E. coli and Klebsiella spp. can be identified through laboratory tests of specimens like urine, pus, blood, and sputum. On Gram stain, both appear as gram-negative rods. On blood agar, E. coli forms smooth, grayish-white colonies, some hemolytic, while Klebsiella forms slimy colonies. On MacConkey agar, E. coli forms red-pink colonies indicating lactose fermentation, and Klebsiella forms mucoid pink colonies. Biochemical tests can further identify E. coli as motile, indole-positive and citrate-negative, while Klebsiella is non-motile, indole-negative and citrate

1. Escherichia coli
&
Klebsiella spp.
Prepared by
Dr. Samira Fattah
PhD in Medical Bacteriology
College of Health Sciences-HMU

2. Laboratory diagnosis:
Specimen:
• Urine.
• Pus
• Blood
• CSF
• Sputum
• Stool for E. coli

3. Gram stain
• Appears as gram negative, rods shape under microscope.

4. On Blood Agar
• E. coli appear as medium sized, smooth, round,
grayish-white colonies on blood agar.
• Some strains produce beta hemolysis.
Klebsiella appearances as slimy
colonies

5. On MacConkey Agar
• E. coli appears as red to pink colonies.
• the red color of the bacterial growth indicates E. coli is able to ferment
lactose sugar in the media.
Mucoid, lactose positive colonies of Klebsiella
pneumoniae on MacConkey agar. Cultivation 37°C,
24 hours.

6. On Eosin Methylene Blue
• On EMB E. coli give a distinctive blue-black colonies with a greenish metallic sheen
(due to the metachromatic properties of the dyes)
• The selective/ inhibitory agents of EMB are the dyes eosin Y and methylene blue.
Methylene blue inhibits the gram + bacteria , while eosin changes color, to a dark
purple, when the medium around the colony becomes acidic.
Klebsiella species produces large, mucoid, pink to purple
colonies with no metallic green sheen on EMB agar

7. • Oxidase test
Oxidase –negative for both of E. coli and Klebsiella
Biochemical test

8. Escherichia coli
motile
Motility test
• Escherichia coli is highly motile
and will show turbidity
throughout the tube.
Klebsiella pneumoniae
Non motile
Biochemical test

9. String test
• A colony that stretches more
than 5 mm using a standard
inoculation loop tests positive
for hypermucoviscosity.
Mucoid colony of Klebsiella pneumoniae .
When colonies were touched with a loop and the
loop lifted vertically from the surface of the agar
plate, mucoid isolates adhered to the loop as it
was lifted from the plate .

10. India ink capsule stain
India Ink Capsule Stain of Klebsiella pneumoniae showing
white capsules surrounding purple cells
-The background will be dark.
-The bacterial cells will be stained purple.
-The capsule (if present) will appear clear against the dark background.

11. API test:
API 20 E is used
E. coli
Indole test is +ve
Gelatinase -ve
H2S production -ve
Citrate -ve
Urease test -ve
Klebsiella
Indol –ve
Gelatine -ve
H2S –ve
Citrate +ve
Urease +ve

12. Serologic test
• For serotyping of E. coli (Epidemiologic information)
• Depend on detection of O and H antigen.

13. Gene Detection by PCR
detecting ID or virulence gene that is specific for E. coli and
Klebsiella