Research ArticleInfectious Disease

Bacterial virulence phenotypes of Escherichia coli and host susceptibility determine risk for urinary tract infections

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Science Translational Medicine  22 Mar 2017:
Vol. 9, Issue 382, eaaf1283
DOI: 10.1126/scitranslmed.aaf1283
  • Fig. 1. Phylogenetic distribution of UAEC strains from rUTI patients.

    The phylogenetic relatedness of the UAEC strains (n = 43; taxon labels in red) was contextualized within the broader phylogeny of reference E. coli strains (n = 46; taxon labels in black) by comparing the single-copy core genes of the strains using the RAxML algorithm. Reference E. coli strains that were associated with urinary disease (for example, cystitis, pyelonephritis, or asymptomatic bacteriuria) are set in boldface. Asterisks indicate UAEC strains chosen as representative isolates for their clonal clusters (fig. S2B). Bootstrap supports are indicated at internal nodes, and bootstrap values >95 have been removed. UAEC strains were found in four out of five E. coli clades (indicated by red and purple bars on the left). Black arrows indicate model UPEC strains commonly used in UTI research.

  • Fig. 2. Carriage of PUFs is enriched in both UAEC and non-UAEC strains from the B2 clade.

    (A) Comparisons between indicated groups were performed using the Mann-Whitney U test. Significant differences were calculated for each group, and significant results are indicated as follows: ***P < 0.001; ****P < 0.0001. (B) Clinical UAEC strains and reference E. coli strains that were not associated with urinary disease (non-UAEC) (x axis) were examined for the presence of 31 PUF sequences (y axis) using the Basic Local Alignment Search Tool (BLAST) and alignment-based searches. The binary presence of PUFs (indicated by black squares) was tallied for each strain and for each PUF (indicated in parentheses on both axes). Two-dimensional hierarchical clustering identified clusters of PUFs that tended to co-occur in UAEC strains (dendrogram along the y axis) and showed that PUF carriage was associated with phylogeny (dendrogram along the x axis; phylogeny indicated in the column labeled Clade).

  • Fig. 3. Increased PUF carriage does not correlate with increased colonization efficiency.

    (A) The colonization efficiencies of 21 representative UAEC strains and the model strain UTI89 from indicated clades were tested in C3H/HeN mice. Bacteria were enumerated from individual harvested bladders at 24 hpi (black boxes). Each UAEC strain was categorized as “deficient” (n = 4; blue stippled outline) at <104 CFU per bladder, “variable” (n = 5; orange outline) above and below 104 CFU per bladder, or “robust” (n = 12; green outline) at >104 CFU per bladder. The black broken horizontal line represents the limit of detection (LOD) of bacteria. Data presented represent the median (gray bar) of mouse inoculations for each strain. (B) No enrichment of PUF carriage, as measured by PUF scores, was found in comparisons of robust, variable, or deficient colonizer strains. Comparisons were performed with Mann-Whitney U test; horizontal solid bars indicate median values. (C) Using Spearman’s rank correlation (ρ statistic indicated at the top), there was no significant correlation between carriage of PUFs and bladder burden with UAEC strains at 24 hpi. Squares indicate the median bladder colonization of B2 (red) or non-B2 (purple) UAEC strains.

  • Fig. 4. Both B2 and non-B2 UAEC strains cause chronic cystitis in mice.

    Incidence of chronic cystitis, defined as persistent high-titer bacteriuria (>104 CFU/ml of urine), high-titer bladder colonization (>104 CFU per bladder), and chronic inflammation at 28 dpi, was measured for a subset of B2 and non-B2 UAEC strains infecting C3H/HeN mice. Both B2 and non-B2 UAEC strains could cause chronic cystitis in mice. PUF scores and incidence of chronic cystitis are indicated for each strain. Horizontal bars indicate median values.

  • Fig. 5. PUF carriage does not increase competitive advantage during coinfection.

    UAEC strains isolated from two patients (9 and 41) with different strain-induced rUTIs were differentially marked with antibiotic resistance markers. C3H/HeN mice were then coinfected with the supplanting strain (9.2p or 41.4p) at an equal dose with the enrollment strain from the same patient (9.1a or 41.1a, respectively). A competitive index was calculated from the ratio of the supplanting strains over their enrollment strains in the urine from each mouse at 24 hpi. Clades and PUF scores are indicated for each strain. Horizontal bars indicate median values.

  • Fig. 6. Differential expression of core genes by UAEC strains distinguishes robust from deficient colonizers.

    (A) PCA was used to cluster UAEC strains based on their expression of 3340 core genes under type 1 pili–inducing culture conditions used to culture bacteria before inoculation into mouse bladders. B2 strains separated from non-B2 strains along PC1, whereas robust colonizers separated from deficient colonizers along PC2. Linear regression identified the expression of 42 core genes under type 1 pili–inducing conditions, as correlated with bladder bacterial burden at 24 hpi in C3H/HeN mice after correction for false discovery (P < 0.1). These 42 genes included pathway genes mediating nutrient uptake, such as lamB (B), and chemotaxis, such as tar (C), shown here as representative examples. Data presented represent averages derived from three independent experiments that were corrected for variance in read depth between samples and gene length.

  • Fig. 7. Colonization efficiency by UAEC strains varies between mouse models.

    Select UAEC strains were inoculated into C3H/HeN mice (black squares and light gray bars) and C57BL/6 mice (black circles and dark gray bars). Bacteria were enumerated from individual harvested bladders at 24 hpi (black squares and circles). Median values for each infection are given (gray bars). Each UAEC infection of the two mouse strains was categorized separately as “deficient” (blue stippled outline), “variable” (orange outline), or “robust” (green outline). Bladder colonization between C3H/HeN and C57BL/6 mice was significantly different for strains 31.1a and 41.1a by Mann-Whitney U test, **P < 0.01.

  • Table 1. Summary of 43 UAEC isolates from women with rUTIs.
    Isolation time pointNumber of isolates
    Enrollment*14
    rUTI*18
    Before recurrence11
    Total43

    *Diagnosed UTI.

    †No UTI diagnosis.

    Supplementary Materials

    • www.sciencetranslationalmedicine.org/cgi/content/full/9/382/eaaf1283/DC1

      Materials and Methods

      Fig. S1. Sample collection and patient time line.

      Fig. S2. Measurement of UAEC gene carriage and nucleotide diversity in core genome.

      Fig. S3. Kidney colonization by B2 and non-B2 UAEC strains.

      Fig. S4. Gut-associated E. coli are poor colonizers in the C3H/HeN mouse model of UTI.

      Fig. S5. IBC formation by B2 and non-B2 UAEC strains.

      Fig. S6. Carriage of PAI IIUTI89 enhances competitive fitness in chronic UTI in C3H/HeN mice.

      Fig. S7. Hemagglutination is correlated to colonization efficiency in both B2 and non-B2 UAEC strains.

      Fig. S8. Differentially expressed genes in UAEC relative to UTI89.

      Fig. S9. Expression of core genes under type 1 pili–inducing culture conditions correlates with bladder colonization in UAEC.

      Fig. S10. Motility does not correlate with colonization efficiency in either B2 or non-B2 UAEC strains.

      Fig. S11. Both B2 and non-B2 UAEC strains are capable of forming persistent bladder reservoirs in C57BL/6 mice.

      Table S1. Clinical information of enrolled patients.

      Table S2. Characteristics of UAEC isolates.

      Table S3. Reference E. coli strains.

      Table S4. List of PUFs.

      Table S5. Gut-associated E. coli strain characteristics.

      Table S6. IBC formation in select UAEC strains at 6 hpi in C3H/HeN mice.

      Table S7. Presence of brnAT genes in robust and deficient colonizer UAEC strains.

      Table S8. Reads mapped to core genome of UAEC strains.

      Table S9. DESeq analysis of UAEC transcription of core genome.

      Table S10. Expression of PUF genes in select UAEC strains.

      Table S11. Linear correlation of core gene expression with bladder colonization.

      Table S12. Functions enriched in genes associated with bladder colonization.

      Table S13. UAEC strain clinical data.

      Table S14. UAEC strain sequencing and genome assembly results.

      References (8286)

    • Supplementary Material for:

      Bacterial virulence phenotypes of Escherichia coli and host susceptibility determine risk for urinary tract infections

      Henry L. Schreiber IV, Matt S. Conover, Wen-Chi Chou, Michael E. Hibbing, Abigail L. Manson, Karen W. Dodson, Thomas J. Hannan, Pacita L. Roberts, Ann E. Stapleton, Thomas M. Hooton, Jonathan Livny,* Ashlee M. Earl,* Scott J. Hultgren*

      *Corresponding author. Email: livny{at}broadinstitute.org (J.L.); aearl{at}broadinstitute.org (A.M.E.); hultgren{at}wusm.wustl.edu (S.J.H.)

      Published 22 March 2017, Sci. Transl. Med. 9, eaaf1283 (2017)
      DOI: 10.1126/scitranslmed.aaf1283

      This PDF file includes:

      • Materials and Methods
      • Fig. S1. Sample collection and patient time line.
      • Fig. S2. Measurement of UAEC gene carriage and nucleotide diversity in core genome.
      • Fig. S3. Kidney colonization by B2 and non-B2 UAEC strains.
      • Fig. S4. Gut-associated E. coli are poor colonizers in the C3H/HeN mouse model of UTI.
      • Fig. S5. IBC formation by B2 and non-B2 UAEC strains.
      • Fig. S6. Carriage of PAI IIUTI89 enhances competitive fitness in chronic UTI in C3H/HeN mice.
      • Fig. S7. Hemagglutination is correlated to colonization efficiency in both B2 and non-B2 UAEC strains.
      • Fig. S8. Differentially expressed genes in UAEC relative to UTI89.
      • Fig. S9. Expression of core genes under type 1 pili–inducing culture conditions correlates with bladder colonization in UAEC.
      • Fig. S10. Motility does not correlate with colonization efficiency in either B2 or non-B2 UAEC strains.
      • Fig. S11. Both B2 and non-B2 UAEC strains are capable of forming persistent bladder reservoirs in C57BL/6 mice.
      • Table S1. Clinical information of enrolled patients.
      • Table S2. Characteristics of UAEC isolates.
      • Table S3. Reference E. coli strains.
      • Table S4. List of PUFs.
      • Table S5. Gut-associated E. coli strain characteristics.
      • Table S6. IBC formation in select UAEC strains at 6 hpi in C3H/HeN mice.
      • Table S7. Presence of brnAT genes in robust and deficient colonizer UAEC strains.
      • Table S8. Reads mapped to core genome of UAEC strains.
      • Legend for table S9
      • Table S10. Expression of PUF genes in select UAEC strains.
      • Legends for tables S11 and S12
      • Table S13. UAEC strain clinical data.
      • Table S14. UAEC strain sequencing and genome assembly results.
      • References (8286)

      [Download PDF]

      Other Supplementary Material for this manuscript includes the following:

      • Table S9 (Microsoft Excel format). DESeq analysis of UAEC transcription of core genome.
      • Table S11 (Microsoft Excel format). Linear correlation of core gene expression with bladder colonization.
      • Table S12 (Microsoft Excel format). Functions enriched in genes associated with bladder colonization.

      [Download Tables S9, S11, and S12]

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