MFHPB-27 Enumeration of Escherichia coli in Foods by the Direct Plating (DP) Method
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6B72876CDE754B88B0643BE2293D5C4E |
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0.02 |
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页数： |
4 |
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日期： |
2012-3-2 |
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Published by: POLYSCIENCE PUBLICATIONS, P.O. Box 1606, Station St-Martin, Laval, Quebec,Canada H7V 3P9. TEL.: 1-800-840-5870. FAX: (450) 688-1930.,Government of Canada Gouvernement du Canada,HPB Method MFHPB-27,September 1997,HEALTH PROTECTION BRANCH,OTTAWA,ENUMERATION OF ESCHERICHIA COLI IN FOODS BY A,DIRECT PLATING (DP) METHOD,R.A. Szabo and E.C.D. Todd,Microbiology Research Division,Bureau of Microbial Hazards, HPB,Postal Locator: 2204A2,Ottawa, Ont., K1A 0L2,1. APPLICATION,This method is applicable to the enumeration of Escherichia coli biotype 1 in foods and food ingredients to,determine compliance with the requirements of Sections 4 and 7 of the Food and Drugs Act. Where an Official,Method for certain products is specified, that method shall be followed. This revised method replaces MFHPB-,27, dated December 1995.,2. DESCRIPTION,It has been used successfully for the enumeration of E. coli in red meats, poultry, pork, sausages, carcass,washings, fish, shellfish, milk, ice cream, cheese, apple and strawberry purées, flour, carrots, potatoes, swede,and green beans (see Section 8). With the exception of bean and alfalfa sprouts in which large numbers of,Klebsiella spp. may preclude an accurate determination of E. coli, the DP method can be used successfully for,the enumeration of E. coli in other foods and food ingredients.,3. PRINCIPLE,The conventional Most Probable Number (MPN) procedure for enumerating E. coli in foods takes 8 to 12 days,to complete, whereas, the Direct Plating (DP) method takes 24 to 30 h. In previous studies comparing these two,methods, the MPN procedure was shown to be less precise and yielded lower counts of E. coli in frozen meat,samples. In addition, the MPN procedure is incapable of enumerating late lactose fermenters and anaerogenic,E. coli which comprise approximately 10% of the Escherichia strains (2). The main disadvantage of the DP,method is its inability to enumerate non-indole producing E. coli which represent 3 to 5% of the E. coli strains,(2); this disadvantage is offset by the brevity of the method and its facility in enumerating anaerogenic E. coli.,4. DEFINTION OF TERMS,See Appendix A of Volume 2.,5. COLLECTION OF SAMPLES,See Appendix B of Volume 2.,MFHPB-27,- 2 - September 1997,6. MATERIALS AND SPECIAL EQUIPMENT,1) A supply of 0.45 mm membrane filters of 85 mm diameter (MF) (Available from Nuclepore Canada,Inc., Toronto, Cat No. 145318).,2) Peptone water (0.1%).,3) Nutrient agar (NA).,4) Tryptone Bile Agar (TBA).,5) Indole reagent.,6) A water-jacketed incubator to maintain a temperature of 44.5EC ± 0.5E or a similarly accurate,incubator.,7) Blender or Colworth Stomacher 400.,7. PROCEDURE,Analyze each sample unit individually. Carry out the test in accordance with the following instructions:,7.1 Handling of Sample Units in the Laboratory,7.1.1 During storage and transport, the following shall apply: with the exception of shelf-stable,products, keep the sample units refrigerated (0-5EC). Sample units of frozen products shall be,kept frozen.,7.1.2 Analyze the sample units as soon as possible after receipt at the laboratory.,7.2 Preparation for Analysis,7.2.1 Clean the surface of the working area with a suitable disinfectant.,7.2.2 Mark clearly the duplicate Petri plates identifying sample, sample unit, dilution, and date of,inoculation.,7.2.3 Have ready sterile peptone dilution water.,7.3 Preparation of Samples,7.3.1 To ensure a truly representative analytical unit agitate liquids or free flowing materials until the,contents are homogeneous. If the sample unit is a solid, obtain the analytical unit by taking a,portion from several locations within the sample unit.,7.3.2 For solid samples, blend or stomach 11 (10) g (the analytical unit) with 99 (90) mL of peptone,water for 2 min. Make decimal dilutions in peptone water. If a low count is expected, blend 11,(10) g with 44 (40) mL of peptone water (1:5). For liquid samples, shake 11 (10) mL with either,99 (90) mL (1:10) or 44 (40) mL of peptone water (1:5). Liquid samples may also be plated,undiluted (see below).,NOTE: Weight or volume in brackets indicates alternate procedure for making dilutions.,7.3.3 Check pH of the food suspension. If the pH is outside the range of 5.5-7.6, adjust pH to 7.0,with sterile 1N NaOH or 1N HCl.,7.3.4 If organisms in the sample might have been stressed by freezing or by other processing,proceed as follows, otherwise go directly to 7.3.9. On the prepoured and dried surface of a NA,plate, place an MF and flatten it against the agar with a sterile glass or plastic spreader. Avoid,trapping air bubbles.,7.3.5 In duplicate, plate 0.5 mL of two consecutive decima……
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