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The accuracy of antimicrobial disk sensitivity testing in urinary tract infections.

Fifty bacterial strains shown resistant to ampicillin, cephalothin or tetracycline by Kirby-Bauer disk diffusion susceptibility testing were isolated from patients with urinary infections. Inhibitory activity in urine of volunteers given these antimicrobial agents and tube dilution sensitivity testing indicated that agar disk diffusion gradients do not provide sufficiently high antimicrobial concentrations to predict accurately clinical efficacy.

Ampicillin↗

Disk diffusion susceptibility testing of Branhamella catarrhalis with ampicillin and seven other antimicrobial agents.

A total of 74 clinical isolates of Branhamella catarrhalis were characterized with respect to their ampicillin, amoxicillin-clavulanate, cephalothin, cefaclor, erythromycin, tetracycline, chloramphenicol, and trimethoprim-sulfamethoxazole MICs and zones of inhibition. Disk diffusion tests were performed according to the guidelines of the National Committee for Clinical Laboratory Standards with two different media (Mueller-Hinton agar and chocolate Mueller-Hinton agar) and plates incubated under two atmospheric conditions (ambient air and 5 to 7% CO2). Optimum disk diffusion test results were obtained with Mueller-Hinton agar plates incubated in ambient air with all eight antimicrobial agents. On the basis of comparisons of MICs versus zones of inhibition, the following zone diameter interpretive criteria were defined for testing B. catarrhalis with disks containing 10 micrograms of ampicillin: greater than or equal to 38 mm, susceptible; 20 to 37 mm, moderately susceptible; less than or equal to 19 mm, resistant. The respective MIC correlates were less than or equal to 0.06, 0.125 to 0.5, and greater than or equal to 1.0 micrograms/ml. Because of the absence of frankly resistant test organisms, it was not possible to make definitive recommendations pertaining to disk diffusion tests with amoxicillin-clavulanate, cephalothin, cefaclor, erythromycin, tetracycline, chloramphenicol, and trimethoprim-sulfamethoxazole. Evidence is presented, however, which suggests that the current National Committee for Clinical Laboratory Standards disk diffusion interpretive criteria for nonfastidious bacteria can be applied to B. catarrhalis, at least as they pertain to the susceptible category with cephalothin, amoxicillin-clavulanate, erythromycin, tetracycline, chloramphenicol, and trimethoprim-sulfamethoxazole. With cefaclor, a zone diameter of greater than or equal to 21 mm was determined to adequately define the susceptible category.

Ampicillin↗

Antibacterial activity of WY-49605 compared with those of six other oral agents and selection of disk content for disk diffusion susceptibility testing.

The in vitro antimicrobial activity of an oral penem, WY-49605, was compared with those of six other oral antimicrobial agents against 598 bacterial isolates representing 51 different species. WY-49605 exhibited good activity against most gram-positive bacteria and members of the family Enterobacteriaceae. It had little activity against nonfermenting gram-negative bacilli, Enterobacter spp., Serratia spp., enterococci, Staphylococcus haemolyticus, and methicillin-resistant Staphylococcus aureus. Its activity was unaffected by the beta-lactamases of Neisseria gonorrhoeae, Haemophilus influenzae, and staphylococci. Disk diffusion susceptibility tests were performed with 5-, 10-, 15-, and 30-micrograms WY-49605 disks. The 5-micrograms disk is recommended, with tentative breakpoints of > or = 16 mm for susceptibility (MIC, < or = 2.0 microgram/ml) and < or = 12 mm for resistance (MIC, > or = 8.0 micrograms/ml).

Anti-Bacterial Agents↗

Detecting erythromycin resistance in Streptococcus pyogenes: reliability of the disk diffusion method and the breakpoint susceptibility testing method. Finnish Study Group for Antimicrobial Resistance (FIRE).

Erythromycin susceptibility of clinical Streptococcus pyogenes isolates was determined at 19 Finnish clinical microbiology laboratories by their routine disk diffusion method and by a screening method adapted from the breakpoint susceptibility testing method. Results obtained at 12 laboratories using 4 major variants of the disk method were further evaluated. From these laboratories, 286 consecutive resistant and 349 consecutive susceptible isolates were sent to the Antimicrobial Research Laboratory, Turku where the MIC of erythromycin was determined. 96% and 97% of the disk results were correct, as compared with MIC results, when general and laboratory-specific breakpoints, respectively, were used. The results of the screening method were comparable to those of the disk method.

Drug Resistance, Microbial↗

Evaluations of antimicrobial susceptibility test proficiency by the College of American Pathologists Survey program. A clarification of quality control recommendations.

The College of American Pathologists (CAP) Interlaboratory Survey Program in microbiology was reviewed for quality control practices of the Special Bacteriology participants (907 laboratories) using the disk agar diffusion antimicrobial susceptibility test method. Data from a questionnaire was compared to performance results on two well recognized quality control strains, e.g., Pseudomonas aeruginosa ATCC 27853 and Streptococcus faecalis ATCC 29212. The results show no significant difference in survey performance between those participants using daily controls (97.78 and 96.99% acceptable results) and those using weekly controls (97.33 and 96.67% acceptable results). Similarly, the number of tests done per work day had little influence on the reported mean zone diameter and subscriber results were comparable to published means from the reference literature. Recommendations are made to use weekly quality control after test proficiency has been repeatedly demonstrated by daily or batch control results. More frequent testing would be required if any reagent lot were replaced or altered. Comparable data on quality control intervals of greater than 7 days was not evaluated due to small numbers of test results.

Cost Control↗

Multilaboratory evaluation of an agar diffusion disk susceptibility test for rapidly growing anaerobic bacteria.

A multilaboratory collaborative study was undertaken to determine whether the anaerobic disk diffusion test of Horn et al. could be performed reproducibly and accurately. Tests with nine different antimicrobial disks were evaluated. Reproducibility of the agar diffusion disk method was similar to that of the reference agar dilution test procedure. The anaerobic disk diffusion procedure was found to be a potentially useful method for testing some antimicrobial agents against rapidly growing anaerobes belonging to the Bacteroides fragilis group. These promising results warrant further investigations and validations.

Bacteria, Anaerobic↗

Antimicrobial activity of LY164846, a new oral cephalosporin, and recommendations for disk diffusion tests.

LY164846 is a new oral cephalosporin with a limited spectrum of antimicrobial activity that includes staphylococci (other than methicillin-resistant), streptococci (other than enterococci), Haemophilus influenzae (beta-lactamase-negative and beta-lactamase-positive), Branhamella catarrhalis (beta-lactamase-negative and beta-lactamase-positive), and Neisseria species (beta-lactamase negative and beta-lactamase-positive). The tentative recommendations for susceptibility breakpoints are less than or equal to 4 micrograms/ml and greater than or equal to 19 mm for susceptible, 8 micrograms/ml and 15-18 mm for intermediate, and greater than or equal to 16 micrograms/ml and less than or equal to 14 mm for resistant.

Cephalosporins↗

Antimicrobial susceptibility testing of aquatic bacteria: quality control disk diffusion ranges for Escherichia coli ATCC 25922 and Aeromonas salmonicida subsp. salmonicida ATCC 33658 at 22 and 28 degrees C.

Quality control (QC) ranges for disk diffusion susceptibility testing of aquatic bacterial isolates were proposed as a result of a multilaboratory study conducted according to procedures established by the National Committee for Clinical Laboratory Standards (NCCLS). Ranges were proposed for Escherichia coli ATCC 25922 and Aeromonas salmonicida subsp. salmonicida ATCC 33658 at 22 and 28 degrees C for nine different antimicrobial agents (ampicillin, enrofloxacin, erythromycin, florfenicol, gentamicin, oxolinic acid, oxytetracycline, ormetoprim-sulfadimethoxine, and trimethoprim-sulfamethoxazole). All tests were conducted on standard Mueller-Hinton agar. With >/=95% of all data points fitting within the proposed QC ranges, the results from this study comply with NCCLS guidelines and have been accepted by the NCCLS Subcommittee for Veterinary Antimicrobial Susceptibility Testing. These QC guidelines will permit greater accuracy in interpreting results and, for the first time, the ability to reliably compare susceptibility test data between aquatic animal disease diagnostic laboratories.

Aeromonas↗

Antimicrobial activity of coumermycin and recommendations for disk diffusion tests with 5- and 15-micrograms disks.

Coumermycin was found to be extremely active against methicillin-susceptible and -resistant Staphylococcus spp. (MIC 90, less than or equal to 0.002 microgram/ml). Vancomycin and coumermycin were equally active against Streptococcus spp. (MIC 90s, 0.5 microgram/ml) and both were superior to fusidic acid (MIC 90, 8.0 micrograms/ml). The enterococci had the highest MICs for all three drugs. Disk diffusion susceptibility tests using either 5 or 15 micrograms coumermycin disks seem reliable. The tentative interpretive breakpoints for testing the Staphylococcus spp. only are: 5 micrograms disk-susceptible greater than or equal to 17 and resistant less than or equal to 13 mm; 15 micrograms disk-susceptible greater than or equal to 20 mm and resistant less than or equal to 16 mm. These zone criteria have approximate coumermycin MIC correlates of less than or equal to 0.12 microgram/ml for susceptible and greater than or equal to 0.5 microgram/ml for resistant.

Aminocoumarins↗

Cefmenoxime (SCE-1365), a new cephalosporin: in vitro activity, comparison with other antimicrobial agents, beta-lactamase stability, and disk diffusion testing with tentative interpretive criteria.

The in vitro activity of cefmenoxime (SCE-1365) was evaluated in a multiphased collaborative investigation. Over 7,500 consecutive clinical isolates were tested in five laboratories, and greater than 90% of the following organisms were inhibited by cefmenoxime at the following concentrations: Enterobacteriaceae and non-enterococcal streptococci, </=0.125 mug/ml; Staphylococcus aureus, </=2.0 mug/ml; and nonfermenting gram-negative bacilli and Bacteroides fragilis group, </=32 mug/ml. Both beta-lactamase-producing and -nonproducing Haemophilus influenzae and Neisseria gonorrhoeae were inhibited by cefmenoxime at </=0.03 mug/ml. The spectrum of cefmenoxime was similar to that of other, newer cephalosporins, particularly cefotaxime. A pronounced inoculum effect was found with some species upon increasing inocula from 10(5) to 10(7) colony-forming units per ml, resulting in an approximate eightfold increase in minimum inhibitory concentrations. Cefmenoxime was bactericidal when tested with inocula of 10(5) colonyforming units per ml, and mean differences between the minimum inhibitory concentration and the minimum lethal concentration were less than one log(2) dilution. No significant hydrolysis of cefmenoxime by five different beta-lactamases was detectable, and cefmenoxime exhibited marked inhibition of type I beta-lactamases. Regression and error rate-bounded analyses of results of disk diffusion and reference broth microdilution susceptibility tests were performed on 421 bacterial isolates, and the following tentative zone size breakpoints are proposed: >/=22 mm, susceptible; </=14 mm, resistant; and 15 to 21 mm, moderately susceptible (indeterminate). These data and cross-resistance studies with other newer cephalosporins indicate marked similarity of in vitro activity within this group of drugs, particularly between cefmenoxime, moxalactam, and cefotaxime. Any one of these could serve as the representative for the disk diffusion testing of this group of drugs if comparable minimum inhibitory concentration breakpoints were used for each drug.

Bacteria↗

Antimicrobial activity of fusidic acid and disk diffusion susceptibility testing criteria for gram-positive cocci.

The in vitro activity of fusidic acid was assessed and was compared with those of cloxacillin, cefamandole, vancomycin, teicoplanin, ofloxacin, ciprofloxacin, pefloxacin, and fleroxacin against 500 gram-positive cocci: 151 Staphylococcus aureus, 197 coagulase-negative staphylococci, and 152 Enterococcus faecalis strains. All clinical isolates were concomitantly tested by disk diffusion and agar dilution procedures as outlined by the National Committee for Clinical Laboratory Standards. The results with fusidic acid were further analyzed by regression line and error rate-bounded methods. With control American Type Culture Collection organisms, the values were within the limits of the National Committee for Clinical Laboratory Standards or published limits. The incidence of resistance to fusidic acid was 0.7% for S. aureus, 2.5% for coagulase-negative staphylococci, and 99.3% for E. faecalis. The correlation coefficient between the results of disk diffusion and agar dilution tests with fusidic acid was 0.90. Current interpretive criteria for susceptibility to fusidic acid (i.e., MIC of < 2 micrograms/ml and inhibitory zone of 20 mm) gave 1% false susceptibility (all strains being E. faecalis). This error rate is practically eliminated if a zone diameter of 21 mm is considered the breakpoint for susceptibility.

Drug Resistance, Microbial↗

Analgesic and antiinflammatory properties of Sideritis lotsyi var. Mascaensis.

The antiinflammatory, analgesic and antimicrobial activities of crude ethanol extracts of Sideritis lotsyi var. mascaensis (Lamiaceae), and chloroform and aqueous fractions were evaluated in mice using paw and ear oedema induced by carrageenan and 12-o-tetradecanoyl-phorbol-acetate (TPA), respectively, as inflammation models, the writhing test induced by acetic acid for evaluating analgesic activity and the disk-diffusion method for testing antimicrobial actions. The results obtained demonstrated significant topical antiinflammatory and analgesic activities for the ethanol extract and chloroform fraction, but no relevant antimicrobial activity against the microorganisms tested.

Acetic Acid↗

Effects of diet formulations containing proteins from different sources on intestinal colonization by Campylobacter jejuni in broiler chickens.

The objective of this study was to compare the effects of 3 diet formulations containing different protein sources (animal, plant, and a combination of animal and plant) on the colonization of Campylobacter jejuni in the gastrointestinal tract of broiler chickens. A freshly isolated strain of C. jejuni (biotype IV, serotype HS O:21, O:29, HL untypable) from a broiler chicken was used to infect 3-day-old chicks that had been free of C. jejuni; 0.5 mL of an inoculum containing 10(8) colony-forming units was administered orally. Shedding of the organism was studied, and C. jejuni in the ceca, jejuni, and crop were enumerated by quantitative culture. The isolates recovered from the birds during the study period of 35 d were characterized and confirmed as C. jejuni by the use of standard methods and underwent biotyping, serotyping, antimicrobial susceptibility testing by disk diffusion and the E-test, and flagellin gene typing. A cyclical pattern of shedding of C. jejuni was observed in all the birds. Colonization was highest in the ceca. The ceca of birds receiving plant-protein-based feed had significantly less colonization then the ceca of birds receiving the other types of feed, whereas the differences in colonization of the jejuni and crops were not significant. Characterization by biotyping, serotyping, and flagellin gene typing showed that 95% of the recovered isolates were identical to the strain used for infecting the chicks. However, with the Lior-HL typing scheme, 74% of the recovered isolates were HL untypable. Antimicrobial resistance testing did not reveal significant differences between the infecting strain and the recovered isolates among the different feed groups.

Administration, Oral↗

Colonization of HIV-infected outpatients in Taiwan with methicillin-resistant and methicillin-susceptible Staphylococcus aureus.

To better understand the epidemiology of bacterial pathogens with particular public health importance in Taiwan, we determined the prevalence of nasal colonization with methicillin-susceptible Staphylococcus aureus (MSSA) and methicillin-resistant S. aureus (MRSA) in a cohort of HIV-infected patients attending two hospital outpatient departments. All HIV-infected patients followed regularly between May and September 1999 were enrolled and cultures of the anterior nares were performed using a dry sponge swab. All confirmed S. aureus isolates underwent antimicrobial susceptibility testing using disk diffusion according to recommendations of the National Committee for Clinical Laboratory Standards. Of a total of 162 outpatients studied, 48 (30%) were found colonized with S. aureus including 39 (24%) colonized with MSSA and 9 (6%) colonized with MRSA. The only factor associated with MSSA colonization was receipt of trimethoprim-sulphamethoxazole which appeared protective (relative risk 0.4, 95% confidence interval [CI(95)] 0.2-0.78, P = 0.006). In contrast, ciprofloxacin use was an independent risk factor for MRSA colonization (conditional odds ratio [OR] 11.9, CI(95) 1.8-77.8, P = 0.010) along with a one-quartile reduction in CD(4) count (OR 3.9, CI(95) 1.1-14.3, P = 0.04). Although MRSA colonization was not associated with hospitalization within the previous three months, the multi-drug resistance pattern of MRSA isolates suggests strains were at some point acquired in the healthcare setting. Our study shows that the rate of S. aureus colonization in Taiwanese HIV-infected outpatients is 30%. Low CD4+ counts are most likely associated with other unmeasured risk factors for MRSA. Antimicrobial use may function alternatively as a protective or risk factor for colonization with S. aureus, depending upon the drugs involved and resistance encountered. Fluoroquinolone use may have an important role in the spread of MRSA from inpatient to outpatient settings.

Adult↗

Survey of antimicrobial susceptibility testing practices of veterinary diagnostic laboratories in the United States.

OBJECTIVE: To describe antimicrobial susceptibility testing practices of veterinary diagnostic laboratories in the United States and evaluate the feasibility of collating this information for the purpose of monitoring antimicrobial resistance in bacterial isolates from animals. DESIGN: Cross-sectional study. PROCEDURES: A questionnaire was mailed to veterinary diagnostic laboratories throughout the United States to identify those laboratories that conduct susceptibility testing. Nonrespondent laboratories were followed up through telephone contact and additional mailings. Data were gathered regarding methods of susceptibility testing, standardization of methods, data management, and types of isolates tested. RESULTS: Eighty-six of 113 (76%) laboratories responded to the survey, and 64 of the 86 (74%) routinely performed susceptibility testing on bacterial isolates from animals. Thirty-four of the 36 (94%) laboratories accredited by the American Association of Veterinary Laboratory Diagnosticians responded to the survey. Laboratories reported testing > 160,000 bacterial isolates/y. Fifty-one (88%) laboratories reported using the Kirby-Bauer disk diffusion test to evaluate antimicrobial susceptibility; this accounted for 65% of the isolates tested. Most (87%) laboratories used the NCCLS (National Committee for Clinical Laboratory Standards) documents for test interpretation. Seventy-five percent of the laboratories performed susceptibility testing on bacterial isolates only when they were potential pathogens. CONCLUSIONS: The veterinary diagnostic laboratories represent a comprehensive source of data that is not easily accessible in the United States. Variability in testing methods and data storage would present challenges for data aggregation, summary, and interpretation.

Animals↗

Risk Factors for Ciprofloxacin Resistance in Bloodstream Infections Due to Extended-Spectrum beta-Lactamase-Producing Escherichia coli and Klebsiella pneumoniae.

The present study was conducted to identify risk factors for ciprofloxacin resistance in bloodstream infections due to extended-spectrum beta -lactamase-producing Escherichia coli and Klebsiella pneumoniae (ESBL-EK). ESBL production was determined in stored E. coli and K. pneumoniae blood isolates from January, 1998, to December, 2002, by National Committee for Clinical Laboratory Standards (NCCLS) guidelines and/or double-disk synergy test. Antimicrobial susceptibility was determined by the disk diffusion test method. A total of 133 patients with ESBL-EK bacteremia were analyzed retrospectively. A total of 80 (60.2%) patients experienced bacteremia due to strains resistant to ciprofloxacin. There were no significant differences in age, sex, and APACHE II score between the ciprofloxacin-resistant group (CIP-R) and-susceptible group (CIPS). The most common primary site of infection in CIP-R was pancreaticobiliary tract infection (46/80, 57.5%) and that in CIP-S was unknown primary site (23/53, 43.4%). Independent risk factors for ciprofloxacin resistance were: prior use of fluoroquinolones (OR, 5.53; 95% CI, 1.56-25.42, p = 0.032), indwelling urinary catheter (OR, 3.68; 95% CI, 1.27-10.67, p = 0.017), and invasive procedure within 72 hr prior to bacteremia (OR, 4.03; 95% CI, 1.44-11.25, p = 0.008). Our data suggest that strategies designed to reduce the ciprofloxacin resistance rate in ESBL-EK strains should focus on limiting the use of fluoroquinolones and minimizing invasive procedures, including insertion of a urinary catheter.

Adolescent↗

Genetic diversity and antibiotic resistance patterns in a campylobacter population isolated from poultry farms in Switzerland.

The diversity and genetic interrelation of Campylobacter jejuni and C. coli isolated from Swiss poultry were assessed by three independent typing methods. Samples were derived prior to slaughter from 100 randomly selected flocks (five birds per flock) raised on three different farm types. The observed flock prevalence was 54% in total, with 50% for conventional and 69% for free-range farms. Birds held on farms with a confined roaming area had the lowest prevalence of 37%. Campylobacter isolates were characterized by amplified fragment length polymorphism (AFLP), restriction fragment length polymorphism of flaA PCR fragments (flaA-RFLP), and disk diffusion testing for eight antimicrobial agents that are commonly used in veterinary or human medicine in Switzerland. Analysis of the genotypic results indicates that the Campylobacter population in Swiss poultry is genetically highly diverse. Nevertheless, occasionally, isolates with identical or nearly identical characteristics were isolated from different farms or farm types in different locations. Genetic typing by AFLP and flaA-RFLP was found to be complementary. The majority of isolates (67%) were susceptible to all tested antibiotics; however, single, double, and triple resistances were observed in 7%, 23%, and 2% of the strains, respectively. There was no correlation between genotype and antibiotic resistance. Surprisingly, sulfonamide resistance was frequently found together with streptomycin resistance. Our findings illustrate the results of common genetic exchange in the studied bacterial population.

Animal Husbandry↗

Performance of the chromogenic medium CHROMagar Staph Aureus and the Staphychrom coagulase test in the detection and identification of Staphylococcus aureus in clinical specimens.

CHROMagar Staph aureus (CSAM) (CHROMagar Microbiology, Paris, France) is a new chromogenic medium designed to enable detection of colonies of Staphylococcus aureus by their pink color. A total of 775 specimens were cultured in parallel on CHROMagar Staph aureus and conventional media. Among the 267 S. aureus strains recovered on at least one medium, 263 were isolated on CSAM medium (sensitivity, 98.5%), and 245 (sensitivity, 91.8%) were isolated on conventional media. The specificity of presumptive identification of S. aureus on the basis of pink colony color on CSAM medium was 97% (493 of 508). This specificity increased to 100% when coagulase detection with the Staphychrom coagulase test was added and to 98.8% when S. aureus surface components were detected by agglutination in the Pastorex Staph Plus test. Susceptibility testing of 67 S. aureus strains, performed in parallel on pink CSAM colonies and on colonies grown on blood agar, gave similar results. Thus, rapid and accurate recognition and identification of S. aureus isolates were achieved with CSAM as the primary isolation medium, followed by the staphylocoagulase Staphychrom test. Antimicrobial susceptibility testing (disk-diffusion method or ATB STAPH System) can be performed directly on pink CSAM colonies.

Chromogenic Compounds↗