Antimicrobial susceptibility testing: general principles and contemporary practices.
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Anti-Infective Agents↗
Biomedical subjects
J H Jorgensen
Publications and source records attributed to J H Jorgensen.
Development of quinolone-resistant Campylobacter fetus bacteremia in human immunodeficiency virus-infected patients.
Campylobacter fetus subspecies fetus has been recognized as a cause of systemic illness in immunocompromised hosts, including relapsing bacteremia in human immunodeficiency virus (HIV)-infected patients. Acquired resistance to quinolone therapy, while reported for a variety of bacteria, including Campylobacter jejuni, has not been previously documented for C. fetus. Two cases of quinolone-resistant C. fetus bacteremia were detected in HIV-infected patients. Cloning and nucleotide sequencing of the C. fetus gyrA gene in the 2 resistant isolates demonstrated a G-to-T change that led to an Asp-to-Tyr amino acid substitution at a critical residue frequently associated with quinolone resistance. In addition, comparison of the pre- and posttreatment isolates from 1 patient documented outer membrane protein changes temporally linked with the development of resistance. Relapsing C. fetus infections in quinolone-treated HIV-infected patients may be associated with the acquisition of resistance to these agents, and this resistance may be multifactorial.
AIDS-Related Opportunistic Infections↗
Evaluation of the Dade MicroScan MICroSTREP antimicrobial susceptibility testing panel with selected Streptococcus pneumoniae challenge strains and recent clinical isolates.
The MicroScan MICroSTREP panel is a recently marketed frozen broth microdilution panel for susceptibility testing of various streptococci, including Streptococcus pneumoniae. The panel contains 10 antimicrobial agents in cation-adjusted Mueller-Hinton broth supplemented with 3% lysed horse blood, similar in concept to the National Committee for Clinical Laboratory Standards (NCCLS) reference broth microdilution method for testing streptococci. A group of 210 isolates of S. pneumoniae were selected to include isolates with previously documented resistance to agents incorporated in the MICroSTREP panel, as well as recent invasive clinical isolates. All isolates were tested simultaneously with the MICroSTREP panel and an NCCLS reference panel whose drug concentrations were prepared to coincide with those of the MICroSTREP panel. Of the 210 isolates, 5 failed to grow in the MICroSTREP panel; 3 of those also did not grow in the reference panel. Essential agreement of MICs determined by the two methods (test MIC +/- one dilution of the reference MIC) was 99.6% overall and ranged from 98.0% with chloramphenicol to 100% with penicillin, ceftriaxone, erythromycin, tetracycline, and vancomycin. There were no very major or major interpretive category errors resulting from the MICroSTREP panel tests. Minor interpretive category errors ranged from 12.2% with cefotaxime and 9.8% with ceftriaxone (due mainly to clustering of MICs for the selected strains near the breakpoints) to 0% with chloramphenicol and vancomycin. These results indicate that the MicroScan MICroSTREP frozen panels provide susceptibility results with pneumococci that are essentially equivalent to results derived by the NCCLS reference broth microdilution procedure.
Anti-Bacterial Agents↗
Reproducibility of broth microdilution and disk diffusion susceptibility tests of nine antimicrobial agents against Streptococcus pneumoniae ATCC 49619.
Collaborative studies documented the reproducibility of broth microdilution susceptibility tests of Streptococcus pneumoniae against nine antimicrobial agents and of disk diffusion tests with six of those drugs. Replicate tests of Streptococcus pneumoniae ATCC 49619 in five different laboratories led to the following provisional quality control limits: cefdinir--0.03 to 0.25 microgram/ml and 26 to 31 mm; cefetamet--0.5 to 2 micrograms/ml and 20 to 25 mm; ciprofloxacin--0.25 to 1 microgram/ml and 20 to 26 mm; clinafloxacin--0.03 to 0.125 microgram/ml and 28 to 34 mm; grepafloxacin--0.06 to 0.5 microgram/ml and 21 to 28 mm; PD131628--0.125 to 0.5 microgram/ml and 24 to 29 mm; clindamycin--0.03 to 0.12 microgram/ml; cefpodoxime--0.03 to 0.12 microgram/ml; and trospectomycin--1 to 4 microgram/ml (disk tests were not evaluate for the latter three drugs).
Anti-Bacterial Agents↗
Laboratory issues in the detection and reporting of antibacterial resistance.
The emergence of antimicrobial resistance among several common bacterial pathogens requires that clinical microbiology laboratories have the ability to promptly and accurately recognize resistance in patients' isolates. Laboratories have several options for performing routine susceptibility testing, including the broth microdilution procedure (with or without instrumentation for test reading), automated instrument systems that provide rapid results, antibiotic gradient diffusion, and disk diffusion procedures. In addition, there are definitive screening tests capable of recognizing resistance to drugs of choice among several common bacterial species based on single drug concentration tests or rapid spot tests. The likely emergence of still newer resistance mechanisms will provide a challenge to clinical microbiologists to devise accurate, yet cost-effective strategies for use in the future.
Anti-Bacterial Agents↗
Activity of quinupristin/dalfopristin and its components against Haemophilus influenzae.
Quinupristin/dalfopristin is an injectable streptogramin antibiotic that is constituted in a 30:70 (w/w) ratio of the two components. Quinupristin and dalfopristin are thought to act synergically by binding to two separate sites on the bacterial 50S ribosomal subunit. The in-vitro activities of the two components separately and together in different ratios were determined for a collection of 100 Haemophilus influenzae strains representing various antimicrobial resistance phenotypes. The NCCLS microdilution susceptibility testing procedure incorporating Haemophilus test medium (HTM) broth was used to determine MICs of quinupristin, dalfopristin and seven other antimicrobial agents. The MIC50 and MIC90 values were 4 and 8, 4 and 16, and 64 and 128 mg/L for quinupristin/dalfopristin (30:70), dalfopristin and quinupristin, respectively. MICs of quinupristin and dalfopristin were also determined in Mueller-Hinton lysed horse blood broth and by HTM agar dilution testing. Compared with HTM broth-derived results, the MICs of quinupristin/dalfopristin and its components were the same or one dilution higher in lysed horse blood and HTM agar incubated in air, and were equivalent or one dilution lower in HTM agar incubated in a CO2 atmosphere. The MICs of quinupristin and dalfopristin separately or together were directly proportional to erythromycin MICs, but were otherwise unaffected by any of the resistance mechanisms represented in these strains. MICs of quinupristin and dalfopristin combined in ratios of 10:90, 70:30 and 90:10 did not differ significantly from those of the 30:70 ratio. Thus, unlike the synergic activity noted against Gram-positive bacteria, the activity of quinupristin/dalfopristin against H. influenzae appears to be due almost entirely to the dalfopristin component of the combination.
Ampicillin↗
Reappraisal of the antimicrobial susceptibilities of Chryseobacterium and Flavobacterium species and methods for reliable susceptibility testing.
Several Flavobacterium species, comprising a heterogeneous group of gram-negative bacilli that are capable of causing opportunistic infections in humans, have recently been reclassified as Chryseobacterium or Myroides species. Intrinsically resistant to a number of antibiotics, these organisms have been reported to be susceptible to vancomycin and certain other drugs that are normally active against gram-positive bacteria. By using the National Committee for Clinical Laboratory Standards (NCCLS) broth microdilution procedure, 58 clinical isolates of former flavobacteria (36 Chryseobacterium meningosepticum isolates, 11 C. indologenes isolates, 3 C. gleum isolates, 4 unspeciated former members of Flavobacterium group IIb, and 4 Myroides odoratum isolates) were tested with 23 antibiotics, including conventional and investigational agents. In addition, the broth microdilution results were compared to those generated by agar dilution, E-test, and disk diffusion for vancomycin and piperacillin-tazobactam. Compared to the NCCLS microdilution results, there were 7.1 and 17.9% very major errors with piperacillin-tazobactam by agar dilution and E-test, respectively. In addition, there were from 12.1 to 48.3% minor errors with both procedures with vancomycin and piperacillin-tazobactam. The very major and minor error rates were unacceptably high with disk testing of piperacillin-tazobactam; the use of enterococcal vancomycin disk breakpoints (zone diameter of > or =17 mm = susceptible) resulted in >20% minor errors but only one very major error. All of the isolates were susceptible to minocycline; over 90% were susceptible to sparfloxacin, levofloxacin, and clinafloxacin; and 88% were susceptible to rifampin. None was susceptible to vancomycin. When Chryseobacterium or Myroides species are isolated from serious infections, susceptibility testing by broth microdilution should be performed and therapy should be guided by those results.
Anti-Bacterial Agents↗
In vitro activities of the oxazolidinone antibiotics U-100592 and U-100766 against Staphylococcus aureus and coagulase-negative Staphylococcus species.
U-100592 and U-100766 are closely related antibiotics of the oxazolidinone class. Their in vitro activities were determined against 100 isolates of Staphylococcus aureus and 100 isolates of coagulase-negative Staphylococcus species by broth and agar dilution test methods. The MICs of both compounds by either test method at which 50 and 90% of isolates are inhibited were 2 and 4 micrograms/ml, respectively, for S. aureus and 1 to 2 micrograms/ml for coagulase-negative staphylococci. Time-kill assay with selected strains indicated a primarily bacteriostatic effect against staphylococci.
Acetamides↗
In vitro susceptibility testing of Borrelia burgdorferi by a dialysis culture method.
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Borrelia burgdorferi Group↗
Controlled clinical laboratory comparison of BACTEC plus aerobic/F resin medium with BacT/Alert aerobic FAN medium for detection of bacteremia and fungemia.
Blood specimens collected from adult patients with suspected sepsis in four medical centers were inoculated into BACTEC Plus/F and BacT/Alert FAN aerobic culture bottles. Both bottles of 7,401 bottle pairs contained the prescribed blood volume of 8 to 12 ml. Bottles were incubated in their respective instruments for a standard 7-day protocol or until the instruments signaled that they were positive. A total of 720 isolates that were judged to represent true infections were recovered from 338 patients; 451 isolates were recovered from both bottles, 143 were recovered from only the Plus/F bottle, and 126 were recovered from only the FAN bottle (P was not significant). Although more Histoplasma capsulatum isolates were recovered from Plus/F bottles (P < 0.005), there were no other statistically significant differences in recovery rates of individual species or groups of organisms between the two systems. Of 329 monomicrobic patient septic episodes, 244 episodes were detected by both blood culture systems, 40 were detected only by the BACTEC system, and 45 were detected only by the BacT/Alert system (P was not significant). There was no significant difference between the two systems in the detection of septic episodes among patients receiving antibiotic therapy at the time of blood cultures. Of the cultures found to be positive within the first 72 h of incubation, detection was on average earlier by the BACTEC system (16.9 h) than by the BacT/Alert system (18.7 h). Larger differences in average time to detection were seen with streptococci (10.7 h by the BACTEC system and 17.9 h by the BacT/Alert system) and yeasts (an average of 29.4 h by the BacT/Alert system versus 37.2 h by the BACTEC system). With the exception of the differences noted above, BACTEC Plus/F aerobic resin and BacT/Alert aerobic FAN blood culture bottles were comparable in their abilities to recover aerobic and facultative organisms.
Adult↗
Development of an antimicrobial susceptibility testing method suitable for performance during space flight.
Very little is known regarding the effects of the microgravity environment of space flight upon the action of antimicrobial agents on bacterial pathogens. This study was undertaken to develop a simple method for conducting antibacterial susceptibility tests during a space shuttle mission. Specially prepared susceptibility test research cards (bioMérieux Vitek, Hazelwood, Mo.) were designed to include 6 to 11 serial twofold dilutions of 14 antimicrobial agents, including penicillins, cephalosporins, a beta-lactamase inhibitor, vancomycin, erythromycin, tetracycline, gentamicin, ciprofloxacin, and trimethoprim-sulfamethoxazole. MICs of the drugs were determined by visual reading of color end points in the Vitek research cards made possible by incorporation of a colorimetric growth indicator (alamarBlue; Accumed International, Westlake, Ohio). This study has demonstrated reproducible susceptibility results in the testing of isolates of Staphylococcus aureus, group A Streptococcus species, Enterococcus faecalis, Escherichia coli (beta-lactamase-positive and -negative strains), Klebsiella pneumoniae, Enterobacter cloacae, and Pseudomonas aeruginosa. In some instances, the MICs were comparable to those determined by a standard broth microdilution method, while in some cases the unique test media and format yielded slightly different values that were themselves reproducible. The proposed in-flight experiment will include inoculation of the Vitek cards on the ground prior to launch of the space shuttle, storage of inoculated cards at refrigeration temperature aboard the space shuttle until experiment initiation, and then incubation of the cards for 18 to 48 h prior to visual interpretation of MICs by the mission's astronauts. Ground-based studies have shown reproducible MICs following storage of inoculated cards for 7 days at 4 to 8 degrees C to accommodate the mission's time schedule and the astronaut's activities. For comparison, ground-based control (normal gravity) MIC values will be generated by simultaneous inoculation and incubation of a second set of test cards in a laboratory at the launch site. This procedure can provide for a safe and compact experiment that should yield new information on the effects of microgravity on the biological activities of various classes of antibiotics.
Aerospace Medicine↗
Serogroup-specific epidemiology of Streptococcus pneumoniae: associations with age, sex, and geography in 7,000 episodes of invasive disease.
A study sample of 7,010 episodes of invasive Streptococcus pneumoniae disease was obtained by combining 13 existing datasets. Disease episodes due to each of 12 pneumococcal serogroups (1, 3-9, 14, 18, 19, and 23) were then compared with episodes in a constant internal control group to describe serogroup-specific variations in disease frequency by age, sex, and geographic origin. The results are presented as odds ratios (with 95% confidence intervals) derived by logistic regression, with adjustment for the major confounders, including dataset of origin. Variation in the male:female ratios between serogroups is small, suggesting that capsular characteristics are an unlikely explanation for the male preference of S. pneumoniae. Serogroups associated with higher nasopharyngeal prevalence (e.g., 19 and 24) are relatively more common in Europe and North American, while the invasive serotypes 1 and 5 are much more common in South America. The custom of reporting serogroup frequencies in two age groups, children and adults, conceals much of the variation in the age distributions across the whole span of life. The reduction of risk associated with serogroups 6, 14, 18, 19, and 23 beyond childhood follows different gradients, being most abrupt in serogroups 14 and most gradual in serogroup 18. The relative risk of disease with serotype 1 declines steadily throughout life, while with serotypes 3 and 8 it increases over middle age. Serogroups 7 and 23 are found unusually frequently in the third decade of life. Because of the wide differences in the epidemiology of individual serogroups of S. pneumoniae, it is questionable whether pneumococcal infection should continue to be classified as a single disease entity.
Adolescent↗
In-vitro susceptibility of Streptococcus pneumoniae to the d- and l-isomers of ofloxacin: interpretive criteria and quality control limits.
The l-isomer of ofloxacin (levofloxacin) was twice as active as ofloxacin and the d-isomer was inactive against 654 Streptococcus pneumoniae isolates. For disc susceptibility tests of pneumococci, 5 micrograms levofloxacin discs can be used with interpretive criteria of < or = 12 mm for resistant (MIC > or = 8.0 mg/L) and > or = 16 mm for susceptible (MIC < or = 2.0 mg/L). For quality control of levofloxacin tests, S. pneumoniae ATCC 49619 should give zones 20-25 mm in diameter and MICs 0.5-2.0 mg/L.
Anti-Infective Agents↗
In vivo activities of ceftriaxone and vancomycin against Borrelia spp. in the mouse brain and other sites.
Borrelia burgdorferi, the agent of Lyme disease, and B. turicatae, a neurotropic agent of relapsing fever, are susceptible to vancomycin in vitro, with an MIC of 0.5 microgram/ml. To determine the activity of vancomycin in vivo, particularly in the brain, we infected adult immunocompetent BALB/c and immunodeficient CB-17 scid mice with B. burgdorferi or B. turicatae. The mice were then treated with vancomycin, ceftriaxone as a positive control, or normal saline as a negative control. The effectiveness of treatment was assessed by cultures of blood and brain and other tissues. Ceftriaxone at a dose of 25 mg/kg of body weight administered every 12 h for 7 to 10 days eliminated cultivable B. burgdorferi or B. turicatae from all BALB/c or scid mice in the study. Vancomycin at 30 mg/kg administered every 12 h was effective in eliminating infection from immunodeficient mice if treatment was started within 3 days of the onset of infection. If treatment with vancomycin was delayed for 7 days or more, vancomycin failed to eradicate infection with B. burgdorferi or B. turicatae from immunodeficient mice. The failure of vancomycin in eradicating established infections in immunodeficient mice was associated with the persistence of viable spirochetes in the brain during antibiotic treatment.
Animals↗
Development of interpretive criteria and quality control limits for macrolide and clindamycin susceptibility testing of Streptococcus pneumoniae.
A six-laboratory collaborative study was conducted to develop MIC and zone diameter quality control limits and interpretive criteria for antimicrobial susceptibility testing of Streptococcus pneumoniae with azithromycin, clarithromycin, dirithromycin, and clindamycin. The MICs of all of the agents plus erythromycin for 302 clinical isolates of pneumococci that had been selected with an emphasis on resistant strains were determined by use of the National Committee for Clinical Laboratory Standards (NCCLS)-recommended broth microdilution procedure. The zone diameters of the isolates were also determined for the same agents except erythromycin by the NCCLS disk diffusion test procedure. Repeated testing of S. pneumoniae ATCC 49619 with different sources and lots of media and disks allowed development of MIC and zone diameter quality control ranges for these agents. Interpretive criteria for the MIC of azithromycin were established and were as follows: susceptible, < or = 0.5 microgram/ml; intermediate, 1 microgram/ml; and resistant, > or = 2 micrograms/ml. The interpretive criteria advocated for the MICs of clarithromycin and clindamycin were as follows: susceptible, < or = 0.25 microgram/ml; intermediate, 0.5 microgram/ml; and resistant, > or = 1 microgram/ml. Comparison of MICs and disk diffusion zone diameters led to the development of interpretive criteria for the zone diameters for azithromycin, clarithromycin, and clindamycin that correlated well with these MIC breakpoints. Testing of this organism collection also led to the reestablishment of the erythromycin MIC breakpoints as being identical to those of clarithromycin, which resulted in equivalent cross-susceptibility and cross-resistance for the three macrolides that are currently marketed in the United States. Thus, the susceptibility of pneumococci to azithromycin and clarithromycin can be predicted accurately by testing only erythromycin in clinical laboratories. This recommendation, as well as the interpretive and quality control criteria that are described, have been accepted by NCCLS and are included in the latest NCCLS susceptibility testing guidelines.
Anti-Bacterial Agents↗
Comparison of inoculation methods for testing enterococci by using vancomycin screening agar.
One hundred four recent clinical isolates of Enterococcus species were screened for vancomycin resistance by using inocula of 10(5) or 10(6) CFU dispensed by pipet and by use of a cotton swab dipped in a 0.5 McFarland standard organism suspension applied to the surface of brain heart infusion agar containing 6 micrograms of vancomycin per ml. The three inoculation methods were equivalent in the detection of nonsusceptible isolates. The use of swab inoculation was convenient and less costly than the use of micropipets.
Agar↗
Tentative criteria for determining the in vitro susceptibilities of Haemophilus influenzae, including quality control parameters, to two fluoroquinolones (grepafloxacin and PD 131628).
Grepafloxacin (OPC 17116) and PD 131628 were evaluated against 150 Haemophilus influenzae isolates to propose susceptibility testing criteria for both broth dilution and disk diffusion procedures using haemophilus test medium. Grepafloxacin-susceptible isolates are defined as those for which minimum inhibitory concentrations (MICs) are < or = 0.06 micrograms/ml and zones of inhibition are > or = 27 mm. PD 131628-susceptible strains of H. influenzae included those that exhibited MICs < or = 0.03 micrograms/ml, and the zones were > or = 32 mm. All MICs were well below concentrations that can be achieved in the blood and tissues of patients treated with either study drug. Criteria for defining a resistant category cannot be determined until strains with elevated MICs are available for study. The proposed quality control guidelines for H. influenzae ATCC 49247 and the disk test are 32-39 mm in diameter for grepafloxacin and 34-42 mm for PD 131628. The preliminary broth microdilution MIC control limits for this strain are 0.002-0.016 micrograms/ml for grepafloxacin and 0.002-0.008 micrograms/ml for PD 131628.
Anti-Infective Agents↗
Clinical and molecular epidemiology of vancomycin-resistant Enterococcus faecium during its emergence in a city in southern Texas.
During a 19-month period from April 1993 to October 1994, 41 isolates of vancomycin-resistant Enterococcus faecium (VREF) were detected in seven different hospitals in a city in southern Texas. A case-control study to determine the risk factors for acquisition was done in the hospital in which the majority of isolates were detected. Pulsed-field gel electrophoresis (PFGE) of whole-cell DNA was used to determine strain identity. Thirty-five (85%) of the 41 VREF isolates were of the vanB phenotype. Of these, 32 (91%) of 35 were the same strain by PFGE typing. The same vanB strain was documented in five different hospitals in the city. In contrast, 4 (67%) of 6 of the vanA phenotype VREF isolates were distinct strains by PFGE typing. Significant risk factors for colonization or infection with VREF were prior exposure to antibiotics (P = .04), the previous use of third-generation cephalosporins (P = .03), and the previous use of parenteral vancomycin (P = .002). Infection-control and antibiotic-utilization measures were implemented to control cross-transmission and selection of VREF isolates. During the emergence of VREF in our city, clonal dissemination of a single strain of vanB VREF among six hospitals was documented. Limited cross-transmission of vanA phenotype VREF isolates occurred, but most vanA VREF isolates were distinct strains selected in individual hospital environments.
Anti-Bacterial Agents↗