Stability and Resilience in the Human Microbiome: Response of Gut Microbiota to Colonic Lavage Laurie Rumker, Dr. Les Dethlefsen, Dr. David Relman Department of Microbiology and Immunology, Stanford University School of Medicine
Microbiota and the Human Body Ecosystem Experimental Design Methodology Study Aims
• = Fecal and Urine ! Aim 1. Compare rou ne unperturbed dynamics to varied • Bacteria contribute 90% of cells, Samples Taken ! responses to lavage; characterize effect of disturbance 99% of genes in human body Colonic • Roles of microbiota in health: Lavage! • Aim 2. Evaluate applica on of ecological principles to human gut - Pathogen resistance microbiota in context of colonic lavage disturbance - Metaboliza on - Xenobio c modifica ons Baseline Stability! Disturbance! Recovery Dynamics! (Weekly Sampling)! (Daily Sampling)! (Weekly Sampling)! • Aim 3. Iden fy features characteris c of resilient communi es - Immune system regula on • Contributes to malnutri on, obesity, heart disease, • Study Subjects: 8 healthy volunteer adult par cipants Body Sites and Predominant inflammatory bowel disease, • Fecal and urine samples at each me point analyzed three ways: Proof of Concept: An bio c Disturbance Taxa of the Microbiota Crohn’s disease, many more (Dethlefsen and Relman, 2007) Fluctua ons of α-Diversity Metrics in Response to an An bio c Series • Marker gene amplified, sequenced 16S rDNA • Conserved iden fica on tag An bio cs An bio cs Research Ques ons Sequencing • Reveals changes in taxonomic 80 composi on, rela ve abundances 60 • How consistently does the microbiome retain its composi on and 40 ac vity over me? 20 • Fragmented, sequenced whole Rela ve Diversity • 0 How easily can the health benefits conferred upon a host by the Shotgun genome microbiota be lost due to a disturbance of the community? 0 50 100 150 200 250 300 350 Metagenomic • Reveals rela ve gene frequencies Day of Sampling Sequencing • Computa onal reassembly associates Phylogene c Diversity Applying Macroecological Theory genes with specific taxa Taxonomic Diversity (Shannon Index, Values *10) OTU Richness (Values/10) • Stability = ability of system to return to equilibrium quickly, with • An bio c series triggered drama c temporary decrease in α- minimal devia on a er temporary disturbance • Iden fies small molecules produced diversity metrics based on 16S rDNA data (Dethlefsen, 2011) • Equilibrium = set of defined values for relevant key variables Metabolomic Mass • Present study expands upon this one with longer mescale, Spectrometry • (Analyses performed by collaborators • Resilience = when challenged with disturbance, ecosystem ability at University College London) greater sampling depth, use of metagenomics and metabolomics to adapt and retain func on, structure, iden ty, and feedbacks • Colonic lavage is less selec ve disturbance causing greater • Disturbance could induce regime shi to new stable state decrease in total biomass
Ecosystem Project Status
Varia on Around Stable Equilibrium Alterna ve Subjects DNA Send Stable State Design Study Recruit Metagenomic Data Processing Sample Extrac on 16S Metabolomics and Obtain and Enroll Sequencing and with QIIME and and from Fecal Sequencing Urine and Fecal Ecosystem Stable States or “Basins of A rac on” IRB Approval Subjects Reassembly Phyloseq Cleanout Samples Samples • Possible predictors of microbiome resilience: - Taxonomic diversity - Intermediate stability week-to-week or day-to-day 3 par cipants currently sampling, 5 par cipants’ samples DNA extracted, 16S sequenced - Func onal redundancy (the Insurance Hypothesis) about to undergo cleanout, will complete on HISeq2500 with 150nt paired-reads, produced - Specific constella on of taxa sampling December 2014 >98 million reads, metagenomics prepara ons ongoing