AOAC Performance Tested Methods

•X-Gal)

• 9 26 VRBFDA-BAM AOACI Method

Committee Guidelines for Validation of Qualitative and Quantitative Food Microbiological OMA

y=1.04x-0.13 (y xFDABAM) (r2)=0.98

85y=0.95x+0.15 r2=0.94

• FDA-BAM

• PTM

The Sanita-kun Coliforms device is a cultural medium for the enumeration of coliform bacteria. The device consists of a non-woven fabric to which a layer of microbial nutrients is deposited in a film. The nutrient film also contains 5-bromo-4-chloro-3-indolyl- -D-galactopyranoside, or X-gal, which is hydrolyzed by -galactosidasefrom coliforms to produce a visible blue dye. The non-woven fabric and nutrient film are constructed on a base film. The basic device (fabric, nutrient and base film) is then inserted between an adhesive backing to stabilize the basic device, and a transparent film. The adhesive backing and the transparent top film serve to deter evaporation, and facilitate inspection of the inoculated device.

Sample solutions containing bacteria are deposited on the fabricportion of the device. Then, the sample solution diffuses throughout the entire pad to dissolve and release the nutrient compound film forming a highly viscous solution. Bacteria migrate to the surface of the fabric where coliforms metabolize the nutrients and X-gal to produce a visible blue dye after incubation, while growth of non-coliforms is inhibited with selective reagents.

Principle

X-Gal

+

1mL

X-Gal

Sanita-kun Coliforms 35 24hours

1mL

1.2. 1mL3.4. 35 245.

20/

Inclusivity ExclusivityBGLB BGLB

Cedecea lapagei 1 + + Bacillus cereus 2 - -Citrobacter 1 + + Bacillus megaterium 1 - -Citrobacter diversus 1 + + Bacillus subtilis 3 - -Citrobacter freundii 2 + + Corynebacterium pyrogenes 1 - -Citrobacter sp. from afood

3 + + Corynebacterium xerosis 1 - -

Enterobacter 2 + + Enterobacter gergoviae 1 - -Enterobacter cloacae 1 + + Enterococcus hirae 1 - -Enterobacterintermedium

1 + + Lactic acid bacteria isolated fromfoods

8 - -

Enterobacter sp. from afood

4 + + Lactobacillus acidophilus 1 - -

Escherichia coli 3 + + Lactobacillus delbruedkii 1 - -Escherichia sp. from afood

3 + + Leuconostoc mesenteroides 1 - -

Klebsiella oxytoca 1 + + Proteus mirabilis 1 - -Klebsiella planticola 1 + + Proteus vulgaris 2 - -Klebsiella pneumoniae 2 + + Providencia alcalifaciens 1 - -Klebsiella terrigena 2 + + Pseudomonas aeruginosa 1 - -Klebsiella sp. from foods 6 + + Rahnella aquatilis 1 - -

: Salmonella Enteritidis 1 - -Salmonella Typhymurium 3 - -Serratia marcescens 2 - +Serratia rubidaea 1 - +Staphylococcus aureus 1 - -Staphylococcus sp. 5 - -Streptococcus sp. 1 - -Yersinia aldovae 1 - -

a)Significantly higher than FDA-BAM (p<0.05)b)Significantly higher than FDA-BAM (p<0.01)

1.00

FDAFDA--BAMBAM

FDAFDA--BAMBAM

FDA-BAM(cfu/mL) RSD (cfu/mL) RSD

FDA-BAM FDA-BAM FDA-BAM FDA-BAMKlebsiella sp.

strain No291 92 0.10 0.08 Enterobacter

aerogenesIFO 13534

149 147 0.09 0.1349 50 0.16 0.14 73 73 0.07 0.112 8 0.25 0.38 17 14 0.29 0.43

Klebsiellaoxytoca JCM

1665

21 27 0.28 0.19 KlebsiellapneumoniaeJCM 1662

194 199 0.03 0.0611 14 0.32 0.21 100 103 0.05 0.11

1 1 1 1 20 21 0.11 0.35Klebsiella sp.strain No53

80 58 0.1 0.07 Klebsiella sp.strain No171

108 118 0.1 0.0943 25 0.07 0.16 59 64 0.17 0.09

7 5 0.3 0.2 13 12 0.31 0.25Klebsiella sp.strain No172

89 74 0.09 0.15 Klebsiella sp.strain No171

53 53 0.15 0.2147 43 0.7 0.16 32 31 0.16 0.26

8 9 0.3 0.3 6 6 0.5 0.2Klebsiella

pneumoniae IID5209

97 105 0.1 0.03 Klebsiella sp.strain No171

98 100 0.04 0.0748 50 0.19 0.14 55 50 0.11 0.0810 7 0.2 0.6 11 12 0.45 0.33

Enterobactersp. strain No8

97 93 0.05 0.06 Escherichiacoli IID 5208

115 121 0.06 0.1956 47 0.12 0.13 67 54 0.06 0.1311 7 0.45 0.3 13 10 0.23 0.1

Enterobacteraerogenes IFO

13534

94 94 0.14 0.15 EnterobacteraerogenesIFO 13534

114 108 0.1 0.1151 47 0.22 0.13 54 47 0.17 0.11

7 11 0.3 0.4 11 10 0.09 0.3Klebsiella sp.

strain No262 52 0.1 0.13 Enterobacter

sp. strain No865 50 0.11 0.08

37 37 0.22 0.16 33 28 0.12 0.2411 7 0.2 0.4 11 6 0.36 0.5

Klebsiella sp.strain No171

67 68 0.13 0.1 Klebsiellaoxytoca JCM

1665

73 70 0.12 0.141 35 0.27 0.23 41 34 0.15 0.09

5 4 0.6 0.5 9 9 0.3 0.2Klebsiella sp.strain No52

34 8 0.12 0.3 Blue letter:Significantly higher than FDA-BAM(P<0.05) 18 1 0.17 1 Red letter:Significantly higher than FDA-BAM(P<0.01)

4 0 0.8 1Citrobacter sp.

strain No20260 62 0.17 0.128 28 0.3 0.21

5 7 0.4 0.6Escherichia sp.

strain No4164 59 0.09 0.136 32 0.08 0.13

7 7 0.4 0.3Klebsiella sp.strain No172

67 58 0.06 0.1236 25 0.11 0.2

7 5 0.1 0.4Klebsiella

pneumoniae IID5209

84 83 0.15 0.0550 43 0.12 0.12

9 7 0.3 0.3Klebsiella

oxytoca JCM1665

120 112 0.07 0.0959 49 0.11 0.1611 9 0.17 0.34

Klebsiella sp.strain No52

53 38 0.13 0.1632 21 0.28 0.29

5 5 0.4 0.4Klebsiella

pneumoniae IID5209

108 94 0.1 0.255 48 0.04 0.19

9 9 0.3 0.4

3