WP 3.1 Impact of Drugs on Microbiomes in Humans and Wastewater Treatment

project description

Human gut and wastewater microbiomes interact with human-targeted and antibiotic drugs in multiple ways, which can have profound consequences for the treatment of patients and the health of ecosystems that receive drugs. Drug-microbe interactions result in the inhibition or stimulation of members of the gut microbiome with multiple consequences for host health, and the spread of antibiotic resistance (caused also by non-antibiotic drugs). Furthermore, human gut and environmental microbiomes biotransform drugs and produce new metabolites with altered bioactivities, which enter wastewater treatment systems and the environment as micropollutants.

Our knowledge of the effects of drugs on human gut microbiomes largely stems from either incubation experiments of single drugs with pure microbial cultures, or from large cohort studies that correlate drug profiles with fecal microbiome composition data. Similarly, biotransformation of drugs by sewage and wastewater treatment microbiomes is mostly either analyzed with isolated microbes or by correlating changes in community composition or metatranscriptomes with biotransformation rates. Here, we hypothesize that drug-induced inhibition/stimulation patterns of microorganisms can vary drastically between pure cultures and complex microbiomes. Furthermore, we postulate that (i) exposure of microbiomes to combinations of drugs (and biotransformation products) results in effects not predictable from experiments with single compounds and that (ii) due to microbiome interaction networks, drug-induced changes in microbiome composition are not sufficient to reveal microbiome members directly affected by the drug.

To investigate these hypotheses, we will use our recently developed D₂O-based activity labeling in combination with stimulated Raman spectroscopy (SRS) and fluorescence in-situ hybridization (FISH) for a single-cell, in-situ, and high-throughput assessment of direct inhibition and stimulation patterns of bacterial and archaeal members of colon, small intestine, and sewage and wastewater treatment microbiomes upon exposure to the most commonly used drugs and drug combinations in Austria. These analyses will be complemented by (i) metagenomics and metatranscriptomics data from incubation experiments and selected patient cohorts to examine the in-vivo direct and indirect effects of drugs on microbiome composition and function as well as the spread of resistance markers, (ii) single- cell bioaccumulation investigations based on chemical imaging, to assess drug availability, and (iii) a biomimetic human gut-on-a-chip system, to study the effects of drugs on gut barrier functionality.

In parallel with the original drugs, we will also investigate the effects of biotransformation products by exposing the drugs to (i) enzyme extracts derived from the systems of interest and (ii) the InfoGest static in-vitro digestion simulation procedure, which simulates transfer through the upper digestive tract. Biotransformation kinetics and pathways will be studied using high-resolution mass spectrometry. To identify enzymes and derive links between enzyme abundance and biotransformation rates, we will develop workflows based on metaproteomics and activity-based protein-profiling. For key enzymes, in- vitro studies and structural characterizations will be conducted to assess substrate specificity, inhibition by drugs and biotransformation products, and potential for biotechnological optimization.

The expected results will substantially improve our understanding of drug-microbiome interactions and reveal the in-situ landscape of drug inhibition/stimulation in drug-affected systems. These insights will provide guidance for personalized therapy (e.g., drug selection and probiotic treatments), drive the (re-)design of effective drugs, and pave the way for innovative technologies (e.g., enzymatic inactivation of antibiotics in hospital wastewaters) to reduce the release of drugs into natural systems.

work package leader