| 1. Background and Objective:Dental caries, one of the most prevalent oral infectious diseases, is caused by a cluster of bacteria involved in the progression of initial plaque biofilms formation and subsequent cariogenesis[1]. Nearly 90 percent of teenagers and young adults are suffering from or had dental caries in USA[2]. A suitable carbohydrate substrate, micro-organisms in dental plaque, a susceptible tooth surface and time are the main factors of dental caries[3]. A variety of bacteria and proteins adhere to tooth surfaces, and biofilm is formed after a period of time. Biofilm structure can not only provides shelter for bacteria, but also increase the resistance of bacteria to antimicrobial agent[4].Thus, the effective dental plaque control is the key to the prevention and treatment of dental caries. The anti-caries capacity of fluoride is determined, but it has several difficulties of popularizing in China[5]. Bacteria are easily developed resistance to traditional antibiotics. Therefore, the development of novel drugs for microorganism is needed.Antimicrobial peptides are the focus of recent research in oral microbiology. Antimicrobial peptides which characterized by small molecular weight, water solubility, heat resistance, non-immunogenic, a broad spectrum antibiotic, are likely to be highly efficient and low toxicity agents [6]. Pleurocidin, α-helix cationic peptides, is extracted from Pleuronectes American. But pleurocidin is high cost in synthesis because it is consisted of 25 amino acids, which also reduces the ability of penetrating biofilms. So it is urgent in developing new peptides. Our research designed pm11, a novel antibacterial peptide, which based on pleurocidin, and verify its bactericidal effect.2. Methods:2.1 Analyzing the effect of pm11 on oral microorganisms using the method of minimal inhibitory concentration(MIC), minimal bactericide concentration(MBC).2.2 Analyzing the effect of pm11 on oral microorganisms by bactericidal kinetics.2.3 Observing the antimicrobial activity of pm11 in saliva environment by inhibition zone in soft agar.2.4 Cultivating biofilm and divided into three groups, the PBS-treated group as a negative control, the positive control group was treated with isopropanol, pm11 treated group as the experimental group, then confocal laser scanning microscopy(CLSM) was performed to analyze the dye of three groups.2.5 Bacteria was divided into two groups, the PBS-treated group as a negative control, pm11 treated group as the experimental group, and scanning electron microscopy(SEM) was performed to observe the change of bacterial membrane.3. Results:3.1 The antimicrobial activity of pm11 varied greatly against different oral microorganisms. MIC values ranged from 2μg/ml to 256μg/ml, and MBC values from 2 μg/ml to higher than 256μg/ml.3.2 The bactericidal kinetics results showed that the more interaction time,the lower surviving rate of bacteria in two hours.3.3 The inhibitory-zone diameters showed a statistical indifference(P<0.05) between the water solution and the sterile saliva solution. And the higher concentration could enhance pm11 antimicrobial activities.3.4 The CLSM results showed dead bacteria mumber increased obviously in pm11- treated group, and dead bacteria had a trend of spreading to the biological membrane inside.3.5 The SEM results showed the shape of bacteria membrane was changed when treated with pm11.4. Conclusion:4.1 Pm11 has a broad spectrum of antimicrobial activities on oral microorganisms,and the antimicrobial effect of pm11 depend on time and concentration.4.2 Pm11 still had antimicrobial effect on S. mutans biofilm.4.3 Pm11 could disrupting the cell membrane of bacteria, leading to out flowing of bacteria contents, which resulted in the bacterial death. |
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