To screen test bacterial isolates for the production of ESBL enzymes, single antibiotic disks containing 30 µg of cefotaxime and 30 µg of ceftazidime should be used. In practice, any of the 3rd generation cephalosporins can be used to screen for ESBL production in bacteria isolates.
The antibiotics discs should be placed aseptically at a distance of about 20mm – 30mm apart on Mueller-Hinton (MH) agar (Oxoid, UK) plate(s) that has been previously inoculated with standardized inoculums(s) of the test bacterium using a sterile swab stick in order to get a confluent growth. Isolates showing inhibition zone diameter (IZD) of ≤ 23 mm should be suspected to produce ESBL.
The MH agar plates should be allowed for about 30 mins for pre-diffusion of the antibiotics. After this, the MH agar plates should be incubated for 18-24 hrs at 37oC. After the incubation time, the zones of inhibition should be measured in millimeter using a meter rule.
ESBL production should be suspected or inferred if any of the test bacteria showed reduced susceptibility or is resistant to any one of the third generation cephalosporins (e.g., cefotaxime 30µg and ceftazidime 30µg) used for the ESBL screening studies.
BRIEF DESCRIPTION OF ESBL
ESBLs (extended spectrum beta-lactamases) are plasmid – mediated beta – lactamases capable of hydrolyzing many beta – lactam antibiotics including third – generation cephalosporins and monobactams, but are yet inhibited by clavulanic acid, a beta – lactamase inhibitor. Since the discovery of the ESBLs in the 1980’s, over 100 different enzymes have been described, and they have become a worldwide health problem affecting many countries of the world with varying prevalence.
ESBLs arise by mutations in genes for common plasmid – mediated beta – lactamases (especially TEM and SHV enzymes) that alter the amino acid configuration of the enzyme near its active site to increase the affinity and hydrolytic ability of the beta – lactamases for oxyimino cephalosporins. ESBLs are detected mostly in Enterobacteriaceae especially Klebsiella pneumoniae, Escherichia coli, and Klebsiella oxytoca. They often contain resistance determinants for other classes of antibiotics like the aminoglycosides, sulfonamides, fluoroquinolones which are readily transmissible from one strain of organism to another and between different species of Gram – negative bacteria.
Apart from the TEM- and SHV- type ESBLs, other classes of ESBLs have also been described including CTX – M type ESBLs, PER- and OXA- type ESBLs, mutations that expand the spectrum of the earlier beta – lactamases (TEM and SHV) usually increase the sensitivity of ESBLs to beta – lactamase inhibitors like clavulanic acid, tazobactam and sulbactam. ESBL – producing organisms are not new, having first been reported in the 1980’s from Germany, and they are now found worldwide in both the community hospital settings. Resistance of Gram – negative bacteria to third – and fourth – generation cephalosporins is mediated by ESBLs, and they are an important cause of treatment failure in patients receiving cephalosporins because these agents are hydrolyzed in vivo by ESBLs when used for therapy especially in patients harbouring ESBL – producing bacteria.
Peter C. Ejikeugwu, Nkechukwu M.I., Ugwu C.M., Iroha I.R and Esimone C.O (2012). Extended-spectrum β-lactamase-producing Escherichia coli isolates from suspected community acquired urinary tract infections. European Journal of Scientific Research. 84(4):565-571.