Details

Autor(en) / Beteiligte

• Deshpande, Aditi
• Wu, Xiaoqian
• Huo, Wenwen
• Palmer, Kelli L
• Hurdle, Julian G

Titel

Chromosomal Resistance to Metronidazole in Clostridioides difficile Can Be Mediated by Epistasis between Iron Homeostasis and Oxidoreductases

Ist Teil von

• Antimicrobial agents and chemotherapy, 2020-07, Vol.64 (8)

Ort / Verlag

United States: American Society for Microbiology

Erscheinungsjahr

2020

Link zum Volltext

Quelle

MEDLINE

Beschreibungen/Notizen

• Chromosomal resistance to metronidazole has emerged in clinical isolates, but the genetic mechanisms remain unclear. This is further hindered by the inability to generate spontaneous metronidazole-resistant mutants in the lab to interpret genetic variations in clinical isolates. We therefore constructed a mismatch repair mutator in nontoxigenic ATCC 700057 to survey the mutational landscape for resistance mechanisms. In separate experimental evolutions, the mutator adopted a deterministic path to resistance, with truncation of the ferrous iron transporter FeoB1 as a first-step mechanism of low-level resistance. Deletion of in ATCC 700057 reduced the intracellular iron content, appearing to shift cells toward flavodoxin-mediated oxidoreductase reactions, which are less favorable for metronidazole's cellular action. Higher-level resistance evolved from sequential acquisition of mutations to catalytic domains of pyruvate-ferredoxin/flavodoxin oxidoreductase (PFOR; encoded by ), a synonymous codon change to putative (xanthine dehydrogenase; encoded by ), likely affecting mRNA stability, and last, frameshift and point mutations that inactivated the iron-sulfur cluster regulator (IscR). Gene silencing of , , or with catalytically dead Cas9 revealed that resistance involving these genes occurred only when was inactivated; i.e., resistance was seen only in the deletion mutant and not in the isogenic wild-type (WT) parent. Interestingly, metronidazole resistance in infection (CDI)-associated strains carrying mutations in was reduced upon gene complementation. This observation supports the idea that mutation in PFOR is one mechanism of metronidazole resistance in clinical strains. Our findings indicate that metronidazole resistance in is complex, involving multigenetic mechanisms that could intersect with iron-dependent and oxidoreductive metabolic pathways.