Conventional antibiotics are facing strong microbial resistance that has recently reached critical levels. This situation is leading to significantly reduced therapeutic potential of a huge proportion of antimicrobial agents currently used in clinical settings. Antimicrobial peptides (AMPs) could provide the medical community with an alternative strategy to traditional antibiotics for combating microbial resistance. However, the development of AMPs into clinically useful antibiotics is hampered by their relatively low stability, toxicity, and high manufacturing costs. In this study, a novel in-house-designed potent ultrashort AMP named RBRBR was encapsulated into chitosan-based nanoparticles (CS-NPs) based on the ionotropic gelation method. The encapsulation efficacy reported for RBRBR into CS-NPs was 51.33%, with a loading capacity of 10.17%. The release kinetics of RBRBR from the nanocarrier exhibited slow release followed by progressive linear release for 14 days. The antibacterial kinetics of RBRBR-CS-NPs was tested against four strains of Staphylococcus aureus for 4 days, and the developed RBRBR-CS-NPs exhibited a 3-log decrease in the number of colonies when compared to CS-NP and a 5-log decrease when compared to control bacteria. The encapsulated peptide NP formulation managed to limit the toxicity of the free peptide against both mammalian cells and human erythrocytes. Additionally, the peptide NPs demonstrated up to 98% inhibition of biofilm formation when tested against biofilm-forming bacteria. Loading RBRBR into CS-NPs could represent an innovative approach to develop delivery systems based on NP technology for achieving potent antimicrobial effects against multidrug-resistant and biofilm-forming bacteria, with negligible systemic toxicity and reduced synthetic costs, thereby overcoming the obstructions to clinical development of AMPs.

The recent upsurge of multidrug resistant bacteria (MDRB) among global communities has become one of the most serious challenges facing health professionals and the human population worldwide. Cationic ultrashort antimicrobial peptides (USAMPs) are a promising group of molecules that meet the required criteria of novel antimicrobial drug development. UP-5, a novel penta-peptide, displayed significant antimicrobial activities against various standard and clinical isolates of MDRB. UP-5 displayed MICs values within the range of (10–15 μM) and (55–65 μM) against Gram-positive and Gram-negative bacteria, respectively. Furthermore, UP-5 displayed antibiofilm activity with minimum biofilm eradication concentration (MBEC) value as equal to twofold higher than MIC value. At the same inhibitory concentrations, UP-5 exhibited very low or negligible toxicity toward human erythrocytes and mammalian cells. Combining UP-5 with conventional antibiotics led to a synergistic or additive mode of action that resulted in the reduction of the MIC values for some of the antibiotics by 99.7% along a significant drop in MIC values of the peptide. The stability profile of UP-5 was evaluated in full mouse plasma and serum with results indicating a more stable pattern in plasma. The present study indicates that USAMPs are promising antimicrobial agents that can avoid the negative characteristics of conventional antimicrobial peptides. Additionally, USAMPs exhibit good to moderate activity against MDRB, negligible toxicity, and synergistic outcomes in combination with conventional antimicrobial agents.

IntroductionAs the development of new antimicrobial agents faces a historical decline, the issue of bacterial drug resistance has become a serious dilemma that threatens the human population worldwide. Antimicrobial peptides (AMPs) represent an attractive and a promising class of antimicrobial agents.AimThe hybridization of AMPs aimed at merging two individual active fragments of native peptides to generate a new AMP with altered physicochemical properties that translate into an enhanced safety profile.Materials and methodsIn this study, we have rationally designed a new hybrid peptide via combining two individual α-helical fragments of both BMAP-27 and OP-145. The resultant peptide, was evaluated for its antimicrobial and antibiofilm activity against a range of microbial strains. The resultant peptide was also evaluated for its toxicity against mammalian cells using hemolytic and anti proliferative assays.ResultsThe antimicrobial activity of H4 revealed that the peptide is displaying a broad spectrum of activity against both Gram-positive and Gram-negative bacteria including standard and multidrug-resistant bacterial strains in the range of 2.5–25 μM. The new hybrid peptide displayed potent activity in eradicating biofilm-forming cells, and the reported minimum biofilm eradication concentrations were equal to the minimum inhibitory concentration values reported for planktonic cells. Additionally, H4 exhibited reduced toxicity profiles against eukaryotic cells. Combining H4 peptide with conventional antibiotics has led to a dramatic enhancement of the antimicrobial activity of both agents with synergistic or additive outcomes.ConclusionOverall, this study indicates the success of both the hybridization and synergism strategy in developing AMPs as potential antimicrobial therapeutics with reduced toxicity profiles that could be efficiently employed to eradicate resistant bacterial strains and enhance the selectivity and toxicity profiles of native AMPs.

There is great interest in the development of antimicrobial peptides as a potentially novel class of antimicrobial agents. Several structural determinants are responsible for the antimicrobial and cytolytic activity of antimicrobial peptides. In our study, a new synthetic peptide analog, AamAP1-Lysine from the naturally occurring scorpion venom antimicrobial peptide AamAP1, was designed by modifying the parent peptide in order to increase the positive charge and optimize other physico-chemical parameters involved in antimicrobial activity. AamAP1-Lysine displayed potent antibacterial activity against Gram-positive and Gram-negative bacteria. The minimum inhibitory concentration was in the range of 5 to 15 µM with a 10 fold increase in potency over the parent peptide. The hemolytic and antiproliferative activity of AamAP1-Lysine against eukaryotic mammalian cells was minimal at the concentration range needed to inhibit bacterial growth. The antibacterial mechanism analysis indicated that AamAP1-Lysine is probably inducing bacterial cell death through membrane damage and permeabilization determined by the release of β-galactosidase enzyme from peptide treated E. coli cells. DNA binding studies revealed that AamAP1-Lysine caused complete retardation of DNA migration and could display intracellular activities in addition to the membrane permeabilization mode of action reported earlier. In conclusion, AamAP1-Lysine could prove to be a potential candidate for antimicrobial drug development in future studies.

Background and purpose: Antimicrobial resistance toward antibiotics is reaching historical unprecedented levels. There is an urgent and imminent need to develop novel antimicrobial alternatives. Antimicrobial peptides could prove to be a successful group of antimicrobials for drug development. Recently, we have designed a novel synthetic peptide named AamAP1-Lysine. The peptide displayed potent wide-spectrum antimicrobial activities against Gram-positive and Gram-negative bacteria. The purpose of this study is to evaluate the antimicrobial effect of combining AamAP1-Lysine with five different conventional antibiotics each representing a distinct mechanism of action in order to explore the possibility of producing a synergistic mode of action against a resistant strain of Gram-positive and a resistant strain of Gram-negative bacteria. Methodology: The antimicrobial activity of AamAP1-Lysine in combination with five different antibiotics were evaluated for their antimicrobial activity employing standard antimicrobial assays, the synergistic activity of the peptide-antibiotic combinations were evaluated using checkerboard technique in addition to real-time time-kill assays. For the antibiofilm studies, the MBEC values were determined by employing the Calgary device. Results: The combination strategy displayed potent synergistic activities against planktonic bacteria in a significant number of peptide-antibiotic combinations. The synergistic activity managed to reduce the effective minimum inhibitory concentration (MIC) concentrations dramatically with some combinations exhibiting a 64-fold decrease in the effective MIC of AamAP1-Lysine individually. Additionally, the combined synergistic activities of the peptide antibiotics were evaluated, and our results have identified two peptide antibiotic combinations with potent synergistic activities against biofilm growing strains of resistant bacteria. Conclusion: Our results clearly indicate that peptide-antibiotic combinations could prove to be a very effective strategy in combatting multidrug-resistant bacteria and biofilm caused infections.

Background and purpose: The world is heading to a post-antibiotic era where the treatment of bacterial infections will not be possible even with well-known last-line antibiotics. Unfortunately, the emergence of multidrug resistant bacterial strains is uncontrollable, and the humanity will face a life-threatening fate unless new antimicrobial agents with new bacterial target sites are promptly developed. Herein, we design a hybrid antimicrobial peptide (B1) from helical parts taken from the parent peptides: LL-37 and BMAP-27. The purpose of this design is to improve the potency and enhance the toxicity profile of the parent peptides. Methods: Rational design was used to hybridize two antimicrobial peptides, in which two helical parts from the bovine analog BMAP-27, and the human cathelicidin LL-37 were used to generate a novel peptide (B1). The physicochemical properties were checked using in silico methods. The antimicrobial activities were tested against nine control and resistant strains of Gram-positive and Gram-negative bacteria. On the other hand, the antibiofilm activities were tested against four resistant strains. The cytotoxicity on mammalian cells was tested using HEK293, and the hemolysis activity was also investigated on human blood. Finally, synergistic studies were performed with four conventional antibiotics against four resistant strains of Gram-positive and Gram-negative bacteria. Results: The new peptide B1 exhibited broad-spectrum activities against all tested strains. The concentration against planktonic cells ranged between 10 and 20 µM. However, 40–60 µM were needed to eradicate the biofilms. B1 showed reduced toxicity toward mammalian cells with minimal hemolysis risk. On the other hand, the synergistic studies showed improved activities for the combined conventional antibiotics with a huge reduction in their minimum inhibitory concentration values. The concentrations of B1 peptide combined with the tested antibiotics were also decreased markedly down to 0.5 µM in some cases. Conclusion: B1 is a hybrid peptide from two cathelicidin peptides. It showed an improved activity compared to parent peptides. The hybridization was successful in this study. It generated a new potent broad-spectrum antimicrobial. The toxicity profile was improved, and the synergism with the convention antibiotics showed promising results.

Current research in the field of antimicrobials is focused on developing novel antimicrobial agents to counteract the huge dilemma that the human population is mainly facing in regards to the rise of bacterial resistance and biofilm infections. Host defense peptides (HDPs) are a promising group of molecules for antimicrobial development as they display several attractive features suitable for antimicrobial activity, including their broad spectrum of activity and potency against bacteria. AamAP1 is a novel HDP that belongs to the venom of the North African scorpion Androctonus amoeruxi. In vitro antimicrobial assays revealed that the peptide displays moderate activity against Gram-positive and Gram-negative bacteria. Additionally, the peptide proved to be highly hemolytic and displayed significantly high toxicity against mammalian cells. In our study, a novel synthetic peptide analogue named A3 was synthetically modified from AamAP1 in order to enhance its activity and toxicity profile. The design strategy depended on modifying the amino acid sequence of AamAP1 in order to alter its net positive charge, percentage helicity and modify other parameters that are involved theoretically in HDPs activity. Accordingly, A3 was evaluated for its in vitro antimicrobial and anti-biofilm activity individually and in combination with four different types of conventional antibiotics against clinical isolates of multi-drug resistant (MDR) Gram-positive bacteria. A3 was also evaluated for its cytotoxicity against mammalian cells. A3 managed to selectively inhibit the growth of a wide range of resistant strains of Gram-positive bacteria. Our results also showed that combining A3 with conventional antibiotics caused a synergistic antimicrobial behavior that resulted in decreasing the MIC value for A3 peptide as low as 0.125 µM. At the concentrations needed to inhibit bacterial growth, A3 displayed minimal mammalian cell toxicity. In conclusion, A3 exhibits enhanced activity and selectivity when compared with the parent natural scorpion venom peptide. The combination of A3 with conventional antibiotics could provide researchers in the antimicrobial drug development field with a potential alternative for conventional antibiotics against MDR bacteria.

The rise of multidrug-resistant bacteria is causing a serious threat to the world’s human population. Recent reports have identified bacterial strains displaying pan drug resistance against antibiotics and generating fears among medical health specialists that humanity is on the dawn of entering a post-antibiotics era. Global research is currently focused on expanding the lifetime of current antibiotics and the development of new antimicrobial agents to tackle the problem of antimicrobial resistance. In the present study, we designed a novel consensus peptide named “Pepcon” through peptide consensus sequence determination among members of a highly homologous group of scorpion antimicrobial peptides. Members of this group were found to possess moderate antimicrobial activity with significant toxicity against mammalian cells. The aim of our design method was to generate a novel peptide with an enhanced antimicrobial potency and selectivity against microbial rather than mammalian cells. The results of our study revealed that the consensus peptide displayed potent antibacterial activities against a broad range of Gram-positive and Gram-negative bacteria. Our membrane permeation studies displayed that the peptide efficiently induced membrane damage and consequently led to cell death through the process of cell lysis. The microbial DNA binding assay of the peptide was found to be very weak suggesting that the peptide is not targeting the microbial DNA. Pepcon induced minimal cytotoxicity at the antimicrobial concentrations as the hemolytic activity was found to be zero at the minimal inhibitory concentrations (MICs). The results of our study demonstrate that the consensus peptide design strategy is efficient in generating peptides.