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Katie Boland DVM, PhD June 15, 2013

1. Vaccination of cattle with Escherichia coli O157:H7-derived proteins results in humoral and cellular immune responses but does not confer protection against subsequent challenge. Katie Boland DVM, PhD June 15, 2013. 2. E. Coli O157:H7. Causes disease in humans Diarrhea (MMRW, 1985)

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Katie Boland DVM, PhD June 15, 2013

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  1. 1 Vaccination of cattle with Escherichia coli O157:H7-derived proteins results in humoral and cellular immune responses but does not confer protection against subsequent challenge Katie Boland DVM, PhD June 15, 2013

  2. 2 E. Coli O157:H7 • Causes disease in humans • Diarrhea (MMRW, 1985) • Hemolytic-uremic syndrome (HUS) (Riley LW, Remis RS, Helgerson SD, et al, 1983) • Renal failure, hemolytic anemia and thrombocytopenia • Pathogenesis • Attaching and effacing (A/E) lesions • Pedestal formation • Dissolution of brush border • Shiga toxins • Prevention is key

  3. 3 Reservoir • Cattle are major source of contamination (Armstrong et al., 1996) • Transient colonization • Variable shedding • Meat • Produce • Water • Recto-anal junction • Lymphoid rich tissue

  4. 4 Pre-processing control • Decontamination • Probiotics • Diet • Antimicrobials Vaccination http://www.chadcompany.com/MVC-006F.JPG

  5. 5 A/E lesions (Gauthier, Finlay 2002)

  6. 6 HYPOTHESES 1: Mucosal immunization of naïve cattle with recombinant intimin induces a more robust adaptive immune response at the recto-anal junction than subcutaneous immunization 2: An induced adaptive immune response to recombinant E. coli O157:H7 proteins is associated with decreased colonization

  7. 7 Trial Setup • Trials 1, 2 and 3 • 3 animals per group • Trial 1: Cholera toxin B subunits (rectal) and Freund’s (SQ) • Trial 2: TLR 7/8 agonist (rectal and SQ) • Trial 3: TLR 4 agonist (rectal and SQ) • Immunized weeks 0, 3, 6

  8. 8 cellular responses Systemic PBMC Responses Local MRLN Responses

  9. 9 Trial 4 • Is the MRNL response targeted? • TLR 7/8 agonist • All get SQ immunization week 0 • Immunized again weeks 8 and 11 • All SQ immunizations on same side • Evaluate MRLNs as well and prescapular LNs

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  13. 13 Serum Antibody responses • All 4 trials: • Significant increases in titers to INT • Significant increases in titers to OVA in SQ groups • Variable responses to OVA in rectal groups • In trial 1 and 4: • Titers in SQ groups significantly increased compared to rectal groups

  14. 14 HYPOTHESES 1: Mucosal immunization of naïve cattle with recombinant intimin induces a more robust adaptive immune response at the recto-anal junction than subcutaneous immunization (reject) 2: An induced adaptive immune response to recombinant E. coli O157:H7 proteins is associated with decreased colonization

  15. 15 Challenge Trials Trials 1 and 2 Trials 3 and 4 Trial 3 10 animals per group Previous oral or rectal exposure Oral or rectal challenge Trial 4 Oral group from trial 3 Rectal challenge • 6 animals per group • Immunized or sham • 4(trial 1) or 3 (trial 2) doses • Challenged • Rectal (trial 1) • Oral (trial 2) Colonization levels followed for 4-7 weeks Evaluated PBMC and serum antibody responses

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  18. 18 Serum Antibody Responses • In trials 1 and 2: • All animals had significant increases in titers to all proteins • No change 2 weeks post-challenge • In trials 3 and 4 • Variable, low responses to all proteins • Tended to be higher in trial 4 • In all trials: • Did not correlate with colonization duration

  19. 19 Challenge Summary

  20. 20 Challenge Conclusions • Immunization and experimental exposure induced lymphoproliferative responses • Immunization induced significant antibody responses • No significant increase in responses after challenge • Neither lymphoproliferative nor serum antibody responses conferred protection against colonization

  21. 21 Overall Summary • Mucosal immunization does not induce a more robust regional immune response compared to SQ immunization • Regional responses overall more robust than systemic and are targeted in SQ immunization • Induced immune responses are not protective against challenge

  22. 22 Thank you! Tovah Kerr Susan Smart Emma Karel Fred Loaiza Lonnie Austin Kevin Lahmers Tim Baszler Tom Besser Wendy Brown Doug Call Esther Trueblood WADDL Pathologists and residents VMP Faculty and staff Graduate Students Officemates Friends Family Alex Beck Andrea Hayles Eric Sutten Kathleen Sutten Allison Vilander Claire Miller Carolyn Bohach HaiqingSheng Claudia Deobald

  23. ELISA Negative control Samples 1:10 1:20 Positive if > average + 2 SD 1:1280 Titer is last positive dilution

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