scholarly journals Escherichia coliH-Genotyping PCR: a Complete and Practical Platform for Molecular H Typing

2018 ◽  
Vol 56 (6) ◽  
Author(s):  
Masaya Banjo ◽  
Atsushi Iguchi ◽  
Kazuko Seto ◽  
Taisei Kikuchi ◽  
Tetsuya Harada ◽  
...  
Keyword(s):  
Low Cost ◽  
Epidemiological Studies ◽  
Pathogenic Potential ◽  
Content Type ◽  
E Coli ◽  
Outbreak Investigations ◽  
Pcr Products ◽  
Wild Strains ◽  
Encoding Genes ◽  
Reference Strains

ABSTRACTInEscherichia coli, more than 180 O groups and 53 H types have been recognized. The O:H serotyping ofE. colistrains is an effective method for identifying strains with pathogenic potential and classifying them into clonal groups. In particular, the serotyping of Shiga toxin-producingE. coli(STEC) strains provides valuable information to evaluate the routes, sources, and prevalence of agents in outbreak investigations and surveillance. Here, we present a complete and practical PCR-based H-typing system,E. coliH-genotyping PCR, consisting of 10 multiplex PCR kits with 51 single PCR primer pairs. Primers were designed based on a detailed comparative analysis of sequences from all H-antigen (flagellin)-encoding genes,fliCand its homologs. The specificity of this system was confirmed by using all H type reference strains. Additionally, 362 serotyped wild strains were also used to evaluate its practicality. All 277 H-type-identified isolates gave PCR products that corresponded to the results of serological H typing. Moreover, 76 nonmotile and nine untypeable strains could be successfully subtyped into any H type by the PCR system. TheE. coliH-genotyping PCR developed here allows broader, rapid, and low-cost subtyping of H types and will assist epidemiological studies as well as surveillance of pathogenicE. coli.

scholarly journals Escherichia coli O-Genotyping PCR: a Comprehensive and Practical Platform for Molecular O Serogrouping

2015 ◽  
Vol 53 (8) ◽  
pp. 2427-2432 ◽  
Author(s):  
Atsushi Iguchi ◽  
Sunao Iyoda ◽  
Kazuko Seto ◽  
Tomoko Morita-Ishihara ◽  
Flemming Scheutz ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Low Cost ◽  
Epidemiological Studies ◽  
Biosynthesis Gene Cluster ◽  
Content Type ◽  
E Coli ◽  
O Antigen ◽  
Promising Tool ◽  
Outbreak Investigations ◽  
Wild Strains

The O serogrouping of pathogenicEscherichia coliis a standard method for subtyping strains for epidemiological studies and enhancing phylogenetic studies. In particular, the identification of strains of the same O serogroup is essential in outbreak investigations and surveillance. In a previous study, we analyzed the O-antigen biosynthesis gene cluster in all knownE. coliO serogroups (A. Iguchi et al., DNA Res, 22:101–107, 2015,http://dx.doi.org/10.1093/dnares/dsu043). Based on those results, we have arranged 162 PCR primer pairs for the identification or classification of O serogroups. Of these, 147 pairs were used to identify 147 individual O serogroups with unique O-antigen biosynthesis genes, and the other 15 pairs were used to identify 15 groups of strains (Gp1 to Gp15). Each of these groups consisted of strains with identical or very similar O-antigen biosynthesis genes, and the groups represented a total of 35 individual O serogroups. We then used the 162 primer pairs to create 20 multiplex PCR sets. Each set contained six to nine primer pairs that amplify products of markedly different sizes. This genetic methodology (E. coliO-genotyping PCR) allowed for comprehensive, rapid, and low-cost typing. Validation of the PCR system using O-serogroup references and wild strains showed that the correct O serogroups were specifically and accurately identified for 100% (182/182) and 90.8% (522/575) of references and wild strains, respectively. The PCR-based system reported here might be a promising tool for the subtyping ofE. colistrains for epidemiological studies as well as for the surveillance of pathogenicE. coliduring outbreaks.

scholarly journals Autotransporter Protein-Encoding Genes of Diarrheagenic Escherichia coli Are Found in both Typical and Atypical Enteropathogenic E. coli Strains

2012 ◽  
Vol 79 (1) ◽  
pp. 411-414 ◽  
Author(s):  
Afonso G. Abreu ◽  
Vanessa Bueris ◽  
Tatiane M. Porangaba ◽  
Marcelo P. Sircili ◽  
Fernando Navarro-Garcia ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Content Type ◽  
E Coli ◽  
Diarrheagenic Escherichia Coli ◽  
Virulence Markers ◽  
Protein Encoding ◽  
Encoding Genes ◽  
Specific Virulence

ABSTRACTAutotransporter (AT) protein-encoding genes of diarrheagenicEscherichia coli(DEC) pathotypes (cah,eatA,ehaABCDJ,espC,espI,espP,pet,pic,sat, andtibA) were detected in typical and atypical enteropathogenicE. coli(EPEC) in frequencies between 0.8% and 39.3%. Although these ATs have been described in particular DEC pathotypes, their presence in EPEC indicates that they should not be considered specific virulence markers.

scholarly journals Additional Og-Typing PCR Techniques Targeting Escherichia coli-Novel and Shigella-Unique O-Antigen Biosynthesis Gene Clusters

2020 ◽  
Vol 58 (11) ◽  
Author(s):  
Atsushi Iguchi ◽  
Hironobu Nishii ◽  
Kazuko Seto ◽  
Jiro Mitobe ◽  
Kenichi Lee ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Strong Association ◽  
Gene Clusters ◽  
Epidemiological Studies ◽  
Content Type ◽  
E Coli ◽  
O Antigen ◽  
Wide Range ◽  
Biosynthesis Gene ◽  
Almost All

ABSTRACT The O-serogrouping of pathogenic Escherichia coli is a standard method for subtyping strains for epidemiological studies and controls. O-serogroup diversification shows a strong association with the genetic diversity in some O-antigen biosynthesis gene clusters. Through genomic studies, in addition to the types of O-antigen biosynthesis gene clusters (Og-types) from conventional O-serogroup strains, a number of novel Og-types have been found in E. coli isolates. To assist outbreak investigations and surveillance of pathogenic E. coli at inspection institutes, in previous studies, we developed PCR methods that could determine almost all conventional O-serogroups and some novel Og-types. However, there are still many Og-types that may not be determined by simple genetic methods such as PCR. Thus, in the present study, we aimed to develop an additional Og-typing PCR system. Based on the novel Og-types, including OgN32, OgN33, and OgN34, presented in this study, we designed an additional 24 PCR primer pairs targeting 14 novel and 2 diversified E. coli Og-types and 8 Shigella-unique Og-types. Subsequently, we developed 5 new multiplex PCR sets consisting of 33 primers, including the aforementioned 24 primers and 9 primers reported in previous studies. The accuracy and specificity of the PCR system was validated using approximately 260 E. coli and Shigella O-serogroup and Og-type reference strains. The Og-typing PCR system reported here can determine a wide range of Og-types of E. coli and may help epidemiological studies, in addition to the surveillance of pathogenic E. coli.

scholarly journals Rapid Identification of Different Escherichia coli Sequence Type 131 Clades

2017 ◽  
Vol 61 (8) ◽  
Author(s):  
Yasufumi Matsumura ◽  
Johann D. D. Pitout ◽  
Gisele Peirano ◽  
Rebekah DeVinney ◽  
Taro Noguchi ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Epidemiological Studies ◽  
Rapid Identification ◽  
Sequence Type ◽  
Fluoroquinolone Resistance ◽  
Pcr Assay ◽  
Content Type ◽  
E Coli ◽  
Multiplex Pcr Assay ◽  
Clade C

ABSTRACT Escherichia coli sequence type 131 (ST131) is a pandemic clonal lineage that is responsible for the global increase in fluoroquinolone resistance and extended-spectrum-β-lactamase (ESBL) producers. The members of ST131 clade C, especially subclades C2 and C1-M27, are associated with ESBLs. We developed a multiplex conventional PCR assay with the ability to detect all ST131 clades (A, B, and C), as well as C subclades (C1-M27, C1-nM27 [C1-non-M27], and C2). To validate the assay, we used 80 ST131 global isolates that had been fully sequenced. We then used the assay to define the prevalence of each clade in two Japanese collections consisting of 460 ESBL-producing E. coli ST131 (2001-12) and 329 E. coli isolates from extraintestinal sites (ExPEC) (2014). The assay correctly identified the different clades in all 80 global isolates: clades A (n = 12), B (n = 12), and C, including subclades C1-M27 (n = 16), C1-nM27 (n = 20), C2 (n = 17), and other C (n = 3). The assay also detected all 565 ST131 isolates in both collections without any false positives. Isolates from clades A (n = 54), B (n = 23), and C (n = 483) corresponded to the O serotypes and the fimH types of O16-H41, O25b-H22, and O25b-H30, respectively. Of the 483 clade C isolates, C1-M27 was the most common subclade (36%), followed by C1-nM27 (32%) and C2 (15%). The C1-M27 subclade with bla CTX-M-27 became especially prominent after 2009. Our novel multiplex PCR assay revealed the predominance of the C1-M27 subclade in recent Japanese ESBL-producing E. coli isolates and is a promising tool for epidemiological studies of ST131.

scholarly journals Genome Signatures of Escherichia coli O157:H7 Isolates from the Bovine Host Reservoir

2011 ◽  
Vol 77 (9) ◽  
pp. 2916-2925 ◽  
Author(s):  
Mark Eppinger ◽  
Mark K. Mammel ◽  
Joseph E. LeClerc ◽  
Jacques Ravel ◽  
Thomas A. Cebula
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Phylogenomic Analysis ◽  
Genotypic Differences ◽  
Pathogenic Potential ◽  
Content Type ◽  
E Coli ◽  
A Genome ◽  
Major Interest ◽  
Host Reservoir

ABSTRACTCattle comprise a main reservoir of Shiga toxin-producingEscherichia coliO157:H7 (STEC). The significant differences in host prevalence, transmissibility, and virulence phenotypes among strains from bovine and human sources are of major interest to the public health community and livestock industry. Genomic analysis revealed divergence into three lineages: lineage I and lineage I/II strains are commonly associated with human disease, while lineage II strains are overrepresented in the asymptomatic bovine host reservoir. Growing evidence suggests that genotypic differences between these lineages, such as polymorphisms in Shiga toxin subtypes and synergistically acting virulence factors, are correlated with phenotypic differences in virulence, host ecology, and epidemiology. To assess the genomic plasticity on a genome-wide scale, we have sequenced the whole genome of strain EC869, a bovine-associatedE. coliO157:H7 isolate. Comparative phylogenomic analysis of this key isolate enabled us to place accurately bovine lineage II strains within the genetically homogenousE. coliO157:H7 clade. Identification of polymorphic loci that are anchored both in the chromosomal backbone and horizontally acquired regions allowed us to associate bovine genotypes with altered virulence phenotypes and host prevalence. This study catalogued numerous novel lineage II-specific genome signatures, some of which appear to be associated intimately with the altered pathogenic potential and niche adaptation within the bovine rumen. The presented extended list of polymorphic markers is valuable in the development of a robust typing system critical for forensic, diagnostic, and epidemiological studies of this emerging human pathogen.

scholarly journals Genomic Diversity and Virulence Profiles of Historical Escherichia coli O157 Strains Isolated from Clinical and Environmental Sources

2014 ◽  
Vol 81 (2) ◽  
pp. 569-577 ◽  
Author(s):  
Lydia V. Rump ◽  
Narjol Gonzalez-Escalona ◽  
Wenting Ju ◽  
Fei Wang ◽  
Guojie Cao ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
The United States ◽  
Genomic Diversity ◽  
Severe Illness ◽  
Pathogenic Potential ◽  
Content Type ◽  
E Coli ◽  
Virulence Markers ◽  
Virulence Potential ◽  
Food Borne

ABSTRACTEscherichia coliO157:H7 is, to date, the majorE. coliserotype causing food-borne human disease worldwide. Strains of O157 with other H antigens also have been recovered. We analyzed a collection of historic O157 strains (n= 400) isolated in the late 1980s to early 1990s in the United States. Strains were predominantly serotype O157:H7 (55%), and various O157:non-H7 (41%) serotypes were not previously reported regarding their pathogenic potential. Although lacking Shiga toxin (stx) andeaegenes, serotypes O157:H1, O157:H2, O157:H11, O157:H42, and O157:H43 carried several virulence factors (iha,terD, andhlyA) also found in virulent serotypeE. coliO157:H7. Pulsed-field gel electrophoresis (PFGE) showed the O157 serogroup was diverse, with strains with the same H type clustering together closely. Among non-H7 isolates, serotype O157:H43 was highly prevalent (65%) and carried important enterohemorrhagicE. coli(EHEC) virulence markers (iha,terD,hlyA, andespP). Isolates from two particular H types, H2 and H11, among the most commonly found non-O157 EHEC serotypes (O26:H11, O111:H11, O103:H2/H11, and O45:H2), unexpectedly clustered more closely with O157:H7 than other H types and carried several virulence genes. This suggests an early divergence of the O157 serogroup to clades with different pathogenic potentials. The appearance of important EHEC virulence markers in closely related H types suggests their virulence potential and suggests further monitoring of those serotypes not implicated in severe illness thus far.

scholarly journals Pathogenic Potential, Genetic Diversity, and Population Structure of Escherichia coli Strains Isolated from a Forest-Dominated Watershed (Comox Lake) in British Columbia, Canada

2014 ◽  
Vol 81 (5) ◽  
pp. 1788-1798 ◽  
Author(s):  
Abhirosh Chandran ◽  
Asit Mazumder
Keyword(s):  
Escherichia Coli ◽  
Drinking Water ◽  
Population Structure ◽  
Virulence Genes ◽  
Repetitive Element ◽  
Fingerprint Analysis ◽  
Pathogenic Potential ◽  
Content Type ◽  
E Coli ◽  
Heat Stable

ABSTRACTEscherichia coliisolates (n= 658) obtained from drinking water intakes of Comox Lake (2011 to 2013) were screened for the following virulence genes (VGs):stx1andstx2(Shiga toxin-producingE. coli[STEC]),eaeand the adherence factor (EAF) gene (enteropathogenicE. coli[EPEC]), heat-stable (ST) enterotoxin (variants STh and STp) and heat-labile enterotoxin (LT) genes (enterotoxigenicE. coli[ETEC]), andipaH(enteroinvasiveE. coli[EIEC]). The only genes detected wereeaeandstx2, which were carried by 37.69% (n= 248) of the isolates. Onlyeaewas harbored by 26.74% (n= 176) of the isolates, representing potential atypical EPEC strains, while onlystx2was detected in 10.33% (n= 68) of the isolates, indicating potential STEC strains. Moreover, four isolates were positive for both thestx2andeaegenes, representing potential EHEC strains. The prevalence of VGs (eaeorstx2) was significantly (P< 0.0001) higher in the fall season, and multiple genes (eaeplusstx2) were detected only in fall. Repetitive element palindromic PCR (rep-PCR) fingerprint analysis of 658E. coliisolates identified 335 unique fingerprints, with an overall Shannon diversity (H′) index of 3.653. Diversity varied among seasons over the years, with relatively higher diversity during fall. Multivariate analysis of variance (MANOVA) revealed that the majority of the fingerprints showed a tendency to cluster according to year, season, and month. Taken together, the results indicated that the diversity and population structure ofE. colifluctuate on a temporal scale, reflecting the presence of diverse host sources and their behavior over time in the watershed. Furthermore, the occurrence of potentially pathogenicE. colistrains in the drinking water intakes highlights the risk to human health associated with direct and indirect consumption of untreated surface water.

scholarly journals Escherichia coli Isolates That Carryvat,fyuA,chuA, andyfcVEfficiently Colonize the Urinary Tract

2012 ◽  
Vol 80 (12) ◽  
pp. 4115-4122 ◽  
Author(s):  
Rachel R. Spurbeck ◽  
Paul C. Dinh ◽  
Seth T. Walk ◽  
Ann E. Stapleton ◽  
Thomas M. Hooton ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Urinary Tract Infection ◽  
Urinary Tract ◽  
Tract Infection ◽  
Virulence Factors ◽  
Neonatal Meningitis ◽  
Content Type ◽  
E Coli ◽  
Core Set ◽  
Encoding Genes

ABSTRACTExtraintestinalEscherichia coli(ExPEC), a heterogeneous group of pathogens, encompasses avian, neonatal meningitis, and uropathogenicE. colistrains. While several virulence factors are associated with ExPEC, there is no core set of virulence factors that can be used to definitively differentiate these pathotypes. Here we describe a multiplex of four virulence factor-encoding genes,yfcV,vat,fyuA, andchuA, highly associated with uropathogenicE. colistrains that can distinguish three groups ofE. coli: diarrheagenic and animal-associatedE. colistrains, human commensal and avian pathogenicE. colistrains, and uropathogenic and neonatal meningitisE. colistrains. Furthermore, human intestinal isolates that encode all four predictor genes express them during exponential growth in human urine and colonize the bladder in the mouse model of ascending urinary tract infection in higher numbers than human commensal strains that do not encode the four predictor genes (P= 0.02), suggesting that the presence of the predictors correlates with uropathogenic potential.

scholarly journals Complete Genome Sequence of Escherichia coli Podophage Penshu1

2019 ◽  
Vol 8 (38) ◽  
Author(s):  
Douglas Pechacek ◽  
Myung Hwangbo ◽  
Russell Moreland ◽  
Mei Liu ◽  
Jolene Ramsey
Keyword(s):  
Escherichia Coli ◽  
Complete Genome Sequence ◽  
Complete Genome ◽  
Coliform Bacteria ◽  
Negative Bacterium ◽  
Gram Negative ◽  
Content Type ◽  
E Coli ◽  
Protein Encoding ◽  
Encoding Genes

Escherichia coli 4s is a Gram-negative bacterium found in the equine intestinal ecosystem alongside diverse other coliform bacteria and bacteriophages. This announcement describes the complete genome of the T7-like E. coli 4s podophage Penshu1. From its 39,263-bp genome, 54 protein-encoding genes and a 179-bp terminal repeat were predicted.

scholarly journals Systematic Nomenclature for GGDEF and EAL Domain-Containing Cyclic Di-GMP Turnover Proteins of Escherichia coli: TABLE 1

2015 ◽  
Vol 198 (1) ◽  
pp. 7-11 ◽  
Author(s):  
Regine Hengge ◽  
Michael Y. Galperin ◽  
Jean-Marc Ghigo ◽  
Mark Gomelsky ◽  
Jeffrey Green ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Molecular Mechanisms ◽  
Bacterial Species ◽  
Content Type ◽  
E Coli ◽  
Diguanylate Cyclases ◽  
Genes Encoding ◽  
Systematic Nomenclature ◽  
K 12 ◽  
Encoding Genes

In recent years,Escherichia colihas served as one of a few model bacterial species for studying cyclic di-GMP (c-di-GMP) signaling. The widely usedE. coliK-12 laboratory strains possess 29 genes encoding proteins with GGDEF and/or EAL domains, which include 12 diguanylate cyclases (DGC), 13 c-di-GMP-specific phosphodiesterases (PDE), and 4 “degenerate” enzymatically inactive proteins. In addition, six new GGDEF and EAL (GGDEF/EAL) domain-encoding genes, which encode two DGCs and four PDEs, have recently been found in genomic analyses of commensal and pathogenicE. colistrains. As a group of researchers who have been studying the molecular mechanisms and the genomic basis of c-di-GMP signaling inE. coli, we now propose a general and systematicdgcandpdenomenclature for the enzymatically active GGDEF/EAL domain-encoding genes of this model species. This nomenclature is intuitive and easy to memorize, and it can also be applied to additional genes and proteins that might be discovered in various strains ofE. coliin future studies.

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