Role of cell wall components of Gram-positive bacteria in bacterial biofilm formation

Gram-positive bacteria;lipoteichoic acid;lipoprotein;biofilm;Lactobacillus;617.6

면역 및 분자미생물 ;

University:서울대학교 대학원

关键词: Gram-positive bacteria;    lipoteichoic acid;    lipoprotein;    biofilm;    Lactobacillus;    617.6;   

Others  :  http://s-space.snu.ac.kr/bitstream/10371/161687/1/000000157903.pdf

美国|英语

来源: Seoul National University Open Repository

【 摘 要 】

ObjectivesStreptococcus gordonii and Enterococcus faecalis are Gram-positive facultative anaerobes present in the mucosal tissues of the human body such as oral cavity, gastrointestinal tracts, and genital tracts. Recently, these bacteria have been reported to cause inflammatory diseases such as refractory apical periodontitis and endocarditis through opportunistic infections. These oral pathogens can form biofilms in the oral cavity, which contribute to chronic inflammatory diseases by having physical and chemical defenses against antibiotics and host immune system during biofilm formation. Although biofilm formation is important for oral pathology, the role of cell wall components of oral pathogen in biofilm formation has not been clearly understood. Treatment of biofilm-associated oral diseases by current antibiotics has limitations since biofilms are resistant to antibiotics and immune cells. Therefore, the development of new effective anti-biofilm agents that prevent and suppress the biofilm formation of oral pathogens are needed. Recently, Lactobacillus species inhibiting bacterial biofilm formation have been suggested as an anti-biofilm agent. However, there is still a lack of study about what components of Lactobacillus species inhibit biofilm formation. Therefore, the aims of this study were (1) to determine the role of cell wall components of Streptococci in the biofilm formation, and (2) to evaluate which cell wall components of Lactobacillus species inhibit E. faecalis biofilm.MethodsThe effects of bacterial cell wall components on biofilm formation were investigated using lipoprotein-deficient strain (Δlgt) and lipoteichoic acid -deficient strain (ΔltaS) of Streptococcus gordonii, Streptococcus mutans, Streptococcus pneumoniae, and Staphylococcus aureus. The biofilm formation was examined by a crystal violet assay and the bacterial growth was examined by measuring the optical density at 600 nm (OD600). AI-2 mRNA expression was examined by reverse transcription-polymerase chain reaction (RT-PCR). The effects of L. plantarum lipoprotein (Lp.Lpp), lipoteichoic acid (Lp.LTA), and peptidoglycan (Lp.PGN) on biofilm formation were investigated by crystal violet assay, confocal laser scanning microscopy (CLSM) using a LIVE/DEAD viability assay. Biofilm on human dentin slices were visualized with a scanning electron microscopy (SEM).ResultsΔlgt of S. gordonii showed higher biofilm formation compared to wild-type strain. In addition, Δlgt of other Streptococci including S. mutans and S. pneumoniae showed augmented biofilm formation in common with S. gordonii, but S. aureus did not. But the growth rate of Δlgt and ΔltaS was not different from that of the wild-type. In addition, S. gordonii Δlgt showed upregulation of mRNA expression of luxS and increased AI-2 level compared to wild-type. L. plantarum culture supernatants containing cell wall components inhibited biofilm formation of E. faecalis. Lp.LTA most effectively inhibited biofilm formation of E. faecalis in a dose-dependent manner among cell wall components of L. plantarum including Lp.LTA, Lp.Lpp, and Lp.PGN. When E. faecalis was incubated with Lp.LTA, the inhibitory effect was first observed at 1 h and lasted up to 24 h. Notably, Lp.LTA did not affect bacterial growth. In addition, purified LTA form various Lactobacillus species including Lactobacillus casei, Lactobacillus rhamnosus GG, Lactobacillus acidophilus inhibited and disrupted biofilm formation of E. faecalis. Among these LTA, Lp.LTA show the most potent inhibition effect on E. faecalis biofilm. D-Alanine is a key component of Lp.LTA for its inhibitory effects on the biofilm formation of E. faecalis based on the result that D-alanine-removed Lp.LTA failed to inhibit biofilm formation. Furthermore, Lp.LTA inhibited clinical isolates of E. faecalis and E. faecalis biofilm on human dentin slices.ConclusionsTaken together, the results show that lipoprotein-deficient Streptococci form biofilms much potently than the wild-type, which might be related with quorum sensing mechanism including AI-2. Lp.LTA effectively inhibits the biofilm formation of E. faecalis and disrupts preformed biofilms implying that LTA from Lactobacillus can be used as an anti-biofilm agent to prevent and treat biofilm-associated diseases in oral cavity.