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

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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.