Application Of Next Generation Sequencing In The Etiological Diagnosis Of Intellectual Disability/Global Developmental Delay

ObjectiveTo explore the genetic etiology and the diagnostic strategy of unexplained Intellectual Disability /Global Developmental Delay(ID/GDD)through Copy Number Variations Sequencing(CNV-seq)and trio Whole Exome Sequencing(trio-WES)based on next generation sequencing,and to evaluate the value of combined testing.Methods:The peripheral blood samples of 65 children with unexplained ID/GDD in the outpatient clinic of Fujian Maternal and Child Health Hospital from June 2016 to December 2019 were collected and sequenced on the Illumina Nova Seq6000(American Illumina Company)sequencer.The raw data were compared with the CRCh37/Hg19 reference genome and compared with the database.The American College of Medical Genetics and Genomics(ACMG)scoring system was used to analyze the pathogenicity of Copy Number Variations(CNVs).The correlation between phenotypes and CNVs detection rate was analyzed by the FISH exact probability method,and the difference of CNVs detection rate among different Developmental Quotient(DQ)/ Intelligence Quotient(IQ)groups was analyzed by the chi-square test.The gene variation sites were detected with trio-WES in negative samples.The pathogenicity was determined by the ACMG Standards and guidelines for the interpretation of sequence variants and the three-factor classification system and verified by sanger sequencing.The characteristics of pathogenic Single Nucleotide Variation(SNV)were analyzed by descriptive analysis,and the difference of clinical phenotype between CNVs positive children and SNV positive children was analyzed by FISH exact probability method.Mc Nemar’s test was used to compare the positive rate between CNV-seq alone and CNV-seq combined with trio-WES Results:A total of 29 cases of positive results were detected in 65 children,of which 22 cases were positive CNVs(22/65,33.8%),of which 23.1% were pathogenic CNVs(15/65),including 2 cases of non-syndromic or undefined pathogenic variants in the database,four cases(4/65,6.2%)of likely pathogenic CNVs,three cases(3/65,4.6%)of Variants of uncertain significance,involving 11 known syndromes.The proportion of CNVs on chromosomes 7,10,14,15,22,and X was higher.Among the 65 cases,there were 12 cases of facial abnormality.Eight cases(8/22,36.4%)in the CNVs positive group;Four cases(4/43,9.3%)in the CNVs negative group and there were significant differences between the two groups.There are 12 cases with the organ deformity in 65 patients,seven cases(7/22,31.8%)in the CNVs positive group,and 5 cases(5/43,11.6%)in the negative group and there was a significant difference between the two groups.There were significant differences in the detection rate of CNVs among different DQ groups.Seven cases were SNV positive,three cases were Autosomal dominant(AD)inheritance,all were de novel variants;Four cases were Autosomal recessive(AR)inheritance,of which 3 cases were compound heterozygous variants and1 case was a homozygous variant.There was no significant difference in clinical phenotype between CNVs positive group and SNV positive group.There was a significant difference in the positive rate of CNV-seq combined tri-WES compared with that of CNV-seq alone.Conclusion:The combined detection with CNV-seq and trio-WES may increase the detection yield,which provides a new way to identify the genetic etiology of ID/GDD.It is a detection strategy worth popularizing at this stage.The high incidence of chromosomal CNVs in the Chinese ID/GDD population may be different from that abroad.The positive rate of CNVs was higher in children with facial abnormalities,organ deformities,and moderate and severe ID/GDD.