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.     

Mixed outbreak ofBordetella pertussis andBordetella parapertussis infection in Finland

The epidemiology of whooping cough in a vaccinated population was studied during an outbreak of paroxysmal cough in an elementary school with 258 pupils in Turku, Finland. Nasopharyngeal specimens for isolation ofBordetella pertussis and/or sera for ELISA detection of antipertussis immunoglobulin M, A and G antibodies were taken from 94 % of children who were prospectively followed for two months.Bordetella pertussis was isolated in six patients, and 17 culture-positive cases withBordetella parapertussis were identified. Patients withBordetella pertussis orBordetella parapertussis were found simultaneously in the same classrooms. Comparison of immunoglobulin M responses toBordetella pertussis andBordetella parapertussis was used for differential diagnosis of these two infections. Twenty-six cases with pertussis and 27 cases with parapertussis were diagnosed. The results of this prospective study suggest thatBordetella parapertussis is a more common etiologic agent of mild respiratory tract infection among vaccinated school-aged children than is generally recognised. The possibility thatBordetella pertussis was converted toBordetella parapertussis during this outbreak is discussed.     

Activity of new macrolides againstBordetella pertussis andBordetella parapertussis

MICs and MBCs of four new macrolides (azithromycin, clarithromycin, dirithromycin and roxithromycin) and two older macrolides (erythromycin and josamycin) forBordetella pertussis andBordetella parapertussis were determined. The activity of the new macrolides was as good as that of erythromycin, while josamycin was slightly less active.Bordetella parapertussis was more resistant thanBordetella pertussis.     

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.     

Role of pertussis toxin in causing symptoms ofBordetella parapertussis infection

Whooping cough can be caused by eitherBordetella pertussis orBordetella parapertussis. Although the two species share an almost complete DNA identity,Bordetella parapertussis does not produce pertussis toxin, which is thought to be the main virulence factor ofBordetella pertussis. In order to elucidate the role of pertussis toxin in causing the typical symptoms of whooping cough, clinical information from 33 patients with culture-positiveBordetella parapertussis infection was collected and compared to that from 331 patients with infection caused byBordetella pertussis. Isolated strains ofBordetella parapertussis lacked pertussis toxin expression, as was demonstrated by negative tests for histamine sensitization. This was further substantiated in vivo by a significantly lower leukocyte count in the parapertussis group as compared to the pertussis group. Frequencies of typical symptoms of whooping cough, such as paroxysmal coughing, whooping and vomiting, were almost identical in the two groups. Nocturnal coughing and contact anamnesis were noted more often in theBordetella pertussis group. Children in the parapertussis group were significantly more often vaccinated with whole-cell pertussis vaccine than children infected withBordetella pertussis. The results indicate that pertussis toxin may not play a decisive role in causing the typical symptoms of whooping cough, such as paroxysmal coughing, whooping and vomiting.     

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 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 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.     

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 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 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 %.     

Shelf life of prepared Bordet-Gengou and Regan-Lowe agar plates for isolation ofBordetella pertussis

The shelf life of prepared agar plates used for the isolation ofBordetella pertussis was studied. They contained Bordet-Gengou agar, Bordet-Gengou agar with cephalexin, Regan-Lowe agar, Regan-Lowe agar with cephalexin, or Regan-Lowe agar with both cephalexin and amphotericin B. Plates stored were compared to freshly prepared control plates for up to a maximum of 18 weeks. They were inoculated with clinical isolates ofBordetella pertussis, either in pure culture, or mixed with a defined oropharyngeal flora. Bordet-Gengou agar plates may be used, with proper storage at 4 °C in airtight-sealed plastic bags, for up to 10 weeks, Regan-Lowe agar plates for up to 14 weeks. Field studies are needed to substantiate our findings.     

Comparison of the Sceptor Pseudomonas Plus MIC Panel with agar dilution for susceptibility testing of Pseudomonas aeruginosa.

The antimicrobial susceptibilities of 100 clinical isolates of Pseudomonas aeruginosa to gentamicin, amikacin, tobramycin, ticarcillin, piperacillin, and ceftazidime were determined by using the Sceptor system (BBL Microbiology Systems, Cockeysville, Md.), and the results were compared with those obtained using the National Committee for Clinical Laboratory Standards reference agar dilution method. Excellent correlation was observed for the aminoglycosides, with greater than 95% agreement within 1 doubling dilution of the reference agar dilution MIC, while ticarcillin and piperacillin showed lower percent agreement values of 91 and 88%, respectively.     

Comparison of E test and agar dilution for antimicrobial susceptibility testing of Stenotrophomonas (Xanthomonas) maltophilia.

Currently recommended dilution test methods for the determination of antimicrobial susceptibility of Stenotrophomonas (Xanthomonas) maltophilia are labor-intensive and often impractical in many clinical laboratories. We compared the E test with the agar dilution method for susceptibility testing of 176 clinical isolates of S. maltophilia against 16 antimicrobial agents. The MICs obtained by E test correlated well with those determined by the agar dilution method, with an overall agreement of 94%. Very major and major errors occurred infrequently (0.6 to 2.9%) when testing beta-lactam agents, tobramycin, trimethoprim-sulfamethoxazole, and fluoroquinolones. The E test was found to be accurate and easy to perform. For most antimicrobial agents tested against S. maltophilia, the E test is an acceptable alternative susceptibility test method.     

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 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.     

Two-site comparison of broth microdilution and semisolid agar dilution methods for susceptibility testing of Cryptococcus neoformans in three media.

This study evaluated the inter- and intralaboratory agreement between results of the semisolid agar dilution and broth microdilution methods of antifungal susceptibility testing of Cryptococcus neoformans. Three media were tested in two laboratories. The drugs tested were amphotericin B, flucytosine, itraconazole, fluconazole, and Schering 39304. Analysis by kappa statistics revealed good agreement between the laboratories for the two methods. The highest level of inter- and intralaboratory agreement was observed in RPMI 1640 with L-glutamine followed by Eagle's minimum essential medium and yeast nitrogen broth. The broth microdilution method appears more suitable than the semisolid agar dilution method for testing cryptococci because of its ease in performance, cost, and simplicity.     

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.