Risk-factors for emerging bloodstream infections caused by extended-spectrum beta-lactamase-producing Escherichia coli.

Risk-factors for bloodstream infections caused by extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli were investigated using an exploratory case-double control study in which 43 cases (70% producing CTX-M enzymes) were compared with: (i) 86 patients with bacteraemia caused by non-ESBL-producing E. coli; and (ii) 86 hospitalised patients. Previous follow-up as an outpatient, urinary catheterisation and use of oxyimino-beta-lactams or fluoroquinolones were independent risk-factors for ESBL-producing E. coli among patients with E. coli bacteraemia, and previous use of oxyimino-beta-lactams or fluoroquinolones were also independent risk-factors among hospitalised patients. These findings may help in identifying patients at greater risk for bloodstream infection caused by ESBL-producing E. coli in endemic areas.

Comparative penetration of lomefloxacin and other quinolones into human phagocytes.

The penetration of lomefloxacin into human polymorphonuclear leukocytes (PMNs) and peritoneal macrophages (PMphis) was evaluated using a fluorometric assay. Lomefloxacin reached high intracellular concentrations into PMNs at extracellular concentrations of 2 and 5 mg/L (cellular to extracellular concentration ratio [C/E] greater than 4). At the same conditions (20 minutes incubation; extracellular concentrations: 2 mg/L) lomefloxacin uptake by human PMNs (C/E: 7.9 +/- 2.6) was slightly higher than those of norfloxacin (C/E 5.1 +/- 1.8), ciprofloxacin (C/E: 6.2 +/- 2.0), and ofloxacin (C/E 7.1 +/- 2.6). Lomefloxacin penetration into human PMphis was significantly lower than PMNs but still with C/E ratios greater than 4. Entry of lomefloxacin into phagocytes was not affected by cell viability but was environmental-temperature dependent. It is concluded that lomefloxacin and the other quinolones evaluated reach high intracellular concentrations in human phagocytic cells.

Single doses of ofloxacin in uncomplicated gonorrhoea.

The clinical efficacies of 2 different single-dose oral treatments of ofloxacin were evaluated in a double-blind, randomised study of 60 males with gonococcal urethritis. 30 patients received a single dose of ofloxacin 100mg and 30 received a single dose of ofloxacin 200mg. The minimal inhibitory concentrations of ofloxacin against all isolates were less than or equal to 0.25 mg/L. Neisseria gonorrhoeae was eradicated from all 50 patients evaluated and clinical cure was achieved in 84%. In total, 8 patients developed post-gonococcal urethritis, although there was a significantly (p less than 0.05) lower rate of post-gonococcal urethritis in the group treated with ofloxacin 200mg. In conclusion, a single oral dose of ofloxacin 100mg could be an alternative treatment for uncomplicated gonorrhoea.

Entry of lomefloxacin and temafloxacin into human neutrophils, peritoneal macrophages, and tissue culture cells.

The uptake of lomefloxacin (difluoroquinolone) and temafloxacin (trifluoroquinolone) by human polymorphonuclear leukocytes (PMNs), peritoneal macrophages (PM phis), and two tissue culture cells (McCoy and Vero) was measured by a fluorometric assay. Both antimicrobials reached high intracellular concentrations in PMNs [cellular to extracellular ratio (C/E) greater than 4], in PM phis (C/E greater than 3) and lower in tissue culture cells (C/E greater than 1) at an extracellular concentration of 5 mg/L. Lomefloxacin uptake by PMNs was more rapid than that of temafloxacin. Entry of both quinolones into PMNs was environmental temperature-dependent, but not affected by cell viability. Ingestion of opsonized Staphylococcus aureus did not affect the ability of PMNs to concentrate these antimicrobials. Ingestion of opsonized zymosan or stimulation with phorbol myristate acetate significantly increased the PMN association of both quinolones, this effect being particularly marked with temafloxacin. It is concluded that both lomefloxacin and temafloxacin are markedly concentrated within human phagocytes and tissue culture cells, although this phenomenon is not dependent on the degree of fluorination of the molecule.

Effect of a siliconized latex urinary catheter on bacterial adherence and human neutrophil activity.

The effect of a siliconized latex urinary catheter on the in vitro adherence and growth of Pseudomonas aeruginosa and Escherichia coli and on the activity of human polymorphonuclear leukocytes (PMNs) was assessed. The adherence of P. aeruginosa to latex catheters was significantly greater than that of E. coli, being 30 times higher at 6 hr of incubation. The survival of E. coli (10(8) CFU/ml) in phosphate-buffered saline (PBS) containing segments of siliconized latex catheters was lower than in the controls. This effect seems to be due to a toxic effect produced by substances eluted into the medium from the catheters, as the viability of E. coli (10(3) CFU/ml) in eluates obtained from the incubation of catheters in PBS (24 hr; 37 degrees C) was significantly lower than in the controls. These phenomena were not observed with P. aeruginosa. The incubation of human PMNs with either catheter segments or eluates did not affect the uptake of opsonized E. coli but significantly decreased the production of superoxide radicals by these phagocytes. It is concluded that the adherence of P. aeruginosa to these catheters is higher than that of E. coli and that the latex urinary catheter used herein elutes substances that are in vitro toxic for E. coli and the oxygen-dependent bactericidal mechanisms of human PMNs.

Effect of plastic catheter material on bacterial adherence and viability.

The kinetics of adherence of single isolates of Staphylococcus aureus, S. epidermidis, Pseudomonas aeruginosa and Escherichia coli to catheters made of polyvinyl chloride (PVC), Teflon, siliconised latex, polyurethane and Vialon was evaluated by a radiometric assay. Radiolabelled bacteria (10(8) cfu/ml) were incubated in vials containing 1-cm lengths of catheter for up to 3 days. The peak of maximal adherence to each biomaterial was reached after 24 h for P. aeruginosa and after 72 h for the other strains. Bacterial adherence to PVC and siliconised latex was significantly higher (2-6 times; p less than 0.05) than to the other biomaterials for all the strains. The lowest values of adherence were observed with polyurethane and Vialon for the staphylococci but with Teflon for E. coli and P. aeruginosa. Bacterial viability and growth was evaluated in eluates obtained from incubation of segments of each catheter in buffer for 24 h. None of the eluates affected the viability of the staphylococci. However, all of them, significantly increased the growth of E. coli and P. aeruginosa with the exception of the eluate from siliconised latex, in which the inoculum count was reduced to an undetectable level for E. coli. We conclude that bacterial adherence to catheters may depend in part on the nature of the biomaterial and that certain substances eluted from the catheters may affect the viability and growth of different micro-organisms.

Kinetics of adherence of mucoid and non-mucoid Pseudomonas aeruginosa to plastic catheters.

The adherence of six non-mucoid and three mucoid strains of Pseudomonas aeruginosa to polyvinyl chloride (PVC), polyurethane (PU) and siliconised latex (SL) was evaluated by a radiometric method and scanning electronmicroscopy. Initially greater numbers of mucoid than non-mucoid strains adhered to all three materials. Hydrophobic non-mucoid strains adhered more efficiently than hydrophilic strains. Numbers of adherent non-mucoid P. aeruginosa cells increased with time, reaching a peak, which was different for each strain, at 1-4 h for PU, 4 h-2 days for SL and 2-3 days for PVC; thereafter a gradual decrease was observed, maximal and final values of adherence being higher with PVC and SL than with PU. Adherence of mucoid strains increased with time in 3-5 days, until a steady state was reached. We conclude that although non-mucoid strains of P. aeruginosa initially colonise plastic surfaces better than mucoid strains, mucoid strains also persist on these surfaces.

Activity of eight antibacterial agents on Staphylococcus epidermidis attached to Teflon catheters.

The presence of a Teflon catheter had no effect on the in-vitro activity of a range of antibacterial agents against slime producing and non-producing Staphylococcus epidermidis strains as determined by a microdilution assay. The susceptibility of S. epidermidis attached to Teflon catheters for 6, 24 and 48 h was also evaluated. MICs for planktonic and attached bacteria were similar. When bacteria attached to Teflon for 6 h were used as inocula, MBC values increased 32-8192-fold for the antibacterial agents tested. Similar results were observed when bacteria attached for 24 and 48 h were used as inocula. The activity of a high concentration (16 x MBC) of these antimicrobial agents against S. epidermidis biofilms in Teflon catheters was evaluated; for five slime non-producing strains, the highest reduction (around 99%) in bacterial viability was produced by cloxacillin and teicoplanin; for the slime producers, the highest effect (99.5% reduction) was shown by amikacin, clindamycin cloxacillin and ciprofloxacin but all cases still showed bacterial counts higher than 10(3) cfu/catheter segment. It is concluded that adherence of S. epidermidis to Teflon catheters decreases the bactericidal activity of the antibacterial agents tested in vitro.

Detection of enterovirulent Escherichia coli associated with diarrhoea in Seville, southern Spain, with non-radioactive DNA probes.

To assess the role of diarrhoeagenic Escherichia coli in Southern Spain, faecal samples from 135 patients with diarrhoea and 40 healthy subjects from Seville, Andalusia, were investigated. In this prospective study, enterovirulent E. coli were identified by hybridisation with five non-radioactive DNA probes specific for enterotoxigenic E. coli (ETEC), enteropathogenic E. coli (EPEC) and verocytotoxin-producing E. coli (VTEC). Probe-positive strains were isolated from four patients (3%) with diarrhoea and from none of the healthy controls. Two patients harboured ETEC and two patients had EPEC probe-positive strains in their faeces. No VTEC were isolated during this study. Salmonella spp. were the most frequently identified enteric pathogens, accounting for 10% of the cases, followed by Campylobacter jejuni (3%) and diarrhoeagenic E. coli (3%). This study indicates that enterovirulent E. coli play a modest role in the aetiology of diarrhoea among the indigenous population of Southern Spain.

Uptake and intracellular activity of ketolide HMR 3647 in human phagocytic and non-phagocytic cells.

OBJECTIVE: To evaluate the uptake of HMR 3647 into human neutrophils (PMNs), human peritoneal macrophages (PMOs) and tissue-cultured cells (epithelial cells and fibroblasts), and to assess the intracellular activity of this drug. METHOD: Cell uptake of HMR 3647 was measured by radiometric assay, as described by Klemper and Styrt. Intracellular activity was determined by incubation for 3 h of PMNs containing bacteria in the presence of HMR 3647. RESULTS: The intracellular concentrations were 130 and 71 times higher than extracellular concentrations in PMNs and PMOs, respectively (extracellular concentrations: 2-25 mg/L). The cellular-to-extracellular concentration ratios (C/E) for tissue-cultured cells were lower than those obtained in phagocytic cells but still greater than 5. The uptake of HMR 3647 was rapid and non-saturable in all cells. HMR 3647 was released slowly from phagocytic cells. HMR 3647 (extracellular concentration: 0.5-10 mg/L) did not significantly reduce the intracellular survival rate of Staphylococcus aureus ATCC 25923 in PMNs. CONCLUSIONS: HMR 3647 reaches intracellular concentrations several times higher than extracellular concentrations within phagocytic and non-phagocytic cells. The slow efflux of this drug from phagocytic cells suggests that these cells may be a vehicle for it, delivering it to sites of infection.

Cytomegalovirus mononucleosis as a cause of prolonged fever and prominent weight loss in immunocompetent adults.

Four immunocompetent adults presented with protracted fever lasting > 6 weeks and severe weight loss, associated with primary cytomegalovirus (CMV) infection. Each patient had spleen enlargement, lymphocytosis and hypertriglyceridaemia, but recovered spontaneously. A further 20 immunocompetent patients with primary CMV infection were also reviewed, and all presented the usual clinical picture of CMV mononucleosis. It was concluded that CMV mononucleosis should be considered in the differential diagnosis in patients with prolonged fever and weight loss if lymphocytosis is present.

Evaluation of the WIDER I system for antimicrobial susceptibility testing of clinical isolates of Haemophilus influenzae and Streptococcus pneumoniae.

The aim of this study was to evaluate the WIDER I system for susceptibility testing of Haemophilus influenzae and Streptococcus pneumoniae. MICs of 12 antimicrobials against 42 H. influenzae and 58 S. pneumoniae strains were determined using 1W MIC panels and compared with those obtained by microdilution. Overall essential agreements were >99%. Very major errors were not detected. Major errors occurred with ampicillin (1.7% H. influenzae). Minor errors were 2.3% (amoxicillin-clavulanate, cefuroxime, chloramphenicol), 7.1% (ampicillin) and 16.7% (clarithromycin) for H. influenzae, and 1.7% (chloramphenicol, erythromycin, meropenem), 3.4% (amoxicillin-clavulanate, cefuroxime, tetracycline) and 8.6% (levofloxacin) for S. pneumoniae. The WIDER I system is a reliable method for susceptibility testing of H. influenzae and S. pneumoniae.

Uptake and intracellular activity of ofloxacin isomers in human phagocytic and non-phagocytic cells.

The penetration and intracellular activity of ofloxacin and its isomers (levofloxacin and D-ofloxacin) into human polymorphonuclear leucocytes (PMN), human peritoneal macrophages (PMphi) and tissue cultured epithelial cells (McCoy) were evaluated. The cellular to extracellular concentration (C/E) values of the three fluoroquinolones were higher than 3.6 and 2.6 in PMN and PMphi, respectively. The C/E ratios in McCoy cells were lower than those in PMN, but still higher than 2.0. The uptake of ofloxacin and its isomers was rapid, non-saturable and reversible. All quinolones (extracellular concentrations: 2, 5 and 10 mg/l) produced a significant reduction of viable intraphagocytic Staphylococcus aureus in phagocytic cells. We concluded that ofloxacin and its isomers reach high intracellular concentrations in phagocytic and non phagocytic cells while remaining active in the former.

Evaluation of in vitro activity of a new fluoroquinolone trovafloxacin (cp-99,219) compared with other anti-anaerobic antimicrobials against members of the Bacteroides fragilis group.

The in vitro activity of a new fluoroquinolone, trovafloxacin (CP-99,219) was compared with that of ten other agents against 100 clinical isolates in the Bacteroides fragilis group. Trovafloxacin was the most active quinolone (MIC(90), 1 microg/ml) followed by sparfloxacin (MIC(90), 8 microg/ml), levofloxacin (MIC(90), 16 microg/ml) and ofloxacin (MIC(90), 32 microg/ml). Ciprofloxacin was the least active quinolone (MIC(90), 64 microg/ml). Metronidazole, chloramphenicol, imipenem and piperacillin/tazobactam, showed excellent activity with an MIC(90) of 1, 8, 0.25 and 16 microg/ml, respectively. Cefoxitin showed good activity and piperacillin was the least active compound. B. vulgatus and B. ovatus were the most resistant species to trovafloxacin among those of the B.fragilis group with an MIC(90) of 4 microg/ml while B. fragilis and B. thetaiotaomicron were the most susceptible (MIC(90), 1 microg/ml).

Evolution of the antimicrobial susceptibility of B. fragilis group at the university hospital of Seville (Spain) between 1977 and 1995.

A susceptibility survey of the B. fragilis group divided into three periods was carried out between 1977 and 1995 at the University Hospital of Seville (Spain) using the agar dilution method. No chloramphenicol, imipenem or meropenem-resistant strains were found. Metronidazole-resistant strains (2%) were isolated only in the first period. The most active beta-lactam drugs were piperacillin and ceftizoxime (resistance rate 16%), followed by ticarcillin mezlocillin and azlocillin (25%) and cefotaxime, cefotetam, and cefmetazol (around 40%). All strains tested were resistant to ampicillin and 4% to ampicillin/sulbactam. Cefoxitin resistance increased from 10% in the first two periods to 21% in the third and that of clindamycin from 12% in 1982 to 29% in 1987 and 50% in 1995.

Effect of three plastic catheters on survival and growth of Pseudomonas aeruginosa.

The effect of polyvinylchloride (PVC), polyurethane (PU) and siliconized latex (SL) catheters on the survival and growth of six non-mucoid and three mucoid strains of Pseudomonas aeruginosa was evaluated. Pseudomonas aeruginosa (1 x 10(8)) was incubated in PBS alone (control) or with 30 1-cm length segments of each catheter and the number of viable microorganisms was determined after 8 h, 1, 2, 5, 7 and 10 days. The presence of PVC catheters significantly favoured the survival and growth of non-mucoid strains in comparison to the control (P less than 0.05 at 5 days, P less than 0.01 at 7 days and thereafter); a similar result was observed with SL catheters (P less than 0.05 at 2 days, P less than 0.01 at 5 days and thereafter). No differences were observed with PU catheters. The number of mucoid microorganisms decreased with time in all controls and suspensions containing segments of catheter, but non-mucoid revertants appeared and quickly increased in the presence of PVC and SL (but not PU) catheters. Eluates of PBS previously containing PVC or SL segments induced a 100- to 500-fold increase in the growth of a non-mucoid strain in comparison with PBS alone. It is concluded that some plastic catheters can release substance(s) that favour the viability of P. aeruginosa.

Effect of polyurethane catheters and bacterial biofilms on the in-vitro activity of antimicrobials against Staphylococcus epidermidis.

The effect of two polyurethane ['Cavafix Certo' (CAV); 'Viacath' (VIA)] catheters on the in-vitro activity of amikacin (AN), clindamycin (CM), cloxacillin (CX), ciprofloxacin (CIP), vancomycin (VA), teicoplanin (TEI) and daptomycin (DAP) against slime producing and non-producing Staphylococcus epidermidis strains was determined using a microdilution assay. None of the antimicrobial agents was significantly affected in the presence of the catheters. The susceptibility of S. epidermidis attached to CAV and VIA catheters was also evaluated. Minimum inhibitory concentration (MIC) values were similar when planktonic and attached bacteria were compared. Minimum bactericidal concentrations (MBCs) markedly increased in the presence of 6 and 48 h bacterial biofilms. These increases in MBC values occurred when either slime producing or non-producing strains were used, and in most cases were higher for CAV catheters than for VIA catheters. This phenomenon was shown not to be due to differences in bacterial adherence. It is concluded that the in-vitro bactericidal activity of certain antimicrobials markedly decreased when bacteria adhered to plastic catheters, but this effect could have been dependent partially on the nature of the catheters.

[Comparison of the activity of different macrolides on strains of Haemophilus influenzae]. / Actividad comparada de diversos macrólidossobre cepas de Haemophilus influenzae. Grupo Español de Estudio de la Actividad de Macrólidos sobre H. influenzae.

In this study we evaluated the activity of macrolides and b-lactam antimicrobials on Haemophilus influenzae isolated in 1998 in eight Spanish cities. A total of 174 clinical isolates were examined. Overall, 29% of the isolates were found to produce b-lactamase. Azithromycin was the most active of the macrolides tested in this study (MIC90 of 4 mg/l); no azithromycin-resistant strains were found. Ampicillin resistance was 29%. We found one strain intrinsically resistant to beta-lactam agents (0.65% overall); and two beta-lactamase-positive strains that were resistant to amoxicillin-clavulanic acid (1.2%). The presence of these strains, while uncommon at present, makes it necessary to test the activity of antimicrobial drugs on H. influenzae.

[In vitro activity of azithromycin against clinical isolates of Acinetobacter baumannii]. / Actividad in vitro de azitromicina frente a aislamientos clínicos de Acinetobacter baumannii.

The activity of azithromycin against 225 clinical strains of Acinetobacter baumannii isolated consecutively from 26 Spanish hospitals in November 2000 was studied. The MICs of azithromycin were determined by microdilution, according to the NCCLS guidelines. The bactericidal activity of azithromycin against 15 clonally unrelated A. baumannii strains with different antimicrobial susceptibility patterns was tested using the subculture method. The killing-curves method was also performed against five strains with different susceptibility to azithromycin. The MIC(50) and MIC(90) of azithromycin were 32 and 64 mg/l, respectively. Moderate bactericidal activity was observed in 14 out of the 15 strains evaluated by the subculture method (MBCs from 1 to 4 dilution steps higher than the MICs) and by the killing-curve method. For three strains the number of CFU/ml was reduced 1 to 1.4 log by concentrations of azithromycin equivalent to 1 and 4 times their MICs. lt is concluded that azithromycin has moderate bactericidal activity against the strains of A. baumannii evaluated.

Clinical experience with cefotaxime (HR-756) in surgical patients.

Twenty adult surgical patients aged between 15 and 83 years (mean:45), 10 of whom had wound infections (one complicated with septicaemia), 2 with septicaemia, 1 with gynecological infection complicated with endocarditis, 5 with urinary tract infections and 2 with lower respiratory tract infections, were treated with parentally-administered cefotaxime. Aetiology was Proteus 7, E. coli 4, Pseudomonas 3, Enterobacter 2, Klebsiella 2 and 2 Serratia. Susceptibility testing was determined by the agar dilution method, with MIC values ranging from 0.01 to 5 microgram/ml, with two urinary isolates of Pseudomonas with MIC of 20 microgram/ml. Clinical responses were excellent in 13 (65 per cent) cases, moderate in 2 (10 per cent) and 3 (15 per cent) failed to respond to therapy. Clinical assessment was not possible in three patients. Bacteriological responses were excellent in 14 (70 per cent) cases, poor in 4 (20 per cent) and in two there was no follow-up. Systemic tolerance was good in all patients except one.