Clavibacter species cause disease on a narrow host range of plants, but their ability to colonize endophytically is broad. There are six pathogens of the genus:
The tomato pathogen is the most well studied, known to carry multiple virulence genes on two plasmids and within a pathogenicity island (PAI). While the plasmid number, size, and gene content vary within and across species, the entire PAI is only found in Cm. Instead, fragments and/or homologs of PAI members can be found in Cc, Cs, and Ci genomes. While functional validation of some of effectors has been done, most of the plant targets are unknown. Even less is known for other pathogens of the genus.
Part of figure 1, Thapa et al., 2019. Annual Review in Phytopathology.
Nanopore MinION next to tomato and Gram-positive actinobacteria.
In order to comprehensively predict effectors, I am generating a robust dataset of high-quality genomes of species not well sampled in NCBI (national database). By using both short and long-read technologies, we can leverage their individual strengths without drastically increasing costs. Long reads (Oxford Nanopore), while more error-prone, can provide scaffolding and span through repetitive regions. The short reads (Illumina), while very accurate for base calling, are prone to incomplete assemblies. These genomes will provide a robust resource for this work as well as future research.
Pathogen adaptation to a given environment can occur through gene gains, losses, and diversification. In my model, Clavibacter pathogens have adapted to a particular host through subtle changes in effector content, which can be predicted through a variety of computational approaches. As I've learned from a wise postdoc, no one computational approach will give you the answer but a variety of robust approaches will provide insight to help you move forward and develop hypotheses to further test and validate. In total, I anticipate being able to predict effectors essential for pathogenicity and host range.
In silico (computational) predictions are only that, predictions. Therefore, I am developing genetic and biochemical assays to be able to better test these predictions (see more info on that here). Previous work supports proteases (proteins that cut up other proteins) and CAZymes (proteins that build, modify, and break carbohydrates, the main component of plant cell walls) as candidates for mediating these processes. Using the tools developed above, I plan to assess the contribution of host specificity in tomato. The completion of these experiments will enable the identification of the first host targets of Clavibacter effectors and their contribution to host range.