A replicator method for the combined determination of minimum inhibitory concentration and minimum bactericidal concentration

Minimal bactericidal concentrations (MBCs) of nine antimicrobial agents were determined for clinical isolates by a replica plating method. Membranes were placed on the antibiotic-containing plates and the organisms replicated onto the membranes. After 18 h incubation the minimal inhibitory concentrations (MICs) were determined, the membranes were transferred to antibiotic-free plates and incubated a further 18 h and the MBCs determined. MICs and MBCs were also determined in broth. The reproducibility of the 'membrane' method and the agreement of these results for MIC and MBC with the agar and/or broth methods was satisfactory for most antibiotics, within one two-fold dilution. With sulphamethoxazole, trimethoprim and co-trimoxazole the results were less satisfactory, especially with Gram-negative rods, but agreement within two dilutions could be achieved.     

The minimum inhibitory and bactericidal concentrations of antibiotics and anti-ulcer agents against Campylobacter pyloridis

The minimum inhibitory and bactericidal concentrations of twenty-two antibiotics, in liquid and solid media, and three anti-peptic ulcer drugs in liquid media were determined against twenty isolates of Campylobacter pyloridis. Camp. pyloridis was very susceptible to most antibiotics, more so than Camp. jejuni and Camp. fetus, but was resistant to nalidixic acid. The MIC90 of cimetidine against Camp. pyloridis was 512 mg/l, and of ranitidine was 6400 mg/l. The difference in susceptibility patterns between Camp. pyloridis and other campylobacters may indicate that Camp. pyloridis is not a member of the Campylobacter genus.     

Comparison of inhibitory and bactericidal activity of antipseudomonal antibiotics against Pseudomonas aeruginosa isolates from cystic fibrosis patients

Using a broth microtiter dilution method, the minimum inhibitory (MIC) and minimum bactericidal concentrations (MBC) of so-called antipseudomonal antibiotics were determined against 79 Pseudomonas aeruginosa isolates from cystic fibrosis patients. On the basis of the MIC values and using DIN breakpoints, the percentual susceptibilities led to the following rank order: imipenem (91%), ciprofloxacin (90%), tobramycin (87%), amikacin (87%), ceftazidime (82%), cefsulodin (76%), piperacillin (71%), azlocillin (62%), followed by gentamicin, ceftriaxone, mezlocillin, netilmicin, and cefotaxime (less than 50%). However, evaluating MBC values according to DIN breakpoints, only ciprofloxacin (82%), tobramycin (77%), amikacin (58%), and imipenem (57%) showed a pronounced antipseudomonal effectiveness. The data indicate that MBC determinations are necessary to evaluate the antibiotic susceptibility of P. aeruginosa rather than MIC determinations, at least in patients with impaired defence mechanisms which require bactericidal therapy.     

Serum bactericidal and inhibitory titres in the management of melioidosis

A retrospective evaluation of the relationship between serum bactericidal and inhibitory titres and treatment outcome in 195 adult Thai patients with severe melioidosis was conducted. Drug regimens included ceftazidime (52% of patients), co-amoxiclav (24%), imipenem (11%) or the conventional four-drug combination (11%). Pre- and 1 h post-dose serum samples were collected after 48-72 h of therapy, and serum inhibitory and bactericidal titrations determined. Median post-dose titres were: bactericidal 1:8 (range 0-1:128) and inhibitory 1:16 (range 0-1:128). Overall mortality was 26% and outcome was not influenced by either inhibitory or bactericidal titres. Pre-dose titres correlated with renal function; renal function was the most important predictor of mortality. Determination of serum inhibitory or bactericidal titres is unhelpful in the management of severe melioidosis.     

Comparison of fractional inhibitory concentration index with response surface modeling for characterization of in vitro interaction of antifungals against itraconazole-susceptible and -resistant Aspergillus fumigatus isolates

Although the fractional inhibitory concentration (FIC) index is most frequently used to define or to describe drug interactions, it has some important disadvantages when used for drugs against filamentous fungi. This includes observer bias in the determination of the MIC and no agreement on the endpoints (MIC-0, MIC-1, or MIC-2 [> or = 95, > or = 75, and > or = 50% growth inhibition, respectively]) when studying drug combinations. Furthermore, statistical analysis and comparisons are troublesome. The use of a spectrophotometric method to determine the effect of drug combinations yields quantitative data and permits the use of model fits to the whole response surface. We applied the response surface model described by Greco et al. (W. R. Greco, G. Bravo, and J. C. Parsons, Pharmacol. Rev. 47:331-385, 1995) to determine the interaction coefficient alpha (ICalpha) using a program developed for that purpose and compared the results with FIC indices. The susceptibilities of amphotericin B (AM), itraconazole (IT), and terbinafine (TB) were tested either alone or in combination against 10 IT-susceptible (IT-S) and 5 IT-resistant (IT-R) clinical strains of Aspergillus fumigatus using a modified checkerboard microdilution method that employs the dye MTT [3-(4,5-dimethyl-2-thiazyl)2,5-diphenyl-2H-tetrazolium bromide]. Growth in each well was determined by a spectrophotometer. FIC indices were determined and ICalpha values were estimated for each organism strain combination, and the latter included error estimates. Depending on the MIC endpoint used, the FIC index ranged from 1.016 to 2.077 for AM-IT, from 0.544 to 1.767 for AM-TB, and from 0.656 to 0.740 for IT-TB for the IT-S strains. For the IT-R strains the FIC index ranged from 0.308 to 1.767 for AM-IT, from 0.512 to 1.646 for AM-TB, and from 0.403 to 0.497 for IT-TB. The results indicate that the degree of interaction is not only determined by the agents themselves but also by the choice of the endpoint. Estimates of the ICalpha values showed more consistent results. Although the absolute FIC indices were difficult to interpret, there was a good correlation with the results obtained using the ICalpha values. The combination of AM with either IT or TB was antagonistic in vitro, whereas the combination of IT and TB was synergistic in vitro for both IT-S and IT-R strains. The use of response surface modeling to determine the interaction of drugs against filamentous fungi is promising, and more consistent results are obtained by this method than by using FIC indices.     

In vitro interaction of flucytosine combined with amphotericin B or fluconazole against thirty-five yeast isolates determined by both the fractional inhibitory concentration index and the response surface approach

Combination therapy could be of benefit for the treatment of invasive yeast infections. However, in vitro interaction studies are relatively scarce and the interpretation of the fractional inhibitory concentration (FIC) index can be contradictory due to various definitions used; not all information on the interaction study is used in the index, and different MIC end points exist for different classes of drugs. Fitting an interaction model to the whole response surface and estimation of an interaction coefficient alpha (IC(alpha)) would overcome these objections and has the additional advantage that confidence intervals of the interaction are obtained. The efficacy of flucytosine (5FC) in combination with amphotericin B (AB) and fluconazole (FCZ) was studied against 35 yeast isolates in triplicate (Candida albicans [n = 9], Candida glabrata [n = 9], Candida krusei [n = 9], and Cryptococcus neoformans [n = 8]) using a broth microdilution checkerboard method and measuring growth after 48 h by a spectrophotometer. The FIC index and IC(alpha) were determined, the latter by estimation from the response surface approach described by Greco et al. (W. R. Greco, G. Bravo, and J. C. Parsons, Pharmacol. Rev. 47:331-385, 1995) by using a computer program developed for that purpose. For the 5FC-FCZ combination, the interactions determined by the IC(alpha) generally were in concordance with the interactions determined by the FIC index, but large discrepancies were found between both methods for the 5FC-AB combination. These could mainly be explained by shortcomings in the FIC approach. The in vitro interaction of 5FC-AB demonstrated variable results depending on the tested Candida isolate. In general, the 5FC-FCZ combination was antagonistic against Candida species, but for some Candida isolates synergism was found. For C. neoformans the interaction for both combinations was highly dependent on the tested isolate and the method used. Response surface approach is an alternative method for determining the interaction between antifungal agents. By using this approach, some of the problems encountered with the FIC were overcome.     

Minimum inhibitory and minimal lethal concentration against Chlamydia trachomatis dependent on the time of addition and the duration of the presence of antibiotics.

The purpose of this study was to investigate the properties of several antimicrobial agents found to be effective against Chlamydia trachomatis and to verify the eradication therapy schedule. The in vitro activities of two quinolones (sparfloxacin, ofloxacin), of three macrolides (azithromycin, erythromycin, clarithromycin) and of a tetracycline (doxycycline) against C. trachomatis were evaluated by several methods for the determination of the minimum inhibitory concentration (MIC) and minimal lethal concentration (MLC). MLC of azithromycin was only 2 times higher than that of MIC. On the other hand, MLCs of other antibiotics were 4-16 times higher than their respective MICs. When all antimicrobial agents were added to the infected culture at different times, we found that the quinolones even at a concentration of 64 microg/ml could not inhibit the formation of inclusion if they were added after 20 h from the start of infection. The corresponding period for macrolides and doxycycline was 24 h. When the antibiotics were removed at 8 h after the start of the infection, all antibiotics except azithromycin and clarithromycin were needed at a concentration much higher than their MLCs to inhibit the formation of inclusion. We consider macrolides, especially azithromycin, to be an excellent anti-C. trachomatis drug because of its lower MICs and MLCs values which were also closer together.     

Effect of beta-lactam antibiotics on migration and bactericidal activity of human phagocytes.

The effect of beta-lactam antibiotics upon some functions of human phagocytes was examined. Penicillins (carbenicillin and piperacillin), thienamycin, and cephalosporins (cefotetan, ceftazidime, and moxalactam) had no effect on either the random or directional migration or on the bactericidal activity of neutrophils and monocytes against Staphylococcus aureus. On the contrary, cefoperazone at therapeutic levels was shown to inhibit neutrophil chemotaxis.     

单过硫酸氢钾复合盐杀菌效果与影响因素研究

摘要 目的 〖HT5"SS〗研究单过硫酸氢钾复合盐消毒剂杀菌效果与影响因素。方法 采用悬液定量杀菌实验方法,对该单过硫酸氢钾复合盐消毒剂的杀菌效果与影响杀菌效果的因素进行观察。结果 用含单过硫酸氢钾复合盐1 000 mg/L的消毒液作用60 min,对悬液内枯草芽孢杆菌黑色变种芽孢杀灭对数值>5.00。用含100 mg/L单过硫酸氢钾复合盐消毒液作用20 min,对悬液内金黄色葡萄菌和大肠杆菌杀灭对数值均>5.00;作用7.5 min,对悬液内铜绿假单胞菌的杀灭对数值>5.00;对白色念珠菌作用20 min杀灭对数值>4.00需要150 mg/L。在10～30℃范围内,温度升高可增强单过硫酸氢钾复合盐杀菌效果;消毒液 pH值超过5.0使该消毒剂杀菌效果下降;在菌悬液内含25%以上的小牛血清可降低该消毒剂杀菌效果。结论 该单过硫酸氢钾复合盐消毒剂具有广谱、高效杀菌效果,作用温度、pH值和有机物对单过硫酸氢钾复合盐杀菌效果均有影响。     

Delayed bactericidal activity of beta-lactam antibiotics against Listeria monocytogenes: antagonism of chloramphenicol and rifampin

Penicillins are considered to be the drugs of choice for the treatment of listeric meningitis, and relapse of infection is rare when treatment is given in appropriate doses for at least 14 days. Despite this, in vitro studies by others have shown that penicillins are bacteriostatic against Listeria spp. We have shown that thienamycin, penicillin G, and ampicillin are the most active beta-lactam antibiotics against Listeria spp. Of 10 strains tested, 9 were killed by less than or equal to 8 micrograms of beta-lactam antibiotics (greater than or equal to 99.9% killing) when subcultures were performed after 48, rather than 24, h of incubation. In contrast, chloramphenicol, erythromycin, doxycycline, and rifampin were bacteriostatic after 48 h of incubation. In time-kill curves, these last drugs antagonized the bactericidal action of penicillins. In view of the inefficiency of opsonization in the cerebrospinal fluid, these antagonistic combinations should probably be avoided in documented or suspected listeric meningitis.     

Evaluation of the bactericidal activity of beta-lactam antibiotics on slowly growing bacteria cultured in the chemostat.

The bactericidal activity of 23 beta-lactam antibiotics was compared in slowly growing bacteria cultured in a chemostat. In an attempt to mimic possible in vivo conditions, slowly growing cultures were produced by limitation of iron, glucose, phosphate, or magnesium. Only select antibiotics remained effectively bactericidal against slowly growing cells. For these compounds, the rate of antibiotic-induced loss of viability was a constant when killing was expressed per generation (in contrast to absolute time) in that slowly growing bacteria were killed proportionately more slowly. Individual antibiotics differed greatly, however, in their specific bactericidal activities against slowly growing cells, i.e., in the absolute degree of killing elicited during exposure of the bacteria to MIC equivalents of the drugs. Specific bactericidal activities varied not only with drug structure but also with the bacterial strains and, to a lesser extent, with the nature of the growth-limiting nutrient. In slowly growing cultures exposure to the low drug concentrations studied here (near MIC) caused killing without detectable lysis. Antibiotics with high specific bactericidal activities were capable of rapidly killing cultures of slowly growing pathogens despite extremely long generation times approaching those reported for in vivo growth rates.     

Alpha 1-antitrypsin: the effect of anticoagulants on the trypsin inhibitory capacity, concentration and phenotype

The effects of anticoagulants on the determination of both trypsin inhibitory capacity and the concentration of alpha 1-antitrypsin measured by radial immunodiffusion, and on the alpha 1-antitrypsin phenotype were investigated. These results were compared with those obtained for serum. The following anticoagulants were investigated: sodium citrate; sodium oxalate; buffered citrate; potassium oxalate/sodium fluoride; sodium heparin; and potassium EDTA. It was found that plasmas from all of the anticoagulants, except sodium heparin, resulted in apparently significant decreases of both trypsin inhibitory capacity and concentration of alpha 1-antitrypsin measured by radial immunodiffusion, relative to serum. These decreases were not simply due to dilution by anticoagulants. Using both acid starch gel electrophoresis followed by immunofixation and isoelectric focusing in agarose, no interference was found in the phenotype determination. It is concluded that serum should be used to measure the trypsin inhibitory capacity or the concentration of alpha 1-antitrypsin by radial immunodiffusion, although plasma is also suitable provided that sodium heparin is used as the anticoagulant. The alpha 1-antitrypsin phenotype can be determined with either serum or any of the plasma. None of the anticoagulants employed in this study was present in excess.     

Bacteriostatic and bactericidal activities of beta-lactam antibiotics enhanced by the addition of low concentrations of gentamicin

The combined effects of low concentrations of gentamicin on certain beta-lactam antibiotics were studied by the agar plate method. the combination of gentamicin with cephalothin produced synergism against 17 of 26 strains of Escherichia coli and 19 of 27 strains of Klebsiella sp. if assessed at the bacteriostatic (minimal inhibitory concentration) level. Synergy against many more strains was apparent when bactericidal concentrations were used. Synergy against most of these strains was observed if bactericidal concentrations with brief exposure times (3 h) to the antibiotics were used for measurement. Additive effects were observed in almost all of the remaining strains. The combination of gentamicin and carbenicillin were synergistic against most strains of Pseudomonas aeruginosa when any bacteriostatic or bactericidal measurement was used as the criterion.