Search

Yersinia pestis is the causative agent of plague. As adequate antibiotic treatment falls short and currently no effective vaccine is available, alternative therapeutic strategies are needed. In order to contribute to solving this problem we investigated the therapeutic potential of the peptide construct LFchimera against the safer-to-handle Y. pestis simulants Yersinia enterocolitica and Yersinia pseudotuberculosis in vitro. LFchimera is a heterodimeric peptide construct mimicking two antimicrobial domains of bovine lactoferrin, i.e. lactoferrampin and lactoferricin. LFchimera has been shown to be a potent antimicrobial peptide against a variety of bacteria in vitro and in vivo. Also Y. enterocolitica and Y. pseudotuberculosis have been shown to be susceptible for LFchimera in vitro. As Yersiniae spp. adhere to and invade host cells upon infection, we here investigated the effects of LFchimera on these processes. It was found that LFchimera has the capacity to inhibit host-cell invasion by Yersiniae spp. in vitro. This effect appeared to be host-cell mediated, not bacteria-mediated. Furthermore it was found that exposure of human HeLa epithelial cells to both LFchimera and the bacterial strains evoked a pro-inflammatory cytokine release from the cells in vitro.

Dental biofilms are formed in a multistep process that is initiated by the adhesion of oral bacteria to the dental hard surface. As dental biofilms are associated with oral diseases their control is necessary in order to maintain oral health. Recently, it was revealed that phytosphingosine (PHS)-treated hydroxyapatite discs showed anti-adhesive activity in a static in vitro biofilm model against Streptococcus mutans. The goal of the present study was to further unravel the anti-adhesive and anti-biofilm properties of PHS in both static and dynamic in vitro biofilm models against a full salivary inoculum. After 3 h under static conditions, bacterial adherence on PHS-treated cover glass slides was reduced by 60% compared to the untreated surface. After 6 and 24 h under static conditions, no significant differences in bacterial adherence were observed between PHS-treated and untreated cover glass slides. However, under dynamic conditions, i.e., the presence of shear forces, virtually no bacterial adherence was observed for up to 16 h on PHS-coated surfaces. Besides, PHS showed a strong bactericidal activity on salivary biofilms. Treatment of a 3- and 6-h statically grown biofilm resulted in a 99 and 94% reduction of viable cells, respectively, which was effectuated within minutes. In principle, these anti-adherence and anti-biofilm properties make PHS a promising candidate ingredient for use in oral care products aimed at oral microbial control., (© 2017 S. Karger AG, Basel.)

Lactoferrin (LF) is an important immune protein in neutrophils and secretory fluids of mammals. Bovine LF (bLF) harbours two antimicrobial stretches, lactoferricin and lactoferampin, situated in close proximity in the N1 domain. To mimic these antimicrobial domain parts a chimeric peptide (LFchimera) has been constructed comprising parts of both stretches (LFcin17-30 and LFampin265-284). To investigate the potency of this construct to combat a set of Gram positive and Gram negative bacteria which are regarded as simulants for biological warfare agents, the effect on bacterial killing, membrane permeability and membrane polarity were determined in comparison to the constituent peptides and the native bLF. Furthermore we aimed to increase the antimicrobial potency of the bLF derived peptides by cationic amino acid substitutions. Overall, the bactericidal activity of the peptides could be related to membrane disturbing effects, i.e. membrane permeabilization and depolarization. Those effects were most prominent for the LFchimera. Arginine residues were found to be crucial for displaying antimicrobial activity, as lysine to arginine substitutions resulted in an increased antimicrobial activity, affecting mostly LFampin265-284 whereas arginine to lysine substitutions resulted in a decreased bactericidal activity, predominantly in case of LFcin17-30.

Introduction: The Porcine Myeloid Antibacterial Peptide (PMAP)-23 is a porcine host defence peptide with strong antibacterial activity against Gram-positive and Gram-negative bacteria, and fungi., Objective: PMAP-23 and truncated/mutated derivatives were tested for antibacterial and immunomodulatory activities to determine core elements of the peptide required for functionality., Methods: PMAP-23 and truncated and/or mutated derivatives were synthesized. Antibacterial activity against Gram positive and negative bacteria was determined using colony counting assays. Cytotoxicity was measured against red blood cells and epithelial cells. Peptide induced cytokine production of epithelial cells was determined by ELISA. LPS neutralization was measured using isothermal titration calorimetry and inhibition of LPS induced cytokine production by macrophages. The effect of peptides on phagocytosis was performed by measuring uptake of fluorescently labelled beads by porcine macrophages., Results: Truncation of the peptide did not lead to a strong reduction in antibacterial activity, but interestingly, all C-terminal truncated forms were strongly inhibited by salt addition, unlike the full length peptide or the two N-terminally truncated peptides. None of the peptides were hemolytic or toxic in concentrations up to 40 μM. Full length PMAP-23 induced IL-8 production in porcine epithelial cells, however, this activity was lost in all truncated peptides. None of the peptides bound LPS and subsequently did not inhibit LPS-induced cytokine production of monocytes. Finally, all PMAP-23 derived peptides reduced the uptake of beads by freshly isolated monocytes., Conclusion: PMAP-23 is mainly antibacterial with only limited immunomodulating capacity; the full length peptide is required for the full spectrum of activities., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.)

This study evaluated the anti-biofilm activity of sphingosine, phytosphingosine (PHS), and sphinganine for: (i) anti-adherence activity on hydroxyapatite (HA) surfaces; and (ii) bactericidal activity on different Streptococcus mutans phenotypes (i.e. planktonic cells and cells from a disrupted biofilm). For this, HA discs treated with sphingolipids were incubated with S. mutans and the number of adherent cells was evaluated by both culture and confocal microscopy. Sphinganine strongly inhibited bacterial adherence by 1000-fold compared with an untreated surface. Phytosphingosine and sphingosine inhibited bacterial adherence by eight- and five-fold, respectively, compared with an untreated surface. On saliva-coated HA, sphinganine and PHS inhibited bacterial adherence by 10-fold. Bactericidal activity of sphingolipids was evaluated by culture. For biofilms, the strongest bactericidal activity was exhibited by sphingosine compared with PHS and sphinganine. At a concentration of 12.5 μg ml(-1) , PHS and sphingosine were profoundly effective against planktonic and disrupted biofilms; and sphinganine reduced the number of cells in planktonic form by 100-fold and those derived from a disrupted biofilm by 1000-fold. Atomic force microscopy studies suggested that mechanical stability does not appear to be a factor relevant for anti-fouling activity. The results suggest that sphingolipids may be used to control oral biofilms, especially those loaded with S. mutans., (© 2015 Eur J Oral Sci.)

Proteases are essential for the proliferation and growth of bacteria, and are also known to contribute to bacterial virulence. This makes them interesting candidates as diagnostic and therapeutic targets for infectious diseases. In this review, the authors discuss the most recent developments and potential applications for bacterial proteases in the diagnosis and treatment of bacterial infections. Current and future bacterial protease targets are described and their limitations outlined.

The porcine cathelicidin PR-39 is a host defence peptide that plays a pivotal role in the innate immune defence of the pig against infections. Besides direct antimicrobial activity, it is involved in immunomodulation, wound healing and several other biological processes. In this study, the antimicrobial- and immunomodulatory activity of PR-39, and N- and C-terminal derivatives of PR-39 were tested. PR-39 exhibited an unexpected broad antimicrobial spectrum including several Gram positive strains such as Bacillus globigii and Enterococcus faecalis. Of organisms tested, only Staphylococcus aureus was insensitive to PR-39. Truncation of PR-39 down to 15 (N-terminal) amino acids did not lead to major loss of activity, while peptides corresponding to the C-terminal part of PR-39 were hampered in their antimicrobial activity. However, shorter peptides were all much more sensitive to inhibition by salt. Active peptides induced ATP leakage and loss of membrane potential in Bacillus globigii and Escherichia coli, indicating a lytic mechanism of action for these peptides. Finally, only the mature peptide was able to induce IL-8 production in porcine macrophages, but some shorter peptides also had an effect on TNF-α production showing differential regulation of cytokine induction by PR-39 derived peptides. None of the active peptides showed high cytotoxicity highlighting the potential of these peptides for use as an alternative to antibiotics.

A truncated version of host defense peptide chicken cathelicidin-2, C1-15, possesses potent, broad spectrum antibacterial activity. A variant of this peptide, F(2,5,12)W, which contains 3 phenylalanine to tryptophan substitutions, possesses improved antibacterial activity and lipopolysaccharide (LPS) neutralizing activity compared to C1-15. In order to improve the proteolytic resistance of both peptides we engineered novel chicken cathelicidin-2 analogs by substitution of l- with D-amino acids and head-to-tail cyclization. Both cyclic and D-amino acid variants showed enhanced stability in human serum compared to C1-15 and F(2,5,12)W. The D-amino acid variants were fully resistant to proteolysis by trypsin and bacterial proteases. Head-to-tail cyclization of peptide F(2,5,12)W resulted in a 3.5-fold lower cytotoxicity toward peripheral blood mononuclear cells. In general, these modifications did not influence antibacterial and LPS neutralization activities. It is concluded that for the development of novel therapeutic compounds based on chicken cathelicidin-2 D-amino acid substitutions and cyclization must be considered. These modifications increase the stability and lower cytotoxicity of the peptides without altering their antimicrobial potency., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Staphylococcus epidermidis is a major cause of nosocomial infections owing to its ability to form biofilms on the surface of medical devices. Biofilms are surface-adhered bacterial communities. In mature biofilms these communities are encased in an extracellular matrix composed of bacterial polysaccharides, proteins and DNA. The antibiotic resistance of bacteria present in biofilms can be up to 1000-fold higher compared with the planktonic phenotype. Host defence peptides (HDPs) are considered to be excellent candidates for the development of novel antibiotics. Recently, we demonstrated that a short variant of the HDP chicken cathelicidin-2, peptide F(2,5,12)W, has potent antibacterial and lipopolysaccharide-neutralising activities. This study reports on the antibiofilm activity of peptide F(2,5,12)W against two strains of S. epidermidis, including a multiresistant strain. Peptide F(2,5,12)W potently inhibited the formation of bacterial biofilms in vitro at a low concentration of 2.5 μM, which is below the concentration required to kill or inhibit growth (minimal inhibitory concentration=10 μM). Moreover, peptide F(2,5,12)W also impaired existing S. epidermidis biofilms. A 4-h challenge of pre-grown biofilms with 40 μM F(2,5,12)W reduced the metabolic activity of the wild-type strain biofilm completely and reduced that of the multiresistant strain biofilm by >50%. It is concluded that F(2,5,12)W prevents biofilm formation and impairs mature S. epidermidis biofilms., (Copyright © 2011 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.)

Host defence peptides (HDPs) are considered to be excellent candidates for the development of novel therapeutic agents. Recently, it was demonstrated that the peptide C1-15, an N-terminal segment of chicken HDP cathelicidin-2, exhibits potent antibacterial activity while lacking cytotoxicity towards eukaryotic cells. In the present study, we report that C1-15 is active against bacteria such as Bacillus anthracis and Yersinia pestis that may potentially be used by bioterrorists. Substitution of single and multiple phenylalanine (Phe) residues to tryptophan (Trp) in C1-15 resulted in variants with improved antibacterial activity against B. anthracis and Y. pestis as well as decreased salt sensitivity. In addition, these peptides exhibited enhanced neutralisation of lipopolysaccharide (LPS)-induced release of pro-inflammatory cytokines in human peripheral blood mononuclear cells (PBMCs). The antibacterial and LPS-neutralising activities of these C1-15-derived peptides are exerted at concentrations far below the concentrations that are toxic to human PBMCs. Taken together, we show that Phe-->Trp substitutions in C1-15 variants enhances the antibacterial and LPS-neutralising activities against pathogenic bacteria, including those that may potentially be used as biological warfare agents., (Copyright (c) 2010 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.)