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1.
Microbiol Resour Announc ; 8(33)2019 Aug 15.
Artículo en Inglés | MEDLINE | ID: mdl-31416858

RESUMEN

The use of culture methods to detect Escherichia coli diversity does not provide sufficient resolution to identify strains present at low levels. Here, we target the hypervariable gnd gene and describe a database containing 534 distinct partial gnd sequences and associated O groups for use with culture-independent E. coli community analysis.

2.
Microbiol Resour Announc ; 8(12)2019 Mar 21.
Artículo en Inglés | MEDLINE | ID: mdl-30938701

RESUMEN

Water from the Hickey Run Tributary of the Anacostia River is being collected quarterly (beginning August 2018) and analyzed to create high-resolution baseline taxonomic profiles of microbiota associated with this important aquatic ecosystem, which has a long history of exposure to residential and commercial effluents from Washington, DC. These United States National Arboretum Microbial Observatory data are available under NCBI BioProject number PRJNA498951.

3.
Foodborne Pathog Dis ; 16(1): 54-59, 2019 01.
Artículo en Inglés | MEDLINE | ID: mdl-30601032

RESUMEN

Enterobacteriaceae producing ß-lactamases have spread rapidly worldwide and pose a serious threat to human-animal-environment interface. In this study, we present the presence of Salmonella enterica (1.3%) and commensal Escherichia coli (96.3%) isolated from 400 environmental fecal dairy cattle samples over 20 farms in Uganda. Among E. coli isolates, 21% were resistant to at least one antimicrobial tested and 7% exhibited multidrug resistance. Four E. coli isolates displayed extended-spectrum beta-lactamase (ESBL)-producing genes, including blaCTX-M-15 (n = 2/4), blaCTX-M-27 (n = 1/4), blaSHV-12 (n = 1/4), and blaTEM-1B (n = 2/4). Whole genome sequencing confirmed the presence of the plasmid-mediated quinolone resistance qnrS1 gene among three ESBL isolates. No statistically significant differences in seasonal prevalence for E. coli and S. enterica among dairy cattle sampling periods were observed. Furthermore, to our knowledge, this is the first report of E. coli carrying blaCTX-M-15, blaCTX-M-27, blaSHV-12, or qnrS1 isolated from dairy cattle in Uganda. We conclude that the presence of globally disseminated blaCTX-M-15 and blaCTX-M-27 warrants further study to prevent further spread. In addition, the presence of fluoroquinolone resistant ESBL-producing E. coli on dairy farms highlights the potential risk among the human-livestock-environment interaction. This study can be used as a baseline for implementation of a more robust national integrated surveillance system throughout Uganda.


Asunto(s)
Farmacorresistencia Bacteriana Múltiple , Infecciones por Escherichia coli/veterinaria , Escherichia coli/aislamiento & purificación , Salmonelosis Animal/microbiología , Salmonella enterica/aislamiento & purificación , Animales , Antiinfecciosos/farmacología , Bovinos , Estudios Transversales , Industria Lechera , Escherichia coli/enzimología , Escherichia coli/genética , Infecciones por Escherichia coli/epidemiología , Infecciones por Escherichia coli/microbiología , Granjas , Heces/microbiología , Femenino , Proyectos Piloto , Plásmidos/genética , Quinolonas/farmacología , Salmonelosis Animal/epidemiología , Salmonella enterica/enzimología , Salmonella enterica/genética , Uganda/epidemiología , beta-Lactamasas/genética
4.
Artículo en Inglés | MEDLINE | ID: mdl-30533715

RESUMEN

Here, we report the genomes of all 72 isolates belonging to the Escherichia coli reference (ECOR) collection. Strains in this collection were isolated from diverse hosts and geographic locations and have been used for more than 30 years to represent the phylogenetic diversity of E. coli.

5.
Artículo en Inglés | MEDLINE | ID: mdl-30533843

RESUMEN

Shigella is a genus of Gram-negative enteric pathogenic bacteria which has four species, Shigella dysenteriae, S. flexneri, S. boydii, and S. sonnei. Shigella species are clinically important bacteria because they cause shigellosis or dysentery. Here we report the genome sequences of 72 Shigella isolates from these four species.

6.
J Food Prot ; 81(8): 1275-1282, 2018 08.
Artículo en Inglés | MEDLINE | ID: mdl-29985068

RESUMEN

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.


Asunto(s)
Proteínas de Escherichia coli , Microbiología de Alimentos , Escherichia coli Shiga-Toxigénica , Virulencia/genética , Proteínas de Escherichia coli/genética , Humanos , Serotipificación , Toxina Shiga , Toxina Shiga I , Escherichia coli Shiga-Toxigénica/aislamiento & purificación , Estados Unidos , United States Food and Drug Administration
7.
Genome Announc ; 6(26)2018 Jun 28.
Artículo en Inglés | MEDLINE | ID: mdl-29954907

RESUMEN

Here, we present the genome sequences of 23 Bifidobacterium isolates from several commercially available dietary supplements and cultured food products. Strains of this genus are natural inhabitants of the mammalian mouth, gastrointestinal tract, and vagina. Some species are considered beneficial to human health.

8.
Genome Announc ; 6(26)2018 Jun 28.
Artículo en Inglés | MEDLINE | ID: mdl-29954914

RESUMEN

Here, we present the genome sequences of 56 isolates of 10 species of the genus Lactobacillus that are considered beneficial components of the gut microbiota. The isolates examined were found in commercially available dietary supplements in the U.S. market.

9.
Genome Announc ; 6(18)2018 May 03.
Artículo en Inglés | MEDLINE | ID: mdl-29724828

RESUMEN

We report here the genome sequences of 55 strains belonging to the genus Escherichia from multiple animal and environmental sources. These strains include representatives of Escherichia albertii, Escherichia fergusonii, and six additional genetically distinct lineages of Escherichia spp., one of which is newly discovered and is being reported for the first time here.

10.
Genome Announc ; 5(50)2017 Dec 14.
Artículo en Inglés | MEDLINE | ID: mdl-29242221

RESUMEN

Pathogenic and nonpathogenic Escherichia coli strains present a vast genomic diversity. We report the genome sequences of 2,244 E. coli isolates from multiple animal and environmental sources. Their phylogenetic relationships and potential risk to human health were examined.

11.
Appl Environ Microbiol ; 83(18)2017 09 15.
Artículo en Inglés | MEDLINE | ID: mdl-28687651

RESUMEN

Shiga toxin-producing Escherichia coli (STEC) strains of the O91:H21 serotype have caused severe infections, including hemolytic-uremic syndrome. Strains of the O91 serogroup have been isolated from food, animals, and the environment worldwide but are not well characterized. We used a microarray and other molecular assays to examine 49 serogroup O91 strains (environmental, food, and clinical strains) for their virulence potential and phylogenetic relationships. Most of the isolates were identified to be strains of the O91:H21 and O91:H14 serotypes, with a few O91:H10 strains and one O91:H9 strain being identified. None of the strains had the eae gene, which codes for the intimin adherence protein, and many did not have some of the genetic markers that are common in other STEC strains. The genetic profiles of the strains within each serotype were similar but differed greatly between strains of different serotypes. The genetic profiles of the O91:H21 strains that we tested were identical or nearly identical to those of the clinical O91:H21 strains that have caused severe diseases. Multilocus sequence typing and clustered regularly interspaced short palindromic repeat analyses showed that the O91:H21 strains clustered within the STEC 1 clonal group but the other O91 serotype strains were phylogenetically diverse.IMPORTANCE This study showed that food and environmental O91:H21 strains have similar genotypic profiles and Shiga toxin subtypes and are phylogenetically related to the O91:H21 strains that have caused hemolytic-uremic syndrome, suggesting that these strains may also have the potential to cause severe illness.


Asunto(s)
Microbiología Ambiental , Infecciones por Escherichia coli/microbiología , Carne/microbiología , Escherichia coli Shiga-Toxigénica/aislamiento & purificación , Animales , Bovinos , Pollos , Ciervos , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Microbiología de Alimentos , Humanos , Filogenia , Serogrupo , Toxina Shiga/metabolismo , Escherichia coli Shiga-Toxigénica/clasificación , Escherichia coli Shiga-Toxigénica/genética , Escherichia coli Shiga-Toxigénica/metabolismo
12.
J Food Prot ; 80(3): 383-391, 2017 03.
Artículo en Inglés | MEDLINE | ID: mdl-28199145

RESUMEN

Shiga toxin-producing Escherichia coli (STEC) of serotype O113:H21 have caused severe diseases but are unusual in that they do not produce the intimin protein required for adherence to intestinal epithelial cells. Strains of serogroup O113 are one of the most common STEC found in ground beef and beef products in the United States, but their virulence potential is unknown. We used a microarray to characterize 65 O113 strains isolated in the United States from ground beef, beef trim, cattle feces, and fresh spinach. Most were O113:H21 strains, but there were also nine strains of O113:H4 serotype. Although strains within the same serotype had similar profiles for the genes that were tested on the array, the profiles were distinct between the two serotypes, and the strains belonged to different clonal groups. Analysis by clustered regularly interspaced short palindromic repeat analysis showed that O113:H4 strains are conserved genetically, but the O113:H21 strains showed considerable polymorphism and genetic diversity. In comparison to the O113:H21 strains from Australia that were implicated in severe disease, the U.S. isolates showed similar genetic profiles to the known pathogens from Australia, suggesting that these may also have the potential to cause infections.


Asunto(s)
Bovinos/microbiología , Serogrupo , Escherichia coli Shiga-Toxigénica/aislamiento & purificación , Animales , Australia , Escherichia coli/aislamiento & purificación , Infecciones por Escherichia coli/veterinaria , Proteínas de Escherichia coli/genética , Carne Roja , Serotipificación , Virulencia/genética
13.
PLoS One ; 11(12): e0167870, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-27930729

RESUMEN

Consumption of fresh bagged spinach contaminated with Shiga toxin-producing Escherichia coli (STEC) has led to severe illness and death; however current culture-based methods to detect foodborne STEC are time consuming. Since not all STEC strains are considered pathogenic to humans, it is crucial to incorporate virulence characterization of STEC in the detection method. In this study, we assess the comprehensiveness of utilizing a shotgun metagenomics approach for detection and strain-level identification by spiking spinach with a variety of genomically disparate STEC strains at a low contamination level of 0.1 CFU/g. Molecular serotyping, virulence gene characterization, microbial community analysis, and E. coli core gene single nucleotide polymorphism (SNP) analysis were performed on metagenomic sequence data from enriched samples. It was determined from bacterial community analysis that E. coli, which was classified at the phylogroup level, was a major component of the population in most samples. However, in over half the samples, molecular serotyping revealed the presence of indigenous E. coli which also contributed to the percent abundance of E. coli. Despite the presence of additional E. coli strains, the serotype and virulence genes of the spiked STEC, including correct Shiga toxin subtype, were detected in 94% of the samples with a total number of reads per sample averaging 2.4 million. Variation in STEC abundance and/or detection was observed in replicate spiked samples, indicating an effect from the indigenous microbiota during enrichment. SNP analysis of the metagenomic data correctly placed the spiked STEC in a phylogeny of related strains in cases where the indigenous E. coli did not predominate in the enriched sample. Also, for these samples, our analysis demonstrates that strain-level phylogenetic resolution is possible using shotgun metagenomic data for determining the genomic relatedness of a contaminating STEC strain to other closely related E. coli.


Asunto(s)
Metagenómica , Toxina Shiga/biosíntesis , Escherichia coli Shiga-Toxigénica/metabolismo , Spinacia oleracea/microbiología , Filogenia , Polimorfismo de Nucleótido Simple , Escherichia coli Shiga-Toxigénica/clasificación , Escherichia coli Shiga-Toxigénica/patogenicidad , Especificidad de la Especie , Virulencia
14.
Parasit Vectors ; 9(1): 389, 2016 07 07.
Artículo en Inglés | MEDLINE | ID: mdl-27387914

RESUMEN

Feline aelurostrongylosis, caused by the metastrongyloid nematode Aelurostrongylus abstrusus, is an important gastropod-borne parasitic lung disease in cats. Infection with A. abstrusus is widespread globally, but the increasing awareness of this parasite and the advent of more sensitive diagnostics have contributed to the apparent increase in its prevalence and geographic expansion. Clinical features may range in severity from subclinical to life-threatening respiratory disease. Parasitological standard techniques, such as visualization of the nematode first larval stage in faecal and respiratory (bronchial mucus or pleural fluid) samples, remain the mainstays of diagnosis. However, diagnosis is evolving with recent advances in serological and molecular testing, which can improve the time to initiation of effective anthelmintic therapy. Despite numerous anthelmintics that are now available as treatment options, the role of host immunity and lifestyle factors in selecting cats that may benefit from more targeted anthelmintic prophylaxis or treatment practice remains unclear and is likely to guide therapeutic choices as newer data become available. This review summarizes the biology, epidemiology, pathophysiology, diagnosis and treatment options currently available for feline aelurostrongylosis.


Asunto(s)
Enfermedades de los Gatos/epidemiología , Metastrongyloidea/aislamiento & purificación , Infecciones por Strongylida/veterinaria , Animales , Enfermedades de los Gatos/diagnóstico , Enfermedades de los Gatos/parasitología , Gatos , Heces/parasitología , Larva , Infecciones por Strongylida/diagnóstico , Infecciones por Strongylida/epidemiología , Infecciones por Strongylida/parasitología
15.
Infect Immun ; 84(8): 2362-2371, 2016 08.
Artículo en Inglés | MEDLINE | ID: mdl-27271741

RESUMEN

Enteroinvasive Escherichia coli (EIEC) is a unique pathovar that has a pathogenic mechanism nearly indistinguishable from that of Shigella species. In contrast to isolates of the four Shigella species, which are widespread and can be frequent causes of human illness, EIEC causes far fewer reported illnesses each year. In this study, we analyzed the genome sequences of 20 EIEC isolates, including 14 first described in this study. Phylogenomic analysis of the EIEC genomes demonstrated that 17 of the isolates are present in three distinct lineages that contained only EIEC genomes, compared to reference genomes from each of the E. coli pathovars and Shigella species. Comparative genomic analysis identified genes that were unique to each of the three identified EIEC lineages. While many of the EIEC lineage-specific genes have unknown functions, those with predicted functions included a colicin and putative proteins involved in transcriptional regulation or carbohydrate metabolism. In silico detection of the Shigella virulence plasmid (pINV), which is essential for the invasion of host cells, demonstrated that a form of pINV was present in nearly all EIEC genomes, but the Mxi-Spa-Ipa region of the plasmid that encodes the invasion-associated proteins was absent from several of the EIEC isolates. The comparative genomic findings in this study support the hypothesis that multiple EIEC lineages have evolved independently from multiple distinct lineages of E. coli via the acquisition of the Shigella virulence plasmid and, in some cases, the Shigella pathogenicity islands.


Asunto(s)
Escherichia coli Enteropatógena/clasificación , Escherichia coli Enteropatógena/genética , Escherichia coli/clasificación , Escherichia coli/genética , Genoma Bacteriano , Genómica , Shigella/clasificación , Shigella/genética , Biología Computacional/métodos , Escherichia coli Enteropatógena/aislamiento & purificación , Genes Bacterianos , Genómica/métodos , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Sistemas de Lectura Abierta , Filogenia , Plásmidos/genética , Virulencia/genética
16.
Appl Environ Microbiol ; 82(14): 4309-4319, 2016 07 15.
Artículo en Inglés | MEDLINE | ID: mdl-27208138

RESUMEN

UNLABELLED: Hybrid isolates of Shiga toxin-producing Escherichia coli (STEC) and enterotoxigenic E. coli (ETEC) encoding heat-stable enterotoxin (ST) are being reported with increasing frequency from a variety of sources. However, information regarding the plasmids that these strains harbor is scarce. In this study, we sequence and characterize a plasmid, p7v, from the STEC/ETEC hybrid strain 7v. Whole-genome phylogenetic analyses of STEC/ETEC hybrid strains and prototype E. coli isolates of other pathotypes placed 7v in the Escherichia sp. cryptic lineage 1 (CL1) clade. The complete plasmid, p7v, was determined to be 229,275 bp and encodes putative virulence factors that are typically carried on STEC plasmids as well as those often carried on ETEC plasmids, indicating that the hybrid nature of the strain extends beyond merely encoding the two toxins. Plasmid p7v carries two copies of sta with identical sequences, which were discovered to be divergent from the sta sequences found in the prototype human ETEC strains. Using a nomenclature scheme based on a phylogeny constructed from sta and stb sequences, the sta encoded on p7v is designated STa4. In silico analysis determined that p7v also encodes the K88 fimbria, a colonization factor usually associated with porcine ETEC plasmids. The p7v sequence and the presence of plasmid-encoded virulence factors are compared to those of other STEC/ETEC CL1 hybrid genomes and reveal gene acquisition/loss at the strain level. In addition, the interrogation of 24 STEC/ETEC hybrid genomes for identification of plasmid replicons, colonization factors, Stx and ST subtypes, and other plasmid-encoded virulence genes highlights the diversity of these hybrid strains. IMPORTANCE: Hybrid Shiga toxin-producing Escherichia coli/enterotoxigenic Escherichia coli (STEC/ETEC) strains, which have been isolated from environmental, animal, and human clinical samples, may represent an emerging threat as food-borne pathogens. Characterization of these strains is important for assessing virulence potential, aiding in the development of pathogen detection methods, and understanding how the hybrid strains evolve to potentially have a greater impact on public health. This study represents, to our knowledge, both the first characterization of a closed plasmid sequence from a STEC/ETEC hybrid strain and the most comprehensive phylogenetic analysis of available STEC/ETEC hybrid genomes to date. The results demonstrate how the mobility of plasmid-associated virulence genes has resulted in the creation of a diverse plasmid repertoire within the STEC/ETEC hybrid strains.


Asunto(s)
Escherichia coli Enterotoxigénica/genética , Plásmidos/análisis , Recombinación Genética , Escherichia coli Shiga-Toxigénica/genética , Animales , ADN Bacteriano/química , ADN Bacteriano/genética , Escherichia coli Enterotoxigénica/clasificación , Escherichia coli Enterotoxigénica/aislamiento & purificación , Genes Bacterianos , Genoma Bacteriano , Humanos , Filogenia , Análisis de Secuencia de ADN , Homología de Secuencia , Escherichia coli Shiga-Toxigénica/clasificación , Escherichia coli Shiga-Toxigénica/aislamiento & purificación
17.
Appl Environ Microbiol ; 82(11): 3384-3394, 2016 06 01.
Artículo en Inglés | MEDLINE | ID: mdl-27037122

RESUMEN

UNLABELLED: Most Escherichia coli strains are nonpathogenic. However, for clinical diagnosis and food safety analysis, current identification methods for pathogenic E. coli either are time-consuming and/or provide limited information. Here, we utilized a custom DNA microarray with informative genetic features extracted from 368 sequence sets for rapid and high-throughput pathogen identification. The FDA Escherichia coli Identification (FDA-ECID) platform contains three sets of molecularly informative features that together stratify strain identification and relatedness. First, 53 known flagellin alleles, 103 alleles of wzx and wzy, and 5 alleles of wzm provide molecular serotyping utility. Second, 41,932 probe sets representing the pan-genome of E. coli provide strain-level gene content information. Third, approximately 125,000 single nucleotide polymorphisms (SNPs) of available whole-genome sequences (WGS) were distilled to 9,984 SNPs capable of recapitulating the E. coli phylogeny. We analyzed 103 diverse E. coli strains with available WGS data, including those associated with past foodborne illnesses, to determine robustness and accuracy. The array was able to accurately identify the molecular O and H serotypes, potentially correcting serological failures and providing better resolution for H-nontypeable/nonmotile phenotypes. In addition, molecular risk assessment was possible with key virulence marker identifications. Epidemiologically, each strain had a unique comparative genomic fingerprint that was extended to an additional 507 food and clinical isolates. Finally, a 99.7% phylogenetic concordance was established between microarray analysis and WGS using SNP-level data for advanced genome typing. Our study demonstrates FDA-ECID as a powerful tool for epidemiology and molecular risk assessment with the capacity to profile the global landscape and diversity of E. coli IMPORTANCE: This study describes a robust, state-of-the-art platform developed from available whole-genome sequences of E. coli and Shigella spp. by distilling useful signatures for epidemiology and molecular risk assessment into one assay. The FDA-ECID microarray contains features that enable comprehensive molecular serotyping and virulence profiling along with genome-scale genotyping and SNP analysis. Hence, it is a molecular toolbox that stratifies strain identification and pathogenic potential in the contexts of epidemiology and phylogeny. We applied this tool to strains from food, environmental, and clinical sources, resulting in significantly greater phylogenetic and strain-specific resolution than previously reported for available typing methods.


Asunto(s)
Escherichia coli/clasificación , Escherichia coli/genética , Técnicas de Genotipaje/métodos , Análisis por Micromatrices/métodos , Epidemiología Molecular/métodos , Serotipificación/métodos , Factores de Virulencia/análisis , Variación Genética , Estados Unidos , United States Food and Drug Administration , Factores de Virulencia/genética
18.
J Food Prot ; 79(10): 1656-1662, 2016 10.
Artículo en Inglés | MEDLINE | ID: mdl-28221838

RESUMEN

More than 470 serotypes of Shiga toxin-producing Escherichia coli (STEC) have been identified, but not all cause severe illness in humans. Most STEC that cause severe diseases can adhere to epithelial cells, produce specific stx subtypes, and belong to certain serotypes; therefore, these traits appear to be critical STEC risk factors. However, testing for these traits is labor intensive, and serotyping is inadequate because of extensive variations among E. coli O and H antigen types. In the present study, the E. coli identification microarray, which tests for over 40,000 E. coli gene targets, was examined for its potential to quickly characterize STEC strains. Analysis of 47 E. coli isolates, including 31 STEC isolates, recovered from 39 foods revealed that the microarray effectively determined the presence or absence of adherence genes and identified the specific eae allele in 3 isolates. The array identified most of the stx subtypes carried by all the isolates but had some difficulties in discerning between stx2a, stx2c, and stx2d because of the genetic similarities of these subtypes. The array determined the O and H types of 68 and 96% of the isolates, respectively, and although most serotypes were unremarkable, a few known pathogenic serotypes were also found. These selected STEC traits provided a scientific basis for assessing the potential health risks of STEC strains and also showed the importance of H typing in determining health risks. However, the diversity of the STEC group, the complexity of virulence mechanisms, and the variation in pathotypes among strains continue to pose challenges to assessing the potential of STEC strains to cause severe illness.


Asunto(s)
Escherichia coli/aislamiento & purificación , Escherichia coli Shiga-Toxigénica/aislamiento & purificación , Infecciones por Escherichia coli , Proteínas de Escherichia coli/genética , Microbiología de Alimentos , Humanos , Serotipificación , Toxinas Shiga , Virulencia/genética
19.
Open Forum Infect Dis ; 2(4): ofv134, 2015 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-26484357

RESUMEN

Shiga toxins (Stx) are commonly produced by Shigella dysenteriae serotype 1 and Stx-producing Escherichia coli. However, the toxin genes have been detected in additional Shigella species. We recently reported the emergence of Stx-producing Shigella in travelers in the United States and France who had recently visited Hispaniola (Haiti and the Dominican Republic). In this study, we confirm this epidemiological link by identifying Stx-producing Shigella from Haitian patients attending clinics near Port-au-Prince. We also demonstrate that the bacteriophage encoding Stx is capable of dissemination to stx-negative Shigella species found in Haiti, suggesting that Stx-producing Shigella may become more widespread within that region.

20.
Appl Environ Microbiol ; 81(23): 8183-91, 2015 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-26386062

RESUMEN

Culture-independent diagnostics reduce the reliance on traditional (and slower) culture-based methodologies. Here we capitalize on advances in next-generation sequencing (NGS) to apply this approach to food pathogen detection utilizing NGS as an analytical tool. In this study, spiking spinach with Shiga toxin-producing Escherichia coli (STEC) following an established FDA culture-based protocol was used in conjunction with shotgun metagenomic sequencing to determine the limits of detection, sensitivity, and specificity levels and to obtain information on the microbiology of the protocol. We show that an expected level of contamination (∼10 CFU/100 g) could be adequately detected (including key virulence determinants and strain-level specificity) within 8 h of enrichment at a sequencing depth of 10,000,000 reads. We also rationalize the relative benefit of static versus shaking culture conditions and the addition of selected antimicrobial agents, thereby validating the long-standing culture-based parameters behind such protocols. Moreover, the shotgun metagenomic approach was informative regarding the dynamics of microbial communities during the enrichment process, including initial surveys of the microbial loads associated with bagged spinach; the microbes found included key genera such as Pseudomonas, Pantoea, and Exiguobacterium. Collectively, our metagenomic study highlights and considers various parameters required for transitioning to such sequencing-based diagnostics for food safety and the potential to develop better enrichment processes in a high-throughput manner not previously possible. Future studies will investigate new species-specific DNA signature target regimens, rational design of medium components in concert with judicious use of additives, such as antibiotics, and alterations in the sample processing protocol to enhance detection.


Asunto(s)
Microbiología de Alimentos/métodos , Metagenómica/métodos , Escherichia coli Shiga-Toxigénica/aislamiento & purificación , Spinacia oleracea/microbiología , Inocuidad de los Alimentos , Límite de Detección , Sensibilidad y Especificidad , Análisis de Secuencia de ADN , Escherichia coli Shiga-Toxigénica/genética
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