Data_Sheet_1_Bacillus subtilis natto Derivatives Inhibit Enterococcal Biofilm Formation via Restructuring of the Cell Envelope.PDF

Enterococcus faecalis is considered a leading cause of hospital-acquired infections. Treatment of these infections has become a major challenge for clinicians because some E. faecalis strains are resistant to multiple clinically used antibiotics. Moreover, the presence of E. faecalis biofilms can make infections with E. faecalis more difficult to eradicate with current antibiotic therapies. Thus, our aim in this study was to investigate the effects of probiotic derivatives against E. faecalis biofilm formation. Bacillus subtilis natto is a probiotic strain isolated from Japanese fermented soybean foods, and its culture fluid potently inhibited adherence to Caco-2 cell monolayers, aggregation, and biofilm production without inhibiting the growth of E. faecalis. An apparent decrease in the thickness of E. faecalis biofilms was observed through confocal laser scanning microscopy. In addition, exopolysaccharide synthesis in E. faecalis biofilms was reduced by B. subtilis natto culture fluid treatment. Carbohydrate composition analysis also showed that carbohydrates in the E. faecalis cell envelope were restructured. Furthermore, transcriptome sequencing revealed that the culture fluid of B. subtilis natto downregulated the transcription of genes involved in the WalK/WalR two-component system, peptidoglycan biosynthesis and membrane glycolipid biosynthesis, which are all crucial for E. faecalis cell envelope synthesis and biofilm formation. Collectively, our work shows that some derivatives present in the culture fluid of B. subtilis natto may be useful for controlling E. faecalis biofilms.

粪肠球菌（Enterococcus faecalis）被认为是医院获得性感染的主要致病菌。此类感染的治疗已成为临床医师面临的重大挑战，因为部分粪肠球菌菌株对多种临床常用抗生素存在耐药性。此外，粪肠球菌生物被膜（biofilms）的存在会使粪肠球菌感染更难通过当前的抗生素疗法予以清除。因此，本研究旨在探究益生菌衍生物对抗粪肠球菌生物被膜形成的效果。纳豆芽孢杆菌（Bacillus subtilis natto）是一种从日本发酵大豆食品中分离得到的益生菌菌株，其培养液可有效抑制粪肠球菌对Caco-2细胞单层的黏附、聚集以及生物被膜的生成，且不会抑制粪肠球菌的生长。通过激光共聚焦扫描显微镜（confocal laser scanning microscopy）观察可见，粪肠球菌生物被膜的厚度显著降低。此外，经纳豆芽孢杆菌培养液处理后，粪肠球菌生物被膜中的胞外多糖（exopolysaccharide）合成水平有所下降。碳水化合物组成分析结果显示，粪肠球菌细胞包膜中的碳水化合物结构发生了重塑。进一步的转录组测序（transcriptome sequencing）结果表明，纳豆芽孢杆菌培养液可下调与WalK/WalR双组分系统（WalK/WalR two-component system）、肽聚糖生物合成（peptidoglycan biosynthesis）以及膜糖脂生物合成（membrane glycolipid biosynthesis）相关的基因转录，而这些通路均对粪肠球菌的细胞包膜合成及生物被膜形成至关重要。综上，本研究表明纳豆芽孢杆菌培养液中的部分衍生物可用于防控粪肠球菌生物被膜。