Application Of Single Nucleotide Polymorphism Microarray Technology In Fetal Genetic Diagnosis Of Ultrasound Abnormalities

Objective To analyze the results of single nucleotide polymorphism array(SNP array)detection of ultrasound abnormal fetal amniotic fluid or cord blood,and to explore the application value of SNP array in fetal genetic diagnosis of ultrasound abnormalities.Materials and Methods 1.We collected 1437 fetal samples with prenatal ultrasound abnormalities which accepted invasive prenatal diagnosis after informed consent for SNP array technique and karyotype analysis from Fujian Provincial Maternity and Children’s Hospital during May 2015 to December 2018.The age of gravida is 16 to 45 years old,and the gestational age is 17 to 35+6 weeks.According to the ultrasound performance,they are divided into ultrasound structural abnormalities and non-structural abnormalities.According to the number of systems involved in ultrasonic structural anomalies and whether non-structural anomalies are combined,they are divided into 4 groups: single system structural anomalies,multiple system structural anomalies,single system structural anomalies combined with non-structural anomalies,multiply system structural anomalies combined with non-structural anomalies.According to the anatomical system of abnormal ultrasound structure,they are divided into 8 groups,including structural abnormalities of nervous system,structural abnormalities of head and face,structural abnormalities ofrespiratory system,structural abnormalitiesof cardiovascular system,structural abnormalities of digestive system,structural abnormalities of urinary system,structural abnormalities of skeletal system and other structural abnormalities(Fetal edema,abnormal tumor and abdominal wall structure).Non-structural abnormalities include ultrasound soft marker,fetal growth restriction and amniotic fluid abnormalities.According to the number of ultrasound non-structural abnormalities,they are divided into three groups: one non-structural abnormalities,two non-structural abnormalities, and≥3 non-structural abnormalities.2.All samples were subjected to chromosome microarray analysis and karyotype detection according to the standard laboratory procedures.To analyze the results of SNP array,database were used to evaluate the pathogenicity of CNV.3.Combined with family genetic analysis and SNP array results which were tested from peripheral blood of fetal parents to make clear the property and source of CNV in variants of uncertain clinical significance(VOUS).4.Using SPSS19.0 software for statistical analysis,analysis of chi-square was applied in the interblock analysis.Only if the P < 0.05,the difference was statistically significant.Results 1.All the 1437 samples were successfully tested for SNP array.1295 cases were negative(90.1%,1295/1437)and 142 cases were positive(9.9%,142/1437).The positive results included 87 cases with pathogenic CNV(6.1%,87/1437)and 55 cases with VOUS(3.8%,55/1437).Pathogenic CNV included 34 cases of aneuploidies(39.1%,34/87),1 case of triploid(1.1%,1/87),45 cases of deletion/repeat,which included 32 cases of micro-deletion/micro-repeat syndrome(36.8%,32/87)and 13 cases of deletion/repeat which were fall short of micro-deletion/micro-repeat syndrome(14.9%,13/87),5 cases of chimera(5.7%,5/87),2 cases of pathogenic single parent diploid(UPD)(2.3%,2 /87).2.The all 1437 samples were performed according to karyotype analysis,three cases were failed to incubate.There were 1343 cases(93.7%,1343/1434)of normal karyotype and 91 cases(6.3%,91/1434)of abnormal karyotype.Among the 91 cases of abnormal karyotype,48 cases(52.7%,48/91)were abnormal in chromosome number,including 47 cases of aneuploidy,which including 17 cases of trisomy 21(36.2%,17/47),and 8 cases of trisomy 18(17.0%,8/47),1 case of trisomy 13(2.1%,1/47),6 cases of sex chromosome abnormalities(12.8%,6/47),5 cases of extra chromosomes(10.6%,5/47)and 10 cases of the number of chimeric types was abnormal(21.1%,10/47).43 cases were abnormal in chromosome structure(47.3%,43/91),including 30 cases of balanced translocation/inversion(69.8%,30/43),4 cases of chimeric structural abnormalities(9.3%,4/43)and 9 cases(20.9%,9/43)of other abnormalities(including partial deletion and partial repeat of chromosomes).3.142(9.9%,142/1437)cases were detected positive in SNP array.91 cases(6.3%,91/1434)were detected positive in karyotype analysis.The SNP array detection rate were increased by 3.6% compared with the karyotype analysis.Among all the 52 cases of pathogenic CNV except aneuploid and triploid,29 cases were not detected in karyotype analysis.SNP array increased the detection rate of pathogenic CNV by 2.0%(29/1437).4.Among the 805 cases of structural abnormalities,716 cases were negative(88.9%,716/805)and 89 cases were positive(11.1%,89/805).The positive results included 55 cases with pathogenic CNV(6.8%,55/807)and 34 cases with VOUS(4.2%,34/807).13.7% of pathogenic CNVwere detected in the multiply system structural anomalies combined with non-structural anomalies,which had the highest detection rate.The following detection rate is the single system structural anomalies combined with non-structural anomalies,multiple system structural anomalies and singlesystem structural anomalies,which were 8.8%,7.7% and 4.2%,the difference was statistically significant(P<0.05).According to the anatomical system where the structural abnormalities occurred,the detection rate of pathogenic CNV was 12.2% of urinary system,10.3% of skeletal system,8.6% of nervous system,7.3% of cardiovascular system,and 5.0% of other structural abnormalities.No pathogenic CNV was detected in structural abnormalities of head and face,structural abnormalities of respiratory system and structural abnormalities of digestive system.5.53 cases(8.4%,53/632)were positive and 579 cases(91.6%,579/632)were negative among the 632 cases of non-structural abnormalities using SNP array.The positive results of 53 cases included 32 cases of pathogenic CNV(5.1%,32/632)and 21 cases of VOUS(3.3%,21/632).The detection rates of pathogenic CNV in onenon-structural abnormalities,twonon-structural abnormalities,and≥3 non-structural abnormalities were 4.6%,4.8%,and 16.7%.There was a significant tendency of increasing from group 1 to group 3(P<0.05).In one non-structural abnormalities,the detection rate of nasal bone dysplasia,FGR,NT thickening,NF thickening,choroid plexus cyst,lateral ventricle widening,intestinal echo enhancement,and fetal growth dysplasia were 8.1%,7.0%,5.6%,4.8%,4.5%,3.3%,3.1%,and 2.7%.No Pathogenic CNV was detected in mild renal pelvis separation,echogenic foci in the heart,abnormal amniotic fluid volume,single umbilical artery,subclavian artery vagus,and venous catheter blood flow spectrum abnormalities.6.As the age of pregnant women increased,the detection rate of chromosome aneuploidy increased,the difference was statistically significant(P<0.05).However,there was no significant correlation between the detection rate of pathogenic CNV and VOUS with the age of pregnant women.7.A total of 55 cases of VOUS were detected in 1437 samples,and the detection rate was 3.8%.Among them,29 cases were traced to parents validation(52.7%)and 26 cases were rejected by parents validation(47.3%).There are 19 cases(34.5%)were likely to be benign,17 cases(30.9%)may be pathogenic,16 cases(29.1%)were unclear clinical significance,and 3 cases were LOH(5.5%)according to the ACMG guidelines.Conclusion 1.Compared with the traditional karyotype analysis,SNP array can significantly improve the detection rate of genetic abnormalities in prenatal diagnosis of ultrasound abnormalities.With the increase number of abnormal ultrasound items,the risk of chromosome copy number variation is also on the rise.Therefore,the application of SNP array technology in fetal detection of prenatal ultrasound abnormalities should be further promoted.2.As the age of pregnant women increased,the detection rate of chromosome aneuploidy increased.However,there was no significant correlation between the detection rate of pathogenic CNV and VOUS with the age of pregnant women.3.In order to define the quantities and origins of CNV,parents’ genetic tests are suggests for the variants of uncertain clinical significance.In the meantime,follow the fetal pregnancy outcome to provide assistance for future genetic counseling.