Last data update: Sep 23, 2024. (Total: 47723 publications since 2009)
Records 1-10 (of 10 Records) |
Query Trace: Strockbine NA [original query] |
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PacBio Genome Sequences of Eight Escherichia albertii Strains Isolated from Humans in the United States.
Lindsey RL , Rowe LA , Batra D , Smith P , Strockbine NA . Microbiol Resour Announc 2019 8 (9) Escherichia albertii is an emerging pathogen that is closely related to Escherichia coli and can carry some of the same virulence genes as E. coli. Here, we report the release of Illumina-corrected PacBio sequences for eight E. albertii genomes. Two of these strains carry Shiga toxin 2f. |
Interlaboratory Evaluation of the U.S. Food and Drug Administration Escherichia coli Identification Microarray for Profiling Shiga Toxin-Producing Escherichia coli.
Patel IR , Gangiredla J , Lacher DW , Mammel MK , Bagi L , Baranzoni GM , Fratamico PM , Roberts EL , Deb Roy C , Lindsey RL , VStoneburg D , Martin H , Smith P , Strockbine NA , Elkins CA , Scheutz F , Feng PCH . J Food Prot 2018 81 (8) 1275-1282 The U.S. Food and Drug Administration Escherichia coli Identification (FDA-ECID) microarray provides rapid molecular characterization of E. coli. The effectiveness of the FDA-ECID for characterizing Shiga toxin-producing E. coli (STEC) was evaluated by three federal laboratories and one reference laboratory with a panel of 54 reference E. coli strains from the External Quality Assurance program. Strains were tested by FDA-ECID for molecular serotyping (O and H antigens), Shiga toxin subtyping, and the presence of the ehxA and eae genes for enterohemolysin and intimin, respectively. The FDA-ECID O typing was 96% reproducible among the four laboratories and 94% accurate compared with the reference External Quality Assurance data. Discrepancies were due to the absence of O41 target loci on the array and to two pairs of O types with identical target sequences. H typing was 96% reproducible and 100% accurate, with discrepancies due to two strains from one laboratory that were identified as mixed by FDA-ECID. Shiga toxin (Stx) type 1 subtyping was 100% reproducible and accurate, and Stx2 subtyping was 100% reproducible but only 64% accurate. FDA-ECID identified most Stx2 subtypes but had difficulty distinguishing among stx2a, stx2c, and stx2d genes because of close similarities of these sequences. FDA-ECID was 100% effective for detecting ehxA and eae and accurately subtyped the eae alleles. This interlaboratory study revealed that FDA-ECID for STEC characterization was highly reproducible for molecular serotyping, stx and eae subtyping, and ehxA detection. However, the array was less useful for distinguishing among the highly homologous O antigen genes and the stx2a, stx2c, and stx2d subtypes. |
Genetic Analysis of Virulence Potential of Escherichia coli O104 Serotypes Isolated From Cattle Feces Using Whole Genome Sequencing.
Shridhar PB , Patel IR , Gangiredla J , Noll LW , Shi X , Bai J , Elkins CA , Strockbine NA , Nagaraja TG . Front Microbiol 2018 9 (MAR) (341) 341 Escherichia coli O104:H4, a Shiga toxin-producing hybrid pathotype that was implicated in a major foodborne outbreak in Germany in 2011, has not been detected in cattle. However, serotypes of O104, other than O104:H4, have been isolated from cattle feces, with O104:H7 being the most predominant. In this study, we investigated, based on whole genome sequence analyses, the virulence potential of E. coli O104 strains isolated from cattle feces, since cattle are asymptomatic carriers of E. coli O104. The genomes of ten bovine E. coli O104 strains (six O104:H7, one O104:H8, one O104:H12, and two O104:H23) and five O104:H7 isolated from human clinical cases were sequenced. Of all the bovine O104 serotypes (H7, H8, H12, and H23) that were included in the study, only E. coli O104:H7 serotype possessed Shiga toxins. Four of the six bovine O104:H7 strains and one of the five human strains carried stx1c. Three human O104 strains carried stx2, two were of subtype 2a, and one was 2d. Genomes of stx carrying bovine O104:H7 strains were larger than the stx-negative strains of O104:H7 or other serotypes. The genome sizes were proportional to the number of genes carried on the mobile genetic elements (phages, prophages, transposable elements and plasmids). Both bovine and human strains were negative for intimin and other genes associated with the type III secretory system and non-LEE encoded effectors. Plasmid-encoded virulence genes (ehxA, epeA, espP, katP) were also present in bovine and human strains. All O104 strains were negative for antimicrobial resistance genes, except one human strain. Phylogenetic analysis indicated that bovine E. coli O104 strains carrying the same flagellar antigen clustered together and STEC strains clustered separately from non-STEC strains. One of the human O104:H7 strains was phylogenetically closely related to and belonged to the same sequence type (ST-1817) as the bovine O104:H7 STEC strains. This suggests that the bovine feces could be a source of human illness caused by E. coli O104:H7 serotype. Because bovine O104:H7 strains carried virulence genes similar to human clinical strains and one of the human clinical strains was phylogenetically related to bovine strains, the serotype has the potential to be a diarrheagenic pathogen in humans. Copyright © 2018 Shridhar, Patel, Gangiredla, Noll, Shi, Bai, Elkins, Strockbine and Nagaraja. |
Implementation of Whole Genome Sequencing (WGS) for Identification and Characterization of Shiga Toxin-Producing Escherichia coli (STEC) in the United States.
Lindsey RL , Pouseele H , Chen JC , Strockbine NA , Carleton HA . Front Microbiol 2016 7 766 Shiga toxin-producing Escherichia coli (STEC) is an important foodborne pathogen capable of causing severe disease in humans. Rapid and accurate identification and characterization techniques are essential during outbreak investigations. Current methods for characterization of STEC are expensive and time-consuming. With the advent of rapid and cheap whole genome sequencing (WGS) benchtop sequencers, the potential exists to replace traditional workflows with WGS. The aim of this study was to validate tools to do reference identification and characterization from WGS for STEC in a single workflow within an easy to use commercially available software platform. Publically available serotype, virulence, and antimicrobial resistance databases were downloaded from the Center for Genomic Epidemiology (CGE) (www.genomicepidemiology.org) and integrated into a genotyping plug-in with in silico PCR tools to confirm some of the virulence genes detected from WGS data. Additionally, down sampling experiments on the WGS sequence data were performed to determine a threshold for sequence coverage needed to accurately predict serotype and virulence genes using the established workflow. The serotype database was tested on a total of 228 genomes and correctly predicted from WGS for 96.1% of O serogroups and 96.5% of H serogroups identified by conventional testing techniques. A total of 59 genomes were evaluated to determine the threshold of coverage to detect the different WGS targets, 40 were evaluated for serotype and virulence gene detection and 19 for the stx gene subtypes. For serotype, 95% of the O and 100% of the H serogroups were detected at > 40x and ≥ 30x coverage, respectively. For virulence targets and stx gene subtypes, nearly all genes were detected at > 40x, though some targets were 100% detectable from genomes with coverage ≥20x. The resistance detection tool was 97% concordant with phenotypic testing results. With isolates sequenced to > 40x coverage, the different databases accurately predicted serotype, virulence, and resistance from WGS data, providing a fast and cheaper alternative to conventional typing techniques. |
Comparison of whole genome sequences from human and non-human Escherichia coli O26 strains.
Norman KN , Clawson ML , Strockbine NA , Mandrell RE , Johnson R , Ziebell K , Zhao S , Fratamico PM , Stones R , Allard MW , Bono JL . Front Cell Infect Microbiol 2015 5 21 Shiga toxin-producing Escherichia coli (STEC) O26 is the second leading E. coli serogroup responsible for human illness outbreaks behind E. coli O157:H7. Recent outbreaks have been linked to emerging pathogenic O26:H11 strains harboring stx 2 only. Cattle have been recognized as an important reservoir of O26 strains harboring stx 1; however the reservoir of these emerging stx 2 strains is unknown. The objective of this study was to identify nucleotide polymorphisms in human and cattle-derived strains in order to compare differences in polymorphism derived genotypes and virulence gene profiles between the two host species. Whole genome sequencing was performed on 182 epidemiologically unrelated O26 strains, including 109 human-derived strains and 73 non-human-derived strains. A panel of 289 O26 strains (241 STEC and 48 non-STEC) was subsequently genotyped using a set of 283 polymorphisms identified by whole genome sequencing, resulting in 64 unique genotypes. Phylogenetic analyses identified seven clusters within the O26 strains. The seven clusters did not distinguish between isolates originating from humans or cattle; however, clusters did correspond with particular virulence gene profiles. Human and non-human-derived strains harboring stx 1 clustered separately from strains harboring stx 2, strains harboring eae, and non-STEC strains. Strains harboring stx 2 were more closely related to non-STEC strains and strains harboring eae than to strains harboring stx 1. The finding of human and cattle-derived strains with the same polymorphism derived genotypes and similar virulence gene profiles, provides evidence that similar strains are found in cattle and humans and transmission between the two species may occur. |
Clinical isolates of Shiga toxin 1a-producing Shigella flexneri with an epidemiological link to recent travel to Hispañiola.
Gray MD , Lampel KA , Strockbine NA , Fernandez RE , Melton-Celsa AR , Maurelli AT . Emerg Infect Dis 2014 20 (10) 1669-77 Shiga toxins (Stx) are cytotoxins involved in severe human intestinal disease. These toxins are commonly found in Shigella dysenteriae serotype 1 and Shiga-toxin-producing Escherichia coli; however, the toxin genes have been found in other Shigella species. We identified 26 Shigella flexneri serotype 2 strains isolated by public health laboratories in the United States during 2001-2013, which encode the Shiga toxin 1a gene (stx1a). These strains produced and released Stx1a as measured by cytotoxicity and neutralization assays using anti-Stx/Stx1a antiserum. The release of Stx1a into culture supernatants increased approximately 100-fold after treatment with mitomycin C, suggesting that stx1a is carried by a bacteriophage. Infectious phage were found in culture supernatants and increased approximately 1,000-fold with mitomycin C. Whole-genome sequencing of several isolates and PCR analyses of all strains confirmed that stx1a was carried by a lambdoid bacteriophage. Furthermore, all patients who reported foreign travel had recently been to Hispaniola, suggesting that emergence of these novel strains is associated with that region. |
Genotypes and virulence characteristics of Shiga toxin-producing Escherichia coli O104 strains from different origins and sources.
Miko A , Delannoy S , Fach P , Strockbine NA , Lindstedt BA , Mariani-Kurkdjian P , Reetz J , Beutin L . Int J Med Microbiol 2013 303 (8) 410-21 Sixty-two Escherichia coli strains carrying the wzxO104-gene from different sources, origins and time periods were analyzed for their serotypes, virulence genes and compared for genomic similarity by pulsed-field gel-electrophoresis (PFGE). The O104 antigen was present in 55 strains and the structurally and genetically related capsular antigen K9 in five strains. The presence of 49 genes associated with enteropathogenic E. coli (EPEC), enteroaggregative E. coli (EAEC) and enterohemorrhagic E. coli (EHEC) was investigated. Fifty-four strains of serotypes O104:H2 (n=1), O104:H4 (n=37), O104:H7 (n=5) and O104:H21 (n=11) produced Shiga-toxins (Stx). Among STEC O104, a close association between serotype, virulence gene profile and genomic similarity was found. EAEC virulence genes were only present in STEC O104:H4 strains. EHEC-O157 plasmid-encoded genes were only found in STEC O104:H2, O104:H7 and O104:H21 strains. None of the 62 O104 or K9 strains carried an eae-gene involved in the attaching and effacing phenotype. The 38 O104:H4 strains formed a single PFGE-cluster (>83.7% similarity). Thirty-one of these strains were from the European O104:H4 outbreak in 2011. The outbreak strains and older O104:H4 strains from Germany (2001), Georgia and France (2009) clustered together at>86.2% similarity. O104:H4 strains isolated between 2001 and 2009 differed for some plasmid-encoded virulence genes compared to the outbreak strains from 2011. STEC O104:H21 and STEC O104:H7 strains isolated in the U.S. and in Europe showed characteristic differences in their Stx-types, virulence gene and PFGE profiles indicating that these have evolved separately. E. coli K9 strains were not associated with virulence and were heterogeneous for their serotypes and PFGE profiles. |
Spread of a distinct Stx2-encoding phage prototype among Escherichia coli O104:H4 strains from outbreaks in Germany, Norway, and Georgia
Beutin L , Hammerl JA , Strauch E , Reetz J , Dieckmann R , Kelner-Burgos Y , Martin A , Miko A , Strockbine NA , Lindstedt BA , Horn D , Monse H , Huettel B , Muller I , Stuber K , Reinhardt R . J Virol 2012 86 (19) 10444-55 Shiga toxin 2 (Stx2)-producing Escherichia coli (STEC) O104:H4 caused one of the world's largest outbreaks of hemorrhagic colitis and hemolytic uremic syndrome in Germany in 2011. These strains have evolved from enteroaggregative E. coli (EAEC) by the acquisition of the Stx2 genes and have been designated enteroaggregative hemorrhagic E. coli. Nucleotide sequencing has shown that the Stx2 gene is carried by prophages integrated into the chromosome of STEC O104:H4. We studied the properties of Stx2-encoding bacteriophages which are responsible for the emergence of this new type of E. coli pathogen. For this, we analyzed Stx bacteriophages from STEC O104:H4 strains from Germany (in 2001 and 2011), Norway (2006), and the Republic of Georgia (2009). Viable Stx2-encoding bacteriophages could be isolated from all STEC strains except for the Norwegian strain. The Stx2 phages formed lysogens on E. coli K-12 by integration into the wrbA locus, resulting in Stx2 production. The nucleotide sequence of the Stx2 phage P13374 of a German STEC O104:H4 outbreak was determined. From the bioinformatic analyses of the prophage sequence of 60,894 bp, 79 open reading frames were inferred. Interestingly, the Stx2 phages from the German 2001 and 2011 outbreak strains were found to be identical and closely related to the Stx2 phages from the Georgian 2009 isolates. Major proteins of the virion particles were analyzed by mass spectrometry. Stx2 production in STEC O104:H4 strains was inducible by mitomycin C and was compared to Stx2 production of E. coli K-12 lysogens. |
Association of nucleotide polymorphisms within the O-antigen gene cluster of Escherichia coli O26, O45, O103, O111, O121, and O145 with serogroups and genetic subtypes.
Norman KN , Strockbine NA , Bono JL . Appl Environ Microbiol 2012 78 (18) 6689-703 Shiga toxin-producing Escherichia coli (STEC) strains are important food-borne pathogens capable of causing hemolytic-uremic syndrome. STEC O157:H7 strains cause the majority of severe disease in the United States; however, there is a growing concern for the amount and severity of illness attributable to non-O157 STEC. Recently, the Food Safety and Inspection Service (FSIS) published the intent to regulate the presence of STEC belonging to serogroups O26, O45, O103, O111, O121, and O145 in nonintact beef products. To ensure the effective control of these bacteria, sensitive and specific tests for their detection will be needed. In this study, we identified single nucleotide polymorphisms (SNPs) in the O-antigen gene cluster that could be used to detect STEC strains of the above-described serogroups. Using comparative DNA sequence analysis, we identified 22 potentially informative SNPs among 164 STEC and non-STEC strains of the above-described serogroups and designed matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF) assays to test the STEC allele frequencies in an independent panel of bacterial strains. We found at least one SNP that was specific to each serogroup and also differentiated between STEC and non-STEC strains. Differences in the DNA sequence of the O-antigen gene cluster corresponded well with differences in the virulence gene profiles and provided evidence of different lineages for STEC and non-STEC strains. The SNPs discovered in this study can be used to develop tests that will not only accurately identify O26, O45, O103, O111, O121, and O145 strains but also predict whether strains detected in the above-described serogroups contain Shiga toxin-encoding genes. |
Multicenter evaluation of a sequence-based protocol for subtyping Shiga toxins and standardizing stx nomenclature
Scheutz F , Teel LD , Beutin L , Pierard D , Buvens G , Karch H , Mellmann A , Caprioli A , Tozzoli R , Morabito S , Strockbine NA , Melton-Celsa AR , Sanchez M , Persson S , O'Brien AD . J Clin Microbiol 2012 50 (9) 2951-63 When Shiga toxin-producing Escherichia coli (STEC) strains emerged as agents of human disease, two types of toxin were identified: Shiga toxin type 1 (Stx1) (almost identical to Shiga toxin produced by Shigella dysenteriae type 1) and the immunologically distinct type 2 (Stx2). Subsequently, numerous STEC strains have been characterized that express toxins with variations in amino acid sequence, some of which confer unique biological properties. These variants were grouped within the Stx1 or Stx2 type and often assigned names to indicate that they were not identical in sequence or phenotype to the main Stx1 or Stx2 type. A lack of specificity or consistency in toxin nomenclature has led to much confusion in the characterization of STEC strains. Because serious outcomes of infection have been attributed to certain Stx subtypes and less so with others, we sought to better define the toxin subtypes within the main Stx1 and Stx2 types. We compared the levels of relatedness of 285 valid sequence variants of Stx1 and Stx2 and identified common sequences characteristic of each of three Stx/Stx1 and seven Stx2 subtypes. A novel, simple PCR subtyping method was developed, independently tested on a battery of 48 prototypic STEC strains, and improved at six clinical and research centers to test the reproducibility, sensitivity, and specificity of the PCR. Using a consistent schema for nomenclature of the Stx toxins and stx genes by phylogenetic sequence-based relatedness of the holotoxin proteins, we developed a typing approach that should obviate the need to bioassay each newly described toxin and that predicts important biological characteristics. |
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