20 research outputs found
BarA-UvrY two-component system regulates virulence of uropathogenic E. coli CFT073.
No full textUropathogenic Escherichia coli (UPEC), a member of extraintestinal pathogenic E. coli, cause ∼80% of community-acquired urinary tract infections (UTI) in humans. UPEC initiates its colonization in epithelial cells lining the urinary tract with a complicated life cycle, replicating and persisting in intracellular and extracellular niches. Consequently, UPEC causes cystitis and more severe form of pyelonephritis. To further understand the virulence characteristics of UPEC, we investigated the roles of BarA-UvrY two-component system (TCS) in regulating UPEC virulence. Our results showed that mutation of BarA-UvrY TCS significantly decreased the virulence of UPEC CFT073, as assessed by mouse urinary tract infection, chicken embryo killing assay, and cytotoxicity assay on human kidney and uroepithelial cell lines. Furthermore, mutation of either barA or uvrY gene reduced the production of hemolysin, lipopolysaccharide (LPS), proinflammatory cytokines (TNF-α and IL-6) and chemokine (IL-8). The virulence phenotype was restored similar to that of wild-type by complementation of either barA or uvrY gene in trans. In addition, we discussed a possible link between the BarA-UvrY TCS and CsrA in positively and negatively controlling virulence in UPEC. Overall, this study provides the evidences for BarA-UvrY TCS regulates the virulence of UPEC CFT073 and may point to mechanisms by which virulence regulations are observed in different ways may control the long-term survival of UPEC in the urinary tract
UreR, the Transcriptional Activator of the Proteus mirabilis Urease Gene Cluster, Is Required for Urease Activity and Virulence in Experimental Urinary Tract Infections
No full textProteus mirabilis, a cause of complicated urinary tract infection, produces urease, an essential virulence factor for this species. UreR, a member of the AraC/XylS family of transcriptional regulators, positively activates expression of the ure gene cluster in the presence of urea. To specifically evaluate the contribution of UreR to urease activity and virulence in the urinary tract, a ureR mutation was introduced into P. mirabilis HI4320 by homologous recombination. The isogenic ureR::aphA mutant, deficient in UreR production, lacked measurable urease activity. Expression was not detected in the UreR-deficient strain by Western blotting with monoclonal antibodies raised against UreD. Urease activity and UreD expression were restored by complementation of the mutant strain with ureR expressed from a low-copy-number plasmid. Virulence was assessed by transurethral cochallenge of CBA mice with wild-type and mutant strains. The isogenic ureR::aphA mutant of HI4320 was outcompeted in the urine (P 0.004), bladder (P 0.016), and kidneys (P ≤ 0.001) 7 days after inoculation. Thus, UreR is required for basal urease activity in the absence of urea, for induction of urease by urea, and for virulence of P. mirabilis in the urinary tract
Uropathogenicity in Rats and Mice of Providencia Stuartii from Long-Term Catheterized Patients
No full text
Phase-contrast photomicrographs of human kidney (HK-2) cells.
No full text<p>Photomicrographs were taken 6 hr later from either untreated control (HK-2 cells) (A), or treated with supernatants from CFT073 WT (B), <i>csrA</i> (C), <i>barA</i> (D), <i>uvrY</i> (E), <i>hlyD</i> (F), <i>barA</i>/p-<i>barA</i> (G), <i>uvrY</i>/p-<i>uvrY</i> (H) and <i>hlyD</i>/p<i>-hlyD</i> (I). The rounding and detachment of cells in the treated monolayer are marked with arrow or arrowhead. The pictures were representative of three individual experiments.</p
Assessment of virulence of uropathogenic Escherichia coli type 1 fimbrial mutants in which the invertible element is phase-locked on or off
No full textType I fimbria is a proven virulence factor of uropathogenic Escherichia coli (UPEC), causing urinary tract infections. Expression of the fimbria is regulated at the transcriptional level by a promoter situated on an invertible element, which can exist in one of two different orientations. The orientation of the invertible element that allows the expression of type 1 fimbriae is defined as "on," and the opposite orientation, in which no transcription occurs, is defined as "off." During the course of a urinary tract infection, we have observed that the infecting E. coli population alternates between fimbriated and nonfimbriated states, with the fimbriated on orientation peaking at 24 h. We propose that the ability of the invertible element to switch orientations during infection is itself a virulence trait. To test this hypothesis, nucleotide sequence changes were introduced in the left inverted repeat of the invertible element of UPEC pyelonephritis strain CFF073 that locked the invertible elements permanently in either the on or the off orientation. The virulence of these mutants was assessed in the CBA mouse model of ascending urinary tract infection at 4, 24, 48, and 72 h postinoculation (hpi). We conducted independent challenges, in which bladders of mice were inoculated with either a single mutant or the wild type, and cochallenges, in which a mutant and the wild type were inoculated together to allow direct competition in the urinary tract. In both sets of experimental infections, the locked-off mutant was recovered from the urine, bladder, and kidneys in significantly lower numbers than the wild type at 24 hpi (P less than or equal to 0.05), demonstrating its attenuation. Conversely. the locked-on mutant was recovered in higher numbers than the wild type at 24 hpi (P less than or equal to 0.05), showing enhanced virulence of this mutant. No significant differences were seen between the mutants and wild type in the urine or the bladder at 48 or 72 hpi. However, the wild type outcompeted the locked-off mutant in the kidneys during the cochallenge experiment at 72 hpi (P = 0.009). Overall, these data suggest that the ability of the invertible element controlling type 1 fimbria expression to phase vary contributes significantly to virulence early (24 hpi) in the course of a urinary tract infection by UPEC an most profoundly influences colonization of the bladder
Visualization of <i>Proteus mirabilis</i> within the Matrix of Urease-Induced Bladder Stones during Experimental Urinary Tract Infection
No full textABSTRACT
The virulence of a urease-negative mutant of uropathogenic
Proteus mirabilis
and its wild-type parent strain was assessed by using a CBA mouse model of catheterized urinary tract infection. Overall, catheterized mice were significantly more susceptible than uncatheterized mice to infection by wild-type
P. mirabilis
. At a high inoculum, the urease-negative mutant successfully colonized bladders of catheterized mice but did not cause urolithiasis and was still severely attenuated in its ability to ascend to kidneys. Using confocal laser scanning microscopy and scanning electron microscopy, we demonstrated the presence of
P. mirabilis
within the urease-induced stone matrix. Alizarin red S staining was used to detect calcium-containing deposits in bladder and kidney tissues of
P. mirabilis
-infected mice.
</jats:p
Mutation in <i>barA</i> or <i>uvrY</i> reduces and <i>csrA</i> increases the cytotoxic effect to human kidney (HK-2) cells.
No full texta<p>Data obtained from three independent experiments with 6 replicates per condition.</p><p>(*, p≤0.01; **, p≤0.001, relative to WT, *** p≤0.0001, relative to <i>barA</i> mutant).</p
Mutation in <i>uvrY</i> gene down regulates inflammatory cytokine expression.
No full text<p>The values are mean ± SD of three independent experiments with triplicate samples.</p><p>The downward arrow indicates down regulation compared to the WT.</p><p>(*, p≤0.05; **, p≤0.01 compared to WT).</p
