Department of Microbiology and Immunology, Uniformed Services University of the Health Sciencesgrid.265436.0, Bethesda, Maryland, USA.
Henry M. Jackson Foundation for the Advancement of Military Medicine, Rockville, Maryland, USA.
Antimicrob Agents Chemother. 2022 May 17;66(5):e0233921. doi: 10.1128/aac.02339-21. Epub 2022 Apr 21.
Enterococcus faecalis, a leading cause of health care-associated infections, forms biofilms and is resistant to many antimicrobial agents. Planktonic-phase E. faecalis is resistant to high concentrations of the enzyme lysozyme, which catalyzes the hydrolysis of -acetylmuramic acid and -acetylglucosamine linkages in peptidoglycan and is also a cationic antimicrobial peptide (CAMP). E. faecalis lysozyme resistance in planktonic cells is stimulated upon activation of the extracytoplasmic function sigma factor SigV via cleavage of the anti-sigma factor RsiV by the transmembrane protease Eep. Planktonically grown E. faecalis lacking is more sensitive than wild-type strains to growth inhibition by lysozyme. This study was initiated to determine whether E. faecalis OG1RFΔ biofilms would be protected from lysozyme. Serendipitously, we discovered that exposure of both E. faecalis OG1RF and OG1RFΔ biofilms to chicken egg white lysozyme resulted in decreases in biofilm cell viability of 3.7 and 3.8 log CFU/mL, respectively. Treatment of biofilms of both strains with recombinant purified human lysozyme was associated with reductions in cell viability of >99.9% for both strains. Lysozyme-treated OG1RF and OG1RFΔ biofilms contained a higher percentage of dead cells by Live/Dead staining and were associated with more extracellular DNA. Heat-inactivated human lysozyme, which was devoid of muramidase activity, as well as the lysozyme-derived CAMP LP9 and the CAMP polymyxin B, decreased biofilm cell viability. These results are consistent with a model in which the CAMP activity, rather than the muramidase activity, of lysozyme causes lysis of E. faecalis biofilm cells despite them having an intact lysozyme resistance-inducing signaling pathway. Finally, lysozyme was also effective in reducing viable biofilm cells of several other E. faecalis strains, including the vancomycin-resistant strain V583 and multidrug-resistant strain MMH594. This study demonstrates the potential for lysozyme to be developed as a novel antibiofilm therapeutic.
屎肠球菌是一种主要的医源性感染病原体，能够形成生物膜并对许多抗菌药物具有耐药性。浮游期屎肠球菌能够抵抗高浓度的溶菌酶，该酶能够催化肽聚糖中 N-乙酰胞壁酸和 N-乙酰葡萄糖胺键的水解，同时也是一种阳离子抗菌肽（CAMP）。通过跨膜蛋白酶 Eep 对反西格玛因子 RsiV 的切割，浮游期细胞中外质功能 sigma 因子 SigV 的激活会刺激屎肠球菌溶菌酶的抗性。与野生型菌株相比，缺乏 的浮游期屎肠球菌对溶菌酶的生长抑制更为敏感。本研究旨在确定屎肠球菌 OG1RFΔ生物膜是否能够免受溶菌酶的保护。偶然的是，我们发现，鸡卵清白溶菌酶处理屎肠球菌 OG1RF 和 OG1RFΔ生物膜都会导致生物膜细胞活力分别降低 3.7 和 3.8 log CFU/mL。用重组纯化的人溶菌酶处理两种菌株的生物膜，均会导致两株菌的细胞活力降低超过 99.9%。用溶菌酶处理的 OG1RF 和 OG1RFΔ生物膜中活/死染色的死细胞比例更高，且含有更多的胞外 DNA。缺乏裂解酶活性的热失活人溶菌酶、溶菌酶衍生的 CAMP LP9 和 CAMP 多粘菌素 B 也会降低生物膜细胞活力。这些结果与一种模型一致，即尽管屎肠球菌具有完整的溶菌酶诱导信号通路，但溶菌酶的 CAMP 活性而不是裂解酶活性会导致生物膜细胞的裂解。最后，溶菌酶也能有效减少其他几种屎肠球菌菌株的生物膜活菌，包括万古霉素耐药菌株 V583 和多药耐药菌株 MMH594。本研究表明，溶菌酶具有开发为新型抗生物膜治疗药物的潜力。
