1. Human infections with E. coli O157:H7 and other Shiga toxin-producing bacteria P. I. Tarr, M.D. Washington University School of Medicine FDA AIDAC Meeting Rockville, MD April 12, 2007

2. Spontaneous Resolution (~85%) HUS (~15%) -3-201234567 Diarrhea Bloody diarrhea Culture + culture HCT < 30% Plts < 150K Cr > ULN Adapted from: Lancet 2005; 365:1073

3. Opportunities About 90% are seen by a physician pre- HUS Toxemic disorder

4. Challenge – Assess therapies to prevent HUS Identify infected patients –Quickly, accurately Low incidence, sporadic epidemiology, rural predominance –Many sites of presentation Narrow window to prevent sequelae –Vascular injury already is underway What is outcome of interest? –HUS vs. severe HUS Informed consent in urgent situation, multi-site randomization

5. Challenges – Use of an approved therapeutic Safety Efficacy Cost Carrying Costs to Institution

6. Diagnostic Challenges Target pathogen: All STEC? Only E. coli O157:H7? USA, Canada, Japan, UK, and South America: E. coli O157:H7 predominant, and nearly exclusive (> 95%), cause of HUS. Pediatrics. 1987;80:37 J Infect Dis. 1990;162:553 J Pediatr. 1998;132:777 J Infect Dis. 2001;183:1063 J Pediatr. 2002;141:172 Foodborne Pathog Dis. 2006;3:88 Epidemiol Infect. 2007 Mar 5 (epub)1-7

7. EIA vs. SMAC, point of care,1998-2001 1626 ER stools, 225 from private practice (all children) 39 signals in Meridian EIA (broth) (in ER) Klein, E, et al, J Peds 2002; 172

8. 39 EIA positives 11 non-O157:H7 STEC (> 1/5 colonies) O103:H2 (4), O118:H16 (2), O26:H11, O111:nm, O111:H8, O121:H19, Orough:H11 (1 each) (1 with C. jejuni) 3 signals, no STEC (20 colonies and broth tested) 25 E. coli O157:H7 Klein, E, et al, J Peds 2002; 172

9. 39 STECs O157 (28) non-O157 (11) HUS18% 0% Bloody 92%50% Laboratory blood 70%22% Klein, E, et al, J Peds 2002; 172

10. Why not test stool for toxin? Difficult analyte Often not there, especially when patient subsequently develops HUS Test misses about 10% of patients infected with E. coli O157:H7 Cornick NA, et. al., J Infect Dis, 2002; 57-63

11. Is EIA signal for real, if SMAC is negative? Prior probability: Probability of disease before performing a diagnostic test If high, then positive result likely to be valid If low, then positive test less likely to be valid

12. EIA+, SMAC no O157 Test applied to: ER Population Bloody diarrheaHighly credible, low risk Painful diarrhea Hospitalized diarrhea Signal from pathogenic STEC (i.e., O26, O45, O103, O111, O113, O118, O121, O145, 177)

13. STEC Testing Test applied to: Chronic diarrhea Infantile diarrheaNot highly credible Non-painful, non-if positive bloody diarrhea No STEC recovered, non-major serogroup isolated

14. Summary of Diagnostic Challenges O157 preferential testing should be performed before action Direct stool tests insensitive Choose population well (i.e., not large volume commercial laboratories) Time to positive critical

15. Spontaneous Resolution (~85%) HUS (~15%) -3-201234567 Diarrhea Bloody diarrhea Culture + culture

16. Bloody Diarrhea Diagnose, then treat, or … Treat all bloody diarrhea (would miss ~20% of E. coli O157:H7 patients, and most (> 90%) won’t have target infection) Syndromic profiling to increase specificity

17. Syndromic Profiling – Bloody Diarrhea as entry criteria Diarrhea for X days –X > 1 –X < 5-7 ≥ 3 bowel movements/day 9 months – 10 years No documented fever in health care setting Abdominal pain, especially when defecating

18. Syndromic Profiling Best in high acuity venues Will miss ~ 20% of cases Will have ~ 2-3:1 false positive rate Must be coupled with expeditious testing If in research setting, needs support for negatives

19. Presymptomatic/Contacts Epidemics, household 10% of cases are 2 0, but few (of many) contacts become 2 0 cases Massive outbreaks on wane at time of “discovery.” Few symptomatic subjects subsequently come to light.

20. Early in Timeline (Pre-Bloody Stage) Contact of known case (but most symptomatic contacts are not infected) Strain – specific replicates

21. OnsestPresentationHUS

22. 10,000 diagnosed cases per annum 2,500 cases < age 10 48 cases < age 10 per week, of whom 7 will develop HUS (15% rate) One child in the US daily develops HUS. Today, two children in the US are in the opportune post-presentation, pre-deterioration, window. Today, 14 children are in that window but will not develop HUS Extrapolations

23. First Encounter High acuity setting (ED) ‘Primed’ point of care Research projects focusing on diarrhea, bloody diarrhea, and E. coli O157:H7 Outstanding microbiology

24. ER KINETICS Inter-defecation interval (O157:H7+): 206 min 70% chance of producing stool on site if in ER for 4 hours

25. CHRMC ED, 1998 - 2001 4799 children with “diarrhea” or “gastroenteritis” 1626 “enrolled” 39 STEC 28 E. coli O157:H7 (every 39 days) 5 HUS (all O157:H7) (every 219 days) Klein, EJ, J Pediatrics (2002) 141:172; Clin Infect Dis (2006) 43:7

26. CHRMC ED, 1998 - 2001 4799 children with “diarrhea” or “gastroenteritis” 1626 “enrolled” (34%) 28 (1.7%) + for O157:H7 372 specimens submitted as cups; 5.4% + 1254 non-cups (largely swabs); 0.6% + Klein, EJ, J Pediatrics (2002) 141:172; Clin Infect Dis (2006) 43:7

27. Target Population Bloody diarrhea → culture + → 22/237 9.3% (miss 6) (once every 7 weeks) Bloody diarrhea → HUS → 4/237 1.7% (miss 1) (once every 39 weeks)

28. Spontaneous Resolution (~85%) HUS (~15%) -3-201234567 Diarrhea Bloody diarrhea Culture + culture Does host injury precede presentation/ diagnosis? Adapted from: Lancet 2005; 365:1073

29. * F 1+2: Thrombin generation before HUS NEJM 2002; 346:23 P<0.01 p<0.001 THROMBIN + F 1+2 FXa PROTHROMBIN NEJM 2002; 346:23

30.  D-dimer Before HUS, as Lesion Evolves * * NEJM 2002; 346:23

31. Fibrin Degradation D-dimer signifies fibrin degradation, infers fibrin presence DDEDDEDDD Cross-linked fibrin polymers D-Dimer Plasmin

32. LABORATORY VALUES - ALL GROUPS NormalUncomp Pre-HUS HCT (%) 36  3 37  3 38  5 Plts (k/mm 3 ) 321  70 317  74 322  97 Cr (mg/dL) 0.4 .1 0.4 .1 0.4 .2 NEJM 2002; 346:23

33. Stx vanishing from Stool DOES SHIGA TOXIN IN STOOL RELATE TO HUS RISK? Stx FrequencyTiter Pre-HUS: 40%320 (160-1280) Uncomplicated: 48% 1689 (160-40 K) At HUS: 16% Cornick, N., J Infect Dis. 2002;186:57

34. Additional Pre-HUS findings and host perturbations Stx not on circulating PMNs (“PMN are not acting as transporter for Stx in the pathogenesis of HUS.”) ( Nephrol Dial Transplant 2007; 22:749 ) PAF activated (Pediatr Nephrol 2002;17:1047) Platelets activated (Blood 2006; 108:167) vWF multimers show signs of shear-induced cleavage (? from nascent thrombi) (Pediatr Res 2001; 49:653)

35. Outcome Definition Hemolysis: Packed cell volume <30%, smear evidence of intravascular erythrocyte destruction Thrombocytopenia: Platelets <150,000 mm -3 Renal insufficiency: Creatinine > ULN age Urinalysis not in criteria (difficulty obtaining a reliable specimen, non-functional data)

36. Outcome Subgroups Mild HUS: transfusions, no dialysis, nonanuric. LOS: 6 days Severe HUS: dialysis, anuria, LOS: 12 days Anuric HUS: best predictor of chronic renal injury ( JAMA 2003; 290:1360; J Pediatr 1991; 118:195 ) Ake, J. A. et al. Pediatrics 2005;115:e673

37. Anuric vs. Non-anuric HUS CHRMC, 1997-2003 16 children with anuric HUS 13 children with non-anuric HUS All E. coli O157:H7 + Variable routes of enrollment (syndromic profiling (2), contact (1), culture + (19), arrive with HUS (7)) All pre-HUS variables examined

38. Copyright ©2005 American Academy of Pediatrics Ake, J. A. et al. Pediatrics 2005;115:e673 Fig 1. Timing of critical events during illness

39. Copyright ©2005 American Academy of Pediatrics Fig 2. Volume and characteristics of fluids that were administered during first 4 days of illness Ake, J. A. et al. Pediatrics 2005;115:e673

40. Multivariate Analysis VARIABLEADJUSTED RELATIVE RISK (95% C.I.) P Age (yr)1.9 (.8-4.4)0.15 Female sex1.5 (.1-19.4)0.77 Pre-HUS antibiotics1.1 (0.1-17.0)0.95 Free water in IVF (mL/m 2 ) 1.0 (.999- 1.001) 0.49 Na in IVF (mmol/m 2 )0.994 (.989-.999).017 Pediatrics. 2005 Jun;115(6):e673-80 Ake, J. A. et al. Pediatrics 2005;115:e673

41. Target Population Bloody diarrhea → culture + → 22/237 9.3% (miss 6) (once every 7 weeks) Bloody diarrhea → HUS → 4/237 1.7% (miss 1) (once every 39 weeks) Bloody diarrhea → anuric HUS → 0/237

42. Four patterns of outcome Tarr, PI, et al, in Yamada, T., Textbook of Gastroenterology, Blackwell, 5 th ed., in press

43. Trial Specifications Diagnose early, accurately Intervene early Support all patients with aggressive volume expansion (IV) Collect data in usable fashion for subsequent users Benefits (what outcomes are to be averted?) to be weighed against risk Address important consent issues re urgent enrollment Might need to address Stx2c Return to Main Menu.

44. Eileen J. Klein, MD, MPH, Jennifer R. Stapp, BS, Carla R. Clausen, PHD, Daniel R. Boster, BS, Joy G. Well, MS, Xuan Qin, PhD, David L. Swerdlow, MD, and Philip I. Tarr, MD. Shiga toxin-producing Escherichia coli in children with diarrhea: A prospective point-of-care study. The Journal of Pediatrics 2002;172-178. Return to Main Menu.