Protein composition and methods for analysing microbiota
Inventors
FIGEYS, Joseph Michel Daniel • STINTZI, Alain Christophe • Mack, David R. • Zhang, Xu • NING, Zhibin
Assignees
Publication Number
US-11175294-B2
Publication Date
2021-11-16
Expiration Date
2037-06-01
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Abstract
A method of isotope-labelling a microbiota sample. It involves providing a first microbiota sample that was obtained from a given source; exposing the first microbiota sample to an isotope enriched medium; and culturing the exposed first microbiota sample in the isotope enriched medium to obtain an isotope-labelled microbiota sample, wherein the isotope labelled metaproteome of the isotope-labelled microbiota sample is taxon specific for taxa present in the first microbiota sample when initially obtained from the given source.
Core Innovation
The invention provides a fast, cost-effective method called stable isotopically labelled microbiota (SILAMi) for metabolic labeling of whole microbiota samples obtained from various sources, including human, animal, soil, plant, water, or industrial settings. SILAMi overcomes previous limitations by achieving high isotope incorporation across diverse and complex microbial populations, including anaerobic species, thereby producing isotope-labelled microbiota protein standards that are taxon-specific and representative of the original microbiota sample composition.
The method involves exposing a microbiota sample obtained from a source to an isotope-enriched medium and culturing it to incorporate the isotope into microbial proteins, resulting in a labelled microbiota sample with an isotope-labelled metaproteome specific to microbes present in the initial sample. Importantly, SILAMi enables accurate and reproducible quantitative metaproteomic analysis by providing an internal isotope-labelled standard that reduces variability across experiments and allows measurement of protein abundance changes in microbiota samples.
The problem addressed is the difficulty in applying existing metabolic labeling techniques to complex microbiota due to species diversity, variable metabolic capacities, oxygen sensitivity, and differing growth rates of microbes, which precluded faithful isotope labeling and quantitative proteomic analysis. The invention solves this by demonstrating that effective isotope labeling of microbiota populations is achievable within a practical timeframe while maintaining taxonomic representativity, thereby enabling improved functional characterization of microbiomes.
Claims Coverage
The patent presents one independent claim detailing a high throughput screening method for metaproteomic analysis of multiple microbiota samples.
High throughput screening of multiple microbiota samples
Culturing a plurality of microbiota samples from human, soil, or animal sources, spiking these cultures with an isotope-labelled microbiota standard, performing metaproteomic analysis to identify changes in microbiomes, selecting microbiomes exhibiting changes, and analyzing selected microbiomes to characterize those changes.
Use of micro-well receptacles for culturing
Culturing the plurality of microbiota samples in micro-well receptacles or plates to facilitate high throughput parallel processing.
Advanced analysis using microbial gene catalogs and iterative database searches
Analyzing metaproteomic data with microbial gene catalogs specific to subject type (human or animal) combined with iterative database search strategies, optionally integrating metagenomic analysis.
Isotope-labelled standard with high enrichment and taxon specificity
Using an isotope-labelled standard with at least 90% average heavy isotopic enrichment rate, taxon-specific for at least 50% to 85% of microbes in the samples, encompassing multiple microbiota types and employing stable and/or radioactive isotopes such as 13C, 14C, 15N, 32S, 35S, 32P, or deuterium.
Applications of metaproteomic screening results
Assessing disease diagnosis, treatment response, remission, screening effects of xenobiotics or compounds (foods, drugs, chemicals, cosmetics, pesticides, consumer products, etc.) on microbiomes, and identifying subject responsiveness to therapies or treatments based on analysis of microbiome changes.
The claim covers a comprehensive high throughput metaproteomic screening method employing isotope-labelled microbiota standards cultured in micro-well formats, combined with sophisticated microbial gene catalog analyses to identify and characterize microbiome changes for diverse applications including diagnostics, treatment monitoring, and compound screening.
Stated Advantages
SILAMi is a fast (approximately 3 days), efficient, and cost-effective approach for metabolic labeling of diverse microbiota populations.
The invention allows accurate, standardized, and reproducible quantitative metaproteomic analysis, reducing variability across experiments.
It enables functional analysis of microbiome responses to treatments and compounds, identifying specific pathways and metabolic process alterations.
The method supports high throughput screening of microbiota samples from multiple individuals or sources in parallel using micro-well plate formats.
SILAMi facilitates disease diagnosis, treatment response assessment, and screening compound effects on microbiomes, aiding precision medicine and microbiome research.
Documented Applications
Quantitative assessment of changes in microbiome protein expression following treatment with compounds such as fructooligosaccharide (FOS) or monosaccharides.
High throughput screening of microbiota samples from humans, animals, or soil to evaluate effects of drugs, xenobiotics, foods, chemicals, cosmetics, packaging components, pesticides, and other consumer products on microbiome composition and function.
Disease diagnosis including intestinal diseases like inflammatory bowel disease (IBD) by measuring protein abundance deviations in microbiome samples.
Assessing treatment response and remission status in patients by analyzing metaproteomic changes in microbiota over time.
Rapid screening platform (RapidAIM) for identifying biota-affecting or biota-altered compounds and stratifying patients' microbiome-mediated responses to therapeutics.
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