Identification of host-specific virulence determinants in the opportunistic enteric pathogen, Pantoea

Pantoea是一种与人类病原菌、肠杆菌和大肠杆菌密切相关的细菌群，但目前许多新分离株在加拿大注册为生防剂，用于管理作物上的植物病害。这一点特别令人担忧，因为许多Pantoea分离物已经被证明本身会引起植物疾病，也会引起人类疾病。Pantoea已被发现在健康和免疫功能低下的成年人中都会导致肺部感染、败血症和感染性关节炎。这项研究计划的目的是促进我们对Pantoea致病能力的了解，以及更广泛地说，病原体如何适应利用不同的宿主，包括人类。这一建议基于这样的假设，即存在使Pantoea在多个宿主中致病的特定疾病决定因素。*第一个目标是通过比较Pantoea ananatis毒力和非毒力菌株的基因来确定候选疾病因素，以确定那些仅存在于强毒菌株中的决定因素。为了评估这些基因中是否有任何基因是在人体温度(37摄氏度)下诱导的，我们将使用转录转录方法检测在30摄氏度和37摄氏度下的基因表达。第二个目标是评估一个宿主定居所需的遗传因素是否需要另一个宿主。这将使用三组突变型的花叶夜蛾来实现，它们被证明在与果蝇、线虫或阿米巴的交往能力方面受到了损害。线虫和阿米巴突变体组的生长将在模型果蝇中进行评估，以确定那些特定于一个宿主的基因，或多个宿主所需的基因。第三个目标将确定许多Pantoea分离株在37℃下表现出的溶血素活性(红细胞溶解)的遗传基础。我们将生成并筛选溶血能力丧失的突变库，从而确定与这种能力有关的基因。第四个目标将评估Pantoea生产的天然抗生素在宿主联合中的作用。由于宿主经常被各种共生细菌定植，病原体可能会通过消除竞争细菌来增强它们确定感染的能力。利用已经感染了病原体金黄色葡萄球菌的果蝇，我们将比较产生抗生素的菌株和抗生素突变体的定植效率，以确定这些天然产物是否可能是增强宿主定植的毒力因素。*这项研究将识别有助于Pantoea在人类身上致病的毒力因素，并将使我们能够评估生控菌株成为病原体的能力。这项研究不仅对生物控制剂的监管具有广泛的影响，而且还将加强我们对细菌如何进化成为致病细菌的理解。

Pantoea is a bacterial group that is closely related to human pathogenic, Enterobacter and E. coli, yet many isolates are currently registered in Canada as biological control agents for managing plant disease on crop plants. This is particularly concerning given that many Pantoea isolates have already been shown to cause plant disease themselves, as well as human disease. Pantoea has been noted to cause pulmonary infections, septicemia, and septic arthritis in both healthy and immunocompromised adults. The objective of this research program is to advance our understanding of the pathogenic ability of Pantoea, and more broadly, how pathogens adapt to exploit different hosts, including humans. This proposal rests on the hypothesis that there are specific disease determinants that enable Pantoea to cause disease in multiple hosts.***     The first objective is to identify candidate disease factors by comparing the genes of virulent and non-virulent isolates of Pantoea ananatis to identify those determinants that are present only in virulent isolates. To evaluate whether any of these genes are induced at human body temperature (37C), we will examine the expression of genes at 30C versus 37C using transcriptomic methods. The second objective is to evaluate whether genetic factors necessary for colonization of one host are needed for another host. This will be achieved using three mutant sets of P. ananatis, which were shown to be impaired in the ability to associate with either the fruit fly, nematode, or amoeba. Growth of the nematode and amoeba mutant sets will be evaluated in the model fruit fly to identify those genes that are specific to one host, or are needed for multiple hosts. The third objective will identify the genetic basis of hemolysin activity (lysis of red blood cells) exhibited by many Pantoea isolates at 37C. We will generate and screen mutant libraries for loss of hemolytic ability, thereby identifying the genes that are responsible for this capability. The fourth objective will evaluate the role of natural antibiotics produced by Pantoea during host association. Because hosts are often colonized by a variety of commensal bacteria, pathogens may enhance their ability to establish an infection by eliminating competing bacteria. Using fruit flies that are already infected with the pathogen Staphylococcus aureus, we will compare the colonization efficiency of an antibiotic-producing isolate to an antibiotic mutant to determine whether these natural products may be virulence factors that enhance host colonization.***     This research will identify virulence factors that contribute to the ability of Pantoea to cause disease in humans, and will allow us to assess the capacity for biological control strains to become pathogens. This research not only has broad implications for the regulation of biological control agents, but will also enhance our understanding of how bacteria evolve to become pathogenic.**