Microbiota

Gut is home to how many microorganisms (bacteria & fungi)

Gut is home to 10¹³-10¹⁴ microorganisms (bacteria & fungi)

How many individual species of microorganisms are there in the gut

15,000-36,000 individual species

Concentration and complexity of microorganisms increases/decreases from proximal to distal gut

Concentration and complexity of microorganisms increases from proximal to distal gut

Most bacteria in the gut are aerobic/anaerobic

In the gut, 99% of bacteria are anaerobes

Where in the GI tract are each of these found:

Lactobacillus, Clostridium, Steptococcus, Helicobacter pylori, Bacteroides, Bifdobacterium

Stomach: Lactobacillus, Steptococcus, Helicobacter pylori

Duodenum: Steptococcus, Lactobacillus

Jujunum: Steptococcus, Lactobacillus

Proximal ileum: Steptococcus, Lactobacillus

Distal ileum: Clostridium, Steptococcus, Bacteroides, others

Colon: Bacteroides, Clostridium, Bifdobacterium, others

The Nobel Prize in Physiology or Medicine in

2005 was awarded jointly to Barry J. Marshall

and J. Robin Warren for their discovery of what

the bacterium Helicobacter pylori and its role in gastritis and peptic ulcer disease.

(Dr. Barry Marshall, an Australian physician, famously infected himself with Helicobacter pylori in 1984 to prove that the bacterium caused peptic ulcers)

(Helicopters can fly & so can Robins)

What important things do gut microbiota produce

Production of vitamins: B12, K, Thiamine, Folate, Biotin

Other than produce vitamins, what do gut microbiota do for us

Regulation of energy balance

Transplanted bacteria treatment for colitis

Bile acid metabolism and bile acid signalling

How do gut microbiota regulate energy balance

A shift in gut microbiota is associated with obesity

In genetically obese mice, a shift in gut microbial ecology occurs

- Bacteroidetes reduced 50%

- Firmicutes increased 50%

Similar shift in Bacteroidetes:Firmicutes occurs in obese versus lean humans

There are some mice that are bred & raised in an incubator. Does this affect their gut bacteria

Yes, they have no bacteria in their GI tract

Effect of no bacteria in the gut of germ free mice

Germ-free mice are resistant to both genetic and diet induced obesity

How would germ free mice react if they mixed with obese mice

Colonisation of germ-free mice with the gut bacteria of conventionally raised obese mice induces obesity in 10-14 days with no increase in food intake

How are germ free (GF) mice protected from diet-induced obesity

by mechanisms that result in changes to fatty acid metabolism:

• stimulate oxidative phosphorylation (Enhanced oxidative phosphorylation means these mice burn more calories, reducing fat accumulation)

• stimulate lipid oxidation (prevents excessive fat accumulation)

-reduce lipid accumulation in skeletal muscle

What infection regularly occurs after antibiotic treatment

Clostridium difficile (C. difficile)

______% of antibiotic associated diarrhoea is caused by C. difficile

15-25%

Clostridium difficile infection causes what

colitis (inflammation of the inner lining of the colon)

How do you treat colitis

antibiotics

Why is C. difficile a considerable health worry for health systems

± 500,000 cases/year in USA

± 30,000 deaths

Costs >US$5 billion/year

How does a c. difficile infection occur

Small outgrowth of C. difficile arises naturally with ageing but is enhanced with antibiotics while other members of the gut flora are reduced

Effect of increased c. difficile on bile acids & effect of bile acids on c. difficile

Increased c. difficile causes bile acid modifications.

Microbially modified bile acids to secondary bile acids are reduced - these normally act to keep C. difficile in a dormant state - c. difficile increases

Effect of unmodified primary bile acids on c. difficile

The unmodified bile acids promote germination of C.difficile

Effect of c. difficile on the gut

C.difficile produce toxins that can destroy the gut cells and reduce other populations of bacteria so that C diff can flourish in the niche

Can faecal transplantation be used to get rid of c. difficile

Faecal microbiota transplantation (FMT) replenish normal microbial functionality to displace C. difficile.

In the last 10 years faecal microbial transplant has been major area of research in treatment of C. difficile

Successful resolution of recurrent infection in up to 75-95% of cases

Basic premise = reestablish a healthy microbiota by transplanting faecal microbes from a healthy donor

How does a faecal microbial transplant work

Faeces collected from donor

– Can be frozen

– Can mixed with saline and transplanted freshly

Transferred to recipient

– Orally (capsule)

– By naso-jejunal infusion

– Into colon

Where are bile acids (BAs) synthesized? & by what

BAs are synthesized from cholesterol, exclusively by the liver

How many mg of BAs (bile acids) are produced daily

500mg

When converting cholesterol to BAs it goes from soluble → insoluble or the other way around

Water-insoluble cholesterol converted into water-soluble compounds

What is the primary function of bile acids

Bile acids convert water-insoluble cholesterol into water-soluble compounds with detergent properties, essential for bile formation and fat absorption

What happens to bile acids after synthesis in the liver

Bile acids are conjugated with glycine or taurine, actively secreted by hepatocytes, stored in the gallbladder, and released into the intestinal lumen in response to a fatty meal.

How do bile acids aid digestion in the small intestine

Bile acids act as detergents, emulsifying fats and facilitating the absorption of dietary fats and lipid-soluble vitamins (A, D, E, K).

How are bile acids reabsorbed

Bile acids are reabsorbed in the terminal ileum via the Apical Sodium Dependent Bile Acid Transporter (ASBT).

What is the enterohepatic circulation of bile acids

It is the cycling of bile acids between the liver and intestine, where 95% are reabsorbed and recycled, while 5% are lost in feces.

(The 5% lost in feces is replaced by new bile acid synthesis in the liver)

What is the most important enzyme in bile acid synthesis

CYP7A1

What regulates CYP7A1 activity

The activity of CYP7A1 is regulated by bacteria in the gut

What are the two pathways for bile acid synthesis? (name & product)

The classical pathway (producing cholic acid [CA]) and the alternative pathway (producing chenodeoxycholic acid [CDCA]).

What happens to primary bile acids after they are synthesized

They are conjugated with glycine (G) or taurine (T) to form bile salts (T(G)CA & T(G)CDCA)

What do gut bacteria do to primary bile acids

Gut bacteria deconjugate bile acids using bile salt hydrolase (BSH) and convert them into secondary bile acids.

Effect of deconjugation on the fate of bile acids

Deconjugated BA can pass through the colon & enter the faeces - it prevents active reuptake from the small intestine via ASBT (apical sodium-dependent bile acid transporter)

Did germ free mice have high/lower levels of bile acids in the GI tract & liver / serum / faeces.

What did this show?

Bile acid levels in germ free mice were higher in the GI tract & liver but lower in serum & faeces.

Microbiota must have an influence on either the synthesis and excretion of BAs

Synthesis of BAs is tightly regulated by what mechanism through the nuclear BA receptor farnoside-X receptor (FXR)

negative feedback

What is FXR and where is it found

FXR(farnoside-X receptor) is a transcription factor that binds the promoter region and initiates the expression of a wide range of target genes

FXR is found in in liver and ileum

Effect of FXR on CYP7A1

Inhibits its expression

This is how the negative feedback works - BAs exert negative feedback of BA synthesis by reducing CYP7A1 activity

Effect of Murine taurine-conjugated primary BAs T- ⍺MCA and T- βMCA on FXR

naturally occurring FXR antagonists - reduce FXR signalling (therefore increase the BA pool) (T-βMCA cannot be metabolized in absence of gut bacteria)

BAs can shape the gut microbiota by……

• promoting the growth of BA-metabolizing bacteria

• -inhibiting the growth of other BA-sensitive bacteria

effect of a Biliary obstruction on microbiota

Biliary obstruction blocks bile flow leading to bacterial overgrowth

BAs have a direct pro/antimicrobial effect

BAs have a direct antimicrobial effect

-detergent effect

-through FXR: induction of antimicrobial agents

What are probiotics

Live microorganisms ingested to confer health benefits upon the host

Requirement for probiotics to work

Must be able to make it through the acidic environment of the stomach in sufficient numbers to influence intestinal microbial composition

Common probiotics

Commonly Lactobacillus ssp.

Bifidobacterium sp

What type of foods are often rich in probiotics

Fermented foods (e.g. Japanese fermented soy beans)

difference between pro & pre biotics

Probiotics are live microorganisms

Prebiotics are a source of food for healthy gut bacteria

What do intestinal bacteria do with prebiotics

Prebiotics cannot be digested and so are fermented by the intestinal bacteria

Effect of prebiotics on gut bacteria

Selectively stimulate the growth and/or activity of intestinal bacteria potentially associated with health and well-being