The Role of Thickened Nuchal Translucency on Copy Number Variations and Pregnancy Outcomes in Northeast Coast of Fujian Province, China: A Comparison Between Traditional Karyotyping and Single Nucleotide Polymorphism Array Analysis

doi:10.21203/rs.3.rs-3194093/v1

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Research Article

The Role of Thickened Nuchal Translucency on Copy Number Variations and Pregnancy Outcomes in Northeast Coast of Fujian Province, China: A Comparison Between Traditional Karyotyping and Single Nucleotide Polymorphism Array Analysis

https://doi.org/10.21203/rs.3.rs-3194093/v1

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Background

Thickened nuchal translucency (NT; ≥2.5 mm) is closely associated with various chromosomal abnormalities, structural abnormalities, and genetic diseases. However, the cutoff value of thickened NT for invasive prenatal diagnosis remains inconsistent, and little research has been conducted on pathogenic copy number variations (CNVs) affecting the outcomes in foetuses with thickened NT. This study aimed to assess the cutoff value for thickened NT to predict aneuploid foetuses and CNVs associated with thickened NT in prenatal diagnosis to determine the characteristics of pathogenic CNVs, which would assist the genetic counselling of women with thickened NT.

Methods

Ninety pregnant women with thickened NT who underwent invasive prenatal diagnosis using traditional karyotyping and single nucleotide polymorphism (SNP) array analysis in diagnostic institutions between January 2021 and March 2023 were included. The accuracy of the thickened NT cutoff value for diagnosing aneuploid abnormalities was assessed using receiver operating characteristic (ROC) curve analysis.

Results

Karyotype analysis identified 15 chromosomal abnormalities. In addition to the 10 chromosomal abnormalities corresponding to routine karyotyping, SNP array analysis identified six patients with CNVs but normal karyotypes. ROC curves demonstrate that the NT measured between 11 and 13^+ 6 weeks was associated with 0.729 area under the curve (AUC; 95% CI: 0.56–0.898, P = 0.019) with the foetal aneuploidy. The ROC curve revealed that the cutoff value of NT was 4.15 mm, which showed 50% sensitivity and 90% specificity for detecting aneuploidies. With increasing NT thickness, the probability of aneuploidy increases in the first trimester.

Conclusion

Aneuploid abnormalities and CNVs are closely related to a thickened NT, which affects pregnancy outcomes. Thickened NT and abnormal CNVs are closely related and are not only associated with chromosome aneuploidy in the first trimester. We recommend that karyotyping and SNP array analysis should be performed for prenatal diagnosis in all foetus with NT thickness > 2.5 mm, regardless of NT thickness > 3.0 or 3.5 mm to avoid the occurrence of child birth with genetic defects due to missing prenatal diagnosis.

Copy number variations

Karyotyping

Pregnancy outcomes

Prenatal diagnosis

ROC curve

Thickened nuchal translucency

In the early 1990s, nuchal translucency (NT) screening was first reported as an ultrasound indicator of chromosomal abnormalities in the first trimester of pregnancy, particularly for detecting Down syndrome (full trisomy of chromosome 21)[1]. Foetal NT is a clinically significant indication for ultrasound screening between 11 and 13^+ 6 weeks of the first trimester. At present, ≥ 3.5 mm NT, which is above the 99th centile irrespective of the gestational age and foetal crown rump length (CRL), is one of the most sensitive indicators of foetal chromosomal abnormalities and a definite indicator for invasive prenatal diagnosis[2]. Foetuses with thickened NT have an increased risk of chromosomal aneuploidy, pathogenic copy number variations (pCNVs), prenatal or perinatal death, and other adverse pregnancy outcomes[3–5]. Moreover, the risk of foetal abnormalities and adverse pregnancy outcomes is proportional to the degree of NT enlargement. When a thickened foetal NT is detected in the first trimester, amniocentesis using an invasive prenatal diagnostic technique is performed to exclude chromosomal defects at approximately 20 weeks of gestation. Recent studies have reported that NT increase is associated with not only chromosomal aneuploidy, but also chromosomal CNVs, cardiac and other structural anomalies, and single-gene disorders (such as Roberts syndrome and KAT6A syndrome)[4–7]. In addition to prenatal screening, NT and maternal age are independent indicators in first trimester[5]. Chromosome anomalies have been found in up to 69% foetuses with 3.0–3.4 mm thick NT and trisomy13, 18, and 21 in the first trimester of gestation[8]. However, the cutoff value for thickened NT in invasive prenatal diagnosis is inconsistent and differs across countries. For example, some countries used cutoff values such as ≥ 2.5, ≥ 3.0, or ≥ 3.5 mm[9]. Inconsistent criteria have resulted in controversial data and analyses, affecting the potential use of NT in prenatal diagnosis.

Standard karyotyping has been considered the “gold standard” to detect chromosomal abnormalities for a long time in the past, which would detect numerical abnormalities and balanced translocations, such as free trisomies, Robertsonian translocations, and large structural abnormalities of chromosomes (> 5–10 Mb) during prenatal diagnosis[10, 11]. However, karyotyping requires a long time for cell culture and karyotype analysis and low resolution (> 5–10 Mb) for cytogenetic detection. Chromosomal microarray analysis (CMA) is used to detect CNVs in the whole genome. This method can be used to detect CNVs of chromosome imbalances, such as aneuploidy and unbalanced rearrangements, particularly chromosome microdeletions and duplications that cannot be detected using standard karyotyping[12, 13].

Ningde is located on the northeastern coast of Fujian Province, with 3.15 million population and 7.29‰ annual birth rate of (The Bureau of Statistics of Ningde, China). Ningde is the largest She-inhabited location in China. Unlike other ethnic groups, the She population have specific traditional lifestyles and genetic backgrounds. The She population have a unique hereditary background, and no reports have focused on CNV analysis in foetuses with increased NT using single nucleotide polymorphism (SNP) array in the patient population in Ningde, China. This study aimed to analyse the CNVs and pregnancy outcomes in foetuses with thickened NT using SNP array to evaluate its application in prenatal diagnosis.

Patient data

Ninety pregnant women with thickened NT, who accepted invasive prenatal diagnosis for traditional karyotyping and SNP array analysis at the diagnostic institutions of the Ningde Municipal Hospital affiliated with the Ningde Normal University between January 2021 and March 2023 were retrospectively analysed. Fetal tissue samples were collected from foetus with thickened NT at a median gestational age of 19^+ 5 weeks (range: 18^+ 1–25^+ 4 weeks) using amniocentesis. The median age at pregnancy was 30 years (range: 18–43 years). Under ultrasound guidance, 30 mL amniotic fluid was extracted from pregnant women, of which 20 mL was used for chromosome karyotyping by culturing in two lines and the remaining 6–10 mL was used for SNP array analysis. All pregnant women underwent genetic counselling and signed informed consent forms prior to the invasive diagnostic procedures. Once pathogenic CNVs in the foetus were confirmed, approximately 2.0 mL peripheral venous blood was collected from the parents to verify inheritance. DNA was extracted using the Gentra Puregene Blood Kit (QIAGEN, Santa Clara, CA, USA). This study was approved by the Ethics Committee of Ningde Municipal Hospital affiliated with Ningde Normal University (NSYKYLL-2023-028).

Cytogenetic analysis

Lymphocytes and amniocytes were analysed by traditional karyotyping using the Giemsa banding technique at 400–550 band resolution.

CMA (SNP array)

According to the manufacturer's instructions, the QIAamp® DNA Blood kits (Qiagen, Valencia, CA, USA) was used to directly extract foetal genomic DNA from uncultured amniotic fluid. DNA was quantified using a NanoDrop spectrophotometer (Thermo Fisher Scientific, Waltham, MA, USA) according to the manufacturer’s instructions. Briefly, foetal DNA was digested, attached to adapters, amplified, purified, fragmented, biotin-labelled, and hybridised using an Affymetrix® CytoscanTM 750 K array system (Affymetrix, Santa Clara, CA, USA) that contains SNP and CNV probes and detects CNVs, mosaics (mosaic proportion > 10%), and loss of heterozygosity (LOH). The SNP array detected imbalances within the kilobase range. The results obtained from the CytoScan arrays were analysed using Chromosome Analysis Suite software (Affymetrix) with annotations of the genome version GRCH37 (hg19)[10]. The detected copy number gains or losses were systematically evaluated by comparing the values available in publicly available databases, including the Database of Genomic Variants (http://projects.tcag.ca/variation/), the DECIPHER database (http://decipher.sanger.ac=/), International Standards for Cytogenomic Arrays (https://www.iscaconsortium.org/), and Online Mendelian Inheritance in Man (OMIM; http://www.omim.org). CNVs were categorised as benign, pathogenic, or variants of uncertain significance (VOUS) according to American College of Medical Genetics guidelines[14, 15]. SNP array analysis of DNA from maternal and paternal blood samples was performed to determine whether the CNVs detected in the foetal samples were de novo or inherited. All de novo CNVs were validated using metaphase fluorescence in situ hybridisation (FISH).

Receiver operating characteristic curve for foetal aneuploidy

To explore the diagnostic efficiency, we assessed whether NT values could be used to predict aneuploidies using ROC curve analysis. The ROC curve is a comprehensive index reflecting the sensitivity and specificity of continuous variables, which is also used to determine the accuracy of the observation variables. The area under the ROC curve displays objectivity in prenatal diagnosis. Generally, a large area under the curve (AUC) reflects high screening efficiency.

Statistical analysis

IBM SPSS Statistics version 20 (IBM Corp., Armonk, NY, USA) was used for the statistical analysis. The chi-square (χ²) test was performed to determine the significance of the differences between the NT values. The diagnostic accuracy of the threshold was assessed using ROC curve analysis for chromosomal aneuploidies. Differences were considered statistically significant at P < 0.05.

A total of 90 women pregnant with foetuses having ≥ 2.5 mm NT, with a median age of 30 years (range: 18–43 years) were included in this study. According to the degree of thickened NT, the pregnant women were divided into four groups: 2.5–2.9, 3.0–3.4, 3.5–3.9, and ≥ 4.0 mm. With a thickened NT, the aneuploidy detection rates were 5.0%, 15%, 5%, and 25%, and those of other abnormal CNVs were 15%, 10%, 0%, and 5%, respectively. The highest detection rate for aneuploidy and abnormal CNVs was in the foetuses with ≥ 4.0 mm NT (up to 25%) and 2.5–2.9 mm NT, respectively. Rearrangements and abnormal CNVs were not detected in the foetuses with 3.5–3.9 mm NT. The detection rate of abnormal CNVs in the foetuses with ≥ 4.0 mm NT was higher (43.75%; 7/16) than that in the foetuses with 3.5–3.9 mm NT (16.22%; 6/37) (χ² = 4.57, P = 0.032). All results are presented in Table 1.

Table 1

Abnormal foetal karyotyping and copy number variations grouped by NT thickness (11–13^+ 6 weeks)
Group	Number of samples, n (%)	Age (mean ± SDs)	Number of abnormal karyotyping and CNVs
Group	Number of samples, n (%)	Age (mean ± SDs)	Aneuploidy, n (%)	rearrangements, n (%)	Abnormal CNVs, n (%)
NT 2.5–2.9 mm	27(30.0)	30.58 ± 5.3	1(5.0)	2(10.0)	3(15.0)
NT 3.0-3.4 mm	37(41.1)	29.53 ± 3.7	3(15.0)	1(5.0)	2(10.0)
NT 3.5–3.9 mm	10(11.1)	30.0 ± 4.7	1(5.0)	0	0
NT ≥ 4.0 mm	16(17.8)	30.6 ± 6.1	5(25.0)	1(5.0)	1(5.0)

NT: Nuchal translucency; chromosomal rearrangements, including duplications, inversions, and balanced translocations.

Cytogenetic analysis of foetuses with thickened NT

Karyotyping and SNP array analysis have identified 16 chromosomal abnormalities, including 11 chromosomal abnormalities with pathogenicity, such as trisomy 18 (Trisomy 18 syndrome; n = 2), trisomy 21 (Down syndrome; n = 8) and 15q duplication syndrome (n = 1), in the 90 foetuses with thickened NT. Four foetuses showed abnormal chromosomal structures, such as pericentric (9)(p12q13) inversion (n = 1), and one foetus had balanced genetic translocation t(4;13)(q31.1;q32). Moreover, two foetuses exhibited chromosomal polymorphisms associated with 1qh + and 21pss (Table 2).

Table 2

Abnormal karyotyping results of foetuses with thickened nuchal translucency
Case	Karyotype	SNP-array results	Pathogenicity classification	Postnatal outcome
1	47,XY,+18	arr[hg19](18)x3	P (Edward syndrome)	TP
2	47,XY,+18	arr[hg19](18)x3	P (Edward syndrome)	TP
3	47,XX,+21	arr[hg19](21)x3	P(Down syndrome)	TP
4	47,XX,+21	arr[hg19](21)x3	P(Down syndrome)	TP
5	47,XX,+21	arr[hg19](21)x3	P(Down syndrome)	TP
6	47,XY,+21	arr[hg19](21)x3	P(Down syndrome)	TP
7	47,XX,+21	arr[hg19](21)x3	P(Down syndrome)	TP
8	47,XX,+21	arr[hg19](21)x3	P(Down syndrome)	TP
9	47,XX,+21	arr[hg19](21)x3	P(Down syndrome)	TP
10	47,XX,+21	arr[hg19](21)x3	P(Down syndrome)	TP
11	46,XX,dup(15)(q11.2q13.1)	arr[hg19]15q11.2q13.1(23,286,423 − 28,526,905)x3	P(15q duplication syndrome)	TP
12	46,XY,t(4;13)(q31.1;q32)	arr[hg19](1–22) × 2,(XX) × 1	Benign	TD
13	46,XY,inv(9)(p12q13)	arr[hg19](1–22) × 2,(XX) × 1	Benign	TD
14	46,XY,inv(9)(p12q13)	arr[hg19](1–22) × 2,(XX) × 1	Benign	TD
15	46,XY,1qh+	arr[hg19](1–22) × 2,(XX) × 1	Benign	TD
16	46,XX,21pss	arr[hg19](1–22) × 2,(XY) × 1	Benign	TD

SNP, single nucleotide polymorphism; TP, termination of pregnancy; TD term delivery

SNP array analysis results

The results of SNP array analysis and normal karyotyping were consistent in 11 foetuses with chromosomal abnormalities. In addition, SNP array analysis identified another six foetuses with abnormal CNVs. Among these, the CNVs in six foetuses were VOUS. The size of abnormal CNVs were 0.3–11.7 Mb. Among these CNVs, five were microdeletions/microduplications and one was a 6p22.3p21.31 LOH (Table 3).

Table 3

SNP array analysis of foetuses with thickened nuchal translucency in normal karyotyping
Case	SNP-array results	Size (Mb)	Inheritance	Pathogenicity classification	Postnatal outcome
1	arr[GRCh37]6p24.3p24.2(10,494,273 − 10,839,759)x1	0.38	De novo	VOUS	TD
2	arr[GRCh37]2q21.1(130,874,614_131,244,620)x1 arr[GRCh37]15q11.2(22,770,422_23,277,436)x1	0.3 0.5	De novo	VOUS	TP
3	arr[GRCh37]7q31.32(122,066,578_123,066,288)x3	0.98	De novo	VOUS	TD
4	arr[GRCh37]21q22.12q22.13(37,223,669 − 38,330,604)x3	1.1	De novo	VOUS	TD
5	arr[GRCh37]18q12.3(42,111,540 − 43,387,120)x3	1.2	De novo	VOUS	TD
6	arr[GRCh37]6p22.3p21.31(23,972,905 − 35,703,669)x2hmz	11.7	De novo	VOUS	TD

SNP, single nucleotide polymorphism; TD, term delivery; hmz, loss of heterozygosity.

Chromosomal aneuploidy and thickened NT

The average maternal age, NT thickness, and CRL for 10 foetuses with aneuploidies were 33.5 years (range: 21–42 years), 3.81 mm (range: 2.9–7.9 mm), and 6.4 mm, respectively (Table 4). Among these, 80% foetuses had Down syndrome. The detection rate for foetal aneuploidy was significantly associated with thickened NT as analysed by an ROC curve with 0.729 AUC (95% CI: 0.56–0.898, P = 0.019). The highest cutoff value of NT for predicting foetal aneuploidies was 4.15 mm, with 50% sensitivity and 90% specificity for detecting aneuploidies (Fig. 1).

Pregnancy outcomes and clinical follow-up

We obtained delivery information concerning 90 pregnancies in women with thickened NT, including 12 cases of pregnancy termination (13.33%; 12/90); among them 10 foetuses had aneuploidies and 2 had abnormal CNVs. Karyotyping and SNP array analysis showed 10 foetuses with aneuploidies, including trisomy 18 syndrome (n = 2) and Down syndrome (n = 8). Down syndrome was the most common type of chromosomal aneuploidy in the foetuses with thickened NT (80%, 8/10). Therefore, NT thickening, particularly > 4 mm NT, and Down syndrome are significantly associated, and the incidence of chromosomal aneuploidy was 33.3%. In addition, SNP array analysis displayed a higher detection rate for chromosomal abnormalities than karyotyping. This study showed that SNP array analysis is an important supplement to karyotyping for foetuses with thickened NT.

Table 4

Maternal and foetal information of 10 foetuses with aneuploidy and thickened nuchal translucency
Case	Maternal age (years)	NT (mm)	CRL (mm)	Aneuploidy	Gestational age (weeks)
A132	35	3.6	6.27	Trisomy 21	12^+ 5
A287	37	5.3	5.5	Trisomy 21	12^+ 1
A291	36	3.0	5.6	Trisomy 21	12^+ 1
A384	42	2.9	6.9	Trisomy 21	13
A456	35	0.6	7.7	Trisomy 21	13^+ 6
A510	41	4.3	6.85	Trisomy 21	13^+ 3
A653	21	3.3	7.1	Trisomy 21	13^+ 2
A705	25	4.2	7.5	Trisomy 21	13^+ 4
A671	28	3.0	5.2	Trisomy 18	12
B202	35	7.9	5.4	Trisomy 18	12

NT: Nuchal translucency; CRL: Crown rump length.

Molecular diagnostic techniques, such as CMA and SNV-seq, increase pCNV detection rate by 4–7% compared with standard karyotyping in patients with thickened NT[16, 17]. CMA is a high-resolution method for genome-wide detection and is recommended as the primary test for prenatal diagnosis[18, 19]. This method can be used to detect the CNVs of chromosomal imbalances such as aneuploidy and unbalanced rearrangements, particularly chromosome microdeletions and duplications[12]. With a normal karyotype, CMA can detect approximately 1% clinically significant microdeletions/microduplications in structurally normal pregnancies and 6% structural anomalies in prenatal diagnostic cases[20]. CMA includes microarray-based comparative genomic hybridisation (aCGH) and SNP arrays. SNP array analysis would detect uniparental disomy (UPD), LOH, and low-level mosaicism[21, 22]. Since the SNP array detects low-level mosaicism, microdeletions/microduplications, LOH, and UPD at the submicroscopic level, it has replaced standard karyotyping as the first-tier test for prenatal diagnosis in some countries[23]. Meanwhile, the American Congress of Obstetricians and Gynecologists (ACOG) recommended CMA as a first-tier prenatal diagnostic test for evaluating the foetuses with one or more major structural abnormalities in 2016[20, 24]. In this study, we performed whole-genome scanning using an SNP array and routine karyotyping of 90 foetuses from women with thickened NT. We attempted to elucidate the chromosomal abnormalities and CNVs to explore the clinical value of applying an SNP array and the cutoff values of NT in pregnant women with thickened NT.

In this study, standard karyotyping of amniotic fluid from 90 women pregnant with foetuses having thickened NT detected 10, 4, and 2 foetuses with chromosomal aneuploidies, structural abnormalities, and normal variation, respectively. SNP array analysis additionally detected six foetuses with abnormal CNVs. The details of the five foetuses with pCNVs are as follows:

Two foetuses had chromosomal microdeletions involving 6p24.3p24.2, 2q21.1, and 15q11.2. Foetus 1 had a 0.38 Mb deletion in the 6p24.3p24.2 region containing four OMIM genes, including GCNT2 and MAK. GCNT2 mutations are related to autosomal dominant blood group (Ii) diseases[25]. MAK homozygous mutation is associated with autosomal recessive Retinitispigmentosa62 disease. The clinical phenotypes include night blindness, retinitis pigmentosa, and pale optic disc[26, 27]. Deletion of a fragment smaller than this fragment is associated with global developmental delay, facial hypotonia, short limbs, prematurity, and short stature. However, the ClinGen database does not provide sufficient evidence for the haplotype dose sensitivity of the genes present in this fragment.

Foetus 2 had 0.3 and 0.5 Mb microdeletions in the 2q21.1 and 15q11.2 regions, respectively, which contained CCDC115 and SMPD4, respectively. Homozygous or compound heterozygous mutations in SMPD4 are associated with autosomal recessive neurodevelopmental disorders with microcephaly, arthrogryposis, and structural brain anomalies. The clinical phenotypes include global developmental delay, hypotonia, impaired mental development, aphasia, progressive microcephaly, and distal skeletal abnormalities. Homozygous or compound heterozygous mutations in CCDC115 are associated with autosomal recessive congenital disorders of glycosylation type II0[28]. The clinical phenotypes include progressive liver failure in infancy, hypotonia, and developmental delay[29, 30]. Previous studies have detected CCDC115 mutation and deletion abnormalities in patients with congenital glycosylation disorders[31]. In this case, the foetus was diagnosed with SMPD4 and CCDC115 deletions. The 0.5 Mb 15q11.2 deletion, deleted TUBGCP5, NIPA1, and BP1–BP2 within the 15q11.2 region. Heterozygous mutations in NIPA1 are associated with autosomal dominant spastic truncations spastic paraplegia 6[32, 33]. A previous study estimated the penetrance of this fragment deletion to be approximately 10.4%, suggesting an incomplete penetrance or expressivity difference[34]. The ClinGen database did not show haplotype dose sensitivity for genes contained in this fragment. Therefore, the clinical significance of these deletions remains unclear. The woman did not accept the risk of uncertainty and decided to terminate the pregnancy at 23^+ 3 weeks.

SNP array analysis detected three foetuses with chromosomal microduplications in the 7q31.32, 21q22.12q22.13, and 18q12.3 regions. Foetus 3 had a 0.98 Mb deletion in the 7q31.32 region, which contains three OMIM genes, including TAS2R16. Mutations in these genes are associated with autosomal dominant β-glycopyranoside-tasting disease with clinical manifestations including β-glycopyranoside taste sensitivity[35]. Evidence from the ClinGen database regarding the three-fold dose sensitivity of the genes contained in this fragment is not clear. Among the TAS2R16 duplication cases in the ClinVar database, all pathogenic/likely pathogenic cases had point mutations or larger duplications. However, its clinical significance remains unclear. Moreover, foetus 4 had a 1.1 Mb microduplication in the 21q22.12q22.13 region, which contains CLDN14, HLCS, and eight other genes. CLDN14 homozygous mutations relate to autosomal recessive hereditary Deafness 29 disease, with the clinical phenotypes including sensorineural hearing loss[36]. HLCS compound homozygous or heterozygous mutations are closely related to autosomal recessive all-carboxylase lack synthetase disease, with clinical phenotypes including shortness of breath, feeding problems, hypotonia, and metabolic acidosis. Therefore, the clinical significance of these micro-duplicates remains unclear.

Foetus 5 had a 1.2 Mb microduplication in the 18q12.3 region, which contains SETBP1 and three other OMIM genes. SETBP1 mutation is associated with mental retardation, autosomal dominant 29 (MRD29), and the clinical phenotype includes short head, pointed chin, ptosis, epilepsy, intellectual disability, and attention deficit hyperactivity disorder[37, 38]. Evidence from the ClinGen database regarding the three-fold dose sensitivity of the genes contained in this fragment is not clear. Moreover, SNP array analysis revealed an 11.7Mb LOH on 6p22.3p21.31 in foetus 6. 6p22.3p21.31 contains VARS2, SKIV2L, and 39 other genes related to autosomal recessive diseases[39, 40]. Studies have reported clinical phenotypes including foetal developmental delays, hyperglycaemia, and dehydration[41]. However, whether the LOH region in this case carries imprinting-related genes is not clear.

In this study, we found that foetuses with Down syndrome had the highest proportion of aneuploidies (80%) among foetuses with thick NT (≥ 2.5 mm). Indications such as age, serological screening, and thick NT would detect 80–90% foetuses with Down syndrome in the first trimester of prenatal screening[5]. The diagnostic yield is approximately 6% and 49% for positive serum screening in foetuses with structural anomalies and ultrasound abnormalities, respectively, in the first trimester[20]. We also determined that thickened NT is a high-risk factor for autosomal aneuploidy and is closely related to CNVs such as microduplication/microdeletion and LOH.

In foetuses with NT > 2.5 mm, the probability of aneuploidy increased with increasing NT thickness, which was 3.7% (1/27), 8.1% (3/37), 10% (1/10), and 31.3% (5/16) for the foetuses with 2.5–2.9, 3.0–3.4, 3.5–3.9, and > 4.0 mm NT, respectively. The cutoff value was 4.15 mm, at which the probability of aneuploidy was 46.2% (6/13) and the specificity was up to 90% by ROC curve analysis. Therefore, karyotyping and SNP array analysis should be performed for prenatal diagnosis in all foetuses with ≥ 2.5 mm NT to avoid missing the formation of abnormal foetus.

Thickened NT is a major risk factor for aneuploidy and abnormal CNVs. Thus, a clear understanding of the incidence of aneuploidy and abnormal CNVs associated with NT thickening may help develop intervention strategies and improve pregnancy outcomes. In this study, we revealed aneuploidies and characteristics of CNVs, including microduplications/microdeletions and LOH, in pregnant women with thickened NT. We recommend that karyotyping and SNP array analysis should be used for prenatal diagnosis in all foetuses with ≥ 2.5 mm NT to avoid the birth of a child with genetic defects due to missed prenatal diagnosis.

Ethics approval and consent to participate

This study was approved by the Ethics Committee of Ningde Municipal Hospital affiliated with the Ningde Normal University(Approval No. NSYKYLL-2023-028). All procedures were applied following the Declaration of Helsinki, as well as international and national laws, guidelines and regulations. Signed informed consent was obtained from all the pregnant women enrolled in this study.

Consent for publication

All authors consented to publish.

Availability of data and materials

All data generated during and/or analyzed during the current study are available upon request by contact the corresponding author.

Competing interests

The authors declare no conflicts of interest.

Funding

This study was supported by the Ningde Natural Science Foundation (2022J06) and Experimental Diagnostic Technology Innovation Team of Birth Defects and Genetic Diseases at Ningde Normal University (2021T10).

Authors' contributions

CL wrote the manuscript. WQ, ZX, HX, LP and CX were responsible for clinical diagnosis and

treatment. DW managed the study. FX designed the study. LJ and FX revised the manuscript. ZX

collected the data. All the authors contributed to the manuscript and approved the final submitted

version.

Acknowledgements

We thank all patients who participated in this study.

Authors' information

1Department of Central Laboratory, Ningde Municipal Hospital Affilliated to Ningde Normal University, Ningde,Fujian,PR China.2Department of Ultrasound, Ningde Municipal Hospital Affilliated to Ningde Normal University, Ningde,Fujian, PR China.3Department of Obstetrics, Ningde Municipal Hospital Affilliated to Ningde Normal University, Ningde,Fujian, PR China.

4School of Clinical Medicine, Fujian Medical University,Fuzhou,Fujian, PR China.5Public Health School of Fujian Medical University,Fuzhou,Fujian, PR China

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No competing interests reported.

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The Role of Thickened Nuchal Translucency on Copy Number Variations and Pregnancy Outcomes in Northeast Coast of Fujian Province, China: A Comparison Between Traditional Karyotyping and Single Nucleotide Polymorphism Array Analysis

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Abstract

Background

Methods

Results

Conclusion

Figures

Background

Methods

Patient data

Cytogenetic analysis

CMA (SNP array)

Receiver operating characteristic curve for foetal aneuploidy

Statistical analysis

Results

Cytogenetic analysis of foetuses with thickened NT

SNP array analysis results

Chromosomal aneuploidy and thickened NT

Pregnancy outcomes and clinical follow-up

Discussion

Conclusions

Declarations

References

Additional Declarations

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