be considered to ensure accurate interpretation of NIPT results. At Eurofins Genoma Laboratories, the combination of advanced algorithms and expert analysis has led to high clinical sensitivity without sacrificing specificity, even for microdeletions. The ability to detect genome-wide chromosomal aberrations with such precision reinforces the clinical value of extended NIPT in managing pregnancies, particularly with genetic counseling to guide patient care.