ENTEROBACTERIACEAE

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ENTEROBACTERIACEAE
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Morphology Identification

• Gram-negative non-spore forming rods. When
  motile, by peritrichous flagella.
• Primarily normal flora of gastrointestinal tract.
  E. coligtKlebsiellagtProteusgtEnterobacter
• Free living, also transient colonizers of skin.
• Facultative anaerobes mixed acid fermentation
• All ferment glucose all reduce nitrates to
  nitrites all oxidase negative.
• Lactose fermentation normal flora positive and
  pathogens negative.
• Primary isolation media include
  eosin-methylene-blue (EMB) and MacConkey agar.
• Differential selective media for specific
  organisms including dyes and bile salts.
  (Salmonella-Shigella (SS) medium, bismuth sulfite
  media.)

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Classification29 genera, over 100 species.

• Escherichia
• Shigella
• Edwardsiella
• Salmonella
• Citrobacter
• Klebsiella
• Enterobacter
• Hafnia
• Serratia

• Proteus
• Providencia
• Morganella
• Yersinia
• Erwinia
• Pectinobacterium

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Antigenic Structure

• Most are motile by peritrichous flagella --H
  antigens.
• Capsule K antigen ( Vi for Salmonella).
• Cell envelope (wall)
• LPS (endotoxin) O antigen.
• various outer membrane proteins.
• Pili - various antigen types, some encoded by
  plasmids

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????(H)
K?Vi??
????(O)
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Opportunistic diseases -Enterobacteriaceae

• septicemia,
• pneumonia,
• meningitis
• urinary tract infections

Citrobacter Enterobacter Escherichia Hafnia Morgan
ella Providencia Serratia
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Enterobacteriaceae gastrointestinal diseases

• Escherichia coli
• Salmonella
• Shigella
• Yersinia entercolitica

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Reiter's syndrome

• Histocompatibility antigen (HLA) B27
• Enterobacteriaceae
• Salmonella
• Shigella
• Yersinia

• Not Enterobacteriaceae
• Campylobacter
• Chlamydia

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Enterobacteriaceae

• community acquired
• otherwise healthy people
• Klebsiella pneumoniae
• respiratory diseases
• prominent capsule
• urinary tract infection
• fecal contamination
• E. coli
• Proteus
• urease (degrades urea)
• alkaline urine

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Enterobacteriaceae

• gram negative facultative anaerobic rods
• oxidase negative (no cytochrome oxidase)

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Feces

• E. coli
• lactose positive
• not usually identified
• lactose positive sp. common, healthy intestine
• Shigella, Salmonella,Yersinia
• lactose negative
• identified

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Enterobacteriaceae

• other sites
• identified biochemically

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Serotypes

• reference laboratory
• antigens
• O (lipopolysaccharide)
• H (flagellar)
• K (capsular)

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Escherichia coli
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Escherichia coli

• Toxins two types of enterotoxin Shiga-type
  toxin Enteroaggregative ST-like toxin
  Hemolysins Endotoxin
• Type III secretion system
• Adhesions colonization factors both pili or
  fimbriae non-fimbrial factors involved in
  attachment. There are at least 21 different
  types of adhesions.
• Virulence factors that protect the bacteria from
  host defenses Capsule/Iron capturing ability
  (enterochelin)
• Outer membrane proteins

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E. coli fimbriae
Type 1
mannose
P

• galactose
• glycolipids
• glycoproteins

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E.coli-urinary tract infection Is the
leading cause of urinary tract infections which
can lead to acute cystitis (bladder infection)
and pyelonephritis (kidney infection).
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E.coli-Meningitis and Sepsis

• Neonatal meningitis is the leading cause of
  neonatal meningitis and septicemia with a high
  mortality rate. Usually caused by strains with
  the K1 capsular antigen.

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Enteropathogenic E. coli

• fever
• infant diarrhea
• vomiting
• nausea
• non-bloody stools
• Destruction of surface microvilli
• loose attachment mediated by bundle forming
  pili (Bfp)
• Stimulation of intracellular calcium level
  
• rearrangement of intracellular actin,

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Enterotoxigenic E. coli

• A watery diarrhea, nausea, abdominal cramps and
  low-grade fever for 1-5 days.
• Travellers diarrhea and diarrhea in children in
  developing countries
• Transmission is via contaminated food or water.

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Enterotoxigenic E. coli

• diarrhea like cholera
• milder
• nursery travellers diarrhea
• caused by LT, ST, or LT/ST.

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Enterotoxigenic E. coli

• Heat labile toxin
• like choleragen
• Adenyl cyclase activated
• cyclic AMP
• secretion water/ions
• Heat stable toxin
• Guanylate cyclase activated
• cyclic GMP
• uptake water/ions

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LT vs ST activity
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E.coli-Enteroinvasive (EIEC)

• The organism attaches to the intestinal mucosa
  via pili
• Outer membrane proteins are involved in direct
  penetration, invasion of the intestinal cells,
  and destruction of the intestinal mucosa.
• There is lateral movement of the organism from
  one cell to adjacent cells.
• Symptoms include fever,severe abdominal cramps,
  malaise, and watery diarrhea followed by scanty
  stools containing blood, mucous, and pus.
• resembles shigellosis

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Enteroinvasive E. coli (EIEC)

• Dysentery
• resembles shigellosis
• elder children and adult diarrhea

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E.coli-c. Enteropathogenic (EPEC)

• Malaise and low grade fever diarrhea, vomiting,
  nausea, non-bloody stools
• Bundle forming pili are involved in attachment to
  the intestinal mucosa.
• This leads to changes in signal transduction in
  the cells, effacement of the microvilli, and to
  intimate attachment via a non-fimbrial adhesion
  called intimin.
• This is a problem mainly in hospitalized infants
  and in day care centers.

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E.coli-d. Enterohemorrhagic (EHEC)

• Hemorrhagic
• bloody, copious diarrhea
• few leukocytes
• afebrile
• hemolytic-uremic syndrome
• hemolytic anemia
• thrombocytopenia (low platelets)
• kidney failure

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Enterohemorrhagic E. coli

• Usually O157H7

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Enterohemorrhagic E. coli

• Vero toxin
• shiga-like
• Hemolysins
• younger than 5 years old,causing hemorrhagic
  colitis

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Enteroaggregative E. coli ????????

• a cause of persistent, watery diarrhea with
  vomiting and dehydration in infants.
• That is autoagglutination in a stacked brick
  arrangement.
• the bacteria adheres to the intestinal mucosa and
  elaborates enterotoxins (enteroaggregative
  heat-stable toxin, EAST).
• The result is mucosal damage, secretion of large
  amounts of mucus, and a secretory diarrhea.

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E.coli-Enteroaggregative (EAggEC)

• Mucous associated autoagglutinins cause
  aggregation of the bacteria at the cell surface
  and result in the formation of a mucous biofilm.
• The organisms attach via pili and liberate a
  cytotoxin distinct from, but similar to the ST
  and LT enterotoxins liberated by ETEC.
• Symptoms incluse watery diarrhea, vomiting,
  dehydration and occasional abdominal pain.

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Various Types of E. coli
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Summary of E.coli strains that cause
gastroenteritis.
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Sanitary significance

• Totoal bacterial number number of bacteria
  contained per ml or gm of the sample the
  standard of drinking water is less than 100.
• Coliform bacteria index the number of coliform
  bacteria detected out per 1000 ml sample the
  standard of drinking water is less than 3

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Escherichia coli

• Genetically E. coli and Shigella are genetically
  highly closely related. For practical reasons
  (primarily to avoid confusion) they are not
  placed in the same genus. Not surprisingly there
  is a lot of overlap between diseases caused by
  the two organisms.
• 1) Enteropathogenic E. coli (EPEC). Certain
  serotypes are commonly found associated with
  infant diarrhea. The use of gene probes has
  confirmed these strains as different from other
  groups listed below. There is a characteristic
  morphological lesion with destruction of
  microvilli without invasion of the organism that
  suggests adhesion is important. Clinically one
  observes fever, diarrhea, vomiting and nausea
  usually with non-bloody stools.
• 2) Enterotoxigenic E. coli (ETEC) produce
  diarrhea resembling cholera but much milder in
  degree. Also cause "travelers diarrhea". Two
  types of plasmid-encoded toxins are produced. a)
  Heat labile toxins which are similar to
  choleragen (see cholera section below). Adenyl
  cyclase is activated with production of cyclic
  AMP and increased secretion of water and ions. b)
  Heat stable toxins guanylate cyclase is
  activated which inhibits ionic and water uptake
  from the gut lumen. Watery diarrhea, fever and
  nausea result in both cases.
• 3) Enteroinvasive E. coli (EIEC) produce
  dysentery (indistinguishable clinically from
  shigellosis, see bacillary dysentery below).
• 4) Enterohemorrhagic E. coli (EHEC). These are
  usually serotype O157 H7. These organisms can
  produce a hemorrhagic colitis (characterized by
  bloody and copious diarrhea with few leukocytes
  in afebrile patients). Outbreaks are often caused
  by contaminated hamburger meat. The organisms can
  disseminate into the bloodstream producing
  systemic hemolytic-uremic syndrome (hemolytic
  anemia, thrombocytopenia and kidney failure).
  Production of Vero toxin (biochemically similar
  to shiga toxin thus also known as "shiga-like")
  is highly associated with this group of
  organisms encoded by a phage. Hemolysins
  (plasmid encoded) are also important in
  pathogenesis.
• As noted above, there are at least 4
  etiologically distinct diseases. However, in the
  diagnostic laboratory generally the groups are
  not differentiated and treatment would be on
  symptomatology. Generally fluid replacement is
  the primary treatment. Antibiotics are generally
  not used except in severe disease or disease that
  has progressed to a systemic stage
  (e.g.hemolytic-uremia syndrome). Two major
  classes of pili are produced by E. coli mannose
  sensitive and mannose resistant pili. The former
  bind to mannose containing glyocoproteins and the
  latter to cerebrosides on the host epithelium
  allowing attachment. This aids in colonization by
  E. coli.

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Shigella
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Shigella

• S. flexneri, S. boydii, S. sonnei, S. dysenteriae
• bacillary dysentery
• shigellosis
• bloody feces
• intestinal pain
• pus

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Genral features

• Pili.
• Most strains can not ferment lactose S. sonnei
  can slowly_ ferment lactose.
• According to O antigen, 4 groups
• Easily causing drug-resistence.

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Shigellosis

• within 2-3 days
• epithelial cell damage

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Shiga toxin

• enterotoxic
• cytotoxic
• inhibits protein synthesis
• lysing 28S rRNA

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Shigella attachment and penetration

• Within 2-3 days
• Epithelial cell damage

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Clinical significance

• man only "reservoir"
• mostly young children
• fecal to oral contact
• children to adults
• transmitted by adult food handlers
• unwashed hands

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Clinical significance

• The infective dose required to cause infection is
  very low (10-200 organisms).
• There is an incubation of 1-7 days followed by
  fever, cramping, abdominal pain, and watery
  diarrhea (due to the toxin)for 1-3 days.
• This may be followed by frequent, scant stools
  with blood, mucous, and pus (due to invasion of
  intestinal mucosa).
• Is is rare for the organism to disseminate.
• The severity of the disease depends upon the
  species one is infected with. S. dysenteria is
  the most pathogenic followed by S. flexneri, S.
  sonnei and S. boydii.

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Immunity

• SIgA.

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Diagnosis of Shigella infection

• Specimen stool.
• Culture and Identification
• Quick immunological methods
• Immunofluorescent ball test
• Coagglutination.

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Prevention

• streptomycin dependent (SD) dysentery vaccine.

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Treating shigellosis

• manage dehydration
• patients respond to antibiotics , Problem of
  drug-resistance
• disease duration diminished

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Shigella

• Shigella (4 species S. flexneri, S. boydii, S.
  sonnei, S. dysenteriae) all cause bacillary
  dysentery or shigellosis, (bloody feces
  associated with intestinal pain). The organism
  invades the epithelial lining layer, but does not
  penetrate. Usually, within 2-3 days, dysentery
  results from bacteria damaging the epithelium
  lining layers of the intestine often with release
  of mucus and blood (found in the feces) and
  attraction of leukocytes (also found in the feces
  as "pus"). Shiga toxin (chromosomally encoded) is
  neurotoxic, enterotoxic and cytotoxic plays a
  role. The toxin inhibits protein synthesis
  (acting on the 80S ribosome and lysing 28S rRNA).
  This is primarily a disease of young children
  occurring by fecal-oral contact. Adults can catch
  this disease from children. However it can be
  transmitted by infected adult food handlers,
  contaminating food. The source in each case is
  unwashed hands. Man is the only "reservoir".
• Patients with severe dysentery are usually
  treated with antibiotics (e.g. ampicillin). In
  contrast to salmonellosis, patients respond to
  antibiotic therapy and disease duration is
  diminished.

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Salmonella

• Salmonellosis may present as one of several
  syndromes including gastroenteritis, enteric
  (typhoid) fever or septicemia.

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The antigenic structures of salmonellae used in
serologic typing
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Salmonella

• 2000 antigenic "types
• disease category
• S. enteritidis
• many serotypes
• S. cholerae-suis
• S. typhi

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Virulence factors

• Endotoxin may play a role in intracellular
  survival
• Capsule (for S. typhi and some strains of S.
  paratyphi)
• Adhesions both fimbrial and non-fimbrial
• Type III secretion systems and effector molecules
  2 different systems may be found
• One type is involved in promoting entry into
  intestinal epithelial cells
• The other type is involved in the ability of
  Salmonella to survive inside macrophages
• Outer membrane proteins - involved in the ability
  of Salmonella to survive inside macrophages
• Flagella help bacteria to move through
  intestinal mucous
• Enterotoxin - may be involved in gastroenteritis
• Iron capturing ability

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Enteric or typhoid fever

• Enteric or typhoid fever occurs when the bacteria
  leave the intestine and multiply within cells of
  the reticuloendothelial system.
• The bacteria then re-enter the intestine, causing
  gastrointestinal symptoms.
• Typhoid fever has a 10-14 day incubation period
  and may last for several weeks.
• Salmonella typhi is the most common species
  isolated from this salmonellosis.
• Human reservoircarrier state common
• Contaminated foodwater supply
• Poor sanitary conditions

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Typhoid

• Septicemia
• -occurs 10-14 days
• lasts 7 days

• gall bladder
• shedding, weeks

• acute phase, gastroenteritis

gastrointenteritis
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Typhoid -Therapy

• Antibiotics
• essential
• Vaccines
• Vi (capsular) antigen protective

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Salmonella gastroenteritis

• Salmonella gastroenteritis is the most common
  form of salmonellosis and generally requires an
  8-48 hour incubation period and may last from 2-5
  days.
• Symptoms include nausea, vomiting and diarrhea
  (non-bloody stool). Salmonella enteritidis is the
  most common isolate.
• poultry??, eggs. no human reservoir
• self-limiting (2 - 5 days)

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Salmonella septicemia

• Salmonella septicemia (bacteremia) may be caused
  by any species but S. cholerae-suis is common.
  This disease resembles other Gram-negative
  septicemias and is characterized by a high,
  remittent fever with little gastrointestinal
  involvement.

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Immunity (S. typhi)

• Vi (capsular) antigen
• protective

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Diagnosis

• A. Specimens
• a) Enteric fever blood, bone marrow, stool,
  urine.
• b) Food poisoning stool, vomitus, suspected
  food.
• c) Septicemia blood.
• B. Culture and identification
• C. Widal test

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Salmonella

• Using appropriate antibodies more than 2000
  antigenic types have been recognized. There
  are, however, only a few types that are commonly
  associated with characteristic human diseases
  (most simply referred to as S. enteritidis, S.
  cholerae-suis and S. typhi).
• Salmonellosis, the common salmonella infection,
  is caused by a variety of serotypes (S.
  enteritidis) and is transmitted from contaminated
  food (such as poultry and eggs). It does not have
  a human reservoir and usually presents as
  gastroenteritis (nausea, vomiting and non-bloody
  stools). The disease is usually self-limiting
  (2-5 days). Like Shigella they invade the
  epithelium and do not produce systemic infection.
  In uncomplicated cases of salmonellosis, which
  are the vast majority, antibiotic therapy is not
  useful. S. cholerae-suis (seen much less
  commonly) causes septicemia after invasion. In
  this case, antibiotic therapy is required. .
• The severest form of salmonella infections
  "typhoid" (enteric fever), caused by Salmonella
  typhi. Although it is one of the historical
  causes of widespread epidemics and still is in
  the third world. The organism is transmitted from
  a human reservoir or in the water supply (if
  sanitary conditions are poor) or in contaminated
  food. It initially invades the intestinal
  epithelium and during this acute phase,
  gastrointestinal symptoms are noted. The organism
  penetrates, usually within the first week, and
  passes into the bloodstream where it is
  disseminated in macrophages. Typical features of
  a systemic bacterial infection are noted. The
  septicemia usually is temporary with the organism
  finally lodging in the gall bladder. Organisms
  are shed into the intestine for some weeks. At
  this time the gastroenteritis (including
  diarrhea) is noted again. The Vi (capsular)
  antigen plays a role in the pathogenesis of
  typhoid. A carrier state is common thus one
  person e.g. a food handler can cause a lot of
  spread. Antibiotic therapy is essential. Vaccines
  are not widely effective and not generally used

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Klebsiella

• NF of GI tract, but potential pathogen in other
  areas
• Virulence factors
• Capsule
• Adhesions
• Iron capturing ability
• Clinical significance
• Causes pneumonia, mostly in immunocompromised
  hosts. Permanent lung damage is a frequent
  occurrence (rare in other types of bacterial
  pneumonia)
• A major cause of nosocomial infections such as
  septicemia and meningitis

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Klebsiella

• K. pneumoniae (Friedlander bacilli) may cause
  primary pneumonia, urinary tract and wound
  infections, bacteremia, meningitis, etc.
• K. rhinoscleromatis pathogen of granumatous
  destruction of nose and pharynx.
• K. ozaenae causes chronic atrophic rhinitis.

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Proteus

• General characteristics swarming phenomenon on
  nonselective agar (P.vulgaris P.mirabilis and
  P.myxofaciens)
• P.vulgaris strains (OX-19, OX-K, OX-2)have common
  antigen with Rickettsia (Weil-Felix test).
• urinary tract infections food poisoning.