• Radiometric Data Showing Results Of Typical Mic Determinations For Machine

Objective: To study the routine use of the E‐test for susceptibility testing of penicillin‐resistant Streptococcus pneumoniae. Methods: A multicenter study of penicillin‐resistant S. Pneumoniae (PRSP) was carried out in Brittany, France (10 general hospitals, and two university hospitals including a coordinating center). Each hospital detected PRSP by the oxacillin (5‐μg) disk method and determined the MICs of penicillin G, amoxicillin and cefotaxime by the E‐test under routine conditions.

All the PRSP strains were collected in a coordinating center and the MICs were checked by the agar dilution method. The classifications obtained from the MICs determined by the E‐test and by the reference method were compared. Results: Between 1 July 1993 and 30 June 1994, 128 PRSP strains were collected. Agreement within 1 log 2 dilution was obtained for only 62% of strains with benzylpenicillin, 72.5% with amoxicillin and 76% with cefotaxime. These data are well below published values. In addition, 52% of the strains found to be penicillin‐resistant by the reference technique were of intermediate resistance according to the E‐test.

There were major differences in the quality of the results obtained by the participating laboratories. Conclusions: There are problems of standardization in the routine use of the E‐test. Microbiologists should therefore take particular care when performing the test and when reading the results, and ensure that reference strains are included in the assay.

INTRODUCTION Pneumococci have become considerably more resistant to benzylpenicillin in recent years -, and 25.1% of the isolates tested in France in 1993 were resistant to this antibiotic. This has led to clinical problems in the treatment of meningitis and acute otitis -. The degree of resistance demonstrated depends upon the specific β‐lactam used, and it is the high resistance of some strains that is mainly responsible for treatment failures ,. In addition, although third‐generation cephalosporins may be indicated in the most severe disorders, the development of strains that are highly resistant to these molecules can jeopardize their effectiveness ,.

It is therefore important to specify the degree of resistance of the strain by determining the minimal inhibitory concentrations (MICs) of individual relevant antibiotics. The technique used should be reliable, easy to perform, rapid and reproducible.

Several studies have shown that the E‐test (AB Biodisk, Solna, Sweden) fulfills these requirements and appears to be suitable for determining the MICs of β‐lactams for Streptococcus pneumoniae -. Hence this technique is now routinely used in many diagnostic laboratories. However, the widespread use of the test raises problems of inter‐laboratory standardization and requires reexamination of the reliability of the test under these new conditions of use. This study was therefore carried out to compare the MICs obtained with the E‐test used under routine conditions in 12 hospital laboratories in Brittany, France, with those obtained by a reference technique of agar dilution performed in a single coordinating laboratory. The results indicate that the E‐test is not as effective under routine conditions as in the controlled trials carried out in reference centers, and that its use may lead to errors of classification.

RESULTS In total, 128 PRSP strains were sent to the coordinating center between 1 July 1993 and 30 June 1994 by eight of the 12 participating hospitals. The MICs of benzylpenicillin for 121 strains, of amoxicillin for 120 strains and of cefotaxime for 119 strains were compared by the two methods. The MICs obtained by the two methods were used to classify the strains into susceptible, intermediate and resistant according to the CASFM criteria.

The MICs obtained by the E‐test were compared to those obtained by the reference test. The MICs were in agreement within ±1 log 2 dilution for benzylpenicillin in 75/121 strains (62%), in 87/120 strains (72.5%) for amoxicillin, and in 90/119 strains (76%) for cefotaxime. Statistically significant differences in the level of discrepancy between the two techniques were shown among the individual centers ( p. aAgreement:% of strains with a difference in MICs ≤ ±1 log 2. bNA: not applicable because not enough strains.

These differences resulted in many strains being classified as susceptible, intermediate or resistant differently, depending on the method used to measure the MIC. The classification based on the MIC obtained by the E‐test was in agreement with that based on the MIC obtained by the reference test for only 66/121 strains (54.5%) tested for benzylpenicillin. The agreements were a little better for amoxicillin (62% of strains) and cefotaxime (68%). Most of the minor errors for benzylpenicillin and amoxicillin occurred around the breakpoint separating intermediate and resistant strains. Because the MICs for cefotaxime were somewhat lower, these minor errors occurred around the threshold between intermediate and susceptible strains. The MICs checked by the CNRP for the 10 strains for which the difference between the two techniques was ≥ ±2 log 2 dilutions for benzylpenicillin were all within ±1 log 2 dilution of the MICs determined by the reference method by the coordinating center, for all three antibiotics tested (data not shown). The E‐tests for benzylpenicillin were checked by the coordinating center on 33 strains having a difference between the E‐test and reference method values of ≥ ±1 log 2 dilution.

They were within ±1 log 2 dilution of the MICs obtained by the reference technique (data not shown).

Abstract The recent upsurge in the incidence of tuberculosis with significant emergence of multidrug-resistant cases has focused on the priority of discovering effective new drugs and on the strategies to augment the potential of existing drugs against Mycobacterium tuberculosis. In the present study, we investigated cerulenin and trans-cinnamic acid, which have recently been shown to augment the activity of various antibiotics against Mycobacterium avium Antimicrob.

Radiometric Data Showing Results Of Typical Mic Determinations For Machine

Agents Chemother. 38 (1994) 2287–2295, to enhance the activity of isoniazid, rifampin, ofloxacin, amikacin and clofazimine against M. The synergy observed was compared with identical combinations using ethambutol, a cell wall-inhibiting drug used in standard antituberculous chemotherapy. The results showed that ethambutol resulted in synergistic activity in 12/30 drug combinations, as compared to 15/36 for cerulenin and 10/18 for trans-cinnamic acid.

This increase in drug activity was even observed with drug-resistant isolates. Use of novel antimicrobials and understanding of their mechanisms of action may be an effective strategy to determine previously undescribed targets for future drug development. Validation of radiometric results by viable count enumeration for drug combinations using rifampin (RIF) and ethambutol (EMB) against five isolates of M. D0, day 0; D6, day 6.

See the legend to for the drug concentrations used. 2.3 Bacterial viable count determination The bacterial viability was determined by plating the bacterial suspensions from individual Bactec vials at the beginning and at the end of the experiments. For this purpose, 0.1 ml of the culture from Bactec vials was taken and serially diluted 10-fold to provide successive dilutions ranging from 10 −1 to 10 −5. Bactericidal activity was determined by plating a 0.1-ml aliquot from each of the 10-fold dilutions on 7H11 agar plates.

CFU counts were assessed after 21 days of incubation at 37°C. The successive dilutions and the minimal plating volume used under our experimental conditions avoided any artefactual decrease of bacterial viable counts because of drug carry-over. The results are expressed as mean viable count±standard error.

2.4 Drug-combination studies These studies were performed using previously published x/ y methodology, as follows; x represents the radiometric GI of a drug combination, which was divided by y, which represents the lowest GI obtained with any of the single drugs used within the combination tested. An x/ y quotient of. All inhibitors and drugs were used at their sub-MIC levels and individual values were chosen on the basis of individual MICs for each drug versus each isolate. Ethambutol (EMB), cerulenin (CER) and trans-cinnamic acid (TCN) were used at a fixed sub-MIC of 0.5, 1.0, and 1.0 µg ml −1. Sub-MICs (µg ml −1) for other compounds for H37Rv, 90–0145, 90–0201, 90–0216, 90–0233, and 92–0492 isolates, respectively, were: isoniazid (INH, 0.01, 0.02, 0.02, 0.01, 0.5, 1.0); rifampin (RIF, 0.05, 0.05, 0.05, 0.05, 0.05, 2.0); ofloxacin (OFLO, 0.25, 0.25, 0.25, 0.25, 0.25, 0.5); amikacin (AMIK, 0.2, 0.2, 0.2, 0.2, 0.2, 0.4); clofazimine (CLOFA, 0.05, 0.05, 0.05, 0.1, 0.1, 0.1). All inhibitors and drugs were used at their sub-MIC levels and individual values were chosen on the basis of individual MICs for each drug versus each isolate.

Ethambutol (EMB), cerulenin (CER) and trans-cinnamic acid (TCN) were used at a fixed sub-MIC of 0.5, 1.0, and 1.0 µg ml −1. Sub-MICs (µg ml −1) for other compounds for H37Rv, 90–0145, 90–0201, 90–0216, 90–0233, and 92–0492 isolates, respectively, were: isoniazid (INH, 0.01, 0.02, 0.02, 0.01, 0.5, 1.0); rifampin (RIF, 0.05, 0.05, 0.05, 0.05, 0.05, 2.0); ofloxacin (OFLO, 0.25, 0.25, 0.25, 0.25, 0.25, 0.5); amikacin (AMIK, 0.2, 0.2, 0.2, 0.2, 0.2, 0.4); clofazimine (CLOFA, 0.05, 0.05, 0.05, 0.1, 0.1, 0.1).

All the antibiotics were used at their sub-MIC levels and individual values were chosen on the basis of individual MICs for each drug versus each isolate. My disney kitchen download for mac. Ethambutol (EMB), cerulenin (CER) and trans-cinnamic acid (TCN) were used at a fixed sub-MIC of 0.5, 1.0, and 1.0 µg ml −1.

Sub-MICs (in µg ml −1) for other compounds for H37Rv, and isolates 90–0145, 90–0201, 90–0216, 90–0233, and 92–0492 were, respectively, isoniazid (INH, 0.01, 0.02, 0.02, 0.01, 0.5, 1.0); rifampin (RIF, 0.05, 0.05, 0.05, 0.05, 0.05, 2.0); ofloxacin (OFLO, 0.25, 0.25, 0.25, 0.25, 0.25, 0.5); amikacin (AMIK, 0.2, 0.2, 0.2, 0.2, 0.2, 0.4); and clofazimine (CLOFA, 0.05, 0.05, 0.05, 0.1, 0.1, 0.1). The detailed x/ y quotient results obtained are shown in the legend to. 2.5 Drugs Ofloxacin (Hoechst-Marion-Roussel, Romainville, France), and clofazimine (Ciba-Geigy, Basel, Switzerland) were kindly provided by their manufacturers, whereas all other drugs were purchased from Sigma, St. Louis, MI, USA. 3 Results and discussion The results obtained are summarized in. Five antituberculous drugs were used in this investigation for combination studies with ethambutol, cerulenin and trans-cinnamic acid. To select the sublethal concentrations to be used, MICs were determined for each of the strains used.

Examination of CFU following exposure to cerulenin and trans-cinnamic acid revealed that trans-cinnamic acid was bacteriostatic even at concentrations as high as 200 µg ml −1, whereas cerulenin was bacteriostatic until 50 µg ml −1, being able to kill ≥2 log of initial inoculum at 150 µg ml −1. On the other hand, ethambutol was bactericidal at 2×MIC concentrations for all three isolates (results not shown).