Fleck, R. C. | Search bioRxiv

bioRxiv2026

Emerging Pathogens in Urinary Tract Infections: Virulence and Phenotypic Characterization of Pseudomonas aeruginosa strains

Urinary tract infections (UTIs) affect a broad patient population and inflict a substantial financial burden on the U.S. healthcare system. While uropathogenic Escherichia coli (UPEC) causes the majority of cases, other pathogens are emerging. Analysis of patient data from our healthcare system in the Gulf Coast region of Alabama revealed that Pseudomonas aeruginosa accounted for 4.0% of UTI cases, roughly double the national average, prompting further investigation into this historically understudied uropathogen. Here, we performed whole-genome sequencing and phenotypic assays on 55 urinary P. aeruginosa isolates to identify key drivers of pathogenicity in the context of UTI. Multilocus sequence typing identified 19 novel sequence types, underscoring the uncharacterized diversity of urinary P. aeruginosa isolates. Serotype O6 was most common and enriched in patients with indwelling catheters, whereas O4 was linked to diabetes mellitus. Antibiotic susceptibility testing (AST) revealed high levofloxacin resistance (30.9%), with 23.6% multidrug-resistant (MDR) and 9.1% extensively drug-resistant (XDR) isolates. Resistance patterns correlated with demographics, including significantly higher meropenem and aztreonam resistance in isolates from African American patients. Phenotypic assays of growth, motility, and biofilm formation revealed negative correlations between antibiotic resistance and virulence. Specific virulence genes predicted enhanced iron acquisition, hemolysis, and colonization potential. Notably, motility and exotoxin profiles emerged as strong predictors of P. aeruginosa ascension in a murine UTI model. Together, these findings provide new biological and clinical insight into P. aeruginosa as a uropathogen and emphasize the need for continued research. IMPORTANCEPseudomonas aeruginosa is an emerging but understudied pathogen in urinary tract infections (UTIs). Given its resilience, adaptability, and the growing threat of multidrug resistance, P. aeruginosa remains a significant challenge in clinical microbiology and infection control. Our data reveal an increased prevalence of P. aeruginosa in our local patient population. In this study, we examined both genotypic and phenotypic traits of clinical isolates and correlated them with colonization in murine models and extensive patient metadata. We identified strong associations between antibiotic resistance patterns and patient demographics. Novel sequence type strains were linked to motility phenotypes in vitro. Additionally, specific flagellar alleles were associated with enhanced murine kidney colonization and recurrent UTIs in patients. These findings provide new insight into the evolutionary adaptations that contribute to P. aeruginosa uropathogenicity and support a more nuanced understanding of its clinical significance.

microbiology↗

bioRxiv2025

Human bladder organoids model urinary tract infection and bacteriophage therapy

Urinary tract infections (UTIs), primarily caused by uropathogenic Escherichia coli (UPEC), are among the most common antibiotic-resistant infections. Despite this, currently available preclinical UTI models lack the breadth of morphotypic and heterogenous cell populations of the human bladder, impairing the development of novel therapies. To address these limitations, we developed human bladder organoids derived from the bladder stem cells of multiple healthy donors which recapitulate cellular diversity of the urothelium. Using bulk and single cell RNA-sequencing, we characterized organoid responses to UPEC and phage exposure individually and in combination to model phage therapy. Although phage minimally affected the uroepithelium in the absence of infection, during UTI, phage treatment reduced bacterial burdens and dampened inflammatory responses and barrier disruption. Collectively, our findings highlight human bladder organoids as a tool for capturing conserved and individual-specific uroepithelial responses to infection while also providing preclinical efficacy and safety testing for therapeutic development.

microbiology↗

bioRxiv2024

Distinct maternofetal immune signatures delineate preterm birth onset following urinary tract infection

Preterm birth is the leading cause of infant mortality resulting in over one million neonatal deaths annually. Maternal urinary tract infection (UTI) during pregnancy increases risk for preterm birth; however, biological processes mediating UTI-associated preterm birth are not well-described. We established a murine maternal UTI model in which challenge with uropathogenic E. coli resulted in preterm birth in about half of dams. Dams experiencing preterm birth displayed excessive bladder inflammation and altered uteroplacental T cell polarization compared to non-laboring infected dams, with no differences in bacterial burdens. Additional factors associated with preterm birth included higher proportions of male fetuses and lower maternal serum IL-10. Furthermore, exogenous maternal IL-10 treatment absolved UTI-associated preterm birth but contributed to fetal growth restriction in this model. Using urine samples from a cohort of human pregnancies with or without UTI, we correlated urinary cytokines with birth outcomes and urine culture status. These analyses yielded a non-invasive, highly predictive three-model system for evaluating preterm birth risk implicating cytokines IL-10, IL-15, IL-1{beta}, and IL-1RA. Our unique bimodal murine model coupled with patient samples provides a platform to investigate immunological and microbial factors governing UTI-associated preterm birth, revealing novel therapeutic opportunities to predict or prevent preterm birth.

immunology↗