Phylogenetic context of Shiga toxin-producing Escherichia coli serotype O26:H11 in England

The increasing use of PCR for the detection of gastrointestinal pathogens in hospital laboratories in England has improved the detection of Shiga toxin-producing Escherichia coli (STEC), and the diagnosis of haemolytic uraemic syndrome (HUS). We aimed to analyse the microbiological characteristics and phylogenetic relationships of STEC O26:H11, clonal complex (CC) 29, in England to inform surveillance, and to assess the threat to public health. There were 502 STEC belonging to CC29 isolated between 2014 and 2019, of which 416 were from individual cases. The majority of isolates belonged to one of three major sequence types (STs), ST16 (n=37), ST21 (n=350) and ST29 (n=24). ST16 and ST29 were mainly isolated from cases reporting recent travel abroad. Within ST21, there were three main clades associated with domestic acquisition. All three domestic clades had Shiga toxin subtype gene (stx) profiles associated with causing severe clinical outcomes including STEC-HUS, specifically either stx1a, stx2a or stx1a/stx2a. Isolates from the same patient, same household or same outbreak with an established source for the most part fell within 5-SNP single linkage clusters. There were 19 5-SNP community clusters, of which six were travel-associated and one was an outbreak of 16 cases caused by the consumption of contaminated salad leaves. Of the remaining 12 clusters, 9/12 were either temporally or geographically related or both. Exposure to foodborne STEC O26:H11 ST21 capable of causing severe clinical outcomes, including STEC-HUS, is an emerging risk to public health in England. The lack of comprehensive surveillance of this STEC serotype is a concern, and there is a need to expand the implementation of methods capable of detecting STEC in local hospital settings.

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Phylogenetic structure of Shiga toxin-producing Escherichia coli O157:H7 from sub-lineage to SNPs

Microbial Genomics ◽

10.1099/mgen.0.000544 ◽

2021 ◽

Author(s):

Timothy J. Dallman ◽

David R. Greig ◽

Saheer E. Gharbia ◽

Claire Jenkins

Keyword(s):

Escherichia Coli ◽

Shiga Toxin ◽

Sequence Similarity ◽

Evidence Base ◽

Point Sources ◽

Single Linkage ◽

Phylogenetic Structure ◽

Content Type ◽

Geographically Dispersed ◽

Cluster Threshold

Sequence similarity of pathogen genomes can infer the relatedness between isolates as the fewer genetic differences identified between pairs of isolates, the less time since divergence from a common ancestor. Clustering based on hierarchical single linkage clustering of pairwise SNP distances has been employed to detect and investigate outbreaks. Here, we evaluated the evidence-base for the interpretation of phylogenetic clusters of Shiga toxin-producing Escherichia coli (STEC) O157:H7. Whole genome sequences of 1193 isolates of STEC O157:H7 submitted to Public Health England between July 2015 and December 2016 were mapped to the Sakai reference strain. Hierarchical single linkage clustering was performed on the pairwise SNP difference between all isolates at descending distance thresholds. Cases with known epidemiological links fell within 5-SNP single linkage clusters. Five-SNP single linkage community clusters where an epidemiological link was not identified were more likely to be temporally and/or geographically related than sporadic cases. Ten-SNP single linkage clusters occurred infrequently and were challenging to investigate as cases were few, and temporally and/or geographically dispersed. A single linkage cluster threshold of 5-SNPs has utility for the detection of outbreaks linked to both persistent and point sources. Deeper phylogenetic analysis revealed that the distinction between domestic UK and imported isolates could be inferred at the sub-lineage level. Cases associated with domestically acquired infection that fall within clusters that are predominantly travel associated are likely to be caused by contaminated imported food.

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The emerging importance of Shiga toxin-producing Escherichia coli other than serogroup O157 in England

Journal of Medical Microbiology ◽

10.1099/jmm.0.001375 ◽

2021 ◽

Vol 70 (7) ◽

Author(s):

Bhavita Vishram ◽

Claire Jenkins ◽

David R. Greig ◽

Gauri Godbole ◽

Kevin Carroll ◽

...

Keyword(s):

Escherichia Coli ◽

Shiga Toxin ◽

Severe Disease ◽

Descriptive Analysis ◽

Fold Increase ◽

Surveillance Data ◽

Bloody Diarrhoea ◽

Content Type ◽

Emerging Risk ◽

Pcr Targeting

Introduction. Shiga toxin-producing Escherichia coli (STEC) can cause severe disease and large outbreaks. In England, the incidence and clinical significance of STEC serogroups other than O157 (non-O157) is unknown due to a testing bias for detection of STEC O157. Since 2013, the implementation of PCR to detect all STEC serogroups by an increasing number of diagnostic laboratories has led to an increase in the detection of non-O157 STEC. Hypothesis/Gap statement. Due to a bias in testing methodologies to select for STEC serogroup O157 in frontline diagnostic laboratories in most countries, very little surveillance data have been previously generated on non-O157 STEC. Aim. Five years (2014–2018) of STEC national surveillance data were extracted and descriptive analysis undertaken to assess disease severity of non-O157 STEC strains. Methods. Data from 1 January 2014 to 31 December 2018 were extracted from the National Enhanced Surveillance System for STEC and analysed. Results. The implementation of Gastrointestinal Polymerase Chain Reaction (GI-PCR) has resulted in a four-fold increase in the detection of non-O157 STEC cases between 2014 and 2018. There were 2579 cases infected with 97 different non-O157 serogroups. The gender distribution was similar amongst STEC O157 and non-O157 STEC cases with 57 and 56 % of cases being female respectively, but a significantly higher proportion of cases (P <0.001) under 5 years of age was observed among STEC O157 (22 %) cases compared to non-O157 STEC (14 %). The most common non-O157 serogroups were O26 (16 %), O146 (11 %), O91 (10 %), O128 (7 %), O103 (5 %) and O117 (3 %). Overall, rates of bloody diarrhoea were highest in O26 (44 %) and O103 (48 %) cases and lowest in STEC O117 cases (17 %). Strains harbouring Shiga toxin stx1a caused the highest proportion of diarrhoea (93 %) and caused the same level of bloody diarrhoea as stx2a (39 %). However, stx2a caused the highest proportion of vomiting (46 %), hospitalisation (49 %) and considerably more HUS (29 %) than other stx profiles. Conclusion. The implementation of PCR targeting stx at diagnostic laboratories has shown that non-O157 STEC, most notably STEC O26, are an emerging risk to public health.

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Metagenomics as a Public Health Risk Assessment Tool in a Study of Natural Creek Sediments Influenced by Agricultural and Livestock Runoff: Potential and Limitations

Applied and Environmental Microbiology ◽

10.1128/aem.02525-19 ◽

2020 ◽

Vol 86 (6) ◽

Cited By ~ 4

Author(s):

Brittany Suttner ◽

Eric R. Johnston ◽

Luis H. Orellana ◽

Luis M. Rodriguez-R ◽

Janet K. Hatt ◽

...

Keyword(s):

Public Health ◽

Escherichia Coli ◽

Antibiotic Resistance ◽

Health Risk ◽

Resistance Genes ◽

Shiga Toxin ◽

Antibiotic Resistance Genes ◽

Agricultural Runoff ◽

Public Health Risk ◽

Content Type

ABSTRACT Little is known about the public health risks associated with natural creek sediments that are affected by runoff and fecal pollution from agricultural and livestock practices. For instance, the persistence of foodborne pathogens such as Shiga toxin-producing Escherichia coli (STEC) originating from these practices remains poorly quantified. Towards closing these knowledge gaps, the water-sediment interface of two creeks in the Salinas River Valley of California was sampled over a 9-month period using metagenomics and traditional culture-based tests for STEC. Our results revealed that these sediment communities are extremely diverse and have functional and taxonomic diversity comparable to that observed in soils. With our sequencing effort (∼4 Gbp per library), we were unable to detect any pathogenic E. coli in the metagenomes of 11 samples that had tested positive using culture-based methods, apparently due to relatively low abundance. Furthermore, there were no significant differences in the abundance of human- or cow-specific gut microbiome sequences in the downstream impacted sites compared to that in upstream more pristine (control) sites, indicating natural dilution of anthropogenic inputs. Notably, the high number of metagenomic reads carrying antibiotic resistance genes (ARGs) found in all samples was significantly higher than ARG reads in other available freshwater and soil metagenomes, suggesting that these communities may be natural reservoirs of ARGs. The work presented here should serve as a guide for sampling volumes, amount of sequencing to apply, and what bioinformatics analyses to perform when using metagenomics for public health risk studies of environmental samples such as sediments. IMPORTANCE Current agricultural and livestock practices contribute to fecal contamination in the environment and the spread of food- and waterborne disease and antibiotic resistance genes (ARGs). Traditionally, the level of pollution and risk to public health are assessed by culture-based tests for the intestinal bacterium Escherichia coli. However, the accuracy of these traditional methods (e.g., low accuracy in quantification, and false-positive signal when PCR based) and their suitability for sediments remain unclear. We collected sediments for a time series metagenomics study from one of the most highly productive agricultural regions in the United States in order to assess how agricultural runoff affects the native microbial communities and if the presence of Shiga toxin-producing Escherichia coli (STEC) in sediment samples can be detected directly by sequencing. Our study provided important information on the potential for using metagenomics as a tool for assessment of public health risk in natural environments.

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Public Health Investigation of Two Outbreaks of Shiga Toxin-Producing Escherichia coli O157 Associated with Consumption of Watercress

Applied and Environmental Microbiology ◽

10.1128/aem.04188-14 ◽

2015 ◽

Vol 81 (12) ◽

pp. 3946-3952 ◽

Cited By ~ 48

Author(s):

Claire Jenkins ◽

Timothy J. Dallman ◽

Naomi Launders ◽

Caroline Willis ◽

Lisa Byrne ◽

...

Keyword(s):

Public Health ◽

Escherichia Coli ◽

United Kingdom ◽

Shiga Toxin ◽

Phage Type ◽

Epidemiological Studies ◽

Runoff Water ◽

Content Type ◽

Phylogenetic Cluster ◽

Close Proximity

ABSTRACTAn increase in the number of cases of Shiga toxin-producingEscherichia coli(STEC) O157 phage type 2 (PT2) in England in September 2013 was epidemiologically linked to watercress consumption. Whole-genome sequencing (WGS) identified a phylogenetically related cluster of 22 cases (outbreak 1). The isolates comprising this cluster were not closely related to any other United Kingdom strain in the Public Health England WGS database, suggesting a possible imported source. A second outbreak of STEC O157 PT2 (outbreak 2) was identified epidemiologically following the detection of outbreak 1. Isolates associated with outbreak 2 were phylogenetically distinct from those in outbreak 1. Epidemiologically unrelated isolates on the same branch as the outbreak 2 cluster included those from human cases in England with domestically acquired infection and United Kingdom domestic cattle. Environmental sampling using PCR resulted in the isolation of STEC O157 PT2 from irrigation water at one implicated watercress farm, and WGS showed this isolate belonged to the same phylogenetic cluster as outbreak 2 isolates. Cattle were in close proximity to the watercress bed and were potentially the source of the second outbreak. Transfer of STEC from the field to the watercress bed may have occurred through wildlife entering the watercress farm or via runoff water. During this complex outbreak investigation, epidemiological studies, comprehensive testing of environmental samples, and the use of novel molecular methods proved invaluable in demonstrating that two simultaneous outbreaks of STEC O157 PT2 were both linked to the consumption of watercress but were associated with different sources of contamination.

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Whole-Genome-Based Public Health Surveillance of Less Common Shiga Toxin-Producing Escherichia coli Serovars and Untypeable Strains Identifies Four Novel O Genotypes

Journal of Clinical Microbiology ◽

10.1128/jcm.00768-19 ◽

2019 ◽

Vol 57 (10) ◽

Cited By ~ 5

Author(s):

Christina Lang ◽

Miriam Hiller ◽

Regina Konrad ◽

Angelika Fruth ◽

Antje Flieger

Keyword(s):

Public Health ◽

Escherichia Coli ◽

Shiga Toxin ◽

Virulence Gene ◽

Whole Genome Sequence ◽

Complete Analysis ◽

Watery Diarrhea ◽

Whole Genome ◽

Content Type ◽

Gene Information

ABSTRACT Shiga toxin-producing Escherichia coli (STEC) and the STEC subgroup enterohemorrhagic E. coli cause intestinal infections with symptoms ranging from watery diarrhea to hemolytic-uremic syndrome (HUS). A key tool for the epidemiological differentiation of STEC is serotyping. The serotype in combination with the main virulence determinants gives important insight into the virulence potential of a strain. However, a large fraction of STEC strains found in human disease, including strains causing HUS, belongs to less frequently detected STEC serovars or their O/H antigens are unknown or even untypeable. Recent implementation of whole-genome sequence (WGS) analysis, in principle, allows the deduction of serovar and virulence gene information. Therefore, here we compared classical serovar and PCR-based virulence marker detection with WGS-based methods for 232 STEC strains, focusing on less frequently detected STEC serovars and nontypeable strains. We found that the results of WGS-based extraction showed a very high degree of overlap with those of the more classical methods. Specifically, the rate of concordance was 97% for O antigens (OAGs) and 99% for H antigens (HAGs) of typeable strains and >99% for stx1, stx2, or eaeA for all strains. Ninety-eight percent of nontypeable OAGs and 100% of nontypeable HAGs were defined by WGS analysis. In addition, the novel methods enabled a more complete analysis of strains causing severe clinical symptoms and the description of four novel STEC OAG loci. In conclusion, WGS is a promising tool for gaining serovar and virulence gene information, especially from a public health perspective.

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Evaluating the Use of Multilocus Variable Number Tandem Repeat Analysis of Shiga Toxin-Producing Escherichia coli O157 as a Routine Public Health Tool in England

PLoS ONE ◽

10.1371/journal.pone.0085901 ◽

2014 ◽

Vol 9 (1) ◽

pp. e85901 ◽

Cited By ~ 17

Author(s):

Lisa Byrne ◽

Richard Elson ◽

Timothy J. Dallman ◽

Neil Perry ◽

Philip Ashton ◽

...

Keyword(s):

Public Health ◽

Escherichia Coli ◽

Tandem Repeat ◽

Shiga Toxin ◽

Variable Number Tandem Repeat ◽

Variable Number ◽

Escherichia Coli O157 ◽

Repeat Analysis

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Cytotoxic and Apoptotic Effects of Recombinant Subtilase Cytotoxin Variants of Shiga Toxin-Producing Escherichia coli

Infection and Immunity ◽

10.1128/iai.00231-15 ◽

2015 ◽

Vol 83 (6) ◽

pp. 2338-2349 ◽

Cited By ~ 13

Author(s):

J. Funk ◽

N. Biber ◽

M. Schneider ◽

E. Hauser ◽

S. Enzenmüller ◽

...

Keyword(s):

Escherichia Coli ◽

Shiga Toxin ◽

Vero Cells ◽

Toxic Activity ◽

Content Type ◽

B Subunits ◽

Induced Apoptosis ◽

The Individual ◽

Apoptotic Effects ◽

Subtilase Cytotoxin

In this study, the cytotoxicity of the recently described subtilase variant SubAB2-2of Shiga toxin-producingEscherichia coliwas determined and compared to the plasmid-encoded SubAB1and the chromosome-encoded SubAB2-1variant. The genes for the respective enzymatic active (A) subunits and binding (B) subunits of the subtilase toxins were amplified and cloned. The recombinant toxin subunits were expressed and purified. Their cytotoxicity on Vero cells was measured for the single A and B subunits, as well as for mixtures of both, to analyze whether hybrids with toxic activity can be identified. The results demonstrated that all three SubAB variants are toxic for Vero cells. However, the values for the 50% cytotoxic dose (CD50) differ for the individual variants. Highest cytotoxicity was shown for SubAB1. Moreover, hybrids of subunits from different subtilase toxins can be obtained which cause substantial cytotoxicity to Vero cells after mixing the A and B subunits prior to application to the cells, which is characteristic for binary toxins. Furthermore, higher concentrations of the enzymatic subunit SubA1exhibited cytotoxic effects in the absence of the respective B1subunit. A more detailed investigation in the human HeLa cell line revealed that SubA1alone induced apoptosis, while the B1subunit alone did not induce cell death.

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Antibiotic-Mediated Modulations of Outer Membrane Vesicles in Enterohemorrhagic Escherichia coli O104:H4 and O157:H7

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00937-17 ◽

2017 ◽

Vol 61 (9) ◽

Cited By ~ 19

Author(s):

Andreas Bauwens ◽

Lisa Kunsmann ◽

Helge Karch ◽

Alexander Mellmann ◽

Martina Bielaszewska

Keyword(s):

Escherichia Coli ◽

Outer Membrane ◽

Shiga Toxin ◽

Membrane Vesicles ◽

Polymyxin B ◽

Outer Membrane Vesicles ◽

Enterohemorrhagic Escherichia Coli ◽

Content Type ◽

E Coli ◽

Major Virulence Factor

ABSTRACT Ciprofloxacin, meropenem, fosfomycin, and polymyxin B strongly increase production of outer membrane vesicles (OMVs) in Escherichia coli O104:H4 and O157:H7. Ciprofloxacin also upregulates OMV-associated Shiga toxin 2a, the major virulence factor of these pathogens, whereas the other antibiotics increase OMV production without the toxin. These two effects might worsen the clinical outcome of infections caused by Shiga toxin-producing E. coli. Our data support the existing recommendations to avoid antibiotics for treatment of these infections.

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