E. coli, fecal coli and E. coli testing methods in food

1 topic content and scope of applicationIn this paper, the test methods of E. coli, fecal E. coli and E. coli in food are specified.this article is applicable to the inspection of aviation food.2 equipment and materialsstraw (1ml, 10ml),
reverse
culture
case (36±1c, 44.5±0.5C), refrigerator, Homogenizers, oscillators, flat dishes, diluted bottles,
balances
,
microscopes
etc. 3
base
and
reagents laurel sulphuric acid Salt tryptoprotein monthly display (LST) broth, yellow lactose bile salt (BGLB) broth, EC broth, yihong mei blue
agar
(EMB), nutritional agar sloping noodles, tryptophan broth, MR-PV culture, Kor Ser'ssbroth, crystalline azithromid agar (VRBA), Butterfield's phosphate buffer dilution, physiological saline, Terran's dyeing solution, Kovacs' indigo substring reagents, methyl red indicators, Voges-proskauer (V-P) reagents, 4 sample preparation 4.1 Representative samples taken in sterile operations are held in sterilized containers. If packaged, wipe the packaging opening with 75% ethanol and sample it.4.2 Preparation sample homogenizertake 25g sample with sterile operation, put it in a sterilized homogenizer cup filled with 225 ml of diluent, and make a sample homogenization of 1-2min at 8000r/min homogeneity. If the sample homogeneity time exceeds 2min, the homogeneity cup should be cooled with ice water.4.3 Diluted sample homogeneity10 ml sample homogenization with a 10 ml sterilization straw to accurately absorb 10 ml of sample homogenizer and place it in a 150 ml dilution bottle containing 90 ml of diluent. Shake quickly and mix the sample to make a sample homogeneity of 1:100.5 Determination of E. coli group 5.1 Determination of E. coli group MPN value 5.1.1 For each sample, select the appropriate three consecutive dilution of the sample dilution. Inoculate three tubes of laurel sulphate tryplin (LST) broth per dilution, 1 ml per tube.5.1.2 Place the inoculation tube at a culture of 48±2h at 36±1 degrees C.5.1.3 Observe the gas production of the test tube: check the inverter for bubble production and record the number of LST broth tubes producing gas in 24h and 48h. If all LST broth tubes are not gas-producing, they can be reported as E. coli-negative, and if gas-producing persons are available, further confirmed tests may be carried out.5.2 Confirmed trial of E. coli 5.2.1 transferred all trachea-producing tubes with inoculation rings with a diameter of 3 mm to the broth tube of BGLB.5.2.2 BGLB broth tube at 36±1 degrees C culture 48±2h.5.2.3 records the number of tracheal tubes produced by all BGLB broth tubes.5.2.4 Results Report: Check the MPN table to report the MPN value of E. coli in each gram (ml) sample by the number of trachea produced by BGLB broth.6 fecal colium group determination 6.1 with the inoculation ring all 48±2h gas-producing LST broth tube culture transferred to EC broth tube.6.2 Place all inoculated EC broth tubes in a 44.5±0.5C water bath within 30min and culture 24±2h. The horizontal surface of the water bath tank should be higher than the broth culture base liquid surface. Positive and negative controls should be given to E. coli, which is known to be positive for 44.5 degrees C, and E. coli or other E. coli bacteria that are not gas-producing at 44.5 degrees C.6.3 records the production of EC broth tubes. The production trachea tested positive for fecal colium and the non-production trachea tested negative for fecal colium.6.4 Results Report: Check the MPN table to report the MPN value of fecal coli in each gram (ml) of samples by number of trachea produced.7 E. coli assay 7.1 The EC broth tube in 6.3 continued to culture 24h cultures from its trachea inoculated on the EMB plate, and cultured 24±2h at 36±1 degrees C±7.2 check the tablet for typical bacterios with a shiny or non-glossy black center.7.3 If there are typical bacterios, select at least 2 typical bacterios from each tablet and, if there are no typical bacterios, select at least 2 suspected places from each plate. Contact the center of the bacteria with an inoculation needle and transfer to the nutritional agar slope, where the culture of 36±1 degrees C is 18-24h.7.4 transfer of beveled cultures to the following cultures for
bio
tests.7.4.1 Tryptophan Broth: After a culture of 24±2h at 36±1 degrees C, plus Kovacs reagent 0.2-0.3 ml, the upper layer appears red as a substitin positive.7.4.2 MR-VP culture: 48±2h after a culture of 36±1 degrees C. In a test tube with sterile operation to remove the culture from 1 ml to 13 mm x 100mm, add 5% phenol ethanol solution 0.6 ml, 40% potassium hydroxide solution 0.2 ml and a little crea acid crystallization, shake the test tube after static 2h, if the appearance of Ir red, positive for VP test.the rest of the MR-VP culture was then cultured with 48h drops plus 5 drops of methyl red solution. If the culture becomes red, it tests positive for methyl red and negative if it changes yellow.7.4.3 Korser' broth: cultured 96h at 36±1 degrees C recorded no growth.7.4.4 LST broth: cultured at 36±1 degrees C 48±2h to see if gas is produced in the test tube.7.4.5 Terraneum staining: take 18h nutritional agar slope culture for Terranean staining. E. coli is Gloran's negative.7.4.6 E. coli and non-E. coli bio-chemical identification is as follows if the emergence of bio-chemical reaction types outside the table above indicates that the culture may be incomplete, should be re-routed separation, if necessary, repeated tests.7.5 Results Report: E. coli is Gram-negative spore-free bacteria, fermented lactose acid production gas, IMViC test for the number of positive tubes of --, LST broth check the MPN table, report the MPN value of E. coli per gram (ml) sample. : This paper refers to SN0169-92 "E. coli groups, fecal E. coli groups and E. coli testing methods in exported food."