Proteomics analysis of urine and catheter-associated biofilms in spinal cord injury patients

Abstract

After spinal cord injury (SCI), use chronic urinary catheters for bladder management is common, making these patients especially vulnerable to catheter-associated complications. Chronic catheterization is associated with bacterial colonization and frequent catheter-associated urinary tract infections (CAUTI). One determinant of infection success and treatment resistance is production of catheter-associated biofilms, composed of microorganisms and host- and microbial-derived components. To better understand the biofilm microenvironment, we performed proteomics analysis of catheter-associated biofilms and paired urine samples from four people with SCI with chronic indwelling urinary catheters. We developed a novel method for the removal of adhered cellular components on catheters that contained both human and microbial homologous proteins. Proteins from seven microbial species were identified including: Escherichia coli, Klebsiella species (spp), Enterococcus spp, Proteus mirabilis, Pseudomonas spp, Staphylococcus spp, and Candida spp. Peptides identified from catheter biofilms were assigned to 4,820 unique proteins, with 61% of proteins assigned to the biofilm-associated microorganisms, while the remainder were human-derived. Contrastingly, in urine, only 51% were assigned to biofilm-associated microorganisms and 4,554 proteins were identified as a human-derived. Of the proteins assigned to microorganisms in the biofilm and paired urine, Enterococcus, Candida spp, and P. mirabilis had greater associations with the biofilm phase, whereas E. coli and Klebsiella had greater associations with the urine phase, thus demonstrating a significant difference between the urine and adhered microbial communities. The microbial proteins that differed significantly between the biofilm and paired urine samples mapped to pathways associated with amino acid synthesis, likely related to adaptation to high urea concentrations in the urine, and growth and protein synthesis in bacteria in the biofilm. Human proteins demonstrated enrichment for immune response in the catheter-associated biofilm. Proteomic analysis of catheter-associated biofilms and paired urine samples has the potential to provide detailed information on host and bacterial responses to chronic indwelling urinary catheters and could be useful for understanding complications of chronic indwelling catheters including CAUTIs, urinary stones, and catheter blockages.

Keywords: Catheter; biofilm; microbial proteins; proteomics; spinal cord injury; urine.

Figure 1

Summary of Identified Proteins. A. Cumulative protein masses annotated by source of origin expressed in terms of log₁₀ of iBAQ. B. Protein distribution by human (yellow) vs microorganism (purple) populations in catheter biofilms (left) and urine (right). The bottom bar graphs show percentage of proteins assigned to microorganism in catheter and urine. C. Bar graphs of protein mass for individual matched catheterized patient urine and biofilm (catheter) samples, and three urine samples collected from non-catheterized patients, broken down by source of origin.

Figure 2

Heat map of microorganism proteins that differ significantly between urine and catheter biofilms (P < 0.01, Np > 1). The intensity of the color represents the concentration change (log₂) from -4 (blue) to 4 (red), determined by label-free quantification.

Figure 3

Treemap of orthogonal protein classification according to TIGRFAM of the three populations of microorganisms that differ most between paired catheter (left) and urine (right) specimens based on protein mass.

Figure 4

Proteins with significant differences in levels between catheter-associated biofilms and paired urine samples. (A) Heatmap of human proteins (P < 0.01, Np > 1) with higher levels in catheter-associated biofilms compared to urine, and (B) higher levels in urine compared to catheter-associated biofilms. (C) Overrepresentation analysis demonstrating functional pathways according to Panther Pathways. Significant Panther Pathways (FDR < 0.05), according to Fisher’s Exact test and corrected using FDR with four or more proteins.