Mi-Ae Jang

Mutations in DDX58, which Encodes RIG-I, Cause Atypical Singleton-Merten Syndrome

Singleton-Merten syndrome (SMS) is an autosomal-dominant multi-system disorder characterized by dental dysplasia, aortic calcification, skeletal abnormalities, glaucoma, psoriasis, and other conditions. Despite an apparent autosomal-dominant pattern of inheritance, the genetic background of SMS and information about its phenotypic heterogeneity remain unknown. Recently, we found a family affected by glaucoma, aortic calcification, and skeletal abnormalities. Unlike subjects with classic SMS, affected individuals showed normal dentition, suggesting atypical SMS. To identify genetic causes of the disease, we performed exome sequencing in this family and identified a variant (c.1118A>C [p.Glu373Ala]) of DDX58, whose protein product is also known as RIG-I. Further analysis of DDX58 in 100 individuals with congenital glaucoma identified another variant (c.803G>T [p.Cys268Phe]) in a family who harbored neither dental anomalies nor aortic calcification but who suffered from glaucoma and skeletal abnormalities. Cys268 and Glu373 residues of DDX58 belong to ATP-binding motifs I and II, respectively, and these residues are predicted to be located closer to the ADP and RNA molecules than other nonpathogenic missense variants by protein structure analysis. Functional assays revealed that DDX58 alterations confer constitutive activation and thus lead to increased interferon (IFN) activity and IFN-stimulated gene expression. In addition, when we transduced primary human trabecular meshwork cells with c.803G>T (p.Cys268Phe) and c.1118A>C (p.Glu373Ala) mutants, cytopathic effects and a significant decrease in cell number were observed. Taken together, our results demonstrate that DDX58 mutations cause atypical SMS manifesting with variable expression of glaucoma, aortic calcification, and skeletal abnormalities without dental anomalies.

Distribution of Nontuberculous Mycobacteria by Multigene Sequence-Based Typing and Clinical Significance of Isolated Strains

ABSTRACT Species identification of nontuberculous mycobacteria (NTM) is challenging due to the increasing number of identified NTM species and the lack of standardized testing strategies. The objectives of this study were to investigate the distribution of NTM species recovered from respiratory specimens by multigene sequence-based typing and to evaluate the clinical significance of identified species. Two hundred thirty-two consecutive clinical NTM isolates were subjected to sequencing of multiple genes, including hsp65, rpoB, and 16S-23S rRNA internal transcribed spacer (ITS) sequence. In addition, clinical data from all patients whose specimens had NTM isolates were analyzed to examine clinical virulence and treatment history. Eighteen strains from 227 isolates from 169 patients were successfully identified at the species level by multigene sequence-based typing. Mycobacterium avium complex and M. abscessus complex made up the majority of isolated NTM (88%; 199/227), followed by M. fortuitum complex (4%; 10/227). The pathogenic potential of NTM differs enormously by species, and M. avium complex and M. abscessus complex revealed especially high levels of virulence compared with findings for other NTM species. The results from our work support M. avium complex and M. abscessus complex being the most common NTM species with highly pathogenic potential isolated from clinical respiratory specimens and could be a good resource for molecular epidemiology of NTM species in South Korea.

Frequency and spectrum of actionable pathogenic secondary findings in 196 Korean exomes.

Purpose:One of the biggest challenges of exome and genome sequencing in the era of genomic medicine is the identification and reporting of secondary findings. In this study we investigated the frequency and spectrum of actionable pathogenic secondary findings in Korean exomes.Methods:Data from 196 Korean exomes were screened for variants from a list of 56 genes recommended by the American College of Medical Genetics and Genomics (ACMG) for return of secondary findings. Identified variants were classified according to the evidence-based guidelines reached by a joint consensus of the ACMG and the Association for Molecular Pathology.Results:Among the 196 exomes, which were from 100 healthy controls and 96 patients with suspected genetic disorders, 11 variants in 13 individuals were found to be pathogenic or likely pathogenic. We estimated that the frequency of actionable pathogenic secondary findings was 7% for the control subjects (7/100) and 6% for the patients with disease (6/96). For one autosomal-recessive disease, four individuals exhibited either one pathogenic or one likely pathogenic variant of the MUTYH gene, leading to a carrier frequency of 2% (4/196).Conclusion:Secondary findings are not uncommon in Korean exomes.Genet Med 17 12, 1007–1011.

Association of CFTR gene variants with nontuberculous mycobacterial lung disease in a Korean population with a low prevalence of cystic fibrosis.

Several lines of evidence suggest that in Caucasian populations, mutations in the cystic fibrosis (CF) transmembrane conductance regulator (CFTR) gene are associated with susceptibility to lung disease caused by nontuberculous mycobacteria (NTM). However, there is little data available in Asian populations, in which the prevalence of CF is very low. Therefore, we investigated this potential relationship in a Korean population. Sixty patients who fulfilled the diagnostic criteria for NTM lung disease were screened for genetic alterations in the CFTR gene by whole-exon resequencing. For all identified CFTR gene variants, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) genotyping was performed. Genotype and haplotype data were compared between 360 patients with NTM lung disease and 446 healthy controls. Among 13 CFTR genetic variants that were found by whole-exon resequencing, Q1352H showed a significantly higher frequency in NTM patients than in controls, giving an odds ratio (OR) of 4.27 (95% confidence interval (CI), 1.43–12.78). A haplotype with Q1352H showed the strongest association with the disease, with an OR of 3.73 (95% CI, 1.50–9.25). Furthermore, all Q1352H alleles were associated with the V allele of the V470M variant. Our results suggest that CFTR gene variants may increase susceptibility to NTM lung disease in the Korean population. Q1352H appears to be strongly related to NTM lung disease susceptibility in the Korean population.

Identification of a Novel De Novo Variant in the PAX3 Gene in Waardenburg Syndrome by Diagnostic Exome Sequencing: The First Molecular Diagnosis in Korea

Waardenburg syndrome (WS) is a clinically and genetically heterogeneous hereditary auditory pigmentary disorder characterized by congenital sensorineural hearing loss and iris discoloration. Many genes have been linked to WS, including PAX3, MITF, SNAI2, EDNRB, EDN3, and SOX10, and many additional genes have been associated with disorders with phenotypic overlap with WS. To screen all possible genes associated with WS and congenital deafness simultaneously, we performed diagnostic exome sequencing (DES) in a male patient with clinical features consistent with WS. Using DES, we identified a novel missense variant (c.220C>G; p.Arg74Gly) in exon 2 of the PAX3 gene in the patient. Further analysis by Sanger sequencing of the patient and his parents revealed a de novo occurrence of the variant. Our findings show that DES can be a useful tool for the identification of pathogenic gene variants in WS patients and for differentiation between WS and similar disorders. To the best of our knowledge, this is the first report of genetically confirmed WS in Korea.

Comparison of two new generation influenza rapid diagnostic tests with instrument-based digital readout systems for influenza virus detection

Abstract Introduction: Influenza rapid diagnostic tests (RDTs) have been developed to supply scientists with more sensitive and specific techniques. Newly developed digital reader-based techniques require test evaluations before their clinical application. Methods: Two types of digital influenza RDTs using a digital readout system and one conventional RDT were compared using 314 nasopharyngeal swabs of influenza. The swabs originated from symptomatic individuals suspected of influenza infection, and the presence of influenza was confirmed with influenza real-time polymerase chain reaction (PCR) testing and influenza subtyping. Methods were the Sofia® Influenza A + B Fluorescence Immunoassay (FIA), which uses a portable fluorescence analyser, the BD Veritor™ System Flu A + B, which uses a colorimetric immunochromatographic method with a reflectance-based measurement digital device, and the SD Bioline assay, which is based on a traditional immunochromatographic method. Results: The Sofia® Influenza A + B system, the BD Veritor™ System Flu A + B and the SD Bioline assay showed sensitivities in relative real-time PCR results of 74.2, 73.0 and 53.9%, respectively, for influenza A, and 82.5, 72.8 and 71.0%, respectively, for influenza B. All three RDTs showed 100% specificities for influenza A and influenza B. The Sofia® Influenza A + B Fluorescence Immunoassay showed sensitive and specific results for the detection of influenza B in contrast to the BD Veritor™ System Flu A + B. The two digital RDTs showed higher sensitivity and specificity than the conventional RDT in the detection of the influenza H3 subtype. Conclusions: Digital-based readout systems for the detection of the influenza virus can be applied for more sensitive diagnosis in clinical settings than conventional RDTs.

Identification of TSC1 and TSC2 Mutations in Korean Patients With Tuberous Sclerosis Complex

Tuberous sclerosis complex is a genetic disorder caused by mutations in the genes TSC1 or TSC2. Studies of these mutations are very rare in Korean populations. A previous study identified mutations in only 30% of patients by denaturing high performance liquid chromatography with sequencing. Here, we sought to determine the mutational frequency in Koreans. Eleven patients who fulfilled the diagnostic criteria for tuberous sclerosis complex were included. All patients underwent sequencing of both TSC genes, and if no mutations were evident, multiplex ligation-dependent probe amplification was performed. Mutations were detected by sequencing in 82% (9/11) of patients: 36.4% (4/11) in TSC1 and 45.5% (5/11) in TSC2. Two patients with no mutations carried large deletions that included exon 1 of TSC1 in one patient and exons 1-15 of TSC2 in the other patient. Mutations were completely identified in the present study. Therefore, mutation rates in Korean patients may not be lower than those in other ethnic groups. Direct sequencing followed by multiplex ligation-dependent probe amplification analysis may constitute a rational approach to identify disease-causing mutations in Korean patients.

Isolation and Identification of Geosmithia argillacea from a Fungal Ball in the Lung of a Tuberculosis Patient

Geosmithia argillacea, an anamorph of Talaromyces eburneus, is a thermophilic filamentous fungus that has a phenotype similar to that of the Penicillium species, except for the creamy-white colonies and cylindrical conidia. Recently, a new genus called Rasamsonia has been proposed, which is to accommodate the Talaromyces and Geosmithia species. Here, we report the first Korean case of G. argillacea isolated from a patient with a fungal ball. The patient was a 44-yr-old Korean man with a history of pulmonary tuberculosis and aspergilloma. The newly developed fungal ball in his lung was removed and cultured to identify the fungus. The fungal colonies were white and slow-growing, and the filaments resembled those of Penicillium. Molecular identification was carried out by sequencing the internal transcribed spacer (ITS) region of the 28S rDNA and the β-tubulin genes. A comparative sequence analysis using the GenBank (http://blast.ncbi.nlm.nih.gov/) database was performed with the basic local alignment search tool (BLAST) algorithm. The results revealed a 97-100% similarity with the G. argillacea ITS sequence. This case should increase awareness among physicians about the pathogenic potential of G. argillacea in humans and help them accurately identify this fungus, because it can be easily confused with Penicillium and Paecilomyces species owing to their similar phenotypic and microscopic characteristics. A molecular approach should be employed to enable accurate identification of G. argillacea.

Mycobacterial Infection after Intravesical Bacillus Calmette-Guërin Treatment for Bladder Cancer: A Case Report

Bacillus Calmette-Guërin (BCG) has been traditionally used as a vaccine against tuberculosis. Further, intravesical administration of BCG has been shown to be effective in treating bladder cancer. Although BCG contains a live attenuated strain of Mycobacterium bovis, complications such as M. bovis BCG infection caused by BCG administration are extremely rare. Here, we report a case of BCG infection occurring after intravesical BCG therapy. A 67-yr-old man presented with azotemia and weight loss. He had been diagnosed with bladder cancer 4 yr back, and had undergone transurethral resection of the bladder tumor and intravesical BCG (Tice strain) therapy at that time. An acid-fast bacterial strain was isolated from his urine sample. We did not detect Mycobacterium tuberculosis protein 64 (MPT-64) antigen in the isolates obtained from his sample, and multiplex PCR and PCR-reverse blot hybridization assay indicated that the isolate was a member of the M. tuberculosis complex, but was not M. tuberculosis. Finally, sequence analysis of 16S ribosomal RNA and DNA gyrase, subunit B (gyrB) suggested that the organism was M. bovis or M. bovis BCG. Although we could not confirm that M. bovis BCG was the causative agent, the results of the 3 molecular methods and the MPT-64 antigen assay suggest this finding. This is an important finding, especially because M. bovis BCG cannot be identified using common commercial molecular genetics tools.

Identification of DCX Gene Mutation in Lissencephaly Spectrum With Subcortical Band Heterotopia Using Whole Exome Sequencing

Malformations of cortical development include a wide range of brain developmental anomalies that commonly lead to developmental delay and epilepsy. Lissencephaly and subcortical band heterotopia are major malformations of cortical development due to abnormal neuronal migration and several genes have been identified including ARX, DCX, LIS1, RELN, TUBA1A, and VLDLR. Traditionally, genetic testing for lissencephaly and subcortical band heterotopia has been done in the order of the probability of detection of mutation according to the radiologic features, but the success rate could be variable with this time-consuming approach. In this study we used whole-exome sequencing to identify mutations in a 5-year-old girl with lissencephaly spectrum with subcortical band heterotopia. After excluding lissencephaly-related genes, one deleterious mutation (NM_178153.2:c.665C > T, p.Thr222Ile) in the DCX gene was identified. Further Sanger sequencing validated the variant in the patient but not in both parents indicating a de novo mutation. The present report demonstrates that whole-exome sequencing may be a useful tool for the identification of mutations in patients with lissencephaly and subcortical band heterotopias as well as malformations of cortical development.