Effect of Rubusoside, a Natural Sucrose Substitute, on Streptococcus mutans Biofilm Cariogenic Potential and Virulence Gene Expression In Vitro.

Dental caries is a biofilm-mediated disease in which Streptococcus mutans is the main pathogenic microorganism, and its incidence is closely related to sucrose. Rubusoside is a natural nonnutritive sweetener isolated from Rubus suavissimus S. Lee. This study was designed to determine the effect of this sucrose substitute on the cariogenic properties and virulence gene expression of S. mutans biofilms. S. mutans was exposed to brain heart infusion (BHI) medium (as a control), 1% sucrose-supplemented medium, 1% rubusoside-supplemented medium, and 1% xylitol-supplemented medium. The growth curve of the biofilm was monitored by crystal violet staining, and the 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 amount of soluble proteins), numbers of CFU, and amounts of intra- and extracellular polysaccharides. Biofilm structural imaging was performed using a scanning electron microscope (SEM). Virulence gene expression (gtfB, gtfC, gtfD, ftf, spaP, gbpB, ldh, atpF, vicR, and comD) was determined by reverse transcription-quantitative PCR. Growth in rubusoside resulted in lower levels of acid production than observed during growth in sucrose, xylitol, and the control, while it also reduced the level of biofilm accumulation and bacterial viability and even reduced the level of production of extracellular polysaccharides. By SEM, the levels of biofilm formation and extracellular matrix during growth in rubusoside were lower than these levels during growth in sucrose and xylitol. From the perspective of virulence genes, growth in rubusoside and xylitol significantly inhibited the expression of virulence genes compared with their levels of expression after growth in sucrose. Among these genes, gtfB, gtfC, gbpB, ldh, and comD downregulation was found with growth in rubusoside compared with their expression with growth in xylitol. Therefore, rubusoside appears to be less potentially cariogenic than sucrose and xylitol and may become an effective sucrose substitute for caries prevention. Further studies are needed to deepen these findings.IMPORTANCE Dental caries is a major public health challenge and places heavy biological, social, and financial burdens on individuals and health care systems. To palliate the deleterious effect of sucrose on the virulence factors of S. mutans, massive commercial efforts have been oriented toward developing products that may act as sucrose substitutes. Rubusoside, a natural sucrose substitute, is a plant extract with a high level of sweetness. Although some studies have shown that rubusoside does not produce acids or inhibit the growth of S. mutans, little attention has been paid to its effect on dental biofilm and the underlying mechanisms. Our study focuses on the effect of rubusoside on the formation and structure of biofilms and the expression of virulence genes. The results confirm that rubusoside can inhibit accumulation, bacterial viability, polysaccharide production by the biofilm, and related gene expression. These results provide further insight into the cariogenicity of S. mutans biofilms and demonstrate a new perspective for studying the impact of sucrose substitutes on caries.

Silica nanoparticles containing nano-silver and chlorhexidine respond to pH to suppress biofilm acids and modulate biofilms toward a non-cariogenic composition.

OBJECTIVES: Dental caries is caused by acids from biofilms. pH-sensitive nanoparticle carriers could achieve improved targeted effectiveness. The objectives of this study were to develop novel mesoporous silica nanoparticles carrying nanosilver and chlorhexidine (nMS-nAg-Chx), and investigate the inhibition of biofilms as well as the modulation of biofilm to suppress acidogenic and promote benign species for the first time. METHODS: nMS-nAg was synthesized via a modified sol-gel method. Carboxylate group functionalized nMS-nAg (COOH-nMS-nAg) was prepared and Chx was added via electrostatic interaction. Minimal inhibitory concentration (MIC), inhibition zone, and growth curves were evaluated. Streptococcus mutans (S. mutans), Streptococcus gordonii (S. gordonii), and Streptococcus sanguinis (S. sanguinis) formed multispecies biofilms. Metabolic activity, biofilm lactic acid, exopolysaccharides (EPS), and TaqMan real-time polymerase chain reaction (RT-PCR) were tested. Biofilm structures and biomass were observed by scanning electron microscopy (SEM) and live/dead bacteria staining. RESULTS: nMS-nAg-Chx possessed pH-responsive properties, where Chx release increased at lower pH. nMS-nAg-Chx showed good biocompatibility. nMS-nAg-Chx exhibited a strong antibacterial function, reducing biofilm metabolic activity and lactic acid as compared to control (p < 0.05, n = 6). Moreso, biofilm biomass was dramatically suppressed in nMS-nAg-Chx groups. In control group, there was an increasing trend of S. mutans proportion in the multispecies biofilm, with S. mutans reaching 89.1% at 72 h. In sharp contrast, in nMS-nAg-Chx group of 25 µg/mL, the ratio of S. mutans dropped to 43.7% and the proportion of S. gordonii and S. sanguinis increased from 19.8% and 10.9 to 69.8% and 56.3%, correspondingly. CONCLUSION: pH-sensitive nMS-nAg-Chx had potent antibacterial effects and modulated biofilm toward a non-cariogenic tendency, decreasing the cariogenic species nearly halved and increasing the benign species approximately twofold. nMS-nAg-Chx is promising for applications in mouth rinse and endodontic irrigants, and as fillers in resins to prevent caries.

Characterisation and skin protection activities of polysaccharides from Schnabelia terniflora.

The bioactivities of crude polysaccharides from leaves (L-Ps) and flowers (F-Ps) of Schnabelia terniflora (Maxim.) P. D. Cantino were studied, and the characteristics of purified fractions were analysed by HPLC, HP-GPC and NMR. L-Ps exhibited strong DPPH radical scavenging activity (IC50 value of 251.53 ± 4.62 µg/mL) and tyrosinase inhibition (IC50 value of 163.52 ± 2.59 µg/mL). However, the maximum moisture absorption (74.67 ± 1.53%) and retention (68.00 ± 3.61%) abilities were observed in F-Ps. Two main fractions separated by DEAE-Sepharose fast flow column from L-Ps were eluted with 0.1 and 0.3 M NaCl, while one main fraction from F-Ps was eluted with 0.1 M NaCl. Purified fractions were obviously different in monosaccharide composition, molecular weight and 1H NMR and 13C NMR spectra. Therefore, the current manuscript can provide an important evidence for the potential development of L-Ps and F-Ps as promising ingredients in cosmetics industry.

The application of mesoporous silica nanoparticles as a drug delivery vehicle in oral disease treatment.

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.

Draft genome sequence of a metallo-ß-lactamase (blaAIM-1)-producing Klebsiella pneumoniae ST1916 isolated from a patient with chronic diarrhoea.

OBJECTIVES: Klebsiella pneumoniae colonisation of the human gastrointestinal tract is a significant risk factor for extraintestinal infections in severely ill patients. Recent reports have indicated the high rate of K. pneumoniae infection resulting from the patient's own gut microbiota. Here we report the draft genome sequence of a multidrug-resistant (MDR) K. pneumoniae strain (SM32) harbouring the blaAIM-1 gene isolated from a patient with chronic diarrhoea in China. METHODS: Whole genomic DNA was sequenced using an Illumina MiSeq platform. The generated reads were de novo assembled using SOAPdenovo v.2.04. All probable coding sequences were predicted by Glimmer v.3.02 and were annotated using information from GenBank, Pfam, Nr, COG, String, GO and KEGG. Resistance-related genes were also further identified. RESULTS: The draft genome of K. pneumoniae SM32, belonging to sequence type (ST) 1916, was assembled into 165 contigs comprising 5238542bp. A total of 5013 protein-coding sequences and several genes associated with resistance to ß-lactams, tetracycline, aminoglycosides, fluoroquinolones and trimethoprim/sulfamethoxazole were preliminarily identified. CONCLUSIONS: MDR K. pneumoniae colonising the human gastrointestinal tract provides a potential reservoir for extraintestinal infections. The genome sequence of K. pneumoniae SM32 will be helpful to reveal the key role of mobile genetic elements in the adaptive translocation and spread of antimicrobial resistance.

Secretory Expression and Characterization of an Acidic Endo-Polygalacturonase Gene from Aspergillus niger SC323 in Saccharomyces cerevisiae.

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.