• Korean Journal of Clinical Laboratory Science
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• v.38 no.3
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• pp.158-165
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• 2006

Although Mycobacterium tuberculosis complex strains remain responsible for the majority of diseases caused by mycobacterial infections worldwide, the increase in HIV infections has allowed for the emergence of other non-tuberculous mycobacteria as clinically significant pathogens. However, Mycobacterium species has a long period of incubation, and requires serious biochemical tests such as niacin, catalase, and nitrate test that are often tedious. The development of rapid and accurate diagnostics can aid in the early diagnosis of disease caused by Mycobacterium. The current DNA amplification and hybridization methods that have been developed target several genes for the detection of mycobacterial species such as hps65, 16S rDNA, rpoB, and dnaj. These methods produce rapid and accurate results. In this study, PCR-restriction fragment length polymorphism analysis(PCR-RFLP) based on the region of the rpoB gene was used to verify the identification of non-tuburculosis Mycobacterium species. A total of 8 mycobacterial reference strains and 13 clinical isolates were digested with restriction enzymes such as Msp I in this study. The results of using this process clearly demonstrated that all 13 specimens were identified by rpoB gene PRA method. The PCR-RFLP method based on the rpoB gene is a simple, rapid, and accurate test for the identification of Mycobacterium.

• Korean Journal of Clinical Laboratory Science
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• v.45 no.2
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• pp.48-53
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• 2013

Recently, the isolation rate of nontuberculous mycobacteria (NTM) in clinical laboratories and the incidence of NTM infections are on the increase in Korea, but there have been only a few studies that reveal the general aspect of NTM isolation or species distribution. Therefore, this study was performed to examine the species identification by PCR-restriction fragment length polymorphism analysis (PRA, PCR-RFLP), and the clinical significance of mycobacterial cultures. PRA was used during the novel region of the rpoB gene and was developed for rapid and precise identification of mycobacteria to the species level. From January 2012 to April 2012, we examined pre-identified nontuberculous mycobacteria (60 species in 3 hospital of Busan-Kyeongnam area). We confirmed 4 (6.6%) Mycobacterium tuberculosis (MTB) and 56 (93.4%) NTM from 60 pre-identified NTM species by multiplex PCR (MolecuTech $MTB-ID^R$ V3, YD Diagnostics, Korea) and PRA (Myco-ID, YD Diagnostics, Korea). The distribution of 56 NTM species were M. intracellulare type I 15 (26.7%), M. avium 14 (25%), M. abscessus 11 (19.5%), M. kansasii type I 3 (5.4%), M. pulveris 2 (3.6%), M. intracellulare type, M. chelonae, M. kansasii type V, M. gallinarum, M. wolinskyi. Respectively, 1 (1.8%) and 6 (10.7%) species were not identified.

• Journal of Korean Society on Water Environment
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• v.36 no.3
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• pp.220-228
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• 2020

Anaerobic ammonium oxidation (AMX) is a cost-efficient biological nitrogen removal process. The coexistence of ammonia-oxidizing bacteria (AOB) in an AMX reactor is an interesting research topic as a nitrogen-related bacterial consortium. In this study, a sequencing batch reactor for AMX (AMX-SBR) was operated with a conventional activated sludge. The AOB in an AMX bioreactor were identified and quantified using terminal restriction fragment length polymorphism (T-RFLP) and real-time qPCR. A T-RFLP assay based on the ammonia monooxygenase subunit A (amoA) gene sequences showed the presence of Nitrosomonas europaea-like AOB in the AMX-SBR. A phylogenetic tree based on the sequenced amoA gene showed that AOB were affiliated with the Nitrosomonas europaea/mobilis cluster. Throughout the enrichment period, the AOB population was stable with predominant Nitrosomonas europaea-like AOB. Two OTUs of amoA_SBR_JJY_20 (FJ577843) and amoA_SBR_JJY_9 (FJ577849) are similar to the clones from AMX-related environments. Real-time qPCR was used to quantify AOB populations over time. Interestingly, the exponential growth of AOB populations was observed during the substrate inhibition of the AMX bacteria. The specific growth rate of AOB under anaerobic conditions was only 0.111 d^-1. The growth property of Nitrosomonas europaea-like AOB may provide fundamental information about the metabolic relationship between the AMX bacteria and AOB.

• Tuberculosis and Respiratory Diseases
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• v.69 no.5
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• pp.331-336
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• 2010

Background: Recently, the rate of infections with non-tuberculous mycobacteria (NTM) has been increasing in Korea. Precise identification of NTM is critical to determination of the pathogen and to target treatment of NTM patients. Methods: Sixty-eight unclassified mycobacteria isolates by rpoB PCR-RFLP assay (PRA) collected in 2008 were analyzed by National Center for Biotechnology Information (NCBI) Basic Local Alignment Search Tool (BLAST) search after sequencing of 16S rRNA, hsp65, rpoB genes. Results: Nineteen strains of 68 isolates were specified as species after sequencing analysis of 3 gene types. We found 3 M. lentifulavum, 5 M. arupense, 4 M. triviale, 4 M. parascrofulaceum, and one M. obuense. One M. tuberculosis and another M. peregrinum were mutated at the Msp I recognition site needed for rpoB PRA. The remaining 49 isolates did not coincide with identical species at the 3 kinds genes. Conclusion: Sequencing analysis of 16S rRNA, hsp65, rpoB was useful for identification of NTM unclassified by rpoB PRA.