Characterization of Clinical Isolates of Enterobacteriaceae from Italy by the BD Phoenix Extended-Spectrum β-Lactamase Detection Method

Abstract

Production of extended-spectrum β-lactamases (ESBLs) is an important mechanism of β-lactam resistance in Enterobacteriaceae . Identification of ESBLs based on phenotypic tests is the strategy most commonly used in clinical microbiology laboratories. The Phoenix ESBL test (BD Diagnostic Systems, Sparks, Md.) is a recently developed automated system for detection of ESBL-producing gram-negative bacteria. An algorithm based on phenotypic responses to a panel of cephalosporins (ceftazidime plus clavulanic acid, ceftazidime, cefotaxime plus clavulanic acid, cefpodoxime, and ceftriaxone plus clavulanic acid) was used to test 510 clinical isolates of Escherichia coli , Klebsiella pneumoniae , Klebsiella oxytoca , Proteus mirabilis , Providencia stuartii , Morganella morganii , Enterobacter aerogenes , Enterobacter cloacae , Serratia marcescens , Citrobacter freundii , and Citrobacter koseri . Of these isolates, 319 were identified as ESBL producers, and the remaining 191 were identified as non-ESBL producers based on the results of current phenotypic tests. Combined use of isoelectric focusing, PCR, and/or DNA sequencing demonstrated that 288 isolates possessed bla _TEM-1 - and/or bla _SHV-1 -derived genes, and 28 had a bla _CTX-M gene. Among the 191 non-ESBL-producing isolates, 77 isolates produced an AmpC-type enzyme, 110 isolates possessed TEM-1, TEM-2, or SHV-1 β-lactamases, and the remaining four isolates (all K. oxytoca strains) hyperproduced K1 chromosomal β-lactamase. The Phoenix ESBL test system gave positive results for all the 319 ESBL-producing isolates and also for two of the four K1-hyperproducing isolates of K. oxytoca . Compared with the phenotypic tests and molecular analyses, the Phoenix system displayed 100% sensitivity and 98.9% specificity. These findings suggest that the Phoenix ESBL test can be a rapid and reliable method for laboratory detection of ESBL resistance in gram-negative bacteria.