E.coli Flashcards
E.coli organism
-rod-shaped, non spore-forming gram-negative, facultative anaerobe
- belongs to the family enterobacteriaceae
- serotyped using: O antigens of LPS, H antigens of flagellin protein
- Produces pilli - the smaller surface projections
And produces peritricus flagella
why is E.coli a model organism
-Can be easily genetically manipulated
- it grows quickly on range of media
- genetic systems well developed
- metabolically versatile
dual aspect of E.coli
E.coli has a dual aspect of being both a commensal organism and a pathogen
e.coli - commensal organism
- colonises GI tract a few hours after birth
- most abundant facultative anaerobe of human intestinal microflora
E.coli - pathogen
– UTI
- Diarrhoea
- haemolytic uraemic syndrome
- septicaemia, pneumonia, meningitis
pathotypes of E.coli
pathotypes - groups of strains from within a single species that cause a specific disease using a common set of virulence factors
- extra-intestinal E.coli
- diarrheagenic pathotypes
- extra-intestinal E.coli
- neonatal meningitis
- uropathogenic
- avian pathogenic
diarrheagenic pathotypes
- enteropathogenic (EPEC)
-enterohaemorrhagic (EHEC)
-enterotoxigenic (ETEC)
-enteroaggregative (EAEC)
-enteroinvasive (EIEC)
site of pathogenic E. coli colonisation
These pathotypes cause different diseases through colonising different sites on the body
- brain: NMEC
-bloodstream: UPEC and NMEC - large bowel: EHEC, EIEC, and EAEC
-kidney: UPEC - small bowel: EPEC, ETEC, DAEC, and EAEC
- bladder: UPEC
site of diaarhoeagenic E.coli colonisation - large intestine
The role of the 7 metre long intestine is to both absorb nutrients but also to form a barrier to toxic substance and bacteria entering the human body: has undulating ridges and villi to increase SA
- Diaryogenic bacteria must colonise this environment, often directly interacting with these polarised epithelial cells
enteropathogenic E.coli
-important cause of infant diarrhoea in developing countries
diarrhoea results from multiple mechanisms:
- active ion secretion
- increased intestinal permeability
- increased inflammation
- a loss of absorptive microvilli
The most characteristic function of EPEC infection is the intimate interaction it can set up with host cells termed the ‘attaching and effacing’ phenotype
pathogenic mechanisms of enteropathogenic E.coli
-EPEC is an ectracellular pathogen initially binding to the host cell surface using bundle forming pili
-following binding EPEC injects a variety of effector proteins into the host cell using a type 3 secretion system - these hijack the cells for the benefit of the bacterium
- the most prominent change in cell structure is the formation of attaching and effacing structures on epithelial cells
what is effacement
effacement is the destruction of the microvilli with production of the pedestal by T3SS effector protein mediated cytoskeletal changes involving acrin polymerisation
enterohaemorrhagic E.coli
EHEC causes: - non bloody or bloody diarrhoea, haemolytic uraemic syndrome
-similar attaching and effacing activity to EPEC
- importantly produces shiga toxin (Stx)
- outbreaks - particularly associated with beef/beef products and farm visits by children
- serotype 0157:H7 most important in US/UK
EHEC Stx as a virulence factor
- an A1B5 type toxin - Stx or veryoctyotoxin
- EHEC Stx is closely related to Shiga toxin produced by shigella species responsible for dysentery
- receptor for B subunit of toxin is glycolipid globotriaosylceramide
- intracellular target of toxin A subunit is ribosomal RNA that is cleaved leading to cessation of protein synthesis
EHEC STX toxin mechanism of action
-Stx A subunit contains the active site glutamic acid at position 167/293 of the AI subunit
- cleavage at trypsin-sensitive region results in an A1 subunit and A2 peptide linked via a disulphide bond
- the b pentamer allows the toxin to bind to cells that express Gb3 glycolipid on their surface
- toxin is taken up by endocytosis, transported in a retrogade manner through the Golgi/ ER and the AI domain released into the cytoplasm
- toxin cleaves 28srNA component of the ribosome halting protein synthesis
EHEC symptoms
in the colon, Stx can cause bloody diarrhoea, hemorrhagic colitis, necrosis and intestinal perforation
- however, Stx can travel in the bloodstream to the kidney
- receptor is found on kidney cells and toxin binding leads to inflammation and potentially HUS with symptoms:
- haemolytic anaemia
- thrombocytopenia
- ultimately acute renal failure
EHEC causing Haemolytic uraemic syndrome (HUS)
- EHEC is the most common cause of HUS
- classic symptoms of HUS present within 2-12 days following onset of diarrhoea in approximately 15% of cases of EHEC - associated gastroentiritis
- children under 5 and the elderly are more likely to progress to HUS
epidemiology of EHEC in the UK in 2018
in 2018, 607 confirmed cases of EHEC O157 were reported in england and wales
- almost 1/3 of cases were hospitalised
- 612 non- O157 EHEC cases
- four EHEC outbreaks reported, involving 55 cases
EPEC/N EHEC pathogenicity island
- Contains 41 genes
This locus is necessary and sufficient for the production of the attaching and effacing phenotype by an ecoli strain - Thus the EPEC lee can be transferred to an ecoli K-12 lab strain and this will now interact with host cells in exactly the same way
○ To remove microvilli and produce the predestal structure
-The 41 lee genes are arranged into 7 transcripitonal units or operons containing mulitple co regulated genes- LEE1 through to LEE7
There are also 4 genes that have their own individual promoters
- LEE1 through to LEE7
locus of enterocyte effacement - secreted translocator proteins
Form the needle and translocon at the Tip of the machinery, interacting with the host
cell
locus of enterocyte effacement - chaperones
Ensure the appropriate effector Proteins are secreted
locus of enterocyte effacement - transcriptional regulators
Ensuring that this machinery is only produced in the appropriate environment
locus of enterocyte effacement - intimin
Present on the bacterial cell surface and acts as an adhesin binding to one of the
Secreted effector proteins Tir
EPEC injects the receptor TIR, into the host cell, that then localises to the cell membrane whilst EPEC also produces the adhesin intimin, which specifically binds to this receptor
ETEC clinical features
- watery diarrhoea sometimes with vomiting and fever
- mild to severe profuse cholera-like illness
ETEC epidemiology
- food and very commonly water-borne infection via faecal-oral route
- a common cause of diarrhoea in low income countries and is often a child’s first symptomatic enteric illness
- repeated rounds of infection are common in the first 2 years o life eventually leading to protective immunity
- infection can be fatal in infants and young chilfdren
estimated 200 million cases of ETEC diarrhoeal disease each year with 50,000 deaths
ETEC adhesins - fimbrial
- over 20 antigenically distinct fimbriae produced
- referred to as colonisation factor antigen (CFA) or coli surface antigen pili
- the majority of ETEC strains isolated from humans express either CFA/I, CFA/II, or CFA/IV
- belong to the chaperone usher family of pili
- critial for virulence
- many of these are uncharacterised
- genes for fimbrial adhesins often on high. molecular weight plasmids that also contain genes for LT/ST toxins
ETEC adhesins - afimbrial - Tia
- 25 kDa outer membrane protein
- mediates attachment to intestinal epithelial cells as well as their invasion
- binds heparin sulphate proteoglycans promoting adherence and invasion of host epithelial cells
ETEC adhesins - afimbrial - toxigenic invasion locus B
- a member of the self-associating autotransporter family
- outer membrane located 104 kDa glycoprotein
- glycosylation required for oligomerisation and adhesion to host cells
ETEC adhesins - afimbrial - ETEC two-partner secretion protein A protein
- transiently located at the tip of ETEC flagella
- bridges across the flagella tip with host cell receptors
- mediates ETEC attachment to epithelial cells
ETEC toxins - heat labile toxins
- plasmid-encoded heterohexameric holotoxin (closely related to cholera toxin)
- single A subunit with 2 domains(A1/A2) linked by disulfide bridges:
- A1: active toxin molecule
- A2: helical anchor to B pentamer - intact A subunit not enzymatically active until nicked to A1, A2
- A1 subunit released by reduction of disulfide bond - pentameric B subunit
- binds to GM1 gangliosides on host cell surface
- triggers endocytosis of holotoxon
ETEC toxins - heat stable toxin
- retains activity following 95 degrees for 60 minutes
- family of small peptide toxins:
- STa and STb families with differing structure and mechanism of action
- STa associated with human disease
- receptor for STa is host cell guanylate cyclase - regulates intracellular cyclic gmp LEVELS
- STa structural mimic of endogenous human peptide suanylin endogenous ligand for guanylate cyclase
- ST produced as a precursor of approx 72 amino acids
- cysteine rich - intramolecular disulphide bonds important part of structure
pathogenic mechanisms of enterotoxigenic E.coli
Sta toxin upregulates guanylyl cyclase which upregulates cyclic GMP levels and that directly upregulates CFTR
A bacteria can adhere through fimbrae on the cell surface and then it produces toxins which is why we get disease
- That has implications because then we get uptake and release of the A subunit
- Then we get this process where we get increased activity of adenyl cyclase and upregulation of cyclic AMP which affects protein kinase, which affects the acitivity of the CFTR transporter - so chloride secretion
- And that causes fluid loss and the watery type diarrhea
enteroaggregative E.coli
- often cause a persistent diarrhoea in children
- binds intestinal cells with no resultant histological changes
- bacterial aggregation key feature of infection
- secrete array of enterotoxins and cytotoxins
Enteroinvasive E.coli
an intracellular pathogen
- related to shigella species
- generally watery diarrhoea can get invasive inflammatory colitis even dysentry
- host cell invasion but dissemination beyond submucosa rare
- virulence associated with plasmid pINV-borne type III secretion system
- VirG protein localises to single pole of cell recruiting and activating N-WASP and Arp2/3 complex for actin polymerisation
- mediates intercellular spread
