A Semi-Quantitative Assay to Measure Glycosaminoglycan Degradation by the Urinary Microbiota

Abstract

Glycosaminoglycans (GAGs) are linear polysaccharides and are among the primary components of mucosal surfaces in mammalian systems. The GAG layer lining the mucosal surface of the urinary tract is thought to play a critical role in urinary tract homeostasis and provide a barrier against urinary tract infection (UTI). This key component of the host-microbe interface may serve as a scaffolding site or a nutrient source for the urinary microbiota or invading pathogens, but its exact role in UTI pathogenesis is unclear. Although members of the gut microbiota have been shown to degrade GAGs, the utilization and degradation of GAGs by the urinary microbiota or uropathogens had not been investigated. In this study, we developed an in vitro plate-based assay to measure GAG degradation and utilization and used this assay to screen a library of 37 urinary bacterial isolates representing both urinary microbiota and uropathogenic species. This novel assay is more rapid, inexpensive, and quantitative compared to previously developed assays, and can measure three of the major classes of human GAGs. Our findings demonstrate that this assay captures the well-characterized ability of Streptococcus agalactiae to degrade hyaluronic acid and partially degrade chondroitin sulfate. Additionally, we present the first known report of chondroitin sulfate degradation by Proteus mirabilis, an important uropathogen and a causative agent of acute, recurrent, and catheter-associated urinary tract infections (CAUTI). In contrast, we observed that uropathogenic Escherichia coli (UPEC) and members of the urinary microbiota, including lactobacilli, were unable to degrade GAGs.

Urinary Glycosaminoglycans are Associated with Recurrent UTI and Urobiome Ecology in Postmenopausal Women

“…13, 55, 56 Indeed, it is known that some uropathogens, such as Streptococcus agalactiae and Proteus mirabilis , can degrade CS. 15 We find it interesting to note that all the negative taxa-GAG associations were observed with CS, the most abundant urinary GAG. 8 Also of note, the presence of A. vaginae and species of Peptoniphilus , which were negatively associated with urinary CS, are known to be a signature of dysbiosis in the vagina.…”

“…16-20 Although species of the most predominant urobiome genus, Lactobacillus , cannot degrade CS, HS, or HA, Lactobacillus crispatus, Lactobacillus salivaris , and Lactobacillus reuteri have been shown to adhere to HeLa cells in a GAG-dependent manner. 15, 18, 21 These observations suggest that the urothelial GAG layer may serve as a scaffolding site for urobiome species.…”

“…14 Interestingly, while UPEC is unable to degrade GAGs, the uropathogenic bacterium Proteus mirabilis degrades CS, suggesting that some uropathogens may utilize the urothelial GAG layer during urinary tract colonization and invasion. 15 Additionally, bacterial genera often observed in the urinary microbiome (urobiome), like Bacteroides and Streptococcus are known to harbor the ability to metabolize GAGs like heparin and HA into smaller disaccharides. [16][17][18][19][20] Although species of the most predominant urobiome genus, Lactobacillus, cannot degrade CS, HS, or HA, Lactobacillus crispatus, Lactobacillus salivaris, and Lactobacillus reuteri have been shown to adhere to HeLa cells in a GAG-dependent manner.…”

Application of metabolomics in urolithiasis: the discovery and usage of succinate

“…The presence of inhibitors on crystal surface can hinder the formation and growth of the crystals. 35 Atomic force microscopy (AFM) of the crystal surface topology (Fig. 4e ) showed that the control and Taurine crystals had the smallest changes in depth (within 3 nm), although there are defects on the crystal surface.…”

Urinary Glycosaminoglycans Are Associated with Recurrent UTI and Urobiome Ecology in Postmenopausal Women

“…Interestingly, while UPEC is unable to degrade GAGs, the uropathogenic bacterium P. mirabilis degrades CS, suggesting that some uropathogens may utilize the urothelial GAG layer during urinary tract colonization and invasion . Additionally, bacterial genera often observed in the urinary microbiome (urobiome), like Bacteroides and Streptococcus, are known to harbor the ability to metabolize GAGs like heparin and HA into smaller disaccharides. − Although species of the most predominant urobiome genus, Lactobacillus, cannot degrade CS, HS, or HA, Lactobacillus crispatus, Lactobacillus salivaris, and Lactobacillus reuteri have been shown to adhere to HeLa cells in a GAG-dependent manner. ,, These observations suggest that the urothelial GAG layer may serve as a scaffolding site for urobiome species.…”