Multidrug-resistant (MDR) Gram-negative bacteria (GNB), such as Acinetobacter and Klebsiella, are responsible for severe hospital-acquired infections. Colistin, despite its toxicity and low tissue penetration, is considered the last resort antibiotic against these microorganisms. Of concern, the use of Colistin has recently been compromised by the emergence of Colistin resistance. Herein, we developed a new formulation consisting of multifunctional chitosan-coated human albumin nanoparticles for the delivery of Colistin (Col/haNPs). Col/haNPs were in vitro characterized for encapsulation efficiency, drug release, stability and cytotoxicity and were evaluated for antibacterial activity against MDR GNB (Acinetobacter baumannii and Klebsiella pneumoniae). Col/haNPs showed sizes lower than 200 nm, high encapsulation efficiency (98.65%) and prolonged in vitro release of Colistin. The safety of the nanoformulation was demonstrated by a negligible cytotoxicity on human fibroblasts and hemolytic activity. Col/haNPs evidenced a high antibacterial effect with a significant decrease in MIC values compared to free Colistin, in particular against Col-resistant strains with a pronounced decline of bacterial growth over time. Moreover, Col/haNPs exhibited an inhibitory effect on biofilm formation that was 4 and 60 fold higher compared to free Colistin, respectively for Colistin susceptible and resistant A. baumannii. Our findings suggest that Col/haNPs could represent a promising Colistin nanocarrier with high antimicrobial activity on MDR GNB.

Herein, we report the in vivo evolution of imipenem/relebactam-resistance in KPC-producing K. pneumoniae (KPC-Kp) isolated from a critically-ill patient treated with multiple combination therapies based on ceftazidime-avibactam or meropenem-vaborbactam. Imipenem/relebactam-resistance was associated to meropenem-vaborbactam cross-resistance and was due to truncated OmpK35 and OmpK36 porins and increased copy of bla _KPC copy number.

This study aims to investigate the prevalence of ceftazidime-avibactam (CZA) resistance among carbapenemase-producing Enterobacterales (CPE) blood culture (BC) isolates and the performance of the main carbapenemase phenotypic detection methods to identify KPC variants associated with resistance to CZA. Non-duplicate CPE strains isolated from BCs during 2018-2020 were tested for antimicrobial susceptibility. Molecular testing was used to identify carbapenemase producers. Strains harbouring blaKPC and expressing CZA MIC ≥ 8 mg/L were investigated by sequencing; subsequentially, five phenotypic carbapenemase detection methods were evaluated on these strains: mCIM, Rapidec Carba NP, Disc Diffusion Synergy test, NG-Test CARBA 5 and RESIST-5 O.O.K.N.V. Overall, CZA resistance rate was high (13.7%) and remained relevant (5.9%) excluding metallo-β-lactamases-producers. All isolates harboring blaKPC mutants (n = 8) were associated with reduced carbapenems MICs and negative results by all the methods based on revelation of enzyme activity. Lateral flow immunoassays failed to detect KPC-31 (n = 4) and KPC-33 (n = 2) but correctly identified KPC-14 (n = 2). Conversely, isolates harboring wild-type KPC genes (n = 3) were associated with high level of CZA resistance, carbapenems resistance and tested positive by all the evaluated methods. In the CZA-based therapies era, molecular blaKPC identification followed by a carbapenem-hydrolysis-based phenotypic assay could be the most reasonable diagnostic algorithm to detect all KPC-producers and identify mutants associated with impaired carbapenemase activity and CZA resistance.