高危新生儿听力和聋病易感基因联合筛查临床研究

目的探讨高危新生儿听力和聋病易感基因联合筛查的临床意义。方法选择2012年5月至2014年2月入住南方医科大学附属佛山市妇幼保健院新生儿重症监护室的920例具有听力损失高危因素的新生儿为研究组,选取同期产后区938例健康新生儿为对照组,于生后检测GJB2基因35del G、176-191del16、235del C、299-300del AT;SLC26A4基因IVS7-2AG、2168AG;线粒体12Sr RNA基因1494CT、1555AG等3个基因8个突变位点;听力筛查复筛未通过者于3月龄行听力学诊断。结果研究组920例检出35例聋病基因携带,3个基因突变的总体携带率3.8%;检出听力障碍34例(3.7%)、其中重度以上听力障碍15例(1.6%);30例(85.7%)聋病基因携带者通过了听力筛查。对照组938例检出21例聋病基因携带,3个基因突变的总体携带率2.2%;检出听力障碍4例(0.4%)、重度以上听力障碍1例(0.1%);17例(1.8%)聋病基因携带者通过了听力筛查。研究组听力损失和重度以上听力损失检出率、聋病基因突变的总携带率及聋病基因携带者的听力筛查通过率与对照组比较差异均有显着性(均P0.05)。结论高危新生儿听力障碍的检出率和聋病基因突变携带率均高于正常新生儿;采用听力和聋病易感基因联合筛查能及时发现常规通过听力筛查但具有耳聋高危因素和迟发性聋病遗传因素的新生儿,对早期干预、定期随访、减少聋病发生具有指导意义。     

西北地区751例新生儿耳聋基因突变筛查

目的初步了解西北地区新生儿常见耳聋基因突变类型和携带率,探讨耳聋基因突变筛查对于辅助诊断和防治新生儿遗传性耳聋的临床价值。方法采集西北地区751例新生儿的足跟血,采用15项遗传性耳聋基因检测试剂盒(微阵列芯片法)对中国人群常见4种耳聋基因15个突变位点进行筛查。结果在751例新生儿中,检测到39例新生儿携带耳聋基因突变,总突变携带率为5.18%。其中GJB2基因突变19例,突变携带率为2.53%;SLC26A4基因突变16例,突变携带率为2.12%;线粒体12SrRNA 1555A>G均质突变4例,突变携带率为0.53%。新疆出生的新生儿279例中,检测到耳聋基因突变11例,突变携带率3.95%;甘肃省出生的新生儿277例中,检测到耳聋基因突变12例,突变携带率4.32%;陕西省出生的新生儿178例中,检测到耳聋基因突变12例,突变携带率6.74%;青海省出生的新生儿17例中,检测到耳聋基因突变4例,突变携带率23.52%。结论西北地区新生儿耳聋基因突变携带率偏高,但是GJB2基因235 del C位点的突变携带率偏低,可能与西北地区的地域特征和人口遗传学特点有关。新生儿耳聋基因筛查对于听力筛查具有很好的互补作用,可以从基因水平发现可能出现迟发性耳聋和药物敏感性耳聋的高危新生儿。     

新生儿重症监护病房新生儿耳聋基因致病变异携带者筛查的横断面调查

背景：我国耳聋发病率高与耳聋基因致病变异的携带率高有关,目前缺乏对NICU新生儿耳聋基因致病变异携带者的筛查数据。 目的：调查NICU新生儿中耳聋基因GJB2 和SLC26A4致病变异的携带率。 设计：横断面研究。 方法：纳入2016年1月至2021年12月在复旦大学附属儿科医院NICU住院、入院日龄≤28 d,且出院前完成高通量测序的新生儿,排除生后耳聋相关基因诊断阳性者。从病历系统中截取患儿的性别、胎龄、出生体重;从测序数据库中提取GJB2 基因和SLC26A4基因的检测结果、患儿人类表型标准用语信息。携带率（%）=杂合致病或可能致病（P/LP）变异例数/总研究对象人数。检索PubMed、Embase和万方数据库,纳入既往报道中国NICU人群、新生儿人群和孕妇人群中GJB2 基因和/或SLC26A4基因P/LP变异携带情况的文献,并行复习。 主要结局指标：GJB2 基因和SLC26A4基因的P/LP变异携带率。 结果：纳入14 924例新生儿,男8 587例（57.5%）,女6 337例,胎龄（35.6±3.7）周,出生体重（2 711.7±887.1）g。携带GJB2 基因P/LP变异的患儿2 009例（13.462%）,共检出18种杂合P/LP变异,其中c.109G>A最常见（10.902%）,其次为c.235del（1.749%）、c.299_300del（0.409%）、c.176_191del（0.154%）、c.508_511dup（0.074%）和c.257C>G（0.034%）。携带SLC26A4基因P/LP变异的患儿305例（2.044%）,共检出31种杂合P/LP变异,携带率最高的6种依次为c.919-2A>G（1.139%）、c.2168A>G（0.181%）、c.1226G>A（0.100%）、c.1229C>T（0.094%）、c.1174A>T（0.080%）和c.1003T>C（0.047%）。 结论：建议将GJB2 基因上的c.109G>A、c.508_511dup和c.257C>G以及SLC26A4基因的c.1003T>C位点纳入NICU新生儿耳聋基因致病变异携带者筛查。     

天津市新生儿听力及耳聋基因联合筛查结果分析

目的 分析天津市新生儿听力及耳聋基因联合筛查情况。方法 选择2018年1月1日—12月31日在天津市全部助产机构出生的新生儿作为研究对象，在知情告知自愿筛查的前提下进行新生儿听力及耳聋基因联合筛查。结果 102 570例新生儿中210例（0.2%）有不同程度听力障碍，其中双耳聋114例，单耳聋96例。72 866例进行耳聋基因筛查，检出耳聋基因变异3 924例（5.4%），其中单基因纯合变异（均质变异）146例，单基因复合杂合变异14例，多基因复合杂合变异68例，单基因杂合变异（异质变异）3 696例。GJB2、SLC26A4、GJB3及线粒体12SrRNA基因的变异检出率分别为4.8%、4.5%、0.4%和0.2%。进行听力学诊断的423例患儿中，听力障碍组耳聋基因变异率高于正常组；210例听力障碍患儿中，双耳聋组的耳聋基因变异率高于单耳聋组；114例双耳聋患儿中，重度极重度聋组的耳聋基因变异率高于轻中度聋组，差异均有统计学意义（P<0.05）。结论 常见耳聋基因筛查，为本地区医疗保健咨询提供参考，降低听力障碍发生率。     

Citrin 缺陷导致的新生儿肝内胆汁淤积症 SLC25A13 基因分析

目的：Citrin 缺陷导致的新生儿肝内胆汁淤积症（NICCD）是一种由SLC25A13基因突变引起的常染色体隐性遗传病,临床可表现为肝内胆汁淤积性黄疸、低出生体重、生长迟缓和低蛋白血症等。本研究通过DNA测序技术探讨中国NICCD患儿 SLC25A13 基因突变类型。方法：针对 SLC25A13 基因的 18 个外显子及其侧翼区碱基序列设计引物,应用 PCR 技术扩增目的片度。PCR 扩增、纯化后直接测序,确定其突变类型。IVS16ins3kb 突变则采用巢式 PCR 和 RT-PCR 进行检测。结果：发现7种SLC25A13基因突变类型,包括851del4、1638ins23、IVS16ins3kb、IVS6+5G>A、c.775C>T（p.Q259X)、c.1505C>T（p.P502L） 和 c.1311C>T（p.C437C）;并确认一种复合突变类型［1638ins23+IVS16ins3kb］。其中c.775C>T（p.Q259X）、c.1505C>T（p.P502L）和 c.1311C>T（p.C437C）为新发现的基因突变类型。在20例NICCD患儿中,6 例为 851del4 纯合突变,7 例为杂合突变,另有 7 例为单一突变类型的杂合子。突变类型以 851del4 为主,占所有突变类型64%;其次为 1638ins23、IVS16ins3kb 和 IVS6+5G>A（分别占15%、12% 和 6%）。结论：851del4突变在 NICCD 患儿中最为常见。     

GJB2基因及其诊断筛查技术研究进展

耳聋基因GJB2定位于13q11-q12,编码connexin26(CX26)蛋白.在常染色体隐形遗传的非综合征性耳聋中,有50%的患者存在着GJB2基因的突变,然而在不同种族中,GJB2基因的突变位点也是不同的.35delG是欧美人群主要的突变形式;突变位点167delT在犹太耳聋人群中多见;而在亚裔人群中,235delC突变占有极大的比例.由于GJB2基因在遗传性耳聋中的特殊地位,因此对于GJB2基因的诊断及筛查技术就显得尤为重要.在新生儿听力筛查基础上,融入耳聋易患基因分子水平筛查,在早期发现和干预先天性听力损失方面发挥着重要作用.     

Citrin缺陷所致新生儿肝内胆汁淤积症SLC25A13基因突变的分子诊断

目的探讨citrin缺陷导致的新生儿肝内胆汁淤积症(NICCD)患儿的SLC25A13基因突变情况。方法选取17例确诊NICCD患儿,应用PCR-RFLP方法检测其SLC25A13基因中8种中国人最常见的突变,并结合常规实验室检查结果进行分析。结果 17例患儿中,6例为SLC25A13基因纯合突变、3例为SLC25A13基因复合杂合突变,8例为SLC25A13基因单杂合突变。共检测出3种突变类型,分别为851del4(73.1%)、1638ins23(11.5%)和IVS6+5GA(15.4%)。17例患儿的出生体质量偏低,存在病理性黄疸,实验室检查改变包括肝功能异常、高胆红素血症、高胆汁酸血症、低蛋白血症、低血糖、凝血功能障碍、高血乳酸和高血氨等,符合NICCD患儿的典型症状。结论 851del4、1638ins23和IVS6+5GA为中国人SLC25A13基因的突变热点。对于检测SLC25A13基因突变PCR-RFLP方法是一项便捷可靠的NICCD分子诊断技术。     

10例原发性肉碱缺乏症新生儿的基因诊断

目的对新生儿原发性肉碱缺乏症(PCD)基因突变谱进行分析,为PCD早期诊断和治疗以及遗传咨询和产前诊断提供理论依据。方法对进行新生儿串联质谱筛查的34 167份滤纸干血片进行血酰基肉碱谱分析,对其中游离肉碱(C0)低于10μmol/L的新生儿及父母行SLC22A5基因测序。结果酰基肉碱谱筛查C0低于10μmol/L的新生儿10例,患儿母亲游离肉碱均未见降低,10例患儿的SLC22A5基因测序检测到10种20个突变位点,其中c.976CT、c.919del G、c.517del C、c.338GA未见报道,生物信息学分析提示高致病风险。结论串联质谱技术结合SLC22A5基因测序有助于PCD早期诊断,新突变的发现丰富了SLC22A5基因突变谱。     

婴儿肝内胆汁淤积症SLC25A13基因突变分析

目的 探讨SLC25A13基因突变在中国婴儿肝内胆汁淤积症患儿中的检出率，初步了解突变患儿血生化及氨基酸谱特征，肝脏活组织病理变化。方法 2003年12月至2006年12月就诊于复旦大学附属儿科医院的婴儿肝内胆汁淤积症患儿，满足本研究入选条件者共115例进行了血氨基酸质谱分析，伴血浆瓜氨酸明显升高的患儿进行SLC25A13基因全部外显子及邻近序列测序，不伴血浆瓜氨酸明显升高的患儿进行SLC25A13基因常见突变851del4（突变Ⅰ）及突变1638ins23（突变Ⅲ）筛查，突变851del4采用实时荧光定量PCR双标记探针法检测，突变1638ins23采用PCR产物直接电泳法检测，对仅检出单个位点突变的筛查对象，继续进行其余已报道的10种突变位点检测。检测结果仍为单个杂合突变的对象进行SLC25A13基因所有外显子区及其邻近序列分析。对确诊突变患儿的临床表现、血生化及血氨基酸特征等进行分析。结果5例伴血瓜氨酸明显升高的患儿共检出突变4例，其中纯合突变851del4/851del4 1例，复合杂合突变851del4/1638ins23 1例，杂合突变851del4 2例；110例不伴血浆瓜氨酸明显升高患儿共检出突变6例，其中纯合突变851del4/851del4 1例，复合杂合突变851del4/1638ins23 1例，杂合突变851del4 4例。115例婴儿肝内胆汁淤积症患儿共检出SLC25A13基因突变10例，占8.7％。突变患儿血生化改变包括胆红素、γ-谷氨酰转移酶以及碱性磷酸酶等明显升高，AST升高较ALT明显。血串联质谱发现5例突变患儿有特征性氨基酸瓜氨酸、苏氨酸及蛋氨酸升高，另5例突变患儿并无血氨基酸改变。10例患儿中有7例行肝脏活组织病理学检查，4例有显著的脂肪变性。结论 SLC25A13基因突变是中国婴儿肝内胆汁淤积症的重要原因之一。肝脏活组织病理、血生化及氨基酸谱等检查对诊断SLC25A13基因突变患儿有重要意义，但最终仍需通过基因检测确诊。     

语前聋患儿连接蛋白30基因突变

目的分析46例语前聋患儿连接蛋白30（Connexin30,Cx30,GJB6）基因del（GJB6-D13S1830）突变。方法收集46例散发的语前聋患儿及听力正常健康对照30例血标本,提取DNA后利用聚合酶链反应（PCR）分析方法,筛查Cx30基因del（GJB6-D13S1830）突变。结果Cx30基因存在del（GJB6-D13S1830）杂合突变3例,余患儿及听力正常者无此突变。结论Cx30基因del（GJB6-D13S1830）突变可能是导致语前聋的原因之一。     

The principles of molecular diagnosis of recessive forms of prelingual non-syndromic hearing loss

The GJB2 gene defects are the most frequent cause of autosomal recessive non-syndromic hearing loss (DFNB1). Epidemiological data suggest that 35delG is the most prevalent mutation found in 88% of mutated alleles. Another mutations - 313del14 was found in 7% of mutated alleles. The other mutations were identified only in single families. Following the analysis of distribution of GJB2 mutations in the Polish population we propose an algorithm for molecular diagnosis of DFNB1. We propose to screen all patients affected with prelingual non-syndromic deafness for 35delG mutation using ASO or multiplex AS-PCR methods. The presence of 35delG on two alleles confirms DFNB1. The identification of heterozygous 35delG mutation requires additional GJB2 analysis including 313del14 mutation detection and en exon 2 direct sequencing. To determinate the frequency of digenic (GJB2/GJB6) background of DFNB we screened 17 patients with heterozygous 35delG mutation for deletion of 342 kb in GJB6 gene. No such mutation was detected in the analyzed group.     

The mutation 35delG of the gene of the connexin 26 is a frequent cause of autosomal-recessive non-syndromic hearing loss in Morocco]

Mutations of the connexin 26 gene, GJB2, are the most common cause of non syndromic autosomal-recessive hearing loss. One of the GJB2 mutations, the 35delG, is recurrent in European and Mediterranean populations with allelic frequency of at least 70% in patients with hearing loss caused by GJB2 impairment. OBJECTIVES: To determine the prevalence of the 35delG mutation in non-syndromic autosomal-recessive deafness in Morocco. PATIENTS AND METHODS: We looked for the 35delG mutation among 25 non-related Moroccan children suffering from an autosomal recessive hearing loss. A screening for GJB2 mutations, and then a search for GJB6 deletions were carried out among patients who do not bear the 35delG. RESULTS: Twelve patients were homozygous for the 35delG mutation. This mutation was responsible for almost half of the hearing loss among our patients (48%). There was no other GJB2 or GJB6 mutation among 13 patients. CONCLUSION: This study underlines the advantages of a systematic search for this mutation among deaf children when environmental causes are considered irrelevant. The identification of this genetic anomaly signs the etiologic diagnosis of deafness, which allows a relevant genetic advice, and a better treatment of patients.     

GJB2 and GJB6 mutations in children with congenital cytomegalovirus infection

Congenital cytomegalovirus (CMV) infection is a leading cause of sensorineural hearing loss (SNHL) in children. Whether connexin mutations are factors in the development of CMV-related hearing loss has not been explored. We examined gap junction protein beta-2 (GJB2) and gap junction protein beta-6 (GJB6) mutations in 149 children with congenital CMV infection and 380 uninfected neonates. Mutations in GJB2 and GJB6 were assessed by nucleotide sequencing and polymerase chain reaction (PCR) methods, respectively. The study population was predominantly African American, and 4.3% of the subjects were carriers of a connexin 26 mutation. The overall frequency of GJB2 mutations was significantly higher in the group of children with CMV infection and hearing loss (21%) compared with those with CMV infection and normal hearing (3%, p = 0.017) and the group of uninfected newborns (3.9%, p = 0.016). Eight previously reported mutations (M34T, V27I, R127H, F83L, R143W, V37I, V84L, G160S), and four novel mutations (V167M, G4D, A40T, and R160Q) were detected. None of the study children had the 342-kb deletion (delGJB6-D13S1830) in GJB6, which suggests that this mutation does not play a role in hereditary deafness in the African American population. Although GJB2 mutations were detected in children with and without CMV-related hearing loss, those with hearing loss had a higher frequency of GJB2 mutations.     

The R245X mutation of PCDH15 in Ashkenazi Jewish children diagnosed with nonsyndromic hearing loss foreshadows retinitis pigmentosa

Usher syndrome is a frequent cause of the combination of deafness and blindness due to retinitis pigmentosa (RP). Five genes are known to underlie different forms of Usher syndrome type I (USH1). In the Ashkenazi Jewish population, the R245X mutation of the PCDH15 gene may be the most common cause of USH1 (Ben-Yosef T, Ness SL, Madeo AC, Bar-Lev A, Wolfman JH, Ahmed ZM, Desnick RK, Willner JP, Avraham KB, Ostrer H, Oddoux C, Griffith AJ, Friedman TB N Engl J Med 348: 1664-1670, 2003). To estimate what percentage of Ashkenazi Jewish children born with profound hearing loss will develop RP due to R245X, we examined the prevalence of the R245X PCDH15 mutation and its carrier rate among Ashkenazi Jews in Israel. Among probands diagnosed with nonsyndromic hearing loss not due to mutations of connexin 26 (GJB2) and/or connexin 30 (GJB6), and below the age of 10, 2 of 20 (10%) were homozygous for the R245X mutation. Among older nonsyndromic deaf individuals, no homozygotes were detected, although one individual was heterozygous for R245X. The carrier rate of the R245X mutation among the normal hearing Ashkenazi population in Israel was estimated at 1%. Ashkenazi Jewish children with profound prelingual hearing loss should be evaluated for the R245X PCDH15 mutation and undergo ophthalmologic evaluation to determine whether they will develop RP. Rehabilitation can then begin before loss of vision. Early use of cochlear implants in such cases may rescue these individuals from a dual neurosensory deficit.     

Genetic deafness in a preterm infant with a critical postnatal course.

OBJECTIVE: We present a case of deafness in a preterm infant with several predisposing factors of an acquired hearing impairment that, however, turned out to have a genetic cause. We describe the severe postnatal course and review the relevant literature. DESIGN: Case report. SETTING: University-based tertiary neonatal intensive care unit. PATIENT: Preterm infant (gestational age, 26/37; wks). MEASUREMENTS AND MAIN RESULTS: A preterm infant exhibited hearing impairment after a complicated clinical course with pneumothoraces, a hemodynamically relevant patent ductus arteriosus, treatment with potentially ototoxic drugs, intraventricular hemorrhage, and periventricular leukomalacia. Despite the absence of a family history for deafness, genetic testing was performed. Surprisingly, genetic analysis revealed the presence of two compound heterozygous mutations in the patient's GJB2 gene as the cause for his early-onset nonsyndromic deafness. CONCLUSION: To elucidate the nature of a hearing disorder, it is worthwhile to consider a genetic cause, despite the fact that it may seem unlikely in a severely sick preterm infant with numerous risk factors for a postnatally acquired hearing impairment and without a positive family history.     

Unauffälliges UNHS-Screening bei einem homozygoten Träger der 35delG-Mutation im Connexin 26 kodierenden gjb2-Gen

Background

Some cases of recessive nonsyndromic deafness are caused by a mutation in the connexin 26 (Cx26) coding GJB2 gene, mostly 35delG mutations in Western countries. Typically Cx26 mutation-associated hearing impairments are permanent, bilateral, and moderate to severe. However, some variations regarding these factors have been reported.

Case report

A male newborn child received a pass in a TEOAE-based universal newborn hearing screening (UNHS). Though the mother was told about the pass, she was suspicious that her child might be hearing impaired. In response further testing was carried out. A bilateral severe hearing impairment was confirmed by the age of 12 weeks with absent DPAOE and no ABR.

Discussion

Assuming a valid UNHS test result, this would be the first case of a rapid progression in a Cx26-associated hearing impairment. Considering the fact that some members of that family were known to be hearing impaired, a two-step screening (OAE/ABR) should have been favored in the first place. Anyway this case stresses the importance of a thorough and timely audiologic examination in cases of parental supposition that the child may not respond to acoustic stimuli properly.