Polymyxins Bind to the Cell Surface of Unculturable Acinetobacter baumannii and Cause Unique Dependent Resistance

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.

Simulations of octapeptin–outer membrane interactions reveal conformational flexibility is linked to antimicrobial potency

“…Unlike previous simulation studies in which the bacterial OM was simplified into a symmetric or asymmetric LPS bilayer with model phospholipid compositions, [23][24][25][26] we utilized the quantitative membrane lipidomics results from polymyxinresistant A. baumannii 5075R to construct a more realistic OM model that allowed for better precision when characterizing interactions with the octapeptins. 19 Analysis of the simulations revealed that a minor structural variation between octapeptin C4 and FADDI-115, namely, the presence of a hydroxyl group on the fatty acyl group of octapeptin C4 (Fig 1A), was sufficient to impact their conformational transitions and resulted in different free energy profiles for OM penetration. These results provided novel atomic-scale insights into the structure-activity relationship of the octapeptins against polymyxin-resistant Gramnegative bacteria.…”

“…33 Notably, the accurate lipid composition of the OM (Fig 1B) was based on our quantitative lipidomics results of polymyxin-resistant A. baumannii 5075R. 19 Initially, the octapeptin molecules were placed above the outer leaflet of the OM using the gmx insert-molecules tool by replacing the overlapping water molecules. 34 TIP3P water molecules and counter-ions (CaCl2) were employed to hydrate and neutralize the simulation systems, respectively.…”

Colistin Dependence in Extensively Drug-Resistant Acinetobacter baumannii Strain Is Associated with ISAjo2 and ISAba13 Insertions and Multiple Cellular Responses

“…Pairwise sequence alignment with Stretcher (EMBL-EBI) [ 45 ] disclosed 100% identity of the VgrG- and Hcp-encoding contigs from Ab-S and Ab-D. According to Zhu et al, mutations in katG , encoding a catalase involved in reactive oxygen species scavenging, affected polymyxin dependence in MRD A. baumannii AB5075 [ 15 ] and inactivation of mrcA , encoding the penicillin-binding protein A1, has been suggested to promote LOS loss [ 7 ]. Nevertheless, no mutations/substitutions/insertions in katG or mrcA were present in Ab-S and Ab-D.…”

“…This was discovered, however, in colistin-resistant A. baumannii obtained after 120 generations of passaging on 10 µg/mL polymyxin B, while we observed the emergence of phenotypically colistin-dependent subpopulation already after 8 passages on colistin [ 38 ]. Recently, a transcriptomic analysis of a colistin-dependent isolate in relation to its parent colistin-susceptible MRD A. baumannii AB5075 revealed 1.7-fold increased expression of mlaD and 2.0-fold decreased expression of pldA in colistin-dependent bacteria [ 15 ]. We believe that this discrepancy can be attributed to the difference between strains.…”

“…Colistin-susceptible A. baumannii strains may develop dependence after exposure to colistin in vitro [ 12 , 13 , 14 ]. Colistin dependence is not rare, as many as 32.9% of colistin-susceptible isolates were able to do so [ 14 ], most of them were otherwise multidrug-resistant and able to cause infection in mice [ 15 ]. Colistin dependence may represent a transition step to resistance and LOS-deficiency, with direct binding of colistin to phosphatidylglycerol-rich areas of the OM in LOS-deficient strains, has been proposed as a key step [ 15 ].…”