Phylogenetic Diversity of the Enteric Pathogen Salmonella enterica subsp. enterica Inferred from Genome-Wide Reference-Free SNP Characters

Timme, Ruth; Pettengill, James B.; Allard, Marc W.; Strain, Errol; Barrangou, Rodolphe; Wehnes, Chris; Kessel, J.S. Van; Karns, Jeffrey S.; Musser, Steven M.; Brown, Eric W.

doi:10.1093/gbe/evt159

Cited by 120 publications

(147 citation statements)

References 46 publications

(83 reference statements)

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“…Given the dynamic nature of CRISPR loci with respect to spacer acquisition, loss, and duplication, we hypothesized that the identification of major S. enterica serotype Enteritidis lineages by CRISPR-MVLST was due to the imprinting of exogenous genetic cues on the CRISPRs that reflect the different ecological origins of major lineages. However, a recent study that included various Salmonella serotypes suggested that such signals might not be phylogenetically informative at the species level due to factors such as horizontal gene transfer and acquisition of common CRISPRs by different lineages (35). Further studies are necessary to investigate the robustness and scope of CRISPR subtyping in detecting ecological and evolutionary patterns of Salmonella and other organisms.…”

Section: Discussionmentioning

confidence: 99%

Comparative Analysis of Subtyping Methods against a Whole-Genome-Sequencing Standard for Salmonella enterica Serotype Enteritidis

et al. 2015

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A retrospective investigation was performed to evaluate whole-genome sequencing as a benchmark for comparing molecular subtyping methods for Salmonella enterica serotype Enteritidis and survey the population structure of commonly encountered S. enterica serotype Enteritidis outbreak isolates in the United States. A total of 52 S. enterica serotype Enteritidis isolates representing 16 major outbreaks and three sporadic cases collected between 2001 and 2012 were sequenced and subjected to subtyping by four different methods: (i) whole-genome single-nucleotide-polymorphism typing (WGST), (ii) multiple-locus variablenumber tandem-repeat (VNTR) analysis (MLVA), (iii) clustered regularly interspaced short palindromic repeats combined with multi-virulence-locus sequence typing (CRISPR-MVLST), and (iv) pulsed-field gel electrophoresis (PFGE). WGST resolved all outbreak clusters and provided useful robust phylogenetic inference results with high epidemiological correlation. While both MLVA and CRISPR-MVLST yielded higher discriminatory power than PFGE, MLVA outperformed the other methods in delineating outbreak clusters whereas CRISPR-MVLST showed the potential to trace major lineages and ecological origins of S. enterica serotype Enteritidis. Our results suggested that whole-genome sequencing makes a viable platform for the evaluation and benchmarking of molecular subtyping methods. Salmonella enterica is currently the most common bacterial foodborne pathogen in the United States, causing over 1 million cases of illnesses annually, including approximately 20,000 hospitalizations and 400 deaths (1). Serotyping is commonly used to subtype strains below the species level for epidemiologic purposes. Salmonella enterica serotype Enteritidis was the serotype most commonly linked to foodborne outbreaks between 1998 and 2008 in the United States, with shell eggs being the major vehicle for foodborne transmission (2). In recent years, S. enterica serotype Enteritidis was also found to cause multistate outbreaks associated with other foods such as ground beef (2012), Turkish pine nuts (2011), and alfalfa and spicy sprouts (2011), in addition to shelled eggs (2010) (3).During outbreak investigations, it is critical to employ subtyping methods capable of distinguishing outbreak isolates from epidemiologically distinct but genetically related bacterial strains. Most S. enterica serotype Enteritidis isolates have been shown to be genetically homogeneous, making it difficult for conventional subtyping methods such as pulsed-field gel electrophoresis (PFGE), the current gold standard for strain-level Salmonella subtyping, to discriminate between strains (4, 5). Among the S. enterica serotype Enteritidis isolates reported to PulseNet (6), approximately 45% display a single PFGE pattern using XbaI (JEGX01.0004), rendering PFGE ineffective in some foodborne outbreak investigations. One strategy to improve subtype resolution is to target hypervariable regions (i.e., regions of the bacterial chromosome with less genetic stability) in the ba...

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Section: Discussionmentioning

confidence: 99%

Comparative Analysis of Subtyping Methods against a Whole-Genome-Sequencing Standard for Salmonella enterica Serotype Enteritidis

et al. 2015

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“…All three proteins are virulence determinants in Salmonella (43)(44)(45)(46). WGS has already proved its usefulness for elucidating the evolutionary diversity of large populations of bacterial isolates (11,47,48). In the specific case of Salmonella, WGS was successfully applied to illuminate the diversity of the pathogen within a vast epidemic episode, allowing highly efficient traceback of clinical and food isolates (4,13).…”

Section: Discussionmentioning

confidence: 99%

“…Large studies based on WGS within S. enterica subspecies (10) and within serovars in S. enterica subsp. enterica (11,12) contributed to the elucidation of Salmonella phylogenetic diversity and also accomplished important steps forward in the area of bacterial disease tracking. Moreover, serovar-specific studies on S. enterica subsp.…”

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confidence: 99%

Differential Single Nucleotide Polymorphism-Based Analysis of an Outbreak Caused by Salmonella enterica Serovar Manhattan Reveals Epidemiological Details Missed by Standard Pulsed-Field Gel Electrophoresis

et al. 2015

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We retrospectively analyzed a rare Salmonella enterica serovar Manhattan outbreak that occurred in Italy in 2009 to evaluate the potential of new genomic tools based on differential single nucleotide polymorphism (SNP) analysis in comparison with the gold standard genotyping method, pulsed-field gel electrophoresis. A total of 39 isolates were analyzed from patients (n ‫؍‬ 15) and food, feed, animal, and environmental sources (n ‫؍‬ 24), resulting in five different pulsed-field gel electrophoresis (PFGE) profiles. Isolates epidemiologically related to the outbreak clustered within the same pulsotype, SXB_BS.0003, without any further differentiation. Thirty-three isolates were considered for genomic analysis based on different sets of SNPs, core, synonymous, nonsynonymous, as well as SNPs in different codon positions, by Bayesian and maximum likelihood algorithms. Trees generated from core and nonsynonymous SNPs, as well as SNPs at the second and first plus second codon positions detailed four distinct groups of isolates within the outbreak pulsotype, discriminating outbreak-related isolates of human and food origins. Conversely, the trees derived from synonymous and third-codon-position SNPs clustered food and human isolates together, indicating that all outbreak-related isolates constituted a single clone, which was in line with the epidemiological evidence. Further experiments are in place to extend this approach within our regional enteropathogen surveillance system. Salmonellosis is a major food-borne disease worldwide, with an estimated 93.8 million cases occurring each year, resulting in 155,000 deaths (1). The European Union summary report on trends and sources of zoonoses, zoonotic agents and food-borne outbreaks (2) indicated that nontyphoid salmonellosis was the second most reported food-borne zoonosis in Europe in 2012, trailing only behind Campylobacter jejuni infection. The 2012 overall notification rate for human salmonellosis in the European Union (EU) was 22.2 episodes per 100,000 population, for a total of 91,034 confirmed cases, with hospitalization and mortality rates of 45.1% and 0.14%, respectively. The highest proportions of Salmonella-positive foodstuff samples were reported for fresh turkey, poultry, and pork at 4.4%, 4.1%, and 0.7%, respectively (2). In order to manage this food-borne infection and to limit its health and economic burdens, surveillance programs have developed and implemented DNA-based subtyping methods to identify outbreaks in a timely manner and to trace infections back to their food sources. Over the past decades, the two most intensively used protocols for Salmonella subtyping have been pulsed-field gel electrophoresis (PFGE) and multilocus variable-number tandemrepeat analysis (MLVA) (3). Unfortunately, these methods rely on just few features of the entire bacterial genome (rare restriction sites for PFGE or few polymorphic loci for MLVA) to assess the relatedness of different isolates. During epidemiological investigations of food-borne outbreaks, this limitation mi...

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“…There are several advantages to using kSNP, which is a reference-free method of identifying genome-wide SNPs. First, the kSNP approach to gathering reference-free SNPs for downstream phylogenetic analysis has been shown to be effective for microbial-scale phylogenomic analysis (26,27). Second, it is not sensitive to assembly error because the putative SNPs are extracted from k-mers (one SNP per 25-mer in our analysis), which effectively eliminates any influence of assembly.…”

Section: Methodsmentioning

confidence: 99%

Whole-Genome Single-Nucleotide-Polymorphism Analysis for Discrimination of Clostridium botulinum Group I Strains

González-Escalona

Timme

Raphael

et al. 2014

Appl Environ Microbiol

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c Clostridium botulinum is a genetically diverse Gram-positive bacterium producing extremely potent neurotoxins (botulinum neurotoxins A through G [BoNT/A-G]). The complete genome sequences of three strains harboring only the BoNT/A1 nucleotide sequence are publicly available. Although these strains contain a toxin cluster (HA ؉ OrfX ؊ ) associated with hemagglutinin genes, little is known about the genomes of subtype A1 strains (termed HA ؊ OrfX ؉ ) that lack hemagglutinin genes in the toxin gene cluster. We sequenced the genomes of three BoNT/A1-producing C. botulinum strains: two strains with the HA ؉ OrfX ؊ cluster (69A and 32A) and one strain with the HA ؊ OrfX ؉ cluster (CDC297). Whole-genome phylogenic single-nucleotide-polymorphism (SNP) analysis of these strains along with other publicly available C. botulinum group I strains revealed five distinct lineages. Strains 69A and 32A clustered with the C. botulinum type A1 Hall group, and strain CDC297 clustered with the C. botulinum type Ba4 strain 657. This study reports the use of whole-genome SNP sequence analysis for discrimination of C. botulinum group I strains and demonstrates the utility of this analysis in quickly differentiating C. botulinum strains harboring identical toxin gene subtypes. This analysis further supports previous work showing that strains CDC297 and 657 likely evolved from a common ancestor and independently acquired separate BoNT/A1 toxin gene clusters at distinct genomic locations.

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Phylogenetic Diversity of the Enteric Pathogen Salmonella enterica subsp. enterica Inferred from Genome-Wide Reference-Free SNP Characters

Cited by 120 publications

References 46 publications

Comparative Analysis of Subtyping Methods against a Whole-Genome-Sequencing Standard for Salmonella enterica Serotype Enteritidis

Comparative Analysis of Subtyping Methods against a Whole-Genome-Sequencing Standard for Salmonella enterica Serotype Enteritidis

Differential Single Nucleotide Polymorphism-Based Analysis of an Outbreak Caused by Salmonella enterica Serovar Manhattan Reveals Epidemiological Details Missed by Standard Pulsed-Field Gel Electrophoresis

Whole-Genome Single-Nucleotide-Polymorphism Analysis for Discrimination of Clostridium botulinum Group I Strains

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