Development of PCR Markers for Diagnosing Sporisorium ellisii Infection within Populations of Andropogon virginicus 

Artur Romanchuk,  Biology             

Faculty Mentor :  Dr. Janet A. Morrison

              Natural populations are not found as collections of individuals, but rather as complex communities based on mutual interactions.  One class of such interactions is the relationships established between pathogens and their hosts. We work to understand such interactions between a parasitic smut fungus, Sporisorium ellisii, and its grass host, Andropogon virginicus. These species are native to the eastern United States, where A. virginicus occupies old fields undergoing early succession.  Sporisorium ellisii infects the ovaries of its hosts, replacing the plant’s future seeds with its own progeny (teliospores). Infection is asymptomatic until it can be identified by inspecting the host plant during early autumn, when healthy plants normally display seeds.

              In order to continue our work regarding interactions of A. virginicus and S. ellisii, we needed to develop a reliable method to diagnose infection. Polymerase chain reaction (PCR) is used routinely to amplify known segments of DNA, and this ability can be used to identify trace amounts of specific DNA sequences within a larger sample of DNA. We seek to use PCR technology along with suitable primer markers to identify fungi-specific DNA within a host plant’s DNA extract, which would prove to be a reliable diagnostic tool.

              During Summer 2007 we concentrated on primer design and optimization of PCR reaction conditions. Initially we screened fungal and plant DNA samples with universal ITS primers, which identify a particular stretch of moderately variable non-coding DNA within the gene for the small ribosomal subunit. These initial screens proved to be unproductive. We also screened fungal and plant DNA with a version of the ITS primers that are more specific for the group of fungi to which S. ellisii belongs, the Basidiomycetes. This proved to be much more successful. We have shown that the basidiomycete-specific primers produce a signal from fungal DNA template, and do not produce one from A. virginicus DNA template. Future investigation will focus on further narrowing the signal producing range of this molecular marker to amplify only with S. ellisii DNA and not other fungi. This will allow rapid and accurate diagnosis within field conditions, and promote further investigation of host-pathogen interactions within this study system.

              Parallel to the molecular work, I shared in the care of experimental and stock plants in the greenhouse, and collected size data on plants from a US-wide sample of populations.

Personal Statement

              Summer SURP 2007 has proved to be an invigorating experience into the world of real research. I have been able to further understand how academic research is carried out on day to day basis, and fine-tune my own hopes and desires for the future. As well as receiving great guidance from my mentor, other faculty members, and my undergraduate colleagues, I also was able to foster my independence in developing and testing ideas.