A report on the prevalence and the whole-genome sequence-based analysis of Shiga-toxin producing Escherichia coli isolated from the recto-anal junction of slaughter-age Irish sheep

2021 ◽  
Author(s):  
Siobhán C. McCarthy ◽  
Guerrino Macori ◽  
Gina Duggan ◽  
Catherine M. Burgess ◽  
Séamus Fanning ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Human Disease ◽  
Pathogenic Bacteria ◽  
Small Ruminants ◽  
Whole Genome Sequence ◽  
Health Concern ◽  
Toxin Gene ◽  
Whole Genome ◽  
Faecal Excretion

Shiga toxin-producing Escherichia coli (STEC) are a diverse group of pathogenic bacteria capable of causing serious human illness and serogroups O157 and O26 are frequently implicated in human disease. Ruminant hosts are the primary STEC reservoir and small ruminants are important contributors to STEC transmission. This study investigated the prevalence, serotypes and shedding dynamics of STEC, including the super-shedding of serogroups O157 and O26, in Irish sheep. Recto-anal mucosal swab samples (N=840) were collected over 24 months from two ovine slaughtering facilities. Samples were plated on selective agars and were quantitatively and qualitatively assessed via real-time PCR for Shiga-toxin prevalence and serogroup. A subset of STEC isolates (N=199) were selected for whole-genome sequencing and analysed in silico . In total, 704/840 (83.8%) swab samples were Shiga-toxin positive following RT-PCR screening, and 363/704 (51.6%) animals were subsequently culture positive for STEC. Five animals were shedding STEC O157 and three of these were identified as super-shedders. No STEC O26 was isolated. Post-hoc statistical analysis showed that younger animals are more likely to harbour STEC and STEC carriage is most prevalent during the summer months. Following sequencing, 178/199 genomes were confirmed as STEC. Thirty-five different serotypes were identified, fifteen of which were not yet reported in sheep. Serotype O91:H14 was the most frequently reported. Eight Shiga-toxin gene variants were reported, two stx 1 and six stx 2 , and three novel Shiga-toxin subunit combinations were observed. Variant stx 1c was the most prevalent, while many strains also harboured stx 2b . Importance Shiga toxin-producing Escherichia coli (STEC) are foodborne, zoonotic pathogens of significant public health concern. All STEC harbour stx , a critical virulence determinant, but it is not expressed in most serotypes. Sheep shed the pathogen via faecal excretion and are increasingly recognised as important contributors in the dissemination of STEC. In this study, we have found that there is high prevalence of STEC circulating within sheep and prevalence is related to animal age and seasonality. Further, sheep harbour a variety of non-O157 STEC, whose prevalence and contribution to human disease has been under investigated for many years. A variety of Stx variants were also observed, some of which are of high clinical importance.

scholarly journals Whole-Genome Analysis of a Shiga Toxin-Producing Escherichia coli O103:H2 Strain Isolated from Cattle Feces

2020 ◽  
Vol 9 (45) ◽  
Author(s):  
Yujie Zhang ◽  
Yen-Te Liao ◽  
Vivian C. H. Wu
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Whole Genome Sequence ◽  
Whole Genome ◽  
Whole Genome Analysis ◽  
Content Type ◽  
Cattle Feces ◽  
Beef Products ◽  
Foodborne Outbreaks ◽  
Enterocyte Effacement

ABSTRACT Shiga toxin-producing Escherichia coli (STEC) serotype O103 is one of the primary pathogenic contaminants of beef products, contributing to several foodborne outbreaks in recent years. Here, we report the whole-genome sequence of a STEC O103:H2 strain isolated from cattle feces that contains a locus of enterocyte effacement (LEE) pathogenicity island.

Escherichia coli O157:H7 Cells Exposed to Lettuce Leaf Lysate in Refrigerated Conditions Exhibit Differential Expression of Selected Virulence and Adhesion-Related Genes with Altered Mammalian Cell Adherence

2016 ◽  
Vol 79 (7) ◽  
pp. 1259-1265 ◽  
Author(s):  
NICOLE M. KENNEDY ◽  
NABANITA MUKHERJEE ◽  
PRATIK BANERJEE
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Mammalian Cells ◽  
Pathogenic Bacteria ◽  
Cold Shock ◽  
Cold Shock Protein ◽  
Toxin Gene ◽  
Adhesion Assay ◽  
Escherichia Coli O157 ◽  
The Impact

ABSTRACT Contamination by and persistence of pathogenic bacteria in ready-to-eat produce have emerged as significant food safety and public health concerns. Viable produceborne pathogens cope with several stresses (e.g., temperature fluctuations and low-temperature storage) during production and storage of the commodities. In this study, we investigated the impact of transient cold shock on Escherichia coli O157:H7 (EcO157) cells in a produce matrix (romaine lettuce leaf lysate). EcO157 cells were exposed to 25°C for 1 h, 4°C for 1 h, and 4°C for 10 min in lettuce lysate. The expression of selected genes coding for virulence, stress response, and heat and cold shock proteins was quantified by real-time quantitative reverse transcription PCR assay. Treated EcO157 cells adhered to MAC-T mammalian cells were enumerated by in vitro bioassay. Expression of the Shiga toxin 1 gene (stx1a) was upregulated significantly (P < 0.05) upon cold shock treatments, but virulence genes related to EcO157 attachment (eaeA, lpfA, and hcpA) were down-regulated. Two key members of the cold shock regulon, cold shock protein (cspA) and gyrA, were significantly induced (P < 0.05) at the refrigeration temperature (4°C). Significant upregulation of an SOS response gene, recA, was also observed. E. coli heat shock regulon member grpE was induced, but a universal stress protein (uspA) was down-regulated at the refrigeration temperatures in lettuce lysate. The adhesion assay revealed a temperature-dependent reduction in the attachment of cold-shocked EcO157 cells. The results of the current study indicate a reduction in the attachment of cold-shocked EcO157 to epithelial cells and higher levels of Shiga toxin gene expression at the molecular level.

scholarly journals Inconsistent PCR detection of Shiga toxin-producing Escherichia coli: Insights from whole genome sequence analyses

PLoS ONE ◽  
2021 ◽  
Vol 16 (9) ◽  
pp. e0257168
Author(s):  
Vinicius Silva Castro ◽  
Rodrigo Ortega Polo ◽  
Eduardo Eustáquio de Souza Figueiredo ◽  
Emmanuel Wihkochombom Bumunange ◽  
Tim McAllister ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Bacterial Genome ◽  
Disease Outbreaks ◽  
Illumina Miseq ◽  
Feedlot Cattle ◽  
Pcr Detection ◽  
Whole Genome Sequence ◽  
Whole Genome ◽  
The Stability

Shiga toxin-producing Escherichia coli (STEC) have been linked to food-borne disease outbreaks. As PCR is routinely used to screen foods for STEC, it is important that factors leading to inconsistent detection of STEC by PCR are understood. This study used whole genome sequencing (WGS) to investigate causes of inconsistent PCR detection of stx1, stx2, and serogroup-specific genes. Fifty strains isolated from Alberta feedlot cattle from three different studies were selected with inconsistent or consistent detection of stx and serogroup by PCR. All isolates were initially classified as STEC by PCR. Sequencing was performed using Illumina MiSeq® with sample library by Nextera XT. Virtual PCRs were performed using Geneious and bacteriophage content was determined using PHASTER. Sequencing coverage ranged from 47 to 102x, averaging 74x, with sequences deposited in the NCBI database. Eleven strains were confirmed by WGS as STEC having complete stxA and stxB subunits. However, truncated stx fragments occurred in twenty-two other isolates, some having multiple stx fragments in the genome. Isolates with complete stx by WGS had consistent stx1 and stx2 detection by PCR, although one also having a stx2 fragment had inconsistent stx2 PCR. For all STEC and 18/39 non-STEC, serogroups determined by PCR agreed with those determined by WGS. An additional three WGS serotypes were inconclusive and two isolates were Citrobacter spp. Results demonstrate that stx fragments associated with stx-carrying bacteriophages in the E. coli genome may contribute to inconsistent detection of stx1 and stx2 by PCR. Fourteen isolates had integrated stx bacteriophage but lacked complete or fragmentary stx possibly due to partial bacteriophage excision after sub-cultivation or other unclear mechanisms. The majority of STEC isolates (7/11) did not have identifiable bacteriophage DNA in the contig(s) where stx was located, likely increasing the stability of stx in the bacterial genome and its detection by PCR.

scholarly journals Evaluation of applicability of DNA microarray–based characterization of bovine Shiga toxin–producing Escherichia coli isolates using whole genome sequence analysis

2017 ◽  
Vol 29 (5) ◽  
pp. 721-724 ◽  
Author(s):  
Stefanie A. Barth ◽  
Christian Menge ◽  
Inga Eichhorn ◽  
Torsten Semmler ◽  
Derek Pickard ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Dna Microarray ◽  
Shiga Toxin ◽  
In Silico ◽  
Whole Genome Sequence ◽  
Whole Genome ◽  
Molecular Surveillance ◽  
Antimicrobial Resistance Genes ◽  
O Antigens

We assessed the ability of a commercial DNA microarray to characterize bovine Shiga toxin–producing Escherichia coli (STEC) isolates and evaluated the results using in silico hybridization of the microarray probes within whole genome sequencing scaffolds. From a total of 69,954 reactions (393 probes with 178 isolates), 68,706 (98.2%) gave identical results by DNA microarray and in silico probe hybridization. Results were more congruent when detecting the genoserotype (209 differing results from 19,758 in total; 1.1%) or antimicrobial resistance genes (AMRGs; 141 of 26,878; 0.5%) than when detecting virulence-associated genes (VAGs; 876 of 22,072; 4.0%). Owing to the limited coverage of O-antigens by the microarray, only 37.2% of the isolates could be genoserotyped. However, the microarray proved suitable to rapidly screen bovine STEC strains for the occurrence of high numbers of VAGs and AMRGs and is suitable for molecular surveillance workflows.

scholarly journals Complete Genome Sequence of Escherichia coli Phage vB_EcoS Sa179lw, Isolated from Surface Water in a Produce-Growing Area in Northern California

2018 ◽  
Vol 6 (27) ◽  
Author(s):  
Yen-Te Liao ◽  
Fang Liu ◽  
Xincheng Sun ◽  
Robert W. Li ◽  
Vivian C. H. Wu
Keyword(s):  
Escherichia Coli ◽  
Surface Water ◽  
Genome Sequence ◽  
Shiga Toxin ◽  
Virulence Gene ◽  
Whole Genome Sequence ◽  
Whole Genome ◽  
Content Type ◽  
E Coli ◽  
Escherichia Coli Phage

We report here the whole-genome sequence of a novel Escherichia coli phage, vB_EcoS Sa179lw, isolated from surface water collected in a produce-growing area. With the presence of a putative eae-like gene that was associated with previous non-O157 Shiga toxin-producing E. coli outbreaks, this phage is a candidate for the study of virulence gene transfer.

scholarly journals Whole-Genome Sequence of Escherichia coli Serotype O157:H7 Strain B6914-ARS

2017 ◽  
Vol 5 (44) ◽  
Author(s):  
Gaylen A. Uhlich ◽  
Erin R. Reichenberger ◽  
Bryan J. Cottrell ◽  
Pina Fratamico ◽  
Elisa Andreozzi
Keyword(s):  
Escherichia Coli ◽  
Disease Control ◽  
Genome Sequence ◽  
Shiga Toxin ◽  
Applied Research ◽  
Whole Genome Sequence ◽  
Whole Genome ◽  
Content Type ◽  
Control And Prevention ◽  
Shiga Toxin Genes

ABSTRACT Escherichia coli serotype O157:H7 strain B6914-MS1 is an isolate from the Centers for Disease Control and Prevention that is missing both Shiga toxin genes and has been used extensively in applied research studies. Here we report the genome sequence of strain B6914-ARS, a B6914-MS1 clone that has unique biofilm properties.

Comparative Evaluation of Genomic and Laboratory Approaches for Determination of Shiga Toxin Subtypes in Escherichia coli

2016 ◽  
Vol 79 (12) ◽  
pp. 2078-2085 ◽  
Author(s):  
CATHERINE D. CARRILLO ◽  
ADAM G. KOZIOL ◽  
AMIT MATHEWS ◽  
NORIKO GOJI ◽  
DOMINIC LAMBERT ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Genome Sequence ◽  
Shiga Toxin ◽  
In Silico ◽  
Whole Genome Sequence ◽  
Whole Genome ◽  
Pcr Method ◽  
In Silico Pcr

ABSTRACT The determination of Shiga toxin (ST) subtypes can be an important element in the risk characterization of foodborne ST-producing Escherichia coli (STEC) isolates for making risk management decisions. ST subtyping methods include PCR techniques based on electrophoretic or pyrosequencing analysis of amplicons and in silico techniques based on whole genome sequence analysis using algorithms that can be readily incorporated into bioinformatics analysis pipelines for characterization of isolates by their genetic composition. The choice of technique will depend on the performance characteristics of the method and an individual laboratory's access to specialized equipment or personnel. We developed two whole genome sequence–based ST subtyping tools: (i) an in silico PCR algorithm requiring genome assembly to replicate a reference PCR-based method developed by the Statens Serum Institut (SSI) and (ii) an assembly-independent routine in which raw sequencing results are mapped to a database of known ST subtype sequence variants (V-Typer). These tools were evaluated alongside the SSI reference PCR method and a recently described PCR-based pyrosequencing technique. The V-Typer method results corresponded closely with the reference method in the analysis of 67 STEC cultures obtained from a World Health Organization National Reference Laboratory. In contrast, the in silico PCR method failed to detect ST subtypes in several cases, a result which we attribute to assembly-induced errors typically encountered with repetitive gene sequences. The V-Typer can be readily integrated into bioinformatics protocols used in the identification and characterization of foodborne STEC isolates.

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