Periodic Reporting for period 1 - DiAMonD (DiAMonD - DIscovering Active MicroOrgaNisms implicated in adverse Drug effects)
Okres sprawozdawczy: 2024-07-01 do 2026-06-30
The DiAMonD (DIscovering Active MicroOrgaNisms implicated in adverse Drug effects) project explored how the human skin microbiome influences adverse reactions to fluoropyrimidine chemotherapy, a class of drugs including 5-fluorouracil (5FU) and capecitabine. Hand–Foot Syndrome (HFS), a painful skin toxicity, is a common side effect of these treatments. The project aimed to identify microbial genes, species, and mechanisms that contribute to either protection from or susceptibility to HFS.
Due to limited patient sample availability, the fellow implemented a controlled experimental-evolution approach using skin microbiomes from healthy donors. This strategy allowed the discovery of key mechanisms of microbial adaptation to 5FU, including the activation of prophages that induce bacterial cell death and mutations in nucleotide metabolism and cell-envelope genes that promote drug tolerance. Exposure to 5FU caused microbiome dysbiosis, leading to community imbalance, collapse of sensitive strains, and selection of resistant lineages. The project demonstrated that microbial community structure, strain-level genetics, and phage activity jointly determine the response to chemotherapy, revealing novel biological insights into microbiome–drug interactions that are relevant for understanding and potentially mitigating chemotherapy-induced skin toxicity.
Due to limited patient sample availability, the fellow implemented a controlled experimental-evolution approach using skin microbiomes from healthy donors. This strategy allowed the discovery of key mechanisms of microbial adaptation to 5FU, including the activation of prophages that induce bacterial cell death and mutations in nucleotide metabolism and cell-envelope genes that promote drug tolerance. Exposure to 5FU caused microbiome dysbiosis, leading to community imbalance, collapse of sensitive strains, and selection of resistant lineages. The project demonstrated that microbial community structure, strain-level genetics, and phage activity jointly determine the response to chemotherapy, revealing novel biological insights into microbiome–drug interactions that are relevant for understanding and potentially mitigating chemotherapy-induced skin toxicity.
Skin microbiome communities from four donors were cultured and exposed to 5FU in replicated experiments lasting one month, with daily transfers and time-series sampling. Metagenomic sequencing (Illumina short reads and PacBio HiFi long reads at time zero) was used to reconstruct microbial genomes (MAGs) and track evolutionary and community dynamics over time.
A total of 50 bacterial strains were isolated and sequenced, providing a culture collection representative of the experimental communities. The fellow analysed genetic changes within populations and identified both mutations and phage-related processes associated with drug tolerance.
Key findings include:
• Prophage induction by 5FU: several bacterial strains carrying dormant phages (prophages) showed reduced survival when exposed to the drug, and laboratory tests confirmed that 5FU can trigger phage activation, leading to bacterial death, whereas a strain carrying a pseudogenized prophage did not undergo induction and therefore survived.
• Adaptive mutations: tolerant strains accumulated mutations in genes involved in nucleotide metabolism (such as upp and tdk) and in cell-envelope regulation (fmtA, bceB, yycHI, acsA), suggesting changes that reduce drug activation or uptake.
• Community collapse and species replacement: some microbial communities collapsed early under 5FU, while others exhibited replacement of sensitive Staphylococcus epidermidis strains by more resistant S. warneri or S. pasteuri.
• Distinct network and genomic responses across communities, revealing how microbial interactions and genetic backgrounds influence adaptation to chemotherapy drugs.
Together, these results provide new evidence that 5FU not only exerts selective pressure on bacterial populations but also induces prophage activity that reshapes microbial community structure — an overlooked mechanism potentially relevant to skin toxicity in patients.
A total of 50 bacterial strains were isolated and sequenced, providing a culture collection representative of the experimental communities. The fellow analysed genetic changes within populations and identified both mutations and phage-related processes associated with drug tolerance.
Key findings include:
• Prophage induction by 5FU: several bacterial strains carrying dormant phages (prophages) showed reduced survival when exposed to the drug, and laboratory tests confirmed that 5FU can trigger phage activation, leading to bacterial death, whereas a strain carrying a pseudogenized prophage did not undergo induction and therefore survived.
• Adaptive mutations: tolerant strains accumulated mutations in genes involved in nucleotide metabolism (such as upp and tdk) and in cell-envelope regulation (fmtA, bceB, yycHI, acsA), suggesting changes that reduce drug activation or uptake.
• Community collapse and species replacement: some microbial communities collapsed early under 5FU, while others exhibited replacement of sensitive Staphylococcus epidermidis strains by more resistant S. warneri or S. pasteuri.
• Distinct network and genomic responses across communities, revealing how microbial interactions and genetic backgrounds influence adaptation to chemotherapy drugs.
Together, these results provide new evidence that 5FU not only exerts selective pressure on bacterial populations but also induces prophage activity that reshapes microbial community structure — an overlooked mechanism potentially relevant to skin toxicity in patients.
The DiAMonD project generated results that go beyond the current state of the art by uncovering new mechanisms through which chemotherapy drugs, particularly 5-fluorouracil (5FU), interact with and reshape bacterial communities. Previous studies had suggested that microbiome alterations could influence chemotherapy outcomes, but the underlying processes were largely unknown. Through a combination of metagenomics, strain isolation, and experimental evolution, the project revealed that 5FU can directly trigger prophage induction in bacterial genomes, leading to lytic activation and bacterial death. Crucially, these effects were demonstrated across multiple Staphylococcus species and strains, including S. epidermidis, S. hominis, and S. pasteuri, each showing distinct responses depending on their prophage complement and genetic background. Strains carrying pseudogenized or non-functional prophages did not undergo activation, highlighting the evolutionary consequences of prophage decay and mutational inactivation within staphylococcal genomes.
Beyond individual strain responses, the project demonstrated that 5FU exposure reshapes microbial community structure and interactions, driving shifts toward disbiotic configurations characterized by altered phage–bacteria dynamics and competitive imbalances. By integrating genomic and ecological perspectives, DiAMonD established a conceptual and experimental framework for understanding microbiome resilience and adaptation under drug pressure. This approach, bridging evolutionary microbiology and biomedicine, represents a significant advancement in how microbiome–drug interactions are studied and interpreted.
The potential impact of these findings extends to clinical, scientific, and societal domains. From a clinical standpoint, identifying microbial and phage factors that modulate 5FU toxicity opens opportunities for predictive diagnostics and microbiome-informed interventions to reduce adverse effects such as Hand–Foot Syndrome. Scientifically, the project contributes new data and hypotheses to the emerging field of chemotherapeutic–microbiome interactions, with implications for oncology, microbiology, and microbial ecology. On a broader level, by elucidating how microbial ecosystems respond to human-introduced chemical stressors, the project supports European priorities on health, innovation, and sustainable biomedical research. Its outcomes promote safer and more personalized therapeutic approaches while advancing our understanding of microbial adaptation in the context of human health and environmental change.
Beyond individual strain responses, the project demonstrated that 5FU exposure reshapes microbial community structure and interactions, driving shifts toward disbiotic configurations characterized by altered phage–bacteria dynamics and competitive imbalances. By integrating genomic and ecological perspectives, DiAMonD established a conceptual and experimental framework for understanding microbiome resilience and adaptation under drug pressure. This approach, bridging evolutionary microbiology and biomedicine, represents a significant advancement in how microbiome–drug interactions are studied and interpreted.
The potential impact of these findings extends to clinical, scientific, and societal domains. From a clinical standpoint, identifying microbial and phage factors that modulate 5FU toxicity opens opportunities for predictive diagnostics and microbiome-informed interventions to reduce adverse effects such as Hand–Foot Syndrome. Scientifically, the project contributes new data and hypotheses to the emerging field of chemotherapeutic–microbiome interactions, with implications for oncology, microbiology, and microbial ecology. On a broader level, by elucidating how microbial ecosystems respond to human-introduced chemical stressors, the project supports European priorities on health, innovation, and sustainable biomedical research. Its outcomes promote safer and more personalized therapeutic approaches while advancing our understanding of microbial adaptation in the context of human health and environmental change.