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Effect of Distance Between the Clavulanate-disk and b-lactam disks in the Double-disk Diffusion Method for the Detection of Extended-spectrum b-lactamase (ESBL) Production

 

Esperanza C. Cabrera

Biology Department
De La Salle University, Taft Avenue, Manila

corresponding author: This email address is being protected from spambots. You need JavaScript enabled to view it.

 

ABSTRACT

Different distances between the clavulanate disk and b-lactam antibiotic disks were tested with the double-disk diffusion method for the detection of extended- spectrum b -lactamase (ESBL) production. Enterobacteriaceae isolates that screened positive for ESBL production using the disk agar diffusion method, based on the criteria set by the National Committee for Clinical Laboratory Standards (NCCLS), were further tested using the phenotypic confirmatory double-disk diffusion assay. Aztreonam, ceftazidime, cefotaxime and ceftriaxone disks were placed at distances 14 mm, 20 mm and 24 mm between sides from the clavulanate disk. The distance of 14 mm was shown to be the most sensitive in detecting ESBL production, with 21 out of the 28 isolates that were positive in the screening test being positive in the confirmatory test. The distance of 20 mm detected only 13 of the 21 ESBL producers. All of these were also detected using the 14 mm distance. The distance of 24 mm failed to detect any ESBL producer with any of the antibiotics. Ceftriaxone detected 95% of the ESBL producers at a distance of 14 mm. This was followed by aztreonam at 86%, cefotaxime at 81% and ceftazidime at 62%. Conjugative plasmids of approximately 23 kb were detected among the ESBL-producing isolates and the transconjugants of those tested. Results suggest the use of 14 mm distance between sides of the clavulanate disk and b -lactam antibiotic disks for the ESBL confirmatory double disk method, with retest at 20 mm for results with overlapping zones. It also recommends the inclusion of aztreonam and ceftriaxone to supplement ceftazidime and cefotaxime in the phenotypic confirmatory test set by NCCLS.

 

INTRODUCTION

Extended-spectrum B-lactamases (ESBL) confer microbial resistance to extended-spectrum b-lactams: cephalosporins with the oxyimino side chain, such as ceftazidime, cefotaxime, and ceftriaxone; and monobactams with the oxyimino side chain, i.e., aztreonam. These extended-spectrum b-lactams . . . . .

 

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