luxS/AI-2密度感应对缓症链球菌生物膜致病力的影响
Biofilm formation and antimicrobial susceptibility of Streptococcus mitis in response to luxS/AI-2 signaling

摘要： 目的 了解luxS基因敲除,自体诱导物-2（AI-2）密度感应信号缺失对缓症链球菌生物膜致病力表型的影响。方法 常规细菌培养、药敏试验、形态学与生化反应鉴定,16S 核糖体DNA（rDNA）序列同源性分析,获得耐药性缓症链球菌临床分离株。同源重组法敲除luxS基因,构建突变株。哈氏弧菌BB170菌株生物发光实验检测AI-2信号;定量分析临床分离株与突变株生物膜形成量与抗菌药敏感性的差异;荧光黄染液、18909荧光增白染液、L7012死/活菌染液染色,激光共聚焦显微镜扫描,了解生物膜结构、胞外多糖、死/活菌分布差异;临床分离株上清液、4,5-二羟基-2,3-戊二酮（DPD）补偿生长实验确认突变株表型改变的原因。实验数据行Kolmogorov-Smirno非参数检验。结果 luxS基因突变株与临床分离株生物膜形成量存在显著性差异（P<0.001）;氨苄西林、环丙沙星、四环素干预条件下,突变株生物膜形成量显著降低。上清液、DPD补偿后,突变株生物膜形成能力和结构可以恢复。突变株生物膜结构变疏松,对氨苄西林、环丙沙星、四环素的敏感性增加。结论 luxS/AI-2密度感应信号缺失,导致耐药性缓症链球菌生物膜形成量下降,抗菌药敏感性增加,致病力表型发生改变。
Abstract: Objective This paper aims to study the influence of luxS gene mutation of Streptococcus mitis on biofilm formation and antimicrobial susceptibility. Methods Culture in blood plate, Gram stain and susceptibility tests, and phy-siological and biochemical tests were first carried out. 16S ribosomal DNA(rDNA) sequence homology analysis of clinical isolate was examined subsequently. The autoinducer-2(AI-2) bioluminescence assay of Vibrio harveyi BB170 strain, multi-drug resistance, and ability of biofilm formation were examined in luxSmutant and clinical isolate. The structure, exopo-lysaccharides, and dead/live cells of biofilm were scanned by confocal laser scanning microscope. Compared with one another, luxS mutant growth recruited by isolate supernatant or 4,5-dihydroxy-2,3-pentanedione(DPD) was experimented. Results luxS gene deletion decreased biofilm formation(P<0.001), loosened structure, and changed antimicrobial suscepti-bility(to ampicillin, ciprofloxacin, tetracycline). However, isolate supernatant or DPD can recover the structure and ability of biofilm formation in luxS mutant. Conclusion luxS/AI- 2 signaling can influence biofilm formation and antimicro-bial susceptibility of S. mitis with multidrug resistance.
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