Mechanisms of probiotic inhibition of pathogenic oral bacteria

 描述（由申请人提供）：益生菌是微生物，当食用时，可对宿主产生积极或有益的影响。在口腔中使用益生菌作为预防性疗法刚刚开始探索龋齿和牙周病。牙龈卟啉单胞菌（Porphyromonas gingivalis，Pg）是牙周病发生发展的重要病原菌。目前对Pg相关牙周炎的治疗主要是基于症状的治疗而不是预防性治疗。我们已经从乳制品中分离出几种德氏乳杆菌益生菌菌株，它们在体外抑制Pg诱导的细胞毒性和Pg生长。此外，我们已经证明，从这些菌株的L。delbreuckii抑制Pg生长。初步测试表明抑制分子 似乎是分子量在30-100 kDa之间的蛋白质。本研究拟从L.通过阴离子/阳离子交换色谱法和质谱法测定delbreuckii。使用这些数据，我们将确定编码抑制因子的基因组位置。我们将克隆和纯化重组抑制因子，以进行进一步的测试。使用先前构建的Pg转座子文库，我们将使用大规模平行测序技术来快速定量筛选Pg中与抑制性上清液和/或重组抑制性蛋白相互作用的基因，以快速鉴定潜在的作用机制。最后，我们将研究临床分离的Pg的敏感性 和其他口腔细菌对L. delbreuckii抑制性上清液和/或纯化的重组抑制性蛋白，以确定L. delbreuckii抑制剂对临床Pg分离株。完成本申请中概述的目标可能会确定一种新的种间抑制机制，并证明我们的L。delbreuckii益生菌菌株和/或其分泌产物。

 DESCRIPTION (provided by applicant): Probiotics are microorganisms that when consumed confer a positive or beneficial effect on the host. Use of probiotics in the oral cavity as preventative therapies are just beginning to be explored for dental caries and periodontal disease. Porphyromonas gingivalis (Pg) is an important organism in the onset and progression of periodontal disease. Current treatment for Pg associated periodontitis is largely based on treatment of symptoms rather than preventative therapies. We have isolated several probiotic strains of Lactobacillus delbreuckii from dairy products that inhibit Pg-induced cytotoxicity and Pg growth in vitro. Furthermore, we have demonstrated that supernatants derived from these strains of L. delbreuckii inhibit Pg growth. Initial testing indicates that the inhibitory molecule appears to be a protein with a molecular weight between 30-100 kDa. We propose to isolate and identify the secreted factor from L. delbreuckii by anion/cation exchange chromatography and mass spectrometry. Using this data, we will identify the genomic location encoding the inhibitory factor. We will clone and purify a recombinant inhibitory factor to carry out further testing. Using a previously constructed transposon library in Pg, we will use massively parallel sequencing technology to quickly and quantitatively screen for genes in Pg that interact with the inhibitory supernatant and/or recombinant inhibitory protein to quickly identify potential mechanisms of action. Lastly, we will examine the susceptibility profile of clinical isolates of Pg and other oral bacteria to L. delbreuckii inhibitory supernatants and/or purified recombinant inhibitory protein in an effort to determine spectrum of action of the L. delbreuckii inhibitor against clinical Pg isolates. Completion of the aims outlined in this application may identify a novel interspecies inhibitory mechanism and demonstrate the feasibility of a clinical application for our L. delbreuckii probiotic strains and/or their secreted products.