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Medycyna Doświadczalna i Mikrobiologia 2017, 69(2): 115-126

Współwystępowanie plazmidowo kodowanych mechanizmów oporności: DHA-7, CTX-M-15, qnrB6 i aac(6’)-Ib-cr u pałeczek Klebsiella pneumoniae wytwarzających nabyte AmpC
[Co-existence of plasmid-mediated resistance: DHA-7, CTX-M-15, qnrB6 and aac(6’)-Ib-cr among acquired AmpC-producing Klebsiella pneumoniae]

Rzeczkowska Magdalena, Piekarska Katarzyna, Wołkowicz Tomasz, Semczuk Katarzyna, Kamińska Teresa, Gierczyński Rafał

STRESZCZENIE

Gram-ujemne pałeczki Klebsiella pneumoniae są jednym z dominujących etiologicznych czynników zakażeń szpitalnych. Często napotykanym problemem w terapii zakażeń wywoływanych przez te pałeczki jest ich narastająca oporność, w tym na antybiotyki ß- laktamowe i fluorochinolony. Celem pracy była ocena współwystępowania wybranych, kodowanych plazmidowo mechanizmów oporności na antybiotyki β-laktamowe i fluorochinolony u pałeczek K. pneumoniae wytwarzających nabyte cefalosporynazy typu AmpC. Przeprowadzone badania wskazują na zdolność szpitalnych szczepów pałeczek K. pneumoniae do nabywania i akumulacji genów kodujących mechanizmy oporności tj. pAmpC, ESBL i PMQR. Fakt akumulowania PMQR, AmpC i ESBL stanowić może formę skutecznej odpowiedzi na presję selekcyjną wynikłą ze skojarzonego stosowania ß‑laktamów i fluorochinolonów w lecznictwie.

ABSTRACT

Introduction: Increasing antimicrobial resistance, especially towards third and fourth generation cephalosporins, aminoglycosides and fluoroquinolones have been reported last decade and poses serious therapeutic problems with treating K. pneumoniae infections in humans. Extended-spectrum β‑lactamases (ESBLs) constitute the predominant mechanism of acquired antibiotic resistance to β‑lactams. During the past few years, increasing occurrence of plasmid-mediated AmpC β-lactamases (pAmpCs) was increasingly reported. Moreover, plasmid-mediated quinolone-resistance (PMQR) has been reported in co-existence with pAmpCs and ESBLs. Therefore, the aim of our study was to analyse the diversity of ESBLs and plasmid-mediated PMQR among K. pneumoniae pAmpC-producing isolates.

Material and Methods: Twenty-two clinical isolates of K. pneumonia resistant to third-generation

cephalosporins were prospectively collected in 3 hospitals in the Warsaw city and suburbs, Poland during period of 3-months. AmpCs and ESBLs were detected by phenotypic methods: sensitivity to cefoxitin, disk potentiation test (DPT) and duble-disk synergy test (DDST). MICs of 8 antimicrobial agents were determined by E-test. pAmpC, ESBL, QNR and QepA genes were detected by PCR and DNA sequencing. The aac(6’)-Ib was detected by PCR, the presence of aac(6’)-Ib-cr gene variant was identified by digestion of the PCR product with BtsCI. PFGE with XbaI endonuclease was performed to investigate the clonality of the tested isolates.

Results: Five of 22 tested isolates of K. pneumoniae produced AmpC β-lactamases of DHA family. All the isolates were found to co-produce extended-spectrum β-lactamases (ESBLs) of CTX-M-15 family. Furthermore, 4 of the AmpC producing isolates were positive for PMQR determinants. Two of them carried sole aac(6‘)-Ib-cr variant gene while the remaining 2 isolates harbored qnrB6 in addition to the aac(6’)-Ib-cr variant. No clonality was found by XbaI-PFGE.

Conclusions: Accumulation of plasmid-mediated AmpC, ESBL and PMQR resistance traits by K. pneumoniae may be the ad hoc strategy to cope with the third generation beta-lactams and fluoroquinolones. Horizontal transfer seem to play the primary role in dissemination of these resistance traits among K. pneumoniae.

Key words: Klebsiella pneumoniae, plasmid-encoded AmpC cephalosporinases, pAmpC, ESBL, PMQR

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