engleski

First report of extended-spectrum beta-lactamases from Bosnia and Herzegovina

Background: The aim of this study was to characterize ESBLs in outpatient clinical isolates, at the first time in Bosnia and Herzegovina. Methods: Fifteen K. pneumoniae strains and 5 E. coli strains were isolated from urine obtained from outpatients with confirmed urinary tract infections. ESBLs were detected by double-disk synergy test and by 8 fold reduction in ceftazidime MIC in the presence of clavulanate. Transferability of resistance to oxymino cephalosporins was tested by conjugation (broth mating method) employing E. coli A15R- stain resistant to rifampicin as recipient. Frequency of transconjugation was expressed relatively to the number of donor cells. Beta-lactamases were characterized by isoelectric focusing on polyacrilamide gel. BlaESBL genes were detected by PCR with primers specific for TEM, SHV and CTX-M beta-lactamases. Plasmids encoding ESBLs were extracted by alkalyne lysis procedure and analysed after electrophoresis in agarose gel. Molecular relatedness between the strains was investigated by PFGE. Results: Ten out 15 K. pneumoniae and 3 out of 5 E. coli strains transferred resistance to E. coli recipient. The frequency of transconjugation ranged from 10-2 to 10-5. All K. pneumoniae strains and their E. coli transconjugants produced a beta-lactamase with the pI of 8.2 typical for SHV-5 beta-lactamase. Donor strains had an additional band with the pI of 7.6 which is a chromosomally encoded SHV-1 in K. pneumoniae. E. coli strains and their respective transconjugants displayed a band with the pI of 5.4 to 6 typical for TEM beta-lactamases. PCR detected blaSHV genes in all K. pneumoniae strains and their transconjugant and blaTEM genes in all E. coli strains. No CTX-M beta-lactamase producers were found. ESBL genes were encoded on the large plasmids of 100 to 150 kb. Conclusions: Not only hospitals serve as ESBL producers reservoir today. SHV-ESBLs awere dominant among K. pneumoniae strains, whereas TEM-ESBL were most prevalent in E. coli isolates contrary to the results obtained by other authors who found CTX-M beta-lactamases to be the most prevalent in outpatient setting. According to isoelectric focusing, SHV-5 beta-lactamases or its derivatives found in K. pneumoniae strains, which are already widespread in middle and east Europe. Further analysis of beta-lactamases by sequencing of ESBL genes would be necessary in order to determine the exact enzyme type.

Extended-spectrum beta-lactamases; Escherichia coli; Klebsiella pneumoniae

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