E.coli O104 H4, Shiga Toxins

Shiga toxins in E.coli O104 H4:

Dr.T.V.Rao MD Shiga toxins in E.coli O 104 H 4 Dr.T.V.Rao MD 1

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 O157:O7 and other enterohemorrhagic E. coli (EHEC ). What are shiga toxins Dr.T.V.Rao MD 2

Shiga-like toxin structure:

The toxin has two subunits—designated A and B—and is one of the AB 5 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 Shiga-like toxin structure Dr.T.V.Rao MD 3

Site 1 of SLT-IB:Gb3 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 . Site 1 of SLT-IB:Gb3 complex Dr.T.V.Rao MD 4

Side chains conserved in Shiga-like toxin family Side chains that vary in Shiga-like toxin family:

Side chains conserved in Shiga-like toxin family Side chains that vary in Shiga-like toxin family Dr.T.V.Rao MD 5

Shiga-like toxins:

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? Dr.T.V.Rao MD 6

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. Major types of shiga toxins differs in Shigella and E.coli Dr.T.V.Rao MD 7

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 SHIGA TOXINs - how it acts Dr.T.V.Rao MD 8

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 . Shigella toxins affects several animals Dr.T.V.Rao MD 9

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. Shiga toxin targets kidney Dr.T.V.Rao MD 10

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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) E. coli that cause human gastrointestinal illness Dr.T.V.Rao MD 12

Pathogenic mechanisms of E.coli:

Pathogenic mechanisms of E.coli Dr.T.V.Rao MD 13

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 E. coli that cause human gastrointestinal illness Dr.T.V.Rao MD 14

Basic pathology:

Small infectious dose (<100 organisms) • Bacteria adhere to host cell membrane and colonize the large intestine • Produce one or more shigatoxins (Stx1, Stx2) which damage endothelial cells Basic pathology Dr.T.V.Rao MD 15

Pathogenesis of E. coli food poisoning:

Relevant strains all produce Shiga-like toxins • particularly O157:H7 • Toxin-mediated cell damage leads to thrombosis in microvasculature site of damage determines pathology Pathogenesis of E. coli food poisoning Dr.T.V.Rao MD 16

Animals are the reservoirs for STEC :

Cattle Other ruminants Other animals especially those who have contact with cattle Animals are the reservoirs for STEC Dr.T.V.Rao MD 17

Cycle of events in spread of stec Ruminants and contamination cycle :

Cycle of events in spread of stec Ruminants and contamination cycle Dr.T.V.Rao MD 18

Cycle of events in spread of stec STEC Ruminants and contamination cycle :

Cycle of events in spread of stec STEC Ruminants and contamination cycle Dr.T.V.Rao MD 19

Major modes of transmission of STEC to humans – how the fecal matter gets to the oral cavity :

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 Dr.T.V.Rao MD 20

The foods LOOKS NICE can cause infections:

The foods LOOKS NICE can cause infections RAW EGGS UNWASHED SALAD CONTAMINATED SURFACES UNDERCOOKED HAND WOUND WARM KITCHEN Dr.T.V.Rao MD 21

Several bacteria spread through contaminated food:

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 Dr.T.V.Rao MD 22

Sequence of events in E. coli O157:H7 infection :

Sequence of events in E. coli O157:H7 infection E. coli O157 ingested 3 - 4 days non-bloody diarrhea, abdominal cramps 5 - 6 days resolution 92% 8% HUS bloody diarrhea 1 - 2 days 80% Mead. Lancet 1998 Dr.T.V.Rao MD 23

Sequence of events in non-O157 STEC infection :

Non-O157 STEC ingested 3 - 4 days 5 - 6 days resolution 98%? bloody diarrhea 1 - 2 days 40% Sequence of events in non-O157 STEC infection non-bloody diarrhea, abdominal cramps rare HUS Dr.T.V.Rao MD 24

Shiga Toxins:

E-coli isn’t 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. Shiga Toxins Dr.T.V.Rao MD 25

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 O157:H7 (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. What are Shiga toxin-producing E. coli ? Dr.T.V.Rao MD 26

How do you get Shiga toxin-producing Escherichia coli?:

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->oral 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. Dr.T.V.Rao MD 27

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 . Dynamics of shiga toxins Dr.T.V.Rao MD 28

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) How the infected manifest.. Dr.T.V.Rao MD 29

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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) Clinical diagnosis supported by laboratory testing Dr.T.V.Rao MD 31

E.coli O104 H4 a toxigenic bacteria:

E.coli O104 H4 a toxigenic bacteria Dr.T.V.Rao MD 32

E.coli O104 H4 an emerging infection :

E.coli O104 H4 an emerging infection Although rare, E. coli O104:H4 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 O104:H4 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 Dr.T.V.Rao MD 33

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. Shiga toxin-producing Escherichia coli ( STEC) current outbreak Dr.T.V.Rao MD 34

New report on E.coli 0104 h4 outbreak :

Large outbreak of Shiga toxin-producing Escherichia coli O104:H4 (STEC O104:H4) 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 Germany’s 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. New report on E.coli 0 104 h 4 outbreak Dr.T.V.Rao MD 35

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. Vegetables as source of E.coli infections Dr.T.V.Rao MD 36

E.coli 0104: h4 resembles E. coli serotype O157:H7:

E. coli serotype O157:H7 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 O157:H7 can survive at low temperatures as well as under acidic conditions, and the infectious dose is relatively small. E.coli 0104: h4 resembles E . coli serotype O157:H7 Dr.T.V.Rao MD 37

E.coli 0104:h4 picked up new genes :

t's observed that E. coli O104:H4 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 O104:H4, tests have shown that it now carries a gene for shigatoxin, which is commonly found in other disease-causing strains of this species. E.coli 0 104: h 4 picked up new genes Dr.T.V.Rao MD 38

genome OF A KILLER: German and Chinese scientists cracked the genetic code behind the strain of Escherichia coli :

genome OF A KILLER : German and Chinese scientists cracked the genetic code behind the strain of Escherichia coli Dr.T.V.Rao MD 39

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 Need for investigation of cases Goal: control spread of disease Dr.T.V.Rao MD 40

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 ). Handing washing the most essential step Dr.T.V.Rao MD 41

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. avoid Dr.T.V.Rao MD 42

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 0104:H4 Supportive care is the primary need Dr.T.V.Rao MD 43

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. Delayed diagnosis leads to outbreaks Dr.T.V.Rao MD 44

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. Why drugs are dangerous Dr.T.V.Rao MD 45

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. Possible prevention of E.coli O104 H4 Dr.T.V.Rao MD 46

Vegetables too are infectious do believe it ???:

Vegetables too are infectious do believe it ??? Dr.T.V.Rao MD 47

Follow me for Articles of Interest on infectious diseases and Microbiology ..:

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Cdc monitoring events :

CDC is monitoring a large outbreak of Shiga toxin-producing Escherichia coli O104:H4 (STEC O104:H4) infections ongoing in Germany. The responsible strain shares virulence characteristics with enteroaggregative E. coli (EAEC). Cdc monitoring events Dr.T.V.Rao MD 49

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