Identification of Significant Pathogenic Nontuberculous Mycobacteria Species from Presumptive TB Patients Using Partial hsp65 Gene Sequencing

Dyshelly Nurkartika Pascapurnama; Nabilla Ghina Zavitri; Raspati Cundarani Koesoemadinata; Adi Imam Cahyadi; Lidya Chaidir

doi:10.2147/IDR.S419956

Identification of Significant Pathogenic Nontuberculous Mycobacteria Species from Presumptive TB Patients Using Partial hsp65 Gene Sequencing

Infect Drug Resist. 2023 Oct 30:16:6923-6930. doi: 10.2147/IDR.S419956. eCollection 2023.

Authors

Dyshelly Nurkartika Pascapurnama¹, Nabilla Ghina Zavitri¹, Raspati Cundarani Koesoemadinata¹, Adi Imam Cahyadi², Lidya Chaidir^{2

3}

Affiliations

¹ Research Center for Care and Control of Infectious Diseases, Universitas Padjadjaran, Bandung, West Java, Indonesia.
² Department of Biomedical Sciences, Faculty of Medicine, Universitas Padjadjaran, Sumedang, West Java, Indonesia.
³ Center for Translational Biomarker Research, Universitas Padjadjaran, Bandung, West Java, Indonesia.

Abstract

Purpose: To date, the diagnosis of nontuberculous mycobacteria (NTM) disease primarily relies on clinical symptoms and radiological features. Our objective was to apply a sequence-based analysis method by using partial gene sequencing of heat shock protein 65 (hsp65) to identify NTM species.

Patients and methods: A total of 32 stored isolates obtained from individuals suspected of having pulmonary NTM infection were subjected to solid Ogawa culture. Genomic DNA from each sample was extracted and used in a conventional polymerase chain reaction (PCR) targeting a specific region of hsp65 gene. Identified amplicons from the PCR were then subjected to targeted sequencing. Analysis of the obtained hsp65 sequence was performed using DNA Baser tool. The consensus sequences obtained were compared to references in the GenBank NCBI database to determine NTM species.

Results: We identified several important NTM species which posses opportunistic characteristics. M. abscessus and M. chelonae are the most frequent NTM species identified in this study (40.63% and 18.75%, respectively). These two species have the potential to cause significant infections in human, ranging from opportunistic pulmonary infection to localized skin infection. Additionally, pathogenic NTM members of M. fortuitum group (MFG), M. avium, M. intracellulare, M. kansasii, and M. celatum were also found among all identified species.

Conclusion: Sequence-based analysis is a promising method for identifying species of NTM. The hsp65 gene has a high discriminatory power to identify opportunistic pathogen NTM species in specimens in Indonesia. Consequently, hsp65 partial gene sequencing is considerable as an alternative and reliable approach for NTM speciation.

Keywords: hsp65 gene; nontuberculous mycobacteria; sequence-based analysis; species identification.