新生儿高通量测序基因筛查的前瞻性研究

A prospective study of genetic screening of 2060 neonates by high-throughput sequencing

目的探索新生儿高通量测序(HTS)基因筛查在新生儿筛查疾病早期诊断中的临床价值。方法选取2021年3月至2021年9月于宁波市妇女儿童医院出生的2060例新生儿为研究对象。对其采用传统串联质谱和荧光免疫分析筛查后,再应用HTS技术对135个筛查疾病相关致病基因的明确致病高频变异位点进行检测,可疑变异位点进行Sanger测序或多重连接探针扩增技术家系验证。结果2060例新生儿中,基因阳性31例,基因携带557例,基因阴性1472例。31例基因阳性新生儿中,G6PD基因变异5例,遗传性非综合征型耳聋相关基因(GJB2、GJB3、MT-RNR1)变异19例,PAH基因变异2例,GAA基因变异1例,SMN1基因变异1例,MTTL1基因变异2例,GH1基因变异1例。结合基因结果和临床资料确诊脊髓性肌肉萎缩症(SMA)1例,糖原贮积症Ⅱ型1例,耳聋2例,G6PD缺乏症5例,额外诊断患儿母亲SMA 1例。2060例新生儿经传统串联质谱检测未发现确诊患者;经传统荧光免疫分析筛查发现G6PD缺乏症5例(基因筛查阳性),甲状腺功能减退症2例(DUOX2基因变异携带者)。宁波地区新生儿基因变异携带率前6位的基因分别为:DUOX2(3.93%)、ATP7B(2.48%)、SLC26A4(2.38%)、GJB2(2.33%)、PAH(2.09%)、SLC22A5(2.09%)。结论新生儿基因筛查技术的疾病检测范围广泛,检出率高,与传统筛查联合可显著提高临床筛查效果,有助于患儿的二级预防、家系成员的疾病诊断和变异携带者的遗传咨询。

Objective To assess the value of genetic screening by high-throughput sequencing(HTS)for the early diagnosis of neonatal diseases.Methods A total of 2060 neonates born at Ningbo Women and Children′s Hospital from March to September 2021 were selected as the study subjects.All neonates had undergone conventional tandem mass spectrometry metabolite analysis and fluorescent immunoassay analysis.HTS was carried out to detect the definite pathogenic variant sites with high-frequency of 135 disease-related genes.Candidate variants were verified by Sanger sequencing or multiplex ligation-dependent probe amplification(MLPA).Results Among the 2060 newborns,31 were diagnosed with genetic diseases,557 were found to be carriers,and 1472 were negative.Among the 31 neonates,5 had G6PD,19 had hereditary non-syndromic deafness due to variants of GJB2,GJB3 and MT-RNR1 genes,2 had PAH gene variants,1 had GAA gene variants,1 had SMN1 gene variants,2 had MTTL1 gene variants,and 1 had GH1 gene variants.Clinically,1 child had Spinal muscular atrophy(SMA),1 had Glycogen storage diseaseⅡ,2 had congenital deafness,and 5 had G6PD deficiency.One mother was diagnosed with SMA.No patient was detected by conventional tandem mass spectrometry.Conventional fluorescence immunoassay had revealed 5 cases of G6PD deficiency(all positive by genetic screening)and 2 cases of hypothyroidism(identified as carriers).The most common variants identified in this region have involved DUOX2(3.93%),ATP7B(2.48%),SLC26A4(2.38%),GJB2(2.33%),PAH(2.09%)and SLC22A5 genes(2.09%).Conclusion Neonatal genetic screening has a wide range of detection and high detection rate,which can significantly improve the efficacy of newborn screening when combined with conventional screening and facilitate secondary prevention for the affected children,diagnosis of family members and genetic counseling for the carriers.