Non-O157:H7 STEC: Point of care, population- based, and clinical laboratory- centered analyses. P. I. Tarr, MD Washington University School of Medicine FSIS USDA October 17, 2007

Perspectives USA Pediatric Systematically collected specimens, in context of patient care

Background Many different E. coli serotypes produce Shiga toxin (Stx) 1 (and its variants), 2 (and its variants), or both. Only a subset has been demonstrated to be pathogenic to humans.

Four groupings of STEC 1.E. coli O157:H7 global; causes epidemics and severe disease, variety of sources, enduring importance easily detected in human specimens, using best microbiological diagnostic practices.

E. coli O157:H7 Sorbitol MacConkey agar

Four groupings of STEC 2. E. coli O157:H - focal (esp. Germany, not USA yet); increasingly recovered in other countries sources not yet discerned less easily detected – because toxin assays needed to find them

Four groupings of STEC 3.Pathogenic non-O157 STEC (e.g., E. coli O26, O111, …) global; variable type distribution; rarely epidemic; epidemiology and sources unclear less easily detected – because one needs toxin assays to find them might be transmitted by food

Four groupings of STEC 4. nonpathogenic STEC: frequent in animals, environment, food.

STEC cause HUS (Hemolytic Uremic Syndrome) Hemolytic anemia (hct < 30%), Thromobocytopenia (plts <150,000/mm 3 ), Renal insufficiency (creatinine > uln for age) Occurs 5-13 days after diarrhea begins

HUS 15% of culture + E. coli O157:H7 patients < 10 yo will develop HUS however, some non-O157 STEC seem particularly virulent (esp. O157:H -, O111, O113), but in aggregate non-O157 STEC are much less likely to cause HUS

Diagnosis in humans E. coli O157:H7: at time of HUS, 2/3 of patients can be negative. If not found at time of HUS, it does not mean that this pathogen was absent.

ca. 2.5 episodes of diarrhea/annum Vast majority are viral, and self-limiting Only about 15 million cultures performed per annum Cumbersome technology STEC in Humans: Challenges and Opportunities

Hospital ER Office Bloody Diarrhea HUS

Hospital ER Office Bloody Diarrhea HUS Lab

Detection and Enumeration Biases Bloody diarrhea and HUS patients are disproportionately cultured Rural disease, and resources needed to isolate pathogens are often not available at points of presentation

Archaic, Disjointed Technology

Hospital ER Office Bloody Diarrhea HUS Lab E. coli O157:H7 Signal Non O157:H7 STEC

Hospital ER Office Bloody Diarrhea HUS Lab E. coli O157:H7 Signal Non O157:H7 STEC State CDC

Hospital ER Office Bloody Diarrhea HUS Lab E. coli O157:H7 Signal Non O157:H7 STEC State CDC 1/494 episodes + for STEC -Vernacchio, et al, Pediatr Infect Dis J 2006; PID J 25:2 and JPGN 2006; 43:52.

Hospital ER Office Bloody Diarrhea HUS Lab E. coli O157:H7 Signal Non O157:H7 STEC State CDC 0/225 children with acute diarrhea -Denno, et al, Pediatr Infect Dis J 2005; 24:142

Hospital ER Office Bloody Diarrhea HUS Lab E. coli O157:H7 Signal Non O157:H7 STEC State CDC 15/1926 -Klein, et al, J Pediatr 2002; 141:172 -Klein, et al, unpublished

EIA vs. SMAC, point of care, ER Stools 39 signals in Meridian EIA (broth) Klein, E, et al, J Peds 2002; 172

39 EIA positives 25 E. coli O157:H7 10 non-O157:H7 STEC 1 non-O157:H7 STEC + C. jejuni 3 no STEC found 3 more stools + for E. coli O157:H7, not detected by toxin testing

39 EIA positives, 11 non-O157:H7 STEC O103:H2 (4) O118:H16 (2) O26:H11 (1) O111:nm (1) O111:H8 (1) O121:H19 (1) Orough:H11 (1)

39 STECs non-O157 (11) O157 (28) Percent of all: 1.7% 0.7% HUS0% 18% Bloody 50% 92% Laboratory blood 22% 70%

Seattle STEC, E. coli O157:H7: 11 non O157:H7: 4 (O26, O111, O121, O177) Klein, et al, unpublished data

Hospital ER Office Bloody Diarrhea HUS Lab E. coli O157:H7 Signal Non O157:H7 STEC State CDC Michigan: Shannon, et al, Emerg Infect Dis 2007; 13:318

Michigan 7 STEC from bloody stools, 6 were E. coli O157:H7 177 E. coli O157:H7 18 non-O157 STEC (O45, O103) 5/177 E. coli O157 not detected by EIA

Pitfalls of focusing on bloody diarrhea Laboratory can’t tell what specimens are really bloody Many (perhaps most) non-O157:H7 STEC associated with non-bloody diarrhea

Hospital ER Office Bloody Diarrhea HUS Lab E. coli O157:H7 Signal Non O157:H7 STEC State CDC

Hospital ER Office Bloody Diarrhea HUS E. coli O157:H7 is predominant (> 95%), cause of post-diarrheal HUS in USA, Canada, Japan, UK, and South America. 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 Mar 5 (epub)1-7 Pre-HUS cultures have highest yield Antibodies to O157 often present when culture is negative, but short- lived (weeks to months) Absence of proof is not proof of absence!

Hospital ER Office Bloody Diarrhea HUS But, HUS offers the best chance to find pathogenic non-O157:H7 STEC (1-5%, vs. <1% in other series)

Hospital ER Office Bloody Diarrhea HUS Lab E. coli O157:H7 Signal Non O157:H7 STEC State CDC

Five LFs, Seattle children, stools probed (colony hybridization) 13 E. coli O157:H7 (all bloody diarrhea), 9 detected by hybridization 5 non-O157:H7 STEC, only 1 had bloody diarrhea, but 3 were hospitalized Serotypes: O26 (2), O85, O103, O111 Bokete TN, et. al. Gastroenterology 1993;

Other 1990s series Falls Church, VA 6 O157:H7 5 non-O157:H7 STEC (2 with Salmonella) Milwaukee, WI 12 O157:H7 3 non-O157:H7 STEC Park, et al, Diagn Microbiol Infect Dis 1996;26:69 Kehl, et al, J Clin Microbiol 1997; 35:2051

St. Louis Children’s Hospital O157:H7 (33) Non-O157:H7 (13) O103:H2(6), O26, O111:NM, O145, O165, O174:H21 (1) No STEC (2) HUS E. coli O157:H7 missed by toxin assay

Falls Church, Atlanta, Salt Lake City July 2005 – September specimens 19 E. coli O157:H7 8 non-O157:H7 STEC Teel, et al, J Clin Microbiol 2007; 45:3377

Hospital ER Office Bloody Diarrhea HUS Lab E. coli O157:H7 Signal Non O157:H7 STEC State CDC

Montana STEC, 1998 – 2000 E. coli O157:H7 (31)non O157:H7 (50) Bloody81%57% ER56%28% Procedure22%16% O26 (20), O73 (1), O103 (2), O118 (1), O121 (15), O145 (1), O165 (1), O177 (1), O181 (1), Orough, ONT (7) Jelacic J, et. al. J Infect Dis, 2003;

Connecticut O157: 278 Non-O157: 125 (60% of toxin + assays yielded non-O157:H7 STEC) O103 (26) O111 (26) O26 (18) O45 (18) 15 other serogroups MMWR 2007;56:29

Connecticut HUS not defined, but: O157:H7Other STEC Bloody diarrhea90%56% Hospitalized45%12% HUS9%0%

Nebraska 7 non-O157:H7 STEC O26 (2) O103 (1) O111 (2) O145, Orough 6 O157:H7 HUS rate not given 1 of 6 O157:H7s not detected by EIA Fey, et al, Emerg Infect Dis 2000: 6:530

HUS (ca. 100 cases) Culture + rate for E. coli O157:H7 in past 15 years (Seattle and St. Louis): ~ 90% positive ~ 5% highly suspect to be positive ~ 4% nothing found

Summary Human exposure to non-O157:H7 STEC probably common, association with disease is rare Human exposure to O157:H7 STEC less common, burden of disease greater

Summary Human diagnostic resources must still focus on E. coli O157:H7 in USA, in children Some non-O157:H7 serotype predominance in USA: O26, O103, O111, O118, O121, O145 Be on the lookout for O113 – potentially quite virulent – in Canada Sorbitol fermenting O157:H - not yet in USA

Questions What is HUS rate for individual serotypes? (series might be biased by focusing on event of HUS) What is source of pathogenic non- O157:H7 STEC?

How should we find these agents? Toxin assays – OK clinically, much background in other settings Stx2 – but Stx1+/Stx2- strains have caused disease eae – must include different alleles, and O113 won’t be detected O antigen target (side chain or rfb loci) for O26, O103, O111, O113, O118, O121, O145