Selection of Culture Media
Urine Cultures Culture Setup
CLS 418/CLS 419
Clinical Laboratory Science Program
Carol Larson MSEd, MT(ASCP)
Common Etiologic Agents of UTI
Pathogens
• Escherichia coli
• Proteus, Klebsiella,
Enterobacter,
Citrobacter, and
Serratia spp.
• Acinetobacter species
• Pseudomonas
aeruginosa and nonfermenting GNR’s
Media to use
• MacConkey (MAC),
or
• EMB agar
• Urine is cultured for detection of the
common UTI pathogens
• Thus in selecting media it is important that
you use media appropriate for isolating the
common etiologic agents
Common Etiologic Agents of UTI
Pathogens
• Enterococcus species
• Streptococcus agalactiae
• Streptococcus pyogenes
• Staphylococcus
saprophyticus
• Staphylococcus aureus
• Yeasts
• Neisseria gonorrhoeae
Media to use
• 5% Sheep blood agar
(BAP)
• [Some labs also use
CNA agar]
• Chocolate agar
– Special request
Selection of Culture Media
• Most laboratories will routinely setup BAP
and MAC
– Please refer to procedure manual for specifics
on what media your clinical site sets up
• Keep in mind contaminants / commensal
flora of the urethra will also grow on these
media
Why is a blood agar plate and
a MacConkey agar plate used
to setup a urine culture?
The common potential pathogens
(enterics, Staphylococcus, and other
GNR’s) will grow on them. MAC can
assist in selecting GNR’s from other
organisms and differentiating between
lactose fermenters and nonfermenters.
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Inoculation of Media
• A colony count is generally performed on
both the BAP and MAC
– Aids in interpretation
– Used to quantitate growth of each colony type
– Setup on MAC to quantitate gram-negative
rods if culture has mixed growth
Colony Count Method
1. Mix specimen well!
2. Place appropriate calibrated loop
vertically into the well mixed urine just
below the surface of a well-mixed urine.
– Do not go to bottom of container, as
additional urine will adhere to the loop stem
Colony Count Method
4. Without flaming or
reentering the
urine, streak
numerous times
perpendicular to
the primary line of
inoculation, being
careful not streak
over the same area
Colony Count Method
• Use a calibrated loop
– 0.001ml for cleancatch midstream urine
and catheterized
specimens
– 0.01 ml for suprapubic
or cystoscopy
specimens
Colony Count Method
3. Quickly deliver the
known volume of urine
to the culture plate by
touching the center of
the plate
and spreading in a
line across the
diameter of the
plate
Colony Count Method
5. Repeat steps 2-4 for other culture
plate(s) as specified in procedure manual
2
Incubation of Media
Why is a colony count
performed on both the blood
agar and MacConkey agar
plates?
The colony count aids in the interpretation
of the culture. The cc on the BAP provides
comparison of colony types (quantitate
each type) and the MAC provides count for
GNR’s.
Review
• Select culture media
– BAP and MAC
– Common etiologic agents of UTI
• BAP
– Incubate at 35ºC in either a
CO2 incubator or ambient
air (refer to procedure)
• MAC
– Incubate at 35ºC in
ambient air
• First read at 18-24 hours
• 2nd read at 48 hours
References:
• Mahon, C.R., Lehman, D.C. & Manuselis,
G., Textbook of Diagnostic Microbiology,
3rd Ed., Saunders Elsevier, 2007.
• Inoculation of media
– Colony count
• Purpose
• Method
• Incubation of media
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