Non-O157 Shiga toxin-producing E. coli: An emerging pathogen of public health importance

Transcription

1 Non-O157 Shiga toxin-producing E. coli: An emerging pathogen of public health importance Public Health Ontario Grand Rounds June 17, 2014 Vanessa G. Allen MD MPH

2 Objectives Outline the microbiology and clinical features of non- O157 Shiga toxin-producing E. coli (STEC) disease Describe the epidemiology of STEC infection Discuss the laboratory tools for the diagnosis of STEC Review different approaches to non-o157 STEC detection and response

3 THE MICROBIOLOGY & CLINICAL FEATURES OF SHIGA-TOXIN PRODUCING E. COLI 3

4 Shiga-toxin Producing E. coli (STEC) Otherwise known as verotoxin producing E. coli A subset of these are enterohemorrhagic E. coli (EHEC) in virtue of additional virulence factors on the locus of enterocyte effacement (LEE) Typing based on somatic (O) and flagellar (H) antigens O157:H7 most commonly identified Sorbitol negative, so easily detected within the laboratory Likely associated with more severe disease Figure from: Mandell, Douglas, and Bennett s Principles and Practice of Infectious Diseases seventh edition, 2010.

5 Virulence Factors Associated with Shiga-toxin Producing E. coli Shiga toxins Stx1 is virtually identical to Shigella dysenteriae toxin type 1 Stx2 similar genetically and functionally (55-57% homology with stx1) STEC may have stx1,stx2, or both Shiga-toxins bind to globotriaosylceramide of endothelial cells and neural cells inhibiting protein synthesis via action on 28S of 60S subunit of the ribosome Sixma TK, et al. J Mol Biol 1993;230:

6 Subset of STEC with Enterohemorrhagic Properties Locus of enterocyte effacement (LEE) 35kb-gene cluster, including the initimin or eae gene Causes effacement of intestinal microvilli Adherence of the bacteria to the epithelial cells Damage to host cell cytoskeleton Perna NT, et al. Infect Immun. 1998;66:

7 Incubation period 3-4 days (1-10 days) Infective dose unknown Likely CFU Incubation Periods and Transmission of STEC Transmission Fecal-oral Less commonly secondary person to person transmission Paton JC et al. Clin Microbiol Review : Karch, H., et al J. Clin. Microbiol. 33:

8 Natural Reservoir of STEC Gastrointestinal tract of cattle and other large herbivorous mammals 46% of 1458 cattle on 100% of 335 farms had VTEC in fecal samples Also seen in sheep, goats, deer Not known to be present in swine or chicken Transmitted directly to humans via fecal oral route Undercooked meat Dairy products Produce Contact with animals Wilson, J. Bet al J. Infect. Dis. 174:

9 Secondary Transmission of STEC Outbreak in Wales, Sept confirmed and probable cases of E. coli O157 Contaminated deli meat distributed to 46 schools Secondary transmission occurred in 22% of 89 households Highest risk if index case < 5 years old (RR 2.08) and has sibling (RR 3.8) HUS in 4/107 siblings NNT (by isolation to prevent 1 case of HUS) = 47 Werber D, et al. Clin Infect Dis Apr 15;46(8):

10 Clinical Spectrum of Disease Caused by STEC Asymptomatic 6% carriage among Canadian dairy farmers Watery diarrhea Abdominal cramps Fever is usually absent Hemorrhagic colitis, 1-2 days later Hemolytic uremic syndrome (6.3-8%) Triad: renal failure, thrombocytopenia and microangiopathic hemolytic anemia Neurological symptoms Wilson, J. Bet al J. Infect. Dis. 174:

11 Hemolytic Uremic Syndrome Occurs in ~5-10% of children with O157 12% death or end-stage renal disease STEC subtypes and HUS Children from Germany and Austria, 207 isolates 57% O157:H7 43% other subtypes O26:H11/H (15%) Sorbitol-fermenting O157:H (10%) O145:H28/H (9%) O103:H2/H (3%) O111:H8/H (3%) Other (3%) Griffin, P. M., et al Epidemiol. Rev. 13:60-9 Gerber A, et al J Infect Dis 186:

12 E. coli O157 vs E. coli non O157: Differential Presence of Stx2 12

13 Association of Stx2 with HUS Byrne L et al. JMM. June

14 Association of Stx2 with HUS and Bloody diarrhea HUS stx1 stx2 stx1 & stx2 eae O111 (10) 1 (10) 0 (0) 9 (90) 9/9 (100) Other (11) 0 (0) 8 (73) 3 (27) 7 (64) Bloody diarrhea O26 (12) 10 (83) 0 (0) 2 (17) 12 (100) O111 (8) 4 (50) 0 (0) 4 (50) 7 (88) O121 (13) 0 (0) 13 (100) 0 (0) 12 (92) O45 (10) 10 (100) 0 (0) 0 (0) 10 (100) J.T. Brooks, et al. J. Infect. Dis. 192 (2005), pp

15 Stx Negative E coli O157:H7 Germany , 611 stool specimens 471 stx positive O stx negative O157 HUS occurred with and without stx Serotype Total # of Strains stx eae HUS Bloody diarrhea Diarrhea O O O157:H O157:H Friedrick AW et al. Clin Infect Dis Jul 1;45(1):39-45.

16 Long Term Sequelae of HUS associated with STEC Infection Five years post Walkerton 19 HUS vs 24 controls Increased microalbuminemia and slightly decreased renal function. Meta-analysis of forty-nine studies of 3476 patients with a mean follow-up of 4.4 years ESRD 12% (0-30%) Proteinuria, hypertension and GFR < 80 ml/min 25% (0-64%) Garg AX, et al. JAMA. 2003;290: Garg AX, et al. Am J Kidney Dis Mar;51(3):

17 Treatment of STEC Supportive Fluids, hemodialysis Antibiotics not recommended Induction of verotoxin release in vitro Increased severity of illness in mouse models Associated with increased HUS in children 71 children 9 (13%) received antibiotics 10 developed HUS 5/10 received abx RR 17.3 on multivariate analysis controlling for duration of illness and WBC Zhang XP, et al. J Infect Dis. 2000;181: Matsushiro A, et al. J Bacteriol. 1999;181: Wong CS, et al. N Engl J Med. 2000;342:

18 Wong CS, et al. N Engl J Med. 2000;342:

19 Meta-analysis of Effect of Antibiotics in E. coli O157 Infection Conclusion: No significant increased risk of HUS Safdar N et al. JAMA

20 Complement Blockers for STEC associated HUS Use of monoclonal C5 antibody, eculizumab Delmas Y, et al. Nephrol Dial Transplant 2014 Lapeyraque A-L. et al. NEJM

21 EPIDEMIOLOGY OF SHIGA-TOXIN PRODUCING E. COLI 21

22 Disease Caused by non -O157 STEC Retrospective US survey Most common is E. coli O157 Estimated at 2X of all other STEC combined More likely associated with bloody diarrhea O26 O111 O103 O121 O45 O165 O118 O91 O153 O146 O174 Other Undetermined J.T. Brooks, et al. J. Infect. Dis. 192 (2005), pp

23 Incidence of E coli O157 vs non O157 STEC Ruth Luna 14 th Annual PulseNet Meeting,

24 Epidemiology United States 44% of STEC in the U.S. were caused by non-o157 strains. STEC is under-reported A 2011 study by the CDC determined that the under-diagnosis multiplier for STEC O157 was under-diagnosed by a factor of 26 whereas the multiplier for non-o157 STEC was 107 The resulting estimated burden for non-o157 STEC that was 1.8 times that of STEC O157 (112,752 versus 63,153, respectively) Using data from the U.S., the annual burden of foodborne diseases due to non-o157 STEC was estimated at 20,523 compared to 12,827 for O157 STEC. Johnson K et al. Clin Infect Dis 2006 Dec 15; 43(12): Scallan, E et al. Emerging Infectious Diseases Jan; 17(2):7-15. Marks, HM et al J. of Food Protec Jan 21;76(6):

25 STEC: Incidence of E coli O157 vs non O157 Serotypes KE Johnson et al, CID

26 Epidemiology of STEC in Ontario 2005 to 2012 An annual average of 246 cases of STEC infection, including HUS range 153 to 344 cases per year Corresponding incidence rates ranged from 1.2 to 2.7 cases per 100,000 On average, ~ 2% of cases reported HUS reported as a complication of STEC Non-O157 STEC: 1-7 cases per year since % of STEC reported One case of HUS as a complication of non-o157 STEC cases The incidence of STEC decreased with increasing age for both males and females with the highest rates occurring in those under the age of 10 years Courtesy of Stephen Moore, Yvonne Whitfield, Lisa Fortuna and Doug Sider 26

27 Study of STEC Prevalence in Ontario Screening of anonymized routine stool samples from 2 hospitals (one rural and one urban), and one private laboratory May-December 2011 May-August 2012 Year (Positive/total) Prevalence O157 Non-O (28/1244) (42%) 12 (58%) 2012 (16/2948) (9%) 10 (91%) Sandra Zittermann et al. In preparation 27

28 Virulence factors of STEC Ontario isolates, O serotype stx1 stx2 stx1 + stx2 eae hly Sorbitol fermentation Total strains O157:H O O103:H O Rough O111:HNM O146:H UND O45:H O1:H O40:H O153:H O28ab:H O5:H NM Total O26 serogroup were O26:H11 (3), O26:H UND, O26:H21. O Rough were O Rough:H21, O Rough:H UND, O Rough:H45 and O Rough: H4. Sandra Zittermann et al. In preparation 28

29 Differential Disease Severity of E. coli O157 and E. coli non O157 Wang X et al. Can J Infect Dis Med Microbiol

30 Risk Factors and Incidence Associated with Non O157 STEC in British Columbia, Wang X et al. Can J Infect Dis Med Microbiol

31 Serotype Distribution of STEC in British Columbia Wang X et al. Can J Infect Dis Med Microbiol

32 EXAMPLES OF IMPORTANT OUTBREAKS CAUSED BY NON-O157 STEC 32

33 Distribution of Cases of HUS in Germany Frank C. et al. NEJM June 22,

34 -C Frank et al. Eurosurveillance. May 26,

35 Epidemiological Curve of the German E coli O104:H4 Outbreak Affecting mostly adults (89%) and women (68.0%) HUS developed in 25.1% of all cases Frank C. Et al NEJM June 22,

36 E coli O104 Confirmed by PCR in Fenugreek Seeds 36

37 Multistate Outbreak of Shiga toxinproducing Escherichia coli O121 Infections Linked to Raw Clover Sprouts, May- June people infected so far, 47% hospitalized, no HUS or deaths 37

38 Increased Detection of Outbreaks with Increased Routine Screening Luna-Gierke RE et al. Epidemiology Infection

39 Details of 46 Outbreaks of non O157 STEC Reported to the CDC (up to 2010) 28% in childcare settings 7% of outbreak patients with stx2+ had HUS, compared to 0.8% in outbreaks that were stx2 negative Luna-Gierke RE et al. Epidemiology Infection

40 E. coli O157 outbreaks in the US (for comparison 350 outbreaks, rate of HUS 1-18% Rangel JM et al. EID

41 INTERMISSION QUESTION 41

42 Question: What organisms are routinely tested for in bacterial stool cultures? Choose one or more 1) Shigella spp. 2) Clostridium perfringens 3) Shiga-toxin producing E. coli 4) Salmonella spp. 5) Yersinia spp. 6) Norovirus Are there any more that are routinely performed?

43 Answer: Organisms are routinely tested for in bacterial cultures of stool Red highlighted are correct answers: 1) Shigella spp. 2) Clostridium perfringens* 3) All verotoxin producing E coli* 4) Salmonella spp. 5) Yersinia spp. 6) Norovirus* * Clostridium perfringens, non-o157 shiga-toxin producing E coli and norovirus tested during outbreaks at PHO Laboratories

44 Performed as part of GI outbreak if samples submitted Answer: Organisms are routinely tested for in bacterial cultures of stool? Typically routine stool cultures are screened for: Salmonella Shigella E. coli O157 Campylobacter Yersinia Others that can be requested include Clostridium difficile Clostridium perfringens Non O157 shiga-toxin producing E. coli Vibrio Aeromonas, Pleisiomonas norovirus PCR parasites NB. These may require different transport media and different culture plates

45 LABORATORY TOOLS FOR THE DETECTION OF NON-O157 STEC 45

46 Laboratory Testing for Non O157 STEC All stools submitted for bacterial culture to Ontario laboratories are tested for E. coli O157 Testing for non-o157 VTEC is not routinely conducted at hospital or private labs in Ontario. In a 2002 survey of Hamilton area physicians the National Studies on Acute Gastrointestinal Illness found that 44.3% of physicians believed that stool samples sent for routine culture would always be tested for non-o157 E coli. 46

47 Options for Testing for Non-O157 STEC Culture Screen: Too cumbersome for routine screening for non O157 Confirmation: Needed for serotyping and molecular fingerprinting Vero cell assay Time consuming and requires considerable expertise ELISAs Several assays available on the market No comprehensive comparative data Used routinely for outbreak specimens at PHO-Laboratories since 2008 PCR Monoplex and multiplex All options costly 47

48 Laboratory Diagnosis of STEC: Culture High burden of organisms early in disease Up to 90% of organisms cultured in stool Isolation procedures for O157 Sorbitol-MacConkey agar (sorbitol non-fermenters) D-glucuronidase negative Cefixime and rhamnose additives Colourless colonies tested with slide or tube agglutination with O157 and H7 antisera or commercial latex reagents Confirmation of toxin production Can be enhanced by immunomagnetic separation Paton JC et al. Clin Microbiol Review : Karch, H., et al J. Clin. Microbiol. 34:

49 Laboratory Diagnosis of STEC: Toxin Assays Tissue culture cytotoxicity assays Vero cells or HeLa cells Sterile filtrate incubated on monolayer for 48 to 72 hours Sensitivity increased by pretreatment with polymyxin Ideally neutralize with monoclonal antibodies EIA for stx1 and stx2 Sandwich technique Less sensitive than cytotoxic assays as screen Can be performed directly on stool specimen But early on can detect VTEC at levels > 1% of fecal flora

50 Molecular Methods for The Detection of non-o157 STEC PCR more sensitive than other assays 754 stool samples during the German outbreak of E. coli O for STEC by culture Only 86 + by EIA Commercial multiplex assays offer future possibilities for testing for multiple pathogens at the same time (including STEC) Current costs of assays are high Gerritzen A et al. Lancet ID

51 The Challenge of PCR Positive & Culture Negative Specimens Couturier, MR et al JCM stools submitted for enteric viruses from for stx1 or stx2 Fifteen has associated isolates, 20% E. coli O157 Decreased yield associated with freeze/thaw cycles and time (earlier years samples were culture negative) 51

52 Role of testing for STEC in HUS 52 patients with post diarrheal HUS , median age 42 months 94.2% had bloody stools 84.8% of those with confirmed E. coli O157 HUS diagnosed a median of 6.5 days (+/- 2.8d) after onset of diarrhea 13/ 52 tested within 2 days of diarrhea 100% positive for E. coli O157 Yield in the remaining was associated with timing of stool sample 91.7% positive for E. coli O157 if tested within 3-6 days of diarrhea 33.3% positive for E. coli O157 if tested => 7 days after onset of diarrhea Tarr PI et al. JID

53 Estimated Costs of Routine Screening of All Stool for Non O157 STEC in Ontario Routine testing for non-o157 E. coli in Ontario for all stools would require considerable investment The estimated rate of acute gastrointestinal illness (AGI) reported in Ontario is 1.2 episodes per person-year Of these, 22% report seeking health care Stool specimen kits were provided to 33% 80% of those with stool kits submitted specimens Estimated 871,200 stool tests performed each year in Ontario. The total estimated cost of testing all acute GI samples for non-o157 STEC would be approximately $8.712 million/year 53

54 APPROACHES FOR DETECTION OF NON-O157 STEC 54

55 Recommend routine testing all stools for STEC both by culture and by Shiga toxin detection Otherwise will miss cases and clusters No marker reliably predicts infection (bloody stools, leukocytosis) 55

56 PROS of Routine Screening 1) As frequent as other bacterial pathogen for which we test Foodnet 2009 data Incidence of O per 100,000; 459 cases Incidence of non-o per 100,000; 264 cases 2) Severe disease can ensue 300 patients, 21 (7%) HUS and 75 (26%) bloody diarrhea 3) selective screening results in missed cases of non-o157 STEC One study showed that 3/15 non O157 visibly bloody Marcon M et al. JCM

57 CONS of Universal Screening 1) Low prevalence disease 2) Costly 3) Little effect on treatment, procedures, secondary transmission 4) Effect of detection of more outbreaks is uncertain Marcon M et al. JCM

58 PROS of Selective Screening 1) Reduce false positives (given low prevalence disease) 50% PPV in one study 2) Balanced with routine screning of pathogens of equal or greater prevalence and severity Tailor to local prevalence If low prevalence, consider on request or selective criteria Possible criteria include: Bloody stool Age <= 5 or => 60 Summer months Hospitalized patients with HUS or compatible clinical history Suspect food Day care Known contact Known outbreak Marcon M et al. JCM

59 Response to CDC Guidelines for STEC Distribution of testing practices in Washington state laboratories, 2010 (n=57) Proportion of stool specimens in Washington state laboratories, 2010 Stigi KA et al. EID

60 Top 6 Serotypes of Non O157 STEC: Adulterants in Certain Food Products in the US Rolled out June 4, 2012 Serotypes included: O26 O45 O103 O111 O121 O145 ortal/fsis/topics/data-collectionandreports/microbiology/ec/testing -program-for-e-coli-o157h7-andnon-o157-stec 60

61 Other Strategies for STEC Testing South Australia (Vally H et al BMC Public Health 2012) STEC screening criteria evidence of blood in the stool a clinical history of blood in the stool requesting doctor queries STEC infection or HUS Similar strategy in BC (personal communication, Dr. Linda Hoang) 61

62 In Ontario, non O157 testing available for outbreak settings and on request at the Public Health Ontario Laboratories Strategy is currently under review 62

63 Approaches for Ontario STEC is a heterogeneous group of organisms with differential risks of severe disaese The likelihood of detection of sporadic cases and small outbreaks of non-o157 STEC in Ontario is currently very low given lack of routine testing The most comprehensive strategy to capture all non-o157 STEC cases and outbreaks is to add this screening to routine stool testing at all front line laboratories This comes at considerable cost Data on prevalence and disease severity and sequelae are lacking Selective criteria such as age, bloody diarrhea, and HUS may capture a subset of cases Ongoing work at Public Health Ontario to provide support to our colleagues for a provincial strategy 63

64 Conclusions STEC is a common foodborne pathogen with considerable morbidity and mortality Unlike E. coli O157, non-o157 E. coli serogroups of STEC are not routinely tested for in the laboratory Data of the current epidemiology and clinical burden of non-o157 STEC in Ontario is limited, but data suggests that many non-o157 STEC serotypes are associated with milder disease Overall, non O157 STEC are associated with decreased presence of virulence factors such as stx2 Heterogeneity of non O157 STEC serotypes Variable jurisdictional approaches have been taken to investigate outbreaks and sporadic disease If non-o157 STEC infection is suspected, samples can be forwarded to the Public Health Ontario Laboratories (PHOL) for testing HUS Hospitalized cases Severe or bloody diarrhea Outbreaks 64

65 Thanks to Sandra Zittermann Stephen Moore Doug Sider Anne Maki Analyn Peralta Yvonne Whitfield Lisa Fortuna Roger Johnson Linda Chui 65

66 THANK YOU. 66