Emergence of CTX-M group 1 ESBL–producing Klebsiella pneumoniae strains in the community

Extended-spectrum β-lactamase (ESBL)-producing Gram negative bacteria are an emergent cause of urinary tract infections in nonhospitalized patients in different countries. Studies on ESBLs in the outpatient populations reported CTX-M ESBLs to be the most prevalent. The aim of this study was the molecular characterization of ESBL-producing K. pneumoniae strains isolated from urine of outpatients in Zagreb region during the last five years. During the five-year study period a total of 2, 651 K. pneumoniae strains were isolated from urine of nonhospitalized patients with significant bacteriuria. ESBL production was detected by double-disk diffusion technique and by >3-dilution reduction in the minimal inhibitory concentration of ceftazidime in the presence of clavulanate. A total of 441 ESBL-producing K. pneumoniae strains (15.5%) were collected and 25 strains were characterised. Double-disk synergy test was used to detect ESBLs. Minimum inihibitory concentrations (MICs) were determined by broth microdilution method according to CLSI. The transferability of cefotaxime resistance was tested by conjugation (broth mating method). PCR was used to detect alleles encoding ESBL enzymes. The genotypes of the strains were compared by analysis of banding patterns generated by pusled-field gel electrophoresis (PFGE) of Xba I-digested genomic DNA. A significant difference in frequencies of ESBL isolates was observed. In the first year of study only 4.9% of isolated strains were ESBL producers, while in the second year 17.3% ESBL strains were detected (p<0.01), and the frequency remain stabile within following years. Out of 17 characterized strains all strains yielded an amplicon with primers specific for SHV β-lactamases and CTX-M β-lactamases. Based on sequencing of blaCTX-M genes enzymes of nine strains were identified as CTX-M 15 β –lactamase and three as CTX-M-14. Isolates were not clonally related. The study demonstrated community-associated emergence of CTX-M 1 β-lactamase–producing K. pneumoniae strains.