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3M Petrifilm SALX Collaborative Study
July 2013
OMA-2013-July-XXX
DRAFT DOCUMENT
18
3.5.2
Negative – no typical colonies present
1
2
3.6
Alternative 3M SALX Confirmation Method
3
3.6.1
Using a fine tip marker, circle presumptive positive colonies (red to brown
4
colonies with discrete yellow zones and/or gas bubbles) on the plate top film.
5
3.6.2
Lift the top film and place a 3M Petrifilm
Salmonella
Express Confirmation Disk
6
onto the gel. Close the film using a gloved hand (while practicing good
7
laboratory technique) and remove air bubbles by gently applying a sweeping
8
motion with even pressure onto the top of the film with the analyst’s fingers.
9
3.6.3
Incubate the plates with confirmation disks at 41.5° ± 1°C for 4 to 5 hours.
10
3.6.4
Observe circled colonies for color change - red/brown to green blue, blue, dark
11
blue or black confirm the colony as
Salmonella
species. No color change, colony
12
is negative.
13
3.6.5
Transfer typical colonies from SALX plate to TSI/LIA slants. Incubate 35
±
1
°
C for
14
24
±
2h.
15
3.6.6
Confirm a minimum of one typical colony per test portion with
16
biochemical/serological procedures prescribed by the USDA/FSIS method. Either
17
the API20E (Official Method 978.24) or the VITEK GN (Official Method 2011.17)
18
will be used as an alternative to the conventional biochemical tests. The
19
somatic (O) and flagellar (H) tests will also be performed.
20
21
3.7
Reference Method Confirmation
22
3.7.1
Transfer 0.5
±
0.05 mL 3M SALX enriched sample into 10 mL tetrathionate (TT-
23
Hajna) broth and 0.1
±
0.02 mL into 10 mL modified Rappaport-Vassiliadis (mRV)
24
broth. Incubate at 42
±
0.5
°
C for 22-24 h.
25
3.7.2
Streak both secondary enrichments onto double modified lysine iron agar
26
(DMLIA) or xylose lysine Tergitol (XLT4) agar and brilliant green sulfa (BGS) agar.
27
Use one loopful of inoculum for each plate. Incubate 35
±
2
°
C for 18-24h. Select
28
colonies. Re-incubate all plates for an additional 18-24 h. Reexamine initially
29
negative plates and pick colonies as above.
30
3.7.3
Transfer typical colonies to TSI/LIA slants. Incubate 35
±
1
°
C for 24
±
2h.
31
3.7.4
Confirm a minimum of one typical colony per sample with
32
biochemical/serological procedures prescribed by the USDA/FSIS method. Either
33
the API20E (Official Method 978.24) or the VITEK GN (Official Method 2011.17)
34
will be used as an alternative to the conventional biochemical tests. The
35
somatic (O) and flagellar (H) tests will also be performed.
36
37
4.0
FDA-BAM Chapter 5 – Dry dog food
38
1.1.1.
Add twenty 25g contaminated test portions and five 25g uncontaminated test portions
39
to 225ml lactose broth. Blend 2 min. Let stand 60±5 min at room temperature. Mix
40
and adjust pH to 6.8±0.2 if necessary. Incubate 24 ± 2h at 35°C.
41
1.1.2.
Transfer 0.1 mL to 10 mL RV broth (prepared from individual ingredients) and 1 mL to 10
42
mL TT broth. Incubate the RV at 42
±
0.2°C for 24
±
2 h. Incubate the TT broth at 35 ±
43
2.0
°
C for 24
±
2 h in a forced air incubator.
44
1.1.3.
Mix and streak 3 mm loopful (10
µ
L) RV broth onto BS, XLD and HE agars. Repeat from
45
TT broth. Incubate at 35
°
C for 24
±
2 h. Follow isolation procedure according to FDA-
46
BAM.
47
OMAMAN-08/Collaborative Study Protocol
March 2014
Expert Review Panel Use Only
AOAC Research Institute
Expert Review Panel Use Only