In order to evaluate their antibacterial activities and toxicities, the cecropins-melittin hybrid antimicrobial peptide, CA(1-7)-M(4-11) (CAM) and CB(1-7)-M(4-11) (CBM), were designed by APD2 database. The recombinant hybrid antimicrobial peptides were successfully expressed and purified in Pichia pastoris. Antimicrobial activity assay showed that both of the two hybrid antimicrobial peptides had strong antibacterial abilities against Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, Klebsiella pneumoniae, Bacillus subtilis, Bacillus thuringiensis, and Salmonella derby. The potency of CAM and CBM to E. coli 25922 were 0.862 and 0.849, respectively, slightly lower than Amp's 0.957. The hemolytic assays indicated CAM and CBM had no hemolytic in vivo and in vitro, and so they had a good application prospect.
Dental caries is a biofilm-mediated disease with Streptococcus mutans as the main pathogenic microorganism, and its incidence is closely related to sucrose. Rubusoside is a natural non-nutritive sweetener isolated from Rubus suavissimu S.Lee. This study was designed to determine the effect of the sucrose substitute on cariogenic properties and virulence gene expression of S. mutans biofilms. S. mutans was exposed to: BHI medium (as a control), BHI+1% sucrose, BHI+1% rubusoside, and BHI +1% xylitol. The growth curve of biofilm was monitored with crystal violet staining and pH was measured every 24 h. After 5 days, the biofilms formed on the glass coverslips were recovered to determine the biomass (dry weight and total soluble proteins), colony-forming units, and intra- and extracellular polysaccharides. Biofilm structural imaging was observed using a scanning electron microscope (SEM). The virulence gene expression (gtfB, gtfC, gtfD, ftf, spaP, gbpB, ldh, atpF, vicR, and comD) was determined by rt-qPCR. Compared to growth in sucrose, xylitol and control, growth in rubusoside resulted in a lower acid production, while also reducing the biofilm accumulation, bacterial viability, and even reducing the production of extracellular polysaccharides. And less biofilm formation and extracellular matrix was observed through SEM in rubusoside than sucrose and xylitol. From the perspective of virulence genes, rubusoside and xylitol significantly inhibited expression of virulence genes compared with sucrose, among which, down-regulation of the gtfB, gtfC, gbpB, ldh, and comD was found in rubusoside than xylitol. Therefore, rubusoside appears to be less potential cariogenicity than sucrose and xylitol, and may become an effective sucrose substitute for caries prevention. Further studies need to deepen these findings. Importance Dental caries, as a major public health challenge, places a heavy biological, social and financial burden on individuals and health care systems. To palliate the deleterious effect of sucrose on the virulence factors of S. mutans, massive commercial products have oriented towards developing sucrose substitutes. Rubusoside, a natural sucrose substitute, is a plant extract with high sweetness. Although some studies have shown that rubusoside do not produce acids and inhibit the growth of S. mutans, little attention has been paid to dental biofilm and underlying mechanisms. Our study focuses on the effect of rubusoside on formation and structure of biofilm, and expression of virulence genes. Results confirms that rubusoside can inhibit accumulation, bacterial viability, polysaccharide production of biofilm, and inhibit related gene expression. These results provide further insight into the cariogenicity of S. mutans biofilm, and demonstrate a new perspective for studying the impact of sucrose substitutes on caries.
The endo-polygalacturonase gene (endo-pgaA) was cloned from DNA of Aspergillus niger SC323 using the cDNA synthesized by overlapping PCR, and successfully expressed in Saccharomyces cerevisiae EBY100 through fusing the α-factor signal peptide of yeast. The full-length cDNA consists of 1,113 bp and encodes a protein of 370 amino acids with a calculated molecular mass of 38.8 kDa. After induction by galactose for 48 h, the activity of recombinant endo-PgaA in the culture supernatant can reach up to 1,448.48 U/mg. The recombinant protein was purified to homogeneity by ammonium sulfate precipitation and gel filtration column chromatography and subsequently characterized. The optimal pH and temperature of the purified recombinant enzyme were 5.0 and 50°C, respectively. The Michaelis-Menten constant (Km) and maximal velocity (Vmax) of the enzyme for pectin were 88.54 μmol/ml and 175.44 μmol/mg/min, respectively. The enzyme activity was enhanced by Ca(2+), Cu(2+), and Na(+), and strongly inhibited by Pb(2+) and Mn(2+). The pectin hydrolysates were mainly galacturonic acid and other oligo-galacturonates. Therefore, these characteristics suggest that the recombinant endo-PgaA may be of potential use in the food and feed industries.
Mesoporous silica nanoparticles (MSNs) hold promise as safer and more effective medication delivery vehicles for treating oral disorders. As the drug’s delivery system, MSNs adapt to effectively combine with a variety of medications to get over systemic toxicity and low solubility issues. MSNs, which operate as a common nanoplatform for the co-delivery of several compounds, increase therapy effectiveness and show promise in the fight against antibiotic resistance. MSNs offer a noninvasive and biocompatible platform for delivery that produces long-acting release by responding to minute stimuli in the cellular environmen. MSN-based drug delivery systems for the treatment of periodontitis, cancer, dentin hypersensitivity, and dental cavities have recently been developed as a result of recent unparalleled advancements. The applications of MSNs to be embellished by oral therapeutic agents in stomatology are discussed in this paper.
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