Acute Porphyromonas gingivalis Subdural Abscess with Brain Abscess in the Left Temporal Lobe: A Case Report and Review of Literature.

Background: Brain abscesses are a rare but serious complication of focal intracerebral infection. Case Description: We present a patient of acute subdural abscess with brain abscess in the left temporal lobe. After craniotomy, combined with the Third Next Generation Sequencing and Gene Diagnosis (TNGS & GD) of abscess, we prescribed sensitive antibiotics; the patient recovers well and the abscess did not recur. Conclusion: For patients with acute subdural abscess, combined craniotomy and the TNGS & GD of abscess could achieve good results.

[Application of Bionano-DNA Full-Length Optical Mapping Technology in Genetic Diagnosis of Hemophilia A Carriers].

OBJECTIVE: To apply Bionano Saphyr visual full-length DNA optical mapping technology to the precise genetic diagnosis of hemophilia A carriers. METHODS: For 2 suspected F8 gene deficiency female carriers who could not be diagnosed by conventional next-generation sequencing technology, the full-length DNA optical mapping technology was used to detect and scan the sample X chromosome full-length visual haplotype characteristic map, which was compared with the normal haplotype. The gene structure variation information of the samples was obtained by compare with DNA atlas library. RESULTS: The average fluorescent marker length of the X chromosome DNA molecular where the F8 gene was located in the two samples was greater than 2.5 Mbp, and the average copy number was greater than 20×. After comparative analysis, one of the samples was a proximal inversion of intron 22 of the F8 gene, and another was an inversion of intron 22 accompanied by multiple deletions of large fragments. CONCLUSIONS: Bionano technology has a good detection rate for gene defects with large length and complex variation. In the absence of a proband or accurate genetic diagnosis results of the proband, the application of this technology to detect the heterozygous complex variant of the F8 gene is of great significance for the prenatal diagnosis and pre-pregnancy diagnosis of hemophilia carriers.

Clinical laboratory genetic diagnosis of Marfan syndrome / 中华检验医学杂志

Objective:To establish the clinical laboratory genetic diagnosis procedures for Marfan syndrome (MFS) and carry out clinical laboratory genetic diagnosis for MFS families.Methods:The second generation high-throughput sequencing was used to sequence and analyze the FBN1 gene of two MFS families who visited to Fuwai Central China Cardiovascular Hospital (Heart Center of Henan People′s Hospital) from January to December 2020, and then Sanger sequencing was used to verify the second generation high-throughput sequencing results. At the same time, the sanger sequencing of mutation sites was performed on normal family members and 100 healthy people to identify the pathogenic mutations of FBN1 gene in the MFS families. The pregnant women of two families were guided for prenatal diagnosis in the second trimester of pregnancy.Results:The clinical laboratory diagnosis of MFS showed that two MFS patients had the pathogenic mutation of c.2560T>C heterozygous mutation and c.6772T>C heterozygous mutation in FBN1 gene, respectively. The mutation was not observed in 100 healthy people and normal members in two families. The prenatal diagnosis showed that there was a heterozygous mutation of FBN1 gene c.2560T>C in the first fetus of the MFS family, which was MFS. There was no mutation in the FBN1 gene in the second fetus of the MFS family, so it was recommended to continue the pregnancy. The results of postpartum follow-up were consistent with the results of clinical laboratory diagnosis.Conclusion:The clinical laboratory genetic diagnosis procedures for MFS have been established successfully, which provides an important reference for clarifying the clinical diagnosis of MFS.

Significance of Gene Diagnosis in Acute Myeloid Leukemia with the Emergence of New Molecular Target Drug Treatment.

Acute myeloid leukemia (AML) is a heterogeneous hematopoietic malignancy accompanied by impaired differentiation and autonomous proliferation of hematopoietic stem cells. Standard induction therapy results in first complete remission among 70% of patients with AML; however, approximately half of these patients relapse and become refractory. Allogeneic hematopoietic cell transplantation is a useful treatment for relapsed and refractory cases. However, transplantation-related mortality is approximately 20%, which is not a low value, and quality of life after transplantation decreases. Therefore, there is a need to stratify the prognosis of each patient and implement this treatment appropriately. Owing to recent advances in genome analysis technology, many gene mutations involved in onset and recurrence of AML have been discovered. These abnormalities and mutations not only have clinical application as prognostic factors and minimal residual disease markers, but they may also contribute to novel molecular targeted drug development. Many new drugs such as first-generation FMS-like tyrosine kinase 3 (FLT3), isocitrate dehydrogenase 1 and 2 (IDH1/2), and B cell lymphoma 2 (BCL2) inhibitors have been developed in the West. In addition, the second-generation FLT3 inhibitors gilteritinib and quizartinib were developed in Japan, and treatment outcomes for patients with AML have improved. However, there is still a large disparity in drug availability between the West, and Japan. As a result, treatment guidelines in the West cannot be applied in the clinical setting in Japan. In this study, we assessed the molecular target drug treatment by gene diagnosis for treatment of AML patients.

Analysis of genotype-phenotype correlation in patients with α-thalassemia from Fujian province, Southeastern China.

BACKGROUND: There is a high carrying rate of α-thalassemia in Fujian province. However, there are few large-scale studies on the correlation between genotype and phenotype in Fujian province. The purpose of this study was to analyze the phenotype and genotype in a cohort of 2923 patients with α-thalassemia in Fujian province, so as to provide reference data for screening and diagnosis of α-thalassemia in Fujian province. METHODS: The genotype of α-thalassemia was detected by PCR reverse dot blot assay, gap-PCR, single PCR, nested PCR, and sequencing. Clinical and hematological indices of 2923 patients were collected, and the correlation between genotype and phenotype was analyzed. RESULTS: Among 10,350 patients, 2923 cases were found with α-thalassemia, with a detection rate of 28.24%. Among them, --SEA /αα was the most common genotype, accounting for 64.80%. In addition, rare α-thalassemia genotypes were detected in Fujian province, including --THAI /αα (0.41%), HKαα/--SEA (0.03%), and the novel α-thalassemia gene mutation CD5 (GCC>ACC) (HGVS named HBA1: c.16G>A) (0.03%). Patients with deletional genotypes of α-thalassemia were found to have higher RBC and lower Hb, MCV, MCH, and HbA2 than patients with non-deletional genotypes of α-thalassemia (p < 0.05). CONCLUSION: The clinical phenotype of α-thalassemia is influenced by molecular mechanisms. HBA1: c.16G>A mutation is a novel mutation that was first reported in Fujian province, which enriches the human hemoglobin mutation spectrum.

The gene diagnosis of neurofibromatosis type I with headache as the main symptom: A case report and review of the literature.

Neurofibromatosis type I (NF1) is an autosomal dominant disease. Some NF1 patients experience atypical clinical manifestations, genetic testing is not widely available, and the types of mutations vary; thus, they are prone to misdiagnosis and missed diagnosis. Although headache is not included in the diagnostic criteria for NF1, the incidence of headache in NF1 patients is not low. We report an NF1 family in which the proband presented with prominent headache and atypical clinical presentation, with limited skin pigmentation. We identified a frameshift mutation (c.1541_1542del, p. Q514Rfs*) in the NF1 gene by whole-exome sequencing of this family, and the patients were diagnosed with NF1. We hope to attract the attention of clinicians to these patients and improve genetic testing as soon as possible to increase the diagnosis rate.

Successful Management of a Young Athlete with Type 2 Long QT Syndrome by Genotype-specific Risk Stratification and Bridging Therapy with a Wearable Cardioverter Defibrillator.

We herein report a 14-year-old boy with repetitive nocturnal syncope related to medication-refractory long QT syndrome (LQTS). Although the use of an implantable cardioverter-defibrillator (ICD) was inevitable to prevent sudden cardiac death, he refused immediate implantation in order to play in a baseball competition six weeks away. Given his genetic diagnosis of type 2 LQTS, which is associated with cardiac events unrelated to exercise, we prescribed a wearable cardioverter defibrillator (WCD) to be donned at night, without limiting his exercise participation. An ICD was implanted after the competition. We successfully performed the preplanned treatment while maximizing the patient's quality-of-life with a WCD and genotype-specific risk stratification.

Combinations of Ghd7, Ghd8, and Hd1 determine strong heterosis of commercial rice hybrids in diverse ecological regions.

Heterosis of grain yield is closely associated with heading date in crops. Gene combinations of the major heading date genes Ghd7, Ghd8, and Hd1 play important roles in enhancing grain yield and adaptation to ecological regions in rice. However, the predominant three-gene combinations for a specific ecological region remain unclear in both three-line and two-line hybrids. In this study, we sequenced these three genes of 50 cytoplasmic male sterile/maintainer lines, 31 photo-thermo-sensitive genic male sterile lines, and 109 restorer lines. Sequence analysis showed that hybrids carrying strong functional alleles of Ghd7 and Hd1 and non-functional Ghd8 are predominant in three-line hybrids and are recommended for rice production in the subtropics around 30°N/S. Hybrids carrying strong functional Ghd7 and Ghd8 and non-functional Hd1 are predominant in two-line hybrids and are recommended for low latitude areas around 23.5°N/S rich in photothermal resources. Hybrids carrying strong functional Ghd7 and Ghd8 and functional Hd1 were not identified in commercial hybrids in the middle and lower reaches of the Yangtze River, but they have high yield potential in tropical regions because they have the strongest photoperiod sensitivity. Based on these findings, two genic sterile lines, Xiangling 628S and C815S, whose hybrids often head very late, were diagnosed with these three genes, and Hd1 was targeted to be knocked out in Xiangling 628S and replaced with hd1 in C815S. The hybrids developed from both modified sterile lines in turn had appropriate heading dates and significantly improved grain yield. This study provides new insights for breeding design to develop hybrids for various regions.

Generalized lipodystrophy type 1 due to compound heterozygous mutation of AGPAT2 gene: One case report and literature review / 中华内分泌代谢杂志

Congenital generalized lipodystrophy type 1 (CGL1) is an autosomal recessive genetic disease caused by mutations in AGPAT2 gene. The main clinical mainifestations include body subcutaneous fat loss, muscle hypertrophy, obvious subcutaneous veins, pseudoacromegaly, hirsutism, and acanthosis nigricans. What′s more, CGL1 is always accompanied by metabolic diseases. Therefore, it is easily misdiagnosed as metabolic syndrome, type 2 diabetes, polycystic ovary syndrome, acromegaly, or Cushing′s syndrome. Meanwhile, it is difficult to distinguish it from partial lipoatrophy syndrome. In this article, we present clinical and molecular characteristics of a patient with CGL1 and review mutations reported in literature to replenish current knowledge about this orphan disease.

Diagnostic and prognostic value of MCM3 and its interacting proteins in hepatocellular carcinoma.

Aberrant DNA replication is one of the driving forces behind oncogenesis. Furthermore, minichromosome maintenance complex component 3 (MCM3) serves an essential role in DNA replication. Therefore, in the present study, the diagnostic and prognostic value of MCM3 and its interacting proteins in hepatocellular carcinoma (HCC) were investigated. By utilizing The Cancer Genome Atlas (TCGA) database, global MCM3 mRNA levels were assessed in HCC and normal liver tissues. Its effects were further analyzed by reverse transcription-quantitative PCR (RT-qPCR), western blotting and immunohistochemistry in 78 paired HCC and adjacent tissues. Functional and pathway enrichment analyses were performed using the Search Tool for the Retrieval of Interacting Genes database. The expression levels of proteins that interact with MCM3 were also analyzed using the TCGA database and RT-qPCR. Finally, algorithms combining receiver operating characteristic (ROC) curves were constructed using binary logistic regression using the TCGA results. Increased MCM3 mRNA expression with high α-fetoprotein levels and advanced Edmondson-Steiner grade were found to be characteristic of HCC. Survival analysis revealed that high MCM3 expression was associated with poor outcomes in patients with HCC. In addition, MCM3 protein expression was associated with increased tumor invasion in HCC tissues. MCM3 and its interacting proteins were found to be primarily involved in DNA replication, cell cycle and a number of binding processes. Algorithms combining ROCs of MCM3 and its interacting proteins were found to have improved HCC diagnosis ability compared with MCM3 and other individual diagnostic markers. In conclusion, MCM3 appears to be a promising diagnostic biomarker for HCC. Additionally, the present study provides a basis for the multi-gene diagnosis of HCC using MCM3.

[Research progress on laboratory diagnosis of drug-induced liver injury].

Drug-induced liver injury (DILI) is one of the common adverse drug reactions in the clinic and is also the main reason for the withdrawal of new drugs from the market. Although the overall incidence rate of DILI is not high; however, it can cause severe adverse outcome and even death, and has become the core cause of acute liver failure in Europe and the United States. In addition, DILI diagnosis is a puzzling problem for clinicians. Drug re-stimulation can be used as the "gold standard" in the diagnosis of DILI, but it may re-induce liver failure, so it cannot be recommended for clinical use. Currently, laboratory diagnostic methods including serum biomarker, genetic testing, scoring scale, and so on are available for DILI.

The broad-spectrum rice blast resistance (R) gene Pita2 encodes a novel R protein unique from Pita.

BACKGROUND: Rice blast is generally considered the most devastating rice disease worldwide. The development of resistant varieties has been proven to be the most economical strategy to control the disease. A cluster of resistant (R) genes on rice chromosome 12 including Pita, Pita2 and Ptr has been studies for decades. However, the relationship between these R genes has not been well established. RESULTS: In this study, we compared the resistance spectra controlled by Pita2 and Pita by testing their monogenic lines (MLs) in four hotspots found in the Philippines and Burundi from 2014 to 2018. The reaction patterns were distinct in two countries and that Pita2-mediated field resistance was relatively prevalent. Pathogenicity tests using 328 single-spore isolates in greenhouse further verified that IRBLta2-Re for Pita2 conferred a relatively broader spectrum resistance than those of Pita. Rough and fine mapping of Pita2 were conducted using F2 and F3 populations derived from IRBLta2-Re [CO] and CO 39 consisting of 4344 progeny to delimit Pita2 in a genomic interval flanked by two markers 12 g18530 and 12 g18920 proximal to the centromere of chromosome 12. Alignment of the markers to the genomic sequence of IR64, which harbors Pita2 verified by genetic analysis, approximately delimited the candidate gene(s) within 313-kb genomic fragment. The two Pita2 suppressive mutants that contain mutations within Pita2 were verified and identified. Comparative sequence analysis in these two mutants further identified that each individual allele contains a single nucleotide substitution at a different position resulting in nonsense and missense mutations in the protein product of LOC_Os12g18729. On the contrary, no sequence mutation was detected in other candidate genes, indicating that mutations in LOC_Os12g18729 were responsible for the loss of function of Pita2. Pita2 encodes a novel R protein unique from Pita, which is exactly identical to the previously cloned Ptr. Moreover, based on the resistance gene analysis of rice varieties and mutants containing Pita, it was found that Pita2 rather than Pita was responsible for the specificity to some differential isolates with AvrPita. The diagnosis and survey of Pita2 in IRRI released varieties showed relatively low frequency, implying a high value of its application for breeding resistant varieties against rice blast via marker assisted selection. CONCLUSION: Our study clarified the relationship between Pita, Pita2 and Ptr. Pita2 is identical to Ptr and distinct from Pita in both sequence and chromosomal location although Pita2 and Pita are genetically linked to each other. The loss of function of Pita2 but not Pita eliminate the specificity to some AvrPita containing isolates, however, the mechanism underlying the recognition between Pita2/Pita and AvrPita remains elusive.

Newborn screening and diagnosis of inborn errors of metabolism: A 5-year study in an eastern Chinese population.

Inborn errors of metabolism (IEMs) can cause intellectual disability or even death in children. To evaluate the disease spectrum and genetic characteristics of IEMs in Jining City of Shandong Province in East China, we used tandem mass spectrometry (MS/MS) technology for IEMs screening combined with genetic analysis. Newborns were screened from July 14, 2014, to December 31, 2018. Amino acid and carnitine contents were detected by MS/MS. According to the results for normal newborns, the reference range of our laboratory was established with the percentile method. The suspected positive newborns were further diagnosed using next-generation sequencing. A total of 514,234 newborns were screened, and 265 were diagnosed with IEMs, with a detection rate of 1:1941. Of the 265 patients, 130 (49.06%) had organic acid disorders, 83 (31.32%) had amino acid disorders, 34 (12.83%) had fatty acid oxidation disorders, and 18 (6.79%) had urea circulatory disorders. PAHD and MMA were the two most common disorders. IEMs-associated genes were identified in 233 patients. Our data indicated that IEMs are never uncommon in Jining, and the disease spectrum and genetic background were clearly elucidated, contributing to the treatment and prenatal genetic counseling of these disorders in the region.

Current Study of the Detection and Treatment Targets of Spinal Tuberculosis.

Spinal tuberculosis is a common manifestation of extrapulmonary tuberculosis and osteoarticular tuberculosis. Common clinical manifestations include constitutional symptoms, back pain, spinal tenderness, paraplegia, and spinal deformities. They are the common causes of paralysis and could increase the mortality in patients. Most cases of spinal tuberculosis remaining undiagnosed, and early clinical symptoms and imaging manifestations lack specificity, which explained the reason why it is difficult to identify from atypical spinal metastases, brucellosis and other diseases. The rate of missed diagnosis and misdiagnosis for spinal tuberculosis is high. If spinal tuberculosis diagnostic targets could be early detected, the therapeutic targets can be effectively treated, which can not only control the progress of the disease and shorten the course of treatment, but also reduce the economic pressure and avoid spinal deformity. Therefore, early diagnosis should be our focus. Comprehensive use of a variety of diagnostic targets can improve the early diagnosis rate of spinal tuberculosis. Here, we review the progress of laboratory, imaging and gene detection in the diagnosis of spinal tuberculosis in recent years.

[Prenatal gene diagnosis of 200 fetuses at high risk of osteogenesis imperfect].

Objective: The authors aim to provide genetic counselling and prenatal gene diagnosis to the families with osteogenesis imperfecta(OI), based on the identification of pathogenetic mutations in large cohort genetic testing. Methods: DNA was extracted from the peripheral blood of parents of the fetuses, and from the villi tissue, amniotic fluid or cord blood of the fetuses using a standard sodium dodecyl sulfate-proteinase K-phenol/chloroform extraction method. PCR combined with Sanger DNA sequencing was performed to validate the pathogenic mutations of 200 fetuses at risk of OI and their parents from 158 families. Allelic analysis of microsatellite markers was applied to exclude the false positive caused by maternal DNA contamination, when both the fetus and the mother harbored the same pathogenic genotype. Results: A total of 83 affected fetuses (83/200, 41.5%) and 12 (12/200, 6.0%) recessive carriers were identified among the 200 fetuses. The 83 affected fetuses included 78 heterozygotes (45 of COL1A1, 32 of COL1A2, one of IFITM5), and 5 compound heterozygotes or homozygotes of recessive OI (two of FKBP10, one of SEC24D, one of WNT1 and one of CRTAP); The 12 recessive carriers included 7 of WNT1, 4 of SERPINF1 and one of SERPINH1. Maternal DNA contamination was excluded from the genomic DNA samples of OI fetuses when their mother with the same affected genotypes. Conclusion: In this study, the authors used an optimized gene diagnosis system of OI to perform prenatal genetic diagnosis to 200 fetuses at high risk of OI, and provided precisely genetic counselling to the OI families.

Application of Next-Generation Sequencing Following Tandem Mass Spectrometry to Expand Newborn Screening for Inborn Errors of Metabolism: A Multicenter Study.

This study explored the effectiveness of expanding newborn screening (NBS) by tandem mass spectrometry (TMS) and gene diagnosis by next-generation sequencing (NGS). First, we described the characteristics of gene variants in Jiangsu Province. We collected clinical data from three NBS centers. All infants followed a unified screening and diagnosis process. After obtaining informed consent, dried blood spots (DBSs) were collected and analyzed by TMS. If the results fell outside of the cut-off value, repeat analysis was performed. If the re-test results remained abnormal, the infant was recalled for further assessment. We performed targeted sequencing using the extended edition panel of inborn errors of metabolism (IEM) to detect 306 genes using the Illumina HiSeq 2500 platform. A total of 536,008 babies underwent NBS by TMS in three NBS centres. In total, 194 cases were eventually diagnosed with various types of inherited metabolic diseases, with an overall incidence of 1/2763. There were 23 types of diseases, including ten amino acid disorders (43.5%), eight organic acidaemias (34.8%) and five fatty acid oxidation defects (21.7%). In these infants, we clearly identified variants of disease-causing genes by next-generation sequencing (NGS). Most had two variants and others had one or three variants: 88% of gene variants were heterozygous and 12% were homozygous. There is a certain incidence of IEM in Jiangsu Province and it is necessary to carry out screening for 27 diseases. Meanwhile, NGS combined with TMS offers an enhanced plan for NBS for IEM.

A Designed Experiment: Polymorphism Analysis of Angiotensin-Converting Enzyme Gene from Human Buccal Epithelial Cells.

For medical students, we combine the laboratory practice with clinical applications by developing biochemical and molecular biology experiments. In this experiment, students first collect their own buccal epithelial cells by a noninvasive mouthwash method. Then, they extract genomic DNA and perform polymerase chain reaction (PCR) to amplify angiotensin-converting enzyme (ACE) gene using genomic DNA as a template. Finally, the polymorphism of ACE gene is observed by electrophoresis. Students not only learn the techniques but also acquire knowledge of the ACE gene polymorphism. By establishing the relationship among ACE polymorphism and high blood pressure and myocardial hypertrophy, students should be able to understand the gene polymorphism and its association with susceptibility to disease. This laboratory practice teaching can also stimulate desire to do scientific research. Experimental results from many individuals can help us determine and analyze the fractions of ACE gene types in Chinese cohorts. Such an experiment strongly activates students and provides a solid foundation for the medical students' future research and clinical application. © 2019 International Union of Biochemistry and Molecular Biology, 47(2): 168-174, 2019.

[The clinical application of gene diagnosis and genetic counseling on hereditary hearing loss].

Objective:To summarize the clinic procedure and experience about gene diagnosis and genetic counseling on hereditary hearing loss, and explore the strategy and principle about gene diagnosis and genetic counseling on hereditary hearing loss.Method:A retrospective analysis was used on the clinical data of 151 cases who aim at genetic counseling. The all cases were divided into 5 groups according to the purpose of genetic counseling, such as the occurrence risk of hearing loss, the etiological analysis, the choice of the intervention way, the examination guidance, the prevention of hearing loss and the usage requirement of Aminogly cosides drugs. The counseling procedure includes the investigation of the etiology and family history, drawing the family pedigree, general physical examination, auditory examination and genetic analysis. Sanger sequencing analysis and/or Targeted nextgeneration sequencing was utilized to detect the deaf-gene mutations. At last, the genetic counseling, fertility guidance and prenatal diagnosis will be made on the basis of the results of gene detection. Result:There are 33 newborns who did not pass the deafgene screening, 9 of them could be diagnosed definitely as hereditary hearing loss, and the other 24 were the carriers of deafgene mutation. Eighty of 104 deaf patients were diagnosed definitely as hereditary hearing loss and the related gene mutation was found. Six objects in the 10 patients with auditory neuropathy are diagnosed as OTOF or SLC17A8 gene mutations before cochlear implantation. Three of 7 reproductive age objects who had family history were recessive deaf-gene carriers, 2 of them carried the same target gene with the mate who receive our fertility guidance and prenatal diagnose. The other 1 object carried the dominant genetic mutation(incomplete dominant heredity). There were 4 pregnant women who did not pass the deaf-gene screening, 1 of them carry the same target gene with the mate. The populations who want to use Aminoglycosides drugs were not diagnosed as carrying any related mitochondrial gene mutation. We carried out the genetic counseling according to the results of gens detection and clinical phenotype.Conclusion:Genetic counseling is based on the different purpose. The analysis of gene diagnosis should be considered to combine with the clinical phenotype. The principle of choosing the objects to make a gene diagnosis includes: ①the all deaf-genes sequencing was applied for the deaf patients. ② the screening target gene sequencing was used for the newborns who did not pass the deaf-gene screening and the mate whose pregnant wife did not pass the deafgene screening. ③the specific target gene sequencing could be used for the patients who has a clear family history or specific phenotype.

Analysis of AIRE gene mutation in a pedigree with autoimmune polyendocrine syndrome type Ⅰ / 临床检验杂志

Objective@#To analyze the mutation of autoimmune regulator ( AIRE ) gene in a pedigree with autoimmune polyendocrine syndrome type Ⅰ (APS-Ⅰ). @*Methods@#The peripheral blood samples from family members were collected for DNA extraction, and then the mutation sites on AIRE gene were screened by PCR and Sanger sequencing. The mutation sites were further verified in 100 healthy persons by the created restriction site PCR (CRS-PCR) and PCR restriction fragment length polymorphism (PCR-RFLP). The effects of mutation on the structure and function of AIRE protein were analyzed with SIFT, PolyPhen-2, Mutation Taster and Antheprot Editor softwares. The effects of mutation on the splicing sites of AIRE mRNA were predicted with Alternative Splice Site Predictor, FruitFly Splice Predictor and SplicePort softwares, and further verified by Sanger sequencing. @*Results@#Two novel heterozygous mutations c.47 C＞G T16R and c.1631-2 A＞T were found in the proband. The c.47 site is highly conserved and homologous in different species. The missense mutation of c.47C＞G changed the secondary structure and hydrophobicity of AIRE protein, and affected its function. The c.1631 -2 A＞T mutation changed the splicing site of AIRE mRNA, and led to the deletion of exon 13. @*Conclusion@#Two novel pathogenic mutations c.47 C＞G T16R and c.1631-2 A＞T are identified in a pedigree with autoimmune polyendocrine syndrome type Ⅰ.

Diagnostic efficacy of blood promoter methylation of Sox10 in intestinal neuronal dysplasia / 国际儿科学杂志

Objective To explore the efficacy of blood promoter methylation of Sox10 gene in diagnosis of intestinal neuronal dysplasia (IND) and to seek a non-invasive genetic diagnosis method based on peripheral blood for diagnosis of IND.Methods Children diagnosed as Hirschsprung disease (HD) or IND from the Shengjing Hospital of China Medical University and the Capital Institute of Pediatrics were enrolled in 2017-2018.The blood and colon specimens were collected from 9 IND,15 HD and 15 controls (the colon trauma cases).The blood promoter methylation of Soxl0 and its expression level in colon were both detected and the correlation between them was analyzed.The diagnostic efficacy of blood promoter methylation of Soxl0 was analyzed by receiver operating characteristics (ROC) curve.Results The blood promoter methylation level at the 32nd locus of Sox10 was 100％ (90％-100％;95％ CI:91％-98％) in the control,80％ (70％-90％;95％CI:65％-90％) in HD and 60％ (50％-80％;95％ CI:52％-82％) in IND.The expression level of Sox10 in the colon was (1.00 ±0.04) in the control,(2.75 ±0.16) in HD and (3.99 ±0.10) in IND.Western blot showed that the expression of Sox10 protein in the colon of the control group,the HD group and the IND group increased,and the difference was statistically significant (P ＜ 0.05).The blood promoter methylation level was negatively correlated with its expression level in colon (r =-0.88).ROC curve indicated area under curve (AUC) of Sox10 methylation in diagnosis of HD was 0.818,with a cut-off value of 85％ and low diagnostic sensitivity.The AUC in IND was 0.907,with a cut-off value of 85％,producing a sensitivity of 88.9％ and a specificity of 93.3％ respectively.Conclusion Blood promoter methylation of Sox10 might be used as a non-invasive method for diagnosis of IND.