Project description:Proteus mirabilis is a leading cause of catheter-associated urinary tract infections (UTIs) and urolithiasis. The transcriptional regulator MrpJ inversely modulates two critical aspects of P. mirabilis UTI progression: fimbria-mediated attachment to the urinary tract, and flagella-mediated motility. Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) was used for the first time in a CAUTI pathogen to probe for in vivo direct targets of MrpJ. ChIP-seq revealed 81 78 direct MrpJ targets, including genes for motility, fimbriae and a type VI secretion system (T6SS), and the putative MrpJ binding sequence ACnCnnnnnnnGnGT.
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.
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.
Project description:Both Aerococcus urinae (Au) and Globicatella sanguinis (Gs) colonize the human urinary tract and are in the Aerococcaceae family. These rarely pathogenic Gram-positive bacteria were identified in polymicrobial urethral catheter biofilms (CBs) using 16S rDNA and proteomic analyses in this study. For confirming the identities, Au and Gs strains were isolated from small blood agar colonies derived from the CB extracts. Longitudinal surveys of clinical urine specimens revealed their persistence in the urinary tract and recolonization of newly replaced catheters. Dominant CB cohabitating organisms were Enterobacteriaceae, especially Proteus mirabilis and Escherichia coli. The proteomes of Gs and Au profiled from the in vivo milieu suggest that their energy metabolisms rely on glycolytic, heterolactic fermentation and peptide catabolic pathways. Several PTS sugar uptake and oligopeptide ABC transport systems were also highly abundant in the in vivo proteomes of Au and Gs, indicative to adaptations to nutrients available in urine and exfoliated urothelial cells (protein and proteoglycan breakdown products). Differences in Au and Gs metabolisms pertained to citrate lyase and glycogen (only in the Gs proteome), use of Xfp to degrade D-xylulose-5’-phosphate, and synthesis pathways for enzyme cofactors pyridoxal 6’-phosphate and 4’-phosphopantothenate (the latter only in the Au proteome). Interestingly, predicted metal ion (ZnuA-like) uptake systems were abundant in Gs but not in Au in vivo. Au expressed two LPXTG-anchored surface proteins, one predicted to have a pilin D adhesion motif. We describe how two microorganisms not previously characterized metabolically adapt to the milieu in the catheterized human urinary tract. Whether they are true pathogens or bystanders in CBs needs further investigation.
Project description:Antimicrobial peptides (AMPs) serve key proposed roles in defending the urinary tract against invading uropathogens, but individual AMPs bearing greatest responsibility for these functions remain largely unknown. We identified RegIIIγ as the most transcriptionally upregulated AMP in bladder transcriptomes following uropathogenic Escherichia coli (UPEC) infection. We confirmed induction of RegIIIγ mRNA during cystitis and pyelonephritis by quantitative RT-PCR. Immunoblotting demonstrates increased bladder and urinary RegIIIγ protein levels following UPEC infection. Immunostaining localizes RegIIIγ protein to urothelial cells of infected bladders and kidneys. Human patients with cystitis and pyelonephritis exhibit increased urine levels of the orthologous HIP/PAP protein. Recombinant RegIIIγ protein does not demonstrate bactericidal activity toward UPEC in vitro, but does kill Staphylococcus saprophyticus in a dose-dependent manner. RegIIIγ knockout and control urinary tracts contain comparable bacterial burden following experimental inoculation of UPEC as well as Gram-positive uropathogens. Thus, while RegIIIγ and HIP/PAP expression occurs in human and murine UTI, their specific functions in the urinary tract remain uncertain.
Project description:Pseudomonas aeruginosa is one of the most frequent pathogen dominant in complicated urinary tract infections (UTI). To unravel the adaptation strategies of P. aeruginosa to the conditions in the urinary tract and to define the underlying regulatory network an artificial growth system mimicking the conditions in the urinary tract was established. Transcriptome analyses were used to investigate the physiological status of P. aeruginosa under this conditions.
Project description:RIVUR Trial participants had Agilent 1M probe and or Nimblegen 2.1M probe aCGH performed on genomic DNA. The study was designed to discover DNA copy number variations in genes critical in kidney/urinary tract development and urinary tract infection susceptibility. Reference DNA used is a single male sample
Project description:Pseudomonas aeruginosa is one of the most frequent pathogen dominant in complicated urinary tract infections (UTI). To unravel the adaptation strategies of P. aeruginosa to the conditions in the urinary tract and to define the underlying regulatory network an artificial growth system mimicking the conditions in the urinary tract was established. Transcriptome analyses were used to investigate the physiological status of P. aeruginosa under this conditions. We performed comparisons to identify genes induced under artificial urinary tract conditions to unravel the adaptive strategies and the underlying regulatory network used by Pseudomonas aeruginosa during urinary tract infections using Affimetrix GeneChips. Pseudomonas aeruginosa wild type strain PAO1 was grown in an artificial in vitro growth system mimicking the conditions in the urinary tract. Therefore, biofilms were grown on the surface of membrane filters placed on agar plates at 37 °C up to the late logarithmic state under aerobic and anaerobic conditions (incubated in an anaerobic beanch). An artificial urine medium (AUM) simulating the averaged urine of an human adult was used as nutrient souce. 10-fold diluted Luria Bertani (LB)-medium was used as reference medium. For growth under oxygen depletion the media were supplemented with 50 mM KNO3 to sustain anaerobic respiration. The biofilms were harveted at this time points and resuspsended in 0.9% (w/v) NaCl. The OD578 of biofilm suspension was 0.8 for all tested conditions. First comparison: Identification of genes induced or repressed under aerobic conditions in the P. aeruginosa wild type PAO1. Here we compared the transcriptome profile of P. aeruginosa PAO1 grown aerobically for 18 h to the late logarithmic phase in biofilms on AUM with the transcriptome profile of the PAO1 strain, which was grown aerobically for 18 h to the late logarithmic phase in biofilms on 10-fold diluted LB. Second comparison: Identification of genes induced or repressed under anaerobic conditions in the P. aeruginosa wild type PAO1. Here we compared the transcriptome profile of P. aeruginosa PAO1 grown anaerobically for 2 days up to the late logarithmic phase in biofilms on AUM supplemented with 50 mM nitrate with the transcriptome profile of the PAO1 strain, which was grown anaerobically for 2 days up to the late logarithmic phase in biofilms on 10-fold diluted LB supplemented with 50 mM nitrate.
Project description:Proteus mirabilis is a primary cause of complicated urinary tract infections (UTI). Surprisingly, iron acquisition systems have been poorly characterized in this uropathogen despite the urinary tract being iron-limited. In this report the transcriptome of strain HI4320, cultured under iron limitation, was examined using microarray analysis. Of genes upregulated at least 2-fold, 45 were statistically significant and comprise 21 putative iron-regulated systems. Two of these systems, PMI0229-0239 and PMI2596-2605, are organized in operons and appear to encode siderophore biosynthesis genes.
2010-09-09 | GSE18051 | GEO
Project description:Genome sequences of two Klebsiella aerogenes strain isolated from patients with urinary tract infection in Uganda