Transcript Exotoxins

Summary of Lecture 20
• Microbe: human interactions on and within the
body are normal.
• Human body is a good habitat for suitable bacteria
(nutrients, temperature, pH, etc.)
• Normal microbe flora is important for general
well-being (e.g. vitamin synthesis).
• But bacteria sometimes do invade (pathogens).
Some parts of human body colonised
by bacteria (normal flora)
•
•
•
•
Skin (Staphylococcus)
Mouth (Streptococcus)
Respiratory tract (Streptococcus)
Gastrointestinal tract
– Diverse flora
– Largest number in large intestine
• Urogenital tract (Lactobacillus)
Infection and Disease
• Infection is any situation leading to
establishment of microbial growth with or
without causing damage to the host.
• Disease results in damage to the host.
• Not all infections cause disease.
BS104 Lecture 21
Entry of the pathogen into the host.
Colonisation and growth. Toxins.
Entry of the pathogen into the host
• Pathogens must gain access to host tissue.
• They use various strategies to establish virulence.
• Virulence - the relative ability of a pathogen to cause
damage to the host - disease
• In order to cause damage, the pathogen needs to
penetrate
– Mucous membranes.
– The skin (through wounds).
– Intestinal epithelium.
• Pathogen is often selective for tissue and host.
– E.g. Neisseria gonorrhoeae adheres to urogenital
epithelial cells.
• Adherence to is often very important for pathogenesis.
EXPOSURE
to pathogens
Microorganisms
and Pathogenesis
ADHERENCE
to skin or mucosa
Further
exposure
at local
sites
•Virulence factors
•Invasiveness
•Toxicity
•Tissue damage
INVASION
through epithelium
COLONIZATION
and
GROWTH
Production of virulence factors
TOXICITY:
toxin effects are local
or systemic
Brock: Figure 28.12
Further
exposure
INVASIVENESS:
further growth at original
site and distant sites
TISSUE DAMAGE, DISEASE
Adherence of pathogens to the host
• Extracellular macromolecules
facilitate bacterial adherence.
• Protein: protein interactions
(pathogen: host).
• Pathogens are selective when
adhering to cells. N.
gonorrhoeae – urogenital
epithelia.
• Polysaccharides produced by
bacteria:
• Dense polymer coat (capsule)
• Loose network of polymer
fibres (glycocalyx).
• Diffuse network of polymer
fibres (slime)
• Attachment and defence
against host immune response
• (a) Vibrio cholerae attaching to
brush border of rabbit villi.
• (b) Escherichia coli infecting
calf brush border villi. Note
capsule
Brock: Figure 28.13
Bacillus anthracis colonies
can be mucoid (slimy) in
appearance
Bacillus anthracis with stained
capsule (bright green) – note
how capsule extends about 1m
From actual cell (cell is about
0.5 m diameter)
Fimbriae and Pili
• Fimbriae and pili – facilitators of
bacterial adherence.
• Important bacterial cell surface
structures.
• They initiate attachment by binding
to glycoproteins on the cell surface.
• Fimbriated Escherichia coli often
cause urinary tract infections.
• One of the best characterised
fimbriae are the type I fimbriae of
enteric bacteria (Escherichia,
Klebsiella, Salmonella and Shigella).
• Type I fimbriae -uniformly
distributed on the cell surface.
Escherichia coli EM
showing type P fimbriae.
Cell is 0.5 m wide
Brock: Figure 28.15
Fimbriae and Pili
•Pili are longer than fimbriae.
•There are less pili found on the surface of the cell.
•Both structures bind to host cell surface glycoproteins:
leads to adherence to animal cells.
Major adherence factors
• E.g. Pathogenic forms of
Escherichia coli make
fimbrial proteins called
colonization factor
antigens (CFA).
• These specifically
adhere to the small
intestine cells.
• Pathogenic E. coli
colonizes and makes
enterotoxin – a protein
released extracellularly
–causes damage to the
small intestine.
Invasion
Invasion: Penetration of the microorganism into
the host cell (through the epithelium) and
subsequently inflicting damage.
Broken skin or mucosal surfaces can be points of
entry for pathogens. Once growth is established
at these sites, colonisation and invasion begin.
May also spread throughout the host via the
circulatory or lymphatic systems.
Colonization and growth
• Invasion of pathogen followed by growth leads to colonisation.
• Microbial growth requires available nutrients and environmental
conditions.
– Carbon source, nitrogen source, trace metals.
– trace metals are scarce (bound to host cell proteins).
– bacteria may make metal chelating compounds so they can
scavenge trace metals.
– Temperature, pH and oxygen affect microbial growth, but the
availability of nutrients is most important.
• Localized (focal) infection
– Boil (Staphylococcus)
• Spread through blood and lymphatic vessels is a general (systemic)
infection.
– Lymphatic vessels  lymph nodes  containment by the immune
system (?)
– Blood  distribution throughout the body
• Bacteraemia often starts as a localized infection.
Infections in sport
• Tetanus - Clostridium tetani.
– C. tetani spores often contaminate sports fields.
– Spores can enter deep wounds and then germinate
– Production of exotoxin (neurotoxin) that travels to
CNS that can cause death
• Other recreational infections: Scrumpox (Herpes
gladitorium, impetigo - Staphylococcus); Wound
infections (sepsis); Waterborne (giardiasis – Giardia
lamblia, leptospirosis); Vectorborne (Lyme disease).
Impetigo http://www.healthhype.com/staphskin-infections.html
Scrumpox http://www.sciencedaily.com/release
s/2008/09/080928210041.htm
Scanning EM of the spiral-shaped bacteria of
the genus Leptospira – causative agents of
leptospirosis. – Janice Carr (CDC –Image
no:138)
Lyme disease infection; (a) deer tick attached
to skin, (b) circular rash associated with lyme
disease (Madigan et al., 2009. Brock Biology
of Microorganisms. pp.1012.
Virulence
• Relative ability of a
pathogen to cause
disease.
• LD50 ; The dose
(number of
bacteria) required
to kill 50% of a
test group.
Virulence
•Attenuation: Decrease or loss of virulence.
An attenuated (weakened) microorganism
would have lost the ability to cause serious
illness, but still able to trigger an immune
response. These microorganisms may cause a
mild or subclinical form of the illness.
•Used in vaccine production.
Toxicity and Invasiveness
Toxicity
• Ability of an organism to
kill by a preformed toxin
that kills host cells.
• Tetanus is a disease
caused by exotoxin
produced by Clostridium
tetani.
• Bacteria stay in
localized wound but
toxin spreads through
the body.
Invasiveness
• Growth of bacterium
in large enough
numbers to inhibit cell
function.
• Streptococcus
pneumoniae produces
no toxin but grows in
large numbers in the
lung.
• Polysaccharide capsule
of the pathogenic
strains prevent
phagocytosis.
• Host responses lead to
pneumonia.
Virulence Factors and Toxins
• Salmonella virulence factors:
– Enterotoxin
– Endotoxin
– Cytotoxin – kills the cells
– Siderophores (iron-chelation, sequestering iron to
aid in growth)
– Attachment (fimbriae)
– Defense (capsule)
Virulence Factors and Toxins
Injectosome
(inv and prg
products form
complex)
Endotoxin in
LPS layer
(fever)
Siderophores
Enterotoxin
(diarrhea)
Virulence
plasmid
Anti-phagocytic
proteins induced
by oxyR
O antigen
(inhibits phagocyte killing)
Brock: Figure 28.17
Type 1 fimbriae
(adherence)
Cytotoxin
(inhibits host cell
protein synthesis;
calcium influx into
host; adherence)
Vi capsule antigen;
Inhibits complement binding
Flagellum (motility)
H antigen (adherence; inhibits
phagocyte killing)
Fibrin clots
• Produced by host at wound sites.
• Some pathogens (e.g. Streptococcus) dissolve clots by
production of hyaluronidase (breaks down hyaluronic
acid in cell cement).
• Other pathogens (e.g. Staphylococcus aureus) promote
clots (e.g. by a coagulase).
• Fibrin then coats bacteria affording protection to the
bacterium. Boils and pimples are the result.
• S. aureus strains that are coagulase-positive are more
virulent than the ones that are negative.
Bacterial toxins
Exotoxins
• Proteins released
(secreted) by bacterium to
outside of cell during its
growth.
• Special secretion
machinery often used
• Highly toxic
• Specific targets usually
distant from the site of
infection.
– Generally no fever
Endotoxins
• Toxic lipopolysaccharides
that are part of the gram
negative bacterial cell wall.
• Released when bacteria
lyse (burst open)
• Large bacterial numbers
needed for toxic effect.
• Weakly toxic
• General targets
– Fever, diarrhoea
Exotoxins
• Proteins that are released by bacteria.
• Soluble and may travel throughout body of
infected person.
• See table 28.4 (next 2 slides).
Three major categories of
exotoxins
1.
2.
Cytolytic toxins (e.g. hemolysin – degradation of
cytoplasmic membrane leading to cell lysis)
AB toxins
1.
2.
3.
4.
3.
Diphtheria toxin (Corynebacterium diphtheriae)
Botulinum toxin (Clostridium botulinum)
Tetanus toxin (Clostridium tetani)
Cholera enterotoxin (Vibrio cholerae)
Superantigen toxins – stimulate the immune
system leading to inflammation
Exotoxins
Hemolysis by Streptococcus
pyogenes
• Blood agar plate.
• Zones of clearing
surrounding bacterial
colonies caused by
hemolysin that is
secreted by bacteria in
colonies.
• Cytolytic toxin.
• Attack host cell wall
Exotoxins
Diptheria toxin
• Produced by Corynebacterium diptheriae.
• Rats and mice are insensitive, but humans, rabbits, birds
are sensitive.
• One toxin molecule can kill a host cell.
• Called an AB toxin.
• Blocks protein synthesis by ADP ribosylation of
elongation factor 2 (EF-2), leading to cell death.
• Only made in strains lysogenized by phage  (therefore
toxicity determined by horizontal gene transfer).
• Phage  injection converts non-toxic, non-pathogenic
strains to pathogenic ones – phage conversion.
Exotoxins
Cytoplasmic membrane
Out
Receptor protein
Key
Diphtheria toxin
Amino acid
ADP ribosylation of EF-2
Ribosome
Normal protein synthesis
Protein synthesis stops
Diphtheria toxin:
Blocks protein synthesis by ADP ribosylation of EF-2.
Brock Figure 28.20
In
Exotoxins
Tetanus and Botulinum toxins are also
A-B toxins
• Produced by Clostridium tetani and Clostridium
botulinum respectively.
• Both microbes are obligate anaerobes.
• Neither microbe is very invasive – pathogenic
effects are due to neurotoxicity.
• C. botulinum grows in badly preserved food.
• Botulism causes death by muscle paralysis resulting
in respiratory failure.
• C. tetani grows in deep body wounds.
• Toxins spread throughout body via the neural cells,
causing spastic paralysis.
Tetanus toxin: blocks glycine release – constant Exotoxins
acetylcholine release.
Inhibitory
interneuron
Inhibition
Excitation signals
from the central
nervous system
Tetanus
toxin
Muscle
Normal
Glycine (G) release from inhibitory interneurons
stops acetylcholine (A) release and allows
relaxation of muscle
Tetanus
Tetanus toxin binds to inhibitory interneurons,
preventing release of glycine (G) and relaxation
of muscle
Brock: Figure 28.22
Botulinum toxin:Blocks neurotransmitter acetylcholine release
Excitation signals
from the central
nervous system
Muscle
Normal
Acetylcholine (A) induces contraction
of muscle fibers
Brock: Figure 28.21
Botulism
Botulinum toxin, , blocks release of A,
inhibiting contraction
Exotoxins
Clostridium tetani
Large gram negative rod
with terminal endospores
(looks like drumsticks)
A soldier dying from tetanus.
Painting by Charles Bell in the Royal
College of Surgeons, Edinburgh.
Exotoxins
Enterotoxins
•
•
•
•
•
Exotoxins that act on small intestine.
Result in excessive fluid secretion.
Cause vomiting and diarrhoea.
Food poisoning (Staphylococcus aureus).
Intestinal pathogens (Vibrio cholerae, Salmonella
enteriditis).
Exotoxins
Cholera toxin
• AB toxin, produced byVibrio cholerae.
• The B fragment directs the toxin to the intestinal
epithelium.The A fragment crosses the cytoplasmic
membrane and activates adenyl cyclase resulting in an
increase of cellular cAMP.
• cAMP is an important mediator in diverse regulatory
systems, including ion balance.
• Increased cAMP levels induce chloride and bicarbonate
ions to be secreted into the intestinal lumen, leading to
an imbalance. This results in excessive amounts of water
production into the intestinal lumen and subsequently
water loss into the small intestine.
• Treatment is by oral solutions containing electrolytes
and other solutes to reverse the ion imbalance.
Endotoxins
• Endotoxins stimulate host to release endogenous
pyrogens (leads to fever).
• Also cause diarrhoea, inflammation etc.
• Large doses can cause death due to haemorrhagic
shock and tissue necrosis.
• Lower in toxicity than exotoxins.
• Endotoxins are the lipopolysaccharides of gramnegative bacteria outer membrane.
Further reading
• Madigan et al., 2009. Brock: Chapter 28.6 to
28.12