Section 1- The Human Microbiome

Question 1

Microbiome

Answer 1

The totality of microbes, including their genetic content (genomes), in a given environment
Live in close association with plants and animals
Usually only consider disease states of microbiome, but it is so much more
Organisms do not live in isolation; have evolved in context of complex communities

Question 2

Microbe

Answer 2

Bacteria, yeast, fungi, virus, etc.
Estimated to outnumber human cells by 3-10 to 1 in human body
Their total genes may exceed human genes by 100-300 to 1 (extreme functional content)
Many have not been cultured in a lab and their role in human physiology and disease is largely unknown
Community dynamic very important

Question 3

Dysbiosis

Answer 3

Disrupted relationship between living organisms

Opposite of symbiosis

Question 4

Human Microbiome Project (HMP)

Answer 4

Major initiative from NIH to catalog human microbiota and initiate the investigation of its role in human health and disease
Shows substantial intra-individual variation at different anatomical sites
Substantial inter-individual variation at the same anatomical site
Temporal stability in individuals at specific sites (stable through different conditions and times
Disruptions can have long-lasting effects

Question 5

Distribution of human microbiota

Answer 5

30% GI tract (most dense microbiome on earth)
Then oral, skin, airway, urogenital, blood, eye
GI tract contains 1000 or more phenotypes
Each individual has a unique diversity of flora (fingerprint), but functions are stable across individuals
Deeper in body, higher ratio of anaerobic microbes

Question 6

Main force in bacterial evolution over past 200 years

Answer 6

Unnatural (anthropogenic) selection

Question 7

How is information about microbial niche populations generated if most have never been cultured in the lab?

Answer 7

rRNA phylotyping
rRNA is the common denominator in all microbial genomes- is the ideal substrate for phylotyping
DNA sequence analysis of individual organisms or populations
Use 16S r RNA- every microbe has and sequence is evolutionarily conserved
Can analyze an entire population at once
A 1% change represents a new species

Question 8

How does one begin to study the microbiome?

Answer 8

Next generation DNA sequencers capable of generating 2-20 gigabases (billion bases) of sequence per run (=1 human)
1000’s of bacteria per run
Metagenomics

Question 9

Metagenomics

Answer 9

Analysis of DNA sequences within a particular environment (gut, nasopharynx, UG tract, environment)
Bacterial speciation within environment based on conserved DNA sequences (rRNA, conserved housekeeping genes)

Question 10

Gut microbiome

Answer 10

Know the most about the gut
Where the immune system begins to develop
Microbes break down foods we can’t
Most bacteria are Firmicutes and Bacteroidetes (>90%)
Substantial diversity between healthy individuals, esp. between infants; converges to more similar later in life
A few Proteobacteria and Actinobacteria

Question 11

Firmicutes

Answer 11

95% are Clostridial species (Botulism, C. diff)
Also Lactobacillus, Mycoplasma, Bacillus, and many butyrate-producers
Function in maintenance and protection of normal intestinal epithelium (produce short chain fatty acids)

Question 12

Bacteroidetes

Answer 12

Added to gut microbiome with solid food introduction
Highly variable species
Bacteroides thetaiotaomicron is most abundant species (opportunistic pathogen)
Function to help digest food

Question 13

Skin microbiome

Answer 13

Exogenous and endogenous factors contribute to variation between individuals over lifetime of the individual
Highly dependent on microenvironment of sampled site (Ex: dry vs moist)

Question 14

Vaginal microbiome

Answer 14

Lactobacillus characteristic of normal healthy reproductive aged women
Growth of non-indigenous organisms like pathogens is restricted
Protection due to low pH (<4.5), lactic acid, antimicrobial compounds, and other things
Human vagina has no single core microbiome; there are multiple that are defined by 5 community groups
Genetic component exists (by race)

Question 15

Ancient vs Modern mother to child microbiome transmission

Answer 15

Ancient: Oral (chewing food), Mammary (breastfeeding), Cutaneous (skin), Vaginal (birth canal)
Modern: Oral, Mammary, Cutaneous, Vaginal + Maternal exposures (Antiseptics, Antibiotics, Diet, Epigenetics), Bottle feeding (formula), Early-life antibiotics, C-section, Early/extensive bathing

Question 16

Changes in human behavior that can alter the microbiome composition

Answer 16

Clean water, C-sections, Preterm antibiotics, Reduced breast feeding, Smaller family size, Widespread antibiotic use, Increased bathing/showering and use of antibacterial soaps, Use of mercury amalgam dental fillings
“Cleaner” not always better

Question 17

Main vaginal bacteria

Answer 17

Lactobacillus

Question 18

Main C-section delivery bacterium

Answer 18

Staphylococcus

Proprionibacterium

Question 19

Main breast milk bacterium

Answer 19

Bifidobacterium

Lactobacillus

Question 20

Main bacteria in solid food

Answer 20

Bacteriodes

Clostridiales

Question 21

Main contributions of microbiota to host

Answer 21

Digestion and fermentation of carbohydrates
Production of vitamins
Development of the gut-associated lymphoid tissues (start of immune system)
Polarization of gut-specific immune responses (ex: inflammation)
Prevention of colonization by pathogens
Gut immune responses induced by commensal populations regulate the composition of the microbiota as well.
The immune system has evolved adaptations that work to contain the microbiota and preserve the symbiotic relationship

Question 22

Microbiota-Induced Maturation of the GI Tract

Answer 22

Changes in gut microbiota promotes substantial changes in gut morphology
Villus architecture (healthy=long)
Crypt depth (healthy=deeper)
Stem cell proliferation
Blood vessel density
Mucus layer properties
MALT maturation

Question 23

Microbiome and Gut Immune System Interactions

Answer 23

Lymphoid structures develop prenatally before bacterial colonization
M cells at apical surfaces sample luminal antigens and bacteria
Dendritic cells present antigens to induce immune cell maturation
Secretion of IgA and AMPs to control bacterial penetration of gut defenses
Dysbiosis can trigger pro-inflammatory responses

Question 24

Differences in gut microbial composition between obese and lean individuals

Answer 24

2 experiments performed
Preliminary human studies indicated that proportion of genetic material from Firmicutes was higher in obese individuals
When placed on carb-restricting diets, bacteroidetes increased over Firmicutes, correlated with decrease in body fat
In mice, microbiota from obese mice was rich in genes encoding enzymes that break down otherwise indigestible polysaccharides; assists host in extracting extra calories from ingested food

Question 25

Leptin

Answer 25

Hormone that signals levels of body fat

Low levels send message to brain to increase caloric intake

Question 26

Glycan Microhabitats and Food Chains in Gut

Answer 26

Food from plants or meat are rich in source-specific glycans
Attacked in distal gut by primary glycan degraders
More soluble glycan fragments digested by secondary glycan degraders, liberating pools of short chain fatty acids (SCFAs) as fermentation products
Similar food chain occurs in mucus layer

Question 27

Short chain fatty acids (SCFAs)

Answer 27

Important for intestinal epithelial homeostasis

Question 28

Effect of antibiotics on gut flora

Answer 28

Direct and Indirect effects
Indigenous microbes form an ecological barrier that prevents ingress of many pathogenic microorganisms
Antibiotic use sometimes leads to development of C. diff-associated colitis, due to loss of intrinsic “colonization resistance” offered by commensal species
Usually followed by decrease in microbiota diversity
Community is resilient and resembles pretreatment state in matter of days or weeks
Different antibiotics have different recovery periods (up to 6 months)
Gut microflora will never return COMPLETELY to normal-some bacteria lost indefinitely

Question 29

Proteobacteria

Answer 29

Have high levels of antibiotic resistance genes

Question 30

Diseases associated with antibiotic overuse

Answer 30

Clostridium difficile-associated diarrhea
Experimental autoimmune encephalopathy
Type 1 diabetes
Type 2 diabetes
Vancomycin-resistant Enterococcus
Obesity

Question 31

Pseudomembranous Enterocolitis

Answer 31

Caused by C. diff overgrowth due to continuous antibiotic treatments
Produces toxin that damages epithelial cells and produces exudative plaques
Fecal transplantation is remedy-only working treatment to replenish healthy microbiome

Question 32

The Disappearing Microbiota Hypothesis

Answer 32

Changing human ecology beginning in the late 1800s has dramatically altered the transmission and maintenance of our indigenous microbiota
The changes have affected its composition
Altered composition affects human physiology and thus disease risk
Loss of ancestral bacterial, usually acquired early in life, is especially important b/c it affects a developmental critical stage
Ex: H. pylori

Question 33

Microbial therapeutics and disease

Answer 33

Goal to create a healthy biome
Preclinical- subtle biological changes occurring
Early stage- disruption of homeostatic microbiota, production of pro-inflammatory metabolites; mucous layer protection becomes thinner as damage accumulates
Late stage- continuous damage leads to breakdown of epithelial barriers

Question 34

Westernized societies and the microbiome

Answer 34

Modern and Western societies associated with dramatic loss of natural human gut microbiome diversity
Causes and consequences challenging to study
Primate study- Captivity “humanizes” the primate microbiome