Download E coli

E coli, Klebsiella and
Enterobacter
Prof M.I.N. Matee
School of Medicine
MUCHS
Escherichia coli
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Normal inhabitant of the G.I. tract
Some strains cause various forms of
gastroenteritis.
Is a major cause of urinary tract infection and
neonatal meningitis and septicemia.
Morphology and culture
characteristics
Morphology
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Gram negative rods
Non-spore former, non-caspulated and motile
Culture characteristics
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N.agar – circular, convex, small colonies
MacConkey medium– rose pink
Eosin Methylene blue – Metallic sheen
colonies
Biochemical reactions
IMViC: ++-TSI: Acid butt/acid slant with gas
production
Antigenic structure
Has O, H, and K antigens.
K1 has a strong association with virulence,
particularly meningitis in neonates.
Antigenic structure.
Complex
•Lipopolisaccharides/Somatic or O antigen
• Heat stable.more than 150 types
•Most external in the cell wall
•detected by bacterial agglutination
•Antibody produced is predominantly IgM
Capsular/K antigen
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Sometimes external to O antigen but
not always
Can be polysaccharides or protein
Flagella /H antigen
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Heat and alcohol labile.
Colicines/Bacteriocines
• Produced by many Gram -ves
• Virus like bactericidal substance
•Active against some other bacteria of similar
or closely related species
E. coli toxins
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Virulence factors
Toxins
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Enterotoxins – produced by enterotoxigenic strains of E. coli
(ETEC). Causes a movement of water and ions from the
tissues to the bowel resulting in watery diarrhea. There are two
types of enterotoxin:
LT – is heat labile and binds to specific Gm1 gangliosides
on the epithelial cells of the small intestine where it
stimulates production of cAMP.
Increased cAMP alters the activity of sodium and chloride
transporters producing an ion imbalance that results in fluid
transport into the bowel.
E. coli toxins
ST – is heat stable and binds to
specific receptors to stimulate the
production of cGMP with the same
results as with LT.
Urinary tract infections
E coli is the most common cause of UTI
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Account for 90% of cases in young women
Symptoms
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Frequency, dysuria, hematuria, pyuria
UTI may lead to bacteremia and sepsis
Nephropathogenic E coli produces
hemolysin
E coli associated diarrhoeal diseases
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
A.Enterotoxigenic E coli ( ETEC)
 Common cause for travelers diarrhoea, and
watery diarrhoea in children.
Colonisation factor facilitates the attachment to
the intestinal epithelium.
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Some ETEC produces heat labile exotoxin LT and
heat stable or either of the toxins
B.Enteropathogenic E coli (EPEC)
 Important cause of diarrhoea in infants of
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developing countries.
Adhere to mucosal cells in small bowel,loss of
microvilli, NONINVASIVE
enter to cell body. result
in watery
diarrhoea.
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Self limiting ,can be chronic.
Normally do not produce toxins
C. Enterohemorrhagic E coli ( EHEC)
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Produce verotoxin which has similarities
to Shiga toxin
Associated with hemorrhagic
colitis, severe form of diarrhoea.
Hemolytic uremic syndrome Disease can
be prevented by thorough cooking.
Escherichia coli O157:H7
This strain produces a powerful toxin, verotoxin, which can
cause severe illness and death.
E coli O157:H7 does not ferment sorbitol and is negative on
sorbitol MacConkey agar
Other Shiga toxin producing serotypes (e.g. O111 and O26)
are also in the family of enterohemorrhagic E. coli and can
cause similar disease.
Hemorrhagic colitis – preventable by thorough cooking of meat
Hemorrhagic colitis(HC) or bloody diarrhea and
abdominal cramps
Lasts for 5-10 days usually
Infection may proceed to Hemolytic Uremic
Syndrome (HUS), a severe cytopathic attack on
the kidneys requiring intensive care and dialysis.
Red blood cells are destroyed requiring blood
tranfusion
D.Enteroinvasive E coli (EIEC)
• Produces disease similar to shigellosis.
•In adults this has been isolated with Shigella
•Commonly affect children
and travelers.
in developing countries,
Disease is due to invasion into mucosal cells of the intestine
multiply inside the cells and destruction /inflammation/ulceration
diarrhoea with blood
EIEC are nonlactose fermenter,or late lactose fermenter
and non motile.
E. Enteroaggregative E coli (EAEC)
Produce acute/chronic diarrhoea in persons in
developing countries.
Sepsis When normal host defense is poor,
sepsis can happen. Common in new born babies
whose IgM level is low.
Treatment of E.coli related diarrhoea
1st Line
Nitrofurantoin
Nalidixic acid
Norfloxacin
Ampicillin
Cotrimoxazole
2nd line
Ciprofloxacin/Ceftriaxone/Cefuroxime
Gentamicin
Meningitis
• E coli and Gp.B Strept. are the leading causes for
meningitis in infants.
•K1 antigen is responsible for meningitis
K1 cross reacts with the Gp.B capsular
polysaccharides of N meningitides.
Pneumonia
25% of gram -ve pneumonia with 50%
mortality
Usually broncho pneumonia
High level of resistance to Ampicillin
/Cotrimoxazole
Klebsiella spp
Morphology and culture
characteristics
Morphology
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Gram negative rods
Non-spore former, caspulated and non-motile
Culture characteristics
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N.agar – mucoid, circular, convex, small
colonies
MacConkey – mucoid, rose pink
Biochemical reactions
IMViC: --++
TSI: Acid butt/acid slant with gas
production
K pneumoniae and K oxytoca Hospital acquired infec
K rhinoscleromatis produces rhinoscleroma,
condition with destructive granuloma of the nose and
pharynx.
K ozenae – progressive atrophy of nasal mucous membrane
Klebsiella
K pneumoniae
Present in respiratory tract and
feces of about 5% of
normal individuals.
Can cause bacterial pneumonia
(3%).
Produce extensive hemorrhagic
necrotising consolidation of lungs.
Enterobacter aerogenus
Capsulated
Free living in the intestine
Cause UTI and sepsis.
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Motile
Citrate +
Ornithine decarboxylase
Produce gas from glucose
Ferments lactose
VP + (like Klebsiella)
Laboratory identification
tests
Diagnosis of Bacterial
Infection
microscopy
unstained or stained with e.g.
Gram stain
Stain
Decolorise
Counterstain
identification by biochemical or
serological tests on pure growth
from single colony
culture
on plates or in broth
sensitivities
by disc diffusion
methods,
breakpoints or
MICs
Serodiagnosis
DNA technologies
Gram negative rods
Lactose fermentor on MCA
Lactose positive Klebsiella sp
E. coli
EMB agar—Selective AND Differential
 Eosine and methylene blue block are dyes that
block the growth of gram (+) bacteria.
Why are gram (-) bacteria more resistant?
 E. coli forms blue-black colonies
with a green metalic sheen. (due to lactose fermentation)
 E. aerogenes forms large mucoid pink to
purple colonies with no metallic sheen.
 Salmonella large forms pink colonies.
Indole production
• Degradation of tryptophan by tryptophanase to produce indole
• Red colour develops after adding a solution containing pdimethylaminobenzaldehyde (Kovacs or Erlichs)
Negative
Positive
Triple Sugar Iron Agar (TSI):
Purpose: To differentiate
bacteria based on their ability
to ferment glucose, lactose
and/or sucrose, and to
reduce sulfur to hydrogen
sulfide.
INTERPRETATION OF TUBES ABOVE
TUBE 1
(UNINOCULATED)
TUBE 2
TUBE 3
TUBE 4
TUBE 5
SLANT
-
A
A
K
K
BUTT
-
A
A
A
A
HYDROGEN
SULFIDE
-
-
-
+
+
GAS
-
-
+
-
-
A=Acidic K=Alkaline
ONPG test
• Determines presence of -galactosidase
• Utilizes o-nitrophenyl-d--galactosidase
• Differentiates late lactose fermenting organisms
Positive
Negative
Methyl Red
This is a qualitative test of the acidity
produced by the growth of a bacterium in
phosphate –buffered glucose peptone water
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E coli produces a pH of about 5, and hence a red
colour is produced after addition of methyl red.
Enterobacter aerogenes the pH never drops so
low, it appears yellow after addition of methyl red.
Voges-Proskauer
This test for production of
acetylmethylcarbinol (AMC) from
carbohydrates by bacteria in glucose
phosphate peptone water
Red colour is produced after addition of
alpha naphthol and KOH
Klebsiella +
E coli -
MR/VP continued
Reading Results:
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MR— a + result is red (indicating pH below 6) and a – result is yellow
(indicating no acid production)
VP—A + result is red after VP reagents are added (indicating the
presence of acetoin) and a – result is no color change.
Methyl Red: left – and right +
VP: left + and right –
IMViC test
IMViC test is a group of four biochemical
tests collectively used for primary
identification of enteric bacteria
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Indole
Methyl red
Voges proskuer
Citrate utilization test
Indole (IMViC tests)
• E. coli (pink/red) +
• E. coli (left side) –
• Kovac’s reagent
detects if tryptophan
has been hydrolyzed
to indol/tryptophanase
Methyl Red (MR) (IMViC tests)
Enterobacter
aerogenes (left) –
E. coli (bright red) +
Reagent: Methyl red
indicator identifies pH
change due to mixed
acid fermentation
Voges – Proskauer (VP)
(IMViC tests)
Enterobacter aerogenes
+(left)
E. coli – (right)
Red colour is produced
after addition of alpha
naphthol and KOH
Klebsiella +
E coli -
Citrate utilization
• Ability to use sodium citrate as sole source of carbon
• Medium has sodium citrate, ammonium salt, bromothymol blue
• Positive test - deep blue colour in 24-48hrs
• Indicates citrate utilization
• Production of alkaline products
E. coli (left green) –
• Enterobacter aerogenes
(right royal blue) +
Positive
Negative
Treatment of enteric bacteria
No specific treatment is available
A number of antibiotics could be used
Marked antibiotic resistance due to
plasmids
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May involve surgical correction
Treatment of shock
Environmental sanitation
References:
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