| Congresso Brasileiro de Microbiologia 2023 | Resumo: 744-1 | ||||
Resumo:Introduction: Ceftazidime-avibactam (CZA) has been used for difficult-to-treat (DTR) Klebsiella pneumoniae infections, highlighting those caused by KPC producers and/or polymyxin B-resistant isolates. Antibiotic tolerance is the ability of a susceptible bacterium (without resistance mechanism) to survive an antibiotic at the minimum inhibitory concentration or higher for extended periods of time. Aims: The aims of this study were to (i) investigate the occurrence of CZA tolerance in DTR K. pneumoniae clinical isolates, previously categorized as susceptible to CZA, (ii) evaluate whether CZA tolerance occurs due to ceftazidime (CAZ) alone or due to the presence of avibactam (AVI) in the CZA combination, (iii) investigate the genetic determinants of resistance and tolerance to antibiotics and evaluate the molecular epidemiology of selected isolates. Material and Methods: Twenty non-clonal carbapenem-resistant KPC-producing K. pneumoniae, isolated over a three-month period in 2022, from bloodstream infections affecting inpatients at a University Hospital in Ribeirão Preto, were studied. CZA tolerance phenotype was investigated in CZA-susceptible (non-CZA-heteroresistant) isolates using Tolerance Disk Test (TDtest). To investigate whether CZA tolerance occurs due to CAZ tolerance or due to the presence of AVI, 10/20 selected isolates were evaluated by disk-diffusion test using CAZ+phenylboronic acid (PBA), aiming KPC inhibition, with subsequent TDtest for CAZ+PBA. Those same 10/20 isolates were also evaluated by whole genome sequencing for epidemiologic purposes, genetic characterization of the resistance to antibiotics, and search for mutations in genes related to beta-lactam tolerance (rpoS, recA, spoT, and relA). Results: Most isolates (17/20, 85%), previously categorized as CZA-susceptible, were detected as CZA-tolerant. One isolate was CZA-resistant, and two others were CZA-susceptible (no tolerance detected). For the 10/20 selected isolates (nine CZA-tolerant and one CZA-resistant), tolerance to CAZ+PBA was also detected, suggesting that CZA tolerance occurs due to CAZ tolerance and not to the presence of AVI, and it was observed when a KPC inhibitor was added to CAZ. These isolates belonged to sequence types 11, 258 and 340; harbored resistance genes such as blaKPC-2 or KPC-33, blaCTX-M-15, 14 or 2, blaOXA-1 or 2, qnrB19, qnrB1, fosA, rmtB, aac(3)-IId, aac(6')-Ib-cr, aac(3)-IIa, aac(6')-Ib3, aph(3'')-Ib, aph(6)-Id, aadA2, tet(D), tet(G); and presented mutations in genes such as ompK36, ompK37, gyrA, parC, rpsJ, pmrB, and mgrB. CZA-tolerant isolates harbored blaKPC-2, and the CZA-resistant isolate harbored blaKPC-33. Regarding tolerance genes, one isolate presented rpoS mutation (S36N). Conclusions: Notably, 85% of isolates were detected as CZA-tolerant. CZA tolerance seems to occur due to CAZ tolerance, instead of the presence of AVI in the CZA combination, however, further investigations are needed to confirm it. The CZA-tolerant isolates and the CZA-resistant isolate harbored resistance genes to different antibiotic classes (outstandingly blaKPC-33 in the CZA-resistant isolate). Mutation in only one gene related to beta-lactam tolerance was found in only one isolate, suggesting the presence of yet unknown beta-lactam tolerance genes. These results are concerning because CZA tolerance could contribute to antibiotic failure as well as to the emergence and spread of CZA-resistant bacteria. Palavras-chave: antibiotic tolerance, antibiotic resistance, carbapenemase, TDtest Agência de fomento:Instituto Nacional de Pesquisa em Resistencia Antimicrobiana (INCT –MCTI/CNPq/CAPES/FAPs nº16/2014) and FAPESP SPRINT (2020/00898-0). This study was financed in part by the CAPES – Brazil – Finance Code 001. CAPES Grants 88887.670254/2022-0 (PROEX) | |||||