E.coli O104 H4 Shiga Toxins

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Shiga toxins in
E.coli O104 H4

• Dr.T.V.Rao MD

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What are shiga toxins

• Shiga toxins are a family of related toxins with
  two major groups, Stx1 and Stx2, whose genes are
  considered to be part of the genome of lambdoid
  prophages The toxins are named for Kiyoshi
  Shiga, who first described the bacterial origin
  of dysentery caused by Shigella dysenteriae. The
  most common sources for Shiga toxin are the
  bacteria S. dysenteriae and the Shigatoxigenic
  group of Escherichia coli (STEC), which includes
  serotype O157O7 and other enterohemorrhagic E.
  coli (EHEC).

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Shiga-like toxinstructure

• The toxin has two subunitsdesignated A and Band
  is one of the AB5 toxins. The B subunit is a
  pentamer that binds to specific glycolipids on
  the host cell, specifically globotriaosylceramide
  (Gb3). Following this, the A subunit is
  internalised and cleaved into two parts. The A1
  component then binds to the ribosome, disrupting
  protein synthesis. Stx-2 has been found to be
  approximately 400 times more toxic (as quantified
  by LD50 in mice) than Stx-1

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Site 1 of SLT-IBGb3 complex

• Gb3 is, for unknown reasons, present in greater
  amounts in renal epithelial tissues, to which the
  renal toxicity of Shiga toxin may be attributed.
  Gb3 is also found in CNS neurons and endothelium,
  which may lead to neurotoxicity.. Stx-2 is also
  known to increase the expression of its receptor
  GB3 and cause neuronal dysfunctions.

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Side chains conserved in Shiga-like toxin
familySide chains that vary in Shiga-like toxin
family
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Shiga-like toxins

• Shiga toxin family
• Shigella dysenteriae-1 Shiga toxin
• Escherichia coli Shiga-like toxins (SLTs)
• SLT-I - nearly identical to Shiga toxin
• SLT-II variants - 60 identity to SLT-I
• AB5 subunit structure
• A-subunit attacks ribosome enzymatically
• related to ricin
• B-subunit binds to cell-surface glycolipid Gb3
  or Gb4
• Drug target A or B?

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Major types of shiga toxins differs in Shigella
and E.coli

• Shiga toxin (Stx) - true Shiga toxin is produced
  by Shigella dysenteriae. Shiga-like toxin 1 and 2
  (SLT-1 and 2 or Stx-1 and 2) - the Shiga toxins
  produced by some E. coli strains. Stx-1 differs
  from Stx by only 1 amino acid. Stx-2 shares 56
  sequence homology with Stx-1.

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SHIGA TOXINs - how it acts

• SHIGA TOXIN S.DYSENTERIAE AND E.COLI O157
• TWO PART TOXIN PART B BINDS TO CELL AND INJECTS
  PART A WHICH CLEAVES A SPECIFIC ADENINE RESIDUE
  IN THE 60S RIBOSOME PREVENTING PROTEIN SYNTHESIS
  AND CAUSING CELL DEATH

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Shigella toxins affects several animals

• The toxin requires highly specific receptors on
  the cells' surface in order to attach and enter
  the cell species such as cattle, swine, and deer
  which do not carry these receptors may harbor
  toxigenic bacteria without any ill effect,
  shedding them in their feces, from where they may
  be spread to humans.

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Shiga toxin targets kidney

• Once shiga toxin reaches a target organ such as
  the kidney, it binds to receptors on cell
  membranes known as globotriaosylceramide or GB3.
  The toxin is then brought inside the cell and
  transported to the Golgi apparatus, endoplasmic
  reticulum and finally to the nuclear membrane.

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E. coli that cause human gastrointestinal illness

• Enteropathogenic (EPEC)
• Enterotoxigenic (ETEC)
• Enteroinvasive (EIEC)
• Other types, less well characterizedShiga
  toxin-producing (STEC), also called
  Enterohemorrhagic (EHEC)

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Pathogenic mechanisms of E.coli
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E. coli that cause human gastrointestinal illness

• Shiga toxin-producing (STEC), also called
  Enterohemorrhagic (EHEC)
• E. coli O157 Serogroup
• Non-O157 Serogroups
• Enteropathogenic (EPEC)
• Enterotoxigenic (ETEC)
• Enteroinvasive (EIEC)
• Other types, less well characterized

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Basic pathology

• Small infectious dose (lt100 organisms)
• Bacteria adhere to host cell membrane and
  colonize the large intestine
• Produce one or more shigatoxins (Stx1, Stx2)
  which damage endothelial cells

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Pathogenesis ofE. coli food poisoning

• Relevant strains all produce Shiga-like toxins
• particularly O157H7
• Toxin-mediated cell damage leads to thrombosis
  in microvasculature site of damage determines
  pathology

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Animals are the reservoirs for STEC

• Cattle
• Other ruminants
• Other animals
• especially those who have contact with cattle

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Cycle of events in spread of stec Ruminants and
contamination cycle
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Cycle of events in spread of stecSTEC Ruminants
and contamination cycle
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Major modes of transmission of STEC to humans

how the fecal matter gets to the oral
cavity

• Food
• cattle products, e.g., beef, raw milk
• food contaminated with cattle or human feces
  e.g., lettuce
• Water
• Drinking water
• Recreational water
• Animal contact
• contact with farm animals, e.g. petting zoos
• contact with farm animals environment
• Person contact
• With the feces of infected persons

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The foods LOOKS NICEcan cause infections

• RAW EGGS
• UNWASHED SALAD
• CONTAMINATED SURFACES
• UNDERCOOKED
• HAND WOUND
• WARM KITCHEN

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Several bacteria spread through contaminated food

• SALMONELLA
• S.TYPHI
• SHIGELLA
• S.DYSENTERIAE
• CAMPYLOBACTER
• VIBRO CHOLERA
• A.HYDROPHILA
• C.PERFRINGENS
• C.BOTULINUM

• P.SHIGELLOIDIES
• S.AUREUS
• B.CEREUS
• EAEC
• EHEC
• EIEC
• EPEC
• OTHER E.COLI
• B.FRAGILIS

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Sequence of events in E. coli O157H7 infection
non-bloody diarrhea, abdominal cramps
bloody diarrhea
Mead. Lancet 1998
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Sequence of events in non-O157 STEC infection
non-bloody diarrhea, abdominal cramps
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Shiga Toxins

• E-coli isnt usually disease- causing, a major
  commensal in humans.
• Shiga toxin is one of the most potent toxins
  known to man, so much so that the Centers for
  Disease Control and Prevention lists it as a
  potential bioterrorist agent
• Most kinds of E. coli bacteria do not cause
  disease in humans, indeed, some are beneficial,
  and some cause infections other than
  gastrointestinal infections, such urinary tract
  infections.

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What are Shiga toxin-producing E. coli?

• Some kinds of E. coli cause disease by making a
  toxin called Shiga toxin. The bacteria that make
  these toxins are called Shiga toxin-producing
  E. coli, or STEC for short. You might hear them
  called Vero cytotoxic E. coli (VTEC) or
  Enterohemorrhagic E. coli (EHEC) these all refer
  generally to the same group of bacteria. The most
  commonly identified STEC in North America is E.
  coli O157H7 (often shortened to E. coli O157 or
  even just O157). When you hear news reports
  about outbreaks of E. coli infections, they are
  usually talking about E. coli O157.

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How do you get Shiga toxin-producing Escherichia
coli?

• Cases and outbreaks of Shiga toxin-producing
  Escherichia coli have been associated with the
  consumption of undercooked beef (especially
  ground beef), raw milk, unpasteurized apple
  juice, contaminated water, red leaf lettuce,
  alfalfa sprouts, and venison jerky. The bacteria
  have also been isolated from poultry, pork and
  lamb. Person-to- person spread, via fecal-gtoral
  transmission, may occur in high-risk settings
  like day care centers and nursing homes. Further
  studies are being done to better understand the
  modes of transmission.

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Dynamics of shiga toxins

• Shiga toxins act to inhibit protein synthesis
  within target cells by a mechanism similar to
  that of ricin toxin produced by Ricinus
  communis.After entering a cell, the protein
  functions as an N-glycosidase, cleaving a
  specific adenine nucleobase from the 28S RNA of
  the 60S subunit of the ribosome, thereby halting
  protein synthesis.

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How the infected manifest..

• Asymptomatic
• Mild/moderate illness (non-bloody watery
  diarrhea, abdominal pain, rarely vomiting, fever
  usually absent)
• Hemorrhagic colitis (grossly bloody diarrhea,
  severe abdominal pain)
• Hemolytic uremic syndrome (HUS)
• Thrombotic thrombocytopenic pupura(TTP)

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Clinical diagnosis supported by laboratory testing

• Gram-negative, rod shaped
• Clinical samples (typically stool)
• Food samples
• Culture
• Identification of the organism
• Routine culture for 0157 does not detect
  non-0157 STEC
• Enzyme Immunoassay (EIA)
• Detection of shigatoxin (Stx1, Stx2)

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E.coli O104 H4 a toxigenic bacteria
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E.coli O104 H4 an emerging infection

• Although rare, E. coli O104H4 has been
  identified before, in 2009 in the Republic of
  Georgia, culture of specimens from two patients
  who were part of a cluster of diarrheal illness
  yielded an E. coli O104H4 strain that was
  similar to the current outbreak strain. That
  strain produced Shiga toxin, but had a different
  molecular fingerprint and was less resistant to
  antibiotics than the current outbreak strain in
  Germany. No clear outbreak was identified in the
  Republic of Georgia, and no food was identified
  as a source of infections

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Shiga toxin-producing
Escherichia coli (STEC)current outbreak

• Shiga toxin-producing Escherichia coli (STEC) can
  cause severe enteric infections and the
  potentially life threatening hemolytic uremic
  syndrome (HUS).  Prompt diagnosis of these
  infections is important to implement early
  clinical management that minimizes the likelihood
  of developing HUS, to reduce the risk of
  transmitting the infection to others, and to
  detect outbreak
• Commonly consumed vegetables are source of spread.

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New report on E.coli 0104 h4 outbreak

• Large outbreak of Shiga toxin-producing
  Escherichia coli O104H4 (STEC O104H4)
  infections ongoing in Germany. The responsible
  strain shares virulence characteristics with
  enter aggregative E. coli (EAEC). As of June 2,
  2011, case counts confirmed by Germanys Robert
  Koch Institute include 520 patients with
  hemolytic uremic syndrome (HUS) a type of
  kidney failure that is associated with E. coli or
  STEC infections and deaths.

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Vegetables as source of
E.coli infections

• While suspicion has fallen on raw tomatoes,
  cucumbers and lettuce as the source of the germ,
  researchers have been unable to pinpoint the food
  responsible. The outbreak is considered the
  third-largest involving E. coli in recent world
  history, and it is already the deadliest. Twelve
  people died in a 1996 Japanese outbreak that
  reportedly sickened more than 9,000, and seven
  died in a Canadian outbreak in 2000.

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E.coli 0104 h4 resembles
E. coli serotype O157H7

• E. coli serotype O157H7 is a rare variety of E.
  coli that produces toxins which are capable of
  inflicting damage to the lining of the intestine.
  These toxins are closely related or identical to
  the toxin produced by Shigella dysenteriae and
  are referred to as Shiga toxins. In very rare
  instances, the infection can progress to
  hemolytic uremic syndrome ("HUS") and kidney
  failure. E. coli O157H7 can survive at low
  temperatures as well as under acidic conditions,
  and the infectious dose is relatively small.

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E.coli 0104h4 picked up new genes

• t's observed that E. coli O104H4 has picked up
  some new genes, almost certainly through
  horizontal gene transfer, in which stretches of
  DNA are picked up from other E. coli strains, or
  possibly different species entirely. Once
  incorporated into the genome, the new genes can
  provide the bacteria with entirely novel
  properties. In the case of E. coli O104H4, tests
  have shown that it now carries a gene for
  shigatoxin, which is commonly found in other
  disease-causing strains of this species.

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genome OF A KILLER German and Chinese scientists
cracked the genetic code behind the strain of
Escherichia coli
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Need for investigation of cases Goal control
spread of disease

• Determine if case is a threat to spread disease
• Food handler
• Daycare attendee or worker
• Involved in direct patient care
• Determine potential exposures/sources
• Food history
• Travel history
• Recreational water exposure
• Animal exposure

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Handing washing the most essential step

• WASH HANDS thoroughly after using the bathroom
  or changing diapers and before preparing or
  eating food. WASH YOUR HANDS after contact with
  animals or their environments (at farms, petting
  zoos, fairs, even your own backyard).

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avoid

• AVOID raw milk, unpasteurized dairy products, and
  unpasteurized juices (like fresh apple cider).
• AVOID swallowing water when swimming or playing
  in lakes, ponds, streams, swimming pools, and
  backyard kiddie pools.

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Supportive care is the primary need

• Antibiotics might increase the risk for HUS in
  patients infected with O157 STEC, and
  antidiarrheal medications might worsen the
  illness.
• True with 0104H4

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Delayed diagnosis leads to outbreaks

• Delayed diagnosis of STEC infections might lead
  to secondary transmission in homes, child-care
  settings, nursing homes, and food service
  establishments and might delay detection of
  multistate outbreaks related to widely
  distributed foods . Outbreaks caused by STEC with
  multiple Serogroups or PFGE patterns have been
  documented.

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Why drugs are dangerous

• Use of antibiotics other Antidiarrheal
  treatments "can actually make the situation
  worse." That's because killing toxin-producing
  bacteria, such as the ones responsible for this
  outbreak, can actually cause them to release more
  toxins. can actually make the situation worse."
  That's because killing toxin-producing bacteria,
  such as the ones responsible for this outbreak,
  can actually cause them to release more toxins.

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Possible prevention of E.coli O104 H4

• COOK meats and poultry thoroughly. Ground beef
  should reach at least 155 degrees F (until the
  meat is no longer pink and juices run clear).
• DO NOT consume raw milk or unpasteurized dairy
  products.
• WASH YOUR HANDS after using the bathroom or
  changing diapers and before preparing or eating
  food.

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Vegetables too are infectious do believe it ???
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Cdc monitoring events

• CDC is monitoring a large outbreak of Shiga
  toxin-producing Escherichia coli O104H4 (STEC
  O104H4) infections ongoing in Germany. The
  responsible strain shares virulence
  characteristics with enteroaggregative E. coli
  (EAEC).

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• Created by Dr.T.V.Rao MD for
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  Professionals in the Development World
• Email
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