Jul 7, 1986 - Thirty-five modifications of Abbott MS-2 growth medium were evaluated in an attempt to reduce the high frequency of false-susceptible errors ...
JOURNAL OF CLINICAL MICROBIOLOGY, Apr. 1987, p. 732-733
Vol. 25, No. 4
0095-1137/87/040732-02$02.00/0 Copyright © 1987, American Society for Microbiology
Attempted Reduction of Abbott MS-2 False-Susceptible Errors for Cefotaxime-Resistant Pseudomonas aeruginosa by Medium Modification D. W. LANDER,1* D. L. JUNGKIND,2 AND L. L. STONE' Crozer-Chester Medical Center, Chester, Pennsylvania 19013,' and Thomas Jefferson University Hospital, Philadelphia, Pennsylvania 191072 Received 7 July 1986/Accepted 12 December 1986
Thirty-five modifications of Abbott MS-2 growth medium were evaluated in an attempt to reduce the high frequency of false-susceptible errors produced by the MS-2 system when testing resistant Pseudomonas aeruginosa against cefotaxime. A hypotonic modification with 0.1% KNO3 reduced the errors from 41 to 20% for 23 resistant isolates. Ail hypertonic modifications proved unsatisfactory. No modification reduced false-susceptible errors to acceptable clinical levels.
The MS-2 system (Abbott Laboratories, North Chicago, Ill.) is an automated instrument intended for use in the clinical microbiology laboratory. By photometrically monitoring the turbidity changes resulting from bacterial multiplication, the MS-2 and similar instruments are used to determine antibacterial susceptibility. However, in the presence of at least some members of the beta-lactam group of antibacterial agents, resistant Pseudomonas aeruginosa isolates may not begin growth within the time allotted for the test cycle. Such organisms are incorrectly interpreted by the instrument as susceptible to these antimicrobial agents. Previously reported data (7) have shown that a high frequency of MS-2 false-susceptible results is likely to occur whenever resistant P. aeruginosa isolates are tested against cefotaxime and moxalactam. Similar results have also been reported with trimethoprim-sulfamethoxazole and carbenicillin (4), and unpublished data from our laboratory also indicate that piperacillin results can be similarly affected. For these reasons it has been suggested that the MS-2 system should not be used for the routine testing of clinical P. aeruginosa isolates for resistance to these problem antimicrobial agents. It has also been reported that a prolonged lag growth phase is the normal pattern when P. aeruginosa is incubated in the presence of beta-lactams (2, 3). It is a logical assumption that any reduction in the lag phase should reduce the frequency of false-susceptible results, and it was the purpose of this investigation to determine whether the growth rates of cefotaxime-resistant strains of P. aeruginosa could be enhanced via broth modifications. The first phase of this study involved modification of the MS-2 culture medium by the addition of various osmotic stabilizers, the intent being to provide optimum stabilization for bacilli with beta-lactam-induced cell wall damage. The three compounds chosen were sodium chloride in concentrations ranging from 0.025 to 0.6 M, magnesium chloride ranging from 0.01 to 0.10 M, and sucrose ranging from 0.05 to 0.5 M. Six different concentrations of each compound were used, for a total of 18 hypertonic modifications, spanning the ranges which have previously proven useful for osmotic stabilization (1, 5, 6).
Eight strains of P. aeruginosa with various cefotaxime susceptibilities were initially tested against the 18 modifications by manual, macroscopic growth rate determination. Of these isolates, two strains had MICs of 16 ,ug/ml, 2 strains had MICs of 32 ,ug/ml, 3 strains had MICs of 64 ,ug/ml, and 1 strain had an MIC of 256 ,ug/ml. Isolates with MICs of 16 to 32 ,ug/ml were considered moderately susceptible, and those with MICs 264 were considered resistant. The testing incorporated a subinhibitory level of cefotaxime, individually adjusted to one-fourth the MIC for each strain. These test tube cultures were visually compared with controls hourly for 12 h, with a final inspection after 24 h. One-half of these modifications failed to enhance growth rates relative to those of the controls and were eliminated. The nine media which appeared to enhance the growth rates of the test organisms were then reevaluated in the MS-2 system by using 10-well research cuvettes (Abbott). The three best concentrations of sodium chloride were 0.025, 0.05, and 0.10 M. For sucrose, the three best concentrations were 0.05, 0.10, and 0.20 M. For magnesium chloride, the three best concentrations were 0.01, 0.02, and 0.04 M. The mean time for growth detection for each organism-medium combination was accurately evaluated by comparison with the growth rate of the organism in an adjacent control well of plain MS-2 broth. None of the nine broths produced substantially superior growth rates compared with growth in plain broth. Testing was therefore expanded to include 27 different combinations of the nine concentrations listed above. Only one of these 36 hypertonic media-a combination of 0.2 M sucrose, 0.1 M sodium chloride, and 0.04 M magnesium chloride-appeared to be uniformly superior in promoting more rapid growth. This single hypertonic medium was further tested against a total of 42 strains of P. aeruginosa with MICs throughout the susceptible, intermediate, and resistant ranges. There was a slight reduction in the frequency of very major errors, from 22% (plain MS-2 broth) to 14%. However, the number of major errors increased from
