Annex D-1: Methodology for Escherichia coli O157:H7 testing of beef products

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There are specific requirements for laboratory methodology used to test for E. coli O157. These requirements apply to all E. coli O157 testing of beef products (e.g.: beef carcasses, beef trim/trim components, ground beef, raw/cooked meat patties and uncooked dry or semi-dry fermented products containing beef) produced in Canadian registered establishments eligible for export to the USA.

Note that the requirements for non-O157 Shiga toxin-producing E. coli verification sampling and testing of beef trimmings for abattoirs eligible for export to the USA are described in Annex D-2.

Industry must ensure that their laboratories and/or private accredited laboratories contracted for testing of the above products are informed of these methods.

Private laboratories must follow these method instructions when their clients make them aware the sample is submitted from a USA eligible establishment:

Acceptable methodology for E. coli O157 testing of beef products

Laboratories testing beef products (e.g.: beef carcasses, beef trim/trim components, ground beef, raw/cooked meat patties and uncooked dry or semi-dry fermented products containing beef) from establishments eligible for export to the USA, have been instructed to test using either of the following methods.

  • The acceptable FSIS methodology for both screening and cultural confirmation for E. coli O157/NM.
    or
  • The approved Compendium screening methods listed in Annex D and cultural confirmation using MFHPB-10.

Testing Procedure

  1. Test five 65 ± 2 g randomly collected sub-samples (total analytical sample size of 325 ± 10 g) that are representative of the entire sample. The 5 sub-samples may be tested individually or composited into one 325 g sample if allowed by the method used. Samples that are negative by the screening method can be reported as negative for E. coli O157. Samples that are presumptive positive by the screening test will proceed to cultural confirmation.
  2. Acceptable methodology can be found in Annex D.
  3. Upon confirmation, if the cultural method confirms the presence of typical, non-sorbitol fermenting E. coli O157 on selective agar, the following steps are required in addition to the requirements outlined in MFHPB-10:
    • It is mandatory to confirm the presence or absence of the toxin with acceptable methodology (e.g., MFLP-83, or equivalent).
    • If the toxin test is negative, test for the presence of verotoxin genes (stx1 and/or stx2) by an acceptable PCR method (e.g., MFLP-62, or equivalent).
    • If the test performed (either the toxin production test or the genetic test) is positive, it will be concluded that the isolate is confirmed positive for the presence of verotoxin producing E. coli O157 and no further confirmatory testing is required.
    • If the toxin tests performed are both negative, test for the presence or absence of the H7 antigen. If the test is positive, it will be concluded that the isolate is a confirmed E. coli O157 positive and no further confirmatory testing is required.
    • If the test for the H7 antigen is negative, test for the presence of the H7 gene (fliC) using acceptable PCR methodology. If the test for the H7 gene is confirmed positive, then the sample is considered confirmed positive for E. coli O157.
    • If the H7 gene is not confirmed (assuming toxin production and toxin gene, as well as H7 serology tests were negative), then the sample is considered to be negative for E. coli O157.

Note - Genetic tests for the presence of toxin genes and/or H7 genes can be done in CFIA laboratories or in private accredited laboratories using methodology approved by the CFIA.

For details regarding shipping isolates to the CFIA for PCR gene testing or for clarification to the methodology instructions above, please contact the CFIA at the following address:

CFIA Laboratory Coordination Division
e-mail: CFIA.LCD-DCL.ACIA@inspection.gc.ca
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