Evaluation of Vitek system for susceptibility testing of Enterococcus faecium isolates

Although imipenem is not a first-line drug for treating enterococcal infection, it could well become a useful drug for treating mixed infections that include enterococci. However, there is no NCCLS guideline for susceptibility testing of imipenem versus enterococci. Moreover, there are no statements to indicate that in vitro susceptibility results for other antimicrobial agents can be used to predict the in vitro activity of imipenem against enterococci. In this study, 52 Enterococcus faecium isolates were collected from patients hospitalized at Kangnam St. Mary's Hospital between March 2002 and December 2002. The sources of the isolates were mainly urine specimens and wounds. For ampicillin, the "major" and "very major" error rates observed with the Vitek system were 0% and 2.0%, respectively. For penicillin, the major and very major error rates observed with the Vitek system were both 0%. For imipenem, the major and very major error rates observed with the Vitek system were 0% and 36.5%, respectively. The MICs of ampicillin and penicillin obtained using the Vitek system were reliable, but that of imipenem was unreliable. In the 52 E. faecium isolates, the in vitro activity of penicillin and ampicillin versus enterococci accurately predicted that of imipenem. Therefore, the MIC of imipenem obtained with the Vitek system must be retested by the agar dilution method, when it disagrees with those of penicillin and ampicillin.     

Comparison of the Cobas-Bact five-hour susceptibility testing system with the NCCLS agar diffusion and dilution methods

The results of susceptibility tests performed by the Cobas-Bact system were compared with those of the NCCLS agar diffusion (Kirby-Bauer) and NCCLS agar dilution methods. A total of 998 clinical isolates were tested against 10 to 18 antimicrobial agents. Essential agreement (comprising full agreement and minor discrepancies) varied from 90.5 % to 99.2 % on comparison of Cobas-Bact with Kirby-Bauer results, depending on the bacterial group (mean for all 998 strains tested 95.7 %). These figures ranged from 91 % to 99.2 % (mean 96.3 %) for the Cobas-Bact/MIC comparison and from 95.2 % to 99.7 % (mean 98.7 %) for the Kirby-Bauer/MIC comparison. The best results were found forEnterobacteriaceae andStaphylococcus aureus, whereas for enterococci and coagulase-negative staphylococci there was a lower rate of essential agreement in all three comparisons. In the case ofPseudomonas aeruginosa there was a good rate of essential agreement but many minor discrepancies, resulting in a disappointing rate of full agreement of between 67.5 % and 78.9 % in the three comparisons. The Cobas-Bact system would appear to provide satisfactory susceptibility test results in most cases, however there are still some major problems in the system which should be resolved.     

Comparison of Etest, Vitek and agar dilution for susceptibility testing of colistin

In total, 172 isolates of Enterobacteriaceae, Acinetobacter spp., Pseudomonas aeruginosa and Stenotrophomonas maltophilia were tested for susceptibility to colistin by agar dilution, Etest and the Vitek 2 system. Isolates with a colistin MIC < or =2 mg/L were considered to be susceptible. Fifty-four (31%) Gram-negative isolates were resistant to colistin. Categorical agreement between agar dilution and Etest was 87%, and between agar dilution and Vitek 2 was 82%. Based on the data obtained, the Vitek 2 system was unreliable for detecting colistin resistance, and results obtained by Etest may require confirmation by a standard MIC susceptibility testing method.     

Comparison of media for agar dilution susceptibility testing ofBordetella pertussis andBordetella parapertussis

Antimicrobial susceptibility testing of the fastidious speciesBordetella pertussis andBordetella parapertussis is not standardized. In an attempt to find the optimal medium for agar dilution testing, the activity of erythromycin againstBordetella pertussis andBordetella parapertussis (34 isolates each) was assessed using homologous broth/agar combinations of Bordet-Gengou, charcoal, Iso-Sensitest (Oxoid) and Mueller-Hinton media. Each medium was supplemented with 5 % and 20 % whole defibrinated horse blood. Mueller-Hinton medium supplemented with 5 % horse blood performed best overall.     

Use of cephalexin-aztreonam-arabinose agar for selective isolation of Enterococcus faecium.

Cephalexin-aztreonam-arabinose agar (CAA), a new selective agar, was examined in comparison with nalidixic acid-colistin agar for the differentiation of Enterococcus faecium from other enterococci and the ability to isolate the organism from feces. Two hundred sixteen enterococcus isolates and a variety of gram-positive and gram-negative control strains were inoculated onto both media. All control strains of E. faecium were easily differentiated from Enterococcus faecalis and Enterococcus durans on the basis of arabinose fermentation on CAA. Differentiation of E. faecium from other enterococci or Streptococcus bovis was not possible on nalidixic acid-colistin agar. Increased isolation of E. faecium was demonstrated on CAA when both media were compared for the isolation of the organism from feces. CAA has been shown to possess excellent differential and selective features allowing the simple and effective isolation of E. faecium from heavily contaminated sites.     

Comparison of microdilution and agar dilution procedures for testing antibiotic susceptibility of Neisseria gonorrhoeae

Studies were run in parallel to compare the broth microdilution method and the chocolate agar dilution method for testing antibiotic susceptibility of Neisseria gonorrhoeae. Six clinically relevant drugs were tested against 23 clinical isolates of N. gonorrhoeae, including several penicillinase-producing, as well as multiply resistant, strains. Results showed that the MIC obtained by the two methods were not significantly different. The microdilution method appears to be a more sensitive system for discriminating penicillinase activity. The microdilution system is a more expedient method for screening new antibacterial agents and is more readily adaptable to new automated equipment.     

Comparison of E-test with agar dilution for determining susceptibility ofStreptococcus pneumoniae to penicillin

Minimum inhibitory concentrations (MICs) of penicillin forStreptococcus pneumoniae were determined by the E-test and the agar dilution method. NinetyStreptococcus pneumoniae strains were tested, of which 16 were resistant, 33 intermediate, and 41 susceptible by agar dilution. By the E-test, 80 (88.9%) strains agreed with these determinations within one log₂ dilution step, and no strains disagreed by more than two dilution steps. Sixty-eight of the 70 strains with discrepant MICs read lower in the E-test, resulting in 15 strains being placed in different susceptibility categories when classified by this test. Exact MICs rather than classification groups should be used to determine appropriate antibiotic therapy, since small differences in MICs determined by different methods can lead to a significant degree of misclassification.     

Comparison of the Etest and a microbroth dilution system (Sceptor) to a reference agar dilution method for susceptibility testing of Bilophila wadsworthia

Objective: To compare the Etest and a microbroth dilution system (Sceptor) to a reference agar dilution method for susceptibility testing of Bilophila wadsworthia.
Method: The susceptibility of 15 clinical isolates of Bilophila wadsworthia was determined by the National Committee for Clinical Laboratory Standards (NCCLS) agar dilution method using triphenyltetrazolium chloride for endpoint determination. The results were compared with the results obtained by the E test and a commercial microbroth dilution system (Sceptor).
Results: Comparison of the MICs obtained by the reference method and the Etest revealed few discrepancies, with piperacillin and metronidazole being the only exceptions. The overall agreement was 70% within one dilution step. The discrepancies did not result in major interpretative errors. The overall essential agreement using susceptibility categories was 98% for the E test and 99% for the microdilution system.
Conclusions: Both methods may be considered as acceptable alternatives for testing individual isolates of B. wadsworthia.     

A PCR assay for identification of Enterococcus faecium.

Enterococcus faecium has recently emerged as a serious nosocomial pathogen. The prevalence and severity of enterococcal infections, the mortality rate from such infections, and the antibiotic resistance of enterococci are often species dependent. Since conventional biochemical methods fail to differentiate E. faecium from certain newly described enterococcal species, a PCR-based assay was developed for the rapid identification of E. faecium.     

Comparison of the agar dilution, tube dilution, and broth microdilution susceptibility tests for determination of teicoplanin MICs.

The purpose of this study was to evaluate the National Committee for Clinical Laboratory Standards agar dilution, tube dilution, and broth microdilution susceptibility tests for the measurement of teicoplanin MICs. The three standardized tests gave equivalent (within a twofold dilution) results with 98.8 to 99.0% of the 508 gram-positive clinical isolates tested, indicating that either method may be used for teicoplanin MIC determination.     

Comparison of antimicrobial susceptibility testing of Campylobacter spp. by the agar dilution and the agar disk diffusion methods

The correlation and the level of agreement between the standardized agar dilution and the agar disk diffusion methods for antimicrobial susceptibility testing of Campylobacter were investigated. A high-level agreement between the two methods was evident for aminoglycosides and fluoroquinolones, while a low-level agreement was observed for other antibiotics.     

Comparison of Etest and agar dilution method for antimicrobial susceptibility testing of Flavobacterium isolates.

The Etest was evaluated as a possible alternative to the standard agar dilution method for susceptibility testing of nine antimicrobial agents against Flavobacterium species. In studies of 100 clinical isolates, the agreement between the MICs (+/-1 log2 dilution) obtained by the two methods was acceptable for cefotaxime, ceftazidime, amikacin, minocycline, ofloxacin, and ciprofloxacin (> 90%). Conversely, the agreement between the results obtained for piperacillin was limited (84%). The overall agreement was 92.5%.     

Comparison of agar dilution, tube dilution, and broth microdilution susceptibility tests for determination of ramoplanin MICs.

Standard broth microdilution (with and without bovine serum albumin [BSA] supplementation), tube dilution, and agar dilution susceptibility tests were compared for determining ramoplanin MICs. With a data base of 246 clinical isolates of gram-positive bacteria from 33 U.S. sites, it was shown that (i) agar and tube dilution susceptibility tests gave essentially the same results (93.9% of the test results were within 1 doubling dilution of equivalence), (ii) broth microdilution susceptibility tests gave results up to 5 doubling dilutions higher than agar or tube assays, and (iii) this data skewing could be reversed by BSA supplementation (final concentration, 0.02%) of the broth microdilution test medium.     

Comparison of three media for agar dilution susceptibility testing ofBordetella pertussis using six antibiotics

Since antimicrobial susceptibility testing of the fastidious speciesBordetella pertussis is not standardized, the most suitable medium for agar dilution testing of this species has not yet been determined. In the present study, Mueller-Hinton, Bordet-Gengou, and Oxoid charcoal agars (each supplemented with 5% horse blood) were evaluated for agar dilution susceptibility testing ofBordetella pertussis against ampicillin, chloramphenicol, ciprofloxacin, doxycycline, erythromycin, and trimethoprim-sulfamethoxazole. Mueller-Hinton agar was the most suitable medium.     

Comparison of agar disk diffusion, microdilution broth, and agar dilution for testing antimicrobial susceptibility of coagulase-negative staphylococci.

A collection of 120 oxacillin-susceptible and 120 oxacillin-resistant coagulase-negative staphylococci (CNS) from six tertiary care hospital laboratories were tested by agar disk diffusion, three microdilution broth systems (Sensititre, Dynatech, and Alpkem), and the Vitek AutoMicrobic system for comparison with reference agar dilution results. The antimicrobial agents tested were oxacillin, cefazolin, cefotaxime, cefuroxime, cefamandole, fusidic acid, rifampin, and vancomycin. Incubation was at 30 or 35 degrees C for 24, 48, and 72 h. The broth media were supplemented with 2% NaCl for some antimicrobial agents, and the agar dilution method was used with and without the addition of 4% NaCl. The CNS were identified to species by the method of Kloos and Schleifer. The results showed a lack of concordance between two hospitals with respect to oxacillin susceptibility testing by agar dilution with no NaCl supplement. The reasons are not clear but may be related to variations in media. The 4% NaCl supplement or extended incubation to 48 h eliminated this difference. The cefazolin and cefotaxime susceptibility results in the agar disk diffusion test were unreliable if accepted at face value. Cefamandole testing correlated well with the reference method regardless of the method used, and salt supplementation is not recommended. Most of the oxacillin-resistant CNS were resistant to the other beta-lactam drugs except cefamandole. Of 22 CNS resistant to cefamandole, 21 were S. haemolyticus.     

Comparison of the E test and a reference agar dilution method for susceptibility testing of anaerobic bacteria

The susceptibility of 146 recent clinical isolates of gram-negative and gram-positive anaerobes was determined by the E test (AB Biodisk) on both Wilkins-Chalgren and PDM ASM II (AB Biodisk) agar. Results of the E test were compared with results obtained by the NCCLS agar dilution method using Wilkins-Chalgren agar. Incubation was for 20 hours and 44 hours in the E test and for 44 hours in the NCCLS method. In general, 44 hour results were more reliable; however, NCCLS readings were made only once after 44 hours. After two days of incubation, 91 % of E test results on Wilkins-Chalgren agar were within one dilution and 98 % within two dilutions of the corresponding NCCLS values; on PDM agar these values were 89 % and 98 %, respectively. Major and very major discrepancies combined were less than 1 %.     

Comparison of agar dilution, broth dilution, disk diffusion, and the E-test for susceptibility testing of penicillin-susceptible and penicillin-resistantStreptococcus pneumoniae

An evaluation to determine the optimal methods for the in vitro susceptibility testing of 41 clinical isolates and the ATCC 49619 strain ofStreptococcus pneumoniae to penicillin was undertaken. No very major or major interpretive errors were observed with the following test methods and media: agar dilution using either Mueller-Hinton medium with lysed horse blood or Haemophilus test medium; broth dilution using cation-adjusted Mueller-Hinton medium with lysed horse blood, Haemophilus test medium, or Todd-Hewitt medium; and the epsilometer test (E-test) using agar containing Mueller-Hinton medium and 5% sheep blood. The disk diffusion method using agar containing Mueller-Hinton medium and 5% sheep blood agar was an effective screening method, requiring confirmation by a dilution susceptibility test method.     

Comparison of E test with agar dilution for antimicrobial susceptibility testing of Neisseria gonorrhoeae.

A collection of 150 Neisseria gonorrhoeae isolates from Africa, where various resistance mechanisms among N. gonorrhoeae isolates are common, was used to the compare E test (AB Biodisk, Solna, Sweden) with agar dilution susceptibility testing. MICs obtained by the E test agreed within 1 log2 concentration by the agar dilution method for 97.5, 97.3, 96.6, 94, and 84.7% of the tested isolates for penicillin, ciprofloxacin, chloramphenicol, tetracycline, and trimethoprim-sulfamethoxazole, respectively. No significant difference in susceptibility categorization was observed between either method. The E test is an attractive alternative to the agar dilution technique and is a more appropriate method for N. gonorrhoeae susceptibility testing in developing countries.     

Evaluation of a novel chromogenic agar medium for isolation and differentiation of vancomycin-resistant Enterococcus faecium and Enterococcus faecalis isolates

The development of reliable and rapid methods for the identification of patients colonized with vancomycin-resistant enterococci (VRE) is central to the containment of this agent within a hospital environment. To this end, we evaluated a prototype chromogenic agar medium (VRE-BMX; bioMérieux, Marcy l'Etoile, France) used to recover VRE from clinical specimens. This medium can also identify isolated colonies as either vancomycin-resistant Enterococcus faecium or Enterococcus faecalis, based on distinct colony colors. We compared the performance of VRE-BMX with bile esculin azide agar supplemented with vancomycin (BEAV). For this study, 147 stool samples were plated on each test medium and examined after 24 and 48 h of incubation. At 24 h, the sensitivity and specificity of each medium were as follows: BEAV, 90.9% and 89.9%, respectively; VRE-BMX, 96.4% and 96.6%, respectively. The positive predictive values (PPV) of VRE-BMX and BEAV at 24 h were 89.8% and 80.7%, respectively. VRE-BMX provided the identification of 10 isolates of vancomycin-resistant E. faecalis and 4 isolates of vancomycin-resistant E. faecium that were not recovered by BEAV. Further, VRE-BMX was capable of identifying patients colonized with both E. faecium and E. faecalis, a feature useful for infection control purposes that is not a function of BEAV. In terms of the recovery of vancomycin-resistant E. faecium and E. faecalis, the sensitivity and PPV were as follows: BEAV, 75.7% and 74.6%, respectively; VRE-BMX, 95.5% and 91.3%, respectively. In this initial evaluation, we found that VRE-BMX provided improved recovery of VRE from stool specimens, with the added advantage of being able to differentiate between vancomycin-resistant E. faecalis and E. faecium. Extending the incubation period beyond 24 h did not significantly improve the recovery of VRE and resulted in decreased specificity.