Bactericidal Efficiency and Modes of Action of the Novel Antimicrobial Peptide T9W against Pseudomonas aeruginosa

Zhu, Xin; Shan, Anshan; Ma, Zhi; Xu, Wei; Wang, Jiajun; Chou, Shuli; Cheng, Baohua

doi:10.1128/aac.04830-14

Cited by 42 publications

(45 citation statements)

References 35 publications

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“…The membrane depolarization caused by both melimine and Mel4 was a concentration dependent event. These results are in agreement with previous studies showing a concentration-dependent depolarizing effect of the helical peptide LL-37 on P. aeruginosa, and PMAP-36, GI24 and melittin on E. coli cell membranes (54, 55). To further confirm whether membrane depolarization was a lethal event for these peptides, viable count was performed.…”

Section: Discussionsupporting

confidence: 93%

Comparative mode of action of antimicrobial peptide melimine and its derivative Mel4 against Pseudomonas aeruginosa

Yasir

Dutta

Willcox

2018

Preprint

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Society for Microbiology annual meetings in 2017. There are no conflicts of interest for any 31 of the authors that could have influenced the results of this work. Prof. Mark Willcox holds 32 the patent for the melimine peptide. 33 2 ABSTRACT 34Melimine and Mel4 are chimeric cationic peptides with broad spectrum antimicrobial 35 activity, and recent investigations have shown that they are highly biocompatible with animal 36 model and human clinical trials. The current study examined the mechanism of action of 37 these two antimicrobial peptides against P. aeruginosa with a series of investigations. 38 Antimicrobial activities were determined by MIC and MBC. Endotoxin neutralization was 39 determined using the LAL assay, effect on the cytoplasmic membrane was evaluated using 40 DiSC(3)-5 and Sytox green stains, and Syto-9 and PI dyes using flow cytometry. Release of 41 cytoplasmic materials (ATP and DNA/RNA) were determined using ATP luminescence and 42 increase in OD 260nm . The ability to lyse bacteria was studied by measuring a decrease in 43 OD 620nm . The MIC of the peptides remained low against P. aeruginosa strains, which showed 44 efficient neutralization of LPS, indicating their role in the anti-pseudomonas and LPS binding 45 activities. Both AMPs rapidly (starting at 30 seconds) depolarized P. aeruginosa cytoplasmic 46 membrane leading to reduction in viability. Melimine was responsible for more ATP release 47 (75%) compared to Mel4 (36%) (P<0.001) following two minutes exposure. For both 48 peptides, Sytox green entered cells after five minutes of incubation. Flow cytometry 49 demonstrated that both the AMPs permeabilized the cell membrane at 30 minutes and 50 followed by increasing permeability. Similar results were found with DNA/RNA release 51 experiments. Overall, melimine showed higher ability of membrane disruption, cell lysis 52 compared to Mel4 (P<0.001). Knowledge regarding mechanism of action of these two AMPs 53 would be helpful in making them as anti-pseudomonas drug. 54KEYWORDS P. aeruginosa, Antimicrobial peptides, mechanism of action, membrane 55 disruption 56 57 3 58 59 Pseudomonas aeruginosa is a metabolically versatile ubiquitous Gram-negative opportunistic 60 pathogen that can cause infections in animals and plants (1). P. aeruginosa is responsible for 61 localized to systemic infections in humans, which can be life threatening. Over the years, P. 62 aeruginosa has become one of the most frequent causative agents of nosocomial infections, 63 associated with substantial morbidity and mortality (2). The current standards of care to treat 64 P. aeruginosa infections are not effective (3) as its outer membrane offers low permeability, 65 which limits the penetration of antibiotics into the bacterial cells thereby increase antibiotic 66 resistance (4, 5). Given the severity of P. aeruginosa infections and the limited antimicrobial 67 arsenal with which to treat them, finding new alternative antimicrobials with unique 68 mechanisms of action is urgently required (6). 69 Antimicr...

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Section: Discussionsupporting

confidence: 93%

Comparative mode of action of antimicrobial peptide melimine and its derivative Mel4 against Pseudomonas aeruginosa

Yasir

Dutta

Willcox

2018

Preprint

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“…8A). Cecropin A2 behaved in a manner similar to that of polymyxin B, which is known to bind LPS and was used as a positive control (28,29). The LPS-binding activity of cecropin A2 was dose dependent, and the peptide was able to displace BODIPY TR cadaverine, confirming that cecropin A2 binds to P. aeruginosa LPS with a high affinity.…”

Section: Zheng Et Al Antimicrobial Agents and Chemotherapymentioning

confidence: 62%

“…The affinities of the test compounds for LPS were determined using the BODIPY TR cadaverine displacement assay (28). Polymyxin B, a peptide antibiotic known to bind and neutralize LPS, was used as the positive control (29), and meropenem was used as the negative control.…”

Section: Methodsmentioning

confidence: 99%

Synergistic Efficacy of Aedes aegypti Antimicrobial Peptide Cecropin A2 and Tetracycline against Pseudomonas aeruginosa

Zheng

Tharmalingam

et al. 2017

Antimicrob Agents Chemother

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The increasing prevalence of antibiotic resistance has created an urgent need for alternative drugs with new mechanisms of action. Antimicrobial peptides (AMPs) are promising candidates that could address the spread of multidrugresistant bacteria, either alone or in combination with conventional antibiotics. We studied the antimicrobial efficacy and bactericidal mechanism of cecropin A2, a 36-residue ␣-helical cationic peptide derived from Aedes aegypti cecropin A, focusing on the common pathogen Pseudomonas aeruginosa. The peptide showed little hemolytic activity and toxicity toward mammalian cells, and the MICs against most clinical P. aeruginosa isolates were 32 to 64 g/ml, and its MICs versus other Gramnegative bacteria were 2 to 32 g/ml. Importantly, cecropin A2 demonstrated synergistic activity against P. aeruginosa when combined with tetracycline, reducing the MICs of both agents by 8-fold. The combination was also effective in vivo in the P. aeruginosa/Galleria mellonella model (P Ͻ 0.001). We found that cecropin A2 bound to P. aeruginosa lipopolysaccharides, permeabilized the membrane, and interacted with the bacterial genomic DNA, thus facilitating the translocation of tetracycline into the cytoplasm. In summary, the combination of cecropin A2 and tetracycline demonstrated synergistic antibacterial activity against P. aeruginosa in vitro and in vivo, offering an alternative approach for the treatment of P. aeruginosa infections.KEYWORDS antimicrobial activity, antimicrobial peptide, cecropin A2, Pseudomonas aeruginosa, tetracycline P seudomonas aeruginosa is a Gram-negative opportunistic bacterial pathogen that causes life-threatening infections with high rates of mortality (1-3). It is associated with nosocomial pneumonia, wound infections, bacteremia, and sepsis among patients with various underlying diseases, including cystic fibrosis, HIV/AIDS, and cancer (4). Antibiotic therapy is challenging in this setting because clinical strains of P. aeruginosa often show extensive intrinsic resistance to a wide range of antimicrobial agents, including tetracyclines, ␤-lactams, aminoglycosides, and fluoroquinolones (5). This intrinsic resistance has generally been attributed to the low membrane permeability of P. aeruginosa, up to 100 times lower than that of Escherichia coli (6). P. aeruginosa can also withstand antibiotics that attack the outer membrane through the efficient deployment of transmembrane efflux pumps, preventing contact between the antibiotics and their intracellular targets (7-11).The ubiquitous and relentless clinical challenge of drug resistance has created a

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“…(4)(5)(6) Many of these AMPs are short peptides which fold into amphiphilic α-helices in contact with the bacterial membrane, (7)(8)(9)(10)(11)(12)(13)(14) leading to aggregation inducing membrane disruption, morphological changes and cell lysis as documented by imaging studies. (15)(16)(17)(18)) Such short AMPs can exert potent effect against multidrug resistant bacteria and their biofilms, (19,20) and show potent activities in vivo. (21) Interestingly, the experimental evidence for α-helix formation in these peptides is mostly based on circular dichroism (CD) and NMR studies.…”

Section: Introductionmentioning

confidence: 99%

X-ray Crystal Structures of Short Antimicrobial Peptides as Pseudomonas aeruginosa Lectin B Complexes

et al. 2019

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Herein we report X-ray crystal structures of 11-13 residue antimicrobial peptides (AMPs) active against Pseudomonas aeruginosa as complexes of fucosylated D-enantiomeric sequences with the P. aeruginosa lectin LecB. These represent the first crystal structures of short AMPs. In twenty-four individual structures of eight different peptides we found mostly α-helices assembled as two-helix or four-helix bundles with a hydrophobic core and cationic residues pointing outside. Two of the analogs formed an extended structure engaging in multiple contacts with the lectin. Molecular dynamics (MD) simulations showed that αhelices are stabilized by bundle formation and suggested that the N-terminal acyl group present in the linker to the fucosyl group can extend the helix by one additional H-bond and increase α-helix amphiphilicity. Investigating N-terminal acylation led to AMPs with equivalent and partly stronger antibacterial effects compared to the free peptide.

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Bactericidal Efficiency and Modes of Action of the Novel Antimicrobial Peptide T9W against Pseudomonas aeruginosa

Cited by 42 publications

References 35 publications

Comparative mode of action of antimicrobial peptide melimine and its derivative Mel4 against Pseudomonas aeruginosa

Comparative mode of action of antimicrobial peptide melimine and its derivative Mel4 against Pseudomonas aeruginosa

Synergistic Efficacy of Aedes aegypti Antimicrobial Peptide Cecropin A2 and Tetracycline against Pseudomonas aeruginosa

X-ray Crystal Structures of Short Antimicrobial Peptides as Pseudomonas aeruginosa Lectin B Complexes

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