Joint EFSA/ECDC technical report: Shiga toxin/verotoxin‐producing Escherichia coli in humans, food and animals in the EU/EEA, with special reference to the German outbreak strain STEC O104

SUMMARYIn August 2015, Public Health England detected an outbreak of Shiga toxin-producing Escherichia coli (STEC) serotype O157:H7 caused by contaminated salad leaves in a mixed leaf prepacked salad product from a national retailer. The implicated leaves were cultivated at five different farms and the zoonotic source of the outbreak strain was not determined. In March 2016, additional isolates from new cases were identified that shared a recent common ancestor with the outbreak strain. A case–case study involving the cases identified in 2016 revealed that ovine exposures were associated with illness (n = 16; AOR 8·24; 95% CI 1·55–39·74). By mapping the recent movement of sheep and lambs across the United Kingdom, epidemiological links were established between the cases reporting ovine exposures. Given the close phylogenetic relationship between the outbreak strain and the isolates from cases with ovine exposures, it is plausible that ovine faeces may have contaminated the salad leaves via untreated irrigation water or run-off from fields nearby. Timely and targeted veterinary and environmental sampling should be considered during foodborne outbreaks of STEC, particularly where ready to eat vegetables and salads are implicated.

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Outbreak of Shiga Toxin–Producing Escherichia coli (STEC) O104:H4 Infection in Germany Causes a Paradigm Shift with Regard to Human Pathogenicity of STEC Strains

Journal of Food Protection ◽

10.4315/0362-028x.jfp-11-452 ◽

2012 ◽

Vol 75 (2) ◽

pp. 408-418 ◽

Cited By ~ 151

Author(s):

LOTHAR BEUTIN ◽

ANNETT MARTIN

Keyword(s):

Escherichia Coli ◽

Hemolytic Uremic Syndrome ◽

Shiga Toxin ◽

Outbreak Strain ◽

Fenugreek Seeds ◽

E Coli ◽

Aggregative Adherence ◽

Uremic Syndrome ◽

Enterocyte Effacement ◽

The Way

An outbreak that comprised 3,842 cases of human infections with enteroaggregative hemorrhagic Escherichia coli (EAHEC) O104:H4 occurred in Germany in May 2011. The high proportion of adults affected in this outbreak and the unusually high number of patients that developed hemolytic uremic syndrome makes this outbreak the most dramatic since enterohemorrhagic E. coli (EHEC) strains were first identified as agents of human disease. The characteristics of the outbreak strain, the way it spread among humans, and the clinical signs resulting from EAHEC infections have changed the way Shiga toxin–producing E. coli strains are regarded as human pathogens in general. EAHEC O104:H4 is an emerging E. coli pathotype that is endemic in Central Africa and has spread to Europe and Asia. EAHEC strains have evolved from enteroaggregative E. coli by uptake of a Shiga toxin 2a (Stx2a)–encoding bacteriophage. Except for Stx2a, no other EHEC-specific virulence markers including the locus of enterocyte effacement are present in EAHEC strains. EAHEC O104:H4 colonizes humans through aggregative adherence fimbrial pili encoded by the enteroaggregative E. coli plasmid. The aggregative adherence fimbrial colonization mechanism substitutes for the locus of enterocyte effacement functions for bacterial adherence and delivery of Stx2a into the human intestine, resulting clinically in hemolytic uremic syndrome. Humans are the only known natural reservoir known for EAHEC. In contrast, Shiga toxin–producing E. coli and EHEC are associated with animals as natural hosts. Contaminated sprouted fenugreek seeds were suspected as the primary vehicle of transmission of the EAHEC O104:H4 outbreak strain in Germany. During the outbreak, secondary transmission (human to human and human to food) was important. Epidemiological investigations revealed fenugreek seeds as the source of entry of EAHEC O104:H4 into the food chain; however, microbiological analysis of seeds for this pathogen produced negative results. The survival of EAHEC in seeds and the frequency of human carriers of EAHEC should be investigated for a better understanding of EAHEC transmission routes.

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Effects of Antibiotics on Shiga Toxin 2 Production and Bacteriophage Induction by Epidemic Escherichia coli O104:H4 Strain

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.06315-11 ◽

2012 ◽

Vol 56 (6) ◽

pp. 3277-3282 ◽

Cited By ~ 112

Author(s):

Martina Bielaszewska ◽

Evgeny A. Idelevich ◽

Wenlan Zhang ◽

Andreas Bauwens ◽

Frieder Schaumburg ◽

...

Keyword(s):

Escherichia Coli ◽

Antibiotic Therapy ◽

Shiga Toxin ◽

The Other ◽

Emerging Pathogen ◽

Outbreak Strain ◽

Content Type ◽

Shiga Toxin 2 ◽

Uremic Syndrome

ABSTRACTThe role of antibiotics in treatment of enterohemorrhagicEscherichia coli(EHEC) infections is controversial because of concerns about triggering hemolytic-uremic syndrome (HUS) by increasing Shiga toxin (Stx) production. During the recent large EHEC O104:H4 outbreak, antibiotic therapy was indicated for some patients. We tested a diverse panel of antibiotics to which the outbreak strain is susceptible to interrogate the effects of subinhibitory antibiotic concentrations on induction ofstx2-harboring bacteriophages,stx2transcription, and Stx2 production in this emerging pathogen. Ciprofloxacin significantly increasedstx2-harboring phage induction and Stx2 production in outbreak isolates (Pvalues of <0.001 to <0.05), while fosfomycin, gentamicin, and kanamycin insignificantly influenced them (P> 0.1) and chloramphenicol, meropenem, azithromycin, rifaximin, and tigecycline significantly decreased them (P≤ 0.05). Ciprofloxacin and chloramphenicol significantly upregulated and downregulatedstx2transcription, respectively (P< 0.01); the other antibiotics had insignificant effects (P> 0.1). Meropenem, azithromycin, and rifaximin, which were used for necessary therapeutic or prophylactic interventions during the EHEC O104:H4 outbreak, as well as tigecycline, neither inducedstx2-harboring phages nor increasedstx2transcription or Stx2 production in the outbreak strain. These antibiotics might represent therapeutic options for patients with EHEC O104:H4 infection if antibiotic treatment is inevitable. We await further analysis of the epidemic to determine if usage of these agents was associated with an altered risk of developing HUS.

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Enhanced Virulence of the Escherichia coli O157:H7 Spinach-Associated Outbreak Strain in Two Animal Models Is Associated with Higher Levels of Stx2 Production after Induction with Ciprofloxacin

Infection and Immunity ◽

10.1128/iai.02361-14 ◽

2014 ◽

Vol 82 (12) ◽

pp. 4968-4977 ◽

Cited By ~ 14

Author(s):

T. Zangari ◽

A. R. Melton-Celsa ◽

A. Panda ◽

M. A. Smith ◽

I. Tatarov ◽

...

Keyword(s):

Escherichia Coli ◽

Animal Models ◽

Shiga Toxin ◽

Hemorrhagic Colitis ◽

Escherichia Coli O157 ◽

Outbreak Strain ◽

Content Type ◽

Intestinal Pathology ◽

Phage Dna ◽

Uremic Syndrome

ABSTRACTShiga toxin (Stx)-producingEscherichia coli(STEC) causes hemorrhagic colitis and the hemolytic-uremic syndrome (HUS). STEC strains may produce Stx1a and/or Stx2a or variants of either toxin. A 2006 spinach-associated outbreak of STEC O157:H7 resulted in higher hospitalization and HUS rates than previous STEC outbreaks. The spinach isolate, strain K3995, contains bothstx2aandstx2c. We hypothesized that the enhanced virulence of K3995 reflects the combination ofstx2alleles (carried on lysogenic phages) and/or the amount of Stx2 made by that strain. We compared the virulence of K3995 to those of other O157:H7 isolates and an isogenic Stx2 mutant in rabbits and mice. We also measured the relative levels of Stx2 produced from those strains with or without induction of thestx-carrying phage. Some rabbits infected with K3995 exhibited intestinal pathology and succumbed to infection, while none of those infected with O157:H7 strain 2812 (Stx1a+Stx2a+) died or showed pathological signs. Rabbits infected with the isogenic Stx2a mutant K3995stx2a::catwere not colonized as well as those infected with K3995 and exhibited no signs of disease. In the streptomycin-treated mouse model, more animals infected with K3995 died than did those infected with O157:H7 strain 86-24 (Stx2a+). Additionally, K3995 produced higher levels of total Stx2 and toxin phage DNA in cultures after phage induction than did 86-24. Our results demonstrate the greater virulence of K3995 compared to other O157:H7 strains in rabbits and mice. We conclude that this enhanced virulence is linked to higher levels of Stx2 expression as a consequence of increased phage induction.

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Outbreak of haemolytic uraemic syndrome due to Shiga toxin-producing Escherichia coli O104:H4 among French tourists returning from Turkey, September 2011

Eurosurveillance ◽

10.2807/ese.17.04.20065-en ◽

2012 ◽

Vol 17 (4) ◽

Cited By ~ 15

Author(s):

N Jourdan-da Silva ◽

M Watrin ◽

F X Weill ◽

L A King ◽

M Gouali ◽

...

Keyword(s):

Escherichia Coli ◽

Shiga Toxin ◽

Haemolytic Uraemic Syndrome ◽

Outbreak Strain ◽

Molecular Analyses ◽

E Coli ◽

Source Of Infection

Eight cases of diarrhoea, including two cases of haemolytic uraemic syndrome (HUS), were identified among 22 French tourists who travelled to Turkey in September 2011. A strain of Escherichia coli O104:H4 stx2-positive, eae-negative, hlyA-negative, aggR-positive, ESBL-negative was isolated from one HUS case. Molecular analyses show this strain to be genetically similar but not indistinguishable from the E. coli O104:H4 2011 outbreak strain of France and Germany. Although the source of infection was not identified, we conclude that the HUS cases had probably been infected in Turkey.

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A Comparison of Shiga-Toxin 2 Bacteriophage from Classical Enterohemorrhagic Escherichia coli Serotypes and the German E. coli O104:H4 Outbreak Strain

PLoS ONE ◽

10.1371/journal.pone.0037362 ◽

2012 ◽

Vol 7 (5) ◽

pp. e37362 ◽

Cited By ~ 40

Author(s):

Chad R. Laing ◽

Yongxiang Zhang ◽

Matthew W. Gilmour ◽

Vanessa Allen ◽

Roger Johnson ◽

...

Keyword(s):

Escherichia Coli ◽

Shiga Toxin ◽

Enterohemorrhagic Escherichia Coli ◽

Outbreak Strain ◽

E Coli ◽

Shiga Toxin 2

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First description of CTX-M-3 extended-spectrum β-lactamase in an outbreak strain of Shiga toxin-producing Escherichia coli O103:H2

International Journal of Antimicrobial Agents ◽

10.1016/j.ijantimicag.2015.12.014 ◽

2016 ◽

Vol 47 (3) ◽

pp. 244-245 ◽

Cited By ~ 2

Author(s):

Jin Seok Kim ◽

Min Ji Kim ◽

Soo Jin Kim ◽

Eunkyung Shin ◽

Kyung-Hwan Oh ◽

...

Keyword(s):

Escherichia Coli ◽

Shiga Toxin ◽

Outbreak Strain ◽

Extended Spectrum

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Comparative Genomics of Recent Shiga Toxin-Producing Escherichia coli O104:H4: Short-Term Evolution of an Emerging Pathogen

mBio ◽

10.1128/mbio.00452-12 ◽

2013 ◽

Vol 4 (1) ◽

Cited By ~ 37

Author(s):

Yonatan H. Grad ◽

Paul Godfrey ◽

Gustavo C. Cerquiera ◽

Patricia Mariani-Kurkdjian ◽

Malika Gouali ◽

...

Keyword(s):

Escherichia Coli ◽

Shiga Toxin ◽

Genomic Islands ◽

Short Time Scale ◽

Short Term ◽

Outbreak Strain ◽

Content Type ◽

E Coli ◽

Term Evolution ◽

Large Outbreak

ABSTRACTThe large outbreak of diarrhea and hemolytic uremic syndrome (HUS) caused by Shiga toxin-producingEscherichia coliO104:H4 in Europe from May to July 2011 highlighted the potential of a rarely identifiedE. coliserogroup to cause severe disease. Prior to the outbreak, there were very few reports of disease caused by this pathogen and thus little known of its diversity and evolution. The identification of cases of HUS caused byE. coliO104:H4 in France and Turkey after the outbreak and with no clear epidemiological links raises questions about whether these sporadic cases are derived from the outbreak. Here, we report genome sequences of five independent isolates from these cases and results of a comparative analysis with historical and 2011 outbreak isolates. These analyses revealed that the five isolates are not derived from the outbreak strain; however, they are more closely related to the outbreak strain and each other than to isolates identified prior to the 2011 outbreak. Over the short time scale represented by these closely related organisms, the majority of genome variation is found within their mobile genetic elements: none of the nine O104:H4 isolates compared here contain the same set of plasmids, and their prophages and genomic islands also differ. Moreover, the presence of closely related HUS-associatedE. coliO104:H4 isolates supports the contention that fully virulent O104:H4 isolates are widespread and emphasizes the possibility of future food-borneE. coliO104:H4 outbreaks.IMPORTANCEIn the summer of 2011, a large outbreak of bloody diarrhea with a high rate of severe complications took place in Europe, caused by a previously rarely seenEscherichia colistrain of serogroup O104:H4. Identification of subsequent infections caused byE. coliO104:H4 raised questions about whether these new cases represented ongoing transmission of the outbreak strain. In this study, we sequenced the genomes of isolates from five recent cases and compared them with historical isolates. The analyses reveal that, in the very short term, evolution of the bacterial genome takes place in parts of the genome that are exchanged among bacteria, and these regions contain genes involved in adaptation to local environments. We show that these recent isolates are not derived from the outbreak strain but are very closely related and share many of the same disease-causing genes, emphasizing the concern that these bacteria may cause future severe outbreaks.

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Nitric Oxide Induced stx2 Expression Is Inhibited by the Nitric Oxide Reductase, NorV, in a Clade 8 Escherichia coli O157:H7 Outbreak Strain

Microorganisms ◽

10.3390/microorganisms10010106 ◽

2022 ◽

Vol 10 (1) ◽

pp. 106

Author(s):

Rim Al Safadi ◽

Michelle L. Korir ◽

Shannon D. Manning

Keyword(s):

Nitric Oxide ◽

Escherichia Coli ◽

Shiga Toxin ◽

Toxin Production ◽

Anaerobic Growth ◽

Escherichia Coli O157 ◽

Similar Increase ◽

Nitric Oxide Reductase ◽

Outbreak Strain ◽

Uremic Syndrome

Escherichia coli O157:H7 pathogenesis is due to Shiga toxin (Stx) production, though variation in virulence has been observed. Clade 8 strains, for instance, were shown to overproduce Stx and were more common among hemolytic uremic syndrome cases. One candidate gene, norV, which encodes a nitric oxide (NO) reductase found in a clade 8 O157:H7 outbreak strain (TW14359), was thought to impact virulence. Hence, we screened for norV in 303 O157 isolates representing multiple clades, examined stx2 expression following NO exposure in TW14359 for comparison to an isogenic mutant (ΔnorV), and evaluated survival in THP-1 derived macrophages. norV was intact in strains representing clades 6–9, whereas a 204 bp deletion was found in clades 2 and 3. During anaerobic growth, NO induced stx2 expression in TW14359. A similar increase in stx2 expression was observed for the ΔnorV mutant in anaerobiosis, though it was not impaired in its ability to survive within macrophages relative to TW14359. Altogether, these data suggest that NO enhances virulence by inducing Stx2 production in TW14359, and that toxin production is inhibited by NorV encoded by a gene found in most clade 8 strains. The mechanism linked to these responses, however, remains unclear and likely varies across genotypes.

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Characteristics of the enteroaggregative Shiga toxin/verotoxin-producing Escherichia coli O104:H4 strain causing the outbreak of haemolytic uraemic syndrome in Germany, May to June 2011

Eurosurveillance ◽

10.2807/ese.16.24.19889-en ◽

2011 ◽

Vol 16 (24) ◽

Cited By ~ 144

Author(s):

F Scheutz ◽

E Møller Nielsen ◽

J Frimodt-Møller ◽

N Boisen ◽

S Morabito ◽

...

Keyword(s):

Escherichia Coli ◽

Shiga Toxin ◽

Haemolytic Uraemic Syndrome ◽

Coli Strain ◽

Screening Tools ◽

International Exchange ◽

Outbreak Strain ◽

Diagnostic Screening ◽

E Coli ◽

Large Outbreak

The Escherichia coli strain causing a large outbreak of haemolytic uraemic syndrome and bloody diarrhoea in Germany in May and June 2011 possesses an unusual combination of pathogenic features typical of enteroaggregative E. coli together with the capacity to produce Shiga toxin. Through rapid national and international exchange of information and strains the known occurrence in humans was quickly assessed. We describe simple diagnostic screening tools to detect the outbreak strain in clinical specimens and a novel real-time PCR for its detection in foods.

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