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Trade Memoranda
T-3-151s - Guidelines for the evaluation of methods to
determine the viability of direct fed microbials and forage additives
containing viable microorganisms
February 15, 1999
Part of the process for registration of direct fed microbials or forage
additives containing viable microorganisms by the Canadian Food Inspection
Agency (CFIA) is the provision by the applicant of an acceptable method of
analysis for the microorganisms in the carrier or filler. The applicant is also
required to provide the necessary data to support reliability of the method.
The following protocol is intended to ensure that the method, data and test
materials provided by the applicant are adequate to allow for expeditious
evaluation, validation and approval of the methodology.
1. Method Format
The purpose of the method submission is to permit the verification of label
claims and to allow the evaluator and analyst to understand and perform the
method exactly as the originator intended. It is therefore necessary that the
method contains all the necessary information presented in a detailed
step-by-step standard operating procedure.
The method submitted must be clear and concisely written, currently in use by
the submitting company, and proven to be specific, selective, reliable, and
accurate for the enumeration of viable microorganisms in formulated products.
Critical control points, specifications for quality control and familiarization
must be included in the methodology.
The following supporting documents for method performance standards must be
included in the submission to properly evaluate the method:
- Description of the microorganism(s) added in the product with the strain
identification and source.
- Specifications on the level(s) of microorganism(s) guaranteed in the
product, expressed in colony forming unit (CFU) per g or ml.
- Enunciation of the scope and principles of the
methodology.
- List of references supporting the methodology.
- Description of the reagents and equipment, along with specific ordering
information and performance criteria.
- Description of the experimental design with complete standard operating
procedures used for the method evaluation. The description should include
information on sample preparation and procedures for the analysis of samples,
controls and standards.
- Identification of the critical control points and specifications, including
information on stability of the reagents and samples, and specification on
potentially existing interfering substances such as non-target organism,
components of the sample matrix, chemical additives, drugs etc.
- Safety information and precautions.
- Any existing information on collaborative studies.
- Any other pertinent information.
- When the submitted method is an alternative to those based on standard
plate counts, such as methods based on ELISA, DNA probing, bioluminescence
etc., additional information including standard curves, specificity and
sensitivity data may be requested depending on the need.
2. Supporting Data
The applicant must provide the following information in support of the
proposed methodology, which includes raw data and statistical analyses from
their own method evaluation (data should be presented in a tabular form):
- Provide analytical results from three production batches, each tested in
triplicate (indicate lot numbers), including calculations of
CFU / g or ml and coefficients of variation for
precision assessment.
- Provide analytical results from various days, analysts, reagent lots or
from various laboratories, including calculations of CFU / g or ml and
coefficients of variation for repeatability and reproducibility
assessments.
- When a reference method is available, provide analytical results, including
calculation of CFU / g or ml from the reference method, to specify the accuracy
of the submitted method in comparison with the reference method.
- Provide analytical data confirming the selectivity of the method resulting
from the proper isolation of the target microorganism. Adequate identification
may be obtained from data on gram stain, cell morphology, biochemical analyses,
patterns on utilization of carbohydrates, or any other valid confirmation
tests.
- When the purpose of the submitted method is the analysis of microbial
contaminants or microorganisms detected at low levels, provide analytical
results establishing limits of detection.
3. CFIA Laboratory Evaluation
When the above requirements are met, CFIA will proceed with a desk review
followed by a laboratory evaluation of the method. For the laboratory
evaluation, the following materials must be supplied by the company.
- An appropriate amount of pure culture or culture mix.
- An appropriate amount of non-inoculated sample matrix, carrier or filler
(minimum 250 g).
- An appropriate amount of product sample (for 5 analyses per batch) from the
same three production batches analysed previously by the company (see 2.
Supporting data).
- Any uncommon media, reagents or supplies that are required in the
methodology.
4. Evaluation Criteria
An adequate method for the enumeration of the microorganism(s) will be
approved following criteria based on specificity, accuracy, precision,
repeatability and reproducibility, taking into consideration the analytical
results provided by the company and those generated by CFIA.
5. CFIA Report
Upon request, CFIA will prepare a report including the following
information:
- Title, name of applicant, identification of the CFIA laboratory and the
evaluators, and date of report.
- Summary statement indicating acceptance or rejection of the method, with
signatures and dates.
- Results of the evaluation of the data provided by the applicant and the
data generated by the CFIA laboratory, with comparisons.
- Tables including raw data, calculations and statistical analysis for the
determination of specificity, accuracy, precision, repeatability and
reproducibility of the submitted method.
- Discussion and conclusion.
6. Changes to Methodology
CFIA must be informed of any modifications of the methodology. The new or
modified method must be submitted to a desk review. The degree
of verification will depend on the changes.
Questions regarding this memorandum may be directed to :
Feed Section
Animal Health and Production Division
Canadian Food Inspection Agency
59 Camelot Drive
Nepean, Ontario
K1A 0Y9
Phone: (613) 225-2342
Fax: (613) 228-6614
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