Transcriptome Analysis of Two Strains of Proteus mirabilis with Swarming Migration Deficiency Isolated from Patients with Urinary Tract Infection.

Two rare strains of Proteus mirabilis with swarming migration deficiency were isolated from urine samples of two patients with urinary tract infections and were named as G121 and G137. Migration experiments showed that P. mirabilis HI4320 had typical migration on blood agar, while G121 and G137 had significantly weakened migration ability. Results of adhesion tests showed that the adhesion ability of G121 and G137 to the bladder epithelial cell line 5637 was significantly reduced. High-throughput sequencing and alignment analysis of the transcriptomes of the three P. mirabilis strains were conducted, with P. mirabilis HI4320 as the reference strain. Reverse transcription quantitative PCR (RT-qPCR) was used to verify differentially expressed genes. Results of transcriptome analysis and RT-qPCR showed that, compared to the HI4320 strain, genes related to flagellum and fimbria formation, dicarboxylate transport, and cystathionine and anthranilate metabolism were down-regulated in G121 and G137, while genes related to iron transport, molybdenum metabolism, and metalloprotease were up-regulated, suggesting that these genes may be involved in the migration ability and epithelial cell adhesion ability of P. mirabilis. These results provide important insight to the search for virulence genes and the screening of new antibacterial targets for P. mirabilis.

Analysis of inflammatory cytokine expression in the urinary tract of BALB/c mice infected with Proteus (P.) mirabilis and enteroaggregative Escherichia (E.) coli (EAEC) strains.

This study aimed to analyze the proinflammatory cytokine mRNA expression in the urinary tract of BALB/c mice infected with bacterial strains with uropathogenic potential. Groups of four 6-week-old female BALB/c mice were intraurethrally inoculated with 5 × 107 colony-forming units (CFU) of P. mirabilis ATCC29906, EAEC O42, P. mirabilis RTX339, or sterile saline (control group) and then sacrificed at 0, 2, 4, 7, or 10 days post-infection (p.i.). Samples were cultured to determine the CFU/mL in urine or CFU/g in the bladders and kidneys. Cytokine expression (tumor necrosis factor (TNF)-α and interleukin (IL)-1ß, -6, and -8) was evaluated in the target organs using real-time PCR and immunohistochemistry; histology was examined with hematoxylin and eosin staining. The results are presented as the means and standard deviations and were compared using one-way ANOVA, with p < 0.05 indicating significant differences. Bacteriuria was not detected in the infected groups; bacterial colonization occurred in the target organs at all time points, but was higher in mice infected with EAEC O42 or P. mirabilis RTX339 at 7 days p.i. The expression of all cytokine mRNAs was seen, but only the levels of the IL-8 protein increased in situ at 7 days p.i. in the P. mirabilis RTX339 and EAEC O42 groups in both organs. Morphological alterations, observed in all of the infected groups, were more prominent in the EAEC O42 and P. mirabilis RTX339 groups. The findings provide insights into the uropathogenicity and inflammatory cytokine expression in the urinary tract of mice infected with three previously untested bacterial strains.

Urine Cytokine and Chemokine Levels Predict Urinary Tract Infection Severity Independent of Uropathogen, Urine Bacterial Burden, Host Genetics, and Host Age.

Urinary tract infections (UTIs) are among the most common infections worldwide. Diagnosing UTIs in older adults poses a significant challenge as asymptomatic colonization is common. Identification of a noninvasive profile that predicts likelihood of progressing from urine colonization to severe disease would provide a significant advantage in clinical practice. We monitored colonization susceptibility, disease severity, and immune response to two uropathogens in two mouse strains across three age groups to identify predictors of infection outcome. Proteus mirabilis caused more severe disease than Escherichia coli, regardless of mouse strain or age, and was associated with differences in interleukin-1ß (IL-1ß), beta interferon (IFN-ß), CXCL5 (LIX), CCL5 (RANTES), and CCL2 (MCP-1). In a comparison of responses to infection across age groups, mature adult mice were better able to control colonization and prevent progression to kidney colonization and bacteremia than young or aged mice, regardless of mouse strain or bacterial species, and this was associated with differences in IL-23, CXCL1, and CCL5. A bimodal distribution was noted for urine colonization, which was strongly associated with bladder CFU counts and the magnitude of the immune response but independent of age or disease severity. To determine the value of urine cytokine and chemokine levels for predicting severe disease, all infection data sets were combined and subjected to a series of logistic regressions. A multivariate model incorporating IL-1ß, CXCL1, and CCL2 had strong predictive value for identifying mice that did not develop kidney colonization or bacteremia, regardless of mouse genetic background, age, infecting bacterial species, or urine bacterial burden. In conclusion, urine cytokine profiles could potentially serve as a noninvasive decision support tool in clinical practice and contribute to antimicrobial stewardship.

Pathogenesis of Proteus mirabilis Infection.

Proteus mirabilis, a Gram-negative rod-shaped bacterium most noted for its swarming motility and urease activity, frequently causes catheter-associated urinary tract infections (CAUTIs) that are often polymicrobial. These infections may be accompanied by urolithiasis, the development of bladder or kidney stones due to alkalinization of urine from urease-catalyzed urea hydrolysis. Adherence of the bacterium to epithelial and catheter surfaces is mediated by 17 different fimbriae, most notably MR/P fimbriae. Repressors of motility are often encoded by these fimbrial operons. Motility is mediated by flagella encoded on a single contiguous 54-kb chromosomal sequence. On agar plates, P. mirabilis undergoes a morphological conversion to a filamentous swarmer cell expressing hundreds of flagella. When swarms from different strains meet, a line of demarcation, a "Dienes line," develops due to the killing action of each strain's type VI secretion system. During infection, histological damage is caused by cytotoxins including hemolysin and a variety of proteases, some autotransported. The pathogenesis of infection, including assessment of individual genes or global screens for virulence or fitness factors has been assessed in murine models of ascending urinary tract infections or CAUTIs using both single-species and polymicrobial models. Global gene expression studies performed in culture and in the murine model have revealed the unique metabolism of this bacterium. Vaccines, using MR/P fimbria and its adhesin, MrpH, have been shown to be efficacious in the murine model. A comprehensive review of factors associated with urinary tract infection is presented, encompassing both historical perspectives and current advances.

Purple Urine Bag Syndrome: A Rare Spot Diagnosis.

Purple urine bag syndrome (PUBS) is a complication of urinary tract infections (UTIs) where catheter bags and tubing turn purple. It is alarming for patients, families, and clinicians; however, it is in itself a benign phenomenon. PUBS is the result of UTIs with specific bacteria that produce sulphatases and phosphatases which lead tryptophan metabolism to produce indigo (blue) and indirubin (red) pigments, a mixture of which becomes purple. Risk factors include female gender, immobility, constipation, chronic catheterisation, and renal disease. Management involves reassurance, antibiotics, and regular changing of catheters, although there are debates regarding how aggressively to treat and no official guidelines. Prognosis is good, but PUBS is associated with high morbidity and mortality due to the backgrounds of patients. Here, we review the literature available on PUBS, present a summary of case studies from the last five years, and propose the Oxford Urine Chart as a tool to aid such diagnoses.

In vitro studies on the role of glycosaminoglycans in crystallization intensity during infectious urinary stones formation.

Proteus mirabilis cause urinary tract infections which are recurrent and can lead to formation of urinary calculi. Both bacterial and the host factors are involved in the development of urolithiasis. To determine the impact of glycosaminoglycans (GAGs) in the formation of P. mirabilis-induced urinary stones, we investigated the in vitro crystallization, aggregation and adhesion of crystals in the presence of GAGs naturally appearing in urine. Crystallization experiments were performed in synthetic urine infected with P. mirabilis in the presence of: hyaluronic acid (HA), heparan sulfate (HS), chondroitin sulfate A, B and C (ChSA, ChSB, ChSC). The intensity of crystallization and aggregation were established by counting particles and phase-contrast microscopy. To analyze the adhesion of crystals, we used normal urothelium and (45)Ca isotope-labeled crystals. In the presence of ChSC, both the size of the crystals formed and their number were higher compared with the control. GAGs increased crystals adhesion to the cells, but only for ChSA this effect was significant. Chondroitin sulfates, which accelerate the first stages of infection-induced stones formation, may play an important role in the pathogenesis of infectious urolithiasis.

Increased serum procalcitonin levels in pregnant patients with asymptomatic bacteriuria.

BACKGROUND: Among the pregnancy urinary tract infections, asymptomatic bacteriuria (ASB) is the most common one. Untreated ASB can progress to pyelonephritis in 30-50% of the patients and can also result in prematurity in 27% of the pregnancy so it needs immediate diagnosis and treatment. In this study, we wanted to evaluate procalcitonin levels, compared to other inflammatory in pregnant women with ASB. METHODS: The study was designed between the period of January 2012 and February 2013 at Sakarya University School of Medicine, Department of Gynecology and Obstetrics. The study population included 30 pregnant patients with asymptomatic bacteriuria and 39 healthy pregnant controls. RESULTS: Mean age was 28 (SD, 5.5) of the study population; mean maternal weight was 70 (SD, 8) kilogram. There were no statically significant differences between the groups according to the routine biochemical parameters, but gestational age was significantly lower in the ASB group compared to the controls (20.4 vs 28.6, respectively; p < 0.001). Serum procalcitonin levels were negative in all of the controls. In ASB group, 9 (30%) patients had procalcitonin levels greater than >0.05 ng/ml and 21(70%) patients had negative procalcitonin levels (Chi-squrae, p < 0.001). The sensitivity and specificity of procalcitonin assay for ASB was calculated as 30% and 100%, respectively. The positive predictive value was 100% and the negative predictive value was 65%. The most frequent microorganisms in the urine culture were Escherichia coli (26 patients, 87%), Proteus mirabilis (3 patients, 10%) and Klebsiella (1 patient, 3%) in the ASB group. We experienced four (44%) recurrences among nine positive procalcitonin in ASB patients after completion of treatment of the first ASB diagnosis. DISCUSSION: Procalcitonin levels were significantly higher in ASB group than the control group and serum procalcitonin levels were higher in pregnant women with recurrent ASB. This finding is an important result revealed that high procalcitonin level can predict the further urinary tract infection risk. Finally, serum procalcitonin levels were normal in healthy pregnant women while other inflammatory markers such as WBC, ESR and CRP levels were higher.

Isolation, identification & characterization of Proteus penneri--a missed rare pathogen.

BACKGROUND & OBJECTIVES: Indole negative Proteus species are invariably incorrectly identified as P. mirabilis, missing isolates of Proteus penneri. P. penneri is an invasive pathogen capable of causing major infectious diseases still seldom reported in individual cases. We report here the isolation, differentiation, characterization and typing of P. penneri from patients with different clinical infections. METHODS: Urine, pus and body fluids collected from patients in intensive care units, wards and out patients departments of a tertiary health care institute from north India were cultured. A total of 61 indole negative Proteus isolates were subjected to extended biochemical tests to differentiate and identify P. penneri from P. mirabilis including failure to produce ornithine decarboxylase (by 0% strains of P. penneri and 100% strains of P. mirabilis) besides P. penneri being uniformly salicin negative, non-utilizer of citrate but ferments sucrose and maltose. Antibiograms and Dienes phenomenon were performed to characterize and type P. penneri isolates besides screening for ß-lactamase production. RESULTS: Eight isolates of P. penneri were identified; four from urine, three from abdominal drain-fluid and one from diabetic foot ulcer. P. penneri was isolated as the sole pathogen in all patients having underlying disease; post-operatively. Swarming was not seen in the first strain on primary isolation and was poor in strain-4. All eight isolates were biochemically homologous but multi-drug resistant (MDR) with resistance to 6-8 drugs (up to 12). ß-lactamase production was seen in three of five isolates while Dienes phenomenon found four distinct types and discriminated strains differing in resistance even with a single drug. INTERPRETATION & CONCLUSIONS: A few additional biochemical tests identified P. penneri isolates; it infected patients with underlying disease and strains were MDR and heterogenous.

Strategy to control catheter encrustation with citrated drinks: a randomized crossover study.

PURPOSE: Nucleation pH is the pH at which Ca and Mg come out of urine to form crystals. If the safety margin between voiding pH and nucleation pH could be increased, it would increase the possibility of an alternative to controlling the activity of urease producing bacteria as a strategy to control catheter encrustation. MATERIALS AND METHODS: We performed a 6-week randomized crossover study in 24 patients with catheter blockage who were randomly allocated to a specific sequence of 3 consecutive available treatments, including increased fluid intake, lemon juice and potassium citrate. Each patient received all available regimens. At the end of each week 24-hour urine samples were analyzed for voiding and nucleation pH, citrate, Ca and Mg. RESULTS: Mean +/- SD nucleation pH increased from 7.45 +/- 0.60 at baseline to 7.93 +/- 0.50, 7.68 +/- 0.64 and 7.96 +/- 0.37 in the lemon juice, increased fluid intake and potassium citrate groups, respectively (p <0.0001). Mean urinary citrate increased significantly (p <0.0001), in particular due to lemon juice and potassium citrate effects. The association between treatment and Ca was weak (p = 0.12) while that of Mg was negative due to lemon (p <0.001). Average increase in the safety margin (nucleation pH minus voiding pH) beyond baseline was 0.84 (95% CI 0.63-1.04), 0.57 (95% CI 0.37-0.78) and 0.41 (95% CI 0.20-0.61) for lemon juice, increased fluid intake and potassium citrate, respectively. A strong treatment effect on the safety margin was apparent even when controlling for study design (p <0.001). CONCLUSIONS: Increased fluid intake with lemon juice may be a simple, inexpensive, effective strategy to control catheter encrustation.

Asymptomatic bacteriuria during pregnancy.

OBJECTIVE: The present study was aimed to test the association between asymptomatic bacteriuria during pregnancy, among patients in whom antibiotic treatment was recommended, and perinatal outcome. Our study was also designed to characterize common bacteria and risk factors associated with asymptomatic bacteriuria during pregnancy. METHODS: A retrospective population-based study comparing all singleton pregnancies of women with and asymptomatic bacteriuria was conducted. Patients with urinary tract infection were excluded from the analysis. Multiple logistic regression model was used to control for confounders. RESULTS: Out of 199,093 deliveries, 2.5% (n = 4890) were in patients with asymptomatic bacteriuria. E. coli was the most common pathogen associated with asymptomatic bacteriuria, representing 78.6% of the cultures with specified growth. Patients with asymptomatic bacteriuria were more likely to deliver preterm (PTD, 13.3%vs. 7.6%, odd ratio (OR) = 1.9, 95% confidence interval CI 1.7-2.0; P < 0.001) and deliver low birth-weight neonates (<2500 g, 13.3%vs. 7.9%, OR = 1.8; 95% CI 1.6-2.0; P < 0.001). Asymptomatic bacteriuria was independently associated with PTD (adjusted OR = 1.6; 95% CI 1.5-1.7; P < 0.001), fertility treatments, hypertensive disorders, recurrent abortions, diabetes mellitus, intrauterine growth restriction, polyhydramnion and oligohydramnion, premature rupture of membranes and labour induction, in a multivariable analysis with backwards elimination. Perinatal mortality rates (1.5%vs. 1.4%; P = 0.707) as well as low 5 min Apgar scores (0.8%vs. 0.6%; P = 0.065) were comparable between the groups. CONCLUSION: Asymptomatic bacteriuria is an independent risk factor for preterm delivery.

Proteus bacteriuria is associated with significant morbidity in spinal cord injury.

STUDY DESIGN: Retrospective chart review. OBJECTIVES: We investigated the morbidity associated with Proteus bacteriuria in a spinal cord injured (SCI) population. SETTING: Michael E DeBakey Veterans Affairs Medical Center in Houston, Texas, USA. METHODS: We reviewed the medical records of all veterans with SCI who received care in our medical center during the past 3 years. Proteus bacteriuria was defined as the growth of Proteus species in any urine culture during the study period. Urinary stones were defined as either renal or bladder calculi. RESULTS: During the study period, 71 of the 501 subjects (14%) had Proteus and 90 (18%) had urinary stones. Twenty-seven percent of the subjects with Proteus had stones, and the association of Proteus with stones was significant (P<0.05). Proteus bacteriuria was likewise associated with complete injury, hospitalization, decubitus ulcers, and history of stones (P<0.001). Subjects using indwelling catheters, either transurethral or suprapubic, were significantly more likely to have Proteus, whereas subjects practising spontaneous voiding and clean intermittent catheterization were significantly less likely to have Proteus. In the 90 patients with stones, Proteus was associated with requiring treatment for stones and having multiple stones (P<0.01). Twenty-five of the 90 patients with stones (28%) required treatment, most often with lithotripsy, and 6 (7%) developed urosepsis. CONCLUSIONS: In persons with SCI, Proteus was found in subjects with a greater degree of impairment who were more likely to be hospitalized, to have decubiti, and to use indwelling catheters. Bacteriuria with Proteus predicted urologic complications in persons with SCI. STATEMENT OF ETHICS: All applicable institutional and governmental regulations concerning the ethical use of human volunteers were followed during the course of this research.

Prospective study of individuals with long-term urinary catheters colonized with Proteus species.

OBJECTIVES: To characterize the variability in the times catheters take to block with encrustation in patients who have Proteus in their urinary flora, and to identify factors responsible for modulating the rate of catheter encrustation and blockage. PATIENTS AND METHODS: Twenty patients were followed prospectively for > or = 12 weeks, with a bacteriological analysis on weekly urine samples. The pH of the voided urine samples and the pH at which crystals formed in them (the nucleation pH) were determined. Catheters were collected and examined for bacterial biofilm and crystal deposition. RESULTS: The time that catheters took to block was 2-98 days. The mean pH of the urine voided by patients designated as slow encrusters (6.9) was not significantly different (P = 0.237) from that of rapid encrusters (7.2). However, patients whose catheters took longer to block had a significantly higher mean nucleation pH (8.1 vs 7.3, P = 0.002) and significantly higher mean safety margin between their nucleation pH and voided pH (1.17 pH units vs 0.13, P = 0.003). CONCLUSION: The variation in the rate of catheter encrustation between individuals infected with Proteus is a function of the difference between the voided pH and the nucleation pH of their urine. The value of nucleation pH of an individual's urine varies widely, suggesting it should be possible to devise strategies to increase this value and thus reduce the rate of encrustation in those with urinary tract colonization by urease-positive bacteria.