Plasmid-mediated fluoroquinolone resistance determinants in Escherichia coli from community uncomplicated urinary tract infection in an area of high prevalence of quinolone resistance

In Italy fluoroquinolones (FQs) are extensively prescribed in empirical therapy of uncomplicated urinary tract infection (UTI) despite recommendations in national guidelines and widespread antibiotic resistance in community. To survey the dissemination of plasmid-mediated quinolone resistance in a peak area of FQs consumption, E. coli strains from 154 community and 41 local hospital patients were collected; low level ciprofloxacin resistance qnrA, qnrB, qnrS, and aac(6)'-Ib-cr genes were screened by PCR and patterns of transferable resistances were determined. Clinical ciprofloxacin resistance in hospital doubled community value, while overall rates of FQ resistance genes were similar (31.6% and 27.8%). Prevalence of aac(6')-Ib-cr gene was 11% in outpatients (21%, inpatients) and risk of harbouring this variant was significantly associated with gentamicin resistance; linkage to ceftazidime resistance was significant (P=0.001) and six out of eight strains produced CTX-M-15 and TEM-1 beta lactamases. In transconjugants, the unique pattern ampicillin/kanamycin-gentamicin/ ESBL + was associated with aac(6')-Ib-cr gene presence and with an increase of ciprofloxacin MIC value. Data highlight the need to monitor the resistance risk factors in the local community to provide clinicians with well-grounded guidelines for UTI therapy.     

Adhesion to a human cell line by Escherichia coli strains isolated during urinary tract infections.

It has been shown that some, but not all, Escherichia coli strains isolated from urine adhere, in vitro, to the surface of uroepithelial or vaginal cells. In the present study, 212 strains, isolated from urine of 212 infected patients, were tested for adhesion by using an in vitro human cell line assay. A variable degree of attachment to the cell monolayer was detected in these strains. From patients with cystitis, only 19 (9.7%) of the 195 strains examined were adherent, whereas 5 (29.4%) of the 17 pyelonephritis strains had similar properties (P less than 0.05). To investigate the incidence of adhesion in the clinical manifestations of urinary tract infection, a sample of patients was picked at random from those with cystitis. During cystitis caused by adhesive bacteria, patients suffer more often from macroscopic hematuria than from dysuria, frequency, or recurrency (P less than 0.05). This study shows that E. coli strains isolated from urine samples possess a strikingly difference in capacity to adhere to a human cell line surface as demonstrated previously with uroepithelial or vaginal cells. Moreover, according to these data, the adhesion of E. coli may be considered as a virulent factor and would play a part in the infection of the urinary tract in humans.     

Evolution of fluoroquinolone resistance among Escherichia coli urinary tract isolates from a French university hospital: application of the dynamic regression model

Escherichia coli urinary tract isolates were collected in 1997-2003 from Nimes University Hospital in order to investigate long-term trends in antibiotic resistance and to explore the relationship between antibiotic use and the emergence of resistance. Time-series analysis (ARIMA models) and dynamic regression models were used to investigate relationships between antibiotic use and resistance to ofloxacin and ciprofloxacin. Significant increases were seen in the frequency of ofloxacin (8.9 to 16.7%) and ciprofloxacin resistance (6.2 to 10.1%) (p < 0.001). Using multivariate dynamic regression analysis, it was found that an increased use of one defined daily dose (DDD)/1000 patient-days for ofloxacin, ciprofloxacin and norfloxacin induced average increases of 0.81%, 0.65% and 0.53% in E. coli ofloxacin resistance (p < 0.01), with average delays of 4, 4 and 6 months, respectively. An increase of 1 DDD/1000 patient-days of ciprofloxacin, ofloxacin and norfloxacin use induced increases of 0.73%, 0.82% and 0.63% in E. coli ciprofloxacin resistance (p < 0.01), with average delays of 4, 4 and 5 months, respectively. The use of nalidixic acid was not associated significantly with an increase in resistance to fluoroquinolones by multivariate analysis.     

Studies on beta-lactamases from Escherichia coli isolated from urinary tract infections

The beta-lactamase types present in 75 ampicillin and carbenicillin resistant E. coli were characterized using isoelectric focusing (IEF). The strains were isolated from patients with urinary tract infections from two geographically different areas of Denmark: 38 strains from Copenhagen and 37 strains from North Jutland. For 19 of the strains from Copenhagen and 18 of the strains from North Jutland, their beta-lactamase activity against nitrocefin and ampicillin, carbenicillin, benzylpenicillin, cloxacillin and cephaloridine was examined by a micro-iodometric and an UV-spectrophotometric assay. The strains from Copenhagen showed greater activity (p less than 0.001) against nitrocefin than the strains from North Jutland. The rate of hydrolysis of ampicillin was greater for the strains from Copenhagen than for the strains from North Jutland. Ninety-three per cent of the strains produced plasmid-mediated beta-lactamases, of which the most prevalent, TEM-1, was produced by 97 per cent of these strains, and OXA-1 by 3 per cent.     

Virulence factors in urinary Escherichia coli strains: phylogenetic background and quinolone and fluoroquinolone resistance

Quinolone- and fluoroquinolone-resistant Escherichia coli strains harbor fewer virulence factors than susceptible strains. The reasons underlying this correlation are incompletely understood. We investigated the phylogenetic background, the presence of the papC, hlyA, and cnf1 (pathogenicity island II_J96-associated), fimA, iss, and iutA genes, and the presence of type 1 fimbriae, P fimbriae, and hemolysin in 243 urinary E. coli isolates resistant only to quinolones (8%), resistant to both quinolones and fluoroquinolones (51%), or susceptible to both drugs (41%). Group B2 accounted for 56% of the isolates, showing a significantly higher prevalence among fluoroquinolone-susceptible strains than among resistant strains (65% versus 50% [P = 0.03]). hly and cnf1 were significantly more associated with susceptibility (P < 0.001) and with group B2 (P < 0.001 for group B2 versus groups A and D). However, within group B2, fluoroquinolone-resistant strains showed lower prevalences of papC, hlyA, and cnf1 than their susceptible counterparts (P < 0.001). In contrast, the incidence of iutA appeared higher for refractory isolates, including group B2, than for susceptible isolates (P < 0.001). Only in group B2 did fluoroquinolone-resistant strains reveal a lesser ability to agglutinate Saccharomyces cerevisiae (7%) than quinolone-resistant (87%) and susceptible (80%) isolates, despite uniform possession of fimA genes. No similar contrast emerged for expression of hemolysin and P fimbriae. Mutations conferring quinolone and fluoroquinolone resistance may thus require a particular genetic background, not strictly correlated with phylogenetic groups. More interestingly, the mutational event itself can affect the expression of type 1 fimbriae, at least in the prevalent and complex B2 strains.     

Association between Escherichia coli with NotI-restriction resistance and urinary tract infections

BackgroundEscherichia coli is the most common cause of urinary tract infections (UTIs). It is widely accepted that uropathogenic E. coli (UPEC) mainly emerge from the distal gut microbiota. Identification of bacterial characteristics that are able to differentiate UPEC from fecal commensal strains will facilitate the development of novel strategies to detect and monitor the spread of UPEC.MethodsFifty fecal commensal, 83 UTI-associated and 40 biliary tract infection (BTI)-associated E. coli isolates were analyzed. The NotI restriction patterns of chromosomal DNA in the isolates were determined by pulse-field gel electrophoresis. The phylogenetic types and the presence of 9 known virulence genes of each isolate were determined by PCR analyses. Additionally, the susceptibilities of the isolates to antibiotics were revealed. Then the associations of NotI resistance with UTI-associated isolates, phylotypes, and antibiotic resistance were assessed.ResultsNotI resistance was correlated with UTI-associated isolates, compared to the fecal isolates. Consistently, NotI-resistant isolates harbored a greater number of virulence factors and mainly belonged to phylotype B2. Additionally NotI resistance was correlated with chloramphenicol resistance among the bacteria. Among the fecal, UTI-associated and BTI-associated groups, the distribution of NotI-resistant group B2 isolates was correlated with UTI-associated bacteria.ConclusionNotI resistance alone is a potential marker for distinguishing fecal strains and UPEC, while the combination of NotI resistance and B2 phylogeny is a candidate marker to differentiate UPEC from fecal and other extraintestinal pathogenic E. coli. Additionally, NotI resistance may be valuable for assessing the potential of chloramphenicol resistance of E. coli.     

Virulence factors in strains of Escherichia coli isolated in urinary tract infections]

In 21 strains of E. coli isolated from patients with cystitis and asymptomatic bacteriuria the authors assessed the group, mannososensitive and mannoresistant agglutination of human, bovine, chick, guinea pig, sheep and pig erythrocytes, the production of haemolysis, colicins, aerobactin, the capacity of strains to induce oedema on mouse paws, the lethal effect in mice, the resistance to tetracycline, streptomycin, chloramphenicol, kanamycin, ampicillin and sulfamethoxidine, the transmission of resistance and sensitivity of strains to the action of fresh human serum. In ten strains the authors recorded also the production of haemolysin and the lethality for mice, in four strains the production of colicin V. production of aerobactin and serum resistance. In three strains the aerobactin production was recorded concurrently with the haemolysin production. In none of the samples P. fimbriae were found.     

Virulence determinants, phylogenetic groups and fluoroquinolone resistance in Escherichia coli isolated from cystitis and pyelonephritis

Aim of the study

The aim of this study is to assess the relation between virulence genotype, phylogenetic group and susceptibility to fluoroquinolones and the urinary tract infection type including pyelonephritis and cystitis due to Escherichia coli.

Materials and methods

Between 2006 and 2007, 129 non-duplicate E. coli isolates from pyelonephritis (n = 56) and cystitis (n = 73) were prospectively collected. The antibiotic susceptibility was done by disk diffusion method. The phylogenetic groups, A, B1, B2 and D and 18 virulence genes were determined by multiplex PCR. Statistical analysis was done with the Pearson χ2 test, Mann-Whitney U-test, Kruskal-Wallis test and stepwise multivariable logistic regression analysis, P values below 0.05 were considered statistically significant.

Results

For the pyelonephritis group, sex ratio was 0.3, the median age for women was 30 years and for men it was 54 years. For the cystitis group, sex ratio was 0.4, the median age for women was 41.5 years and for men it was 67.8 years. Significant statistical correlations were found between pyelonephritis isolates and susceptibility to ciprofloxacin (P = 4 10^–5), papG allele II (P = 2 10^–6), hlyA (P = 10^–03), iroN (P = 0.04), iha (P = 0.03) and ompT (P = 0.03) virulence genes, high virulence score (P = 0.008) and B2 phylogenetic group (P = 0.03). In multivariate logistic regression analysis, papG II as predictor of pyelonephritis, no correlation could be established for the cystitis group.

Conclusion

Our findings argue for a direct link between pyelonephritis, virulence factors, susceptibility to fluroquinolones and B2 phylogenetic group among uropthogenic E. coli.     

Alpha-hemolytic strains of Escherichia coli isolated in urinary tract infections in small children]

The authors focused attention on testing of biological properties of alpha-haemolytic strains of E. coli in infections of the urinary pathways in children and on the antibody response against alpha-haemolysin. The strains belonged into serogroups 02, 04, 06, 018, 075, 0112. The majority of strains had fimrial adhesins, i.e. P fimbriae, or fimbriae type 1. Alpha-haemolysin one of the factors of UPEC virulence induces anti-alphahaemolysin formation in serum (ANTI AH). Titres of ANTI AH in sera of children with acute uroinfection were from 1:32 to 1:1024. Lower titres were recorded in infections of the lower urinary pathways higher ones in renal infections, whereby the dynamics of ANTI AH titres depend on the development of the disease.     

Uropathogenic Escherichia coli outer membrane antigens expressed during urinary tract infection

Uncomplicated urinary tract infection (UTI) caused by uropathogenic Escherichia coli (UPEC) represents a prevalent and potentially severe infectious disease. In this study, we describe the application of an immunoproteomics approach to vaccine development that has been used successfully to identify vaccine targets in other pathogenic bacteria. Outer membranes were isolated from pyelonephritis strain E. coli CFT073 cultured under conditions that mimic the urinary tract environment, including iron limitation, osmotic stress, human urine, and exposure to uroepithelial cells. To identify antigens that elicit a humoral response during experimental UTI, outer membrane proteins were separated by two-dimensional gel electrophoresis and probed using pooled antisera from 20 CBA/J mice chronically infected with E. coli CFT073. In total, 23 outer membrane antigens, including a novel iron compound receptor, reacted with the antisera and were identified by mass spectrometry. These antigens also included proteins with known roles in UPEC pathogenesis, such as ChuA, IroN, IreA, Iha, IutA, and FliC. These data demonstrate that an antibody response is directed against these virulence-associated factors during UTI. We also show that the genes encoding ChuA, IroN, hypothetical protein c2482, and IutA are significantly more prevalent (P < 0.01) among UPEC strains than among fecal-commensal E. coli isolates. Thus, we suggest that the conserved outer membrane antigens identified in this study could be rational candidates for a UTI vaccine designed to elicit protective immunity against UPEC infection.     

Fluoroquinolone resistance of Escherichia coli at a cancer center: epidemiologic evolution and effects of discontinuing prophylactic fluoroquinolone use in neutropenic patients with leukemia

The aim of the present study was to investigate the epidemiologic evolution of fluoroquinolone resistance of E. coli clinical isolates from patients admitted to a hematology-oncology service where fluoroquinolone prophylaxis during neutropenia was recommended as the standard of care for many years but was then discontinued in a trial conducted in patients with acute leukemia. Fluoroquinolones had been shown to decrease the incidence of gram-negative bacteremia in cancer patients with neutropenia, yet it was thought that the emergence of resistance in Escherichia coli and other gram-negative bacteria may have caused a progressive lack of efficacy of fluoroquinolone prophylaxis. Epidemiologic surveillance of fluoroquinolone resistance of E. coli clinical isolates at our cancer center since 1992 showed a continuing influx of new clones not previously observed in the population of cancer patients, an increase in the number of cancer patients per year colonized and/or infected by fluoroquinolone-resistant E. coli (1992–1994, 10–16 patients; 1995–1997, 24–27 patients), and a resistance rate of >50% among E. coli bloodstream isolates of hematology-oncology patients. A 6-month fluoroquinolone prophylaxis discontinuation intervention trial in 1998 suggested that despite increasing resistance among E. coli isolates, fluoroquinolone prophylaxis in acute leukemia patients was still effective in the prevention of gram-negative bacteremia (incidence rates, 8% during the pre-intervention period vs. 20% after discontinuation; p<0.01). The resumption of fluoroquinolone prophylaxis in acute leukemia patients thereafter decreased the incidence of gram-negative bacteremia to the pre-intervention level (9%; p=0.03), while the proportion of in vitro fluoroquinolone resistance in E. coli bacteremia isolates again increased (from 15% during the intervention period to >50% in the post-intervention period). Relative rates of resistance thus were a poor indicator of the potential clinical benefits associated with fluoroquinolone prophylaxis in cancer patients.     

Clinical and molecular epidemiology of quinolone-resistant Escherichia coli isolated from urinary tract infection

Since the use of fluoroquinolone antibiotic in clinical practice was introduced about a decade ago, quinolone-resistant E. coli (QREC) strains are being isolated with increasing frequency. From 1996 to 2000, 297 cases of urinary tract infection (UTI) due to QREC were observed in our hospital; 218 episodes (73.5%) were community acquired. The incidence of QREC UTIs increased steadily from 14.4% to 21.3% during 5 years when we compared the clinical characteristics of 60 QREC UTI with those of 80 quinolone-susceptible E. coli UTIs. Significant differences in susceptibility to various antibiotics were observed between the QREC and QSEC strains of E. coli. The multidrug resistance rate of QREC was much higher (38.3%) than those of quinolone susceptible isolates (18.8%). Prior fluoroquinolone use (p = 0.05), old age (p = 0.001), and a vegetative state (p = 0.03) were the independent risk factors for the acquisition of QREC UTI. The outcome of E. coli UTI is dependent on quinolone resistance. Thirty-day mortality was higher in QREC UTI patients, probably due to aggravation of underlying illness, but not quinolone resistance. On the basis of PFGE analysis, although some clustering was found in the hospital, genomic diversity was found among both the community and nosocomial strains. The increased frequency of QREC UTIs is thus not due to transmission of resistant strains but probably results from the selection of resistant strains from the endogenous flora of patients.     

Biological cost of fluoroquinolone resistance in Escherichia coli implicated in polyclonal infection

Polyclonal Escherichia coli strains were isolated in a transplanted patient who experienced successive septic shocks. Fluoroquinolone susceptible and resistant strains were corresponding to different PFGE fragment profiles. The gyrA S83L mutation was associated with a reduction in biological fitness. Resistant strain was selected by a long-term single use of ofloxacin.     

Multidrug-resistant pathogenic Escherichia coli isolated from wild birds in a veterinary hospital

Wild birds are carriers of Escherichia coli. However, little is known about their role as reservoirs for extra-intestinal pathogenic E. coli (ExPEC). In this work we investigated E. coli strains carrying virulence genes related to human and animal ExPEC isolated from free-living wild birds treated in a veterinary hospital. Multidrug resistance was found in 47.4% of the strains, but none of them were extended-spectrum beta-lactamase producers. Not only the virulence genes, but also the serogroups (e.g. O1 and O2) detected in the isolates of E. coli have already been implicated in human and bird diseases. The sequence types detected were also found in wild, companion and food animals, environmental and human clinical isolates in different countries. Furthermore, from the 19 isolates, 17 (89.5%) showed a degree of pathogenicity on an in vivo infection model. The isolates showed high heterogeneity by pulsed-field gel electrophoresis indicating that E. coli from these birds are clonally diverse. Overall, the results showed that wild birds can be reservoirs and/or vectors of highly pathogenic and multidrug-resistant E. coli that have the potential to cause disease in humans and poultry.     

Virulence factors in Escherichia coli urinary tract infection.

Uropathogenic strains of Escherichia coli are characterized by the expression of distinctive bacterial properties, products, or structures referred to as virulence factors because they help the organism overcome host defenses and colonize or invade the urinary tract. Virulence factors of recognized importance in the pathogenesis of urinary tract infection (UTI) include adhesins (P fimbriae, certain other mannose-resistant adhesins, and type 1 fimbriae), the aerobactin system, hemolysin, K capsule, and resistance to serum killing. This review summarizes the virtual explosion of information regarding the epidemiology, biochemistry, mechanisms of action, and genetic basis of these urovirulence factors that has occurred in the past decade and identifies areas in need of further study. Virulence factor expression is more common among certain genetically related groups of E. coli which constitute virulent clones within the larger E. coli population. In general, the more virulence factors a strain expresses, the more severe an infection it is able to cause. Certain virulence factors specifically favor the development of pyelonephritis, others favor cystitis, and others favor asymptomatic bacteriuria. The currently defined virulence factors clearly contribute to the virulence of wild-type strains but are usually insufficient in themselves to transform an avirulent organism into a pathogen, demonstrating that other as-yet-undefined virulence properties await discovery. Virulence factor testing is a useful epidemiological and research tool but as yet has no defined clinical role. Immunological and biochemical anti-virulence factor interventions are effective in animal models of UTI and hold promise for the prevention of UTI in humans.