Lecture 4 - Microbiome Flashcards
What is microbiota?
- We are not alone - contact with numerous microorganisms daily
- Most microbes are not pathogenic
– Balancebetweencommunities - Some are found natively on/in our body
- Some are introduced suddenly
- ‘Our other genome’ (Nature, 2010)
- ‘Your inner ecosystem’ (Scientific American, 2012)
“signify the ecological community of commensal, symbiotic and pathogenic microorganisms that literally share our body space“
-> Bacteria (bacteriome), viruses (virome), fungi (mycobiome), ‘tiny other organisms’ (e.g., helminths, protozoans)
-> Microbiome = Bacteriome + Virome + Mycobiome + tiny others
Size of the microbiota
Most microbiomes are 10-100x smaller than human cells (mitochondria size)
- About 1014 cells
- About same number as human cells (1.3x) Sender et al. 2016 PLoS Biol; Nature News Jan 2016
- 150x more genes
- About 0.2-2 kg of the human body mass
- ‘Forgotten organ’
Human microbiome
- Associated with diseases
– Diabetes
– Rheumatic disorder
– Multiple sclerosis
– Parkinson
– Neurodermatitis
– Psoriasis
– Autism
– … - Cause or consequence / link microbial biomarker with causation
– Helicobacter pylori associated peptic ulceration and gastric cancer
– Clostridium difficile infection-associated diarrhea - Symbiosis between host and microbiota (not only in human)
What is a healthy (gut) microbiome?
Complex combination of environmental, genetic & lifestyle factors
“One person’s healthy microbiome might not be healthy in another context - it’s a tricky concept”
“There won’t be “the” healthy microbiome, just like there’s no perfect genome”;
“There could be multiple healthy configurations.”
’Our findings suggest that gut taxonomic signatures can predict health status, and highlight how data sharing efforts can provide broadly applicable discoveries.’
A predictive index for health status using species- level gut microbiome profiling
– Gupta et al. (Nat. Comm. 2020)
Body site differences
‘Diversity of the human microbiome is concordant among measures, unique to each individual, and strongly determined by microbial habitat.’
Cultural differences
Differences in the faecal microbial communities of Malawians, Amerindians and US adults.
Human gut microbiome
- ‘Gut feeling’ (vagus nerve)
- Heavily influenced by what you eat
– Rapidly reacts to changes in diet
– Leveloffat ,carbohydrates, fiber, sugar, artificial sweeteners,… - Changes with age
- Responses to stress
- Altered in presence of diseases, antibiotics, …
-> Mostly influenced by environmental factors and not genetics
GENUS LEVEL
Diversity changes after birth
PHYLUM LEVEL
Diversity changes through lifetime
Food & Gut microbiome
-> Modulation of the gut microbiome
- High-fiber diet changes microbiome function
- Fermented-food diet increases microbiome diversity
- Calorie restriction leads to increase in metabolic health BUT bacterial diversity decreases (C. difficile enrichment)
Food & Gut microbiome
-> Modulation of the gut microbiome
- High-fiber diet changes microbiome function
- Fermented-food diet increases microbiome diversity
- Calorie restriction leads to increase in metabolic health BUT bacterial diversity decreases (C. difficile enrichment)
Gut brain axis
- Bacteria produce
– Short-chain fatty acids
– Secondary bile acids
– Tryptophan metabolites - Problem
1. Intestinal barrier
2. Blood-Brain barrier
-> Some products can cross the barriers - Products
– Oxytocin (L. reuteri)
– GABA (L. rhamnosus)
– Brain-derived neurotrophic factor (BDNF; B. longum NCC3001)
– SCFA (SCFA-producing bacteria)
Human skin microbiome
- Skin – border to the outer world
- Microorganisms are protective
- Very different regional composition
– Moist - inter-digital
– dry - forearm, lower leg
– Sebaceous - nose, neck, shoulders
Microbiome sequencing
- Commonly used marker: 16S rRNA gene
– ~1.3 kbp long
– Conserved secondary structure
-> prevents the sequence from accumulating too much diversity
– Nine hypervariable regions (V1-V9)
– Bacteria and archaea (but different primers)
– Next generations sequencing -> V1-V2, V3-V4 or V4 - Marker for fungi: 18S rRNA, ITS
- Marker for eukaryotes: COI (Cytochrome C oxidase subunit I)
1) Sampling
- 16S rRNA Gene
2) DNA isolation
- PCR amplification
3) Sequencing
4) fastq
5) Analysis (Preprocessing, Taxonomic assignment, Statistical analysis)
Data Preprocessing
- Sequencing data usually noisy and redundant
- pre processing steps
-> quality filtering (remove bad quality bases/reads)
-> merge overlaps
-> remove chimeric sequences
-> data dereliction (to reduce computational burden)
-> cluster data into (zero-radius) operational taxonomic units (OTUs) or amplicon sequence variants (ASVs)
-> Taxonomic assignment
-> OTU/ASV table
-> Phylogenetic tree reconstruction - (Popular) Software
-> Mothur
-> Dada2
-> USEARCH, vsearch
-> FastTree (fast phylogenetic tree reconstruction)
Biodiversity
‘variety within and among life forms on a site, ecosystem, or landscape’
1. Richness (R or S)
* Number of groups of genetically or functionally related individuals
-> number of species
2. Evenness (E)
* Relative abundance of different species
- Biodiversity can be measured at different scales
– Alpha diversity - Diversity within a given community
species richness, species evenness or combination of both
– Beta diversity - Partitioning of biological diversity among communities number of species shared between two communities
– Gamma diversity - Landscape diversity within a region
– (Bio-) Diversity: Low dimensional summary of communities
Biodiversity and microbial world
- Species concept
– Based on evolutionary theory
– Introduced by Ernst Mayr 1944
‘species is a group of organisms that remain phenotypically similar because of recombination’
Particularly problematic with bacteria and archaea
* Reproduce asexually
* Horizontal gene transfer (HGT) between distantly related species
-> Generally, do not talk about species but operational taxonomic units (OTUs)
* 97% sequence similarity threshold for 16S rRNA
– An OTU may be associated with several species
* Amplicon sequence variant (ASV; dada2) or Zero-radius OTUs (e.g., UNOISE3/USEARCH) (denoised sequencing reads)
– One species, multiple sequences (ASVs)
