Dalbavancin exposure in vitro selects for dalbavancin-non-susceptible and vancomycin-intermediate strains of methicillin-resistant Staphylococcus aureus

Abstract

Objectives: Dalbavancin is a lipoglycopeptide active against methicillin-resistant Staphylococcus aureus (MRSA). Its long half-life (8.5e16 days) allows for once-weekly or single-dose treatments but could prolong the mutant selection window, promoting resistance and cross-resistance to related antimicrobials such as vancomycin. The objective of this study was to evaluate the capacity of post-distributional pharmacokinetic exposures of dalbavancin to select for resistance and cross-resistance in MRSA. Methods: We simulated average, post-distributional exposures of single-dose (1500 mg) dalbavancin (fCmax 9.9 mg/mL, b-elimination t 1/2 204 h) in an in vitro pharmacokinetic/pharmacodynamic (PK/PD) model for 28 days (672 h) against five MRSA strains and one methicillin-susceptible strain (MSSA). Samples were collected at least daily, and surviving colonies were enumerated and screened for resistance on drug-free and dalbavancin-supplemented medium respectively. Isolates from resistance screening plates were subjected to whole-genome sequencing (WGS) and susceptibly testing against dalbavancin, vancomycin, daptomycin, and six b-lactams with varying penicillin-binding protein (PBP) affinities. Results: Dalbavancin was bactericidal against most strains for days 1e4 before regrowth of less susceptible subpopulations occurred. Isolates with eight-fold increases in dalbavancin MIC were detected as early as day 4 but increased 64e128-fold in all models by day 28. Vancomycin and daptomycin MICs increased 4e16-fold, exceeding the susceptibly breakpoints for both antibiotics; b-lactam MICs generally decreased by two-to eight-fold, suggesting a dalbavancineb-lactam seesaw effect, but increased by eight-fold or more in certain isolates. Resistant isolates carried mutations in a variety of genes, most commonly walKR, apt, stp1, and atl. Conclusions: In our in vitro system, post-distributional dalbavancin exposures selected for stable mutants with reduced susceptibility to dalbavancin, vancomycin, and daptomycin, and generally increased susceptibility to b-lactams in all strains of MRSA tested. The clinical significance of these findings remains unclear, but created an opportunity to genotype a unique collection of dalbavancin-resistant strains for the first time. Mutations involved genes previously associated with vancomycin intermediate susceptibility and daptomycin non-susceptibility, most commonly walKR-associated genes.

WITHDRAWN: Molecular Insight Into the Mutation Within Critical Zinc2+-Binding Site in the PAS Domain of WalK in Vancomycin-Intermediate Resistant Staphylococcus Aureus

“…We also observed that one nsSNP (H271D) within the Zn 2+ -binding site (residue 271) was detected in a VISA strain 12 .…”

“…In the present study, an overview of the protein structure mapping of several previously reported nsSNPs in VISA strains indicated that WalK nsSNPs frequently occurred in the PAS CYTO domain 5,12,[27][28][29]33 . However, we observed that nsSNP could occur in every WalK protein domain.…”

“…was associated with increased resistance to vancomycin in the VISA strain 12 . Indeed, in contrast to WalK-H271Y substitution, WalK-H271D substitution in Zn 2+ -binding led to increased vancomycin resistance.…”

“…Recently, structural and functional analyses of Zn 2+ -binding residue 271 within the PAS domain of S. aureus WalK showed that the Zn 2+ -binding site regulates S. aureus WalKR 10 . On the other hand, a substitution mutation in H271 was recently reported in a VISA strain 12 . In B. Subtilis, residue D274 was con rmed to be related to the regulatory pathways of peptidoglycan synthesis 13 .…”

Varied Contribution of Phospholipid Shedding From Membrane to Daptomycin Tolerance in Staphylococcus aureus

“…As such, most studies on daptomycin resistance point to development of mutations in genes that control membrane lipid metabolism and/or lead to changes in surface charge, membrane fluidity, or both, such as mprF, cls, pgsA, and the dlt operon, which reduces binding of daptomycin to the cell membrane or prevents disruption of the membrane by daptomycin (Yang et al, 2009;Peleg et al, 2012;Mishra and Bayer, 2013;Bayer et al, 2014;Cafiso et al, 2014;Mishra et al, 2014;Bayer et al, 2015;Hines et al, 2017;Jiang et al, 2019). Mutations in two-component regulatory systems that regulate cell wall and cell membrane metabolism, such as vraSR and walKR (Friedman et al, 2006;Mehta et al, 2012;Werth et al, 2021), have also been shown to contribute to daptomycin resistance. We previously applied a novel multidimensional lipidomic method to characterizing the detailed lipid profile changes associated with MRSA strains that have developed resistance to daptomycin and found overall greatly decreased levels of PGs in a resistant strain with mutations in both pgsA and mprF (Hines et al, 2017) and greatly elevated levels of lysylphosphatidylglycerols (lysylPGs) and cardiolipins (CLs) in a strain with only an mprF mutation (Hines et al, 2020).…”