Food Safety and Inspection Service A review of laboratory-based regulatory and response activities.

Transcription

1 Food Safety and Inspection Service A review of laboratory-based regulatory and response activities.

2 Objective Provide an overview of recently implemented regulatory testing programs for non-o157 shiga-toxin producing E. coli and new standards for Salmonella and Campylobacter in young chickens and turkeys

3 Laboratory Activities Process 100,000 specimens Microbiology, chemistry, species identification National and international Support monitoring and enforcement programs ISO Accredited

4 Updates Overall program changes Salmonella and campylobacter L. monocytogenes E. coli O157:H7 Non-O157:H7

5 Program Structure HACCP verification component Prevalence Baseline Follow-up program Special interest projects

6 Salmonella / Campy programs HACCP verification programs Sample sets New std for salmonella and campylobacter Chicken Turkey Salmonella 5/51 4/56 Campylobacter 8/51 3/56 Two sets for each plant to get categorized Serotype differentiation Target for-cause follow up testing

7 ARS - FDA FERN Collaboration Send DNA prep from the broiler samples we receive to North Carolina Ag lab to evaluate using the PLEX ID instrument for identification of Salmonella strains using PCR and mass spectrophotometry NARMS testing for ATB resistance

8 Raw Red Meat Programs HACCP verification E. coli O157:H7 (ground, trim, components) Salmonella (swabs C/B, S/H, hogs) Pathogen-based vs. product-based Test one product for multiple targets Ground or trim or components tested for multiple pathogens Increase analytical portion size to 325g Target for-cause follow up testing

9 Increase sample analytical portion to Listeria monocytogenes programs HACCP verification Random and risk based program Baseline 4-year cycle census of plants under inspection For cause - Intensified Verification

10 Non-O157:H7 STEC programs Parallel the O157:H7 program Start with trim and trim component program Expand to ground as resources permit Timeline for results equal to O157 program Method preview

11 FSIS E. coli O157:H7 and Non-O157 STEC Method Comparison Day 0 Day 1 Day 2 PCR Screen POTENTIAL O157 STEC FSIS Method Samples Collected from Plant And sent to Lab Sample Prepared in Enrichment broth Samples incubated at 42⁰C For hours + - Confirm Negative O157 specific Immunomagnetic bead capture Beads plated onto Chromogenic agar Real Time PCR Screen for stx and eae genes POTENTIAL Non-O157 STEC - FSIS Method Samples Collected from Plant And sent to Lab Sample Prepared in Enrichment broth Samples incubated at 42⁰C For 22 hours + - Confirm Negative Real Time PCR Screen for target O groups + - O-group specific Immunomagnetic bead capture Beads plated onto Chromogenic agar Confirm Negative

12 FSIS E. coli O157:H7 and Non-O157 STEC Method Day 3 Day 4 Day 5 Aggluntination test on Typical colonies (O157) SBA O157 and H7 Agglutination Test or wzy, flic PCR Toxin ELISA or stx PCR Analysis extends past day 5 if restreaking is required to isolate colonies. + SBA - PRESUMPTIV E + - Biochemical Confirmation + Confirm Negative Confirmed Negative Confirm Positive Aggluntination (if available) test on Typical colonies + SBA SBA stx, eae, wzy PCR Analysis extends past day 5 if restreaking is required to isolate colonies. - PRESUMPTIV E Agglutination Test or wzy, PCR - Biochemical Confirmation + Confirm Negative Confirmed Negative Confirm Positive

13 ARS Collaboration FSIS enrichment broths will be tested for the features described in Prevalence and Characterization of Non-O157 Shiga Toxin Producing Escherichia coli Isolated from Commercial Ground Beef in the United States article.(bosilevac and Koohmaraie, 2011) Serotype (O:H)* Stx subtype +/-, Stx2 subtype +/- Eae subtypes Ehx +/- Stx subtype,stx2 subtype Suba +/- Presence of genetic markers within virulence associated O-islands of STEC isolates (e.g. nle)

14 Questions?

15 KEY: CDC/PulseNet = ARS/VetNet = Field Labs = NVSL = OSEL = LIMS (NADS) = START 1: Clinical clusters are added to WebBoard by PulseNet and member laboratories. New cluster PFGE patterns are retrieved by VetNet PulseNet Cluster information is uploaded to LIMS PulseNet Cluster PFGE patterns are compared to VetNet isolate patterns from previous 90 days (request BlnI) Matches to recent Clusters Matches to PulseNet Clusters Search 1. An establishment s isolate s pattern is compared to patterns from a recognized cluster of human illnesses, and matches are determined. Matches to old and new Clusters of CDC interest OSEL forwards list of matches to PulseNet Clusters to PulseNet OSEL flags clusters of CDC interest VetNet pattern names are assigned to all XbaI PFGE patterns (BlnI by VetNet as needed) VetNet is checked for PFGE patterns that match confirmed PulseNet patterns (BlnI by VetNet as needed) PulseNet names are added to the VetNet database, indicating that the isolate has a match in PulseNet. VetNet Frequency Calculated for pattern Pattern frequency ranked VetNet Pattern Search, VetNet Pattern History VetNet Pattern frequency, rank and distribution over time. Search 2. PFGE pattern frequency and history within VetNet is determined. PulseNet /CDC searches VetNet Database for matches to recent clinical isolates at discretion of PulseNet/ CDC Other Tests Isolates to ARS Isolates run with XbaI on PFGE Gel PulseNet Quarterly Report is uploaded to LIMS PulseNet Quarterly Report VetNet Database PFGE information added to LIMS data-fields LIMS Database LIMS checks against list of common clinical strains Matches to Common PulseNet patterns Search 3. An establishment's isolate's pattern is compared to common PulseNet isolate patterns, and matches are determined. Isolates/Plugs Archived START 2: Field laboratories confirm isolates to Salmonella genus. Isolates to NVSL NVSL serotypes isolates PulseNet Database OSEL performs 60-day PulseNet Pattern Search and 360-day PulseNet Pattern History on newly named VetNet isolates OSEL Uploads pattern search and history results to LIMS Matches to recent PulseNet patterns Search 4. An establishment's isolate's pattern is compared to recent PulseNet isolate patterns, and matches and pattern history are determined. Research Publications Data Analyzed LIMS Database (ARS Accessible) Collaborative Research Publications Collaborative work Data Filtered Data Analyzed LIMS Database (ARS Inaccessible) Establishment checked for previous instances of pulsetype Matches to PFGE Patterns previously associated with establishment or associated cooperation. Search 5. An establishment's isolate's pattern is compared to isolate patterns from samples previously collected at that establishment or associated cooperation, and matches are determined. 15