Francisco Vassiliepe Sousa Arruda

Casbane diterpene as a promising natural antimicrobial agent against biofilm-associated infections

Croton nepetaefolius is a native plant from northeastern Brazil that belongs to the Euphorbiaceae family. The biological action of this plant has been extensively explored, being the secondary metabolites responsible for its properties alkaloids, diterpenes, and triterpenes. This study aimed to evaluate the ability of casbane diterpene (CD), isolated from the ethanolic extract of C. nepetaefolius, to inhibit microbial growth and biofilm formation of several clinical relevant species (bacteria and yeasts). It was found that CD possessed biocidal and biostatic activity against the majority of the species screened, with minimal active concentrations ranging between 125 and 500 µg/mL. In addition, it was observed that biofilm formation was inhibited even when the planktonic growth was not significantly affected. In conclusion, CD showed potential to be a natural tool for the treatment of diseases caused by different infectious microorganisms.

Antimicrobial activity of the synthetic peptide Lys-a1 against oral streptococci

The peptide LYS-[TRP(6)]-Hy-A1 (Lys-a1) is a synthetic derivative of the peptide Hy-A1, initially isolated from the frog species Hypsiboas albopunctatus. According to previous research, it is a molecule with broad antimicrobial activity. The objective of this study was to evaluate the antimicrobial activity of the synthetic peptide Lys-a1 (KIFGAIWPLALGALKNLIK-NH2) on the planktonic and biofilm growth of oral bacteria. The methods used to evaluate antimicrobial activity include the following: determination of the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) in microtiter plates for growth in suspension and quantification of biomass by crystal violet staining and counting of colony forming units for biofilm growth. The microorganisms Streptococcus oralis, Streptococcus sanguinis, Streptococcus parasanguinis, Streptococcus salivarius, Streptococcus mutans and Streptococcus sobrinus were grown in Brain Heart Infusion broth at 37°C under atmospheric pressure with 10% CO2. The peptide was solubilized in 0.1% acetic acid (v/v) at various concentrations (500-1.9 μg mL(-1)). Chlorhexidine gluconate 0.12% was used as the positive control, and BHI culture medium was used as the negative control. The tested peptide demonstrated a remarkable antimicrobial effect, inhibiting the planktonic and biofilm growth of all strains tested, even at low concentrations. Thus, the peptide Lys-a1 is an important source for potential antimicrobial agents, especially for the control and prevention of microbial biofilms, which is one of the most important factors in cariogenic processes.

Effect of Lectins from Diocleinae Subtribe against Oral Streptococci

Surface colonization is an essential step in biofilm development. The ability of oral pathogens to adhere to tooth surfaces is directly linked with the presence of specific molecules at the bacterial surface that can interact with enamel acquired pellicle ligands. In light of this, the aim of this study was to verify inhibitory and antibiofilm action of lectins from the Diocleinae subtribe against Streptococcus mutans and Streptococcus oralis. The inhibitory action against planctonic cells was assessed using lectins from Canavaliaensi formis (ConA), Canavalia brasiliensis (ConBr), Canavalia maritima (ConM), Canavalia gladiata (CGL) and Canavalia boliviana (ConBol). ConBol, ConBr and ConM showed inhibitory activity on S. mutans growth. All lectins, except ConA, stimulated significantly the growth of S. oralis. To evaluate the effect on biofilm formation, clarified saliva was added to 96-well, flat-bottomed polystyrene plates, followed by the addition of solutions containing 100 or 200 µg/mL of the selected lectins. ConBol, ConM and ConA inhibited the S. mutans biofilms. No effects were found on S. oralis biofilms. Structure/function analysis were carried out using bioinformatics tools. The aperture and deepness of the CRD (Carbohydrate Recognition Domain) permit us to distinguish the two groups of Canavalia lectins in accordance to their actions against S. mutans and S. oralis. The results found provide a basis for encouraging the use of plant lectins as biotechnological tools in ecological control and prevention of caries disease.

Effect of Algae and Plant Lectins on Planktonic Growth and Biofilm Formation in Clinically Relevant Bacteria and Yeasts

This study aimed to evaluate the abilities of plant and algae lectins to inhibit planktonic growth and biofilm formation in bacteria and yeasts. Initially, ten lectins were tested on Staphylococcus epidermidis, Staphylococcus aureus, Klebsiella oxytoca, Pseudomonas aeruginosa, Candida albicans, and C. tropicalis at concentrations of 31.25 to 250 μg/mL. The lectins from Cratylia floribunda (CFL), Vatairea macrocarpa (VML), Bauhinia bauhinioides (BBL), Bryothamnion seaforthii (BSL), and Hypnea musciformis (HML) showed activities against at least one microorganism. Biofilm formation in the presence of the lectins was also evaluated; after 24 h of incubation with the lectins, the biofilms were analyzed by quantifying the biomass (by crystal violet staining) and by enumerating the viable cells (colony-forming units). The lectins reduced the biofilm biomass and/or the number of viable cells to differing degrees depending on the microorganism tested, demonstrating the different characteristics of the lectins. These findings indicate that the lectins tested in this study may be natural alternative antimicrobial agents; however, further studies are required to better elucidate the functional use of these proteins.

Effect of the lectin of Bauhinia variegata and its recombinant isoform on surgically induced skin wounds in a murine model.

Lectins are a structurally heterogeneous group of highly specific carbohydrate-binding proteins. Due to their great biotechnological potential, lectins are widely used in biomedical research. The purpose of the present study was to evaluate the healing potential of the lectin of Bauhinia variegata (nBVL) and its recombinant isoform (rBVL-1). Following surgical creation of dorsal skin wounds, seven groups of mice were submitted to topical treatment for 12 days with lectin, d-galactose, BSA and saline. The animals were anesthetized and euthanized on POD 2, 7 and 12 in order to evaluate the healing potential of each treatment. The parameters considered included wound size, contraction rate, epithelialization rate and histopathological findings. Wound closure was fastest in animals treated with rBVL-1 (POD 7). nBVL was more effective than the controls. All skin layers were reconstructed and keratin deposition increased. Our findings indicate that the lectin of Bauhinia variegata possesses pro-healing properties and may be employed in the treatment of acute skin wounds.

Immunostimulatory activity of ConBr: a focus on splenocyte proliferation and proliferative cytokine secretion

Lectins constitute a class of glycoproteins, which are capable of selectively and reversibly binding to carbohydrates, distinguishing small structural differences in complex oligosaccharides. Studies have shown that the binding of lectins to cell-surface carbohydrates can lead to various effects such as cellular proliferation, histamine release and cytokine production. Canavalia brasiliensis lectin (ConBr) is a (D-mannose) D-glucose lectin. In this study, murine splenocytes were cultured to determine the effect of ConBr on cell proliferation, nitric oxide (NO) release and cytokine secretion. In addition, cellular viability assays were performed to evaluate any mitogenic activity induced by this lectin. ConBr significantly increased cell proliferation with minimal cell damage. This lectin was able to induce an increased production of cytokines such as IL-2, IL-6 and IFN-γ and a decreased production of IL- 10. The release of NO was also observed. The results of this study indicate that ConBr could potentially be used as an immunomodulator.

Correlation between Enterococcus faecalis biofilms development stage and quantitative surface roughness using atomic force microscopy.

Biofilms are assemblages of microorganisms and their associated extracellular products at an interface and typically with an abiotic or biotic surface. The study of the morphology of biofilms is important because they are associated with processes of biofouling, corrosion, catalysis, pollutant transformation, dental caries, drug resistance, and so forth. In the literature, biofilms have been examined by atomic force microscopy (AFM), which has proven to be a potent tool to study different aspects of the biofilm development on solid surfaces. In this work, we used AFM to investigate topographical changes during the development process of Enterococcus faecalis biofilms, which were generated on sterile cellulose nitrate membrane (CNM) filters in brain heart infusion (BHI) broth agar blood plates after 24, 36, 72, 192, and 360 h. AFM height images showed topographical changes due to biofilm development, which were used to characterize several aspects of the bacterial surface, such as the presence of extracellular polymeric substance, and the biofilm development stage. Changes in the development stage of the biofilm were shown to correlate with changes in the surface roughness as quantified through the mean roughness.

Biological Applications of Plants and Algae Lectins: An Overview

More than 120 years ago, Peter Hermann Stillmark in his doctoral thesis presented in 1888 to the University of Dorpat, gave the earliest step in the study of proteins that have a very interesting feature: the ability to agglutinate erythrocytes. These proteins were initially referred as to hemagglutinins or phytoagglutinins, since they were originally isolated from extracts of plants [1]. The first hemagglutinin isolated by Stillmark was extracted from seeds of the castor tree (Riccinus communis) and was named ricin [2]. This hemagglutinin was strongly used by Paul Ehrlich as model antigens for immunological studies [2,3].

Purification, characterization and partial sequence of a pro-inflammatory lectin from seeds of Canavalia oxyphylla Standl. & L. O. Williams.

Recent studies have shown that lectins are promising tools for use in various biotechnological processes, as well as studies of various pathological mechanisms, isolation, and characterization of glycoconjugates and understanding the mechanisms underlying pathological mechanisms conditions, including the inflammatory response. This study aimed to purify, characterize physicochemically, and predict the biological activity of Canavalia oxyphylla lectin (CoxyL) in vitro and in vivo. CoxyL was purified by a single‐step affinity chromatography in Sephadex® G‐50 column. Sodium dodecyl sulfate polyacrylamide gel electrophoresis showed that the pure lectin consists of a major band of 30 kDa (α‐chain) and two minor components (β‐chain and γ‐chain) of 16 and 13 kDa, respectively. These data were further confirmed by electrospray ionization mass spectrometry, suggesting that CoxyL is a typical ConA‐like lectin. In comparison with the average molecular mass of α‐chain, the partial amino acid sequence obtained corresponds to approximately 45% of the total CoxyL sequence. CoxyL presented hemagglutinating activity that was specifically inhibited by monosaccharides (D‐glucose, D‐mannose, and α‐methyl‐D‐mannoside) and glycoproteins (ovalbumin and fetuin). Moreover, CoxyL was shown to be thermostable, exhibiting full hemagglutinating activity up to 60°C, and it was pH‐sensitive for 1 h, exhibiting maximal activity at pH 7.0. CoxyL caused toxicity to Artemia nauplii and induced paw edema in rats. This biological activity highlights the importance of lectins as important tools to better understand the mechanisms underlying inflammatory responses. Copyright

Characterization of Isoforms of the Lectin Isolated from the Red Algae Bryothamnion seaforthii and Its Pro-Healing Effect

Lectins are a structurally heterogeneous group of proteins that have specific binding sites for carbohydrates and glycoconjugates. Because of their biotechnological potential, lectins are widely used in biomedical research. The present study aimed to evaluate the healing potential of the lectin isolated from the marine red alga Bryothamnion seaforthii (BSL). The lectin was purified using ion exchange chromatography with DEAE cellulose and characterized using tandem mass spectrometry. For healing tests, skin wounds were induced in the dorsal thoracic region of mice. These animals were randomly divided into three groups and subjected to topical treatment for 12 days with BSL, bovine serum albumin and 150 mM NaCl. To evaluate the potential of each treatment, the animals were anesthetized and sacrificed on days 2, 7 and 12, respectively. The parameters evaluated included the wound area, the proportion of wound closure and the histological diagnosis. The wound closure was more effective with BSL (Postoperative Day 7 and 12) than controls. The luminal epithelium was completely restructured; the presence of collagen in the dermis and the strongly active presence of young skin annexes demonstrate the potential of treatment with BSL compared with controls. Our findings suggest that BSL has pro-healing properties and can be a potential medical process in the treatment of acute wounds.