Uropathogens Preferrentially Interact with Conditioning Film Components on the Surface of Indwelling Ureteral Stents Rather than Stent Material

Abstract

Despite routine implementation in urology, indwelling ureteral stents pose as a nidus for infection. Conditioning film accumulates on stents, which prime pathogen adhesion, promoting infectious biofilm formation. However, the extent to which conditioning film components play a role in facilitating bacterial adhesion and biofilm formation remains largely unknown. Here, we examined the interaction of previously identified stent-bound conditioning film components (fibrinogen, uromodulin, and albumin) with bacterial uropathogens. Cytoscopically removed stents were incubated with common uropathogens (Escherichia coli, Enterococcus faecalis, and Staphylococcus aureus). Immunofluorescent double staining was performed to study the localization of uropathogens relative to stent-bound conditioning film proteins. Conditioning film components were identified on the external stent surface with some deposition in the inner lumen. Bacteria co-localized with fibrinogen, uromodulin, and albumin, suggesting a potential mechanism for stent-associated infections. Here, we determine strong co-localization between common uropathogenic bacterial species with prominent conditioning film components on ureteral stents. Further functional validation of interactions amongst these uropathogens and conditioning film proteins may enhance clinical management for stent-associated infections and development of improved stent technologies.

Keywords: biofilm; ureteral stent; urinary tract infection.

Figure 1

Biofilm formation on a device.

Figure 2

Illustration of an indwelling ureteral stent anchored in place via characteristic “pig curls” in the kidney and bladder. Indwelling ureteral stents are designed to bridge an obstruction and maintain adequate urine flow by dilating the ureter and creating a space between the device and the ureteral wall. Obstruction can be caused by a kidney stone, ingrowth of tissue (stricture), or postoperative inflammation.

Figure 3

Representative scanning electron micrographs of ureteral stents with bacterial biofilm illustrating the random distribution of bacterial colonization on the surface of stents that were indwelling in patients. All images have a magnification of 5000×. (A) Enterococcus faecalis, (B) Escherichia coli, (C) Staphylococcus aureus (Additional SEM images please see Supplementary Materials).

Figure 4

Confocal microscopy reveals bacteria localized to stent surface and lumen (axial sections of the stent). (A) Control stent incubated with no bacteria. Panels (B–D) are representative images illustrating co-localization of E. coli. Panel (B) shows the distribution of albumin (Green Fluorecent Protein (GFP)-tagged anti-albumin antibody), while Panel (C) shows the distribution of E. coli (Red Fluorescent Protein (RFP)-tagged anti-E. coli antibody), and Panel (D) is the merged image showing co-localization (yellow). Arrows indicate bacterial colonization on the internal and external stent surfaces. Magnification 20×.

Figure 5

Representative confocal microscopy images showing the distribution of common conditioning film components on indwelling stents and co-localization of E. faecalis with these components. The distribution of fibrinogen and albumin is not uniform across the stent surface, while that of uromodulin, the most common protein found in urine, has a broader distribution.

Figure 6

Representative confocal microscopy images illustrating the presence of encrustation on the surface of indwelling stents. Albumin (A) was found to be part of the encrustation with which E. coli (B) was found to co-localize (C) following adhesion experiments. In addition, encrustation was found to contain uromodulin with which S. aureus was found to co-localize (D). Albumin and Uromodulin were detected using GFP-tagged anti-albumin and anti-uromodulin antibodies, respectively, while E. coli and S. aureus were detected using RFP-tagged anti-E. coli and anti-S. aureus antibodies, respectively.

Figure 7

Confocal microscopy images showing co-localization of S. aureus (red) with fibrinogen (green) (A), (B) E. coli (red) with albumin (green), and (C) E. faecalis (red) with uromodulin (green). Deconvolution processing of the images provides a clearer view of the co-localization and verifies the nonuniform distribution of conditioning film components on the stent surface as well as the nonuniform interaction of uropathogens with conditioning film components.

Figure 8

Illustration of the interaction between bacteria and the deposited urinary conditioning film or encrustation on the surface of indwelling stents. Bacterial adhesion to conditioning film components occurs via the interaction of adhesins and conditioning film proteins, while adhesion to encrustation is facilitated via conditioning film proteins and the uneven surface of the crystals.