Nubia L. Ramos

Estrogen Supports Urothelial Defense Mechanisms

Estrogen supports urothelial defense against infection by increasing the expression of antimicrobial peptides and by protecting epithelial integrity. Urothelial Defense Tug-of-War It is well known that urinary tract infections (UTIs) are common in women, but which women are at more risk of UTIs is still far from certain, with some studies suggesting that it’s younger women, and others showing evidence of increased risk after menopause. Now, a study by Lüthje and coauthors suggests that both of those views may be partially correct and demonstrates the mechanisms for each. To understand how estrogen contributes to UTI pathogenesis, the authors examined cells from the urothelium (bladder lining) of menstruating women and older, postmenopausal subjects, studying the postmenopausal women before and after a 2-week period of estrogen supplementation. Then, to determine the mechanistic basis for their observations, they studied urothelial cells in an estrogen-depleted mouse model and in vitro. Thus, they demonstrated that estrogen has a protective effect on the urothelium, stimulating production of antimicrobial peptides and strengthening the attachment between urothelial cells. At the same time, the authors found that estrogen also increases the amount of bacteria taken up inside the urothelial cells. On the basis of the findings of Lüthje et al., one can conclude that young women may experience a greater incidence of UTIs because the high-estrogen environment increases the risk of bacterial invasion of the urothelium. Conversely, postmenopausal women may have more difficulty fighting off the infections because of their decreased production of antimicrobial peptides and diminished integrity of the urothelial lining. Additional studies of human and animal subjects will be needed to learn more about the molecular mechanism of estrogen’s effects on the urothelium and understand what determines the balance between its pro- and anti-UTI effects. However, this line of research does advance us closer to understanding and eventually helping prevent the problem of recurrent UTIs in women. Epidemiological data imply a role of estrogen in the pathogenesis of urinary tract infections (UTIs), although the underlying mechanisms are not well understood. However, it is thought that estrogen supplementation after menopause decreases the risk of recurrent infections. We sought to investigate the influence of estrogen on host-pathogen interactions and the consequences for UTI pathogenesis. We analyzed urothelial cells from menstruating and postmenopausal women before and after a 2-week period of estrogen supplementation, and also studied the influence of estradiol during Escherichia coli UTI in a mouse infection model. Important findings were confirmed in two human urothelial cell lines. We identified two epithelial defense mechanisms modulated by estrogen. Estrogen induced the expression of antimicrobial peptides, thereby enhancing the antimicrobial capacity of the urothelium and restricting bacterial multiplication. In addition, estrogen promoted the expression and redistribution of cell-cell contact–associated proteins, thereby strengthening the epithelial integrity and preventing excessive loss of superficial cells during infection. These two effects together may prevent bacteria from reaching deeper layers of the urinary tract epithelium and developing reservoirs that can serve as a source for recurrent infections. Thus, this study presents some underlying mechanisms for the beneficial effect of estradiol after menopause and supports the application of estrogen in postmenopausal women suffering from recurrent UTI.

Genetic relatedness and virulence gene profiles of Escherichia coli strains isolated from septicaemic and uroseptic patients

We investigated the relationship between clonality and virulence factors (VFs) of a collection of Escherichia coli strains isolated from septicaemic and uroseptic patients with respect to their origin of translocation. Forty septicaemic and 30 uroseptic strains of E. coli were tested for their phylogenetic groupings, genetic relatedness using randomly amplified polymorphic DNA (RAPD), biochemical fingerprinting method (biochemical phenotypes [BPTs]), adherence to HT-29 cells and the presence of 56 E. coli VF genes. Strains belonging to phylogenetic groups B2 and D constituted 93% of all strains. Fifty-four (77%) strains belonged to two major BPT/RAPD clusters (A and B), with cluster A carrying significantly (P = 0.0099) more uroseptic strains. The degree of adhesion to HT-29 cells of uroseptic strains was significantly (P = 0.0012) greater than that of septicaemic strains. Of the 56 VF genes tested, pap genes was the only group that were found significantly (P < 0.0001) more often among uroseptic isolates. Phylogenetic group B2 contained a significantly higher number of strains carrying pap genes than those in group D. We conclude that uroseptic E. coli are clonally different from septicaemic strains, carry more pap genes and predominantly adhere more to the HT-29 cell model of the gut.

Prevalence and Persistence of Escherichia coli Strains with Uropathogenic Virulence Characteristics in Sewage Treatment Plants

ABSTRACT We investigated the prevalence and persistence of Escherichia coli strains in four sewage treatment plants (STPs) in a subtropical region of Queensland, Australia. In all, 264 E. coli strains were typed using a high-resolution biochemical fingerprinting method and grouped into either a single or a common biochemical phenotype (S-BPT and C-BPT, respectively). These strains were also tested for their phylogenetic groups and 12 virulence genes associated with intestinal and extraintestinal E. coli strains. Comparison of BPTs at various treatment stages indicated that certain BPTs were found in two or all treatment stages. These BPTs constituted the highest proportion of E. coli strains in each STP and belonged mainly to phylogenetic group B2 and, to a lesser extent, group D. No virulence genes associated with intestinal E. coli were found among the strains, but 157 (59.5%) strains belonging to 14 C-BPTs carried one or more virulence genes associated with uropathogenic strains. Of these, 120 (76.4%) strains belonged to seven persistent C-BPTs and were found in all four STPs. Our results indicate that certain clonal groups of E. coli with virulence characteristics of uropathogenic strains can survive the treatment processes of STPs. These strains were common to all STPs and constituted the highest proportion of the strains in different treatment tanks of each STP.

Population structure and uropathogenic virulence-associated genes of faecal Escherichia coli from healthy young and elderly adults.

We investigated the population structures of faecal Escherichia coli in 30 healthy young adults (13 males and 17 females) aged between 20 and 45 years and 29 elderly adults (14 females and 15 males) aged between 65 and 77 years. In all, 1566 strains were typed with the PhPlate system and grouped into biochemical phenotypes (BPTs). Strains with shared BPTs were further typed using randomly amplified polymorphic DNA analysis. Forty-four per cent of the strains were shared between two or more age and gender groups. Elders had a significantly higher (P<0.001) number of BPTs (mean±standard error 3.3±0.27) than younger groups (1.82±0.27). Phylogenetic affiliation and virulence-associated genes (VAGs) of the strains showed that more than 80 % of the strains belonging to dominant types belonged to phylogroups B2 and D. Amongst dominant BPTs, phylogenetic group A was significantly associated with females (P<0.0001), and elders were more likely to carry group D (P<0.0124). Elderly males had a higher prevalence of VAGs than young males (P<0.0001) and young females (P<0.0005). We conclude that there is a lower prevalence of E. coli with uropathogenic properties in healthy young adults than in elders.

Characterisation of uropathogenic Escherichia coli from children with urinary tract infection in different countries

Uropathogenic Escherichia coli (UPEC) carry many virulence factors, including those involved in long-term survival in the urinary tract. However, their prevalence and role among UPEC causing urinary tract infection (UTI) in children is not well studied. To further understand the virulence characteristics of these bacteria, we investigated the prevalence of antibiotic resistance, antigen 43 genes, curli and cellulose among UPEC in children from different countries. Isolates (n = 337) from five countries were tested for antibiotic susceptibility, phylogenetic groups, prevalence of flu, fluACFT073, fluBCFT073, curli and cellulose. High prevalence of multidrug resistance and extended spectrum beta lactamase production was found among Iranian and Vietnamese isolates. Resistance was associated with phylogenetic group D while group B2 was associated with fluACFT073 and fluBCFT073. Fewer Iranian isolates carried fluACFT073, curli and cellulose. fluBCFT073 was most prevalent among Slovak isolates. Ampicillin and amoxicillin/clavulanic acid resistance was prevalent among fluACFT073- and fluBCFT073-positive Australian, Iranian and Swedish isolates. Lack of curli and cellulose was associated with resistance among Vietnamese isolates. We conclude that major differences exist in the prevalence of antibiotic resistance among UPEC from different countries. Associations observed between resistance and virulence factors may, in different ways, promote the long-term survival of UPEC in the urinary tract.

Uropathogenic Escherichia coli Isolates from Pregnant Women in Different Countries

ABSTRACT Urinary tract infection (UTI) is common during pregnancy and can be associated with negative outcomes for both the mother and fetus. Increased risk of infection among these patients has been attributed to physiological changes, and less focus has been placed on Escherichia coli, the most frequent causative agent. We investigated the virulence properties of isolates causing UTI in pregnant women in Sweden, Uganda, and Vietnam, as well as nonpregnant women in Sweden. Although phylogenetic group B2 was the most prevalent group, more Ugandan isolates belonged to group B1, associated with commensal strains, than isolates from other countries. Adherence to and invasion of urothelial cells, key events in the infection process, were low among group B1 isolates from pregnant Swedish women compared to those from nonpregnant patients. Similar levels of adherence and invasion were seen in isolates from pregnant women in Uganda and Vietnam. More biofilm was formed by group B2 isolates than by those belonging to group B1 and by Ugandan group B2 isolates than by those from pregnant Swedish and Vietnamese women. The antigen 43a-encoding gene, fluA CFT073, was most prevalent among Ugandan isolates. Expression of the biofilm components, curli and cellulose, was low among all isolates. Multidrug resistance was more common among isolates from Uganda and Vietnam than among those from Swedish patients. We suggest that while bacterial virulence properties play an important role in UTI during pregnancy, physiological changes in the host may contribute more to the incidence of infection caused by less virulent E. coli.

Host species-specific translocation of Escherichia coli.

The purpose of this paper is to investigate the rate of translocation of Escherichia coli strains in different experimental/animal models. Four proficient translocating E. coli strains isolated from mesenteric lymph nodes (MLNs) and/or the blood of rats (strains KIC-1 and KIC-2), from a fatal case of pancreatitis (HMLN-1) and from pigs (PC-1 isolated in this study) were tested for their ability to translocate across two host species and the Caco-2 cell line as a model of the human gut epithelium. HMLN-1 was found in the MLNs of all 15 pigs tested. This strain, however, did not translocate in any rats and only colonised the caecum of four rats in small numbers. HMLN-1 and PC-1 were the dominant translocating strains in Caco-2 cells compared to KIC-1 and KIC-2, which were found to translocate at a lower rate in pigs and in Caco-2 cells. The rate of translocation of PC-1 in rats was also very low compared to KIC-1 and KIC-2. We suggest that, in studies aiming to investigate the mechanism of translocation of E. coli strains isolated from humans, rats may not be an appropriate animal model and that the Caco-2 cells or pigs are more suitable in vitro and in vivo models, respectively.

Decoctions from Citrus reticulata Blanco seeds protect the uroepithelium against Escherichia coli invasion.

ETHNOPHARMACOLOGICAL RELEVANCE Traditional usage suggests Citrus reticulata Blanco seeds have beneficial effects against infection. The purpose of this study was to investigate the effect of Citrus reticulata on the uroepithelium and to determine the mechanisms responsible for protection against urinary tract infection (UTI). MATERIALS AND METHODS Human bladder cell lines T24 and 5637 were employed in a cell culture infection model to determine the effects of Citrus reticulata treatment on Escherichia coli adherence and invasion of the uroepithelium. β1 integrin and caveolin-1 mRNA expression was assessed using RT real-time PCR. β1 integrin protein expression was confirmed by Western Blot. The effect of Citrus reticulata on bacteria was investigated using antibacterial sensitivity, yeast agglutination and biofilm assays. RESULTS Citrus reticulata treatment decreased β1 integrin expression and reduced bacterial invasion while adhesion of uroepithelial cells was not affected. Caveolin-1 expression was not influenced either and Citrus reticulata did neither exhibit any direct antimicrobial effect nor interfered with type 1 fimbriae binding. CONCLUSIONS Our results show that Citrus reticulata has a protective effect on the uroepithelium as seen by reduced bacterial invasion of uroepithelial cells. These properties suggest that seeds from Citrus reticulata may have therapeutic potential in preventing UTI.

The impact of vitamin D on the innate immune response to uropathogenic Escherichia coli during pregnancy

Urinary tract infections are highly common during pregnancy, and can cause serious complications for the mother and baby. Vitamin D, predominantly obtained from the sunlight, is known to have an effect on the urothelium, with immunomodulatory capacity against Escherichia coli infection. However, its influence at this site remains to be further explored. This study therefore investigated its impact during pregnancy in a population of women who have the possibility of adequate year-round sun exposure. Serum from pregnant Ugandan women (n = 32) in each trimester of pregnancy, from women after delivery (n = 29) and from never-pregnant controls (n = 25) was collected. 25-Hydroxyvitamin D (25-OHD), cathelicidin LL-37, human β-defensin 2, interleukin (IL)-8 and soluble CD14 serum concentrations were measured by chemiluminescence immunoassay or ELISA. The ability of serum to inhibit E. coli growth was tested. The immunomodulatory capacities of these serum samples and 1,25-dihydroxyvitamin D3 were investigated in urothelial cells. Increases in 25-OHD and LL-37 levels were observed as pregnancy progressed, peaking in the third trimester. Serum 25-OHD levels were higher in multigravidae than in primigravidae, and correlated positively with maternal age. IL-8 levels were lower in the third trimester than in the first trimester, increased after delivery, but remained below those of never-pregnant women. Similarly, soluble CD14 concentrations increased after delivery. As gestation advanced, serum had an increased capacity to inhibit E. coli growth. In vitro, it modulated the IL-8 response to infection in a vitamin D concentration-dependent manner. Our findings demonstrate that increasing vitamin D levels as pregnancy advances modulate the innate immune system towards a protective response to infection.