2012-08-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/644903摘要：感染性心內膜炎(Infective endocarditis)是因細菌黏附在受損的心臟瓣膜造成的疾病，其中由口腔內共生菌之草綠色鏈球菌（viridans streptococci）是造成不同年齡層病患亞急性感染性心內膜炎(subacute infective endocarditis)的主要致病菌。目前研究認為當草綠色鏈球菌經由口腔內傷口進入血流，會造成暫時性的菌血症，血流中的細菌會黏附到受損的心臟瓣膜上，形成贅疣（vegetation）進而引發感染性心內膜炎的發生。當菌血症持續存在時間越長，越有利細菌黏附上受傷瓣膜，引發感染性心內膜炎。在臨床上，對於高危險群病患預防感染性心內膜炎多會給予高劑量抗生素以減少外科手術過程中菌血症的發生。然而，預防性抗生素投藥在預防感染性心內膜炎的效果仍未有定論。本實驗室初步成果顯示在菌血症大鼠模式中，給予高劑量抗生素預防投藥，卻無法抑制或減緩菌血症嚴重程度。實驗室先前研究首先發現受到血漿刺激後可增加草綠色鏈球菌在大鼠體內的存活率(Mol Microbiol. 2009; 74:888–902)，此外，最新的研究中也證明血漿可大幅提升細菌對多種抗生素的抵抗能力(J Infect Dis. 2011; in press)。我們的研究成果顯示當細菌進入血流受到血漿刺激後可調控基因表現增加細菌抵抗宿主免疫攻擊與抗生素的能力。進一步利用微陣列基因體技術(microarray)檢測細菌受血漿刺激後基因表現的改變，發現一對細菌雙分子調控系統LytST 中的LytT 調節蛋白(response regulator)是負責細菌受血漿刺激的主要調控因子之一。因此本三年計畫目的要找出引發感染性心內膜炎草綠色鏈球菌受到血漿刺激後，LytT是如何增加細菌在血液中的存活與抵抗抗生素的分子調控機制。1. 鑑定細菌雙分子調控系統的LytT調節蛋白會調控的下游基因。2. 研究受LytT調節蛋白調控的基因抵抗宿主免疫攻擊與抵抗抗生素的分子機制。3. 篩選可抑制LytTS 雙分子調控系統訊息傳遞之藥物用以增加抗生素效用。在這個計畫中，選擇轉糖鏈球菌作為實驗的模型系統是由於(1)國內感染科醫師研究指出該菌為引發菌血症草綠色鏈球菌中引發國人感染性心內膜炎比例最高之菌株。(2)國外臨床研究調查顯示該菌可引發嚴重型心內膜炎。(3)該菌的染色體已被完整的定序而且又有方便操作的基因剔除及互補系統進行分子生物相關研究。本計畫對於臨床預防及治療心內膜炎的重要性在於隨著抗藥性菌株的增加，抗生素臨床治療心內膜炎益發不易，藉由了解細菌以何種分子機制影響在血液中存活與抵抗抗生素的能力，可有助發展增加抗生素效用或有效殺菌的非抗生素類的其他藥物。並篩選可抑制雙分子調控系統訊息之藥物以增進抗生素之效用。<br> Abstract: Infective endocarditis(IE) is the infectious disease resulting from the bacterial adhesiononto the injured valve and viridans streptococci are the major pathogens in causing subacuteinfective endocarditis in general populations. Bacteremia of viridans streptococci is causedby trauma or dental procedures in the oral cavity and the circulating bacteria can adhere tothe damaged valve to form a compact structure, vegetation, and further induce infectiveendocarditis. Moreover, the prolonged time period of bacteraemia is in favor of theadherence onto the vegetation. Clinically, the prophylactic usage of high dose antibiotics isalways prescribed to devoid the bacteremia in the high risk patients under surgery. However,the prophylactic effect of antibiotics in preventing IE remains uncertain. In our preliminaryresults, the prophylaxis with high dose antibiotics had no effect on the prevention ofbacteraemia in rat model. Besides, we previously reported that plasma stimulation couldenhance the bacterial survival rate in vivo. (Mol Microbiol. 2009; 74:888–902) Furthermore,plasma could contribute to multiple antibiotics tolerance of viridans streptococci in ourrecent results. (J Infect Dis. 2011; in press) Our results suggested that bacterial generegulation in response to the plasma stimulation confers the resistance to host immuneresponse and tolerance to multiple antibiotics. Preliminarily, we have identified a twocomponent system LytST response regulator LytT, which plays an important role in thebacterial gene regulation in response to plasma stimulation from microarray. Therefore, wepropose to investigate the molecular mechanisms in response to plasma stimulation thatregulates bacterial survival in vivo and the tolerance to multiple antibiotics. The specificaims of this three-year project are:1. To identify the downstream genes regulated by response regulator LytT in response toplasma stimulation2. To investigate the plasma-induced mechanism of LytT-governed regulation inconfronting host immune response and bacterial tolerance to multiple antibiotics3. Selection of the possible inhibitors in LytTS to enhance the efficacy of antibioticsIn this project, we will use S. mutans as the model system for three reasons. First, S.mutans is responsible for the highest incidence of endocarditis in bacteremia-associatedinfection in Taiwan. Second, S. mutans could induce severe-type quadric-valvularendocarditis in clinical cases. Third, its complete genomic information and well-establishedgenetic manipulations are advantageous in the research. The significance of this project is tofurther understand the whole-picture mechanism of multiple antibiotics tolerance in vivo.Following with the occurrence of multiple antibiotics resistant bacteria, antibiotics therapybecomes more and more difficult to treat bacterial infection. Through studying themolecular mechanism to enhance bacterial survival rate in vivo and to tolerate multipleantibiotics, it renders great benefits to develop new agents with synergistic enhancementcombined with antibiotics. More precisely, the selection of inhibitors in TCS signaling willbe performed to identify the possible agents to confer the synergistic effect in combinationwith antibiotics.感染性心內膜炎細菌雙分子系統抗生素抵抗性infective endocarditistwo component systemantibiotics tolerance