3M Interpretation EC (Panduan Pembacaan Koloni E Coli Dan Coliform) PDF

3
Petrifilm
Interpretation Guide
E.coli/Coliform Count Plate

This guide familiarizes you with results on 3M Petrifilm E. coli/Coliform Count
plates. For more information, contact the official 3M Microbiology Products
representative nearest you.
Petrifilm E. coli/Coliform Count (EC) plates contain Violet Red Bile (VRB) nutrients, a cold-water-soluble gelling agent, an
indicator of glucuronidase activity, and an indicator that facilitates colony enumeration. Most E. coli (about 97%) produce
beta-glucuronidase which produces a blue precipitate associated with the colony. The top film traps gas produced by the
lactose fermenting coliforms and E. coli. About 95% of E. coli produce gas, indicated by blue to red-blue colonies associated
with entrapped gas on the Petrifilm EC plate (within approximately one colony diameter).
AOAC INTERNATIONAL and U.S. FDA Bacteriological Analytical Manual (BAM) define coliforms as gram-negative rods which
produce acid and gas from lactose during metabolic fermentation. Coliform colonies growing on the Petrifilm EC plate
produce acid which causes the pH indicator to make the gel color darker red. Gas trapped around red coliform colonies
indicates confirmed coliforms.
The identification of E. coli may vary by country

(see Reminders for Use section for incubation times
and temperatures):
AOAC INTERNATIONAL validated method
E. coli = 49 (blue colonies with gas)
Total coliform = 87 (red and blue colonies with gas)
Do not use this plate alone for the detection of E. coli O157. Like most other E. coli/coliform media, this plate will not
specifically indicate whether any O157 strain is present.
3M Petrifilm
E. coli/Coliform Count Plate
1
1
No growth = 0
E. coli count = 13
Notice the changes in gel color in figures 2 through 8.

As the E. coli or coliform count increases, the color
of the gel turns to dark red or purple-blue.
Total coliform count = 28
Background bubbles are a characteristic of the gel

and are not a result of E. coli or coliform growth.
See square 1.
Back
Lighting
The counting range for the total population on Petrifilm

EC plates is 15150.
Do not count colonies that appear on the foam barrier
because they are removed from the selective influence
of the medium. See circle 1.
Front
Lighting
E. coli count = 3
E. coli count = 17
Any blue in a colony (blue to red-blue) indicates the

presence of E. coli. Front lighting will enhance the
detection of blue precipitate formed by a colony.
Estimated total coliform count = 150
Circle 1 shows a red-blue colony counted using back

lighting. Circle 2 shows the same colony with front
lighting. The blue precipitate is more evident in circle 2.
The circular growth area is approximately 20 cm2.

Estimates can be made on plates containing greater than
150 colonies by counting the number of colonies in one
or more representative squares and determining the average
number per square. Multiply the average number by 20 to
determine the estimated count per plate.
TNTC (Too Numerous to Count)
To obtain a more accurate count, dilute the sample further.
Actual count ~ 106
Actual count ~ 108
Petrifilm EC plates with colonies that are TNTC

have one or more of the following characteristics:
many small colonies, many gas bubbles, and a
deepening of the gel color from red to purple-blue.
High concentrations of E. coli may cause the growth

area to turn purple-blue.
Presumptive E. coli count ~ 8
Actual count ~ 108
Estimated total coliform count ~ 108
When high numbers of non-coliform organisms such as

Pseudomonas are present on Petrifilm EC plates, the gel
may turn yellow.
When high levels of coliforms are present (>10 8),

some strains of E. coli may produce less gas and
blue colonies may be less definitive. Count all blue
colonies without gas and/or blue zones as presumptive
E. coli. Pick blue colonies without gas and confirm
if necessary.
Bubbles
10
11
Food particles are irregularly shaped and are not

associated with gas bubbles.
Bubble patterns may vary. Gas may disrupt the

colony so that the colony outlines the bubble.
See circles 1 and 2.
Artifact bubbles may result from improper
inoculation or from trapped air within the sample.
They are irregularly shaped and are not associated
with a colony. See circle 3.
3
1
4
2
10
12
Examples 110 show various bubble patterns
associated with gas producing colonies. All should
be enumerated.
3Petrifilm E. coli / Coliform Count Plates
Reminders for Use
For detailed WARNING, CAUTIONS, DISCLAIMER OF WARRANTIES / LIMITED REMEDY, LIMITATION OF 3M LIABILITY,
STORAGE AND DISPOSAL information, and INSTRUCTIONS FOR USE see Products package insert.
Storage
Petrifilm
Petrifilm
Petrifilm
Store unopened packages at 8C

(46F). Use before expiration date on
package. In areas of high humidity
where condensate may be an issue,
it is best to allow packages to reach
room temperature before opening.
To seal opened package, fold end

over and tape shut.
Keep resealed package at 25C

(77F) and <50%RH. Do not
refrigerate opened packages.
Use Petrifilm plates within one
month after opening.
Add appropriate quantity of one of the

following sterile diluents: Butterfield's
phosphate buffer (IDF phosphate
buffer, 0.0425 g/L of KH2PO4
adjusted to pH 7.2), 0.1% peptone
water, peptone salt diluent (ISO
method 6887), buffered peptone
water (ISO method 6887-1), saline
solution (0.85-0.90%), bisulfite-free
letheen broth, or distilled water.
Blend or homogenize sample per

current procedure.
Sample Preparation
Prepare a 1:10 or greater dilution of

food product. Weigh or pipette food
product into an appropriate container
such as a stomacher bag, dilution
bottle, Whirl-Pak bag, or other
sterile container.
Adjust pH of the diluted sample

between 6.5 and 7.5.
for acid products, use 1N NaOH,
for alkaline products, use 1N HCI.
Do not use buffers containing

citrate, bisulfite, or thiosulfate;
they can inhibit growth.
Inoculation
Place Petrifilm plate on level

surface. Lift top film.
With pipette perpendicular to

Petrifilm plate, place 1 mL of sample
onto center of bottom film.
Carefully ROLL top film down to

avoid entrapping air bubbles.
Do NOT let top film drop.
RIDGE side
FLAT side
With FLAT side down, place

spreader on top film over inoculum.
11
GENTLY apply pressure on

spreader to distribute inoculum over
circular area before gel is formed.
Do not twist or slide the spreader.
12
Lift spreader. Wait a minimum of

one minute for gel to solidify.
15
Colonies may be isolated for further

identification. Lift top film and pick
the colony from the gel.
Interpretation
Incubation
<70F
13
Incubate plates with clear side up in

stacks of no more than 20. It may be
necessary to humidify incubator to
minimize moisture loss.
14
Petrifilm plates can be counted on a

standard colony counter or other
illuminated magnifier. Refer to the
Interpretation Guide section when
reading results.
Incubation time and temperature varies by

method. Most common approved methods:
AOAC Official Method 991.14
for coliforms: incubate 24h 2h at 35C 1C
for E.coli: incubate 48h 2h at 35C 1C
AOAC Official Method 998.08
for E.coli (for meat, poultry and seafood):
incubate 24h 2h at 35C 1C
NMKL method (147.1993)
for coliforms: incubate 24h 2h at 37C
for E.coli: incubate 48h 2h at 37C
Additional Comments
Questions? U.S., Call 1-800-328-6553, Canada, call 1-800-265-1840 x6574

for technical service.
To order Petrifilm plates in the U.S., call 1-800-328-1671
Latin America / Africa and Asia Pacific regions, call 1-651-733-7562.
Microbiology Products
3M Canada
3M Europe
3M Center Bldg. 275-5W-05

St. Paul, MN 55144-1000
USA
1-800-228-3957
www.3M.com/microbiology
Email:[email protected]
Post Office Box 5757

London, Ontario N6A4T1
Canada
1-800-563-2921
Laboratoires 3M Sant
Boulevard de lOise
95029 Cergy Pontoise Cedex
France
33 1 30 31 85 71
Recycled Paper
40% pre-consumer
10% post-consumer
Petrifilm is a trademark of 3M.

Whirl-Pak is a registered trademark
of NASCO.
Printed in U.S.A.
3M 2001
70-2008-4574-4 (1211) DPI

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How does the Petrifilm plate work in detecting E. coli and coliforms?

How does the Petrifilm plate work in detecting E. coli and coliforms?

How should samples be prepared and inoculated for use with the Petrifilm plate?

How should samples be prepared and inoculated for use with the Petrifilm plate?

3

E.coli/Coliform Count Plate

The identification of E. coli may vary by country

E. coli/Coliform Count Plate

Notice the changes in gel color in figures 2 through 8.

Total coliform count = 28

Background bubbles are a characteristic of the gel

The counting range for the total population on Petrifilm

Any blue in a colony (blue to red-blue) indicates the

Estimated total coliform count = 150

Circle 1 shows a red-blue colony counted using back

The circular growth area is approximately 20 cm2.

TNTC (Too Numerous to Count)

To obtain a more accurate count, dilute the sample further.

Actual count ~ 106

Actual count ~ 108

Petrifilm EC plates with colonies that are TNTC

High concentrations of E. coli may cause the growth

Presumptive E. coli count ~ 8

Actual count ~ 108

Estimated total coliform count ~ 108

When high numbers of non-coliform organisms such as

When high levels of coliforms are present (>10 8),

Total coliform count = 3

Total coliform count = 78

Food particles are irregularly shaped and are not

Bubble patterns may vary. Gas may disrupt the

3Petrifilm E. coli / Coliform Count Plates

Reminders for Use

Store unopened packages at 8C

To seal opened package, fold end

Keep resealed package at 25C

Add appropriate quantity of one of the

Blend or homogenize sample per

Prepare a 1:10 or greater dilution of

Adjust pH of the diluted sample

Do not use buffers containing

Place Petrifilm plate on level

With pipette perpendicular to

Carefully ROLL top film down to

With FLAT side down, place

GENTLY apply pressure on

Lift spreader. Wait a minimum of

Colonies may be isolated for further

Incubate plates with clear side up in

Petrifilm plates can be counted on a

Incubation time and temperature varies by

Questions? U.S., Call 1-800-328-6553, Canada, call 1-800-265-1840 x6574

To order Petrifilm plates in the U.S., call 1-800-328-1671

Latin America / Africa and Asia Pacific regions, call 1-651-733-7562.

3M Center Bldg. 275-5W-05

Post Office Box 5757

Petrifilm is a trademark of 3M.

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