Disc diffusion-based screening tests for extended-spectrum β-lactamases in Haemophilus influenzae

Abstract

To establish a suitable screening test for the detection of extended-spectrum beta-lactamases (ESBLs) in Haemophilus influenzae. Double disc diffusion tests using cephalosporin discs adjacent to an amoxicillin/clavulanate 3 microg disc, prediffusion tests, where an amoxicillin/clavulanate 3 microg disc was prediffused before being replaced with a cephalosporin disc, and combination cephalosporin/clavulanate discs were used to screen recombinant H. influenzae expressing cloned ESBLs. Methods initially determined to be suitable were validated by testing a range of clinical strains. Enhanced zones around cephalosporin discs due to synergy with an adjacent amoxicillin/clavulanate disc were not reliably demonstrated with the recombinant H. influenzae strains expressing ESBLs. Cefotaxime 5 microg or 30 microg and cefpodoxime 10 microg discs prediffused with an amoxicillin/clavulanate 3 microg disc, and cefotaxime 30 microg/clavulanate 10 microg and cefpodoxime 10 microg/clavulanate 1 microg combination discs all gave zone sizes > or = 5 mm greater than the corresponding plain disc for all recombinant ESBL-producing strains. None of the clinical strains gave enhanced zones. Cefotaxime 5 or 30 microg or cefpodoxime 10 microg discs prediffused with an amoxicillin/clavulanate 3 microg disc, or cefotaxime 30 microg/clavulanate 10 microg and cefpodoxime 10 microg/clavulanate 1 mug combination discs are all suitable for screening H. influenzae isolates for the presence of ESBLs. ESBL-producing strains should show an increase in zone diameter of > or = 5 mm compared with the corresponding plain cephalosporin disc.