A novel method for the detection of extended-spectrum b-lactamases, metallo-b-lactamases and AmpC producing Gram-negative bacilli in a single test: the CICA beta-test

Abstract number: 1733_7

M'Zali F., Arpin C., Dubois V., Andre C., Quentin C.

Objectives: Multidrug resistant Gram-negative bacilli are emerging pathogens. Infections caused by these organisms represent a clinical challenge. Currently, evidence related to infectious diseases with Gram-negative rods with extended spectrum b-lactamases (ESBLs) or metallo-b-lactamases (MBLs) or AmpC is accumulating. A rapid, sensitive and reliable method for their early detection is critical. The Cica beta-Test (Kanto Chemical, Japan; Mast, UK) is a new method which uses strips impregnated with a chromogenic cephalosporin (HMRZ-86) coupled with each of clavulanic acid for ESBL detection, boronic acid for AmpC detection and mercapto-acetic-acid and EDTA for detection of MBLs. The aim of this study was to evaluate the Cica beta-Test against a large panel of clinical strains known to harbour these enzymes singly or in combination.

Methods: A collection of 100 epidemiologically distinct clinical isolates from French hospitals and nursing homes were examined. The collection includes strains of the Enterobacteriaecae family, Pseudomonads and Acinetobacter spp. The clinical strains produce previously fully characterised b-lactamases of TEM, SHV, CTX-M, VEB, PER, CMY, GES, VIM, IMP and OXA families. All isolates were re-tested using the Cica beta-Test. The test consists on paper strips each specific for the detection of ESBLs, MBLs, AmpC and penicillinase producing organisms. Only ESBLs and MBLs are able to hydrolyze the b-lactam ring in HMRZ-86, resulting in a colour change from yellow to red. The test organism is grown overnight on Muller Hinton plate. One drop of HMRZ-86 is dispensed on each strip, then one to two colonies of the test organism is spread on the filter pad of the Cica-Beta-Test strip. The test is read after 2 to 15 min.

Results: All mechanisms of resistance in the clinical strains were correctly identified. Presence of combination of mechanisms was also picked up notably (CTX-M +AmpC), (VIM-2 + SHV-5), (VEB-1 +AmpC). In addition, the Cica beta-Test provides us with rapid information on the resistance mechanism(s) (maximum 15 min).

Conclusion: We describe here a technically simple method for the detection of ESBL, MBL and AmpC producing Gram-negative bacilli. The test proved of high sensitivity and specificity and provides useful information for targeted therapy. We recommend this test as a routine screening method for the detection and identification of mechanisms of resistance in Gram-negative bacteria.