Project description:Metagenome-assembled genomes (MAGs) have revealed the existence of novel bacterial and archaeal groups and provided insight into their genetic potential. However, metagenomics and even metatranscriptomics cannot resolve how the genetic potential translates into metabolic functions and physiological activity. Here, we present a novel approach for the quantitative and organism-specific assessment of the carbon flux through microbial communities with stable isotope probing-metaproteomics and integration of temporal dynamics in 13C incorporation by Stable Isotope Cluster Analysis (SIsCA). We used groundwater microcosms labeled with 13CO2 and D2O as model systems and stimulated them with reduced sulfur compounds to determine the ecosystem role of chemolithoautotrophic primary production. Raman microspectroscopy detected rapid deuterium incorporation in microbial cells from 12 days onwards, indicating activity of the groundwater organisms. SIsCA revealed that groundwater microorganisms fell into five distinct carbon assimilation strategies. Only one of these strategies, comprising less than 3.5% of the community, consisted of obligate autotrophs (Thiobacillus), with a 13C incorporation of approximately 95%. Instead, mixotrophic growth was the most successful strategy, and was represented by 12 of the 15 MAGs expressing pathways for autotrophic CO2 fixation, including Hydrogenophaga, Polaromonas and Dechloromonas, with varying 13C incorporation between 5% and 90%. Within 21 days, 43% of carbon in the community was replaced by 13C, increasing to 80% after 70 days. Of the 31 most abundant MAGs, 16 expressed pathways for sulfur oxidation, including strict heterotrophs. We concluded that chemolithoautotrophy drives the recycling of organic carbon and serves as a fill-up function in the groundwater. Mixotrophs preferred the uptake of organic carbon over the fixation of CO2, and heterotrophs oxidize inorganic compounds to preserve organic carbon. Our study showcases how next-generation physiology approach like SIsCA can move beyond metagenomics studies by providing information about expression of metabolic pathways and elucidating the role of MAGs in ecosystem functioning.
Project description:Groundwater samples were collected from five wells in Alberta, Canada. The sampling location and time are indicated in the table below. 13 to 29 genomes were assembled from each metagenome. Proteomic analyses were performed to investigate which genomes and genes were expressed in each well. Sample ID Location Latitude (NAD 83) Longitude (NAD 83) Sampling date 19GWC19026 218 Cluny 50.85 -112.84 30/07/2019 19GWC19028 114 Ross Creek 49.99 -110.46 31/07/2019 19GWC19045 265 Metiskow 52.42 -110.61 18/09/2019 19GWE00050 991 Cynthia 56.22 -117.81 04/09/2019 19GWE00515 33 Fort McMurray 56.98 -111.85 17/09/2019