Obtaining a Peptide Complex with Antibacterial Effect from Donor Leukocytes

Ultrasound exposure can increase the membrane permeability of human neutrophil granulocytes containing microbubbles without causing complete cell destruction

Octominin: An antimicrobial peptide with antibacterial and anti-inflammatory activity against carbapenem-resistant Escherichia coli both in vitro and in vivo

Macroalgae are currently being explored as novel and sustainable sources of bioactive compounds. Ulva prolifera is a major member of the green algae, as it includes various bioactive compounds, including carotenoids, fucoidans, and phlorotannins, with various biological activities in pharmaceutical, nutraceutical, cosmeceutical, and functional foods industries. In the current work, the rbcL gene products were utilized in identifying U. prolifera molecularly. The antibacterial and antioxidant activities of the hexane, ethyl acetate, and methanol extracts of U. prolifera were carried out In vitro. The total antioxidant capacity method was utilized in assessing the antioxidant activity. Estimation of nutritional value of U. prolifera showed 5.4±0.04%, 1.7±0.04% and 43±0.5% dry weight for total proteins, lipids, and carbohydrates, respectively. The total antioxidant capacity of algal hexane, ethyl acetate, and methanol extract was 0.97±0.09, 1.23±0.04 and 1.63±0.09 mg equivalent ascorbic acid/gdw, respectively. Investigation of the antibacterial activity of U. prolifera against Staphylococcus aureus and Aeromonas hydrophila showed that the recorded diameter of the inhibition zones for the methanolic extract was 1.1cm. The antibacterial effect of hexane extract against Edwardsiella tarda was high which had a diameter of 1.5cm. Of interest, the ethyl acetate extract of U. prolifera showed potent antibacterial activity against the three bacteria, including Vibrio anguillarum, Aeromonas hydrophila, and Edwardsiella tarda. These results indicate that macroalgae U. prolifera contain bioactive substances with potential antioxidant and antibacterial effects.

BackgroundEssential oils are becoming increasingly popular in medicinal applications because of their antimicrobial effect. Thymus vulgaris L. (Lamiaceae) is a well-known and widely cultivated medicinal plant, which is used as a remedy for cold, cough and gastrointestinal symptoms. Essential oil content of thyme is responsible for its antimicrobial activity, however, it has been reported that the chemical composition of essential oils influences its biological activity. In order to explore flowering phenophases influence on the chemical composition of thyme essential oil and its antibacterial and anti-biofilm activity, plant materials were collected at the beginning of flowering, in full bloom and at the end of flowering periods in 2019.MethodsEssential oils from fresh and dried plant materials were distilled and analyzed with gas chromatography-mass spectrometry (GC-MS) and gas chromatography-flame ionization detection (GC-FID). The antibacterial activity was performed by broth microdilution and thin layer chromatography-direct bioautography (TLC-DB) assays and the anti-biofilm effect by crystal violet assay, respectively. Scanning electron microscopy was applied to illustrate the cellular changes of bacterial cells after essential oil treatment.ResultsThymol (52.33–62.46%) was the main component in the thyme essential oils. Thyme oil distilled from fresh plant material and collected at the beginning of flowering period exerted the highest antibacterial and anti-biofilm activity against Haemophilus influenzae, H. parainfluenzae and Pseudomonas aeruginosa.ConclusionThe different flowering periods of Thymus vulgaris influence the antibacterial and anti-biofilm activity of its essential oils, therefore, the collection time has to be taken into consideration and not only the full bloom, but the beginning of flowering period may provide biological active thyme essential oil.

Xylocandin is a complex of novel peptides with potent antifungal activity that is produced by Pseudomonas cepacia ATCC 39277. The complex was isolated from the fermentation broth by extraction with butanol-methanol, 9:1, followed by collection of the precipitate formed upon concentration of the solvent extract. Purification was effected by chromatography on reversed phase and size exclusion gels followed by TLC on silica gel. These techniques afforded eight components: A1, A2, B1, B2, C1, C2, D1 and D2. A mixture of the two closely related components, xylocandins A1 and A2, displayed potent anticandidal and antidermatophytic activities in vitro. The activity was diminished by the presence of serum or vaginal washings. No antibacterial activity was demonstrable.

Dendritic Cells, Tolerance Induction and Transplant Outcome

Natural products are used as alternative drugs in traditional medicine to treat infection and inflammation and relieve pain. Heartwood of Cassia garettiana Craib has been investigated as an ingredient in Thai traditional medicine for anti-HIV protease, but there is no report on its antibacterial and anti-inflammatory activities. The objectives of this study were to investigate the anti-inflammatory and antibacterial activities, time-kill profile, and main active constituents of an ethanolic extract of C. garettiana heartwood. The study followed the generally accepted experimental design. All tests were investigated in triplicate. The heartwood of C. garettiana was extracted by maceration with 95% EtOH. The antibacterial activity of the extract and its chemical constituents were determined by their MIC values using resazurin as an indicator. Time-kill profile was determined at 0, 2, 4, 6, 8, 10, 12, and 24 hrs and expressed as log CFU/mL. The anti-inflammatory activity of the extract and its chemical components was investigated by their inhibiting effect on IL-6 and TNF-α production by ELISA. The ethanolic extract was analyzed for its chemical constituents by HPLC technique. The ethanolic extract showed both dose- and time-dependent bactericidal effects against Staphylococcus aureus, methicillin-resistance Staphylococcus aureus, Staphylococcus epidermidis, Escherichia coli, Pseudomonas aeruginosa, Salmonella Typhi, Salmonella Typhimurium, Klebsiella pneumoniae, and Shigella dysenteriae with MIC values of 312.5, 312.5, 312.5, 1,250, 2,500, 625, 625, 2,500, and 625 μg/mL, respectively. It showed an inhibiting effect on IL-6 production at concentrations of 12.5 to 100 μg/mL. The main active chemical constituent of C. garettiana was piceatannol that showed antibacterial activity against all test bacteria except P. aeruginosa. C. garettiana showed a broad spectrum of antibacterial activity against both Gram-negative and Gram-positive bacteria. Piceatannol and resveratrol from the plant strongly inhibited IL-6 production. Based on these results, we concluded that the ethanolic extract of C. garettiana showed both an antibacterial activity and inhibition of IL-6. Piceatannol is the active constituent of the extract and showed anti-inflammatory and antibacterial activities against Gram-negative and Gram-positive bacteria.

Group IIA secreted phospholipase A(2) (sPLA2) is known to display potent Gram-positive bactericidal activity in vitro and in vivo. We have analyzed the bactericidal activity of the full set of recombinant murine and human groups I, II, V, X, and XII sPLA2s on Listeria monocytogenes, Staphylococcus aureus, and Escherichia coli. The rank order potency among human sPLA2s against Gram-positive bacteria is group IIA > X > V > XII > IIE > IB, IIF (for murine sPLA2s: IIA > IID > V > IIE > IIC, X > IB, IIF), and only human group XII displays detectable bactericidal activity against the Gram-negative bacterium E. coli. These studies show that highly basic sPLA2s display potent bactericidal activity with the exception of the ability of the acidic human group X sPLA2 to kill Gram-positive bacteria. By studying the Bacillus subtilis and S. aureus bactericidal potencies of a large panel of human group IIA mutants in which basic residues were mutated to acidic residues, it was found that: 1) the overall positive charge of the sPLA2 is the dominant factor in dictating bactericidal potency; 2) basic residues on the putative membrane binding surface of the sPLA2 are modestly more important for bactericidal activity than are other basic residues; 3) relative bactericidal potency tracks well with the ability of these mutants to degrade phospholipids in the bacterial membrane; and 4) exposure of the bacterial membrane of Gram-positive bacteria by disruption of the cell wall dramatically reduces the negative effect of charge reversal mutagenesis on bactericidal potency.

Although biocides have been used for a century, the number of products containing biocides has recently increased dramatically with public awareness of hygiene issues. The antimicrobial efficacy of biocides is now well documented; however, there is still a lack of understanding of their antimicrobial mechanisms of action. There is a wide range of biocides showing different levels of antimicrobial activity. It is generally accepted that, in contrast to chemotherapeutic agents, biocides have multiple target sites within the microbial cell and the overall damage to these target sites results in the bactericidal effect. Information about the antimicrobial efficacy of a biocide (i.e. the eta-value) might give some useful indications about the overall mode of action of a biocide. Bacteriostatic effects, usually achieved by a lower concentration of a biocide, might correspond to a reversible activity on the cytoplasmic membrane and/or the impairment of enzymatic activity. The bacteriostatic mechanism(s) of action of a biocide is less documented and a primary (unique?) target site within the cell might be involved. Understanding the mechanism(s) of action of a biocide has become an important issue with the emergence of bacterial resistance to biocides and the suggestion that biocide and antibiotic resistance in bacteria might be linked. There is still a lack of understanding of the mode of action of biocides, especially when used at low concentrations (i.e. minimal inhibitory concentration (MIC) or sublethal). Although this information might not be required for highly reactive biocides (e.g. alkylating and oxidizing agents) and biocides used at high concentrations, the use of biocides as preservatives or in products at sublethal concentrations, in which a bacteriostatic rather than a bactericidal activity is achieved, is driving the need to better understand microbial target sites. Understanding the mechanisms of action of biocides serves several purposes: (i) it will help to design antimicrobial formulations with an improved antimicrobial efficacy and (ii) it will ensure the prevention of the emergence of microbial resistance.

The bone infection osteomyelitis (typically caused by Staphylococcus aureus) usually requires a multistep procedure – long term administration of high-dose systemic antibiotics combined with surgical debridement and bone grafting. However, the disease remains notoriously difficult-to-treat due to the poor penetration of systemic antibiotics into the necrotic bone, antibiotic resistance, and the steep decline in antibiotic discovery. Therefore, osteomyelitis treatment has a dual challenge: ensuring an effective and non-toxic dose of antimicrobial, while ensuring bone regeneration is stimulated. Thus, the overall aim of this thesis was to develop a one-step tissue engineering-based treatment strategy for osteomyelitis that combines local, controlled release of non-antibiotic antibacterials with a regenerative collagen-based scaffold to facilitate bone repair.In the study presented in Chapter 2 of this thesis, a number of non-antibiotic antibacterial agents (specifically chitosan, copper nanoparticles, silver nanoparticles, zinc nanoparticles, copper chloride salt, silver nitrate salt, and zinc chloride salt) were screened as potential agents for osteomyelitis infection treatment based on their antibacterial effect on three clinically relevant bacterial species (Staphylococcus aureus, Staphylococcus epidermidis, and Escherichia coli) while retaining viable numbers of mammalian cells. The results demonstrate that there is a fine balance between the two, and that in particular chitosan and the metal salts, copper chloride and zinc chloride, may have potential for osteomyelitis infection treatment due to their superior antibacterial activity potential while maintaining mammalian cell viability.In the study presented in Chapter 3 of this thesis, the optimal antibacterial metal salts identified from the screening process in Chapter 2 were incorporated into 3D collagen/chitosan-based scaffolds via both (i) direct incorporation and (ii) through a chitosan microparticle controlled delivery system, with the aim of minimising toxicity and prolonging bioactivity. The results demonstrated that the two scaffold types were effective in providing different metal salt release quantities and/or profiles which in turn influenced the antibacterial activity of the scaffolds against S. aureus. It was also foundthat the scaffolds did not elicit a significant toxic effect towards mammalian cells, some scaffolds supported osteogenesis, and all copper-incorporated scaffolds enhanced angiogenesis.In the study presented in Chapter 4 of this thesis, a range of silver-doped hydroxyapatites were successfully produced and incorporated into collagen scaffolds. The results demonstrate that the composite silver-doped hydroxyapatite /collagen scaffolds demonstrate enhanced microarchitectural and mechanical properties vs. the collagen control, whilst also demonstrating potent antibacterial activity. Although the scaffolds showed toxicity towards mammalian cells in vitro, an assessment in an in vivo environment may reveal the true cytocompatibility of the scaffolds, as shown previously in the literature for similar scaffolds. Further fine-tuning of the dose of silver-doped hydroxyapatite or its bioactivity through e.g. size or shape changes may also be required to maximise the potential of this nascent system.In Chapter 5 a second multifunctional material – copper-doped bioactive glass – was tested as an alternative to silver-doped hydroxyapatite due to its potential to enhance both osteogenesis and angiogenesis while retaining antibacterial potency. In addition to promoting osteogenesis and angiogenesis in vitro, the scaffolds developed were capable of significant antibacterial activity. Most promisingly, when tested in an in vivo chick embryo ex ovo model, the copper-doped bioactive glass scaffolds were not only biocompatible, showing no signs of toxicity, but also demonstrated the same pattern of enhanced osteogenesis and angiogenesis as the in vitro studies.Collectively, the research in this thesis presents a number of potential single-stage solutions for osteomyelitis treatment. Such strategies potentially reduce the need for antibiotics and bone grafting, reducing hospital stays and costs. In addition, the platform systems developed in Chapters 3 – 5 might be further modified and used for the controlled delivery of an array of antimicrobial and therapeutic metal ions depending on the intended application, making them attractive candidates for other indication beyond bone including soft tissue applications such as wound healing.

Purpose: This study aims to confirm the physiological activity of cytotoxicity, antioxidant effects, and antibacterial effects of 9 kinds phytoncide extracts were investigated and the possibility of their application as cosmetic materials was studied.Methods: The 9 kinds natural products related to phytoncide extract were selected, extracted, and used as samples. Safety was confirmed through MTT assay, and antioxidant effect through DPPH radical scavenging activity evaluation and antibacterial effect through disc diffusion assay, minimum inhibitory concentration (MIC), and minimum bacteriological concentration (MBC) were evaluated.Results: All 9 kinds phytoncide extracts were confirmed to be safe against Fibroblast cytotoxicity at a concentration of 3.00% or less. The DPPH radical scavenging ability was similar in all phytoncide extracts, and it was confirmed that there was an antioxidant effect. The antibacterial effect did not show strong antibacterial activity against gram-negative bacteria, yeast and fungi as a whole, but it showed excellent antibacterial activity against gram-positive bacteria. Dendranthema zawadskii (Herb.) Tzvelev, Cupressus sempervirens showed antibacterial activity only on gram-positive bacteria. Juniperus chinensis showed excellent antibacterial activity against gram-positive bacteria, yeast. Angelica tenuissima Nakai showed high antibacterial activity against fungi rather than bacteria.Conclusion: These results show that excellent antibacterial and antioxidant effects were confirmed in Dendranthema zawadskii (Herb.) Tzvelev, Cupressus sempervirens, Juniperus chinensis and Angelica tenuissima Nakai at a concentration of 3.00% or less. It is judged that highly likely to be use as a cosmetic material with natural ingredients that replace synthetic preservatives or antioxidants.

To generate oregano essential oil, the leaves and flowering tops of the Origanum vulgare plant go through the process known as steam distillation. This essential oil is known for its antibacterial activity. Bacterial biofilms are microbial communities attached to inert surfaces or tissues and encapsulated in complex matrices. Planktonic bacteria reversibly attach to surfaces, form microcolonies, and generate polymeric matrices around biofilms. Bacteria in biofilms provide bacteria with a safer way to reproduce and survive. This research tests the antibacterial activity and effect on the biofilm formation of Oregano essential oil. The antibacterial activity and effect on biofilm formation were tested against five bacterial strains, including Escherichia coli ATCC 14169, Escherichia coli ATCC 25922, Staphylococcus aureus NCTC 12393, Staphylococcus aureus ATCC 25923, and Staphylococcus aureus ATCC 6538. The concentrations of oil that were used in this research were 100%(v/v), 75%(v/v), 50%(v/v), and 25%(v/v). The best antibacterial effect was achieved against Staphylococcus aureus NCTC 12393 at 25%(v/v) of oil concentration. While performing the experiment, a variety of oregano oil concentrations had significant results for further tests to be performed.

The increasing concern over multidrug resistance in pathogens has led to an ongoing search for novel antibiotics derived from soil actinobacteria. In this current investigation, actinobacteria were isolated from the rhizosphere of bamboo plants collected within the Megamalai forest of the Western Ghats in the Theni zone of Tamil Nadu, India. These actinobacteria were subjected to characterization, and their growth conditions were optimized to enhance the production of bioactive compounds. To assess antibacterial properties, the isolated Actinobacteria underwent testing using the agar plug method. The strain exhibiting notable antibacterial activity underwent further characterization through 16s rRNA gene sequencing and subsequent phylogenetic analysis. Employing response surface methodology (RSM), cultural conditions were fine-tuned. Bioactive compounds were extracted from the culture medium using ethyl acetate, and their antibacterial and antioxidant effects were evaluated through disc diffusion and DPPH radical scavenging methods, respectively. Ethyl acetate extracts were analyzed by using FT-IR and GC-MS techniques. In total, nine strains of Actinobacteria were isolated from the rhizosphere soil of bamboo. Among these, strain BS-16 displayed remarkable antibacterial activity against three strains: Staphylococcus aureus (19 mm), Bacillus subtilis (12 mm), and Streptococcus pyogenes (10 mm). This strain was identified as Streptomyces sp. The optimal conditions for bioactive compound production were determined as follows: malt extract (10 g), yeast extract (5 g), dextrose (5 g), pH 6.5, and temperature 30 °C. After a 7-day incubation period, the results showed a 6% increase in production. The ethyl acetate fraction derived from strain BS-16 exhibited dose-dependent antibacterial and antioxidant activities. FT-IR and GC-MS analyses revealed the presence of active compounds with antibacterial effects within the extract. Consequently, further investigation into the BS-16 strain holds promise for scaling up the production of bioactive compounds possessing antibacterial and antioxidant properties.

Nanoparticles synthesized through biological processes are utilized in cancer treatment, drug delivery systems, nanosensors, photoimaging, cosmetics, and medical applications. Extracts from Nigella sativa seeds have been traditionally used for centuries to treat numerous diseases. Current studies focus on the antidiabetic, anticancer, antibacterial, antifungal, anti-inflammatory, antiulcer, immune-enhancing, and hypoglycemic effects of these seeds. In this study, it was aimed to synthesize titanium nanoparticles by green synthesis method using N. sativa seeds and to investigate the effects of these nanoparticles on cytotoxicity, antibacterial, and wound healing activity. N. sativa seeds were extracted with distilled water and titanium nanoparticles were synthesized using titanium (IV) dioxide precursor. UV-VIS, SEM-EDX, FTIR, and XRD were used to characterize the nanoparticles. Then, the cytotoxic effects of the nanoparticles on HT-29 cells and wound healing effects on BJ and Caco-2 cells were investigated. Also, antibacterial activities were tested using Escherichia coli and Staphylococcus aureus. The cytotoxicity analysis demonstrated that cell viability decreased in a time- and dose-dependent manner. The lowest IC50 value was obtained at 72 hours with a value of 61.10±3.33 µg/mL. In the scratch assay, it was determined that 2.5 and 5 µg/mL titanium nanoparticle application showed no positive effect on wound healing. The MIC value for both bacteria was calculated as 0.125 mg/mL, and the inhibition zone diameters were measured as 10.81 mm against E. coli and 10.52 mm against S. aureus bacteria. This study has shown that titanium nanoparticles produced using N. sativa have potential for use in health and biomedical fields.

: This study evaluated the antibacterial effects of a mixture of Sophorae radix and Glycyrrhiza uralensisFischer (1 : 1) ethanol extracts (SGE) on mice infected with Streptococcus (S.) pyogenes. The minimum inhibitoryconcentration (MIC) and minimum bactericidal concentration of SGE necessary for antibacterial effects against S.pyogenes were 20µg/mL. Based on the time-kill curves for S. pyogenes, SGE was effective at 4× MIC after 16 h.On Day 12 after challenge, the survival rate of mice treated with 2.0 mg/kg SGE was 60%. In conclusion, SGE hadpotent in vitro and in vivo antibacterial activities against S. pyogenes.Keywords : crude extracts, medicinal plants, mice, Streptococcus pyogenes, treatment efficacy Streptococcus (S.) pyogenes, a group A streptococcus, is anaerobic, spherical, gram-positive extracellular bacterium thatis associated with a variety of mucosal and invasive humaninfections [3]. These bacteria cause various diseases rangingfrom mild and quite frequent non-invasive infections of theupper respiratory tract and skin to severe invasive infectionsthat include necrotizing fasciitis and streptococcal toxicshock syndrome [4]. The most important modes of S. pyogenes transmission arevia respiratory droplets, hand contact with nasal dischargeand skin contact with impetigo lesions [1]. In addition, S.pyogenes can be spread to cattle and then back to humansthrough raw milk as well as contaminated food sources suchas salads, milk and eggs [7]. In a previous study [16] that analyzed for antimicrobialsusceptibility against S. pyogenes isolates in scarlet feverpatients between 2003 and 2011 in China, 100, 97.0, and89.4% of the 74 S. pyogenes isolates were found to be resis-tant to erythromycin, clindamycin and tetracycline, respec-tively. The emergence of antibiotic resistance has led to thesearch for new, safe and effective antimicrobial agents fromalternative natural resources. Medicinal herbs as alternativesto antibiotics are attracting considerable attention by manyresearchers, and many such herbs have a long history ofmedicinal use in Asia [14]. Many of these herbs are oftenused in combination to increase their effects. These herbscontaining bioactive components have many potential clini-cal and therapeutic applications in modern medical care [11].In addition, antimicrobial agents can be derived from herbs,and more than 1,340 plants have been reported to have anti-microbial effects [15].Sophora (S.) flavescens which has antibacterial, antiinflam-matory, antipyretic, antiulcerative and antineoplastic effectshas been used traditionally as medicinal herbs in the treat-ment of jaundice, leucorrhea, carbuncles, pyogenic infec-tions of the skin, scabies, enteritis, and dysentery [8, 9].Glycyrrhiza (G.) uralensis Fischer is one of the most gener-ally used herbal medicines in the world and has been provedto possess antiinflammatory, liver protection, antibacterial,and anticancer activity, which are attributed to the containedtriterpenoid saponins and flavonoids [5]. This study evaluated the antibacterial activity of a combi-nation of ethanol extracts from the medicinal herbs, S. flave-scens and G. uralensis Fischer, against S. pyogenes. Inaddition, a preparation of the herbs was fed to mice infectedwith S. pyogenes in order to determine its therapeutic poten-