Oral Colonization of Aggregatibacter in Primates

描述（由申请人提供）：放线菌聚合杆菌（Aa）与年轻人的局部侵袭性牙周炎（LAP）密切相关。此外，Aa具有多种毒力性状，与LAP中发生的致病事件一致。我们的团队一直在研究与Aa诱导的疾病起始相关的基因，并以此编码负责Aa附着、定植、持久性和龈下存活的毒力特征。到目前为止，我们已经将两个Aa自转运体黏附素基因（aae和apiA）的功能与Aa附着于上皮的特异性相关，并表明这些黏附素（以及白质毒素）在旧大陆（OW）灵长类动物和人类中具有物种特异性。这种特异性使得OW灵长类动物成为研究与Aa感染相关的早期事件并着眼于制定预防策略的理想模型。本R21应用程序由两个目的组成，旨在比较两种Aa菌株（一种来自人类，一种来自恒河猴）的口腔定植和持久性，这两种Aa菌株被引入OW灵长类动物的口腔。在将每个菌株引入猴口中后的28天内，将在不同时间检查其模式、附着水平和定植。目的1将首先描述Rh猴口腔中Aa的地形和数量水平，然后比较两种Aa菌株（一种人和一种猴）的定植和持久性。在放置之前，猴子将接受洗牙和预防以及氯己定治疗。动物将在煎饼中喂食Aa，并在喂食后28天分析定植和持久性。在牙齿上定植并持续存在的Aa等于或大于1 × 102 /mL菌落的Aa将被选择用于Aim 2。目的2将评估BECs、舌头和牙齿上发现的Aa的位置、水平和时间，比较未接种和野生型接种的Aa与aae和apiA突变的Aa， aae/apiA、flp（纤原外蛋白）和ltx在28天内的双敲除。这些实验将揭示这些基因在Aa附着于BECs （aae和apiA）、牙齿定植（flp）和OW灵长类动物口腔龈下存活（ltx）方面的重要性。建立该模型的实用性，不仅可以让我们剖析附着因子，还可以揭示其他Aa基因的免疫调节因子，因为它们影响Aa的发病机制，并着眼于预防策略。

DESCRIPTION (provided by applicant): Aggregatibacter actinomycetemcomitans (Aa) is intimately associated with Localized Aggressive Periodontitis (LAP) in young adults. Further, Aa possesses a variety of virulence traits that are consistent with pathogenic events that occur in LAP. Our group has been studying genes that are related to Aa-induced disease initiation and as such encode virulence traits responsible for Aa attachment, colonization, persistence, and subgingival survival. Thus far we have ascribed functions for two Aa autotransporter adhesin genes (aae and apiA) that are related to the specificity of Aa attachment to epithelium and have shown that these adhesins (as well as Leukotoxin) show species specificity for Old World (OW) primates and humans. This specificity makes the OW primate an ideal model for studying early events related to Aa infection with an eye toward development of preventive strategies. This R21 application consists of two Aims and is designed to compare oral colonization and persistence of two Aa strains (one from humans and one from Rhesus [Rh] monkeys) that are introduced into the oral cavity of OW primates. Each strain will be examined for its pattern, level of attachment, and colonization at different times over a 28-day period following introduction into the mouth of the monkey. Aim 1 will describe the topographical and quantitative level of Aa found in the mouths of Rh monkeys initially and then compare colonization and persistence of the two strains of Aa (one human and one monkey). Before placement, monkeys will receive scaling and prophylaxis plus chlorhexidine treatment. Animals will be fed Aa in a pancake and colonization and persistence will be analyzed 28 days after feeding. The Aa that colonizes and persists on teeth at a level equal to or greater than 1 x 102 /mL colonies of Aa will be selected for use in Aim 2. Aim 2 will assess the location, level and timing of Aa found on BECs, tongue and teeth comparing un-inoculated and wild type inoculated Aa to Aa with mutations in aae and apiA, a double knockout of aae/apiA, flp (the fibrillar outer protein) and ltx over a 28-day period. These experiments should reveal the importance of these genes in relation to Aa attachment to BECs (aae and apiA), to tooth colonization (flp) and to subgingival survival (ltx) in the oral cavity of OW primates. Establishing the utility of this model should allow us not only to dissect out attachment factors but also to unravel immune modulation factors from other Aa genes as they affect Aa pathogenesis in the future with an eye toward preventive strategies. PUBLIC HEALTH RELEVANCE: Aggregatibacter actinomycetemcomitans is intimately associated with Localized Aggressive Periodontitis in young children, which can result in premature loss of teeth. Coincidentally, A. actinomycetemcomitans is also found in the mouths of Old World primates such as Rhesus (Rh) monkeys. It has been shown that A. actinomycetemcomitans attaches to tissues and kills defense cells in humans and Rh monkeys in a very similar manner. Therefore the Rh monkey provides us with an ideal model in our efforts to unravel some of the mysteries related to the earliest stages of A. actinomycetemcomitans-induced disease. It is also known that A. actinomycetemcomitans produces several factors that are similar to many other bacteria that cause severe infections in man. Therefore, studying A. actinomycetemcomitans in the mouth of Rh monkeys can provide a model that can help us understand how A. actinomycetemcomitans-induced infections develop in a real world environment. This understanding may also have application for other mucosal diseases. We have developed a relationship with the Northeast Primate Research Center and have designed a study that will enable us to place this bacterium in the mouths of Rh monkeys. This study design will allow us to examine how genes direct the way in which A. actinomycetemcomitans attaches to tissues and initiates disease. While attachment is recognized as the first step in infection of skin surfaces that include surfaces like the gums, survival depends on the ability of the bacteria to defend itself against host defense cells. This proposal is designed to examine how certain A. actinomycetemcomitans genes influence early events such as attachment and survival below the gum line. Use of this primate model will allow us to develop ways to interfere with attachment and survival in the hope that we can devise strategies to prevent infections without the need to resort to the use of antibiotics. )