| Congresso Brasileiro de Microbiologia 2023 | Resumo: 941-1 | ||||
Resumo:The treatment of infections caused by Enterobacterales producing metallo-β-lactamases (MBL) and/or co-producing MBL and KPC is challenging. The combination of ceftazidime-avibactam (CZA) and aztreonam (ATM) has been an alternative and the in vitro evaluation of this synergism allows optimization of the therapeutic regimen. Checkerboard and time-kill tests are time-consuming techniques which are not indicated for clinical microbiology laboratories. Therefore, we developed and standardized a new semi-quantitative methodology, the microelution, to quickly assess the susceptibility of Enterobacteriaceae to CZA/ATM. For the proof of concept, 5 carbapenem-resistant Enterobacterales (producing NDM (2) or co-producing KPC and NDM (3)) plus Escherichia coli ATCC 25922 (negative control) were used. Stock solutions were prepared by eluting antibiotic discs to reach the following concentrations: (i) ATM 32 μg/mL (further diluted into secondary solutions, 16 and 8 μg/mL); (ii) CZA 120/80 μg/mL. Eight wells of a microtiter plate were used with different concentrations of antibiotic(s): (1) growth control; (2) CZA 6/4 μg/mL; (3) ATM 4 μg/mL; (4) ATM/CZA 4/6/4 μg/mL; (5) ATM 8 μg/mL; (6) ATM/CZA 8/6/4 μg/mL; (7) ATM 16 μg/mL; (8) ATM/CZA 16/6/4 μg/mL. Distinct inoculums were evaluated (3.0, 1.0 and 0.5 McFarland, diluted 1:100, 1:50, 1:10, 1:5 and 1:2). To visualize bacterial growth, resazurin (0.02%) was used and its addition was evaluated at different times (before incubation; after 2h and 3h of incubation). Test was read every 15 minutes for up to 1 hour after prior incubation. Color change, from blue to pink, indicated resistance to the combination/concentration of the antibiotic(s). Results were compared to broth microdilution for CZA, ATM and ATM/avibactam. The most accurate results were obtained using McFarland's 0.5 inoculum diluted 1:10 and with the addition of resazurin after 3h of incubation. At higher concentrations of bacteria, the inoculum effect compromised the specificity, while fewer bacteria needed more time and/or were less sensitive. We did not observe any discordant results. Moreover, except for one isolate that demanded 60 min to color change on the ATM 16 μg/mL well, all positive results were obtained within 45 min after addition of resazurin. These preliminary results indicated that the semi-quantitative methodology based on disc microelution is reliable and warrants further work using an adequate number of isolates. Palavras-chave: aztreonam, ceftazidime-avibactam, Enterobacterales, microelution | |||||