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PubMed2005

Assessment of accuracy of disk diffusion tests for the determination of antimicrobial susceptibility of common bovine mastitis pathogens: a novel approach.

A novel approach was used to assess disk diffusion accuracy for determination of antibiotic susceptibility of various bovine mastitis pathogens (Escherichia coli, Staphylococcus aureus, Staphylococcus chromogenes, and Streptococcus dysgalactiae). MIC and disk diffusion diameters were compared for 587 bovine mastitis bacterial isolates collected in Israel and 3,186 drug-organism combinations. Results were analyzed by ROC curves, Bayesian statistics, and standard descriptive methods. Low correlation was observed between results of disk diffusion and MIC for S. dysgalactiae and all antimicrobial agents, S. aureus and erythromycin and neomycin, and E. coli and gentamicin, neomycin, and polymyxin B. On a few occasions in which correlation was satisfactory, accepted susceptibility breakpoints to some of the antimicrobial agents resulted in high discrepancies with MIC results and new breakpoints were suggested-e.g., 21 mm for S. aureus susceptibility to penicillin G instead of 29 mm recommended by the National Committee for Clinical Laboratory Standards (NCCLS) and <21 mm, resistant, 21-25 mm, intermediate, and >25 mm, susceptible for susceptibility of E. coli to trimethoprim/sulfamethoxazole. Thus, this approach enabled determination of the most accurate breakpoints that best fitted the specific prevalence of susceptibility in Israel. Thus, we suggest its adoption by microbiology diagnostic laboratories for the provision of accurate antimicrobial susceptibility results when using the disk diffusion test.

Animals↗

PubMed2000

Quality control guidelines for disk diffusion and broth microdilution antimicrobial susceptibility tests with seven drugs for veterinary applications.

This multicenter study proposes antimicrobial susceptibility (MIC and disk diffusion methods) quality control (QC) parameters for seven compounds utilized in veterinary health. Alexomycin, apramycin, tiamulin, tilmicosin, and tylosin were tested by broth microdilution against various National Committee for Clinical Laboratory Standards (NCCLS)-recommended QC organisms (Staphylococcus aureus ATCC 29213, Enterococcus faecalis ATCC 29212, Streptococcus pneumoniae ATCC 49619, Escherichia coli ATCC 25922, and Pseudomonas aeruginosa ATCC 27853). In addition, disk diffusion zone diameter QC limits were determined for apramycin, enrofloxacin, and premafloxacin by using E. coli ATCC 25922, P. aeruginosa ATCC 27853, and S. aureus ATCC 25923. The results from five or six participating laboratories produced >/=99.0% of MICs and >/=95.0% of the zone diameters within suggested guidelines. The NCCLS Subcommittee for Veterinary Antimicrobial Susceptibility Testing has recently approved these ranges for publication in the next M31 document.

Aminoglycosides↗

PubMed2005

Gastroenteritis in US Marines during Operation Iraqi Freedom.

BACKGROUND: Approximately 83,000 US Marines participated in the opening phase of Operation Iraqi Freedom in Spring 2003. A Navy Preventive Medicine laboratory was set up in Ad Diwaniyah, Iraq, to provide clinical diagnostic support for Marine medical units during a period of repositioning in south-central Iraq. METHODS: Specimen collection boxes were sent to >30 primary care medical stations handling 500-900 personnel each. The laboratory had capability to detect many different disease agents, especially those causing febrile illness. Diarrheal stool diagnostic evaluation included plating and biochemical identification, antigen serologic testing, fluorescent antibody antigen detection, disk diffusion antimicrobial susceptibility testing, enzyme immunoassay, and reverse-transcriptase polymerase chain reaction for norovirus (NV). Confirmation and sequencing work for NV was done at Cincinnati Children's Hospital Medical Center (Ohio). RESULTS: By far the most common reason for infectious disease sick call visits was gastrointestinal illness; no other symptoms had equivalent impact. An enteropathogen was detected in 57 (44%) of 129 stool samples, with NV detected in 30 stool samples (23%) obtained from 14 different battalion or similar-sized units; next in frequency were Shigella flexneri and Shigella sonnei, which were isolated from 26 stool samples (20%) obtained from 15 units. Sequencing the NV RNA polymerase gene demonstrated that NV strains represented 7 genetic clusters, including 2 strains from genogroup I and 5 from genogroup II. Ciprofloxacin was effective in vitro against most bacterial agents, but neither doxycyline (which was taken daily as the antimalarial prophylaxis dose) nor trimethoprim-sulfamethoxazole were effective. CONCLUSIONS: Multiple strains of Shigella species and NV predominated, probably because they do not require a large inoculum to cause infection. Otherwise, personnel remained free of infectious illness during this phase of the conflict, because other infectious agents were rare or absent.

Anti-Bacterial Agents↗

PubMed2001

Comparison and evaluation of antimicrobial susceptibility testing of enterococci performed in accordance with six national committee standardized disk diffusion procedures.

Studies were conducted to compare and evaluate antimicrobial susceptibility test results for enterococci obtained by six national committee disk diffusion procedures. Variations in the incidence of isolates in resistance categories and errors were associated with the use of ciprofloxacin, gentamicin, nitrofurantoin, rifampin, and teicoplanin in a number of committee procedures. Results indicate that laboratories performing disk diffusion antimicrobial susceptibility testing may have problems correctly identifying resistance in enterococci with agents used to combat infections and that it may be difficult to compare resistance data for surveillance purposes.

Anti-Bacterial Agents↗

PubMed2006

Comparison of disk diffusion, VITEK 2, and broth microdilution antimicrobial susceptibility test results for unusual species of Enterobacteriaceae.

We compared the antimicrobial susceptibility testing results generated by disk diffusion and the VITEK 2 automated system with the results of the Clinical and Laboratory Standards Institute (CLSI) broth microdilution (BMD) reference method for 61 isolates of unusual species of Enterobacteriaceae. The isolates represented 15 genera and 26 different species, including Buttiauxella, Cedecea, Kluyvera, Leminorella, and Yokenella. Antimicrobial agents included aminoglycosides, carbapenems, cephalosporins, fluoroquinolones, penicillins, and trimethoprim-sulfamethoxazole. CLSI interpretative criteria for Enterobacteriaceae were used. Of the 12 drugs tested by BMD and disk diffusion, 10 showed >95% categorical agreement (CA). CA was lower for ampicillin (80.3%) and cefazolin (77.0%). There were 3 very major errors (all with cefazolin), 1 major error (also with cefazolin), and 26 minor errors. Of the 40 isolates (representing 12 species) that could be identified with the VITEK 2 database, 36 were identified correctly to species level, 1 was identified to genus level only, and 3 were reported as unidentified. VITEK 2 generated MIC results for 42 (68.8%) of 61 isolates, but categorical interpretations (susceptible, intermediate, and resistant) were provided for only 22. For the 17 drugs tested by both BMD and VITEK 2, essential agreement ranged from 80.9 to 100% and CA ranged from 68.2% (ampicillin) to 100%; thirteen drugs exhibited 100% CA. In summary, disk diffusion provides a reliable alternative to BMD for testing of unusual Enterobacteriaceae, some of which cannot be tested, or produce incorrect results, by automated methods.

Anti-Bacterial Agents↗

PubMed1988

Antimicrobial activity and disk diffusion susceptibility testing of U-76,253A (R-3746), the active metabolite of the new cephalosporin ester, U-76,252 (CS-807).

Compound U-76,253A (R-3746), the active metabolite sodium salt of the prodrug ester U-76,252 (CS-807), was demonstrated to be active against members of the family Enterobacteriaceae with 82 and 85% of strains inhibited by less than or equal to 2.0 and less than or equal to 4.0 micrograms/ml, respectively. In addition, U-76,253A inhibited all strains of Branhamella catarrhalis, Haemophilus influenzae, pathogenic Neisseria spp., oxacillin-susceptible Staphylococcus aureus, beta-hemolytic streptococci, and pneumococci at less than or equal to 4.0 micrograms/ml. Pseudomonas spp., Acinetobacter spp., enterococci, and oxacillin-resistant staphylococci were resistant to U-76,253A. This U-76,253A antimicrobial activity and spectrum was generally superior to that of comparison orally administered cephems (cefaclor, cefuroxime, and cefixime) and the amoxicillin-clavulanic acid combination. Tests with beta-lactamase-producing isolates indicated that U-76,253A was bactericidal and that its MICs were only influenced by high inoculum concentrations (10(7) CFU/ml) against type Ia and IVc enzyme-producing strains. Preliminary disk diffusion interpretive zone criteria were calculated for 10- and 30-micrograms U-76,253A disks and several possible susceptible MIC breakpoints. The absolute interpretive agreement between MICs and zone diameters ranged from 87.8 to 95.6%. Final selection of interpretive criteria awaits further U-76,252 pharmacokinetic information.

Anti-Bacterial Agents↗

PubMed1996

Antimicrobial susceptibility testing of Helicobacter pylori. Comparison of E-test, broth microdilution, and disk diffusion for ampicillin, clarithromycin, and metronidazole.

The optimal method for the determination of the minimum inhibitory concentration (MIC) of antimicrobials against Helicobacter pylori has not been established. The epsilometer agar diffusion gradient test (E-Test; AB Biodisk, Solna, Sweden) was compared with broth microdilution, the reference method, and disk diffusion for the antimicrobial susceptibility testing of 122 clinical isolates of H. pylori to ampicillin, clarithromycin, and metronidazole. Isolates were considered to be resistant when the MIC values was > 8 micrograms/ml for either ampicillin or metronidazole and > 2 micrograms/ml for clarithromycin. For an individual isolate, the MICs for ampicillin and clarithromycin determined by broth microdilution and the E-test were highly reproducible, with replicate results being within +/- 1 log2 dilution. The correlation between the MICs determined by E-test and broth microdilution was excellent for both ampicillin and clarithromycin (90.1% and 88.5% were within +/- log2 dilution, and 98.3% and 96.7% of the values were within +/- 2 log2 dilution, respectively). In no instance did the interpretation of "sensitive" or "resistant" differ. Conversely, only 70.5% of the E-test results of metronidazole were within +/- 1 log2 dilution of the broth microdilution results. In addition, 15 (12.3%) of the H. pylori isolates interpreted as resistant by the E-test were sensitive by the broth microdilution method. All discrepancies occurred when the E-test MIC values fell between 8 and 32 micrograms/ml. The results of the ampicillin and clarithromycin disk diffusion assay correlated 100% with the results of the broth microdilution. However, these data suggest that when the E-test MIC results of metronidazole yield values between 8 and 32 micrograms/ml, the MIC should be reevaluated by another method.

Ampicillin↗

PubMed1993

Antimicrobial activity and disk diffusion susceptibility testing of Ro 40-6890, the active metabolite of the new cephalosporin ester, Ro 41-3399.

Ro 40-6890, the active metabolite of Ro 41-3399, was tested against 391 gram-negative and gram-positive clinical isolates. Ro 40-6890 was active against members of the family Enterobacteriaceae, Moraxella catarrhalis, pneumococci, other Streptococcus spp., and methicillin-susceptible staphylococci. Preliminary disk diffusion interpretive zone criteria were calculated for 5-, 10-, and 30-micrograms Ro 40-6890 disks and several possible MIC susceptibility breakpoints. We recommend the use of the 5-micrograms disk and suggest that the following zone diameters be used as breakpoints in clinical trials: susceptible, > or = 21 mm (MIC, < or = 1 microgram/ml); intermediate, 18 to 20 mm (MIC, 2 micrograms/ml); and resistant, < or = 17 mm (MIC, > or = 4 micrograms/ml).

Anti-Bacterial Agents↗

PubMed2006

Antimicrobial-resistant patterns of Escherichia coli and Salmonella strains in the aquatic Lebanese environments.

This study is the first to be conducted in Lebanon on the isolation and molecular characterization and the antimicrobial resistance profile of environmental pathogenic bacterial strains. Fifty-seven samples of seawater, sediment, crab, and fresh water were collected during the spring and summer seasons of 2003. The isolation of Escherichia coli and Salmonella using appropriate selective media revealed that 94.7% of the tested samples were contaminated with one or both of the tested bacteria. The polymerase chain reaction (PCR) was then used to identify the species of both bacteria using various sets of primers. Many pathogenic E. coli isolates were detected by PCR out of which two were identified as O157:H7 E. coli. Similarly, the species of many of the Salmonella isolates was molecularly identified. The confirmed isolates of Salmonella and E. coli were then tested using the disk diffusion method for their susceptibility to four different antimicrobials revealing high rates of antimicrobial resistance.

Animals↗

PubMed1998

An inexpensive and reliable method for routine identification of staphylococcal species.

The aim of this study was to develop a simple, reliable, and inexpensive in-house system for routine species identification of staphylococci in clinical practice. The system combines 15 key tests (including carbohydrate fermentation) performed in micro-well strips and antimicrobial disk diffusion susceptibility tests performed on standardised paper disk method antibiotic sensitivity medium agar. Twenty-eight staphylococcal reference strains belonging to 18 different species were correctly identified using this in-house system. A total of 291 clinical staphylococci isolates were evaluated with the in-house system and a conventional identification scheme. The in-house system identified 281 (96.6%) of these 291 isolates. Eleven different species were recognised. The five species most frequently identified were Staphylococcus epidermidis (48.6%), Staphylococcus aureus (27.8%), Staphylococcus haemolyticus (8.2%), Staphylococcus hominis (5.7%), and Staphylococcus warneri (5.3%). There was an agreement of 86.3% between the species identification obtained with the in-house system and the conventional identification scheme. All coagulase-negative isolates initially identified as species other than Staphylococcus epidermidis as well as indistinctly identified isolates were also evaluated with a commercial identification system. The agreement between species identification obtained with the in-house system and the commercial system for 101 identified isolates was 73%. Several isolates that were difficult to distinguish with the conventional scheme and/or the commercial system were identified with the aid of the antimicrobial susceptibility test included in the in-house system. The described test scheme should be of value for identification of clinically significant staphylococci species.

Agar↗

PubMed1999

Comparison of agar dilution, disk diffusion, MicroScan, and Vitek antimicrobial susceptibility testing methods to broth microdilution for detection of fluoroquinolone-resistant isolates of the family Enterobacteriaceae.

Fluoroquinolone resistance appears to be increasing in many species of bacteria, particularly in those causing nosocomial infections. However, the accuracy of some antimicrobial susceptibility testing methods for detecting fluoroquinolone resistance remains uncertain. Therefore, we compared the accuracy of the results of agar dilution, disk diffusion, MicroScan Walk Away Neg Combo 15 conventional panels, and Vitek GNS-F7 cards to the accuracy of the results of the broth microdilution reference method for detection of ciprofloxacin and ofloxacin resistance in 195 clinical isolates of the family Enterobacteriaceae collected from six U.S. hospitals for a national surveillance project (Project ICARE [Intensive Care Antimicrobial Resistance Epidemiology]). For ciprofloxacin, very major error rates were 0% (disk diffusion and MicroScan), 0.9% (agar dilution), and 2.7% (Vitek), while major error rates ranged from 0% (agar dilution) to 3.7% (MicroScan and Vitek). Minor error rates ranged from 12.3% (agar dilution) to 20.5% (MicroScan). For ofloxacin, no very major errors were observed, and major errors were noted only with MicroScan (3.7% major error rate). Minor error rates ranged from 8.2% (agar dilution) to 18.5% (Vitek). Minor errors for all methods were substantially reduced when results with MICs within +/-1 dilution of the broth microdilution reference MIC were excluded from analysis. However, the high number of minor errors by all test systems remains a concern.

Anti-Infective Agents↗

PubMed1982

Interlaboratory performance of disk agar diffusion and dilution antimicrobial susceptibility tests, 1979-1981. A summary of the microbiology portion of the College of American Pathologists (CAP) surveys.

The antimicrobial susceptibility test results of the 1979-1981 College of American Pathologists (CAP) Microbiology Surveys are summarized for the disk diffusion and dilution test methods. Improved interpretive accuracy (97.5%) was detected for the Special Bacteriology participants compared with those reported for 1972 and 1977-1978. The performance accuracy rates for the Comprehensive and Basic Surveys subscribers was 95.8% and 93.7%, respectively (all surveys accuracy rates was 95.2% from 196,775 responses). The dilution methods were graded since 1980 and showed an acceptable or good performance rate of 98.0%. Most reported MICs were from commercial systems, especially those performed in frozen broth microdilution systems. Eighteen per cent of all susceptibility tests are now reported as MICs. Quality control data shows a trend away from costly daily control practices without compromise of test accuracy. Several problem areas continue to plague these methods: (1) poor predictive value of disk results for enterococci; (2) availability of low potency disks with suboptimal accuracy rates, e.g., the 50 micrograms carbenicillin disk; (3) the release of more new drugs requiring individual testing beyond the capacity of the test systems; (4) vague recommendations for reporting and testing certain bacterial species, e.g., methicillin-resistant staphylococci and penicillin-resistant pneumococci; and (5) the lack of contemporary modification in FDA package insert quality control and interpretive criteria guidelines that might reflect new test or therapeutic information. Overall, the 1979-1981 surveys results show improved performance to a high level of predictive and interpretive accuracy among all subscriber groups coupled with a shift toward the more informative quantitative dilution methodologies.

Agar↗

PubMed1986

Disk diffusion susceptibility testing of two macrolide antimicrobial agents: revised interpretive criteria for erythromycin and preliminary guidelines for roxithromycin (RU 965).

The 15-micrograms erythromycin disk was twice evaluated for interpretive accuracy against 417 and then 266 strains of gram-positive cocci, Neisseria meningitidis, and Haemophilus influenzae by using the criteria suggested by the National Committee for Clinical Laboratory Standards. These studies suggest a revision of streptococcal and Staphylococcus sp. interpretive guidelines to criteria (greater than or equal to 23 mm = susceptible, less than or equal to 13 mm = resistant) that are more compatible with in vivo erythromycin concentrations. It is also recommended that zone diameters be modified for H. influenzae (greater than or equal to 23 mm = susceptible, less than 22 mm = resistant) and that meningococci not be tested. A wide moderately susceptible category (1.0 to 4.0 micrograms/ml) would primarily include enterococcus strains and those organisms that would then respond only to parenteral administration of erythromycin. Roxithromycin (RU 965 or RU 28965), a new oxime ether erythromycin derivative, was also evaluated by investigator-prepared 15-micrograms disks and later with 30- and 60-micrograms commercial disks. Although roxithromycin was comparable to erythromycin in activity and regression line statistics, changes in the susceptible disk criteria were necessary because of superior roxithromycin serum concentrations and a longer serum half-life. Preliminary susceptible breakpoint criteria were greater than 21 mm = susceptible, 10 to 20 mm = indeterminate, and less than or equal to 9 mm = resistant. By using the recommended interpretive criteria for both macrolides, less than 98% absolute agreement was obtained, therefore suggesting the application of a spectrum class concept.

Erythromycin↗

PubMed2006

Resistance to erythromycin and rising penicillin MIC in Streptococcus pyogenes in India.

There has been worldwide resurgence in the incidence of Streptococcus pyogenes infection and its sequelae. S. pyogenes remains uniformly susceptible to penicillin, and it is speculated that its minimum inhibitory concentration (MIC) has not changed during the past 70 years. The purpose of the present study was to determine the occurrence and pattern of resistance to penicillin and erythromycin amongst clinical isolates of S. pyogenes. A total of 34 clinical strains of S. pyogenes were identified by standard procedures. Antimicrobial susceptibility was analyzed by the Kirby-Bauer method of disk diffusion, and the E-test method was used to determine the MIC to penicillin and erythromycin. All the strains were sensitive to penicillin, clindamycin and vancomycin on disk diffusion. Ten (29.4%) strains were resistant to erythromycin. The pattern of macrolide resistance observed was M type. By the E-test method, 7 (20.6%) strains were penicillin nonsusceptible and 6 (17.6%) were erythromycin resistant. We concluded that surveillance of its susceptibility pattern is crucial to monitoring the development of antibiotic resistance in S. pyogenes.

Disk Diffusion Antimicrobial Tests↗

PubMed1999

[Antimicrobial resistance testing of H. pylori epsilometer test and disk diffusion test].

The resistance of Helicobacter pylori to antibiotics leaves great influence on treatment outcome. Routine antimicrobial susceptibility testing of H. pylori should be a simple, inexpensive and standardized method to be adopted. Agar and broth dilution techniques are difficult to perform and not practical. E-test and disk diffusion methods have the advantage of allowing the visualization of resistant subpopulations of bacteria within zones of inhibitions. We studied the susceptibility results of 246 H. pylori isolates against metronidazole and clarithromycin using E-test and disk diffusion test. The results of disk diffusion test were well correlated with these of E-test. Our results showed that disk method is cost effective and simple for the screening for antimicrobial resistance of H. pylori.

Clarithromycin↗

PubMed2006

Multicenter studies of tigecycline disk diffusion susceptibility results for Acinetobacter spp.

Acinetobacter sp. isolates having multidrug resistance (MDR) patterns have become common in many medical centers worldwide, limiting therapeutic options. A five-center study tested 103 contemporary clinical Acinetobacter spp., including MDR strains, by reference broth microdilution and disk diffusion (15-mug disk content) methods against tigecycline. Applying U.S. Food and Drug Administration tigecycline breakpoint criteria for Enterobacteriaceae (susceptibility at < or =2 microg/ml [< or =1 microg/ml by the European Committee on Antimicrobial Susceptibility Testing]; disk diffusion breakpoints at > or =19 mm and < or =14 mm) to Acinetobacter spp. led to an unacceptable error rate (23.3%). However, an adjustment of tigecycline disk diffusion breakpoints (susceptible/resistant) to > or =16/ < or =12 mm reduced intermethod errors to an acceptable level (only 9.7%, all minor).

Acinetobacter↗

PubMed1994

Comparison of direct and standardized testing of infected urine for antimicrobial susceptibilities by disk diffusion.

A total of 14,272 urine specimens were examined over one year to determine the validity of direct antimicrobial agent susceptibility testing against ampicillin, amoxicillin-clavulanic acid, cephalothin, gentamicin, norfloxacin, and trimethoprim. A comparison between direct and standardized disk diffusion tests was made for a total of 1,106 urine specimens containing > or = 10(5) organisms per ml in pure culture. There were 5,821 individual organism-antimicrobial agent challenges compared for the two testing methods, and there was complete agreement of susceptibility category in 5,492 comparisons (94.3%). Initially, discordant results were reduced from 5.7 to 2.1% when the intermediate category was considered susceptible. Intralaboratory variation was assessed by testing another 453 organisms by the standard National Committee for Clinical Laboratory Standards (NCCLS) method on two consecutive days; there was complete agreement in 96.1% of comparisons. When results of direct and standardized testing were simply classified as susceptible or resistant, there was 1.1% discordance. When simple same-day tests were used together with predictable patterns of susceptibility and resistance, 536 (48.5%) of 1,106 isolates could be identified satisfactorily to the genus or species level. For laboratory reporting purposes, the direct method is equivalent to the standard method when the urine being tested is infected with > or = 10(5) organisms of a single type per ml. The presence or absence of preexisting antimicrobial agents in urine did not appreciably influence the results. This procedure allows the earlier reporting of susceptibility results and facilitates less expensive identification of many organisms. Costs and benefits need to be determined in each institution.

Anti-Bacterial Agents↗

PubMed2006

[Comparison of disk-diffusion method and PCR for detection of methicillin resistance in Staphylococcus aureus strains].

The aim of the study was to compare the disk-diffusion (oxacillin 1 microg, cefoxitin 30 microg) method and PCR for detection of methicillin-resistance in S. aureus. The investigation were carried out on 120 S. aureus strains isolated from clinical materials of patients hospitalized in the University Hospital at the L. Rydygier Collegium Medicum in Bydgoszcz, University of Nicolaus Copernicus in Toruń. Of the 120 S. aureus strains tested, 60 (50%) were mecA-positive by PCR. Consistency of results between oxacillin disk-difussion method and PCR amounted 92.5% and cefoxitin disk-diffusion method and PCR--98.3%. The oxacillin disk-difussion method falsely identified 3 (2.5%) strains as MSSA (sensitivity 95.0%) and 4 strains as MRSA (specificity 93.3%) in comparison with PCR. The cefoxitin disk-diffusion method falsely identified 2 (1.6%) strains as MSSA (sensitivity 96.7%) and there were no false resistant results (specificity 100%). Our results showed that in disk-diffusion tests, cefoxitin is a better than oxacillin for the identification of MRSA.

Bacterial Proteins↗