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Objectives: In order to evaluate host defense peptides (HDPs) HHC-10 and synoeca-MP activity in in vitro osteoclastogenesis process and in vivo induced apical periodontitis, testing the effect of molecules in the inflammatory response and in apical periodontitis size/volume after root canal treatment. Materials and methods: In vitro osteoclastogenesis was assessed on bone marrow cell cultures extracted from mice, while in vivo endodontic treatment involved rats treated with Ca(OH)2 or HDPs. In vitro osteoclasts were subjected to TRAP staining, and in vivo samples were evaluated by radiographic and tomographic exams, as well as histologic analysis. Results: None of the substances downregulated the in vitro osteoclastogenesis. Nevertheless, all treatments affected the average of apical periodontitis size in rats, although only teeth treated with HDPs demonstrated lower levels of the inflammatory process. These results demonstrated the in vivo potential of HDPs. Radiographic analysis suggested that HHC-10 and synoeca-MP-treated animals presented a similar lesion size than Ca(OH)2-treated animals after 7-day of endodontic treatment. However, tomography analysis demonstrated smaller lesion volume in synoeca-MP-treated animals than HHC-10 and Ca(OH)2-treated animals, after 7 days. Conclusions: These molecules demonstrated an auxiliary effect in endodontic treatment that might be related to its immunomodulatory ability, broad-spectrum antimicrobial activity, and possible induction of tissue repair at low concentrations. These results can encourage further investigations on the specific mechanisms of action in animal models to clarify the commercial applicability of these biomolecules for endodontic treatment. Clinical significance: HDPs have the potential to be adjuvant substances in endodontic therapy due to its potential to reduce inflammation in apical periodontitis. [ABSTRACT FROM AUTHOR]

‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬This study aims to evaluate the in vitro antimicrobial and immunomodulatory activities and cytotoxicity of chlorhexidine (CHX) and synoeca‐MP peptide alone or in combination against Pseudomonas aeruginosa. The antimicrobial property was evaluated by the determination of minimal inhibitory concentration, minimum bactericidal concentration, and planktonic bacteria and biofilm inhibition. Immunomodulatory activity was determined by enzyme‐linked immunosorbent assay and nitric oxide production by the Griess reaction method. According to the results, synoeca‐MP combined with CHX demonstrated antimicrobial effectiveness compared with its isolated use, in addition to immunomodulatory activity (upregulating MPC‐1 and tumor necrosis factor‐α and downregulating nitric oxide and interleukin‐10). In this context, it is expected that the substances, together, could be capable of controlling bacterial infection and dissemination, besides potentiating macrophages' immune response against the studied microorganism. Moreover, reducing the CHX concentration by the addition of synoeca‐MP peptide may, in a beneficial way, minimize the undesirable effects of both, CHX and synoeca‐MP in a clinical setting. [ABSTRACT FROM AUTHOR]

An important aspect related to infectious pathogens is their exceptional adaptability in developing resistance, which leads to a perpetual challenge in the discovery of antimicrobial drugs with novel mechanisms of action. Among them, antimicrobial peptides (AMPs) stand out as promising anti-infective molecules. In order to overcome the high costs associated with isolation from natural sources or chemical synthesis of AMPs we propose the expression of Pa-MAP 2, a polyalanine AMP. Pa-MAP 2 was fused to an ELP-intein tag where the ELP (Elastin-like polypeptide) was used to promote aggregation and fast and cost-effective isolation after expression, and the intein was used to stimulate a controlled AMP release. For these, the vector pET21a was used to produce Pa-MAP 2 fused to the N-termini region of a modified Mxe GyrA intein followed by 60 repetitions of ELP. Purified Pa-MAP 2 showed a MIC of 25 μM against E. coli ATCC 8739. Batch fermentation demonstrated that Pa-MAP-2 can be produced in both rich and defined media at yields 50-fold higher than reported for other AMPs produced by the ELP-intein system, and in comparable yields to expression systems with protease or chemical cleavage. [ABSTRACT FROM AUTHOR]

The increasing number of multidrug resistance microorganisms is an alarming threat, and their rapid detection is essential to prevent nosocomial, foodborne, or waterborne infections. Many peptides derived from the venom of wasp Synoeca surinama have antimicrobial activity against Gram-positive and Gram-negative bacteria. Synoeca-MP, an antimicrobial peptide (AMP) from mastoparan family, seems to increase bacterial membrane permeability, promoting cytotoxicity and membrane disruption. Here Synoeca-MP was evaluated as biorecognition element tethered over chitosan-coated magnetic nanoparticles (Fe 3 O 4 -Chit). The transducing layer of the biosensor was developed from the self-assembling of 4-mercaptobenzoic acid (4-MBA) monolayer onto gold substrate. Atomic force microscopy (AFM) analyses confirmed the biointeraction between AMP and different pathogens membranes. The fabrication and performance of the biosensing assembly were characterized by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). Detection of Enterococcus faecalis (G+) , Klebsiella pneumoniae (G-) , Pseudomonas aeruginosa (G-), and Candida tropicalis was assessed in a recognition range from 101 to 105 CFU.mL−1. An instrumental limit of detection of 10 CFU.mL−1 was obtained for each specimen. However, the device presented a preferential selectivity towards Gram-negative bacteria. The proposed biosensor is a sensitive, fast, and straightforward platform for microbial detection in aqueous samples, envisaged for environmental monitoring applications. • A new electrochemical biosensor with high sensitivity towards Gram-negative species is presented. • The biosensor combines chitosan-coated magnetic nanoparticles with the antimicrobial peptide Synoeca-MP. • The sensor detected bacteria and fungi species with a LOD of 101 CFU mL−1. • The platform presented high sensitivity towards K. pneumoniae. • This biosensor stands out as a promising tool for fast microbial detection in liquid samples. [ABSTRACT FROM AUTHOR]

• The biosensor can detect and differentiate among various Candida yeast species. • CLAVMO is a suitable biological probe that can be used to detect and differentiate Candida species. • Limits of detection (LODs) between 2 and 3 CFU mL-1 were obtained. Yeast biotechnologies are a cornerstone to modern lifestyle in all spheres of society, as their products are essential for the economy and industrial activities. The life cycle of many yeasts such as Candida alternates between diplophase and haplophase. Both ploidies can exist as stable cultures; however, it has been shown that wild haploid yeast efficiently cross in poor environments, whereas wild diploid species hybridize predominantly in rich nutrient surroundings. Some Candida species have had their ploidy engineered to produce various important biomolecules useful to the detergent, food, pharmaceutical and other industries. Several works show that MALDI-TOF MS has become an ideal platform for ploidy analysis. Recent reports have shown that antimicrobial peptides (AMP) present enhanced hydrophobic interaction towards cell membranes. In this work, the authors explored a modified AMP named Clavanin A (CLAVMO) as an unprecedented bioreceptor capable of differentiating the molecular composition of the cell wall of Candida spp. A thin film of poly(3-thiophene acetic acid) (PTAA) was electropolymerized to avail the covalent immobilization of amino-functionalized titanium dioxide nanoparticles (TiO 2 NPs) whose electrochemical properties are excellent to provide a discernable redox response. Electrochemical impedance spectroscopy (EIS) and atomic force microscopy (AFM) were used to characterize the biosensor assembly and the biorecognition process. Additionally, MALDI-TOF was employed to confirm ploidy of Candida strains used in this work. Our results suggest that TiO 2 NPs_CLAVMO biosensor is a suitable biological probe that can be used to detect and differentiate Candida species based on their ploidy. For instance, C. Albicans and C. tropicalis , which are close related species known for being diploid presented higher impedimetric response than haploid species C.krusei and C. glabrata. The proposed biosensor stands out as a useful alternative for highly sensitive differentiation of Candida yeast cells, without the intricacies of MALDI-TOF sample preparation. The biodetection event and its electrochemical evaluation took less than 20 min to perform. The platform showed stability for over a week, a linear detection range between 101-106 CFU mL-1 and limits of detection (LODs) between 2 and 3 CFU mL-1. [ABSTRACT FROM AUTHOR]

Discovering antibiotic molecules able to hold the growing spread of antimicrobial resistance is one of the most urgent endeavors that public health must tackle. The case of Gram-negative bacterial pathogens is of special concern, as they are intrinsically resistant to many antibiotics, due to an outer membrane that constitutes an effective permeability barrier. Antimicrobial peptides (AMPs) have been pointed out as potential alternatives to conventional antibiotics, as their main mechanism of action is membrane disruption, arguably less prone to elicit resistance in pathogens. Here, we investigate the in vitro activity and selectivity of EcDBS1R4, a bioinspired AMP. To this purpose, we have used bacterial cells and model membrane systems mimicking both the inner and the outer membranes of Escherichia coli, and a variety of optical spectroscopic methodologies. EcDBS1R4 is effective against the Gram-negative E. coli, ineffective against the Gram-positive Staphylococcus aureus and noncytotoxic for human cells. EcDBS1R4 does not form stable pores in E. coli, as the peptide does not dissipate its membrane potential, suggesting an unusual mechanism of action. Interestingly, EcDBS1R4 promotes a hemi-fusion of vesicles mimicking the inner membrane of E. coli. This fusogenic ability of EcDBS1R4 requires the presence of phospholipids with a negative curvature and a negative charge. This finding suggests that EcDBS1R4 promotes a large lipid spatial reorganization able to reshape membrane curvature, with interesting biological implications herein discussed. [ABSTRACT FROM AUTHOR]