Effect of permeabilizing agents on antibacterial activity against a simple Pseudomonas aeruginosa biofilm

In 1994, the Malm?-Klaipeda twinning programme was approved by the World Federation of Hemophilia. One of the first steps in the collaboration has been to set up a registry of the haemophilia patients in the Klaipeda area. In order to collect important clinical data the patients have been examined jointly by experts on haemophilia from the two centres. Seventeen out of 25 patients with severe haemophilia known at the Klaipeda centre were examined and compared to a matched cohort of patients from the Malm? centre. The main differences between the cohorts were that home treatment was not available to the Klaipeda patients, they received less treatment in general, had higher joint scores and more frequent bleeds. The pattern of transmission of blood-borne virus was very similar, with a high prevalence of hepatitis C antibodies. We conclude that the twinning programme between Malm? and Klaipeda has resulted in several achievements, including training of staff and a necessary inventory of the patients. This should not only form a suitable platform for the future development of haemophilia care in Lithuania, but could also serve as an example for liaisons between other haemophilia centres.     

Evaluation of the E test for antimicrobial susceptibility testing of Pseudomonas aeruginosa isolates from patients with long-term bladder catheterization

The E test was evaluated in comparison with reference agar methods (National Committee for Clinical Laboratory Standards) for the susceptibility testing of 248 Pseudomonas aeruginosa isolates from bladder-catheterized patients against nine antibiotics. The E-test MICs correlated well with those determined by the agar dilution and disk diffusion reference methods (88 and 92.5% within 1 log2 dilution step, respectively), confirming that the E test is a reliable method for the determination of MICs of antibiotics for catheterization-associated P. aeruginosa isolates.     

Detection of gyrA mutations among 335 Pseudomonas aeruginosa strains isolated in Japan and their susceptibilities to fluoroquinolones

The authors report two elderly men with diffuse meningeal enhancement 13 and 21 years following insertion of ventriculojugular shunts. Lumber puncture documenting low CSF pressures suggests that diffuse meningeal enhancement in patients with long-standing ventricular shunts may be secondary to dural venous dilatation rather than meningeal fibrosis. The authors theorize that these elderly patients, neither of whom had postural headache, may be less subject to brain "sag" because of decreased brain weight.     

Canadian Pseudomonas aeruginosa susceptibility study from 48 medical centers: focus on ciprofloxacin

The carbapenem-induced endotoxin release was evaluated using experimental models of gram-negative bacterial sepsis in Wistar rats. Infections with Escherichia coli, Serratia marcescens, Klebsiella pneumoniae, Pseudomonas aeruginosa, Proteus vulgaris and Proteus mirabilis resulted in an increase of the plasma endotoxin concentration after treatment with ceftazidime and carbapenems including imipenem, panipenem, meropenem and biapenem. Except for P. aeruginosa, the plasma endotoxin concentrations after carbapenem treatment were significantly lower than those after ceftazidime treatment. It is noteworthy that treatment of P. aeruginosa sepsis with meropenem or biapenem induced significantly more endotoxin release than other carbapenems and the endotoxin concentrations induced by these carbapenems reached those of ceftazidime treatment. The plasma endotoxin concentrations appeared to correlate with the reduction of platelet counts and the elevation of both glutamic oxaloacetic transaminase and glutamic pyruvic transaminase values.     

Penicillin-resistant mechanisms in Pseudomonas aeruginosa: binding of penicillin to Pseudomonas aeruginosa KM 338

A comparison of the binding of radioactive penicillin G to whole cells and the membrane fraction derived from Pseudomonas aeruginosa KM 338 was made. This organism has intrinsic resistance to penicillin. The binding to the membrane fraction which catalyzed peptidoglycan synthesis followed saturation type kinetics and saturation was achieved at approximately 2 nmol of penicillin G per ml, whereas binding to the whole cells was entirely of the nonsaturation type. The binding of carbenicillin to the membrane fraction was determined by competition between radioactive penicillin G and unlabeled carbenicillin for the binding sites. It was bound at the same sites in almost the same manner. When whole cells were pretreated with high concentration of unlabeled penicillin G or carbenicillin, the subsequent binding of radioactive penicillin G to the membrane fraction from carbenicillin-treated cells was entirely nonspecific, but with penicillin G-pretreated cells it was still specific. There was apparently specific binding of radioactive penicillin G to ethylenediaminetetraacetate-treated cells. P. aeruginosa KM 338 had an extremely low activity of beta-lactamase compared with other enzyme-producing organisms. This enzyme from P. aeruginosa KM 338 was of the cephalosporinase type. These data indicate that penicillin resistance of P. aeruginosa KM 338 may be a consequence of the development of a permeability barrier which prevents the antibiotic from reaching its sites of action in the cytoplasmic membrane.     

Effect of addition antimicrobial agents to denture reliners

This study aimed at evaluating the incorporation of two antimicrobial drugs (nystatin and polynoxylin) as regards: the effect of the liner on the activity of the drug, determination of the least effective concentration of each drug and its duration of action, as well as assessment of the effect of the drug on the mechanical properties and the chemical composition of the liner. Results showed that nystatin added to denture liners in three different concentrations by weight (3%, 5%, 10%) acted effectively against Candida albicans, and that there was a direct relationship between concentration of Nystatin and its duration of action. The inhibitory effect of nystatin (10%) lasted for at least 32 weeks (end of study period). Furthermore, this concentration did not affect the strength properties of the liner. On the other hand, polynoxylin inhibited a number of strains of bacteria and Candida only in high concentrations (40-60%), and these concentrations adversely affected the strength properties of the liner.     

Involvement of bacterial migration in the development of complex multicellular structures in Pseudomonas aeruginosa biofilms

Detailed knowledge of the developmental process from single cells scattered on a surface to complex multicellular biofilm structures is essential in order to create strategies to control biofilm development. In order to study bacterial migration patterns during Pseudomonas aeruginosa biofilm development, we have performed an investigation with time-lapse confocal laser scanning microscopy of biofilms formed by various combinations of colour-coded P. aeruginosa wild type and motility mutants. We show that mushroom-shaped multicellular structures in P. aeruginosa biofilms can form in a sequential process involving a non-motile bacterial subpopulation and a migrating bacterial subpopulation. The non-motile bacteria form the mushroom stalks by growth in certain foci of the biofilm. The migrating bacteria form the mushroom caps by climbing the stalks and aggregating on the tops in a process which is driven by type-IV pili. These results lead to a new model for biofilm formation by P. aeruginosa.     

Determination of the in vitro susceptibility of 220 Mycobacterium fortuitum isolates to ten antimicrobial agents

The authors investigated the in vitro susceptibility to antimicrobial agents of 220 Mycobacterium fortuitum isolates originating from clinical samples (14) of patients attending the Hospital Universitario de Canarias and Hospital del Tórax, and from environmental sources (206): 3 from sea water, 10 from the water supply and 193 from sewage. The Minimum Inhibitory Concentration (MIC) was calculated using the broth microdilution method with Mueller-Hinton Broth without supplement. Amikacin was the most efficacious antimicrobial agent against all the isolates of M. fortuitum with an MIC which was considerably lower than its critical concentration. The good results achieved with amikacin in vitro are confirmed by those obtained in vivo, with patients infected with M. fortuitum. No significant difference was found in the efficacy of amikacin and ofloxacin against all the isolates assayed.     

Italian survey on comparative levofloxacin susceptibility in 334 clinical isolates of Pseudomonas aeruginosa

A national survey on susceptibility patterns of 334 Pseudomonas aeruginosa isolates from intensive care units and hematology and oncology wards from 13 Italian hospitals compared the in vitro activity of levofloxacin, an injectable oral fluoroquinolone, to those of ciprofloxacin, ofloxacin, ceftazidime, imipenem, amikacin, and gentamicin. Amikacin and imipenem had the best susceptibility profiles. The activity of levofloxacin was superior to those of the other quinolones and was comparable to that of ceftazidime. The effect of levofloxacin in vitro on P. aeruginosa clinical isolates suggests that further clinical investigations are warranted.     

Influence of growth media on potential virulence factors of Pseudomonas aeruginosa

Potential virulence factors of three Pseudomonas aeruginosa strains after growth in three complex media (CM) and in one mineral medium (MM) were evaluated. Cell surface hydrophobicity demonstrated by adherence of bacteria to xylene as well as enzymatic activity (elastase, protease, lipase) of the strains grown in CM varied with composition of CM and with strain. All strains cultivated in CM showed higher hydrophobicity and higher elastase, protease and lipase (with the exception of one strain) activity in comparison with bacteria incubated in MM. Even no production of elastase was detected in the strains after growth in MM. Motility of bacteria was affected by culture media the least. In vitro composition of growth media influenced some potential virulence factors of P. aeruginosa.     

Organisms isolated from dogs and cats with anaerobic infections and susceptibility to selected antimicrobial agents

OBJECTIVE: To determine the prevalence of obligate anaerobic bacteria in bacterial infections in dogs and cats and susceptibility to selected antimicrobial agents. DESIGN: Case series. SAMPLE POPULATION: Specimens from 1,267 dogs and 243 cats. PROCEDURE: Standard anaerobic and aerobic bacterial culture methods were used. Anaerobic isolates were tested for susceptibility to selected antimicrobial agents. RESULTS: Obligate anaerobic bacteria were isolated from 199 (15.7%) and 69 (28.4%) specimens obtained from dogs and cats, respectively. More than half of the specimens that contained obligate anaerobic bacteria were from draining tracts (exclusively dogs), pleural fluid, abscesses, bones, the respiratory tract, or the abdominal cavity. The most commonly isolated obligate anaerobic bacteria (approx 70% of all isolates) were Bacteroides spp, Peptostreptococcus spp, Fusobacterium spp, and Porphyromonas spp. Eighty percent of the specimens that contained obligate anaerobic bacteria also contained facultative anaerobic or aerobic organisms. The organisms most commonly isolated in association with obligate anaerobic bacteria were members of the family Enterobacteriaceae (Escherichia coli was the most common), Pasteurella spp, and Staphylococcus intermedius. Ninety-seven obligate anaerobic isolates were tested for susceptibility to ampicillin, amoxicillin-clavulanic acid, chloramphenicol, clindamycin, and metronidazole. All were susceptible to amoxicillin-clavulanic acid and chloramphenicol, and most were susceptible to metronidazole. Only 71% of the Bacteroides isolates were susceptible to ampicillin, and only 83% were susceptible to clindamycin. Only 80% of the Clostridium isolates were susceptible to clindamycin, but all were susceptible to ampicillin. CLINICAL IMPLICATIONS: Data on sites and conditions from which anaerobic bacteria are commonly isolated, along with results of susceptibility testing, may be useful in designing antimicrobial treatment regimens.     

Ceftazidime, gentamicin, and rifampicin, in combination, kill biofilms of mucoid Pseudomonas aeruginosa

In continuous flow biofilm cultures in medium resembling cystic fibrosis bronchial secretions, Pseudomonas aeruginosa was not eradicated from biofilms by 1 week of treatment with high concentrations of ceftazidime and gentamicin, to which the strains were sensitive on conventional testing. The addition of rifampicin, which has little activity against the strains as measured by the minimum inhibitory concentration, led to the apparent elimination of the bacteria from the biofilms. The effect was not strain specific.     

Effect of inoculum size on antimicrobial susceptibility of Helicobacter pylori

An agar dilution assay was used to assess the effect of inoculum size and culture period on the susceptibility of 15 clinical isolates of Helicobacter pylori to ampicillin, erythromycin, tetracycline, chloramphenicol, metronidazole and tinidazole. The mean MIC of the isolates increased 2.2- to 21.2-fold as the inoculum size progressed from 10(3) to 10(7) cfu/spot. Identical results were noted when isolates were maintained for two or four days prior to testing. Inoculum size should be carefully controlled when assessing the in vitro susceptibility of Helicobacter pylori.     

Association of CD4+ T cell-dependent keratitis with genetic susceptibility to Pseudomonas aeruginosa ocular infection

The role of T lymphocytes in susceptibility to Pseudomonas aeruginosa corneal infection was studied in inbred C57Bl/6 (B6) beta2-microglobulin+/+ (beta2m+/+) and beta2m-/- knockout (KO) mice on a B6 genetic background. The corneas of both B6 and KO mice perforated by 7 days postinfection (p.i.). Histopathology revealed a similar inflammatory response characterized by an infiltration of polymorphonuclear neutrophilic leukocytes by 24 h p.i. in both groups of mice. CD4+ and CD8+ (latter absent in KO) T cells were present in cornea by 3 days p.i., and by 5 days, IL-2R-positive cells were positively immunostained. Corneas of B6 beta2m+/+ mice depleted of CD4+ T cells and infected with P. aeruginosa did not perforate at 7 days p.i. vs mice depleted of CD8+ T cells or treated with an irrelevant mAb. Neutralization of IFN-gamma before infecting B6 mice prevented corneal perforation and was associated with a lower delayed-type hypersensitivity than in B6 mice similarly treated with an irrelevant mAb. These data provide evidence that a CD4+ T cell (Th1)-dominated response following P. aeruginosa corneal infection is associated with genetic susceptibility and corneal perforation in inbred B6 mice.     

Influence of zinc on Pseudomonas aeruginosa susceptibilities to imipenem

Serial dilution susceptibility testing of imipenem against 59 clinical isolates of Pseudomonas aeruginosa, conducted simultaneously on single lots of Difco and BBL Mueller-Hinton agar (MHA), resulted in MICs for 90% of strains tested of 8 and 16 micrograms/ml, respectively. MICs for Escherichia coli, Klebsiella pneumoniae, and Pseudomonas spp. were also higher on BBL MHA. Quantification of the cation content of the two MHAs by atomic absorption spectroscopy demonstrated that the zinc concentration in BBL MHA was 15 times greater than that measured in Difco MHA (2.61 and 0.17 micrograms/ml, respectively). Concentrations of calcium, magnesium, iron, manganese, and copper in the two agars were similar. Addition of zinc to Difco MHA resulted in increases in MICs of imipenem for P. aeruginosa but not in the MICs of ceftazidime or cefpirome for P. aeruginosa (P < 0.01). A lesser zinc effect was seen on the activity of imipenem against E. coli, K. pneumoniae, and Pseudomonas spp. The activities of ceftazidime and cefpirome were similar on both MHAs when tested against all gram-negative organisms in this study. Thus, the effect of zinc in MHA was clearly demonstrated by a significant increase in the MICs of imipenem for P. aeruginosa, and, to a lesser extent, for other gram-negative bacilli.     

In vivo effect of growth-inhibitor produced by Pseudomonas aeruginosa isolates and anti-pseudomonal drugs on model infection due to Pseudomonas aeruginosa

Pseudomonas aeruginosa Nos. 1 and 5, each co-existing growth-inhibitor-producing and -nonproducing cells, were used in this study. An equal number of both cells (each 10(8) CFU/mouse) was challenged intraperitoneally to mice, and these cells in the heart blood and kidneys of mice were determined. Furthermore, the effect of piperacillin, ceftazidime and sisomicin on the cell distribution in mice was studied in the model infection due to P. aeruginosa Nos. 1 and 5. As a control experiment both cells of P. aeruginosa No. 1 were each challenged intraperitoneally at a dose of 10(8) CFU/mouse to mice of two groups, but there were no marked differences between the two types in cell counts of the heart blood or kidneys 9 hours after challenge. When a concomitant challenge of both cells (each 10(8) CFU/mouse) was performed in mice, the number of growth-inhibitor-producing cells of the heart blood and kidneys was about 100 times greater than that of the non-producing cells. These in vivo results were well comparable to the previous in vitro results and indicated that the inhibitor affected the invasion of the non-producing bacteria in the body in the model infection due to P. aeruginosa isolates consisting of the two types of cells. Similar results were obtained in mice with the model infection due to P. aeruginosa No. 5. Anti-pseudomonal drugs such as piperacillin (50 mg/mouse) and ceftazidime (50 mg/mouse) and sisomicin (1 mg/mouse) were given intramuscularly to mice infected concomitantly with both cells of P. aeruginosa No. 1.(ABSTRACT TRUNCATED AT 250 WORDS)     

Novel approaches to the discovery of antimicrobial agents

OBJECTIVE: To examine the cost-effectiveness of prenatal carrier screening for cystic fibrosis. METHODS: A cost-benefit equation was developed that was based on the hypothesis that the cost of prenatal diagnosis required to diagnose and prevent one case of cystic fibrosis should be equal to or less than the lifetime cost generated from the birth of a neonate with cystic fibrosis. The formula was adjusted because a woman's positive or negative carrier status remains unchanged, thus eliminating the need for testing in subsequent pregnancies. The formula was manipulated to identify the optimal cost per screening test, as well as the net cost savings per prenatally diagnosed case of cystic fibrosis for various racial or ethnic groups. Sensitivity analyses included some key assumptions regarding the cost per screening test ($50-150), patient screening acceptance rates (25-100%), and therapeutic abortion rates (50-100%). RESULTS: Assuming therapeutic abortion rates of 50-100%, the net savings per prenatally diagnosed case of cystic fibrosis are $58,369-$382,369 among whites. Given the previously reported patient screening acceptance rates of 50-78%, the overall annual cost savings in the United States for whites are $161-251 million. However, the screening program was not found to be cost-effective for blacks, Asians, or Hispanics. CONCLUSION: Under most assumptions and sensitivity analyses, a prenatal cystic fibrosis-carrier screening program appears to be cost-effective.     

Iron-limited biofilms of Candida albicans and their susceptibility to amphotericin B

A perfused biofilm fermentor, which allows growth-rate control of adherent microbial populations, was used to assess whether the susceptibility of Candida albicans biofilms to antifungal agents is dependent on growth rate. Biofilms were generated under conditions of glucose limitation and were perfused with drugs at a high concentration (20 times the MIC). Amphotericin B produced a greater reduction in the number of daughter cells in biofilm eluates than ketoconazole, fluconazole, or flucytosine. Similar decreases in daughter cell counts were observed when biofilms growing at three different rates were perfused with amphotericin B. In a separate series of experiments, intact biofilms, resuspended biofilm cells, and newly formed daughter cells were removed from the fermentor and were exposed to a lower concentration of amphotericin B for 1 h. The susceptibility profiles over a range of growth rates were then compared with those obtained for planktonic cells grown at the same rates under glucose limitation in a chemostat. Intact biofilms were resistant to amphotericin B at all growth rates tested, whereas planktonic cells were resistant only at low growth rates (相似文献