Written by Joanna Fyans at the University of Dundee & SCRI. This is the report from a BSPP Junior Fellowship. Click here to read more/apply for one yourself.
My recent research visit to the lab of Prof Rosemary Loria at Cornell University was made possible by the generous award of a BSPP Junior Research Fellowship, providing me with an excellent opportunity to investigate two novel areas of research that had arisen as part of my PhD.
The main focus of my project is to examine how the protein export systems of Streptomyces scabies, the causative organism of common scab of potato, contribute to the pathogenicity of the organism. As the Loria group at Cornell University is widely acknowledged as the leading research laboratory working on S. scabies, having developed and perfected many techniques pertinent to the study of this plant pathogen, I was ideally situated to undertake the required work. I was able to participate in discussions and lab meetings which provided me with useful insight into the mechanisms that plant pathogenic Streptomycetes use to cause disease, potential ways to approach investigation of these and knowledge into current research taking place in the field. I was given access to all facilities and expertise available not only in the Loria lab but also the department as a whole.
My project focuses on two protein export systems; the twin-arginine translocation pathway (Tat) and the Type VII secretion system (Esx), as such it was my intention to investigate aspects of each of these systems.
Although we have recently published that multiple virulence factors are secreted by the Tat pathway in S. scabies, specific roles for any of these are yet to be elucidated. I therefore aimed to investigate the role of one of the proteins, a putative transcription factor, that is potentially secreted by this system. I approached this by attempting to construct a chromosomal knockout mutant in the gene encoding this protein which would then be analysed further. However, despite having prepared the necessary genetic construct I was unsuccessful in isolating this mutant. Taken together with the fact that this protein is highly conserved in other Streptomycetes (including some non-pathogenic species) we believe that the gene is essential. This precluded further work I had planned to carry out at Cornell into the function of this protein.
My time became largely focussed on my second aim, which was to investigate the role of the Type VII system in the virulence of S. scabies. To our knowledge this system is completely uncharacterised in any plant pathogen but is known to be essential for virulence in other Gram positive pathogens such as Mycobacterium tuberculosis and Staphylococcus aureus, so we hypothesised that it may play a role in S. scabies virulence. I was guided in the setting up of sterile plant culture systems, something I had limited experience in and had great difficulty implementing in Dundee, and was able to use these to subject S. scabies Type VII mutant strains to virulence testing. Due to the wide host range of S. scabies the Loria lab utilises a number of model and crop plants so I was able to gain experience in carrying out assays using Arabidopsis, tobacco, radish and potato plants. I also worked together with members of the Loria group to adapt the existing assays and assist in development of new ones with view to gaining increased sensitivity and allowing earlier detection of disease symptoms. Additionally, I learned the method for preparation of thaxtomin extracts (a phytotoxin produced by plant pathogenic Streptomycetes that is essential for disease development) from cultures for quantitative analysis by HPLC.
Despite not finding a role for the Type VII system in S. scabies virulence, my time at Cornell was extremely fruitful and allowed me to collect a large amount of infection data will contribute significantly to my thesis. We also anticipate that the results of this work will form part of a publication in the near future. During my time at Cornell I also had the opportunity to present a seminar of our collaborative work on the Tat system of S. scabies to the department of plant pathology and plant-microbe biology which provided the perfect opportunity to meet and interact with many members of the faculty.
I would like to thank Rose for giving me a warm welcome as well as Dr Isolde Francis and Sara Carpenter for taking time out of their own work to supervise my work ensuring that my visit was not only scientifically productive but also a great personal experience.
University of Dundee & SCRI