Prospective cohort study of microbial and inflammatory events immediately preceding Escherichia coli recurrent urinary tract infection in women.

BACKGROUND: A prospective cohort study was conducted to characterize the temporal sequence of microbial and inflammatory events immediately preceding Escherichia coli recurrent urinary tract infection (rUTI). METHODS: Women with acute cystitis and a history of UTI within the previous year self-collected periurethral and urine samples daily and recorded measurements of urine leukocyte esterase, symptoms, and sexual intercourse daily for 3 months. rUTI strains were characterized by pulsed-field gel electrophoresis and genomic virulence profiling. Urinary cytokine levels were measured. RESULTS: There were 38 E. coli rUTIs in 29 of 104 women. The prevalence of periurethral rUTI strain carriage increased from 46% to 90% during the 14 days immediately preceding rUTI, with similar increases in same-strain bacteriuria (from 7% to 69%), leukocyte esterase (from 31% to 64%), and symptoms (from 3% to 43%), most notably 2-3 days before rUTI (P<.05 for all comparisons). Intercourse with periurethral carriage of the rUTI strain also increased before rUTI (P=.008). Recurrent UTIs preceded by bacteriuria, pyuria, and symptoms were caused by strains less likely to have P fimbriae than other rUTI strains (P=.002). CONCLUSIONS: Among women with frequent rUTIs, the prevalences of periurethral rUTI strain carriage, bacteriuria, pyuria, and intercourse dramatically increase over the days preceding rUTI. A better understanding of the pathogenesis of rUTI will lead to better prevention strategies.

Urinary tract infections in women with type 1 diabetes mellitus: survey of female participants in the epidemiology of diabetes interventions and complications study cohort.

PURPOSE: We determined the prevalence of and risk factors for urinary tract infection in women with type 1 diabetes, and compared the prevalence of cystitis to that in nondiabetic women. MATERIALS AND METHODS: Women enrolled in the Epidemiology of Diabetes Interventions and Complications study were surveyed at year 10 as part of the Uro-EDIC study to assess the prevalence of cystitis and pyelonephritis in the preceding 12 months. Multivariate logistic regression models including measures of glycemic control and vascular complications of type 1 diabetes were used for risk factor analyses. The prevalence of cystitis in Uro-EDIC women was compared to that in a nondiabetic subset of women participants in the National Health and Nutrition Examination Survey III (NHANES III). RESULTS: A total of 550 women participated in the Uro-EDIC survey. The prevalence of cystitis and pyelonephritis in the preceding 12 months was 15% and 3%, respectively. Duration of diabetes, hemoglobin A1C, retinopathy, neuropathy, nephropathy, composite vascular complication score and intensive glycemic therapy during the Diabetes Control and Complications Trial, and Diabetes Control and Complications Trial cohort were not associated with cystitis or pyelonephritis. Sexual activity was associated with increased cystitis risk (adjusted OR 8.28; 95% CI 1.45, 158.32; p = 0.01). The adjusted prevalence of cystitis was 19.1% in Uro-EDIC women and 23.1% in NHANES III participants (adjusted OR 0.78; 95% CI 0.51, 1.22; p = 0.28). CONCLUSIONS: In Uro-EDIC women sexual activity rather than measures of diabetes control and complications was the main risk factor for urinary tract infection. The prevalence of cystitis was similar to that in nondiabetic women participants in NHANES III.

Cranberry products inhibit adherence of p-fimbriated Escherichia coli to primary cultured bladder and vaginal epithelial cells.

PURPOSE: Cranberry proanthocyanidins have been identified as possible inhibitors of Escherichia coli adherence to uroepithelial cells. However, little is known about the dose range of this effect. Furthermore, it has not been studied directly in the urogenital system. To address these issues we tested the effect of a cranberry powder and proanthocyanidin extract on adherence of a P-fimbriated uropathogenic E. coli isolate to 2 new urogenital model systems, namely primary cultured bladder epithelial cells and vaginal epithelial cells. MATERIALS AND METHODS: E. coli IA2 was pre-incubated with a commercially available cranberry powder (9 mg proanthocyanidin per gm) or with increasing concentrations of proanthocyanidin extract. Adherence of E. coli IA2 to primary cultured bladder epithelial cells or vaginal epithelial cells was measured before and after exposure to these products. RESULTS: Cranberry powder decreased mean adherence of E. coli IA2 to vaginal epithelial cells from 18.6 to 1.8 bacteria per cell (p <0.001). Mean adherence of E. coli to primary cultured bladder epithelial cells was decreased by exposure to 50 mug/ml proanthocyanidin extract from 6.9 to 1.6 bacteria per cell (p <0.001). Inhibition of adherence of E. coli by proanthocyanidin extract occurred in linear, dose dependent fashion over a proanthocyanidin concentration range of 75 to 5 mug/ml. CONCLUSIONS: Cranberry products can inhibit E. coli adherence to biologically relevant model systems of primary cultured bladder and vaginal epithelial cells. This effect occurs in a dose dependent relationship. These findings provide further mechanistic evidence and biological plausibility for the role of cranberry products for preventing urinary tract infection.

Ultraviolet radiation alters maize phyllosphere bacterial diversity.

Epiphytic bacteria are subjected to very stressful environments, including UV radiation. Bacterial assemblages on Zea mays (maize) leaves exposure were examined with and without UV-B radiation. Culture-independent molecular techniques were utilized for bacterial identification, diversity analysis and selection of putative UV exposure marker sequences. Few sequences corresponded to previously characterized phyllosphere bacteria. There was a strong tendency toward increased 16S rDNA sequence diversity in UV samples. Overall community structure was assessed using denaturing gel gradient electrophoresis; significant alterations in community structure were found in comparisons of phyllosphere bacterial samples from control and solar UV-B exposed plants.

Intraspecific comparisons reveal differences in the pattern of ultraviolet radiation responses in four maize (Zea mays L.) varieties.

Four maize (Zea mays L.) varieties were examined for ultraviolet radiation-induced changes in leaf rolling, biomass, fluctuating leaf asymmetry and DNA damage. Short-term dose-response curves for each response were constructed and responses in each line compared. The four varieties each exhibited a different pattern of tolerance and reactivity, ranging from B73, which was tolerant in all four measures, to TS1, which was affected in DNA damage levels and leaf rolling but unaffected in biomass accumulation and fluctuating leaf asymmetry. The pattern of ultraviolet radiation responses allows us to narrow the possibilities for the source of the defect in reactive varieties. The four varieties tested include inbred parents that have been used to construct recombinant inbred lines and a variety that is found in the background of the engineered RescueMu transposon mutagenesis lines. These dose-response curves and variety comparisons provide the foundation for genetic dissection of the mechanisms of ultraviolet radiation responses in maize.

A prospective study of asymptomatic bacteriuria in sexually active young women.

BACKGROUND: Asymptomatic bacteriuria is common in young women, but little is known about its pathogenesis, natural history, risk factors, and temporal association with symptomatic urinary tract infection. METHODS: We prospectively evaluated 796 sexually active, nonpregnant women from 18 through 40 years of age over a period of six months for the occurrence of asymptomatic bacteriuria (defined as at least 10(5) colony-forming units of urinary tract pathogens per milliliter). The women were patients at either a university student health center or a health maintenance organization. Periodic urine cultures were taken, daily diaries were kept, and regularly scheduled interviews were performed. Escherichia coli strains were tested for hemolysin, the papG genotype, and the ribosomal RNA type. RESULTS: The prevalence of asymptomatic bacteriuria (the proportion of urine cultures with bacteriuria in asymptomatic women) was 5 percent (95 percent confidence interval, 4 percent to 6 percent) among women in the university group and 6 percent (95 percent confidence interval, 5 percent to 8 percent) among women in the health-maintenance-organization group. Persistent asymptomatic bacteriuria with the same E. coli strain was rare. Symptomatic urinary tract infection developed within one week after 8 percent of occasions on which a culture showed asymptomatic bacteriuria, as compared with 1 percent of occasions when asymptomatic bacteriuria was not found (P<0.001). Asymptomatic bacteriuria was associated with the same risk factors as for symptomatic urinary tract infection, particularly the use of a diaphragm plus spermicide and sexual intercourse. CONCLUSIONS: Asymptomatic bacteriuria in young women is common but rarely persists. It is a strong predictor of subsequent symptomatic urinary tract infection.

Epithelial cell layer thickness and immune cell populations in the normal human vagina at different stages of the menstrual cycle.

OBJECTIVE: The aim of our study was to examine vaginal tissue during 3 phases of the menstrual cycle for the number of cell layers and epithelial immune cells. STUDY DESIGN: Vaginal biopsies were performed during 3 phases of the normal menstrual cycle (menstrual, days 1-5; preovulatory, days 7-12; and postovulatory, days 19-24) in 74 subjects. A subset of women had vaginal tissues stained with specific monoclonal antibody markers for Langerhans cells (CD1a), macrophages (KP1), T and B lymphocytes (CD4, CD8, CD21) and neutrophils (CD15). The number of cell layers and the number of immune cells in the vaginal tissue biopsy specimen were determined by a single observer who was blinded to clinical data. RESULTS: At 3 phases of the normal menstrual cycle, the mean number of epithelial cell layers underwent a small but statistically significant decrease from 27.8 +/- 0.7 on days 1-5 and 28.1 +/- 0.6 on days 7-12 to 26.0 +/- 0.7 on days 19-24 of the cycle (P =.01). Nonovulating women had a reduced mean epithelial cell layer count on days 7-12 (23.7 +/- 1. 4) compared with the epithelial cell layer count in ovulating women (28.8 +/- 0.7; P =.005). No significant changes were observed in the mean number per high-power field of Langerhans cells, macrophages, CD4 or CD8 lymphocytes, and neutrophil cell populations during the 3 phases of the cycle. B lymphocytes were not observed in the vaginal tissues. CONCLUSION: A small but statistically significant reduction in the number of vaginal epithelial cells was observed over the menstrual cycle. This reduction is not likely to be clinically significant. Immune cell populations in the vaginal tissues appeared stable throughout the menstrual cycle.

Risk factors for recurrent urinary tract infection in young women.

To define host factors associated with an increased risk of recurrent urinary tract infection (RUTI), a case-control study was conducted in 2 populations: university women and health maintenance organization enrollees. Case patients were 229 women 18-30 years old with RUTIs; control subjects were 253 randomly selected women with no RUTI history. In a multivariate model, independent risk factors for RUTI included recent 1-month intercourse frequency (odds ratio [OR], 5.8; 95% confidence interval [CI], 3.1-10.6 for 4-8 episodes), 12-month spermicide use (OR, 1.8; 95% CI, 1.1-2.9), and new sex partner during the past year (OR, 1.9; 95% CI, 1.2-3.2). Two newly identified risk factors were age at first urinary tract infection (UTI)

Influence of the normal menstrual cycle on vaginal tissue, discharge, and microflora.

The objective of this study was to examine genital tissue, vaginal fluid, and vaginal microbial flora at 3 phases of the menstrual cycle in asymptomatic women. Vaginal examinations were performed 3 times in 74 women: at the menstrual phase (days 1-5), the preovulatory phase (days 7-12), and the postovulatory phase (days 19-24). Flora of 50 women without bacterial vaginosis (BV) was analyzed separately from flora of 24 women with BV. The volume of vaginal discharge increased and the amount of cervical mucus decreased over the menstrual cycle. Among subjects without BV, the rate of recovery of any Lactobacillus changed little (range, 82% to 98%; P = .2); however, a small increase occurred in the rate of recovery of heavy (3+ to 4+ semiquantitative) growth of Lactobacillus over the menstrual cycle (P = .04). A linear decrease occurred in the rate of recovery of heavy growth of any non-Lactobacillus species, from 72% at days 1-5 to 40% at days 19-24 (P = .002). A linear decrease also occurred in the rate of recovery of Prevotella species, from 56% on days 1-5 to 28% on days 19-24 (P =. 007), while a small linear increase occurred in the rate of recovery of Bacteroides fragilis (P=.05). Among subjects with BV, the only significant change was an increase in the rate of recovery of Lactobacillus, from 33% at days 1-5 to 54% at days 19-24 (P = .008). Among all subjects, the rate of recovery of heavy growth of Lactobacillus increased over the menstrual cycle and, in contrast, the concentration of non-Lactobacillus species tended to be higher at menses, which is evidence that the vaginal flora becomes less stable at this time.

Effect of Staphylococcus aureus bacteria and bacterial toxins on meningeal permeability in vitro.

BACKGROUND AND OBJECTIVES: Epidural catheterization is associated with a significant bacterial colonization rate and occasionally frank infection. During epidural space infection, decreased analgesia despite increased epidural opioid doses has been described. One possible explanation for this observation is that bacterial infection decreases meningeal permeability. The purpose of the study was to determine whether Staphylococcus aureus bacteria, the most common organism causing epidural space infection, or S. aureus toxins alter meningeal permeability. METHODS: Spinal meninges of M. nemestrina monkeys were mounted in a previously established in vitro diffusion cell model and exposed to S. aureus toxins A, B, and F. Simultaneous transmeningeal fluxes of mannitol and sufentanil were measured before and after toxin exposure and compared to controls. In a second series of experiments, diffusion cells were inoculated with live S. aureus bacteria in suspension and the permeability of sufentanil was investigated. RESULTS: Staphylococcus aureus toxin-A increased the transmeningeal flux of mannitol but not sufentanil. Toxins B and F did not alter the meningeal permeability of either drug. Inoculation with live S. aureus bacteria increased the transmeningeal flux of sufentanil by 115+/-21% (P = .032). CONCLUSIONS: These data demonstrate that S. aureus alpha-toxin and live S. aureus bacteria can increase meningeal permeability. Thus, clinical observations of decreased epidural analgesia in the face of bacterial infection cannot be explained by decreased meningeal permeability.

The P histo-blood group-related glycosphingolipid sialosyl galactosyl globoside as a preferred binding receptor for uropathogenic Escherichia coli: isolation and structural characterization from human kidney.

The P histo-blood group-related glycosphingolipid, sialosyl galactosyl globoside (SGG), has recently been implicated as a preferred binding receptor for uropathogenic Escherichia coli [Stapleton, A. E., Stroud, M. R., Hakomori, S., and Stamm, W. E. (1998) Infect. Immun. 66, 3856-3861]. We report here the purification and complete structural characterization of SGG from normal human kidney. Using metabolically [35S]-labeled E. coli as a probe, a monosialylated glycosphingolipid was isolated to homogeneity. The glycosphingolipid was purified by a combination of high-performance liquid chromatography and preparative high-performance thin-layer chromatography and its structure unambiguously elucidated by 1H NMR, electrospray ionization mass spectrometry, and methylation analysis. Its primary structure was shown to be identical to a previously characterized, developmentally regulated, globo-series glycolipid thought to be unique to human teratocarcinoma. The significance of this structure as a unique receptor in human kidney for uropathogenic E. coli and its role in the pathogenesis of urinary tract infections are discussed.

Inverse association of H2O2-producing lactobacilli and vaginal Escherichia coli colonization in women with recurrent urinary tract infections.

Women with recurrent urinary tract infection (UTI) often demonstrate persistent vaginal colonization with Escherichia coli. Since strains of lactobacilli that produce hydrogen peroxide inhibit the growth of E. coli, the absence of these strains may predispose to E. coli colonization and to UTI. To test this hypothesis, vaginal introital cultures were obtained from 140 women, 65 with recurrent UTI (case-patients) and 75 without (controls). Vaginal E. coli colonization was significantly more frequent in case-patients than controls (35% vs. 11%; P < .001) and in women without H2O2-positive lactobacilli than in women with (odds ratio [OR], 4.0; P = .01). Spermicide use was associated with greater risk of vaginal E. coli colonization (OR, 12.5; P < .001) and with absence of H2O2-positive lactobacilli (OR, 2.9; P = .04). The inverse association between H2O2-positive lactobacilli and vaginal E. coli colonization remained in case-patients after controlling for spermicide use (OR, 6.5; P = .02). Thus, absence of H2O2-positive lactobacilli may be important in the pathogenesis of recurrent UTI by facilitating E. coli introital colonization.

The globoseries glycosphingolipid sialosyl galactosyl globoside is found in urinary tract tissues and is a preferred binding receptor In vitro for uropathogenic Escherichia coli expressing pap-encoded adhesins.

Women with a history of recurrent Escherichia coli urinary tract infections (UTIs) are significantly more likely to be nonsecretors of blood group antigens than are women without such a history, and vaginal epithelial cells (VEC) from women who are nonsecretors show enhanced adherence of uropathogenic E. coli isolates compared with cells from secretors. We previously extracted glycosphingolipids (GSLs) from native VEC and determined that nonsecretors (but not secretors) selectively express two extended globoseries GSLs, sialosyl galactosyl globoside (SGG) and disialosyl galactosyl globoside (DSGG), which specifically bound uropathogenic E. coli R45 expressing a P adhesin. In this study, we demonstrated, by purifying the compounds from this source, that SGG and DSGG are expressed in human kidney tissue. We also demonstrated that SGG and DSGG isolated from human kidneys bind uropathogenic E. coli isolates expressing each of the three classes of pap-encoded adhesins, including cloned isolates expressing PapG from J96, PrsG from J96, and PapG from IA2, and the wild-type isolates IA2 and R45. We metabolically 35S labeled these five E. coli isolates and measured their relative binding affinities to serial dilutions of SGG and DSGG as well as to globotriaosylceramide (Gb3) and globotetraosylceramide (Gb4), two other globoseries GSLs present in urogenital tissues. Each of the five E. coli isolates bound to SGG with the highest apparent avidity compared with their binding to DSGG, Gb3, and Gb4, and each isolate had a unique pattern of GSL binding affinity. These studies further suggest that SGG likely plays an important role in the pathogenesis of UTI and that its presence may account for the increased binding of E. coli to uroepithelial cells from nonsecretors and for the increased susceptibility of nonsecretors to recurrent UTI.

Cytotoxicity of hemolytic, cytotoxic necrotizing factor 1-positive and -negative Escherichia coli to human T24 bladder cells.

Approximately one-half of Escherichia coli isolates from patients with cystitis or pyelonephritis produce the pore-forming cytotoxin hemolysin, a molecule with the capacity to lyse erythrocytes and a range of nucleated cell types. A second toxin, cytotoxic necrotizing factor 1 (CNF1), is found in approximately 70% of hemolytic, but rarely in nonhemolytic, isolates. To evaluate the potential interplay of these two toxins, we used epidemiological and molecular biologic techniques to compare the cytotoxicity of hemolytic, CNF1(+), and CNF1(-) cystitis strains toward human T24 bladder epithelial cells in vitro. A total of 29 isolates from two collections of cystitis-associated E. coli were evaluated by using methylene blue staining of bladder monolayers at 1-h intervals after inoculation with each strain. Most (20 of 29) isolates damaged or destroyed the T24 monolayer (less than 50% remaining) within 4 h after inoculation. As a group, CNF1(+) isolates from one collection (11 strains) were less cytotoxic at 4 h than the CNF1(-) strains in that collection (P = 0.009), but this pattern was not observed among isolates from the second collection (18 strains). To directly evaluate the role of CNF1 in cytotoxicity of hemolytic E. coli without the variables present in multiple clinical isolates, we constructed mutants defective in production of CNF1. Compared to the CNF1(+) parental isolates, no change in cytotoxicity was detected in these cnf1 mutants. Our results indicate that CNF1 does not have a detectable effect on the ability of hemolytic E. coli to damage human bladder cell monolayers in vitro.

Urovirulence determinants in Escherichia coli strains causing prostatitis.

To define the urovirulence properties of Escherichia coli strains producing prostatitis, E. coli strains isolated from men with acute (7 strains) or chronic (23) prostatitis were compared with E. coli isolates from women with pyelonephritis (30), acute cystitis (60), or complicated urinary tract infection (UTI; 30). Strains from prostatitis patients were significantly more likely to express hemolysin than were strains causing complicated UTI (73% vs. 43%; P = .02) and more often demonstrated hybridization with the cytotoxic necrotizing factor-1 (CNF-1) probe (63%) than did strains from women (44%-48%). P fimbrial expression was highest among pyelonephritis (73%) and prostatitis strains (53%) and lowest among E. coli from women with complicated UTI (23%) and cystitis (30%; P < .05, prostatitis strains vs. either of the latter 2 groups). Results suggest that E. coli strains producing prostatitis generally possess urovirulence profiles similar to those of strains from women with acute uncomplicated pyelonephritis and that hemolysin and CNF-1 are especially prevalent in prostatitis strains.

Characteristics and prevalence within serogroup O4 of a J96-like clonal group of uropathogenic Escherichia coli O4:H5 containing the class I and class III alleles of papG.

The recent discovery of a geographically dispersed clonal group of Escherichia coli O4:H5 that includes prototypic uropathogenic strain J96 prompted us to determine the prevalence of J96-like strains within serogroup O4 and to further assess the characteristics of such strains. We used O:K:H;F serotyping, PCR-based genomic fingerprinting, pulsed-field gel electrophoresis (PFGE), multilocus enzyme electrophoresis (MLEE), and PCR detection of the three papG alleles and of the cytotoxic necrotizing factor 1 (cnf1) and aerobactin (aer) gene sequences to characterize the 15 O4 strains among 336 E. coli isolates from three clinical collections (187 from mixed-source bacteremia, 75 from urosepsis, and 74 from acute cystitis). J96-like strains constituted approximately half of the O4 strains, or 2% of the total population. In contrast to other O4 strains, the J96-like strains characteristically exhibited specific group III capsular antigens, the H5 flagellar and F13 fimbrial antigens, a distinctive PCR genomic fingerprint, the class III papG allele (plus, in 50% of strains, the enigmatic class I papG allele), and cnf1 but lacked aer. A subset of these strains was remarkably homogeneous with respect to all these characteristics and exhibited a distinctive PFGE fingerprint and MLEE pattern. These findings clarify the epidemiological relevance of J96 as a model extraintestinal pathogen, provide further evidence of the class I papG allele outside of strain J96, and offer insights into the evolution of E. coli serogroup O4.

An Arabidopsis photolyase mutant is hypersensitive to ultraviolet-B radiation.

Photolyases are DNA repair enzymes that use energy from blue light to repair pyrimidine dimers. We report the isolation of an Arabidopsis thaliana mutant (uvr2-1) that is defective in photorepair of cyclobutylpyrimidine dimers (CPDs). Whereas uvr2-1 is indistinguishable from wild type in the absence of UV light, low UV-B levels inhibit growth and cause leaf necrosis. uvr2-1 is more sensitive to UV-B than wild type when placed under white light after UV-B treatment. In contrast, recovery in darkness or in light lacking photoreactivating blue light results in equal injury in uvr2-1 and wild type. The uvr2-1 mutant is unable to remove CPDs in vivo, and plant extracts lack detectable photolyase activity. This recessive mutation segregates as a single gene located near the top of chromosome 1, and is a structural gene mutation in the type II CPD photolyase PHR1. This mutant provides evidence that CPD photolyase is required for plant survival in the presence of UV-B light.

Solar Ultraviolet-B Radiation Affects Seedling Emergence, DNA Integrity, Plant Morphology, Growth Rate, and Attractiveness to Herbivore Insects in Datura ferox.

To study functional relationships between the effects of solar ultraviolet-B radiation (UV-B) on different aspects of the physiology of a wild plant, we carried out exclusion experiments in the field with the summer annual Datura ferox L. Solar UV-B incident over Buenos Aires reduced daytime seedling emergence, inhibited stem elongation and leaf expansion, and tended to reduce biomass accumulation during early growth. However, UV-B had no effect on calculated net assimilation rate. Using a monoclonal antibody specific to the cyclobutane-pyrimidine dimer (CPD), we found that plants receiving full sunlight had more CPDs per unit of DNA than plants shielded from solar UV-B, but the positive correlation between UV-B and CPD burden tended to level off at high (near solar) UV-B levels. At our field site, Datura plants were consumed by leaf beetles (Coleoptera), and the proportion of plants attacked by insects declined with the amount of UV-B received during growth. Field experiments showed that plant exposure to solar UV-B reduced the likelihood of leaf beetle attack by one-half. Our results highlight the complexities associated with scaling plant responses to solar UV-B, because they show: (a) a lack of correspondence between UV-B effects on net assimilation rate and whole-plant growth rate, (b) nonlinear UV-B dose-response curves, and (c) UV-B effects of plant attractiveness to natural herbivores.