Uncovering the genetic basis for host-specific adaptation and opportunism in the bacterial pathogen, Pantoea

了解细菌病原体的宿主范围和潜在毒力对我们预测和控制新发和再发传染病的能力至关重要。我的研究项目的目的是利用跨界细菌群Pantoea来提高我们对机会性病原体的理解。Pantoea是研究机会寄主关联进化的理想研究系统。它最初作为一种植物致病群而受到关注，但其成员后来成为皮下和肺部感染以及严重败血症的致病因子。然而，尚不清楚的是，是否某些泛亚物种对特定宿主变得越来越专门化，或者是否所有物种都具有引起机会性感染的能力。这一建议基于这样的假设，即在某些泛菌物种中存在促进宿主定植和毒力的遗传决定因素。第一个目标是使用全基因组数据来建立一个完全确定的泛古菌系统发育。我们将测序96个新的基因组，代表12个不同的Pantoea物种，包括昆虫，植物和人类的病原体，以及环境菌株。我们将利用全基因组信息，结合核心基因的平均核苷酸同一性和遗传距离建立系统发育树，并划定物种边界。这种系统发育将建立进化框架，以评估毒力是谱系特异性的，还是独立进化的。第二个目标是对96个测序菌株在蜡虫模型中进行毒力测定，这将评估不同菌株定殖宿主和引发感染的能力。蜡虫感染试验将包括注射细菌接种剂，然后评估48小时内蜡虫的黑化和死亡率。第三个目标将是使用比较基因组学确定候选毒力因子，然后对这些基因进行诱变以评估其在毒力中的作用。比较基因组学平台EDGAR将用于鉴定最毒菌株的共同基因集，这些基因集将代表与宿主定植有关的基因。这些基因的一个子集将被选择并在最毒的菌株中通过同源重组单独诱变，并在蜡虫中定性和定量地评估所产生的突变体的毒力。通过使用我们的系统发育提供的进化框架，我们可以阐明这些毒力能力是谱系特异性的还是独立进化的。Pantoea为研究机会主义的进化提供了一个独特的机会，这将使我们更好地理解驱动宿主特异性毒性和疾病的进化过程，以及新感染因子的出现。

Understanding the host range and virulence potential of bacterial pathogens is central to our ability to predict and control emerging and reemerging infectious diseases. The objective of my research program is to advance our understanding of opportunistic pathogens using the cross-kingdom bacterial group, Pantoea. Pantoea serves as the ideal study system for examining the evolution of opportunistic host association. It initially gained attention as a plant pathogenic group, but its members have since emerged as the causal agents of subcutaneous and pulmonary infections, and severe septicemia. What is unclear, however, is whether some Pantoea species have become increasingly more specialized to specific hosts, or whether all have the capacity for causing opportunistic infections. This proposal rests on the hypothesis that there are genetic determinants in some Pantoea species that facilitate host colonization and virulence. The first aim is to use whole genome data to establish a fully resolved Pantoea phylogeny. We will sequence 96 new genomes representing 12 different species of Pantoea, including pathogens of insects, plants, and humans, as well as environmental strains. We will use whole genome information together with average nucleotide identity and genetic distance of core genes to create a phylogenetic tree and delineate species boundaries. This phylogeny will establish the evolutionary framework for evaluating whether virulence is lineage specific, or whether it evolved independently. The second aim is to carry out virulence assays of the 96 sequenced strains in the waxworm model, Galleria mellonella, which will evaluate the ability of different strains to colonize the host and initiate infection. Infection assays with waxworms will involve injection with bacterial inoculum, followed by an assessment of waxworm melanization and mortality over a 48 hour period. The third objective will be to identify candidate virulence factors using comparative genomics, and then mutagenize these genes to assess their role in virulence. The comparative genomics platform, EDGAR, will be used to identify common gene sets across the most virulent strains, which will represent genes involved in host colonization. A subset of these genes will be selected and mutagenized individually by homologous recombination in the most virulent strains, and the virulence of the resulting mutants evaluated in the waxworm both qualitatively and quantitatively.  By using the evolutionary framework provided by our phylogeny, we can elucidate whether these virulence capabilities are lineage-specific or if they evolved independently. Pantoea provides a unique opportunity to study the evolution of opportunism, which will lead to a better understanding of the evolutionary processes that drive host-specific virulence and disease, and the emergence of new infectious agents.