A CLEARER PATH FOR PATHOGEN TESTING

HOST Bill McDowell Editorial Director, Meatingplace MODERATOR Tom Johnston Editor, Meatingplace

WHAT IS ACHIEVABLE IN PATHOGEN TESTING? Gary R. Acuff Professor, Food Microbiology Head, Department of Animal Science Gary R. Acuff Professor, Food Microbiology Head, Department of Animal Science

Poll Question What is really achievable in pathogen reduction? 1. It depends upon how much contamination is present Logs. 3. A scientific endpoint is needed. 4. Complete elimination of pathogens.

Pathogens Enterohemorrhagic Escherichia coli O157:H7 Non-O157 Salmonella Campylobacter? Focus on EHECs Enterohemorrhagic Escherichia coli O157:H7 Non-O157 Salmonella Campylobacter? Focus on EHECs

Source Cattle Gastrointestinal tract Anything in close proximity to feces Hide Hooves Dust Water Cattle Gastrointestinal tract Anything in close proximity to feces Hide Hooves Dust Water

Control Opportunities Genomics Livestock production & feedlot Handling and well-being issues Slaughter Fabrication Retail Consumer, foodservice Genomics Livestock production & feedlot Handling and well-being issues Slaughter Fabrication Retail Consumer, foodservice

Control Opportunities Genomics Livestock production & feedlot Handling and well-being issues Slaughter Fabrication Retail Consumer, foodservice Genomics Livestock production & feedlot Handling and well-being issues Slaughter Fabrication Retail Consumer, foodservice Where are the hazards most effectively addressed?

Control Opportunities Genomics Livestock production & feedlot Handling and well-being issues Slaughter Fabrication Retail Consumer, foodservice Genomics Livestock production & feedlot Handling and well-being issues Slaughter Fabrication Retail Consumer, foodservice Where are the hazards most effectively addressed?

Consumer Education

Carcass Interventions Temperature Chemical Temperature Chemical

Laboratory Challenge Parallel evaluations Marker pathogens in fecal material Salmonella serotype Typhimurium Escherichia coli O157:H7 Non-inoculated fecal material Indicator organisms Parallel evaluations Marker pathogens in fecal material Salmonella serotype Typhimurium Escherichia coli O157:H7 Non-inoculated fecal material Indicator organisms Feces plus Pathogens

Log Reduction of E. coli O157:H7 and S. Typhimurium on Beef Carcass Surfaces

Log Reduction of E. coli O157:H7 and Coliforms on Beef Carcass Surfaces

What Is Realistic? Elimination? Reduction? The real issue: What remains? And where is it? Elimination? Reduction? The real issue: What remains? And where is it?

Trimming and grinding processes will distribute any remaining pathogens.

Unfortunate Event Adulterant declaration Well-intended (possibly), but short-sighted Zero doesnt exist in bacterial enumeration Unintended consequence - difficult to measure improvement or control Adulterant declaration Well-intended (possibly), but short-sighted Zero doesnt exist in bacterial enumeration Unintended consequence - difficult to measure improvement or control

What is really achievable? 1. Does it really matter? 2. It all has to be gone. 3. Attempt to achieve what is required. 4. Absence is unreasonable (impossible, illogical, unscientific, other favorite adjective). 1. Does it really matter? 2. It all has to be gone. 3. Attempt to achieve what is required. 4. Absence is unreasonable (impossible, illogical, unscientific, other favorite adjective).

An Endpoint is Needed How can one aim for a target that does not exist? Or one that constantly changes? How can one aim for a target that does not exist? Or one that constantly changes?

Food Safety Objective The maximum frequency and/or concentration of a hazard in a food at the time of consumption that provides or contributes to the appropriate level of protection.

Food Safety Objective H 0 - R + I FSO FSO = Food Safety Objective H 0 = Initial level of the hazard R = Total (cumulative) reduction of the hazard I = Total (cumulative) increase of the hazard FSO, H 0, R and I are expressed in log 10 units H 0 - R + I FSO FSO = Food Safety Objective H 0 = Initial level of the hazard R = Total (cumulative) reduction of the hazard I = Total (cumulative) increase of the hazard FSO, H 0, R and I are expressed in log 10 units

Food Safety Objective H 0 - R + I FSO 3 - R R -2 R -5 R 5

5-Log Reduction H 0 - R + I FSO 3 - R R -2 R -5 R

Reduction of E. coli O157:H7 and S. Typhimurium on Beef Surfaces by Sequential Pre- and Post-chill Lactic Acid Sprays

Microbiological Testing In the absence of an FSO, it is necessary to establish default criteria. Safe Harbor In the absence of an FSO, it is necessary to establish default criteria. Safe Harbor

Microbiological Testing Where are we going? Is there an end? Is more sampling the answer? Where are we going? Is there an end? Is more sampling the answer?

Microbiological Testing In the ICMSF scheme for managing risk, two uses for criteria are identified To validate that control measures meet performance criteria 2. To determine acceptability when no more effective means of providing assurance is available (i.e., in the absence of knowledge that HACCP has been properly applied) In the ICMSF scheme for managing risk, two uses for criteria are identified To validate that control measures meet performance criteria 2. To determine acceptability when no more effective means of providing assurance is available (i.e., in the absence of knowledge that HACCP has been properly applied)

Microbiological Testing In the ICMSF scheme for managing risk, two uses for criteria are identified To validate that control measures meet performance criteria 2. To determine acceptability when no more effective means of providing assurance is available (i.e., in the absence of knowledge that HACCP has been properly applied) In the ICMSF scheme for managing risk, two uses for criteria are identified To validate that control measures meet performance criteria 2. To determine acceptability when no more effective means of providing assurance is available (i.e., in the absence of knowledge that HACCP has been properly applied) If there is a way out, this may be it.

Poll Question Results What is Really Achievable in Pathogen Reduction? 1. It depends upon how much contamination is present Logs. 3. A scientific endpoint is needed. 4. Complete elimination of pathogens.

Provide a scientific endpoint and the industry will achieve the necessary reduction.

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CURRENT BEEF INDUSTRY SAMPLING AND TESTING PROGRAMS FOR E coli O157:H7 Timothy P. Biela EVP Food Safety & Quality AFA Foods

Poll Question Which best describes your position? Packer Processor Other Are you currently testing the following products for E coli O157? TrimYESorNO Ground Beef YESor NO

What are we doing and why? Carcass Testing – USDA Mandated carcass testing Boneless Beef Trim Testing – N60 Sampling and Testing – Sub-lot size varies from 1 to 5 combo bins Raw Ground Beef Testing – Commercial versus Retail Testing Programs

Food Safety Verification for O157 Verification of HACCP and Food Safety Program for control of Enteric Pathogens – Must define the sublot – Must define the sample size and frequency of sampling – Must define the actions to be taken in the event of a positive result

Total-N60 Platform The Food Safety Umbrella Sample units of n=60 per lot provides robust statistical basis. Statistical confidence of N60 testing is at least 95.0% probability that ECH7 will be detected if present [assuming a population prevalence rate of 5.0% or higher]. Surface Slice vs SubSurface Tissue Surface Area – N60 examines substantially more total surface area of external surface tissue vs other trim or final-grind sampling methods. Dilution Effect – Thin surface slices vs sterile underlying tissue or ground meat gives increased probability of detection. 5X more sensitive than finished ground beef sampling. Lab method sensitivity – capable of detecting 15 cfu/375g, 100% of the time. 1:5 enrichment media [vs 1:10 standard] improves sensitivity. PCR-BAX initial screening using modified-enhanced methodologies provides +90% accuracy. This slide courtesy of others. For Example only.

N60 Sampling 60 individual samples are taken from the lot Preference is to sample surface material Samples should be taken randomly across the entire stream or population of meat Various sizes are used; 1x3, 2x5, etc…. Lot sample size must be a minimum of 375 g The entire sample should be analyzed

References Beef Industry Food Safety Council Best Practices – USDA FSIS – International Commission on Microbiological Specifications for Foods (ICMSF) –

Microbiological Testing Programs must have : A robust sampling plan that is designed to meet the microbiological testing objectives with a high degree of confidence. The adequacy of the sampling plan should be evaluated by an independent third party. In order to ensure the interpretability of testing results, the sampling plan must be implemented correctly. Based on the results of microbiological testing, actions must be taken. (Plan / Do / Check / Act)

Key Elements of Sampling Plans Define what constitutes a sample, Define the number of samples to be collected, Define how samples will be collected, Define the number of samples it will collect, Define the frequency of sample collection, Define the procedures used to analyze samples and, Define the criteria for signaling an out-of-control process

Key Elements of Sampling Plans (2) Ensure that samples collected are representative of the entire population; minimize bias, Ensure that the sample size is sufficient to provide the desired level of confidence. Ensure the sampling plan addresses the fact that pathogens are heterogeneously distributed Ensure that sampling considers the variability in prevalence rates over time (seasonal variation).

Food Safety Verification for O157 Example #1 Major Retailer w Grinding Operation: – One sample every two hours of production. – Samples are analyzed with PCR/DNA. – Positive products are diverted to further processing. – Positives are bracketed from one hour in front of first positive to end of the day. – Positive rates range from.4 to.7%.

Food Safety Verification for O157 Example #2 Major Integrated Fed Beef Packer – Format varies from one sample per hour to one sample every fifteen minutes to screening pre- ground products prior to packaging. – Samples are analyzed with PCR/DNA. – Positives are bracketed from one hour in front of first positive to one behind and subjected to intensified sampling. – Positive rates range from.4 to.7%.

Food Safety Verification for O157 Example #3 Major Quick Service Restaurant – One sample taken for every batch of ground beef formulated. – Samples are composited for four batches. – Composites are sampled and a 25 gram analyte is tested. – Positives are bracketed from one hour in front of first positive to end of the day. – Positive rates have ranged from 0 to.2%.

HACCP and Risk Assessment for Raw Ground Beef Products Retail versus Commercial Ground Beef – Retail Consumers; Families No identified platform for cooking No verification of cooking controls – Commercial Quick Serve and Casual Dining Specific platforms for cooking and controls

Commercial Testing Event Summary Example

Commercial Testing Event Summary Example (2)

Summary Sampling and testing for pathogens is difficult. The key to an effective microbiological testing program is the sampling plan. Since it is not possible to conduct 100% testing, one must use a sample to draw inferences about the entire population. As indicated in the draft USDA Compliance Guideline, the sampling plan should be designed to provide a high probability of finding E. coli O157:H7, if it is present, and identifying a process that is out of control.

Summary (2) Due to the heterogeneous distribution of pathogens in meat products, microbiological testing cannot guarantee that product is free of contamination and should not be used for product acceptance. Microbiological testing should be used to verify process control and should occur on an ongoing basis as part of the Plan-Do- Check-Act cycle. Consistent and uniform sampling and testing methodologies for microbiological testing of beef manufacturing trimmings should be developed and implemented with the primary objective of protecting public health. N60 sampling should be the minimum sampling protocol for beef manufacturing trimmings in all federally inspected meat and poultry establishments.

Poll Question Results Are you currently testing trim or raw ground beef products for E coli O157? TrimYESorNO Ground Beef YESor NO PackerYES orNO ProcessorYESorNo

Ted Brown Senior Food Scientist Cargill R&D Center BUILDING THE LABORATORY RELATIONSHIP

POLL QUESTION Lateral flow users: are you using a lateral flow or antigen/antibody based test that is specific for E. coli O157:H7? Yes No

Discussion Focus Methodology Selection Laboratory / Establishment Relationship

Before you Decide that Testing is necessary and on a Methodology Remember: Testing is a PART of a Food Safety System

60 NRC July 14, 2010 Monitoring & Verification Lotting What does the sample represent? Sampling How will the sample be collected? Analysis What target will be analyzed? Decision Making What decisions will be made based on the data?

Methodology General Indicators (APC) Generic E. coli Shiga toxin producing E. coli E. coli O157 E.coli O157:H7 DNA typing PROGRAM ACTION

Test Methodology Options

Gold Standard > Time to results 72h to Negative High skill Stopping point defines reported result O157:H7 O157 Cultural Less skill Typically target O157 Further testing is necessary for O157:H7 > enrichment times than DNA based methods Antigen Medium skill Typically target O157:H7 Some kits may be O157 specific < time to result DNA

Basis for Method Selection… Test Result Application Approvals Business Based Considerations Fit For Use

Test Method Application Will the method provide the information needed to support my program objectives? Do you know if the reported result for your method is O157 or O157:H7 or something entirely different?

Approvals Methods should be evaluated by an independent body Not all approvals are created equal Approval alone should not be the basis to make a method selection

Business Based Considerations Goal is to find the target bug This is not just a price based decision This is an organization/business specific question with specific organization/business based answers

Fit for Use Do I have the right tool for the job? – Is my method validated for the matrix as I am going to use it Analytical unit size Enrichment/Incubation Detection limit Are there any variations from the method as it was validated compared to my use?

Laboratory Relationship Is not: Only About Cost Data Inquisition Is: Investment Trust, Confidence and Engagement Mutual understanding of Fit for use

Testing is a PART of a Food Safety System. Not all methods are created equal: O157 positive result, does not mean that it is O157:H7 specifically Who or what will the results effect? What will you do with the results? Good, bad, or Ugly? Method must be fit for use when selected and as used A lab relationship is an investment A CLEARER PATH FOR PATHOGEN TESTING

POLL QUESTION RESULTS Lateral flow users: are you using a lateral flow or antigen/antibody based test that is specific for E. coli O157:H7? Yes No

QUESTIONS & ANSWERS

FOR MORE INFORMATION: Gary Acuff: Timothy Biela: Ted Brown: 3M: Christine Aleski,Global Marketing Development Manager Bill McDowell: Tom Johnston: Webinar recording and PowerPoint presentation will be ed to you within 48 hours. For more information: