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Urinary microbiota and metabolome in pediatric vesicoureteral reflux and scarring

ABSTRACT: We enrolled girls and boys aged zero to nine years presenting to a pediatric urologist for recurrent urinary tract infection (UTI) or renal scarring (decreased uptake on a nuclear renal scan) or grade 3-5 vesico ureteral reflux (VUR). Exclusion criteria included other urogenital abnormalities, medical renal disease, immunodeficiency, syndromes associated with VUR, acute UTI, persistent UTI (ongoing positive urine culture 1-3 weeks after completing a treatment course), or global renal atrophy on imaging. At one patient visit, a urine specimen was collected for 16S and metabolomic analysis.

ORGANISM(S): Human Homo Sapiens

TISSUE(S): Urine

DISEASE(S): Urinary Tract Infection

SUBMITTER: Saurav Sarma

PROVIDER: ST001649 | MetabolomicsWorkbench | Fri Jan 01 00:00:00 GMT 2021

REPOSITORIES: MetabolomicsWorkbench

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Project description:The enteric bacterium Proteus mirabilis is a common cause of complicated urinary tract infections. In the study, microrarrays were used to analyze P. mirabilis gene expression in vivo from experimentally infected mice. Urine was collected at 1, 3, and 7d postinfection, and RNA was isolated from bacteria in the urine for transcriptional analysis. Across 9 microarrays, 471 genes were upregulated and 82 were downregulated in vivo compared to in vitro broth culture. Genes upregulated in vivo encoded MR/P fimbriae, urease, iron uptake systems, amino acid and peptide transporters, pyruvate metabolism, and portions of the TCA cycle. Flagella were downregulated. Ammonia assimilation gene glnA (glutamine synthetase) was repressed in vivo while gdhA (glutamate dehydrogenase) was upregulated in vivo. Contrary to our expectations, ammonia availability due to urease activity in P. mirabilis did not drive this gene expression. A gdhA mutant was growth-deficient in minimal medium with citrate as the sole carbon source, and loss of gdhA resulted in a significant fitness defect in the mouse model of urinary tract infection. Unlike Escherichia coli, which represses gdhA and upregulates glnA in vivo and cannot utilize citrate, the data suggest that P. mirabilis uses glutamate dehydrogenase to monitor carbon-nitrogen balance, and this ability contributes to the pathogenic potential of P. mirabilis in the urinary tract. Voided urine from female CBA/J mice infected with Proteus mirabilis was collected and pooled in RNA stabilizing reagent (RNAprotect). Urine was collected at 1, 3, and 7 d postinfection. RNA was isolated from urine and log-phase LB cultures, converted to cDNA, and labeled with CyDye. Three arrays were completed per time point (9 arrays total). Slides were scanned with a ScanArray Express microarray scanner (Perkin Elmer) at 10 μm resolution and quantified using ScanArray Express software. Resulting data were normalized by total intensity and median spot intensities were identified using MIDAS (v. 2.22) software.

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Urinary microbiota and metabolome in pediatric vesicoureteral reflux and scarring

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