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Minimum inhibitory concentration determinations for various antimicrobial agents against 1570 bacterial isolates from turkey poults.

Minimum inhibitory concentrations (MICs) were determined for 1570 bacteria from eight geographic locations (1204 Escherichia coli, 231 other enteric gram-negative bacilli [including Citrobacter spp., Enterobacter spp., Klebsiella spp., Proteus spp., and Salmonella spp.], 31 Pseudomonas spp., 18 coagulase-positive staphylococci, 26 coagulase-negative staphylococci, and 55 streptococci and enterococci) by the National Committee for Clinical Laboratory Standards broth microdilution procedure. Antimicrobial agents tested included ampicillin, ceftiofur, enrofloxacin, erythromycin, florfenicol, gentamicin, neomycin, spectinomycin, sulfamethazine, tetracycline, and trimethoprim/sulfadiazine. Against the E. coli strains tested, ceftiofur, enrofloxacin, gentamicin, and trimethoprim/sulfadiazine were the most active compounds with MIC at which 50% of the strains are at or below (MIC50) = 0.5, < or = 0.03, 0.5, and 0.13 microg/ml, respectively, and MIC at which 90% of the strains are at or below (MIC90) = 1.0, 0.13, 32.0, and 2.0 microg/ml, respectively. Ampicillin, florfenicol, neomycin, and spectinomycin were the next most active compounds against the E. coli strains, with MIC50 = 4.0, 4.0, 16.0, and 16.0 microg/ml, respectively. MIC90 values for these compounds against E. coli strains were > 32.0, 8.0, 512.0, and > 128.0 microg/ml, respectively. The remaining compounds exhibited limited, strain-dependent activity against the E. coli strains tested. As with the E. coli, enrofloxacin, ceftiofur, and trimethoprim/sulfadiazine were also the most active compounds against the 231 other enteric organisms tested, with MIC50 < or = 1.0 microg/ml for all of these genera. The remaining compounds exhibited limited activity against these genera. Against the gram-positive cocci tested, ampicillin, enrofloxacin, ceftiofur, and trimethoprim/sulfadiazine were most active, whereas the remaining compounds exhibited strain-dependent activity. When MIC data for E. coli were summarized separately, differences were observed between the geographic locations for the various antimicrobial agents. In conclusion, ceftiofur, enrofloxacin, and trimethoprim/sulfadiazine were the most active of the compounds tested against all of the bacterial strains.

Animals↗

Effects of sub-minimum inhibitory concentrations of gatifloxacin on the inhibition of Staphylococcus aureus and Escherichia coli adherence.

The aim of this study was to investigate the capacity of subinhibitory concentrations of the newly developed fluoroquinolone antibiotic gatifloxacin (CAS 160738-57-8) to interfere with the mechanism of bacterial adhesion. Human buccal epithelial cells were incubated with different strains of both Staphylococcus aureus and Escherichia coli, grown in the presence of subinhibitory gatifloxacin concentrations varying from 1/2 MIC (minimum inhibitory concentration) to 1/128 MIC. A significant decrease was observed in the adherence of S. aureus and E. coli to buccal cells at drug MICs of 1/2 to 1/32 and 1/2 to 1/64, respectively. A large number of filamentous forms of different lengths and shapes were observed in the case of E. coli, whereas there was an abnormal increase in the diameter of the cells in the case of S. aureus. The interpolation of these pharmacodynamic findings with the pharmacokinetic curve indicate that the effect of subinhibitory concentrations of gatifloxacin can prolong its antimicrobial effects against S. aureus and E. coli for as long as 30 h and 37 h, respectively, after the MIC values have been reached.

Anti-Infective Agents↗

Effects of sub-minimum inhibitory concentration antibiotic levels and temperature on growth kinetics and outer membrane protein expression in Mannheimia haemolytica and Haemophilus somnus.

The objective of this study was to determine the effects of sub-minimum inhibitory concentrations (sub-MICs) of 2 veterinary antibiotic preparations, chlortetracycline (CTC) and chlortetracycline-sulfamethazine (CTC + SMZ), on growth kinetics and outer membrane protein expression in Mannheimia haemolytica and Haemophilus somnus at normal and febrile body temperatures. Sub-minimum inhibitory concentrations of both antibiotics reduced the growth rates of M. haemolytica and H. somnus. Growth of both species was not inhibited when grown at 41 degrees C compared to 37 degrees C. There was no detectable consistent effect of antibiotic or temperature on outer membrane protein expression for either species. Our study indicates that sub-MIC levels of CTC and CTC + SMZ markedly impair growth of clinical M. haemolytica and H. somnus isolates, potentially allowing more effective host clearance during infection.

Anti-Bacterial Agents↗

A new method for determining the minimum inhibitory concentration of essential oils.

A new microdilution method has been developed for determining the minimum inhibitory concentration (MIC) of oil-based compounds. The redox dye resazurin was used to determine the MIC of a sample of the essential oil of Melaleuca alternifolia (tea tree) for a range of Gram-positive and -negative bacteria. Use of 0.15% (w/v) agar as a stabilizer overcame the problem of adequate contact between the oil and the test bacteria and obviated the need to employ a chemical emulsifier. A rapid version of the assay was also developed for use as a screening method. A comparison of visual and photometric reading of the microtitre plates showed that results could be assessed without instrumentation; moreover, if the rapid assay format was used, rigorous asepsis was not necessary. Accuracy of the resazurin method was confirmed by plate counting from microwells and MIC values were compared with results obtained using an agar dilution assay. The MIC results obtained by the resazurin method were slightly lower than those obtained by agar dilution.

Gram-Negative Bacteria↗

A study of the minimum inhibitory concentration and mode of action of oregano essential oil, thymol and carvacrol.

AIMS: The minimum inhibitory concentration (MIC) of oregano essential oil (OEO) and two of its principle components, i.e. thymol and carvacrol, against Pseudomonas aeruginosa and Staphylococcus aureus was assessed by using an innovative technique. The mechanism of action of the above substances was also investigated. METHODS AND RESULTS: The applied technique uses 100-well microtitre plate and collects turbidimetric growth data. To produce the inhibition profiles, a wide range of concentrations were tested for each of the three compounds, as well as for carvacrol-thymol mixtures. Following a specific mathematical analysis of the observed inhibition profiles from all compounds, it was suggested that mixtures of carvacrol and thymol gave an additive effect and that the overall inhibition by OEO can be attributed mainly to the additive antimicrobial action of these two compounds. Addition of low amounts of each additive: (a) increased permeability of cells to the nuclear stain EB, (b) dissipated pH gradients as indicated by the CFDA-SE fluorescent probe irrespective of glucose availability and (c) caused leakage of inorganic ions. CONCLUSION: Mixing carvacrol and thymol at proper amounts may exert the total inhibition that is evident by oregano essential oil. Such inhibition is due to damage in membrane integrity, which further affects pH homeostasis and equilibrium of inorganic ions. SIGNIFICANCE AND IMPACT OF THE STUDY: The knowledge of extent and mode of inhibition of specific compounds, which are present in plant extracts, may contribute to the successful application of such natural preservatives in foods, since certain combinations of carvacrol-thymol provide as high inhibition as oregano essential oil with a smaller flavour impact.

Cell Membrane↗

Comparison of drug sensitivity testing with microdilution quantitative minimum inhibitory concentration and the Autobac I system.

Antibiotic susceptibility studies were compared by the quantitative minimum inhibitory concentration and Autobac I. Four hundred common isolates from clinical materials including both gram-negative and -positive organisms were evaluated. Results demonstrate that the Autobac I method can provide reliable semiquantitative estimation of bacterial susceptibility that correlates well with an acceptable standard procedure. The specific advantages of the Autobac I system in addition to the reliability demonstrated are speed, objectivity of interpretation, and ability to provide a printed result. These advantages can result in increased speed of reporting, which may reduce hospital stay and patient morbidity as well as providing needed information more rapidly in critically ill patients with bacterial infections.

Ampicillin↗

Minimum inhibitory concentrations of 20 antimicrobial agents against Staphylococcus aureus isolated from bovine intramammary infections in Japan.

Minimum inhibitory concentrations (MICs) of 20 antimicrobial agents were determined against 51 isolates of Staphylococcus aureus from bovine intramammary infections. Fourteen (27.4%) isolates were resistant to benzylpenicillin, but none of the isolates was resistant to cloxacillin, nafcillin, or cephems. Among aminoglycosides, gentamicin was the most active, with an MIC50 of 0.2 microg/ml, followed by kanamycin, with an MIC50 of 0.78 microg/ml. Five isolates (9.8%) were resistant to dihydrostreptomycin, three isolates (5.9%) to kanamycin and two isolates (3.9%) to gentamicin. Resistance to erythromycin was observed in two isolates (3.9%). Tylosin was less active than erythromycin, with MIC50s of 1.56 microg/ml versus 0.39 microg/ml, but none of the isolates was resistant to this antibiotic. Oxytetracycline MICs were situated in the range of 0.39-1.56 microg/ml for 48 susceptible isolates. Although 19 (37.3%) isolates were resistant to one or more antimicrobial agents, a single resistance pattern was most frequent: benzylpenicillin (12 isolates), dihydrostreptomycin (two isolates) and kanamycin (one isolate). There were no isolates resistant to antimicrobial agents such as methicillin, lincomycin, clindamycin, chloramphenicol, florfenicol and virginiamycin, which have not been approved for use in cattle husbandry in Japan.

Animals↗

Increased minimum inhibitory concentrations with anaerobiasis for tobramycin, gentamicin, and amikacin, compared to latamoxef, piperacillin, chloramphenicol, and clindamycin.

Minimum inhibitory concentrations (MICs) were determined under both routine aerobic and anaerobic conditions for a total of 93 organisms representing nine genera. MICs for the aminoglycosides amikacin, gentamicin, and tobramycin were significantly increased under anaerobic conditions. Tobramycin was most sensitive to the loss of antimicrobial activity with anaerobiasis. MICs for staphylococci were increased by a higher factor than were MICs for gram-negative rods, but even within the latter group increases in MICs for Proteus species were greater than for Salmonella, Klebsiella and Escherichia coli. No change of anaerobic versus aerobic activity was seen for latamoxef, piperacillin, chloramphenicol, or clindamycin.

Amikacin↗

Comparison of minimum inhibitory concentrations to methicillin in heterogeneous and homogeneous methicillin resistant Staphylococcus aureus.

OBJECTIVE: To determine the minimum inhibitory concentration (MIC) to methicillin and compare it with the type of methicillin resistance in methicillin resistant Staphylococcus aureus (MRSA) isolated from clinical samples, and to evaluate the usefulness of beta-lactamase resistant penicillins such as cloxacillin for treatment of minor MRSA infections. DESIGN: Sixty-five strains of MRSA isolated from clinical samples were studied. Homogeneous and heterogeneous resistance to methicillin of these strains was determined by an efficiency of plating technique. The MICs to methicillin of all homogeneously resistant strains and an equal number of heterogeneously resistant strains were determined by the 'E' test. SETTING: General Hospital, Colombo; De Soysa Maternity Hospital for Women; Lady Ridgeway Hospital for Children. RESULTS: 171 strains of S aureus were isolated from specimens processed in the laboratory of the Department of Microbiology, Faculty of Medicine Colombo, during a 14-month period. Seventy (40.9%) were methicillin resistant. Fifty-three MRSA strains (81.5%) were heterogeneously resistant to methicillin. Only 12 of 65 strains were homogeneously resistant. Nine of 12 of the homogeneously resistant strains were from surgical wards. MICs for methicillin in the two groups were compared. Six of 12 (50%) homogeneously resistant strains had very high levels of resistance to methicillin (MIC > 200 micrograms/ml). In contrast the MICs of all the heterogeneously resistant strains ranged from 16 micrograms/ml to 96 micrograms/ml. CONCLUSIONS: The majority of clinical isolates of MRSA from hospitals in Colombo, Sri Lanka were heterogeneously resistant to methicillin with lower MICs for methicillin than the homogeneously resistant strains. Thus, beta-lactamase resistance penicillins such as cloxacillin may have a place in the treatment of minor MRSA infections. Surveillance studies to determine if there is an increase of homogeneously methicillin resistant MRSA are required.

Adult↗

Effect of protein on ramoplanin broth microdilution minimum inhibitory concentrations.

Ramoplanin is a glycolipodepsipeptide antibiotic active against Gram-positive bacteria. We observed that microdilution minimum inhibitory concentrations (MICs) were higher than those obtained in glass tubes or by agar dilution. Initial studies showed that these differences disappeared when 30% bovine serum was added to the broth. Further studies showed that addition of 0.01% bovine serum albumin (BSA) to the broth lowered the microdilution MICs for staphylococci, streptococci, and enterococci by four- to 32-fold. This phenomenon occurred in several commonly used growth media and in different types of commercially available microtiter trays. Precoating of the microtiter wells with a dilute solution of BSA (0.02%) had the same effect. It seems likely that ramoplanin adsorbs to plastic surfaces and is lost from solution, and that protein masks the sites of adsorption. Ramoplanin MICs may be reliably determined by broth microdilution if a small amount of protein is added to the diluent.

Adsorption↗

Minimum inhibitory concentrations for selected antimicrobial agents against Fusobacterium necrophorum isolated from hepatic abscesses in cattle and sheep.

Minimum inhibitory concentrations for 35 antimicrobial agents against 100 Fusobacterium necrophorum isolates from hepatic abscesses in sheep and cattle were determined. Twelve of the thirteen beta-lactam antibiotics tested inhibited growth of 100% of strains tested. Of the remaining antimicrobial agents, extensive susceptibility was found for: spiramycin, josamycin, lincomycin, tylosin, oxytetracycline, chlortetracycline, rufloxacin, metronidazole, cotrimoxazole, sulfadimethoxine, virginiamycin and fosfomycin.

Animals↗

Beta-lactamase testing of staphylococci in a commercial broth microdilution minimum inhibitory concentration system.

Beta-Lactamase testing of clinical isolates of staphylococci may be performed in commercial broth microdilution minimum inhibitory concentration plates, using a chromogenic cephalosporin reagent directly in a well containing a noninhibitory concentration of a semisynthetic penicillin which serves as an inducer. A total of 115 staphylococcal isolates tested in 0.25 to 0.50 microgram of methicillin per ml in Mueller-Hinton broth showed 100% correlation with beta-lactamase tests performed on Mueller-Hinton agar, using 1-microgram oxacillin disk as an inducer.

Cephalosporins↗

[Antibiotic sensitivity of forty-four strains of group EF4 bacteria: study of minimum inhibitory concentrations using the agar dilution method].

EF4 bacteria are found in animal saliva and may contaminate bite wounds. Minimum inhibitory concentrations of 36 antimicrobials against 44 EF4 strains were determined using dilution in Mueller-Hinton agar. EF4 bacteria were susceptible to aminopenicillins, carboxypenicillins, ureidopenicillins, third-generation cephalosporins, fluoroquinolones, rifampicin, and trimethoprime-sulfamethoxazole. Susceptibility was intermediate for penicillin G, low for macrolides and variable for aminoglycosides. EF4 bacteria were resistant to lincomycin and trimethoprime. Routine prophylactic treatment of bite-induced Pasteurella infections using an aminopenicillin or a cycline also protects against EF4 infection of the wound.

4-Quinolones↗

Guidelines and recommendations for antimicrobial minimum inhibitory concentration (MIC) testing against veterinary mycoplasma species. International Research Programme on Comparative Mycoplasmology.

The absence of standardised procedures for minimum inhibitory concentration (MIC) testing of antimicrobial agents against veterinary mycoplasma and ureaplasma species (Mollicutes) has made it difficult to compare results originating from different laboratories. This report, prepared on behalf of the International Research Programme on Comparative Mycoplasmology (IRPCM), offers guidelines and recommendations for veterinary MIC testing of these organisms in an effort to rectify this problem. The subjects discussed include suitable media for broth and agar MIC assays, storage and preparation of antimicrobial agents, standardisation of mycoplasma inocula for MIC tests, validation of equipment, incubation conditions, and determination of MIC end points. A standard medium for all veterinary mycoplasma MIC tests cannot currently be recommended, owing to the diversity of nutritional requirements of different mycoplasma species. Instead mycoplasma broths or agars giving optimal growth of specific mycoplasmas or ureaplasmas are recommended, as suboptimal growth may lead to falsely low MIC results. The importance of using standardised mycoplasma inocula, for assays using either solid or liquid media is stressed. The growth phase may be less important as lag phase and logarithmic phase cultures of Mycoplasma gallisepticum, M. synoviae, M. bovis and M. hyopneumoniae have given very similar results in liquid MIC assays. The liquid method of Tanner and Wu and the agar method described by Hannan et al. are compared and described in detail. Methods for calculating MIC50s and MIC90s are described and the interpretation of results discussed. Methods for assessing mycoplasmacidal (MMC) activity of antimicrobial agents are also described. Adoption of these guidelines should lead to more consistent MIC results being obtained between laboratories.

Animals↗

Antimicrobial resistance in gram-positive bacteria: the myth of the MIC. Minimum inhibitory concentration.

Although much progress has occurred in the fight to treat infection, resistant bacteria increasingly are becoming problematic. The minimum inhibitory concentration is easily determined, yet its definition, clinical application, and relevance are confusing. In addition, errors can occur during in vitro susceptibility testing. The mechanism of resistance for different organisms is discussed as are the pitfalls of susceptibility testing and the prescription of antimicrobial agents.

Anti-Bacterial Agents↗

[Evaluation of a newly developed broth microdilution test method to determine minimum inhibitory concentrations (MICs) of antimicrobial agents for mycobacteria].

We developed a new broth microdilution antimycobacterial susceptibility test for determination of minimum inhibitory concentration (MICs) using an air-dried microplate containing serially diluted antimicrobial agents and the modified Middlebrook 7H9 broth. The eight agents included were streptomycin (SM), isoniazid (INH), rifampicin (RFP), ethambutol (EB), kanamycin (KM), levofloxacin (LVFX), sparfloxacin (SPFX) and clarithromycin (CAM). Serial dilutions of the agents (128 micrograms/ml to 0.125 micrograms/ml) were prepared in microplates, and were reconstituted by inoculation of 0.2ml of cell suspensions (approximately 3 x 10(5) cells/ml). The test plates were incubated at 36 degrees C in 5% CO2, and the growth endpoints were read visually after 5-day, 7-day and 10-day incubations. Four ATCC reference strains, Mycobacterium tuberculosis, M. avium, M. kansasii and M. intracellulare, were repeatedly tested at three sites. Of 480 determination against eight agents, 455 (94.8%), 470 (97.9%) and 455 (94.8%) of the MICs read after 5-day, 7-day and 10-day incubations fell within 3log2 dilutions, respectively. The MICs gradually elevated during the incubation, however those of 7-day incubation were highly precise and easily determined. A total of 160 clinical isolates of M. tuberculosis and 114 of nontuberculous mycobacteria were tested against eight agents. As for the primary drugs (SM, INH, RFP and EB), most isolates of M. tuberculosis were highly susceptible with MIC90, < or = micrograms/ml. Both LVFX and SPFX were also active. The MICs against nontuberculous mycobacteria distributed in a wide range, and the activities of RFP, LVFX, SPFX and CAM were more potent. These results demonstrate this newly developed test method to be a practical, rapid, quantitative and nonradiometric alternative for the determination of MICs in clinical mycobacteriology laboratories.

Antitubercular Agents↗

Direct application of Etest to gram-positive cocci from blood cultures: quick and reliable minimum inhibitory concentration data.

The Etest was applied directly to 100 Gram-positive bacterial strains from blood cultures to measure their minimum inhibitory concentration (MIC). Results showed 100% concordance of MIC data between a direct Etest method and the standard Etest method for Streptococcus pneumoniae, beta-hemolytic streptococci, and viridans group streptococci. In addition, direct Etest for Staphylococcus aureus, coagulase negative staphylococci, Enterococcus faecium, and Enterococcus faecalis showed 83 to 100% correlation with standard Etests. These data indicate that the Etest is useful to obtain MIC data on Gram-positive cocci (especially streptococci) directly from positive blood cultures. The advantages of a direct Etest are two fold: MIC results can be obtained 24 hours earlier than standard methods and a more representative population of the bacterial isolate is tested.

Anti-Bacterial Agents↗

Rapid determination of minimum inhibitory concentrations of antimicrobial agents by the Autobac method: a collaborative study.

Four laboratories collaborated in an evaluation of the Autobac minimum inhibitory concentration (MIC) test system. The MICs of ranges of MICs determined in this system were compared with the MICs obtained with a microtube modification of the International Collaborative Study broth dilution technique. A total of 1,260 strains, mostly recent clinical isolates and including multiresistant strains, were tested by the four laboratories against 10 antibiotics; 9,360 separate MIC determinations were made. There was an overall agreement of approximately 95% between the two methods. Levels of agreement below 80% were obtained with only 4 of the 104 antibiotic-species pairs. In only one of the four major organism groups (staphylococci and penicillin G) was agreement less than 85%. There was a symmetrical distribution of MIC differences between the two methods. Tests with 56 selected strains were performed in each of four laboratories in an inter- and intra-laboratory reproducibility study. Both methods showed a standard deviation (both inter- and intra-laboratory) of one-half of a twofold dilution step. The Autobac method was actually less variable than the reference method and had equivalent reproducibility. This was particularly true when the Autobac system was operated so that the results generated permitted calculations of MICs via regression analysis.

Anti-Bacterial Agents↗