In vitro antimicrobial susceptibilities of multidrug-resistant Proteus mirabilis coproducing TEM and CMY-16 beta-lactamases

Coexistence of different resistance mechanisms often leaves very few therapeutic options. In this study we characterized the types of β-lactamases that coexist in multidrug-resistant (MDR) P mirabilis isolates from University Hospital Centre Split, and determined in vitro antimicrobial susceptibilities for this type of strain. Methods: Consecutive non-repeated P mirabilis isolates were collected from different clinical samples during a 1-year period (Apr 2013- Apr 2014), based on the resistance to third-generation cephalosporins and cefoxitin. Double-disk synergy test was performed for detection of extended-spectrum β-lactamase (ESBL), and cefoxitin Hodge test for detection of AmpC β-lactamase. Genes encoding beta-lactamases were detected by polymerase chain reaction (PCR) and identified by DNA sequencing. Clonal relatedness of isolates was analyzed by Enterobacterial Repetitive Intergenic Consensus (ERIC)-PCR fingerprinting. Antimicrobial susceptibilities to a wide range of antimicrobial agents were tested by disk diffusion method, and antibiotics that have shown activity were used for determination of minimal inhibitory concentrations (MICs) by gradient strip method. In both methods, the results were interpreted according to EUCAST clinical breakpoints. Results: A total of 109 isolates were analysed. Double-disk synergy test and cefoxitin Hodge test were positive for all of them, and presence of respective genes was confirmed by PCR. DNA sequencing revealed coexistence of TEM-116 and CMY-16 β-lactamases. ERIC-PCR demonstrated clonal relatedness of the isolates. All isolates were multidrug-resistant, with 100% resistance to amoxicillin/clavulanate, cephalosporins except the fourth generation, quinolones and trimethoprim-sulfamethoxazole. Seventeen isolates were susceptible to amikacin, and only 6 of them to all aminoglycosides. Piperacillin/tazobactam and cefepime showed variable degrees of resistance: 52.3% isolates were susceptible to piperacillin/tazobactam in disk diffusion test, and 55.1% were susceptible in gradient strip test. However, the gradient test showed a slightly higher percentage of resistant isolates (16.3% vs. 10.1%), at the expense of those who were intermediate in disk diffusion test. For cefepime, 62.6% isolates were susceptible, 24.3% intermediate and 13.1% resistant in disk diffusion test, while 61.8% were susceptible, 23.5% intermediate and 14.7% resistant in gradient strip method. All isolates were 100% susceptible to carbapenems. Conclusions: This study shows that carbapenems should be the drug of choice for serious infections caused by MDR P mirabilis coproducing TEM-116 and CMY-16 β-lactamases. Piperacillin/tazobactam and cefepime are less reliable agents for therapy of serious infections and should be checked with gradient strip method before considering as a treatment option, with attention paid to pharmacokinetic and pharmacodynamic targets. Acquisition of carbapenemase genes, along with an intrinsic resistance to tigecycline and colistin, indicates the possibility of development MDR P mirabilis into a pan-resistant strain.