ORIGINAL RESEARCH article
Front. Microbiol.
Sec. Antimicrobials, Resistance and Chemotherapy
Volume 16 - 2025 | doi: 10.3389/fmicb.2025.1629464
This article is part of the Research TopicImpacts of Metal and Xenobiotic-Induced Stress on Antibiotic Resistance in Microbial CommunitiesView all 6 articles
Differential Impacts of Hemin and Free Iron on Amoxicillin Susceptibility in Ex vivo Gut Microbial Communities
Provisionally accepted
- 1Brown University, Providence, United States
- 2Saint Louis University School of Medicine, Saint Louis, United States
- 3Hasbro Children's Hospital, Providence, United States
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The rise of antibiotic-resistant infections worldwide has created a need to enhance the efficacy of existing antibiotics. Modification of metabolism has been shown to potentiate antibiotic lethality. In this study, we employed a novel ex vivo microbiome culture approach to study the effects of different forms of iron on amoxicillin susceptibility. Synthetic and human stool-derived microbiota were cultured and treated with amoxicillin, with growth monitored by optical density. These samples were sequenced using an Oxford nanopore long-read 16S rRNA V4-V9 approach and computationally defined using the Emu algorithm. The validity of this pipeline was confirmed with consortia, murine cecal content, and a human stool sample. The stool-derived community was then cultured for 24h with ranging concentrations of either hemin, FeSO4, or FeCl3 and concurrent amoxicillin dosage, then profiled to identify the effects of different forms of iron on amoxicillin susceptibility. Alpha diversity, beta diversity, and normalized relative abundances confirmed the efficacy of the selected ex vivo pipeline, allowing for ~77% species retention over 24 h. Treatment of communities with hemin protected Bacteroides, Escherichia-Shigella, Parabacteroides, and Parasutterella against amoxicillin, while two forms of free iron did not. This ex vivo pipeline enables reproducible assessment of how metabolic modulators like hemin alter amoxicillin susceptibility, highlighting a link between iron-sequestering genera and antibiotic-protection. Future mechanistic insights may support hemin-based strategies to boost antibiotic efficacy.
Keywords: antibiotic susceptibility, ex vivo gut microbiome culture, iron metabolism, Microbial metabolism, Metabolic modulation, Beta lactam antibiotics, Hemin, 16S rRNA sequencing
Received: 15 May 2025; Accepted: 30 Sep 2025.
Copyright: © 2025 Pagano, Bemis, Rehman, Shapiro and Belenky. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
* Correspondence: Peter Belenky, peter_belenky@brown.edu
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