| Maternal serological prenatal screening is to detect specific serum markers in pregnant women’s peripheral blood serum and combine with the pregnant women’s age,gestational age and weight data to infer whether the fetus is at risk of chromosomal abnormalities.It is mainly used for screening for trisomy 21/18 syndrome and open neural tube defects,referred to as Down’s screening for short.Down’s screening is currently the first-line screening method used in China.Scholars at home and abroad had a large number of studies on Down’s screening.In recent years,studies had shown that Down’s screening can also indicate non-21/18 trisomy chromosomal abnormalities,but no consensus had been reached due to lack of datas.Non-21/18 trisomy chromosomal abnormalities includs autosomal trisomy syndrome except to trisomy 21/18,sex chromosome abnormalities(SCA)and chromosome copy number variation(CNVs).Follow-up prenatal diagnosis in people at high risk and critical risk of Down’s screening is dominated by chromosomal karyotype testing,but it is also recognized that there is a risk of missing fetal CNVs if only chromosomal karyotype testing is performed.Chromosome microarray analysis(CMA)or Low depth genome-wide copy number variation test(CNVseq)can be used to detect these diseases.At present,genetic counseling for pregnant women with high risk of Down’s screening will also inform them of the relevant risks,and CMA or CNVseq testing is recommended.However,specific data of non-21/18 trisomy chromosome abnormalities in the fetus in this population cannot be provided.ObjectiveThe detection of various fetal chromosomal abnormalities in pregnant women with high or critical risk of Down’s screening was statistically analyzed,especially the incidence of non-21/18 trisomy chromosomal abnormalities,by analyzing the results of CNVseq or CMA validation,so as to provide data support for the selection of subsequent prenatal diagnosis techniques for pregnant women with high risk and critical risk of Down screening.Materials and methods1 Study subjectsA total of 3028 pregnant women,who were admitted to the Third Affiliated Hospital of Zhengzhou University and required CMA or CNVseq for amniotic fluid examination due to high risk or critical risk of Down’s screening were selected.According to the risk results of Down’s screening,they were divided into high risk group(2555 cases)and critical risk group(473 cases).All the cases were spontaneous conception and single pregnancy,and no fetal ultrasound abnormality was suggested before prenatal diagnosis.Factors that might affect maternal serum markers were excluded,such as genetic history and family history of chromosomal abnormalities,prepregnancy underlying diseases(such as hypertension,diabetes,blood diseases,liver and kidney insufficiency,and malignant tumors).2 Methods2.1 Collection of amniotic fluid specimensAfter the result of the pregnant woman’s blood routine,blood coagulation,four infectious disease,electrocardiogram examination and routine ultrasound examination of the fetus were normal and the informed consent was signed,the amniocentesis was performed with the assistance of B-ultrasound,and 8-15 ml of amniotic fluid was extracted under sterile conditions.Avoid blood contamination when collecting samples.Store the samples in a refrigerator at 4℃ for later use.2.2 Extraction and detection of genomic DNAThe maternal blood contamination was identified by STR site comparison method.After the maternal blood contamination was confirmed,the Cytoscan 750K chip from Affymatrix Company was used for CMA detection.Enzyme digestion,PCR,labeling,hybridization chip scanning and other operations were performed according to the chip instructions.Genomic DNA from amniotic fluid was extracted automatically by magnetic bead method.After extraction,50ng DNA were used to construct DNA library.A large-scale parallel sequencing platform,NextSeq CN500,was used to detect low depth copy number variation in whole-genome DNA..2.3 The data analysis of CNVseq or CMASequencing data were collated and compared with the HG19 genome sequence to find out the Copy Number Variants(CNVs)on the sample genome.The pathogenicity of CNVs was analyzed by referring to databases such as DGV(http://projiects.tcag.ca/variation),DECIPHER(http://decipher.sanger.ac.uk/),,ISCA(ht tps://www.iscaconsortium.org/),Genereviews(https://www.generreviews.org/),Clingen(https://dosage.clinicalgenome.org/),and OMIM(http://www.omim.org).According to ACMG standards in 2020,the results were divided into five grades:pathogenic,likely pathogenic,uncertain significance,likely benign and benign,but the CNVs included in the study excluded benign CNVs.3 Statistical analysisSPSS 23.0 was used for statistical analysis.Quantitative data conforming to the normal distribution were represented.Two independent samples t-test was used for comparison between groups.Qualitative data were represented by number of cases and percentage,χ2 test was used for comparison between groups.α=0.05 was used as the test level.Results1 Clinical features of studied subjectsA total of 3,028 pregnant women aged were included in the study,including 2555 pregnant women in high-risk group,accounting for 84.38%;The critical risk group was 473 cases,accounting for 15.62%.There was no significant difference in the age and gestational age between the two groups(P>0.05).2 Analysis of fetal chromosomal abnormal results in Down’s screening of pregnant women at high or critical risk(1)A total of 107 cases of pathogenic or likely pathogenic fetal chromosomal abnormalities were found in 3028 pregnant women who underwent CMA or CNVseq for high risk or critical risk of Down’s screening,including 95 cases in the high risk group,including 37 cases of CNVS,5 cases of sex chromosomal abnormalities,1 case of trisomy 16 syndrome,and 52 cases of trisomy 21/18 syndrome.There were 12 cases in the critical risk group,including 7 cases of CNVs,2 cases of sex chromosome abnormalities,and 3 cases of trisomy 21/18 syndrome.(2)In the high-risk group,the overall detection rate for pathogenic or likely pathogenic chromosomal abnormality was 3.72%,for non-21/18 trisomy abnormality was 1.68%,for CNVs was 1.45%,and for sex chromosome abnormality was 0.20%;In the critical risk group,the overall detection rate of pathogenic or likely pathogenic chromosomal abnormality was 2.54%,the detection rate of non-21/18 trisomy chromosome abnormality was 1.90%,the detection rate of CNVS was 1.48%,and the detection rate of sex chromosome abnormality was 0.42%.Chi-square test of the two groups showed that there was no significant difference in the detection rate of above chromosomal abnormalities between the high-risk group and the critical risk group(P>0.05).(3)The detection rate of trisomy 21/18 in the high-risk group was 2.04%,and that of the critical risk group was 0.63%.Chi-square test of the two groups indicated that the detection rate of trisomy 21/18 in the high-risk group was significantly higher than that in the critical risk group,with statistical significance(P<0.05).(4)According to the ACMG standard in 2019,pathogenic CNVs accounted for 26.42%(28/106)of the 106 cases,likely pathogenic CNVs accounted for 15.09%(16/106),and CNVs with unclear clinical significance accounted for 54.71%(58/106).Among pathogenic CNVs,there are currently 22 cases of pathogenic syndrome,and one of them contains two pathogenic syndromes,while the remaining 6 cases involve segments that do not include known pathogenic syndrome.3 Comparison of the detection rates of 21/18 trisomy syndrome and pathogenic or prepathogenic CNVsThe detection rate of fetal pathogenic or likely pathogenic CNVs was 1.45%(44/3028)in all pregnant women enrolled in the study.The detection rate of trisomy 21 of 18 was 1.82%(55/3028).Chi-square test showed that there was no significant difference between the detection rate of Down’s screening for trisomy 21/18 syndrome and that of pathogenic or likely pathogenic CNVs(P>0.05).ConclusionThe detection rate of non-trisomy 21/18 chromosome abnormality was basically consistent with that of trisomy 21/18 syndrome in populations at the high risk or critical risk of Down’s screening.Therefore,the risk of various chromosomal abnormalities should be informed when genetic counseling is conducted for this part of the population,and it is suggested that CMA or CNVseq technology should be included as detection technology in subsequent prenatal diagnosis,so as to avoid the birth of fetuses with non-21/18 trisomy chromosomal abnormalities. |
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