Dataset Information

Pyrosequencing-based comparative genome analysis of the nosocomial pathogen Enterococcus faecium and identification of a large transferable pathogenicity island.

ABSTRACT:

Background

The Gram-positive bacterium Enterococcus faecium is an important cause of nosocomial infections in immunocompromized patients.

Results

We present a pyrosequencing-based comparative genome analysis of seven E. faecium strains that were isolated from various sources. In the genomes of clinical isolates several antibiotic resistance genes were identified, including the vanA transposon that confers resistance to vancomycin in two strains. A functional comparison between E. faecium and the related opportunistic pathogen E. faecalis based on differences in the presence of protein families, revealed divergence in plant carbohydrate metabolic pathways and oxidative stress defense mechanisms. The E. faecium pan-genome was estimated to be essentially unlimited in size, indicating that E. faecium can efficiently acquire and incorporate exogenous DNA in its gene pool. One of the most prominent sources of genomic diversity consists of bacteriophages that have integrated in the genome. The CRISPR-Cas system, which contributes to immunity against bacteriophage infection in prokaryotes, is not present in the sequenced strains. Three sequenced isolates carry the esp gene, which is involved in urinary tract infections and biofilm formation. The esp gene is located on a large pathogenicity island (PAI), which is between 64 and 104 kb in size. Conjugation experiments showed that the entire esp PAI can be transferred horizontally and inserts in a site-specific manner.

Conclusions

Genes involved in environmental persistence, colonization and virulence can easily be aquired by E. faecium. This will make the development of successful treatment strategies targeted against this organism a challenge for years to come.

SUBMITTER: van Schaik W

PROVIDER: S-EPMC2858755 | biostudies-literature | 2010 Apr

REPOSITORIES: biostudies-literature

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Publications

Pyrosequencing-based comparative genome analysis of the nosocomial pathogen Enterococcus faecium and identification of a large transferable pathogenicity island.

van Schaik Willem W Top Janetta J Riley David R DR Boekhorst Jos J Vrijenhoek Joyce E P JE Schapendonk Claudia M E CM Hendrickx Antoni P A AP Nijman Isaäc J IJ Bonten Marc J M MJ Tettelin Hervé H Willems Rob J L RJ

BMC genomics 20100414

<h4>Background</h4>The Gram-positive bacterium Enterococcus faecium is an important cause of nosocomial infections in immunocompromized patients.<h4>Results</h4>We present a pyrosequencing-based comparative genome analysis of seven E. faecium strains that were isolated from various sources. In the genomes of clinical isolates several antibiotic resistance genes were identified, including the vanA transposon that confers resistance to vancomycin in two strains. A functional comparison between E. ...[more]

PMID: 20398277

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Project description:Enterococcus faecium is a gut commensal of humans and animals but is also listed on the WHO global priority list of multidrug-resistant pathogens. Many of its antibiotic resistance traits reside on plasmids and have the potential to be disseminated by horizontal gene transfer. Here, we present the first comprehensive population-wide analysis of the pan-plasmidome of a clinically important bacterium, by whole-genome sequence analysis of 1,644 isolates from hospital, commensal, and animal sources of E. faecium Long-read sequencing on a selection of isolates resulted in the completion of 305 plasmids that exhibited high levels of sequence modularity. We further investigated the entirety of all plasmids of each isolate (plasmidome) using a combination of short-read sequencing and machine-learning classifiers. Clustering of the plasmid sequences unraveled different E. faecium populations with a clear association with hospitalized patient isolates, suggesting different optimal configurations of plasmids in the hospital environment. The characterization of these populations allowed us to identify common mechanisms of plasmid stabilization such as toxin-antitoxin systems and genes exclusively present in particular plasmidome populations exemplified by copper resistance, phosphotransferase systems, or bacteriocin genes potentially involved in niche adaptation. Based on the distribution of k-mer distances between isolates, we concluded that plasmidomes rather than chromosomes are most informative for source specificity of E. faeciumIMPORTANCEEnterococcus faecium is one of the most frequent nosocomial pathogens of hospital-acquired infections. E. faecium has gained resistance against most commonly available antibiotics, most notably, against ampicillin, gentamicin, and vancomycin, which renders infections difficult to treat. Many antibiotic resistance traits, in particular, vancomycin resistance, can be encoded in autonomous and extrachromosomal elements called plasmids. These sequences can be disseminated to other isolates by horizontal gene transfer and confer novel mechanisms to source specificity. In our study, we elucidated the total plasmid content, referred to as the plasmidome, of 1,644 E. faecium isolates by using short- and long-read whole-genome technologies with the combination of a machine-learning classifier. This was fundamental to investigate the full collection of plasmid sequences present in our collection (pan-plasmidome) and to observe the potential transfer of plasmid sequences between E. faecium hosts. We observed that E. faecium isolates from hospitalized patients carried a larger number of plasmid sequences compared to that from other sources, and they elucidated different configurations of plasmidome populations in the hospital environment. We assessed the contribution of different genomic components and observed that plasmid sequences have the highest contribution to source specificity. Our study suggests that E. faecium plasmids are regulated by complex ecological constraints rather than physical interaction between hosts.

Project description:Enterococci are among the leading causes of hospital-acquired infections in the United States and Europe, with Enterococcus faecalis and Enterococcus faecium being the two most common species isolated from enterococcal infections. In the last decade, the proportion of enterococcal infections caused by E. faecium has steadily increased compared to other Enterococcus species. Although the underlying mechanism for the gradual replacement of E. faecalis by E. faecium in the hospital environment is not yet understood, many studies using genotyping and phylogenetic analysis have shown the emergence of a globally dispersed polyclonal subcluster of E. faecium strains in clinical environments. Systematic study of the molecular epidemiology and pathogenesis of E. faecium has been hindered by the lack of closed, complete E. faecium genomes that can be used as references.In this study, we report the complete genome sequence of the E. faecium strain TX16, also known as DO, which belongs to multilocus sequence type (ST) 18, and was the first E. faecium strain ever sequenced. Whole genome comparison of the TX16 genome with 21 E. faecium draft genomes confirmed that most clinical, outbreak, and hospital-associated (HA) strains (including STs 16, 17, 18, and 78), in addition to strains of non-hospital origin, group in the same clade (referred to as the HA clade) and are evolutionally considerably more closely related to each other by phylogenetic and gene content similarity analyses than to isolates in the community-associated (CA) clade with approximately a 3-4% average nucleotide sequence difference between the two clades at the core genome level. Our study also revealed that many genomic loci in the TX16 genome are unique to the HA clade. 380 ORFs in TX16 are HA-clade specific and antibiotic resistance genes are enriched in HA-clade strains. Mobile elements such as IS16 and transposons were also found almost exclusively in HA strains, as previously reported.Our findings along with other studies show that HA clonal lineages harbor specific genetic elements as well as sequence differences in the core genome which may confer selection advantages over the more heterogeneous CA E. faecium isolates. Which of these differences are important for the success of specific E. faecium lineages in the hospital environment remain(s) to be determined.

Project description:BackgroundVancomycin-resistant Enterococcus faecium (VREfm) is a leading cause of nosocomial infection. Here, we describe the utility of whole-genome sequencing in defining nosocomial VREfm transmission.MethodsA retrospective study at a single hospital in the United Kingdom identified 342 patients with E. faecium bloodstream infection over 7 years. Of these, 293 patients had a stored isolate and formed the basis for the study. The first stored isolate from each case was sequenced (200 VREfm [197 vanA, 2 vanB, and 1 isolate containing both vanA and vanB], 93 vancomycin-susceptible E. faecium) and epidemiological data were collected. Genomes were also available for E. faecium associated with bloodstream infections in 15 patients in neighboring hospitals, and 456 patients across the United Kingdom and Ireland.ResultsThe majority of infections in the 293 patients were hospital-acquired (n = 249) or healthcare-associated (n = 42). Phylogenetic analysis showed that 291 of 293 isolates resided in a hospital-associated clade that contained numerous discrete clusters of closely related isolates, indicative of multiple introductions into the hospital followed by clonal expansion associated with transmission. Fine-scale analysis of 6 exemplar phylogenetic clusters containing isolates from 93 patients (32%) identified complex transmission routes that spanned numerous wards and years, extending beyond the detection of conventional infection control. These contained both vancomycin-resistant and -susceptible isolates. We also identified closely related isolates from patients at Cambridge University Hospitals NHS Foundation Trust and regional and national hospitals, suggesting interhospital transmission.ConclusionsThese findings provide important insights for infection control practice and signpost areas for interventions. We conclude that sequencing represents a powerful tool for the enhanced surveillance and control of nosocomial E. faecium transmission and infection.

Project description:A variety of methods is used for a molecular typing of Enterococcus spp. and related gram-positive bacteria including macrorestriction analysis using pulsed-field gel electrophoresis (PFGE), ribotyping, rapid amplification of polymorphic DNA (RAPD), and amplified fragment length polymorphism (AFLP). To test the influence of transferable determinants on the outcome of different typing methods commonly used for enterococci, we established a homogenous strain collection of 24 transconjugants resulting from filter matings with antibiotic-resistant Enterococcus faecium. As expected, AFLP, RAPD, and PFGE all identified our model bacteria as strongly related. However, distinct differences in the resolving and discriminatory power of the tested methods could be clearly addressed. In PFGE, 22 of 24 transconjugants possessed less than a three-band difference to the recipient pattern and would be regarded as strongly related. Three different RAPD PCRs were tested; in two reactions, identical patterns for all transconjugants and the recipient were produced. One RAPD PCR produced an identical pattern for 18 transconjugants and the recipient and a clearly different pattern for the remaining 6 transconjugants due to a newly appearing fragment resulting from acquisition of the tetL gene. AFLP clusters all transconjugants into a group of major relatedness. Percent similarities were highly dependent on the method used for calculating the similarity coefficient (curve-based versus band-based similarity coefficient). Fragment patterns of digested plasmids showed the possession of nonidentical plasmids in most transconjugants. PFGE still could be recommended as the method of choice. Nevertheless, the more-modern AFLP approach produces patterns of comparable discriminatory power while possessing some advantages over PFGE (less-time-consuming internal standards). Plasmid fingerprints can be included to subdifferentiate enterococcal isolates possessing identical macrorestriction and PCR typing patterns.

Dataset Information

Pyrosequencing-based comparative genome analysis of the nosocomial pathogen Enterococcus faecium and identification of a large transferable pathogenicity island.

Background

Results

Conclusions

Publications

Pyrosequencing-based comparative genome analysis of the nosocomial pathogen Enterococcus faecium and identification of a large transferable pathogenicity island.

Similar Datasets

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