Journal Article

Phenotypic detection of extended-spectrum and AmpC β-lactamases by a new spot-inoculation method and modified three-dimensional extract test: comparison with the conventional three-dimensional extract test

M. Shahid, Abida Malik, Mithlesh Agrawal and Sanjay Singhal

in Journal of Antimicrobial Chemotherapy

Published on behalf of British Society for Antimicrobial Chemotherapy

Volume 54, issue 3, pages 684-687
Published in print September 2004 | ISSN: 0305-7453
Published online September 2004 | e-ISSN: 1460-2091 | DOI: http://dx.doi.org/10.1093/jac/dkh389
Phenotypic detection of extended-spectrum and AmpC β-lactamases by a new spot-inoculation method and modified three-dimensional extract test: comparison with the conventional three-dimensional extract test

Show Summary Details

Preview

Objectives: To develop an easy, rapid and reproducible spot-inoculation method for phenotypic detection of extended-spectrum β-lactamases (ESBLs) and AmpC β-lactamases and to make the existing three-dimensional extract test more convenient for use in routine diagnostic laboratories.

Methods: ESBL and AmpC producing and non-producing isolates of Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa, as identified by the conventional three-dimensional extract test, were used to evaluate the modified procedures. Whole bacterial cells and freeze–thaw preparations, as β-lactamase sources, were strategically applied to culture plates near ceftazidime and cefoxitin discs on a lawn inoculum of E. coli ATCC 25922. Technical variations of the test included placing the β-lactamase-containing inoculum into slits, wells and trenches, or onto the surface as spots at varying distances from the discs, and adding clavulanate or cloxacillin to the three-dimensional inoculum to confirm the presence of ESBLs and AmpC β-lactamases, respectively.

Results: All the methods adopted for ESBL and AmpC detection by using the whole bacterial cells gave positive results. However, the best results were given by the spot-inoculation method. In modifications using the enzymic extracts, the enhanced growth of surface organisms was better appreciated in the designed modifications compared with the conventional methods.

Conclusions: The method described here is simple and cost-effective. Furthermore, up to 16 isolates may be tested on a single culture plate, thus it is a less labour-intensive and more economic technique than other reported phenotypic methods.

Keywords: direct-spot test; well-extract test; modified enzymic extraction; modifications; user-friendly method

Journal Article.  3017 words.  Illustrated.

Subjects: Medical Oncology ; Critical Care

Full text: subscription required

How to subscribe Recommend to my Librarian

Users without a subscription are not able to see the full content. Please, subscribe or login to access all content.