Metronidazole-resistant Clostridioides difficile: genomic and transcriptomic traits acquired under in-vitro metronidazole induction

Teng Xu; Fenfen Zhou; Li Wang; Shi Wu; Haihui Huang

doi:10.1016/j.ijantimicag.2022.106570

Metronidazole-resistant Clostridioides difficile: genomic and transcriptomic traits acquired under in-vitro metronidazole induction

Int J Antimicrob Agents. 2022 May;59(5):106570. doi: 10.1016/j.ijantimicag.2022.106570. Epub 2022 Mar 13.

Authors

Teng Xu¹, Fenfen Zhou¹, Li Wang¹, Shi Wu², Haihui Huang³

Affiliations

¹ Institute of Antibiotics, Huashan Hospital, Fudan University, Shanghai, China; Key Laboratory of Clinical Pharmacology of Antibiotics, National Health and Family Planning Commission, Shanghai, China.
² Institute of Antibiotics, Huashan Hospital, Fudan University, Shanghai, China; Key Laboratory of Clinical Pharmacology of Antibiotics, National Health and Family Planning Commission, Shanghai, China. Electronic address: wu_shi@fudan.edu.cn.
³ Institute of Antibiotics, Huashan Hospital, Fudan University, Shanghai, China; Key Laboratory of Clinical Pharmacology of Antibiotics, National Health and Family Planning Commission, Shanghai, China. Electronic address: huanghaihui@fudan.edu.cn.

PMID: 35296432
DOI: 10.1016/j.ijantimicag.2022.106570

Abstract

Decreased effectiveness of metronidazole for the treatment of Clostridioides difficile infection has been documented. One reason for this is that levels of metronidazole in the colon are generally low; therefore, a modest increase in the minimum inhibitory concentration of metronidazole for C. difficile may result in an insufficient therapeutic concentration. Due to the lack of efficient genetic manipulation tools for C. difficile strains, the resistance mechanism is largely unknown. In this study, a metronidazole-resistant strain (SH182IR) was acquired by in-vitro induction with metronidazole from a clinical metronidazole-heteroresistant strain (SH182), and the genomic and transcriptional changes were investigated through whole-genome sequencing and RNA-seq. The morphology of the two strains was studied by transmission electron microscopy, and the roles of drug efflux pumps in metronidazole resistance were determined by inhibition assay. Genomic analysis showed that the ferrous iron transporter feoB3 was truncated in SH182IR, indicating that feoB3 contributed to the metronidazole resistance of C. difficile. RNA-seq analysis showed that genes involved in peptidoglycan synthesis, efflux pumps and metronidazole reductive action were expressed differentially between the two strains. Further cell imaging confirmed that cell wall thickness was significantly greater in SH182IR. The efflux pump inhibitor test showed that addition of reserpine or cyanide 3-chlorophenylhydrazone reduced metronidazole resistance in SH182IR, thus proving the role of efflux pumps in metronidazole resistance. These results found an association between genomic variation and metronidazole resistance in C. difficile, and show that metronidazole resistance in C. difficile is multi-factorial, involving metronidazole metabolism, cell wall thickness and efflux pumps. These findings will help improve knowledge and understanding of metronidazole resistance of C. difficile.

Keywords: Clostridioides difficile; Mechanism; Metronidazole; Resistance; Sequence.

MeSH terms

Anti-Bacterial Agents / pharmacology
Anti-Bacterial Agents / therapeutic use
Clostridioides
Clostridioides difficile* / genetics
Clostridium Infections* / drug therapy
Genomics
Humans
Metronidazole / pharmacology
Metronidazole / therapeutic use
Microbial Sensitivity Tests
Transcriptome

Substances

Anti-Bacterial Agents
Metronidazole