Quantifying acquisition and transmission of Enterococcus faecium using genomic surveillance


Nosocomial acquisition and transmission of vancomycin-resistant Enterococcus faecium (VREfm) is the driver for E. faecium carriage in hospitalized patients, which, in turn, is a risk factor for invasive infection in immunocompromised patients. In the present study, we provide a comprehensive picture of E. faecium transmission in an entire sampled patient population using a sequence-driven approach. We prospectively identified and followed 149 haematology patients admitted to a hospital in England for 6 months. Patient stools (n = 376) and environmental swabs (n = 922) were taken at intervals and cultured for E. faecium. We sequenced 1,560 isolates (1,001 stool, 559 environment) and focused our genomic analyses on 1,477 isolates (95%) in the hospital-adapted clade A1. Of 101 patients who provided two or more stool samples, 40 (40%) developed E. faecium carriage after admission based on culture, compared with 64 patients (63%) based on genomic analysis (73% VREfm). Half of 922 environmental swabs (447, 48%) were positive for VREfm. Network analysis showed that, of 111 patients positive for the A1 clade, 67 had strong epidemiological and genomic links with at least one other patient and/or their direct environment, supporting nosocomial transmission. Six patients (3.4%) developed an invasive E. faecium infection from their own gut-colonizing strain, which was preceded by nosocomial acquisition of the infecting isolate in half of these. Two informatics approaches (subtype categorization to define phylogenetic clusters and the development of an SNP cut-off for transmission) were central to our analyses, both of which will inform the future translation of E. faecium sequencing into routine outbreak detection and investigation. In conclusion, we showed that carriage and environmental contamination by the hospital-adapted E. faecium lineage were hyperendemic in our study population and that improved infection control measures will be needed to reduce hospital acquisition rates.

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Fig. 1: Study participants and E. faecium culture positivity.
Fig. 2: E. faecium within host diversity.
Fig. 3: Frequency and time span of E. faecium subtypes.
Fig. 4: E. faecium transmission network.

Data availability

The whole-genome sequences from this study have been deposited at the European Nucleotide Archive (www.ebi.ac.uk/ena) under the study nos. PRJEB12937, PRJEB13191 and PRJEB13192. Individual accession nos. and isolate metadata are listed in Supplementary Data 1. Supplementary Data 2 includes the genetic and epidemiological links characterized in this study. Source data are provided with this paper.


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We thank the nurses and healthcare workers on both wards at CUH for assistance with sample collection and the ward matrons, A. Green and C. Cowling, for their support. We thank L. Chaparadza and R. Swayne for assistance with sample and clinical data collection. We thank L. Drumright, A. Chaudhry and the EPIC and Clinical Informatics teams for providing patient movement data. We thank the library construction, sequencing and Pathogen Informatics teams at the Wellcome Trust Sanger Institute for assistance with Illumina sequencing. The flocked swabs were donated by Copan Italia SpA. The present study presents independent research supported by the Health Innovation Challenge Fund (WT098600, HICF-T5-342), a parallel funding partnership between the Department of Health and the Wellcome Trust. The views expressed in this article are those of the author(s) and not necessarily those of the Department of Health or Wellcome Trust. T.G. is a Wellcome Trust Research Training Fellow (103387/Z/13/Z). F.C. is a Wellcome Trust Sir Henry Postdoctoral Fellow (201344/Z/16/Z). C.L. is a Wellcome Trust Sir Henry Postdoctoral Fellow (110243/Z/15/Z). M.E.T. is a Clinician Scientist Fellow supported by the Academy of Medical Sciences, the Health Foundation and the National Institute for Health Research (NIHR) Cambridge Biomedical Research Centre. S.J.P. is an NIHR Senior Investigator.

Author information




T.G. and S.J.P. designed the study, wrote the study protocol and case record forms, obtained ethical and research and development approvals for the study, and supervised the data collection. M.E.T. supported ethics approvals. T.G., C.L. and C.C. were responsible for collecting samples, and clinical and epidemiological data. T.G., C.L., B.B. and P.N. isolated and identified E. faecium. B.B. and P.N. undertook susceptibility testing and extracted genomic DNA. N.M.B. and D.A.E. provided access to E. faecium cultures in the routine diagnostic microbiology laboratory and expert opinion relating to infection control. T.G. and F.C. undertook the epidemiological and bioinformatic analyses with contributions from J.P. and K.R. B.B. undertook susceptibility testing to disinfectants with contributions from E.M.H. J.P. supervised the genomic sequencing. F.C. and S.J.P. wrote the first draft of the manuscript. S.J.P. supervised and managed the study. All authors had access to the data, and read, contributed and approved the final manuscript.

Corresponding authors

Correspondence to Francesc Coll or Sharon J. Peacock.

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Competing interests

N.M.B. is on the advisory board for Discuva Ltd. S.J.P. is a consultant to Specific Technologies. S.J.P., J.P. and F.C. are consultants for Next Gen Diagnostics. The remaining authors declare no competing interests.

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Extended data

Extended Data Fig. 1 E. faecium stool culture positivity during study.

Diagram showing E. faecium positivity in patients who provided stool samples within (left branch) or after (right branch) 48 hours from index admission. Subsequent boxes show numbers of patients positive or negative for AREfm and VREfm, and, for patients screened at least twice, whether their positivity status changed, suggestive of E. faecium acquisition. A total of 40 cases acquired E. faecium based on culture, either by acquiring any type of E. faecium after being negative for it (17 and 15 patients in the left and right arms, respectively) or VREfm after being already positive for AREfm (4 and 4 patients in the left and right arms, respectively). Abbreviations: AREfm, ampicillin-resistant E. faecium (which may be vancomycin susceptible or resistant); VREfm, vancomycin-resistant E. faecium.

Extended Data Fig. 2 Histogram of pairwise SNP differences between isolates of the same and different subtypes.

Histogram of pairwise SNP differences between 943 clade A1 isolates from stool samples. SNP differences between isolates from the same subtype are shown in dark grey, and between isolates in different subtypes in light grey. Source data

Extended Data Fig. 3 Chlorhexidine and isopropanol susceptibility among selected E. faecium isolates.

Chlorhexidine and isopropanol susceptibility testing results for a subset of phylogenetically representative E. faecium isolates (n=24 biologically independent samples) from the two major subtypes (15A/ST80 (n=3) and 47A/ST78 (n=3)), rest of subtypes in ‘clade A1’ (n=8) and ‘basal’ isolates (n=10) to clade A1. Each dot denotes the median MIC value (panels a and b) or median reduction in colony forming units (CFU) (panel c) across three independent replicates for each isolate tested. In the boxplots, the lower and upper hinges correspond to the first and third quartiles (the 25th and 75th percentiles), and the middle horizontal line to the median. P-values for two-tailed, unpaired Mann-Whitney are showed as NS (non-significant, P > 0.05), * (P < 0.05) or ** (P < 0.01). Source data

Extended Data Fig. 4 Exemplars of E. faecium transmission clusters.

Each row represents the hospital admission period(s) of patients with the exception of the top four rows, which show different environmental sources. Ward of admission is denoted as A or B, and the room numbered and color-coded. Visits to other hospital wards or areas are colored in grey. Positivity results for stool and environmental samples are shown as circles and squares, respectively. Blunt lines and arrowed lines are drawn to point to the putative sources of index and acquired subtypes respectively, the numbers adjacent to these lines indicating the minimum genetic distance observed between connected samples, which ranged from 0 to 6 SNPs. Solid and dotted lines denote strong and weak epidemiological links, respectively. (a) Exemplar of transmission cluster in the same ward (subtype 49A – ST1454). Strong genetic and epidemiological links point to transmission of this subtype in different rooms of ward B among patients D040, D037, D036, D044 and D041. Strong links to the hospital environment, including communal bathrooms and medical devices, suggest their involvement as reservoirs for onward transmission to patients. (b) Exemplar of transmission cluster spanning both hematology wards and involving 7 patients (subtype 26B – ST80). Strong genetic and epidemiological links point to transmission of this subtype in room A3 among patients C015, C023, C009 and D021, followed by spread in different rooms of ward B among patients D021, D022, D010 and D045.

Extended Data Fig. 5 Midpoint rooted maximum likelihood tree based on SNPs in the core genes of 1,560 E. faecium isolates.

E. faecium genomes (1,001 stool, 559 environmental) labeled by clade (B, A2, and A1), commonest sequence types (STs) (only those with more than 10 isolates shown), van genotype, source, ward of origin and month of isolation. Scale bar, ~10,000 SNPs. Source data

Supplementary information

Supplementary information

Supplementary Methods and Tables 1–5.

Reporting Summary

Supplementary Data 1

Isolate metadata, including European Nucleotide Archive accessions.

Supplementary Data 2

Details on transmission of subtypes and epidemiological links.

Source data

Source Data Fig. 2

Genetic diversity captured within stool samples and subtypes in the same patient.

Source Data Fig. 3

Subtype assignation and source for all clade A1 isolates.

Source Data Fig. 4

Cytoscape-compliant network files.

Source Data Extended Data Fig. 2

Pairwise SNP distances between clade A1 stool isolates.

Source Data Extended Data Fig. 3

Isopropanol and chlorhexidine MICs, and isopropanol tolerance measurements, for selected E. faecium isolates.

Source Data Extended Data Fig. 5

Maximum likelihood phylogenetic tree including all 1,560 E. faecium isolates.

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Gouliouris, T., Coll, F., Ludden, C. et al. Quantifying acquisition and transmission of Enterococcus faecium using genomic surveillance. Nat Microbiol 6, 103–111 (2021). https://doi.org/10.1038/s41564-020-00806-7

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