a Medical Microbiology Division, 251 MRC, Department of Pathology, University of Iowa College of Medicine, Iowa City, IA 52242, USA; b Laboratório Especial de Microbiologia Clínica, Division of Infectious Diseases, Universidade Federal de São Paulo, São Paulo, Brazil
![]() |
Abstract |
---|
![]() ![]() ![]() ![]() ![]() ![]() |
---|
![]() |
Introduction |
---|
![]() ![]() ![]() ![]() ![]() ![]() |
---|
In the community and hospital settings the aetiology of UTIs and the antimicrobial susceptibility of urinary pathogens have been changing over the years.7,8 Factors such as the changing patient population, extensive use and misuse of antimicrobial agents, could all contribute to changes in the microbial profile of urinary tract isolates.9
Most cases of UTI in the hospital setting are initially treated empirically based on the frequency of potential pathogens, local antimicrobial resistance rates and illness severity. The use of inappropriate empirical therapy was found to be a predictor of mortality in patients who had bacteraemia originating from a urinary tract source.5 Consequently, the establishment of worldwide antimicrobial resistance surveillance systems seems to be an important step in detecting the emergence of resistance patterns, helping in the selection of the most efficacious empirical therapy at the local level, and supporting the implementation of preventive measures. The SENTRY Antimicrobial Surveillance Program has been monitoring antimicrobial resistance worldwide among selected types of infection since January 1997.10 The selection of the participant medical centres was based on the principle that they should be sentinel laboratories in their respective regions. The main objective of the present study was to evaluate the potency and spectrum of antimicrobial agents tested against UTI pathogens isolated in hospitalized patients in Latin America in the second year of the SENTRY Antimicrobial Surveillance Program (1998). We also determined the mechanism of resistance to quinolones among the ciprofloxacin-resistant Escherichia coli.
![]() |
Materials and methods |
---|
![]() ![]() ![]() ![]() ![]() ![]() |
---|
A total of 434 bacterial UTI isolates collected from Latin American patients were evaluated in this study. Only one isolate per patient was included. Ten Latin American laboratories participated in the study, eight were hospital based and independent laboratories served the two remaining medical centres. The laboratories were distributed throughout six countries (nine cities) including São Paulo, Rio de Janeiro and Florianópolis in Brazil; Buenos Aires and San Isidro in Argentina; Santiago (two sites) in Chile; Medellin in Colombia; Caracas in Venezuela; and Mexico City in Mexico. Nine of the 10 sites were identical to those participating in the 1997 study. In 1998 the Uruguayan centre was replaced by a Venezuelan centre. Each participant laboratory contributed approximately 50 strains, consecutively collected in the period from January to March 1998. The participant medical centres were directed by protocol to collect isolates from consecutively hospitalized patients with symptomatic UTI confirmed by urine culture (colony count > 105 cfu/mL). A summary description of demographic data such as patient's age, gender, ward and hospitalization in intensive care unit was obtained. The mode of acquisition, community or nosocomial, was also noted. The isolates were identified to the species level by the participant centre and sent to the monitoring laboratory (University of Iowa, Iowa City, IA, USA) for identification confirmation and reference antimicrobial susceptibility testing.
Susceptibility testing
Antimicrobial susceptibility testing was performed and results interpreted using the reference broth microdilution method as described by the NCCLS.11 Antimicrobial agents were obtained from their respective USA manufacturers for testing. The activities of ciprofloxacin, ofloxacin, gatifloxacin and trovafloxacin against the 23 ciprofloxacin-resistant E. coli were also evaluated by Etest (AB Biodisk, Solna, Sweden). Quality control was performed using strains from the American Type Culture Collection (ATCC). Isolates of Klebsiella pneumoniae, Proteus mirabilis and E. coli with increased MICs (2 mg/L) of ceftazidime and/or ceftriaxone and/or aztreonam were considered as possible extended-spectrum ß-lactamase (ESBL)-producing phenotypes according to NCCLS criteria.12,13 The ESBL phenotype was confirmed by additional tests using Etest strips (AB Biodisk), containing the ß-lactam substrate (aztreonam, cefotaxime, cefpodoxime, ceftazidime) with or without clavulanate at a fixed concentration of 2 mg/L. The variations of the MIC of the ß-lactam alone and of ß-lactamclavulanate combination were compared. A reduction of the ß-lactam MIC of more than 2 log2 dilutions (>four-fold) in the presence of clavulanate indicated ESBL production.14
Molecular methods
The ESBL-producing isolates and selected multi-resistant isolates were genotyped by ribotyping. An isolate was considered multi-resistant if it was resistant to at least five of the following antimicrobial agents: imipenem (MIC 8 mg/L), ceftazidime (MIC
32 mg/L), cefepime (MIC
32 mg/L), ciprofloxacin (MIC
4 mg/L), penicillin ß-lactamase inhibitor combinations (MIC
128 mg/L), tetracycline (MIC
8 mg/L) and trimethoprimsulphamethoxazole (MIC > 2 mg/L). Ribotyping was performed using the RiboPrinter Microbial Characterization System (E. I. duPont de Nemours, Wilmington, DE, USA) according to the manufacturer's instructions. The patterns were electronically imaged, stored and compared. Pattern comparison was based on both position and signal intensity. Isolates with coefficients of similarity > 0.9 were considered to have the same ribotype profile.15
The strains with identical ribotype profiles were also typed using pulsed-field gel electrophoresis (PFGE). PFGE was performed using the restriction endonucleases SpeI and SmaI for Enterobacteriaceae and Acinetobacter spp. isolates, respectively, as previously described.16 Analysis of PFGE patterns was performed by visual inspection of photographs of ethidium bromide-stained gels. The isolates were classified as identical if they had the same bands, and as similar if they differed by one to three bands. Similar patterns were grouped under the same DNA type (subtypes) and isolates differing by more than three bands were considered to represent distinct DNA types.
Twenty-three isolates were selected from 58 ciprofloxacin-resistant E. coli (MICs > 8 mg/L) for evaluation of the mechanisms of resistance to quinolones. Each centre contributed at least one isolate, except for centres 39 and 49. The molecular characterization of quinolone resistance was determinated by amplification of the gyrA and parC genes by PCR followed by sequencing of the respective amplicons. Briefly, PCR amplification of the genes was performed as follows: stock quantities of master mixes were prepared robotically. For the parC gene, 50 pmol/L of oligonucleotide primers, sense 5'-TCTGAACTGGGCCTGAATG-3' (19-mer) and antisense 5'-CGTTCACCAGCAGGTTAG-3' (18-mer), amplified a 344 bp fragment. For the gyrA gene the primers, sense 5'-CCGTCGCGTACTTTACGC-3' (18-mer) and antisense 5'-CGTTCACCAGCAGGTTAG-3' (18-mer), amplified a 384 bp fragment.17 The PCR mixtures contained 200 mmol dATP, dTTP, dGTP and dCTP; 50 mM NH4Cl; 1.5 mM MgCl2 and 10 mM TrisHCl buffer pH 9.0 M at room temperature.18 Aliquots of 45 µL were robotically loaded into a 96-well polycarbonate plate (Corning, Corning, NY, USA), which was kept at 4°C in a mini-refrigerator. Evaporation and contamination were avoided by utilizing 35 µL light mineral oil as an overlay above the 15 µL of target template DNA.19 Taq polymerase (2.5 U) (Promega, Madison, WI) were added to the master mix immediately before the assembling procedure by the robotic system. Thermal cycling conditions were as follows: 10 min at 95°C (initial DNA denaturing step); 1 min at 94°C (DNA denaturing), 1 min at 58°C (annealing), 3 min at 72°C (extension) cycling 35 times. PCR products were analysed on an agarose gel. Detection was accomplished by staining the products with ethidium bromide. Purification of the PCR products for cycle sequencing (Sanger technique) was accomplished with a QIAquick PCR purification kit (QIAGEN, Valencia, CA, USA). Quantification of the purified DNA was performed with a Hitachi U2000 spectrophotometer, which was interfaced with the XP robotic system to avoid manual contact with amplified DNA. Additional electrophoretic separation ensured proper purification followed by adequate DNA dilution and mixing with the sense primer for the sequencing reaction with subsequent detection on an ABI sequencer.
![]() |
Results and discussion |
---|
![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
The antimicrobial potency and spectrum for 22 selected antimicrobial agents against the five most frequent UTI pathogens are summarized in Table II. For E. coli isolates, the best coverage was achieved with carbapenems (100.0%), followed by amikacin (98.1%) and extended-spectrum cephalosporins (95.896.2%). Piperacillintazobactam showed the highest activity (MIC50, 4 mg/L) and susceptibility rate (88.5%) among the penicillinß-lactamase inhibitor combinations. In spite of the high potency of the fluoroquinolones against E. coli isolates, a high resistance rate to ciprofloxacin was observed. Our study also made evident the existence of cross-resistance between ciprofloxacin and the newest quinolones such as levofloxacin and gatifloxacin.
|
Against P. aeruginosa isolates, although meropenem (MIC50, 2 mg/L), imipenem (MIC50, 4 mg/L) and cefepime (MIC50, 8 mg/L) have been the most potent compounds, the highest percentage of susceptibility was found for piperacillintazobactam (80.6%) followed by meropenem (77.8%). The low susceptibility rate (52.8%) displayed by imipenem might be due to dissemination of multi-resistant clones in the medical centres evaluated. The production of ß-lactamases coupled with loss of carbapenem-specific porins (OprD) could be the mechanisms involved in carbapenem resistance. Generally, severe P. aeruginosa infections are treated empirically with an association of a ß-lactam with an aminoglycoside; however, the low susceptibility rates to aminoglycosides could limit the effectiveness of such combined therapy in Latin America. Among the P. mirabilis isolates evaluated, low susceptibility (70.0%) to fluoroquinolones was also observed. Aztreonam, cefoxitin, ceftazidime and imipenem were active against all P. mirabilis isolates. Enterobacter spp. were resistant to the majority of ß-lactam drugs, and although cefepime showed higher activity than the other cephalosporins tested, only the carbapenems were able to inhibit the growth of 100.0% of Enterobacter spp. isolates.
The overall antimicrobial susceptibility of the UTI pathogens isolated in Latin America was meropenem (98.1%) > imipenem (95.9%) > cefepime (89.6%) > amikacin (87.7%) > ceftazidime (85.7%) > aztreonam (83.8%) > ceftriaxone (81.6%) > piperacillintazobactam (77.5%) > tobramycin (75.1%) and gentamicin (74.9%). The fluoroquinolone susceptibility rates ranged from 71.3% (grepafloxacin, data not shown) to 74.2% (gatifloxacin). The high resistance rates to fluoroquinolones among UTI pathogens in Latin America is of great concern, since these antimicrobial agents are agents frequently used in the treatment of UTIs.
Of 34 enteric bacilli (18 E. coli, 12 K. pneumoniae and four P. mirabilis isolates) identified as ESBL phenotypes using the NCCLS criteria,11 only 20 were characterized as possible ESBL producers by the Etest strip (Table IV). The performance of various ß-lactams in detecting ESBL-producing isolates was evaluated, since different types of ESBL can have different preferred ß-lactam substrates.12 Cefotaxime exhibited the highest sensitivity (90.0%) and detected the greatest number of ESBL producers among the isolates evaluated, while aztreonam exhibited the lowest sensitivity rate (65.0%). Although ceftazidime (85.0%) displayed a lower sensitivity rate than cefotaxime, it was able to detect two isolates that were not detected by any other ß-lactam. It has been reported that ceftazidime is the preferred substrate for most ESBLs isolated in the USA and Europe.12,27 However, our results agree with previous Latin American studies that reported cefotaxime as the preferred substrate in Latin America.28,29 Based on the degradation of preferred substrates, tests for detection of ESBL-producing isolates have been developed. This finding suggests that the use of both substrates (ceftazidime and ceftriaxone) would allow the detection of most ESBL-producing isolates emerging in Latin America.
|
|
As discussed previously, a high percentage of fluoroquinolone-resistant E. coli was observed in this surveillance study. Ciprofloxacin-resistant E. coli isolates were reported by all Latin American medical centres, except centres 39 and 49. The high ciprofloxacin resistance rate observed could be attributed in part to a higher prevalence of ciprofloxacin-resistant E. coli detected in one medical centre (25 of 58 ciprofloxacin-resistant E. coli isolates were obtained from medical centre 45). The resistance rates to fluoroquinolones vary from one country to another, and depend on local epidemiological factors. Generally, in the United States and Canada, E. coli isolates from patients with UTI display >95% susceptibility to fluoroquinolones.22,23,32 However, a dramatic increase in the prevalence of fluoroquinolone resistance has been reported recently, in both community and nosocomial settings in several European countries.3335 In the present study, approximately 70.0% of the E. coli resistant to quinolones were collected from community-acquired UTIs. It could reflect the overuse of the quinolones for treatment of community-acquired UTI. Some authors have advocated that quinolone resistance is higher in developing countries than in developed nations because of the use of less active quinolones, such as nalidixic acid, and/or the use of low dosages of more potent compounds such as ciprofloxacin resulting in selection of mutant isolates.35 The use of short-term treatment with quinolones for UTI, which has been encouraged by some authors, could also have been a contributory factor for selection of mutant isolates.36 On the other hand, other authors have recommended that the use of quinolones should be reduced or at least rationalized, principally among the UTIs, in order to save this potent class of antibiotics and avoid the development of resistance among the Enterobacteriaceae.32
Mutations in gyrA have commonly been localized near the amino terminus of the encoded protein, at positions 83 and 87.37,38 The vast majority of E. coli isolates evaluated in this study (Table V) demonstrated double mutations in gyrA at positions 83 (leucine replacing serine) and 87 (asparagine or tyrosine replacing aspartic acid). Among E. coli isolates, mutations in the parC gene contribute to quinolone resistance, but only in the presence of gyrA mutations. All evaluated isolates also demonstrated mutations in the parC gene at positions 80 or 84. At position 80, isoleucine or arginine replaced the amino acid serine, while in position 84 glutamic acid was replaced by lysine. Different point mutations were detected in the isolates from centre 41, probably because this laboratory receives isolates from three different hospitals. Etest detected high-level resistance to ciprofloxacin (MIC
32 mg/L) in all E. coli isolates with double mutations in gyrA and a single mutation in parC. In general, these isolates also displayed high MICs for the most recently tested fluoroquinolones. However, the isolate that exhibited a single mutation in the gyrA and parC genes demonstrated low-level resistance to ciprofloxacin (MIC, 6 mg/L) and trovafloxacin (MIC, 4 mg/L), but remained susceptible to gatifloxacin (MIC, 1.5 mg/L). These results indicate that multi-step mutations are predominantly responsible for the emergence of high-level resistance to fluoroquinolones in the Latin American Centres monitored. For this reason, recognition of strains with low-level resistance is important. In this manner, the selection of high-level resistant isolates might be avoided and the activity of the newest quinolones might be preserved.
|
![]() |
Acknowledgments |
---|
SENTRY Latin America Study Group includes: H. S. Sader (BrazilLatin America Coordinator); J. Sampaio (Laboratório Lâmina, Rio de Janeiro, Brazil); C. Zoccoli (Laboratório Médico Santa Lúzia, Florianópolis, Brazil); J. M. Casellas (Centro de Estudios en Antimicrobianos, San Isidro, Argentina); J. Smayevsky (Microbiology Laboratory CEMIC, Buenos Aires, Argentina); V. Prado (Faculdad de Medicina de Chile, Santiago, Chile); E. Palavecino (Universidad Catolica del Chile, Santiago, Chile); J. A. Robledo (Corporation Para Investigaciones Biologicas, Medellin, Colombia); J. Sifuentes-Osornio (Instituto Nacional de la Nutricion, Ciudad del Mexico, Mexico); and M. Guzman-Blanco (Caracas, Venezuela).
![]() |
Notes |
---|
![]() |
References |
---|
![]() ![]() ![]() ![]() ![]() ![]() |
---|
2 . National Nosocomial Infections Surveillance (NNIS) report, data summary from October 1986April 1996. Issued May 1996. (1996). American Journal of Infection Control 24, 3808.[ISI][Medline]
3 . Warren, J. W. (1997). Catheter-associated urinary tract infections. Infectious Disease Clinics of North America 11, 60922.[ISI][Medline]
4 . Krieger, J. N., Kaiser, D. L. & Wenzel, R. P. (1983). Urinary tract etiology of bloodstream infections in hospitalized patients. Journal of Infectious Diseases 148, 5762.[ISI][Medline]
5 . Bishara, J., Leibovici, L., Huminer, D., Drucker, M., Samra, Z., Konisberger, H. et al. (1997). Five-year prospective study of bacteraemic urinary tract infection in a single institution. European Journal of Clinical Microbiology and Infectious Diseases 16, 5637.[ISI][Medline]
6 . Platt, R., Polk, B. F., Murdock, B. & Rosner, B. (1982). Mortality associated with nosocomial urinary-tract infection. New England Journal of Medicine 307, 63742.[Abstract]
7 . Neu, H. C. (1992). Urinary tract infections. American Journal of Medicine 92, Suppl. 4A, S6370.[Medline]
8 . Jones, R. N. (1996). Impact of changing pathogens and antimicrobial susceptibility patterns in the treatment of serious infections in hospitalized patients. American Journal of Medicine 100, Suppl. 6A, S312.[Medline]
9 . Brosnema, D. A., Adams, J. R., Pallares, R. & Wenzel, R. P. (1993). Secular trends in rates and etiology of nosocomial urinary tract infections at a university hospital. Journal of Urology 150, 414 16.[ISI][Medline]
10
.
Pfaller, M. A., Jones, R. N., Doern, G. V. & Kugler, K. (1998). Bacterial pathogens isolated from patients with bloodstream infection: frequencies of occurrence and antimicrobial susceptibility patterns from the SENTRY antimicrobial surveillance program (United States and Canada, 1997). Antimicrobial Agents and Chemotherapy 42,176270.
11 . National Committee for Clinical Laboratory Standards. (1997). Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria That Grow AerobicallyFourth Edition: Approved Standard M7-A4. NCCLS, Wayne, PA.
12
.
Bush, K., Jacoby, G. A. & Medeiros, A. A. (1995). A functional classification scheme for ß-lactamases and its correlation with molecular structure. Antimicrobial Agents and Chemotherapy 39, 121133.
13 . National Committee for Clinical Laboratory Standards. (1998). Performance Standards for Antimicrobial Susceptibility TestingEighth Informational Supplement: Approved Standard M100-S8. NCCLS, Wayne, PA.
14 . Cormican, M. G., Marshall, S. A. & Jones, R. N. (1996). Detection of extended-spectrum ß-lactamase (ESBL)-producing strains by Etest ESBL screen. Journal of Clinical Microbiology 34, 18804.[Abstract]
15 . Pfaller, M. A., Wendt, C. & Hollis, R. J. (1996). Comparative evaluation of an automated ribotyping system versus pulsed-field gel electrophoresis for epidemiological typing of clinical isolates of Escherichia coli and Pseudomonas aeruginosa from patients with recurrent gram-negative bacteremia. Diagnostic Microbiology and Infectious Disease 25, 18.[ISI][Medline]
16 . Pfaller, M. A., Hollis, R. J. & Sader, H. S. (1992). Chromosomal restriction fragment analysis by pulsed-field gel electrophoresis. In Clinical Microbiology Procedures Handbook Vol. 2. (Isenberg, H. D., Ed.), pp. 10.5c.112. American Society for Microbiology, Washington, DC.
17 . Georgiou, M., Muñoz, L. R., Román, F., Cantón, R., Gómez-Lus, R., Campos, J. et al. (1996). Ciprofloxacin-resistant Haemophilus influenzae strains possess mutations in analogous positions of GyrA and ParC. Antimicrobial Agents and Chemotherapy 40, 17414.[Abstract]
18 . Blanchard, M. M., Taillon-Miller, P., Nowotny, P. & Nowotny, V. (1993). PCR buffer optimization with uniform temperature regimen to facilitate automation. PCR Method and Applications 2, 23440.
19
.
Wilke, W. W., Sutton, L. D. & Jones, R. N. (1995). Automation of polymerase chain reaction tests to achieve acceptable contamination rates. Clinical Chemistry 41, 6223.
20 . Weber, G., Riesenberg, K., Schlaeffer, F., Peled, N., Borer, A. & Yagupsky, P. (1997). Changing trends in frequency and antimicrobial resistance of urinary pathogens in outpatient clinics and a hospital in Southern Israel, 19911995. European Journal of Clinical Microbiology and Infectious Diseases 16, 8348.[ISI][Medline]
21 . Sader, H. S., Jones, R. N., Winokur, P. L., Pfaller, M. A., Doern, G. V., Barrett, T. & The SENTRY Study Group, Latin America. (1999) Antimicrobial susceptibility of bacteria causing urinary tract infections in Latin American hospitals: Results from the SENTRY antimicrobial surveillance program (1997). Clinical Microbiology and Infection 5, 47887.[Medline]
22 . Sader, H. S., Gales, A. C., Schomberg, L., Jones, R. N. & SENTRY Participants Group. (1998). Comparative evaluation of frequencies and antimicrobial susceptibilities of pathogens isolated from urinary tract infection (UTI) in North America (NA) and Latin America (LA): Results of the SENTRY Antimicrobial Surveillance Program, 1997. In Program and Abstracts of the Thirty-Eighth Interscience Conference on Antimicrobial Agents and Chemotherapy, San Diego, CA, 1998. Abstract E-39, p. 44. American Society for Microbiology, Washington, DC.
23 . Barnett, B. J. & Stephens, D. S. (1997). Urinary tract infection: an overview. American Journal of Medical Science 314, 2459.[ISI]
24 . Martinez-Martinez, L., Hernandez-Alles, S., Alberti, S., Tomas, J. M., Benedi, V. J. & Jacoby, G. A. (1996). In vivo selection of porin-deficient mutants of K. pneumoniae with increased resistance to cefoxitin and extended-spectrum cephalosporins. Antimicrobial Agents and Chemotherapy 40, 3428.[Abstract]
25 . Pangon, B., Bizet, C., Bure, A., Pichon, F., Phillipon, A., Regnier, B. et al. (1989). In vivo selection of a cephamycin-resistant, porin-deficient mutant of Klebsiella pneumoniae producing a TEM-3 ß-lactamase. Journal of Infectious Diseases 159, 10056.[ISI][Medline]
26 . Jenks, P. J., Hu, Y. M., Danel, F., Mehtar, S. & Livermore, D. M. (1995). Plasmid-mediated production of class I (AmpC) ß-lactamase by two Klebsiella pneumoniae isolates from the UK. Journal of Antimicrobial Chemotherapy 35, 2356.[ISI][Medline]
27 . Chanal, C. M., Poupart, M.-C., Sirot, D. L., Labia, R., Sirot, J. L. & Cluzel, R. A. (1992). Nucleotide sequences of CAZ-2, CAZ-6, CAZ-7 ß-lactamases genes. Antimicrobial Agents and Chemotherapy 36, 181720.[Abstract]
28 . Bauernfeind, A., Casellas, J. M., Goldberg, M., Holley, M., Jungwirth, R., Mangold, P. et al. (1992). A new plasmidic cefotaximase from patients infected with Salmonella typhimurium. Infection 20, 15863.
29 . Gales, A. C., Bolmstrom, A., Jones, R. N., Reis, A. O. & Sader, H. S. (1997). Prevalence, antimicrobial susceptibility and molecular typing of extended-spectrum beta-lactamase-producing Klebsiella pneumoniae from Brazil. In Program and Abstracts of the Thirty-Seventh Interscience Conference on Antimicrobial Agents and Chemotherapy, Toronto, Canada. Abstract D-13, p. 85. American Society for Microbiology, Washington, DC.
30 . Jacoby, J. A. & Carreras, I. (1990). Activities of beta-lactam antibiotics against Escherichia coli strains producing extended-spectrum beta-lactamases. Antimicrobial Agents and Chemotherapy 34, 85862.[ISI][Medline]
31 . Sader, H. S., Mendes, C. F., Pignatari, A. C. & Pfaller, M. A. (1996). Use of macrorestriction analysis to demonstrate interhospital spread of multiresistant Acinetobacter baumannii in Sao Paulo, Brazil. Clinical Infectious Diseases 23, 6314.[ISI][Medline]
32 . Thomson, K. S., Sanders, W. E. & Sanders, C. C. (1994) USA resistance patterns among UTI pathogens. Journal of Antimicrobial Chemotherapy 33, Suppl. A, 915.[ISI][Medline]
33 . Kresken, M., Hafner, D., Mittermayer, H., Verbist, L., Bergogne-Berezin, E., Giamarellou, H. et al. (1994). Prevalence of fluoroquinolone resistance in Europe. Infection 22, Suppl. 2, S908.[ISI][Medline]
34 . Aubert, G., Levy, P. P., Ros, A., Meley, R., Meley, B., Bourge, A. et al. (1992). Changes in the sensitivity of urinary pathogens to quinolones between 1987 and 1990 in France. European Journal of Clinical Microbiology and Infectious Diseases 11, 4757.[ISI][Medline]
35 . Acar, J. F. & Goldstein, F. W. (1997). Trends in bacterial resistance to fluoroquinolones. Clinical Infectious Diseases 24, Suppl. 1, S6773.[ISI][Medline]
36 . Margariti, P. A., Astorri, A. L. & Mastromarino, C. (1997). Urinary tract infections: risk factors and therapeutic trends. Recenti Progressi in Medicina 88, 658.[Medline]
37 . Ouabdesselam, S., Hooper, D. C., Tankovic, J. & Soussy, C. J. (1995). Detection of gyrA and gyrB mutations in quinolone-resistant clinical isolates of Escherichia coli by single-strand conformational polymorphism analysis and determination of levels of resistance conferred by two different single gyrA mutations. Antimicrobial Agents and Chemotherapy 39, 166770.[Abstract]
38 . Truong, Q. C., Ouabdesselam, S., Hooper, D. C., Moreau, N. J. & Soussy, C. J. (1995). Sequential mutations of gyrA in Escherichia coli associated with quinolone therapy. Journal of Antimicrobial Chemotherapy 36, 10559.[Abstract]
Received 24 May 1999; returned 13 September 1999; revised 18 October 1999; accepted 26 October 1999