engleski

The first detection of plasmid-mediated AmpC beta-lactamase in multidrug-resistant Proteus mirabilis isolates from University Hospital Centre Split, Croatia

Objectives: P mirabilis is one of the important causes of nosocomial infections. A progressive increase of β-lactam resistance has emerged in this species worldwide. Resistance due to plasmid-mediated AmpC enzymes is less common than extended-spectrum β-lactamase (ESBL) production, but harder to detect and contributes to a broader spectrum of β-lactam resistance. The aims of this study were to detect the presence of AmpC β-lactamase in multidrug-resistant P mirabilis strains isolated from inpatients at the University Hospital Centre Split (southern Croatia), identify the type of AmpC enzyme, and determine genetic relatedness of isolates. Methods: P mirabilis isolates were collected according to the antimicrobial susceptibility pattern which included a clear resistance to third-generation cephalosporins and cefoxitin, determined routinely by EUCAST disk diffusion method during a 6-month survey period (Apr- Sep 2013). The combination-disk test using cefotaxime and ceftazidime alone and in combination with clavulanic acid was performed for detection of ESBL, while modified Hodge test using cefoxitin disk was performed for detection of AmpC β-lactamase production. Detection of plasmid AmpC genes was performed by polymerase chain reaction (PCR). Presence of respective genes was confirmed by DNA sequencing. The clonal relationship between isolates was analysed by pulsed field gel electrophoresis (PFGE) of SmaI-digested genomic DNA, and visual interpretation of patterns. Results: Fifty-three non-repeated P mirabilis isolates were collected. The strains were isolated from various clinically important specimens, obtained from patients hospitalized in 17 different wards. Beside the β-lactam resistance, all of them were also resistant to other routinely tested antibiotics, with an exception of cefepime, carbapenems, piperacillin-tazobactam variably, and amikacin rarely. For all isolates, the both phenotypic tests were positive, indicating the combined expression of ESBL and AmpC β-lactamase. In all isolates, the presence of AmpC gene was confirmed by PCR and DNA sequencing, and identified as a CMY-type (CIT family). With an exception of one dominant indistinguishable clone, isolates differed in PFGE type, indicating the possibility of both clonal and plasmid spread of resistance. Conclusions: This is the first detection of AmpC gene in P mirabilis in Croatia so far. Further analysis will determine the exact CMY-type of enzymes in the near future. The coexistence of ESBLs and AmpC enzymes significantly reduces treatment options, especially because plasmids often carry multiple other resistances. This, along with possible further acquisition of resistance by other mechanisms (AmpC overexpression, outer membrane porin loss, acquired carbapenemase genes), and intrinsic resistance of P mirabilis to tygecycline and colistin, turns this pathogen to be one of the major concerns in nosocomial infections. It should be monitored in surveillance studies, and involved genes should be identified in order to control the movement of resistance mechanisms. These findings will be followed by more comprehensive research and coordinated surveillance in Croatia.

Proteus mirabilis; multidrug resistance; AmpC β-lactamase; CMY-type

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