Antibiotics are losing effectiveness as bacteria become resistant to conventional drugs. To find new alternatives, antimicrobial peptides (AMPs) are rationally designed with different lengths, charges, hydrophobicities (
- Award ID(s):
- 2115790
- NSF-PAR ID:
- 10412117
- Date Published:
- Journal Name:
- Frontiers in Microbiology
- Volume:
- 13
- ISSN:
- 1664-302X
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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H ), and hydrophobic moments (μH), containing only three types of amino acids: arginine, tryptophan, and valine. Six AMPs with low minimum inhibitory concentrations (MICs) and <25% toxicity to mammalian cells are selected for biophysical studies. Their secondary structures are determined using circular dichroism (CD), which finds that the % α‐helicity of AMPs depends on composition of the lipid model membranes (LMMs): gram‐negative (G(−)) inner membrane (IM) >gram‐positive (G(+))> Euk33 (eukaryotic with 33 mol% cholesterol). The two most effective peptides, E2‐35 (16 amino acid [AA] residues) and E2‐05 (22 AAs), are predominantly helical in G(–) IM and G(+) LMMs. AMP/membrane interactions such as membrane elasticity, chain order parameter, and location of the peptides in the membrane are investigated by low‐angle and wide‐angle X‐ray diffuse scattering (XDS). It is found that headgroup location correlates with efficacy and toxicity. The membrane bending modulusK Cdisplays nonmonotonic changes due to increasing concentrations of E2‐35 and E2‐05 in G(–) and G(+) LMMs, suggesting a bacterial killing mechanism where domain formation causes ion and water leakage. -
Lu, Hua (Ed.)SPLUNC1 (short palate lung and nasal epithelial clone 1) is a multifunctional host defense protein found in human respiratory tract with antimicrobial properties. In this work we compare the biological activities of four SPLUNC1 antimicrobial peptide (AMP) derivatives using paired clinical isolates of the Gram-negative (G(-)) bacteria Klebsiella pneumoniae, obtained from eleven patients with/without colistin resistance. Secondary structural studies were carried out to study interactions between the AMPs and lipid model membranes (LMMs) utilizing circular dichroism (CD). Two peptides were further characterized using x-ray diffuse scattering (XDS) and neutron reflectivity (NR). A4-153 displayed superior antibacterial activity in both G(-) planktonic cultures and biofilms. NR and XDS revealed that A4-153 (highest activity) is located primarily in membrane headgroups, while A4-198 (lowest activity) is located in hydrophobic region. CD revealed that A4-153 is helical while A4-198 has little helical character, demonstrating that helicity and efficacy are correlated in these SPLUNC1 AMPs.more » « less
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null (Ed.)Abstract Background Though many plant defensins exhibit antibacterial activity, little is known about their antibacterial mode of action (MOA). Antimicrobial peptides with a characterized MOA induce the expression of multiple bacterial outer membrane modifications, which are required for resistance to these membrane-targeting peptides. Mini-Tn 5-lux mutant strains of Pseudomonas aeruginosa with Tn insertions disrupting outer membrane protective modifications were assessed for sensitivity against plant defensin peptides. These transcriptional lux reporter strains were also evaluated for lux gene expression in response to sublethal plant defensin exposure. Also, a plant pathogen, Pseudomonas syringae pv. syringae was modified through transposon mutagenesis to create mutants that are resistant to in vitro MtDef4 treatments. Results Plant defensins displayed specific and potent antibacterial activity against strains of P. aeruginosa . A defensin from Medicago truncatula , MtDef4, induced dose-dependent gene expression of the aminoarabinose modification of LPS and surface polycation spermidine production operons. The ability for MtDef4 to damage bacterial outer membranes was also verified visually through fluorescent microscopy. Another defensin from M. truncatula , MtDef5, failed to induce lux gene expression and limited outer membrane damage was detected with fluorescent microscopy. The transposon insertion site on MtDef4 resistant P. syringae pv. syringae mutants was sequenced, and modifications of ribosomal genes were identified to contribute to enhanced resistance to plant defensin treatments. Conclusions MtDef4 damages the outer membrane similar to polymyxin B, which stimulates antimicrobial peptide resistance mechanisms to plant defensins. MtDef5, appears to have a different antibacterial MOA. Additionally, the MtDef4 antibacterial mode of action may also involve inhibition of translation.more » « less
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Background Antimicrobial resistance is a growing concern in canine
Staphylococcus pseudintermedius dermatitis. Treatment with rifampicin (RFP) is considered only in meticillin‐resistant and multidrug‐resistantS. pseudintermedius (MDR‐MRSP).Hypothesis/Objectives To determine an optimal RFP dosing for MDR‐MRSP treatment without induction of RFP resistance and identify causal mutations for antimicrobial resistance.
Methods and materials Time–kill assays were performed in a control isolate and three MDR‐MRSP isolates at six clinically relevant concentrations [32 to 1,024 × MIC (the minimum inhibitory concentration)]. Whole‐genome resequencing and bioinformatic analysis were performed in the resistant strains developed in this assay.
Results The genomic analysis identified nine antimicrobial resistance genes (ARGs) in MDR‐MRSP isolates, which are responsible for resistance to seven classes of antibiotics. RFP activity against all four isolates was consistent with a time‐dependent and bacteriostatic response. RFP resistance was observed in six of the 28 time–kill assays, including concentrations 64 × MIC in MDR‐MRSP1 isolates at 24 h, 32 × MIC in MDR‐MRSP2 at 48 h, 32 × MIC in MDR‐MRSP3 at 48 h and 256 × MIC in MDR‐MRSP3 at 24 h. Genome‐wide mutation analyses in these RFP‐resistant strains discovered the causal mutations in the coding region of the
rpoB gene.Conclusions and clinical relevance A study has shown that 6 mg/kg per os results in plasma concentrations of 600–1,000 × MIC of
S. pseudintermedius . Based on our data, this dose should achieve the minimum MIC (×512) to prevent RFP resistance development; therefore, we recommend a minimum daily dose of 6 mg/kg for MDR‐MRSP pyoderma treatment when limited antibiotic options are available. -
null (Ed.)Antimicrobial resistance is a world-wide health care crisis. New antimicrobials must both exhibit potency and thwart the ability of bacteria to develop resistance to them. We report the use of synthetic ionophores as a new approach to developing non-resistant antimicrobials and adjuvants. Most studies involving amphiphilic antimicrobials have focused on either developing synthetic amphiphiles that show ion transport, or developing non-cytotoxic analogs of such peptidic amphiphiles as colistin. We have rationally designed, prepared, and evaluated crown ether-based synthetic ionophores (‘hydraphiles’) that show selective ion transport through bilayer membranes and are toxic to bacteria. We report here that hydraphiles exhibit a broad range of antimicrobial properties and that they function as adjuvants in concert with FDA-approved antibiotics against multi-drug resistant (MDR) bacteria. Studies described herein demonstrate that benzyl C 14 hydraphile (BC 14 H) shows high efficacy as an antimicrobial. BC 14 H, at sub-MIC concentrations, forms aggregates of ∼200 nm that interact with the surface of bacteria. Surface-active BC 14 H then localizes in the bacterial membranes, which increases their permeability. As a result, antibiotic influx into the bacterial cytosol increases in the presence of BC n Hs. Efflux pump inhibition and accumulation of substrate was also observed, likely due to disruption of the cation gradient. As a result, BC 14 H recovers the activity of norfloxacin by 128-fold against resistant Staphylococcus aureus . BC 14 H shows extremely low resistance development and is less cytotoxic than colistin. Overall, synthetic ionophores represent a new scaffold for developing efficient and non-resistant antimicrobial-adjuvants.more » « less