A7 & A8 - Microbiota-Bacterial Pathogen Interactions I & II

How does the intestinal microbiota form a protective barrier?

• Nutritional competition

  • Microbiota will complete with pathogenic bacteria for nutrients, therefore preventing pathogenic bacteria growth

• Physical exclusion to intestinal niches

  • This can be done by taking up all potential binding sites as well as by producing AMP to destroy pathogens

• Induction of host immune responses

  • Microbiota can induce increased AMP and Mucin production as well as inducing intestinal lymphocytes and macrophages

Which commensal bacteria takes up carbohydrates used by the pathogen Citrobacter rodentium?

Bacillus thuringiensis (Bt)

• This leads to nutritional exclusion of the pathogen

How can SCFAs influence Salmonella growth?

Can downregulate virulence genes in Salmonella, inhibiting its growth

How can SCFAs influence Enterohaemorrhagic Escherichia coli (EHEC)?

SCFA can inhibit the growth of EHEC.

How many EHEC cells are needed to cause disease?

Only 10 EHEC cells are needed to cause disease

How many carbons does Acetate have?

2 carbon molecule

How many carbons does Propionate have?

3 carbon molecule

How many carbons does Butyrate have?

4 carbon molecule

How do commensal E. Coli strains prevent the growth of related Enterohaemorrhagic Escherichia Coli (EHEC)?

Commensal E. Coli strains can produce antimicrobial peptides (like colcin) which directly inhibit EHEC growth

What can antimicrobial peptides produced by commensal bacteria do?

They can kill or block the growth of competing bacteria (such as pathogenic bacteria)

How are bile salts used by the microbiota?

Bile salts are produced in the liver and are delivered to the duodenum via the bile duct

The microbiota can convert these bile salts into secondary bile acids which are antibacterial molecules

What is Indole?

A microbial metabolite of tryptophan

What occurs to Indole exposed Salmonella?

They are

• Less invasive for epithelila cells and macrophages in vitro

• Does not impact upon survival within macrophages

• Colonise mice at reduced levels

What is the mechanism of action of Indole on Salmonella?

Reduces expression of Salmonella Pathogenicity Island-1 (SPI-1) genes which are important for invasion

How can SCFAs also prevent Salmonella invasion?

SCFAs can repress Salmonella Pathogenicity Island-1 (SPI-1) gene expression.

• This inhibits Salmonella invasion

What happens to HeLa cells treated with Indole in vitro?

They become more resistant to Salmonella invasion.

Mono-colonisation of GF mice with B. fragilis promotes what?

Promotes CD4 T-cell development (specifically Treg subsets)

What does colonisation with Segmented filamentous bacteria promote?

Promotes CD4 differentiation into Th17 cells.

What is the function of the gut mucus layer?

To keep the microbiota at a ‘safe distance’ from epithelial cells.

• This is to prevent the microbiota from penetrating the epithelial layer

How can the microbiota metabolites sometimes enhance bacterial virulence?

Some bacteria have adapted to utilise microbiota metabolites to enhance growth

• E.g.

  • Succinate promotes C. Difficle

  • Hydrogen jump starts Salmonella growth

  • Fucose and Succinate can enhance EHEC virulence

What microbiota metabolite can Salmonella use to jump start its growth?

Hydrogen produced by the microbiota

What can C. Difficle use to enhance colonisation?

Microbiota produced succinate from sialic acid cleavage.

What are four key ways pathogens can overcome microbiota-mediated colonisation resistance?

• Use of microbiota derived metabolites

• Use of alternative nutrients or niches

• Promotion of host inflammation to weaken epithelial barrier

• Production of toxins to lyse/inhibit commensal bacteria

Clostridium difficile colonises where in the gut?

Colonises the large intestine

How many toxins does Clostridium difficile produce and what are they called?

Two toxins

TcdA and TcdB

C. Difficile catabolises _____ as a carbon source.

C. Difficile catabolises sialic acid as a carbon source

How can antibiotic treatment enhance C. Difficile expansion?

Antibiotics can:

• Reduce microbiota composition which decreases colonisation resistance allowing C. Difficile to dominate

• Enhance production of microbiota-derived Succinate and release of host-derived sialic acid

  • These both promote C. Difficile expansion

Ruminococcus obeum is a key antagonistic component of Cholera infections. How does it do this?

Ruminococcus obeum produces Quorum sensing signal molecule autoinducer-2 (AI-2) which represses V. Cholerae colonisation factors

What is Quorum sensing?

The ability of bacteria to behave multicellular despite being unicellular

• Can regulate gene expression in accordance with population density

How does Vibrio cholerae kill completing bacteria?

Has a type 6 secretion system (T6SS)

How does the microbiota prevent decrease Vibrio cholerae T6SS killing?

Intestinal microbiota can modify bile acids into deoxycholic acid

• Deoxycholic acid decreases expression of T6SS which prevents Vibrio cholerae killing competing E. coli bacteria

________ neurones of the enteric nervous system synthesise noradrenaline

Adrenergic neurones of the enteric nervous system synthesise noradrenaline

Noradrenaline is inactivated by host conjugation with glucuronic acid.

How can the microbiota reactivate noradrenaline?

Microbiota-produced enzymes (glucuronidases) can deconjugate glucuronic acid from noradrenaline.

This reactivates noradrenaline and active noradrenaline levels rise

Bacterial detection of noradrenaline causes what?

Causes expression of virulence genes.

Salmonella are facultative intracellular pathogens. What does this mean?

Means they can grow outside cells but can also thrive inside them if it gives them an advantage

Food poisoning if caused by what Salmonella strains?

Non-Typhoidal Salmonella (NTS) strains

Salmonella SPI-1 and SPI-2 encode what?

Encode two distinct type three secretory systems (T3SS)

What is the structure of T3SS?

Composed of a basal body, needle complex and translocon

• This is used to inject effectors into host cells to manipulate host cell biology

Which SPI, SPI-1 or SPI-2, is associated with intracellular survival of Salmonella inside macrophages?

SPI-2 T3SS

How can intestinal inflammation can lead to dysbiosis?

Causes:

• Reduces growth of obligate anaerobes

  • These cannot thrive with oxygen present

• Increases growth of facultative anaerobes

  • These can thrive with or without oxygen

How can inflammation alter nutrient availability?

IL-22 can

• Increase expression of FUT2 gene which increases α(1,2)-fucosylation of mucus carbohydrates 

• The microbiota can release this fucose and utilise it

How does inflammation promote availability of alternative electron acceptors?

Pro-inflammatory cytokines can

• Activate epithelial cell DUOX2 gene which generates hydrogen peroxide

• Activate expression of Nos2 gene which encodes nitric oxide synthase (iNOS) in epithelial cells which generates nitric oxide

• Both hydrogen peroxide and nitric oxide are election acceptors

Patients with IBD usually have raised nitric acid gases in their colon.

True or False.

True.

Patients with Crohn’s or Ulcerative Colitis show raised nitric acid gases due to increased DUOX2 expression.

True or False?

False.

Patients with Crohn’s or Ulcerative Colitis show raised hydrogen peroxide due to increased DUOX2 expression.

How can hydrogen peroxide and nitric acid cause Enterobacteriaceae growth?

They can react to form Nitrate

Enterobacteriaceae can reduce Nitrate and use it for respiration allowing for a growth bloom.

How can intestinal inflammation aid invading pathogens rather than protecting the gut?

Inflammation causes an increase in hydrogen peroxide and nitric acid which causes Nitrate to form

Enterobacteriaceae can utilise Nitrate but the obligate anaerobic microbiota cannot leading to a Enterobacteriaceae growth bloom

This means bacteria like Salmonella can rapidly grow during inflammation of the gut

Why does Salmonella deliberately elicit an inflammatory response in the gut?

Inflammation in the gut can lead to the production of nitrate which Salmonella can use but the obligate anaerobic microbiota cannot.

This allows Salmonella to have a growth bloom, allowing for more effective invasion