Epidemiology of Nontuberculous Mycobacteria in Nanjing and MAB_0540 Mutations Associated with Clofazimine Resistance in Mycobacterium abscessus

Abstract

Background

Nontuberculous mycobacteria (NTM) are easily misdiagnosed as multidrug-resistant tuberculosis (MDR-TB), and treatment drugs are very limited. The main objective of our study was to evaluate the minimal inhibitory concentration (MIC) in vitro of bedaquiline (BDQ), clofazimine (CFZ), linezolid (LZD), delamanid (DLM), and pretomanid (PA-824) for treatment of M. abscessus and M. intracellulare. Furthermore, we determined whether MAB_1448, MAB_4384, MAB_2299c, MAB_1483, MAB_0540, rplD, rplC, and rrl were related to drug resistance to provide an experimental basis for the use of these five drugs in the treatment of NTM.

Methods

We identified sample characteristics of epidemics in 550 patients with suspected NTM infection in Nanjing from 2019 to 2021 using the PCR-reverse spot hybrid method. Furthermore, we evaluated the MIC of BDQ, CFZ, DLM, LZD, and PA-824 against 155 clinical isolates of NTM using the microbroth dilution method. The resistant isolates were sequenced using Sanger sequencing.

Results

The top three dominant species of NTM distributed in Nanjing were M. intracellulare, M. avium, and M. abscessus. Notably, the proportion of M. abscessus infections increased. The proportion of M. abscessus increased from 12% in 2019 to 18% in 2021. Demographic analysis showed that female infection rates were substantialy greater than male for M. abscessus (P=0.017, <0.05). Our results demonstrate that NTM are highly sensitive to bedaquiline and clofazimine in vitro. However, delamanid and pretomanid had little effect on M. abscessus and M. intracellulare. In addition, we found 30–41 nucleotide deletion mutations and some novel point mutations in the MAB_0540 gene of M. abscessus that are resistant to clofazimine.

Conclusion

Bedaquiline, clofazimine, and linezolid were more successful in vitro treatments against M. abscessus and M. intracellulare. The MAB_0540 mutation may be associated with resistance of M. abscessus to clofazimine.

Keywords:

• nontuberculous mycobacteria
• minimal inhibitory concentration
• gene mutations

Abbreviations

NTM, nontuberculous mycobacteria; MDR-TB, multi-drug resistant tuberculosis; MIC, minimal inhibitory concentration; BDQ, bedaquiline; CFZ, clofazimine; LZD, linezolid; DLM, delamanid; PA-824, pretomanid; MTC, Mycobacterium tuberculosis complex; CI, confidence interval; HIV, Human immunodeficiency virus; NTM-PD, nontuberculous mycobacteria pulmonary.

Data Sharing Statement

Data relating to this study are contained and presented in this document. Other materials are available from the corresponding author on reasonable request.

Ethics Approval and Informed Consent

The study protocol was approved by Nanjing Center for Disease Control and Prevention Ethics Committee (approval number: PJ2020-A001-04; approval date: 17 August, 2020). Patient information collected in the case system did not contain name, address or other personal information, so the patient’s written informed consent was exempt. The study was conducted in accordance with the Declaration of Helsinki.

Author Contributions

All authors made a significant contribution to the work reported, whether that is in the conception, study design, execution, acquisition of data, analysis and interpretation, or in all these areas; took part in drafting, revising or critically reviewing the article; gave final approval of the version to be published; have agreed on the journal to which the article has been submitted; and agree to be accountable for all aspects of the work.

Disclosure

The authors declare that they have no competing interests.

Additional information

Funding

This work was supported by Medical Science and Technology Development Foundation, Nanjing Department of Health (GrantZKX19048).