sbm 2044: lecture 10 aims: to provide brief introduction to e. coli: a versatile pathogen overview...
TRANSCRIPT
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SBM 2044: Lecture 10
AIMS: To provide
• Brief introduction to E. coli: a versatile pathogen
• Overview of Enterotoxigenic E. coli (ETEC)
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Types of pathogenic E. coli
Enterotoxigenic E. coli (ETEC)Enteroaggregative E. coli (EAEC)Diffusely adhering E. coli (DAEC)
Enteropathogenic E. coli (EPEC)Enterohaemorrhagic E. coli (EHEC)
Enteroinvasive E. coli (EIEC)
Uropathogenic E. coli (UPEC)
Septic E. coli strains
Cholera-likewatery diarrhoea
Colitis orhaemorrhagic colitis
Dysentery
Urinary tract/pyelonephritis
Septicemia/meningitis
Intestinal
Extraintestinal
Disease
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First recognised in 1987 as distinct from other ETEC
EAEC
Known virulence factors:
AAF - Aggregative adherence fimbriae
EAST-1 – EAEC heat stable toxin (similar to ETEC Sta)
Pet – Plasmid-encoded enterotoxin mucus release
Pic – role in intestinal colonisation? - various activities (mucinase, serum resistance, haemagglutinin)
Characteristic adhesion pattern in vitro reflects bacteria- bacteria (in addition to bacteria-host) adhesion
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EAEC
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Strains displaying a distinct adhesion pattern
Enteroaggregrative E. coli (EAEC)
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Diffusely adhering E. coli (DAEC)
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DAEC
• Doubts about their importance as pathogens
• Like EAEC and ETEC, appear to be heterogeneous group of strains, sharing certain common factors
• Four different adhesins identified – F1845 fimbriae & 3 non-fimbrial adhesins
• No significant information on potential toxins
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Enteropathogenic E. coli (EPEC)
First associated with diarrhoea in 1940s
No detectable enterotoxin
Major cause of watery diarrhoea in infants (< 6 months)
• Developing countries - endemic
• Developed countries – sporadic outbreaks in nurseries, paediatric wards, day centres, etc
1970s – first seen by EM to produce unique ‘attaching & effacing’ effects on enterocytes - producing unique lesions, now called A/E lesions
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1. Initial (non-intimate) attachment
2. Intimate adhesion + effacement
3. Formation of pedestals
EPEC interactions with epithelial cells
Studies on mechanisms very limited until late 1980s
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1. Initial (non-intimate) attachment
2. Intimate adhesion + effacement
3. Formation of pedestals
EPEC interactions with epithelial cells
Involved: Intimin (EaeA) [OM protein]
BFP Actinrearrangements
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Anti-EspA
Filaments ‘connecting’EPEC & host cells
Knutton et al (1998) EMBO Journal 17: 2166-2176
Activation of EPEC Type III secretion
Immuno-gold antibodies
Filaments composed of EspA
EM
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Host cell membrane
EPEC OM
EPEC IM
EPEC Type III secretion filamentEspB/D pore
Esc R,S,T,U,V,N,DIM-associated ‘machinery’
EscC
EscJ
Deduced from studies combining: Mutants
protein-protein binding
Labelling with specific Ab
Electron microscope
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Similar ‘needles’ in EPEC, but Salmonella lack ‘filament’
Kubori et al (1998) Science 280: 602-605
Salmonella Type III ‘needles’
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EPEC: Kenny et al (1997) Cell 91: 511 - 520
Receptor for intimin (adhesin) is NOT a host cell protein
Remarkable discovery:
It is an EPEC protein
Translocated into host cell, phosphorylated, & inserted into the host cell membrane
Tir: Translocated intimin receptor
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EPEC: Model of A/E Lesion Formation
Initial adhesion (via Bfp )
Activation of Type III secretion
Actin polymerizationpedestal formation
Tir + ? signals translocated into host cell Tir- P
Intimate adhesion by EaeA to Tir- P
Ca++
Details of signalling still unclear
filaments EspB/D pore
Quorum sensing ?
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Enterohaemorrhagic E. coli (EHEC)
1982: two cases of severe food-poisoning (hamburgers) in USA associated with rare serotype of E. coli - O157:H7
1983: E. coli O157:H7 produces a Shiga-like toxin
1986: Like EPEC, O157:H7 produce A/E lesions
‘Enterohaemorrhagic E. coli’ (EHEC) first used in 1987 to describe what seemed to be a new type of pathogenic E. coli
Main difference with EPEC is production of Shiga-toxin
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Origin of E. coli O157:H7 ?
First isolated in US in 1982 - now worldwide
O157:H7 isolates throughout world are single clone that emerged relatively recently - closely related to a much less virulent EPEC clone
Quantitative population genetic studies (MLEE)
Both SLT-I & SLT-II genes encoded by bacteriophages, - could facilitate their transmission into new strains
O157:H7 probably emerged when an EPEC strain (alreadycapable of producing A/E lesions & diarrhoea) acquired slt phages, resulting in a dramatic increase in virulence
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A1
SSA2
Shiga-toxinsTypical A-B subunit toxins
A1 subunit - N-glycosidase
Gb3-rich cells (particularly sensitive to STx) include
Pentameric B subunit (5 x 7kDa)
• vascular endothelial cells
B
27kDa
4kDa
Receptor: glycolipid called Gb-3
• absorptive enterocytes• kidney endothelial cells
haemorrhagediarrhoearenal failure
Hydrolyses eucaryotic cell 28S rRNA
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STx: Entry via RME retrograde transport via Golgi & ER
From: Groisman
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E. coli Shiga-toxins (STx)
Local effects of toxin in colon
Very serious – often fatal.
Role in disease
Haemolytic uraemic syndrome (HUS)
(10% - 20% cases)
haemorrhagediarrhoea
Absorption – systemic effects Renal failure
S. dysenteriae E. coli producing highhigh levels of STx
Strongly associated:
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E. coli O157:H7
• Clinical reports that therapy with certain antibiotics increased severity of O157 infections
Some antibiotics (e.g. quinolones) inhibit DNA replication.
WHY ?
higher levels of STx in gut
DNA damage SOS response
Induces lysogenic phage
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Urinary tract infections
(community acquired)
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Uropathogenic E. coli (UPEC)
source: UPEC in colon - harmless commensals
Urethra
Bladder
Kidneys
Bloodstream
Ascending infection
UrineFlushing
Removesnon-adherent
bacteria
Ureter
septicemia
pyelonephritis
ureteritis
cystitis
urethritis
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Uropathogenic E. coli - virulence mechanisms
Adhesion - essential to avoid removal by urine
Survival - studies limited to some aspects only
Toxicity inflammation
• iron-acquisition (siderophores)
• capsules (evasion of host defences)- more important in kidney than lower UT- critical if organism enters bloodstream
• LPS (endotoxin)• -haemolysin (membrane damaging toxin)• cytotoxic necrotizing factor 1(CNF-1)
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E. coli -haemolysin (HlyA)
Produced by about:
Membrane-damaging toxin:
• 50% strains from upper UTI – most tissue damage
• Produces small hydrophilic pores in mammalian cells• high concentrations ion leakage osmotic lysis
• lower concentrations more subtle cytotoxic effects
Suggests strong association with UTI
• 30% strains from lower UTI
• 10% GI strains
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E. coli -haemolysin (HlyA)
• Role in kidney damage supported by studies in mouse models:
Parent strain expressing P fimbriae + HlyA
colonized bladder + kidneys 66% mice died
Isogenic mutant expressing P fimbriae, but not HlyA
colonized as above, but no apparent damage or deaths
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Cytotoxic necrotizing factor 1 (CNF-1)
Large (110,000Da) protein, produced by ca 30% UPEC
Studies in vitro:
No direct evidence for role in UTI
• epithelial cells: rearrangements of actin filaments & membrane ruffling
• bladder cells: reduced migration & proliferation – - impede repair of damage bladder ??