Blood test revealed evidence of infection with S' pyogenes - PowerPoint PPT Presentation

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Blood test revealed evidence of infection with S' pyogenes

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She has a history of eczyma (on her cheeks and arms) and ... Fibrin deposition -shearing of RBC. Hemolytic anemia. Tiny blood clots -ischemia. Damage to bowel ... – PowerPoint PPT presentation

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Title: Blood test revealed evidence of infection with S' pyogenes


1
Case 1
  • Blood test revealed evidence of infection with
    S. pyogenes
  • (blood test is ASOT, antibody titers against
    Streptolysin-O)

2
Streptococcus pyogenes
3
Soft tissue infections caused by S. pyogenes
4
Streptolysin O
  • Single chain, multidomain protein
  • Secreted by Streptococcus pyogenes
  • Binds cholesterol and oligomerizes x 7
  • After formation of the heptamer, a
    conformational change is presumed to cause
    penetration of the cytoplasmic membrane
  • SLO is not internalized and is not enzymatically
    active
  • Allows ion and water leakage, ultimately causing
    cell death
  • SLO pore is also a channel for translocation of
    Spn (and possibly other virulence factors)
  • Result is cell death and tissue destruction -
    presumably facilitates spread and resistance to
    phagocytosis

5
Pore-forming Mechanism
  • TMHs contain 3 ?-helices
  • Unfold to form transmembrane a-hairpins
  • Oligomer (40-50 monomers) forms large b-barrel
    pore
  • SLO, PFO similar mechanisms

TMH1
TMH2
Adapted from Rajesh Ramachandran et al. Nature
Structural Biology 9, 823 - 827 (2002)
6
Cholesterol-DependentCytolysins
Adapted from Rossjohn et al. Cell, Vol.
89, 685 - 692 (1997)
  • Pore-forming proteins
  • Extensive family 20 molecules from
    Gram-positives (Arcanobacterium, Bacillus,
    Clostridium, Listeria, Streptococcus)
  • 30-60 amino acid identity
  • Cholesterol-dependency for prepore to pore
    conversion
  • 20-30 nm diameter pores

7
Streptococcus pyogenes - Pathophysiology
Fe
3






Streptolysin S Streptolysin O Streptokinase Hyalur
onidase DNAase Pyrogenic exotoxins
M protein F protein
Capsule M protein C5a peptidase
8
Case 2
Baby R.A. 3 mos infant with 1 week of
constipation, decreased activity, weak cry, and
now seems floppy. Other history parents
sometimes put honey on his pacifier.
  • Noted to have decreased muscle tone in all
    extremities, poor feeding, no bowel movements.
  • Muscle weakness increased over next week.
  • Noticed to look a littlebluish, blood gases
    revealed low oxygen, high CO2 required
    mechanical ventilation.
  • Stool culture positive for

Clostridium botulinum
9
Infantile Botulism
10
Clostridium botulinum
11
Botulinum Toxin
  • The most toxic agent known. Medically useful.
    Potential biowarfare agent.
  • Secreted by Clostridium botulinum, either after
    ingestion or innoculation - skin trauma.
  • 7 serologically distinct family members BoNT A
    to G.
  • Comprised of heavy and light chain linked by
    disulfide bond (A-B toxin).
  • Heavy chain mediates receptor binding. Light
    chain is enzymatically active.
  • Bind to gangliosides (low affinity) and
    synaptobrevins (high affinity)
  • Target cell is peripheral motor neuron
    (stimulates muscle contraction by release of
    acetylcholine into neuromuscular junction).

12
Botulinum Toxin (2)
  • Internalized by receptor-mediated endocytosis.
  • Upon acidification, light chain (A domain) is
    translocated to cytoplasm.
  • Light chain is a zinc metalloprotease with high
    specificity for synaptobrevin (SNAP-25).
  • Prevents fusion of synaptic vesicles with
    membrane, prevents acetylcholine release.
  • Results in weakness progressing to paralysis.
  • (Note that tetanus toxin TeNT has almost
    identical structure and enzymatic activity, but
    it is transported to inhibitory neurons in the
    spinal cord - blocking their inhibitory effect on
    motor neurons. Results in tetany - spastic
    paralysis)

13
Structure of Botulinum Toxin
Protease domain (blue)
Translocation domain (green)
Binding domains (red and yellow)
14
Synaptic vesicle fusion with nerve terminus via
SNARE proteins
15
Effect of botulinum toxin on synaptic vesicle
fusion
16
Case 3
D.R. - 5 y.o. admitted with bloody diarrhea x 1
week--gt anemia, low platelet count, decreased
urine output. Family cooked hamburgers two days
before D.R. became ill.
  • Acute kidney failure - required dialysis
  • RBC damage (hemolytic anemia) - required multiple
    transfusions
  • Platelet consumption - bleeding from IV lines
  • Pancreatitis - weeks of IV nutrition
  • Inflammation of the brain - hallucinations,
    decreased level of consciousness
  • Stool culture positive for

E. coli O157H7
17
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18
Shiga-like Toxins
  • Released by enterohemorrhagic E. coli (E.H.E.C.),
    especially E. coli O157H7, ingested in
    contaminated foods.
  • AB5 structure
  • Toxin released only upon lysis of bacterium
  • Binds to glycolipid receptor on host cells
  • Endothelial cells are predominant target cells -
    esp. in kidneys

19
Shiga-like Toxins (2)
  • Transported in retrograde direction to the ER
  • A subunit translocated into cytoplasm
  • RNA-N-glycosylase activity depurinates 28S
    ribosome - results in inhibition of protein
    synthesis
  • Ultimately causes cell death
  • Since endothelial cells are predominantly
    damaged, coagulation cascade is activated and
    numerous small thrombi form

20
Hemolytic Uremic Syndrome
Fibrin deposition -shearing of RBC
Hemolytic anemia
Tiny blood clots -ischemia
Damage to bowel Damage to kidneys
21
Hemolytic Uremic Syndrome
  • anemia, low platelet count, and kidney failure

22
Case 4
XX. 48 year old cattle rancher from Texas
admitted with difficulty breathing, chest pain
and coughing up blood. Within 12 hrs he
developed rapidly progressive hypotension and
coma. The only exposure history was disposal of
a cow found dead on his farm 10 days ago.
  • There was blood and PMN in his bronchial fluid.
    Chest X-ray did not show pneumonia, but he had an
    enlarged lymph node in chest.
  • Laboratory investigations revealed he had
    meningitis and gram stain was positive for long,
    thin Gram negative organisms.
  • Blood and spinal fluid cultures subsequently grew

Bacillus anthracis
23
Pulmonary Anthrax
24
Anthrax Lethal Factor
  • One of two anthrax toxin A domains that share a
    common B domain. The other A domain is edema
    factor (EF)
  • Released by B. anthracis along with PA and EF
  • PA binds receptor as a monomer, is
    proteolytically cleaved (to 63 kD) and
    oligomerizes x 7
  • PA receptor is a widely expressed 369 aa protein
    - initially called ATR (anthrax toxin receptor),
    now identified as tumor endothelial marker-8 (
    TEM8 )
  • Recently, a second cellular receptor for PA was
    identified as the human capillary morphogenesis
    protein 2 (CMG2)
  • LF (or EF) binds to PA heptamer

25
Anthrax Lethal Factor (2)
  • LF-PA7 is internalized by receptor mediated
    endocytosis
  • Upon acidification of endocytic vesicle, LF is
    translocated thru PA channel
  • LF is a Zn-metalloprotease - cleaves MAPKK and
    possibly other substrates
  • Major cellular target is macrophage (target
    MAPKK, rather than ATR)
  • Upon MAPKK cleavage, inflammatory cytokine
    expression is markedly upregulated, and
    macrophage dies.
  • Resuls is septic shock and impaired phagocytosis
    of B. anthracis

26
Internalization of LF and PA
Heptamer of protective antigen (PA)
27
Pathogenesis of Anthrax
28
Case 4
  • M.V. 17 yr old girl with 2 days of fever,
    malaise, and bright red rash. Muscles ache all
    over. Felt dizzy when got out of bed.
  • In ER, had blood pressure of 70/30.
  • Admitted to hospital. Hypotension did not
    respond to IV fluids. BP dropped to 56 systolic.
  • Transferred to the ICU. Unable to keep oxygen
    saturation above 85 by mask.
  • Intubated and paralyzed to assist ventilation and
    started on pressor medications to support blood
    pressure.
  • When placing a urinary catheter, the nurses
    noticed a thick vaginal discharge. Cultures were
    obtained.

29
Stapylococcus aureus
TSST-1
30
Toxic Shock Syndrome Toxin-1
  • Single-chain, multi-domain protein (but NOT a
    pore-forming toxin)
  • Secreted by some isolates of S.aureus - may only
    be colonizing host
  • Silmultaneously binds MHC class II and T-cell
    receptor (TCR)
  • Target cells are macrophages (and other antigen
    presenting cells) T-cells
  • Causes crosslinking and non-specific activation
    of T-cells (not internalized, is not
    enzymatically active)
  • Secretion of inflammatory cytokines
  • Vasodilation and capillary leak cause rash,
    conjunctivitis, hypotension, fluid in lungs
  • Inflammation and dysregulated blood flow causes
    abnormal function of essentially all organs

31
Toxic Shock Syndrome Toxin-1
TSST-1
32
Case 6
  • A.H. was a 16 year old girl who presented to
    another hospital with a one day history of fever
    and malaise (feeling tired and bad). Examination
    was unremarkable.
  • Bloodwork, including blood culture, was obtained.
    She was observed for four hours then discharged
    to home late in the evening.
  • The next morning her father found her without
    pulse or respiration and she could not be
    resuscitated. There was a diffuse rash
    consisting of large purple macules.
  • The initial blood culture was subsequently
    positive for

Neisseria meningitidis
33
Neisseria meningiditis
34
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35
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36
Endotoxin
  • LPS released by gram negative organisms
  • Binds LPS-binding protein (LBP) and presented to
    CD14 which complexes with TLR-4. LPS can also
    bind directly to TLR-4
  • LPS is not internalized (and is not
    enzymatically active)
  • Activation of TLR-4 ultimately results in
    activation and translocation of NF-KB into
    nucleus
  • Expression of inflammatory cytokines TNFa, IL-1b,
    and others
  • Like TSST-1, causes vasodilation, abnormal blood
    flow to organs, resulting in septic shock,
    multi-organ system failure
  • Endothelial cells particularly affected, activate
    abnormal thrombosis and multiple small clots
    (especially in the skin, which causes the rash)

37
Structure of Gram-negative LPS Endotoxin
38
Pathogenesis of Septic Shock
ARDS adult respiratory distress
syndrome DICdiffuse intravascular coagulation
(multiple tiny blood clots) MOSFmulti-organ
system failure (esp. kidney, liver, intestinal
failure)
39
N. meningitidis - Pathophysiology
Fe
3
Adherence
Invasion
Intracellular
Nutrient
Immune
Toxin






survival
acquisition
evasion
production
Iron scavengers
Endotoxin (LPS)
Capsule IgA proteases
Opa proteins
Pilus
40
Biologic Effects of Bacterial Toxins
  • Damage membranes
  • Inhibit protein synthesis
  • Activate second messenger pathways
  • Damage cytoskeleton
  • Arrest cell cycle
  • Inhibit neurotransmitter release
  • Induce inflammation

41
Mechanisms of Bacterial Toxin Action
  • Pore formation
  • ADP-ribosyltransferase activity
  • Glucosyltransferase activity
  • N-glycosidase activity
  • Adenylcyclase activity
  • Protease activity
  • Deamidase activity
  • DNA damage
  • Superantigen properties
  • Biological effect depends on target molecule.
  • Clinical effect depends on target cell
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