[Molecular characterization of ESBL-producing Klebsiella pneumoniae isolates from intensive care patients].
OBJECTIVE
The study aimed at the assessment of the prevalence of ESBL-positive isolates of Klebsiella pneumoniae in intensive care patients and their molecular biology analysis.


METHODS
Over a 5-month period, Klebsiella pneumoniae strains were isolated from patients hospitalized at the Department of Anaesthesiology and Resuscitation of the University Hospital in Olomouc. For each isolate, an antibiogram was performed by the standard microdilution method and the production of ESBL was determined by the modified double-disk synergy test. PCR was used to demonstrate the presence of the blaTEM and blaSHV genes. The isolates producing SHV- and TEM-types of beta-lactamases were typed using the restriction fragment length polymorphism (RFLP) method to identify the most common mutations responsible for the development of an ESBL phenotype. Similar or identical isolates were determined by pulsed-field gel electrophoresis (PFGE) of DNA fragments cleaved by the XbaI restriction endonuclease.


RESULTS
A total of 67 isolates of Klebsiella pneumoniae were obtained. In 13 of them, the production of ESBL was detected and the presence of the blaSHV gene was confirmed by PCR. Restriction cleavage by NheI revealed mutations at position 238 in all SHV-positive PCR products. The restriction analysis did not confirm the presence of the gene encoding TEM-type extended-spectrum beta-lactamase. Molecular biology typing by PFGE detected the presence of 11 different strains.


CONCLUSIONS
In the observed group of intensive care patients, the prevalence of ESBL-positive strains of Klebsiella pneumoniae reached 19.4 %. The analysis of SHV and TEM products of PCR by the RFLP method showed the prevalence of SHV-type ESBL. Overall, 84.6 % of the strains had unique restriction profiles. The results suggest both high levels of hygienic and epidemiological measures at the monitored department and rational antibiotic policy.
Chromá
Magdalena
M
Institute of Microbiology, Faculty of Medicine UP Olomouc, Czech Republic. magdalena.chroma@centrum.cz
Kolár
Milan
M
Marek
Oldrich
O
Koukalová
Dagmar
D
Sauer
Pavel
P
D003428:Cross Infection / Q000382:microbiology; D006801:Humans; D007362:Intensive Care Units; D007710:Klebsiella Infections / Q000382:microbiology; D007711:Klebsiella pneumoniae / Q000201:enzymology* / Q000235:genetics / Q000302:isolation & purification; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D018440:beta-Lactam Resistance; D001618:beta-Lactamases / Q000096:biosynthesis* / Q000235:genetics
D001618:beta-Lactamases