Virulence Profiling and Molecular Typing of Shiga Toxin-Producing E. coli (STEC) from Human Sources in Brazil

Since no recent data characterizing Shiga toxin-producing E. coli (STEC) from human infections in Brazil are available, the present study aimed to investigate serotypes, stx genotypes, and accessory virulence genes, and also to perform pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST) of 43 STEC strains recovered from 2007 to 2017. Twenty-one distinct serotypes were found, with serotype O111:H8 being the most common. However, serotypes less frequently reported in human diseases were also found and included a hybrid STEC/ETEC O100:H25 clone. The majority of the strains carried stx1a as the sole stx genotype and were positive for the eae gene. Regarding the occurrence of 28 additional virulence genes associated with plasmids and pathogenicity islands, a diversity of profiles was found especially among the eae-harboring strains, which had combinations of markers composed of up to 12 distinct genes. Although PFGE analysis demonstrated genetic diversity between serotypes such as O157:H7, O111:H8, O26:H11, O118:H16, and O123:H2, high genetic relatedness was found for strains of serotypes O24:H4 and O145:H34. MLST allowed the identification of 17 distinct sequence types (STs) with ST 16 and 21 being the most common ones. Thirty-five percent of the strains studied were not typeable by the currently used MLST approach, suggesting new STs. Although STEC O111:H8 remains the leading serotype in Brazil, a diversity of other serotypes, some carrying virulence genes and belonging to STs incriminated as causing severe disease, were found in this study. Further studies are needed to determine whether they have any epidemiological relevance.

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Genotype Analysis of Escherichia coliStrains Isolated from Children and Chickens Living in Close Contact

Applied and Environmental Microbiology ◽

10.1128/aem.65.2.472-476.1999 ◽

1999 ◽

Vol 65 (2) ◽

pp. 472-476 ◽

Cited By ~ 31

Author(s):

S. Kariuki ◽

C. Gilks ◽

J. Kimari ◽

A. Obanda ◽

J. Muyodi ◽

...

Keyword(s):

Gel Electrophoresis ◽

Genomic Dna ◽

Genetic Relatedness ◽

Close Contact ◽

Multidrug Resistant ◽

Pulsed Field Gel Electrophoresis ◽

Pfge Analysis ◽

E Coli ◽

Susceptibility Profiles ◽

Endonuclease Digestion

ABSTRACT Escherichia coli isolates from rectal swabs from 62 chickens and stools from 42 children living in close contact with chickens on the same farms in Kiambu district, Kenya, were compared for their genetic relatedness. Antibiotic susceptibility profiles broadly categorized isolates from the children and from the chickens into two separate clusters: the majority (144; 85.5%) of the E. coli isolates from children were multidrug resistant, while the majority (216; 87.1%) of the E. coli isolates from chickens were either fully susceptible or resistant only to tetracycline. Sixty- and 100- to 110-MDA plasmids were found to encode the transferable resistance to co-trimoxazole and tetracycline.HindIII restriction endonuclease digestion of the 60- and 100- to 110-MDA plasmids produced four distinct patterns for isolates from children and three distinct patterns for isolates from chickens.XbaI digestion of genomic DNA followed by pulsed-field gel electrophoresis (PFGE) analysis produced 14 distinct clusters. There were six distinct PFGE clusters among the isolates from children, while among the isolates from chickens there were seven distinct clusters. Only one PFGE cluster contained isolates from both children and chickens, with the isolates displaying an approximately 60% coefficient of similarity. This study showed that although several different genotypes of E. coli were isolated from children and chickens from the same farms, the E. colistrains from these two sources were distinct.

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Sorbitol non-fermenting shiga toxin-producingEscherichia coliin cattle on smallholdings

Epidemiology and Infection ◽

10.1017/s0950268814000351 ◽

2014 ◽

Vol 143 (1) ◽

pp. 94-103 ◽

Cited By ~ 4

Author(s):

M. Z. ISLAM ◽

J. P. CHRISTENSEN ◽

P. K. BISWAS

Keyword(s):

Escherichia Coli ◽

Polymerase Chain Reaction ◽

Shiga Toxin ◽

Gel Electrophoresis ◽

Virulence Genes ◽

Pulsed Field Gel Electrophoresis ◽

Chain Reaction ◽

E Coli ◽

Faecal Samples ◽

Polymerase Chain

SUMMARYWe investigated faecal samples collected from the rectum of 518 cattle on 371 randomly selected smallholdings in Bangladesh for the presence of sorbitol non-fermenting (SN-F) shiga toxin-producingEscherichia coli(STEC). The SN-F isolates were tested for the presence ofrfbO157,stx1, stx2, eaeandhlyAgenes by polymerase chain reaction (PCR). Seven SN-F isolates lacking these genes were profiled by pulsed-field gel electrophoresis (PFGE) to verify their clonality. SN-FE. coliwas identified in 44 [8·5%, 95% confidence interval (CI) 6·4–11·2] samples; of these, 28 (5·4%, 95% CI 3·8–7·7) had shiga toxin-producing strains, although only two carried therfbO157 gene. Thirteen isolates carried thehlyAgene while 18 harboured theeaegene. Based on PFGE, six pulsotypes were observed among the seven isolates that had no virulence genes. To the best of our knowledge this is the first report on shiga toxin-producingE. colifrom direct rectal faecal samples of cattle on smallholdings.

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Prevalence and Genetic Characterization of Shiga Toxin-Producing Escherichia coli Isolates from Slaughtered Animals in Bangladesh

Applied and Environmental Microbiology ◽

10.1128/aem.00854-08 ◽

2008 ◽

Vol 74 (17) ◽

pp. 5414-5421 ◽

Cited By ~ 43

Author(s):

Mohammad A. Islam ◽

Abdus S. Mondol ◽

Enne de Boer ◽

Rijkelt R. Beumer ◽

Marcel H. Zwietering ◽

...

Keyword(s):

Escherichia Coli ◽

Shiga Toxin ◽

Virulence Genes ◽

Genetic Characterization ◽

Clonal Diversity ◽

Pulsed Field Gel Electrophoresis ◽

E Coli ◽

Phage Types ◽

Distinct Restriction

ABSTRACT To determine the prevalence of Shiga toxin (Stx)-producing Escherichia coli (STEC) in slaughter animals in Dhaka, Bangladesh, we collected rectal contents immediately after animals were slaughtered. Of the samples collected from buffalo (n = 174), cows (n = 139), and goats (n = 110), 82.2%, 72.7%, and 11.8% tested positive for stx 1 and/or stx 2, respectively. STEC could be isolated from 37.9%, 20.1%, and 10.0% of the buffalo, cows, and goats, respectively. STEC O157 samples were isolated from 14.4% of the buffalo, 7.2% of the cows, and 9.1% of the goats. More than 93% (n = 42) of the STEC O157 isolates were positive for the stx 2, eae, katP, etpD, and enterohemorrhagic E. coli hly (hly EHEC) virulence genes. STEC O157 isolates were characterized by seven recognized phage types, of which types 14 (24.4%) and 31 (24.4%) were predominant. Subtyping of the 45 STEC O157 isolates by pulsed-field gel electrophoresis showed 37 distinct restriction patterns, suggesting a heterogeneous clonal diversity. In addition to STEC O157, 71 STEC non-O157 strains were isolated from 60 stx-positive samples from 23.6% of the buffalo, 12.9% of the cows, and 0.9% of the goats. The STEC non-O157 isolates belonged to 36 different O groups and 52 O:H serotypes. Unlike STEC O157, most of the STEC non-O157 isolates (78.9%) were positive for stx 1. Only 7.0% (n = 5) of the isolates were positive for hly EHEC, and none was positive for eae, katP, and etpD. None of the isolates was positive for the iha, toxB, and efa1 putative adhesion genes. However, 35.2% (n = 25), 11.3% (n = 8), 12.7% (n = 9), and 12.7% (n = 9) of the isolates were positive for the lpf O113, saa, lpfA O157/01-141, and lpfA O157/OI-154 genes, respectively. The results of this study provide the first evidence that slaughtered animals like buffalo, cows, and goats in Bangladesh are reservoirs for STEC, including the potentially virulent STEC strain O157.

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Diversity and relatedness of Shiga toxin-producingEscherichia coliandCampylobacter jejunibetween farms in a dairy catchment

Epidemiology and Infection ◽

10.1017/s0950268815002782 ◽

2015 ◽

Vol 144 (7) ◽

pp. 1406-1417 ◽

Cited By ~ 4

Author(s):

H. IRSHAD ◽

A. L. COOKSON ◽

C. M. ROSS ◽

P. JAROS ◽

D. J. PRATTLEY ◽

...

Keyword(s):

Population Structure ◽

Shiga Toxin ◽

Bacterial Culture ◽

Spatial Relationship ◽

Pulsed Field Gel Electrophoresis ◽

Chain Reaction ◽

E Coli ◽

Faecal Samples ◽

Polymerase Chain ◽

Sequence Types

SUMMARYThe aim of this study was to examine the population structure, transmission and spatial relationship between genotypes of Shiga toxin-producingEscherichia coli(STEC) andCampylobacter jejuni, on 20 dairy farms in a defined catchment. Pooled faecal samples (n= 72) obtained from 288 calves were analysed by real-time polymerase chain reaction (rtPCR) forE. coliserotypes O26, O103, O111, O145 and O157. The number of samples positive forE. coliO26 (30/72) was high compared toE. coliO103 (7/72), O145 (3/72), O157 (2/72) and O111 (0/72). EighteenE. coliO26 and 53C. jejuniisolates were recovered from samples by bacterial culture.E. coliO26 andC. jejuniisolates were genotyped using pulsed-field gel electrophoresis and multilocus sequence typing, respectively. AllE. coliO26 isolates could be divided into four clusters and the results indicated thatE. coliO26 isolates recovered from calves on the same farm were more similar than isolates recovered from different farms in the catchment. There were 11 different sequence types ofC. jejuniisolated from the cattle and 22 from water. An analysis of the population structure ofC. jejuniisolated from cattle provided evidence of clustering of genotypes within farms, and among groups of farms separated by road boundaries.

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Prevalence of beta-lactam drug-resistance genes in commensal Escherichia coli contaminating ready-to-eat lettuce

10.1101/824516 ◽

2019 ◽

Author(s):

Ningbo Liao ◽

Julia Rubin ◽

Yuan Hu ◽

Hector A. Ramirez ◽

Clarissa Araújo Borges ◽

...

Keyword(s):

Escherichia Coli ◽

Drug Resistance ◽

Resistance Genes ◽

Beta Lactamase ◽

Beta Lactam ◽

E Coli ◽

Class A ◽

Distinct Sequence ◽

Human Infections ◽

Sequence Types

ABSTRACTThe objective of this study was to evaluate the prevalence of antibiotic resistance and beta-lactam drug resistance genes in Escherichia coli isolated from ready-to-eat lettuce, obtained from local supermarkets in Northern California. Bags of lettuce were purchased from 4 chain supermarkets during three different periods—Oct 2018–Jan 2019, Feb 2019–Apr 2019 and May 2019–July 2019. From 91 packages of lettuce, we recovered 34 E. coli isolates from 22 (24%) lettuce samples. All E. coli isolates were genotyped by multilocus sequence typing (MLST), and we found 15 distinct sequence types (STs). Five of these genotypes (ST2819, ST4600, ST2432, ST1198 and ST5143) have been reported to cause infection in humans. Twenty (59%) E. coli isolates were found resistant to at least one of the antibacterial drugs. They included resistance to ampicillin (AMP, 85%) and ampicillin/sulbactam (SAM, 50%), cefoxitin (FOX, 40%) and cefuroxime (CXM, 35%). We found 8 (40%) of 20 beta-lactam resistant E. coli isolates to carry blaCTX-M; 5 (25%) tested positive for blaSHV, while only 4 (20%) tested positive for blaTEM. Additionally, we identified a class A broad-spectrum beta-lactamase SED-1 gene, blaSED, reported by others in Citrobacter sedlakii isolated from bile of a patient. This study found that a large proportion of fresh lettuce carry beta-lactam drug-resistant E. coli, which could serve as a reservoir for drug resistance genes that could potentially enter pathogens to cause human infections.

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Extended-spectrum β-lactamases, transferable quinolone resistance, and virulotyping in extra-intestinal E. coli in Uruguay

The Journal of Infection in Developing Countries ◽

10.3855/jidc.6918 ◽

2016 ◽

Vol 10 (01) ◽

pp. 43-52 ◽

Cited By ~ 13

Author(s):

Rafael Vignoli ◽

Virginia García-Fulgueiras ◽

Nicolás F Cordeiro ◽

Inés Bado ◽

Verónica Seija ◽

...

Keyword(s):

Gel Electrophoresis ◽

Virulence Genes ◽

Pulsed Field Gel Electrophoresis ◽

Quinolone Resistance ◽

Chain Reaction ◽

E Coli ◽

Extended Spectrum ◽

Polymerase Chain ◽

Sequence Types ◽

M 14

Introduction: To characterize extended-spectrum β-lactamases (ESBLs) and plasmid-mediated quinolone resistance (PMQR) genes in Escherichia coli isolates obtained from extra-intestinal samples in three Uruguayan hospitals. Methodology: Fifty-five ESBL-producing E. coli isolates were studied. Virulence genes, ESBLs, and PMQR genes were detected by polymerase chain reaction. ESBL-producing isolates were compared by pulsed-field gel electrophoresis. Multi-locus sequence typing was also performed on 13 selected isolates. Results: Thirty-seven isolates harbored blaCTX-M-15 (67.3%), eight blaCTX-M-2 (14.6%), five blaCTX-M-14 (9.1%), three carried both blaCTX-M-2 and blaCTX-M-14, one blaCTX-M-9, and one blaCTX-M-8. Among the CTX-M-15 producers, 92% belonged to sequence types ST131 and ST405, and carried aac(6’)Ib-cr as well. Isolates harboring blaCTX-M-2, blaCTX-M-14, blaCTX-M-9, or blaCTX-M-8 were found to be genetically unrelated. Conclusions: The successful dissemination of CTX-M-15-producing E.coli isolates seems to be linked to the spreading of high-risk clones and horizontal gene transfer. A trade-off between carrying more antibiotic resistance and less virulence-related genes could partially account for the evolutionary advantages featured by successful clones.

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Analysis of Yersinia pseudotuberculosis isolates recovered from deceased mammals of a German zoo animal collection

Journal of Clinical Microbiology ◽

10.1128/jcm.03125-20 ◽

2021 ◽

Author(s):

J.A. Hammerl ◽

N vom Ort ◽

A Barac ◽

C Jäckel ◽

L Grund ◽

...

Keyword(s):

Gel Electrophoresis ◽

Virulence Genes ◽

Yersinia Pseudotuberculosis ◽

Pulsed Field Gel Electrophoresis ◽

Single Type ◽

Virulence Plasmid ◽

Wild Animals ◽

Pfge Analysis ◽

Zoo Animals ◽

Sequence Types

Yersinia (Y.) pseudotuberculosis is an important pathogen for both humans and animals. It can infect livestock as well as pets and wild animals. During the last years, a number of reports describe the isolation of Y. pseudotuberculosis from zoo animals, mainly birds and mammals, for which the infection was mostly lethal. Between 2005 and 2019, there were at least 17 cases of deceased mammals belonging to five different species at the Zoo Wuppertal, Germany, which suffered from a Y. pseudotuberculosis infection. Since only scarce information exists on properties of Y. pseudotuberculosis from zoo animals, we characterized eight isolates, covering all infected species, in detail. All of them are members of biotype 1, but belong to five serotypes, five sequence types (STs) and even seven cgMLST types. Using Pulsed-Field Gel Electrophoresis (PFGE) analysis and whole-genome sequencing (WGS), seven isolates could be discriminated from each other. They differ significantly regarding their virulence genes and mobile genetic elements. While the virulence plasmid pYV existed in all serotypes (five isolates), a complete high pathogenicity island (HPI) was detected only in the serotypes O:1a, O:1b and O:13 (four isolates), but not in O:2a and O:2b. Similarly, the content of other plasmids and prophages varies greatly between the isolates. The data demonstrates that the deceased mammals were infected by seven individual isolates and not by a single type predominating in the zoo animals.

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Set of virulence genes and genetic relatedness of O113 : H21 Escherichia coli strains isolated from the animal reservoir and human infections in Brazil

Journal of Medical Microbiology ◽

10.1099/jmm.0.015263-0 ◽

2010 ◽

Vol 59 (6) ◽

pp. 634-640 ◽

Cited By ~ 17

Author(s):

Luis Fernando dos Santos ◽

Kinue Irino ◽

Tânia Mara Ibelli Vaz ◽

Beatriz Ernestina Cabilio Guth

Keyword(s):

Escherichia Coli ◽

Virulence Genes ◽

Genetic Relatedness ◽

High Mass ◽

Human Pathogens ◽

Animal Reservoir ◽

E Coli ◽

Virulence Markers ◽

Human Infections ◽

Animal Isolates

Escherichia coli strains of serotype O113 : H21 are commonly described as belonging to a Shiga toxin (Stx)-producing E. coli (STEC) pathotype worldwide. Albeit this STEC serotype is frequently identified among cattle and other domestic animals, to the best of our knowledge no human infections associated with STEC O113 : H21 have been registered in Brazil to date. Here, we report the virulence profile and genetic relatedness of a collection of O113 : H21 E. coli strains mainly isolated from the animal reservoir aimed at determining their potential as human pathogens. The strains from the animal reservoir (n=34) were all classified as STEC, whereas the few isolates recovered so far from human diarrhoea (n=3) lacked stx genes. Among the STEC, the stx 2d-activatable gene was identified in 85 % of the strains that also carried lpfA O113, iha, saa, ehxA, subAB, astA, cdt-V, espP, espI and epeA; the human strains harboured only lpfA O113, iha and astA. All the strains except one, isolated from cattle, were genetically classified as phylogenetic group B1. High mass plasmids were observed in 25 isolates, but only in the STEC group were these plasmids confirmed as the STEC O113 megaplasmid (pO113). Many closely related subgroups (more than 80 % similarity) were identified by PFGE, with human isolates clustering in a subgroup separate from most of the animal isolates. In conclusion, potentially pathogenic O113 : H21 STEC isolates carrying virulence markers in common with O113 : H21 clones associated with haemolytic uraemic syndrome cases in other regions were demonstrated to occur in the natural reservoir in our settings, and therefore the risk represented by them to public health should be carefully monitored.

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Presence of β-Lactamase-producing Enterobacterales and Salmonella Isolates in Marine Mammals

International Journal of Molecular Sciences ◽

10.3390/ijms22115905 ◽

2021 ◽

Vol 22 (11) ◽

pp. 5905

Author(s):

Olivia M. Grünzweil ◽

Lauren Palmer ◽

Adriana Cabal ◽

Michael P. Szostak ◽

Werner Ruppitsch ◽

...

Keyword(s):

Escherichia Coli ◽

Marine Mammals ◽

Virulence Genes ◽

Multidrug Resistant ◽

Healthcare Sector ◽

Whole Genome ◽

E Coli ◽

High Prevalence ◽

Lactamase Gene ◽

Sequence Types

Marine mammals have been described as sentinels of the health of marine ecosystems. Therefore, the aim of this study was to investigate (i) the presence of extended-spectrum β-lactamase (ESBL)- and AmpC-producing Enterobacterales, which comprise several bacterial families important to the healthcare sector, as well as (ii) the presence of Salmonella in these coastal animals. The antimicrobial resistance pheno- and genotypes, as well as biocide susceptibility of Enterobacterales isolated from stranded marine mammals, were determined prior to their rehabilitation. All E. coli isolates (n = 27) were screened for virulence genes via DNA-based microarray, and twelve selected E. coli isolates were analyzed by whole-genome sequencing. Seventy-one percent of the Enterobacterales isolates exhibited a multidrug-resistant (MDR) pheno- and genotype. The gene blaCMY (n = 51) was the predominant β-lactamase gene. In addition, blaTEM-1 (n = 38), blaSHV-33 (n = 8), blaCTX-M-15 (n = 7), blaOXA-1 (n = 7), blaSHV-11 (n = 3), and blaDHA-1 (n = 2) were detected. The most prevalent non-β-lactamase genes were sul2 (n = 38), strA (n = 34), strB (n = 34), and tet(A) (n = 34). Escherichia coli isolates belonging to the pandemic sequence types (STs) ST38, ST167, and ST648 were identified. Among Salmonella isolates (n = 18), S. Havana was the most prevalent serotype. The present study revealed a high prevalence of MDR bacteria and the presence of pandemic high-risk clones, both of which are indicators of anthropogenic antimicrobial pollution, in marine mammals.

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Seasonal prevalence and characterization of Shiga toxin-producing Escherichia coli on pork carcasses at three steps of the harvest process at two commercial processing plants in the US

10.1101/2020.07.15.205773 ◽

2020 ◽

Author(s):

Ivan Nastasijevic ◽

John W. Schmidt ◽

Marija Boskovic ◽

Milica Glisic ◽

Norasak Kalchayanand ◽

...

Keyword(s):

Escherichia Coli ◽

Foodborne Pathogens ◽

Shiga Toxin ◽

Severe Disease ◽

Control Point ◽

Plate Count ◽

Seasonal Effect ◽

E Coli ◽

Significant Control ◽

Pork Products

ABSTRACTShiga toxin (stx) -producing Escherichia coli (STEC) are foodborne pathogens that have a significant impact on public health, with those possessing the attachment factor intimin (eae) referred to as enterohemorrhagic E. coli (EHEC) associated with life threatening illnesses. Cattle and beef are considered typical sources of STEC, but their presence in pork products is a growing concern. Therefore, carcasses (n=1536) at two U.S. pork processors were sampled once per season at three stages of harvest (post-stunning skins; post-scald carcasses; chilled carcasses) then examined using PCR for stx and eae, aerobic plate count (APC) and Enterobacteriaceae counts (EBC). Skins, post-scald, and chilled carcasses had prevalence of stx (85.3, 17.5, and 5.4%, respectively), with 82.3, 7.8, and 1.7% respectively, having stx and eae present. All stx positive samples were subjected to culture isolation that resulted in 368 STEC and 46 EHEC isolates. The most frequently identified STEC were serogroup O121, O8, and O91(63, 6.7, and 6.0% of total STEC, respectively). The most frequently isolated EHEC was serotype O157:H7 (63% of total EHEC). Results showed that scalding significantly reduced (P < 0.05) carcass APC and EBC by 3.00 and 2.50 log10 CFU/100 cm2 respectively. A seasonal effect was observed with STEC prevalence lower (P < 0.05) in winter. The data from this study shows significant (P < 0.05) reduction in the incidence of STEC (stx) from 85.3% to 5.4% and of EHEC (stx+eae) from 82.3% to 1.7% within slaughter-to-chilling continuum, respectively, and that potential EHEC can be confirmed present throughout using culture isolation.IMPORTANCESeven serogroups of Shiga toxin-producing Escherichia coli (STEC) are responsible for most (>75%) cases of severe illnesses caused by STEC and are considered adulterants of beef. However, some STEC outbreaks have been attributed to pork products although the same E. coli are not considered adulterants in pork because little is known of their prevalence along the pork chain. The significance of the work presented here is that it identifies disease causing STEC, enterohemorrhagic E. coli (EHEC), demonstrating that these same organisms are a food safety hazard in pork as well as beef. The results show that most STEC isolated from pork are not likely to cause severe disease in humans and that processes used in pork harvest, such as scalding, offer a significant control point to reduce contamination. The results will assist the pork processing industry and regulatory agencies to optimize interventions to improve the safety of pork products.

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