Abstract
Multidrug‐resistant Acinetobacter baumannii is a top‐priority pathogen globally and polymyxins are a last‐line therapy. Polymyxin dependence in A. baumannii (i.e., nonculturable on agar without polymyxins) is a unique and highly‐resistant phenotype with a significant potential to cause treatment failure in patients. The present study discovers that a polymyxin‐dependent A. baumannii strain possesses mutations in both lpxC (lipopolysaccharide biosynthesis) and katG (reactive oxygen species scavenging) genes. Correlative multiomics analyses show a significantly remodeled cell envelope and remarkably abundant phosphatidylglycerol in the outer membrane (OM). Molecular dynamics simulations and quantitative membrane lipidomics reveal that polymyxin‐dependent growth emerges only when the lipopolysaccharide‐deficient OM distinctively remodels with ≥ 35% phosphatidylglycerol, and with “patch” binding on the OM by the rigid polymyxin molecules containing strong intramolecular hydrogen bonding. Rather than damaging the OM, polymyxins bind to the phosphatidylglycerol‐rich OM and strengthen the membrane integrity, thereby protecting bacteria from external reactive oxygen species. Dependent growth is observed exclusively with polymyxin analogues, indicating a critical role of the specific amino acid sequence of polymyxins in forming unique structures for patch‐binding to bacterial OM. Polymyxin dependence is a novel antibiotic resistance mechanism and the current findings highlight the risk of ‘invisible’ polymyxin‐dependent isolates in the evolution of resistance.
Highlights
A number of genes associated with oxidative stress responses were differentially expressed in untreated 5075D compared to untreated 5075S, including the reduced expression of ferric uptake repressor fur and the increased expression of genes related to siderophore synthesis and transport, heme generation, glutathione biosynthesis, sulfate uptake and assimilation, and thiol/disulfide homeostasis maintenance (Figure S4, Supporting Information); all these results indicated a defense against
Further simulations with polymyxin B nonapeptide (PMBN) showed that PMBN patch bound to 5075D outer membrane (OM) and reduced the lateral diffusion coefficient to 3.24 × 10−8 cm2 s−1 (Figure 5D; and Movie S2, Supporting Information); these results showed that the polymyxin diaminobutyric acid (Dab) residues played major roles in membrane stabilization
We explored if polymyxin dependence exists in other Gram-negative bacteria (e.g., Pseudomonas aeruginosa and Klebsiella pneumoniae); no single polymyxin-dependent strain could be selected in either species
Summary
A colistin-dependent isolate 5075D was evolved from the wildtype A. baumannii AB5075 (labeled as 5075S below, see the Experimental Section). Strain 5075R possessed 3 nonsynonymous substitutions (pmrBG315D, ABUW_0507A91P, and rpoBF915L), while 5075D had an integration of ISAba (1180-bp in length) at nucleotide position 395 of lpxC (Figures S1 and 2A, Supporting Information) and 4 nonsynonymous substitutions (ABUW_0135S25Stop, mrcAM129T, katGR609G, and rpoBF915L) These changes were present in 100% of reads, making the involvement of heterogeneity unlikely. Transposon insertion in ABUW_0135, mrcA, katG, and rpoB were all susceptible to polymyxins (MIC ≤ 1 mg L−1, Tables S1 and S2, Supporting Information), indicating that the inactivation of these individual genes did not confer polymyxin-dependent resistance. Our previous study showed that IS insertions in lpxC abolished its function, led to LPS loss and high-level resistance to polymyxins.[13] katG encodes a catalase and A. baumannii lacking katG showed increased susceptibility to H2O2.[21] Interestingly, 2000704 (3 of 13). The lpxC mutation was essential in the acquisition of polymyxin dependence in 5075D, while the katG mutation is not essential but played an important role here
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
