Ołdak Ewa, Trafny Elzbieta A
Department of Microbiology and Epidemiology, Military Institute of Hygiene and Epidemiology, Kozielska 4, 01-163, Warsaw, Poland.
Antimicrob Agents Chemother. 2005 Aug;49(8):3281-8. doi: 10.1128/AAC.49.8.3281-3288.2005.
Pseudomonas aeruginosa proteases are considered important virulence factors which damage host tissues and interfere with host antibacterial defense mechanisms. P. aeruginosa biofilm cells are not completely killed by antibacterials, and therefore this study addresses the question whether ciprofloxacin attenuates the virulence of biofilm communities by abolishing their secretion of proteases. The surviving cells of the colony biofilms studied, despite their cyclical exposure to four doses of ciprofloxacin at bactericidal concentrations (one dose a day), still secreted active proteases to the environment surrounding the biofilms. The biofilm cells secreted elastase B (LasB) over the duration of the experiments as confirmed by Western immunoblot analysis. The colony biofilms did not secrete LasA-a protease with staphylolytic activity. The same profiles on zymogram gels with gelatin were observed for the proteases secreted by both ciprofloxacin-exposed and unexposed (control) biofilms. Total proteolytic activities of the colony biofilms studied were significantly reduced after exposure to ciprofloxacin at bactericidal concentrations-after 96 h of exposure they dropped to 38% for the strain intermediate resistant to ciprofloxacin and to 65% for the strain highly resistant to the antibiotic, relative to the control biofilms. The surviving cells of the colony biofilms after their release into a fresh medium displayed transient increased resistance to ciprofloxacin compared to their planktonic counterparts.
铜绿假单胞菌蛋白酶被认为是重要的毒力因子，可损害宿主组织并干扰宿主的抗菌防御机制。铜绿假单胞菌生物膜细胞不会被抗菌药物完全杀死，因此本研究探讨了环丙沙星是否通过消除其蛋白酶分泌来减弱生物膜群落的毒力这一问题。尽管所研究的菌落生物膜存活细胞每天接受一剂杀菌浓度的四剂环丙沙星周期性处理，但它们仍向生物膜周围环境分泌活性蛋白酶。如蛋白质免疫印迹分析所证实，生物膜细胞在实验期间持续分泌弹性蛋白酶B（LasB）。菌落生物膜不分泌具有葡萄球菌溶解活性的蛋白酶LasA。环丙沙星处理组和未处理组（对照组）生物膜分泌的蛋白酶在明胶酶谱凝胶上呈现相同的图谱。在接受杀菌浓度的环丙沙星处理后，所研究的菌落生物膜的总蛋白水解活性显著降低——暴露96小时后，对环丙沙星中度耐药的菌株降至38%，对该抗生素高度耐药的菌株降至65%，相对于对照生物膜而言。菌落生物膜的存活细胞在释放到新鲜培养基后，与浮游对应细胞相比，对环丙沙星的耐药性短暂增加。
