Metronidazole: an update on metabolism, structure–cytotoxicity and resistance mechanisms Original paper

What was reviewed?

The review examined the metabolic pathways, cytotoxic effects, and resistance profiles of metronidazole across microbial and mammalian systems. The authors synthesized decades of research, drawing from in vitro assays, murine models, and clinical microbiology to clarify how metronidazole’s reductive activation generates transient intermediates responsible for its antimicrobial and potential cytotoxic effects. Importantly, they explored how these processes interact with microbial physiology, redox balance, and drug resistance mechanisms, providing an advanced understanding of how this common antimicrobial agent exerts both therapeutic and off-target effects—especially in the gut microbiome.

Who was reviewed?

The review incorporated findings from bacterial isolates (both clinical and laboratory strains), murine models, and in vitro cellular systems. It emphasized studies involving obligate anaerobes like Clostridium difficile, Bacteroides fragilis, and Helicobacter pylori, as well as facultative anaerobes such as E. coli. It also referenced experiments in germ-free rodents to illustrate microbiome-dependent metabolism and toxicokinetics. Human pharmacokinetic data complemented these findings, particularly regarding drug absorption, tissue distribution, and metabolite excretion. The authors also discussed mutants with defective DNA repair pathways, shedding light on host and microbial responses to reductively activated metronidazole.

What were the most important findings?

The review clarified that metronidazole functions as a prodrug, requiring reductive activation by microbial enzymes under low-oxygen conditions. Key enzymes involved include pyruvate:ferredoxin oxidoreductase (PFOR), ferredoxin, and flavodoxin, primarily in anaerobic bacteria. The reductive pathway generates unstable metabolites such as nitroso and hydroxylamine intermediates, which can damage microbial DNA either directly or indirectly by altering redox balance and inhibiting DNA repair enzymes like ribonucleotide reductase.

Crucially for the microbiome field, the authors demonstrated that gut bacteria play a critical role in metabolizing metronidazole, with activation occurring mainly in the caecum, home to dense anaerobic populations. In germ-free rodents, the typical reductive metabolites of metronidazole are absent, confirming microbiota dependence. Moreover, metronidazole alters mucosal structure, modulates innate immune responses (increasing IL-25, RegIII), and disrupts colonization resistance, allowing temporary overgrowth of opportunistic pathogens like C. difficile, E. coli, and Citrobacter rodentium. These shifts emphasize the drug’s broad but non-neutral impact on microbial ecology.

The review also offered a detailed mechanistic explanation of resistance pathways, including efflux pumps, reduced drug uptake, inactivation by nim genes, and shifts in energy metabolism that avoid drug activation. Although high-level resistance remains uncommon, the review noted rising multidrug-resistant (MDR) strains in Bacteroides and Prevotella in certain global regions. Importantly, many resistance phenotypes occur independently of nim genes, challenging the assumption that nim carriage alone predicts resistance.

What are the implications of this review?

This review establishes a nuanced model of metronidazole action and resistance, reinforcing its clinical efficacy while highlighting its non-selective microbiome impact and potential for cytotoxicity beyond target pathogens. For clinicians and microbiome researchers, the key implication is that while metronidazole remains an essential tool for treating anaerobic infections, it disrupts beneficial taxa and weakens colonization resistance, potentially increasing the risk for pathogen overgrowth and inflammation.

Moreover, the identification of strain-specific resistance mechanisms, often independent of nim genes, supports more individualized microbiome-aware therapy, ideally guided by antibiograms and molecular screening. The review also emphasized the need for developing next-generation nitroimidazoles with pathogen-specific uptake or activation strategies to reduce collateral damage to the microbiome. In the microbiome signatures context, this review reinforces the need to document both taxa-specific alterations and host immunological shifts following metronidazole use to refine therapeutic strategies.