Improving microbial fitness in the mammalian gut by in vivo temporal functional metagenomics |
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| Authors: | Stephanie J Yaung Luxue Deng Ning Li Jonathan L Braff George M Church Lynn Bry Harris H Wang Georg K Gerber |
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| Affiliation: | 1. Program in Medical Engineering and Medical Physics, Harvard‐MIT Division of Health Sciences and Technology, Massachusetts Institute of Technology, Cambridge, MA, USA;2. Department of Genetics, Harvard Medical School, Boston, MA, USA;3. Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, MA, USA;4. Center for Clinical and Translational Metagenomics, Department of Pathology, Brigham & Women's Hospital, Harvard Medical School, Boston, MA, USA |
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| Abstract: | Elucidating functions of commensal microbial genes in the mammalian gut is challenging because many commensals are recalcitrant to laboratory cultivation and genetic manipulation. We present Temporal FUnctional Metagenomics sequencing (TFUMseq), a platform to functionally mine bacterial genomes for genes that contribute to fitness of commensal bacteria in vivo. Our approach uses metagenomic DNA to construct large‐scale heterologous expression libraries that are tracked over time in vivo by deep sequencing and computational methods. To demonstrate our approach, we built a TFUMseq plasmid library using the gut commensal Bacteroides thetaiotaomicron (Bt) and introduced Escherichia coli carrying this library into germfree mice. Population dynamics of library clones revealed Bt genes conferring significant fitness advantages in E. coli over time, including carbohydrate utilization genes, with a Bt galactokinase central to early colonization, and subsequent dominance by a Bt glycoside hydrolase enabling sucrose metabolism coupled with co‐evolution of the plasmid library and E. coli genome driving increased galactose utilization. Our findings highlight the utility of functional metagenomics for engineering commensal bacteria with improved properties, including expanded colonization capabilities in vivo. |
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| Keywords: | commensal fitness functional metagenomics microbiota next‐ generation sequencing synthetic biology |
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