Exotoxins

Question 1

What is a pathogen

Answer 1

A microorganism capable of causing

disease.

Question 2

what does pathogenicity mean

Answer 2

The ability of an infectious agent

to cause disease.

Question 3

what does virulence mean

Answer 3

The quantitative ability of an agent

to cause disease.

Question 4

what does toxigenicity mean

Answer 4

The ability of a microorganism to
produce a toxin that contributes to the
development of disease.

Question 5

What are exotoxins

Answer 5

–Heterogeneous group of proteins produced
and secreted by living bacterial cells.
–Produced by both gram negative and gram
positive bacteria.
–Cause disease symptoms in host during
disease.
–Act via a variety of diverse mechanisms.

Question 6

difference between gram positive and gram negative

Answer 6

gram positive: thick peptidoglycan layer and no outer lipid membrane
gram negative: thin peptidoglycan layer and have a lipid membrane

Question 7

some activities of exotoxins

Answer 7

• Evade immune response
• Enable biofilm formation
• Enable attachment to host cells.
• Escape from phagosomes

Question 8

in staphylococcus aureus what does haemolytic toxins cause

Answer 8

cause cells to lyse by forming pores

Question 9

in staphylococcus aureus what does phenol soluble modulins (PSM) cause

Answer 9

aggregate the lipid bilayer of host cell causing lysis

Question 10

Genetics of exotoxins

Answer 10

• Can be encoded by chromosomal genes e.g. Shiga toxin in Shigella dysenteriae, TcdA & TcdB in C. difficile
• Many toxins coded by extrachromosomal genes
—• Plasmids — Bacillus anthracis toxin, tetanus toxin
—• Lysogenic bacteriophage — e.g. streptococcal pyrogenic exotoxins in Scarlet Fever, Diphtheria toxin.

Question 11

What are the 3 classification of exotoxins

Answer 11

1. Membrane Acting Toxins — Type I
2. Membrane Damaging Toxins — Type Il
3. Intracellular Toxins —Type Ill

Question 12

How does type 1 exotoxins acts, interfere and target

Answer 12

-Act from without the cell.
-Interfere with host cell signaling by inappropriate activation of host cell receptors.
-Target receptors include
• Guanylyl cyclase which increase intracellular cGMP
• Adenyl cyclase which increase intracellular cAMP
• Rho proteins
• Ras proteins

Question 13

ways type 2 exotoxins work

Answer 13

–Insert channels into host cell membrane.
• ß sheet toxins e.g. S.aureus a — toxin, y toxin, PVL
• a helix toxins — e.g. diphtheria toxin
–Enzymatical damage e.g. S. aureus ß- haemolysin, PSM
–Receptor mediated
–Receptor Independent

Question 14

what are the 2 usual components of type 3 exotoxins

Answer 14

AB toxins :
B= receptor binding and translocation function
A= toxigenic(enzymatic)
there can be single or multiple B units shown as AB subscript n

Question 15

what are the 4 common enzymatic component A of type 3 exotoxins

Answer 15

• ADP — ribosyl transferases - e.g. Exotoxin A of Pseudomonas aeruginosa, pertussis toxin.
• Glucosyltransferases — e.g. TcdA and TcdB of Clostridium difficile
• Deamidase — e.g. dermonecrotic toxin of Bordetella pertussis.
• Protease — e.g. Clostridial neurotoxins: botulism & tetanus
• Adenylcyclase - e.g. EF toxin of Bacillus anthracis

Question 16

how does superantigen induce inflammatory cytokine release

Answer 16

non specific bridging of the MHC Class Il and T- cell receptor leading to cytokine production. E.g.
Staphylococcal Exfoliative Toxin A, Toxic
Shock Syndrome Toxin 1 (T SST 1)

Question 17

what 2 things can you use to inactivate toxins to create toxoids

Answer 17

formaldehyde or glutaraldehyde

Question 18

what are toxoids

Answer 18

inactive proteins but still highly immunogenic -form the basis for vaccines

Question 19

Microbiology of clostridium difficile (6 points)

Answer 19

• gram-positive bacillus. 
• anaerobic. 
• spore-forming. 
• toxin-producing. 
• can be carried 
asymptomatically in the 
gut. 
• 3 toxins.

Question 20

Epidemiology of clostridium difficile (4 points)

Answer 20

• Common hospital acquired infection
worldwide.
• Spread by ingestion of spores — remain
dormant in environment.
• Coloniser of the human gut up to 5% in
adults. (commensal)
• Risk factors— antibiotic use, age, antacids
& prolonged hospital stay.

Question 21

how does antibiotic worsen Clostridium difficile

Answer 21

• Thought to act by disrupting the microbial ecosystem within the gut.
• Antibiotics provide a competitive advantage to spore
forming anaerobes over non spore forming anaerobes.
• Allows C. difficile colonisation and growth.
• All antibiotics have potential for causing disease.

Question 22

C. difficile Pathogenesis

Answer 22

–Cytotoxin A - TcdA coded by tcdA gene
–Cytotoxin B — TcdB coded by tcdB gene
–Binary toxin — C. diff transferase (CDT) — minor role in disease
–Tcd A and Tcd B — Type Ill AB toxins.
The A component of toxins are glycosylating enzymes.

Question 23

What are the cytopathic and cytotoxic effects of C. difficile

Answer 23

• Patchy necrosis with neutrophil infiltration
• Epithelial ulcers
• Pseudomembranes— leucocytes, fibrin, mucous, cell debris.

Question 24

Diagnosis of C.difficile

Answer 24

• Clinical signs and symptoms
• Raised white cell count in blood.
• Detection of organisms and toxins in stool
–• 2 phase test :
I. Glutamate dehydrogenase — detects if C. difficile
organism present.
2. Toxin enzyme linked immunosorbentassay
(ELISA) for TcdA and TcdB toxins.
• Detection of tcdA and tcdb genes — PCR
• Colonoscopy — pseudomembranous colitis

Question 25

C. difficile treatment

Answer 25

• Treatment dependent on severity and presence of surgical complications • Ideally removal of offending antibiotic — not always possible • Antibiotics fidaxomicin or metronidazole or vancomycin • Surgery — partial, total colectomy • Recurrent — faecal transplant.

Question 26

what is Verocytotoxin Escherichia | coli (VTEC) disease

Answer 26

``` • VTEC, or Shiga-toxin (Stx) producing E. coli (STEC) can cause disease mild to life threatening disease. • Stx carried by some E. coli — most commonly 0157:H7 • Identified usually by growth on sorbitol MacConkey agar (SMac) — does not ferment sorbitol and hence is clear. • Other less common types not identified using SMac. ```

Question 27

Epidemiology of VTEC

Answer 27

• E. coli 0157:H7 naturally colonizes the gastrointestinal tracts of cattle who are generally asymptomatic. • Transmission • Predominantly via consumption of contaminated food and water • Person to person, particularly in child day-care facilities, and from • Animal to person. E.g. in petting zoos, dairy farms, or camp grounds. • Very low infectious dose

Question 28

Pathogenesis Toxin of VTEC

Answer 28

``` • Toxin — Shiga like toxin (SLT) = shigatoxin (Stx) = verocytotoxin (VTEC) • Stx, Stxl, Stxla, lc, Id Stx2a, 2c, 2d — variations in a.a. sequence • Gene carried on lysogenic bacteria. • Type Ill exotoxin — AB5 • Enzymatic component A = N-Glycosidase • Bound to 5 B subunits ```

Question 29

Mechanism of VTEC

Answer 29

• Bind to receptor globotriaosylceramide Gb3 or globotetraosylceramide (Gb4) on host cell membrane • Bound toxin internalised by receptor mediated endocytosis. • Carried by retrograde trafficking via the Golgi apparatus to the endoplasmic reticulum. • The A subunit is cleaved off by membrane bound proteases • Once in the cytoplasm Al and A2 disassociate • Al binds to 28S RNA subunit— blocks protein synthesis

Question 30

STEC Pathogenesis

Answer 30

• STEC closely adheres to the epithelial cells of the gut mucosa. • The route by which Stx is transported from the intestine to the kidney and other tissues is debated, possibly polymorphonuclear neutrophils (PMNs) • Bind to glomerular endothelial cells of kidney, cardiovascular and central nervous system. • Very high levels of Gb3 in kidney so kidneys most affected. • Thought that Stx favours inflammation resulting in microvascular thrombosis and inhibition of fibrinolysis.

Question 31

STEC Disease

Answer 31

``` • Can be severe and life threatening • Children < 5 years greatest risk • Abdominal cramps, watery or bloody diarrhoea — may not be present • Haemolytic uraemic syndrome • Anaemia • Renal Failure • Thrombocytopaenia • Less common are neurological symptoms . lethargy, • severe headache, • convulsions, • encephalopathy. ```

Question 32

STEC Diagnosis and Treatment

Answer 32

``` • Diagnosis • Clinical signs and symptoms • Haematological and biochemical evidence. • Stool culture — Growth on SMac • PCR for Stx genes • Treatment • Supportive including renal dialysis and blood product transfusion • Antibiotics have little to no role ```