KS J ISO 16654-2007 Microbiology of food and animal feeding stuffs－Horizontal method for the detection of Escherichia coli O157《食品和动物饲料的微生物学 检测大肠杆菌0157的水平方法》.pdf

1、 KSKSKSKS KSKSKSK KSKSKS KSKSK KSKS KSK KS KS J ISO 16654 O157 KS J ISO 16654 ：2007 (2012 ) 2007 10 31 http:/www.kats.go.krKS J ISO 16654：2007 : ( ) ( ) () () () ( ) : (http:/www.standard.go.kr) ： ：2002 11 30 ：2007 10 31 ：2012 12 28 ： 20120814 ： ( 02-509-7270) (http:/www.kats.go.kr). 10 5 , . KS J I

2、SO 16654：2007 (2012 ) O157 Microbiology of food and animal feeding stuffsHorizontal method for the detection of Escherichia coli O157 2001 ISO 16654, Microbiology of food and animal feeding stuffs Horizontal method for the detection of Escherichia coli O157 , . ISO 16654 ISO/TC34 (SC9) . 1 O157 . ,

3、. 2 . . ( ) . KS H ISO 68871, , 10 1： 10 KS H ISO 7218, 3 . 3.1 O157 (Escherichia coli O157) , (indole) O157 1 (sorbitol) O157 CTSMAC (5.2 ) . 2 . KS J ISO 16654：2007 4 O157 ( A ) . a) 6 1218 (41.51) mTSBN(modified tryptone soya broth with noboviocin) b) O157 (immunomagnetic particles) c) CTSMAC (ce

4、fixime tellurite sorbitol MacConkey agar) 2 , (immunomagnetic particles) 2 d) O157 CTSMAC 2 O157 . 5 , KS H ISO 7218 . 5.1 ： mTSB(modified tryptone soya broth with noboviocin：mTBSN) 1 5.1.1 mTBS (modified tryptone soya broth) 5.1.1.1 (enzymatic digest of casein) 17.0 g (enzymatic digest of soya) 3.0

5、 g D()D()glucose 2.5 g 3(bile salts No. 3) 1.5 g (sodium chloride) 5.0 g (K 2 HPO 4 ) 4.0 g 1 000 mL 5.1.1.2 . , pH (6.6 ) pH 25 7.40.2 pH . (6.7 ) . 121 15 (6.1 ) . 5.1.2 (novobiocin solution) 2 KS J ISO 16654：2007 5.1.2.1 (novobiocin) 0.45 g 100 mL 5.1.2.2 . . 5.1.2.3 , mTBS (5.1.1 ) 225 mL 900 mL

6、 (5.1.1 ) 1 mL 4 mL . mTBS 1 L 20 mg. 5.2 1 ：CTSMAC (cefixime tellurite sorbitol MacConkey agar) 2 5.2.1 5.2.1.1 (enzymatic digest of casein) 17.0 g (enzymatic digest of animal tissues) 3.0 g (sorbitol) 10.0 g 3(bile salts No. 3) 1.5 g (sodium chloride) 5.0 g (neutral red) 0.03 g (crystal violet) 0.

7、001 g 9 18 g a(agar) 1 000 mL a . 5.2.1.2 , . pH (6.6 ) pH 25 7.10.2 pH . 121 15 (6.1 ) . 5.2.2 (potassium tellurite solution) 5.2.2.1 3 KS J ISO 16654：2007 (potassium tellurite for bacteriological use) 0.25 g 100.0 mL 5.2.2.2 . 1 . 5.2.3 (cefixime solution) 5.2.3.1 (cefixime) 5.0 mg 100.0 mL 5.2.3.

8、2 . . 1 (32) . 5.2.4 5.2.4.1 (5.2.1 ) 1 000 mL (5.2.2 )(potassium tellurite solution) 1.0 mL (5.2.3 )(cefixime solution) 1.0 mL 5.2.4.2 44 47 (6.5 ) (5.2.1 ) 4447 . 1 000 mL 1 mL 1 mL . (6.15 ) 15 mL . 2.5 mg/L 0.05 mg/L. 2550 (6.2 ) . . 30 . , 2 (32) . 4 KS J ISO 16654：2007 5.3 2 CTSMAC , O157 . 25

9、50 (6.2 ) . . 30 . , 2 (32) . 5.4 5.4.1 (meat extract) 3.0 g (peptone) 5.0 g 9 18 g a(agar) 1 000 mL a . 5.4.2 , . , pH 25 7.00.2 pH . (6.7 ) . 121 15 (6.1 ) . 5.4.3 4447 (6.5) 15 mL (5.4.2). 2550 (6.2 ) . . 30 . , 2 (32) . 5.5 / 5.5.1 5 KS J ISO 16654：2007 (tryptone) 10.0 g (sodium chloride) 5.0 g

10、DL(DLtryptophan) 1.0 g 1 000 mL 5.5.2 , . , pH 25 7.50.2 pH(6.6 ) . (6.7 ) 5 mL . 121 15 (6.1 ) . 5.6 (Kovacs indole reagent) 5.6.1 4(4-dimethylaminobenzaldehyde) 5.0 g 2-1-(2-methylbutan-1-ol) -1-(pentan-1-ol) 75.0 mL ( 201.181.19 g/mL) 25.0 mL 5.6.2 , 4447 (6.5 ) 4 . . (32) . . 5.7 O157 (anti-Esch

11、erichia coli O157 immunomagnetic particles) O157 . . . 5.8 ：0.01 mol/L, pH 7.2 (wash buffer：modified phosphate buffer, 0.01 mol/L, of pH 7.2) 5.8.1 (sodium chloride) 8.0 g (potassium chloride) 0.2 g ()disodium hydrogen phosphate(anhydrous) 1.44 g 6 KS J ISO 16654：2007 ()potassium dihydrogen phosphat

12、e(angydrous) 0.24 g (polyoxyethylene sorbitanmonolaurate) 0.2 mL (Tween 20 ) 1 000 mL 5.8.2 . , pH 25 7.20.2 pH(6.6 ) . (6.7 ) . 121 (6.1 ) 15 . . . 5.9 (normal saline solution) 5.9.1 (sodium chloride) 8.5 g 1 000 mL 5.9.2 . (6.7 ) . 121 15 (6.1 ) . 5.10 O157 (Escherichia coli O157 antiserum) O157 .

13、 . 6 (KS H ISO 7218 ) . 6.1 () / ( ) KS H ISO 7218 . 6.2 2550 . 7 KS J ISO 16654：2007 6.3 (371) . 6.4 (41.51) . 6.5 4447 . 6.6 pH- 25 0.01 pH 0.1 pH . 6.7 , . 6.8 . 6.9 (total-delivery graduated pipettes) 0.1 mL 0.5 mL 1 mL 10 mL . 6.10 / / 6.11 (mechanical air-displaced pipettors) 10 L 20200 L 6.12

14、 (magnetic rack) (magnetic separator) (6.13 ) . 6.13 . 1.5 mL . . 6.14 (, ) 1520 r/min . 8 KS J ISO 16654：2007 6.15 90 mm 140 mm . 6.16 (vortex mixer) 7 . . . , . 8 . . 9 ( A ) 9.1 KS H ISO 68871 . ISO 6887 . 1/10( ) mTSBN 41.5 (6.4 ) (mTBSN)(5.1)9x mL 9 x g x g x mL . (9.3 ) . 9.2 9.1 41.5 6 1218(,

15、 1824 ) (6.4 ). (IMS) 6 18 . 9.3 (IMS) 9.3.1 6 , 1218 . 9 KS J ISO 16654：2007 . . 9.3.2 . . . (6.12 ) (5.7 ) / . (9.2 ) . (6.13 ) (5.7 ) 20 L . 1 mL . 10 1220 r/min (6.14 ) . 9.3.3 (6.12 ) (9.3.2 ) 180 . . (6.10 ) . (5.8 ) 1 mL . . 180 . . . . (5.8 ) 100 L . . 9.4 O157 9.4.1 (6.11 ) CTSMAC (5.2) (9.

16、3.3 ) 50 L (5.3 ) 50 L . (6.10 ) . 10 KS J ISO 16654：2007 37 1824 CTSMAC (5.2 ) (6.3 ) . 2024 O157 . 50 L O157 , . 9.4.2 O157 CTSMAC 1 mm. O157 . 9.5 , O157 . 9.5.1 9.4 5 . 5 , . (5.4 ) . 37 1824 (6.3 ). 9.5.2 9.5.3 . 9.5.2 ： / (5.5 ) (9.5.1 ) . 24 37 (6.3 ). Kovac (5.6 ) 1 mL 10 . . / . 9.5.3 9.5.3

17、.1 O157 . 11 KS J ISO 16654：2007 9.5.3.2 (5.9) . (6.10 ) (9.5.1 ) . 3060 . . , . 9.5.3.3 O157 (9.5.1) 9.5.3.2 O157 (5.10 ) . 1 , . 9.5.3.4 O157 , O157H7 . 9.6 (reference laboratory) . 10 10.1 O157 . . 10.2 O157 O157 . 11 V/mL M/g O157 . 12 12 KS J ISO 16654：2007 . a) b) ( ) c) d) e) f) . 13 KS J ISO

18、 16654：2007 A () (x g x mL) 41.5 (9.1 ) mTSBN(5.1 ) 9x mL 41.5 (9.2 ) 6 1218 O157 (9.3 ) (9.3 ) 0.1 mL (9.4 ) 50 L CTSMAC (5.2 ) (5.3 ) 1824 37 5 5 O157(9.5.3 ) (9.5 ) (9.5.2 ) (9.5.1 ) () (9.6 ) 14 KS J ISO 16654：2007 15 1 DOYLE M.P. and SCHOENI J.L. Appl. Environ. Microbiol., 53, 1987, pp. 2394239

19、6 2 ZADIK P.M., CHAPMAN P.A. and SIDDONS C.A. J. Med. Microbiol., 39, 1993, pp. 155158 3 KS H ISO 68872, , 10 2： 4 KS H ISO 68873, , 10 3： 5 KS H ISO 68874, , 10 4： , , 6 ISO 68875, Microbiology of food and animal feeding stuffsPreparation of test samples, initial suspension and decimal dilutions for microbiological examinationPart 5：Specific rules for the preparation of the test