Shining light on the "dark metaproteome"
A substantial portion of gut microbial activity—often referred to as the “dark metaproteome”—has remained invisible to science. Traditional metaproteomic approaches struggle to capture proteins from low-abundance organisms, many of which play critical yet underappreciated roles in shaping microbial ecosystems and influencing host health.
uMetaP addresses this sensitivity gap by combining state-of-the-art mass spectrometry (timsTOF Ultra with DIA-PASEF) and novoMP, a purpose-built de novo sequencing pipeline designed to rigorously control false discoveries in complex spectral landscapes.
By going beyond conventional database searches and harnessing the full depth of available data from mass spectrometry analyses, uMetaP expands both taxonomic detection and functional annotation, capturing protein-level signals from bacteria, archaea, fungi, viruses, and host tissues that were previously undetectable. Feng Xian, first author of the paper, explains: "Our new sequencing pipeline provides enhanced visibility of these organisms. It opens the door to studying underappreciated microbial players and their contributions to health and disease at an unprecedented resolution."
Record sensitivity: down to parts-per-million microbial signals
To evaluate the detection limits in a realistic setting, the team conducted controlled "spike-in" experiments by adding two stable isotope-labeled bacterial colonies (Ligilactobacillus murinus and exogenous Salinibacter ruber) to mouse fecal samples. uMetaP successfully detected as few as 10,000 bacterial cells per 10 mg of feces, a detection threshold in the parts-per-million range relative to an estimated 10 billion native gut microbes. This level of performance marks a major step forward for metaproteomics, enabling the detection of rare microbial signatures that may disproportionately influence host–microbiome dynamics and diseases.
From mice with gut injuries to relevance for human diseases
Applying uMetaP to a transgenic mouse model with a colonic injury driven by mitochondrial dysfunction, researchers tracked changes in the microbiome and the hosts responses over time. They captured an early and pronounced increase in Bacteroides caecimuris and mapped microbial metabolic pathways linked to the injury. Comparative analysis of the mouse data with Crohn’s disease patient transcriptomes revealed hundreds of overlapping molecules, strengthening the relevance of the findings to human diseases and reinforcing uMetaP’s potential in bridging experimental and clinical research.
Introducing the "druggable metaproteome"
Going beyond discovery, the team introduced the concept of a druggable metaproteome—a map that integrates microbial functions, host protein hubs, and curated drug–gene interaction data to identify proteins and pathways that could be targeted pharmacologically. "By systematically combining metaproteomic profiles from the inflamed gut with host and microbial annotations – and the known drug–target relationships – we identified over 200 drug–protein interaction candidates", says David Gómez-Varela, Director of the Center of Excellence for Metaproteomics and main author of the study. "This includes not only current treatments for inflammatory bowel disease, but also novel targets and drugs with potential for repurposing, expanding the current therapeutic landscape massively."
The results also highlighted functional convergence between host and microbial networks—suggesting opportunities for dual-target strategies that modulate both sides of the host–microbiome interface. Taken together, this framework lays the foundation for using metaproteomic data to guide precision drug discovery in complex diseases like inflammatory bowel disease.
Original publication
Feng Xian, Malena Brenek, Christoph Krisp, Elisabeth Urbauer, Ranjith Kumar Ravi Kumar, Doriane Aguanno, Tharan Srikumar, Qixin Liu, Allison M. Barry, Bin Ma, Jonathan Krieger, Dirk Haller, Manuela Schmidt & David Gómez-Varela. Ultra-sensitive metaproteomics redefines the dark metaproteome, uncovering host-microbiome interactions and drug targets in intestinal diseases. In Nature Communications (https://www.nature.com/articles/s41467-025-61977-7)
Scientific Contact:
David Gomez Varela PhD
Center of Excellence for Metaproteomics University of Vienna - Bruker Daltonics
Division of Pharmacology & Toxicology
Department of Pharmaceutical Sciences
1090, Josef-Holaubek-Platz 2 (UZA II)
+43-1-4277-55361
david.gomez.varela@univie.ac.at
