My dissertation was focused on two pathogens that affect strawberries. The first was Botrytis cinerea, and the second was Neopestalotiopsis. 

The major issue with Botrytis is that it has high genetic diversity and prolific sporulation. The pathogen can overcome pesticides quickly by mutation, multiple times within a single season. In my dissertation, my first objective was to develop what is called a high-resolution melting, or HRM, assay to identify specific mutations that confer resistance to different fungicides so growers are better informed which fungicides to spray based on the type of Botrytis they have. The protocol that I developed uses spores, avoiding the DNA-extraction step and just running the HRM. 

The second objective was to study the resistance frequency of a fungicide called fludioxonil. This fungicide is pretty much considered a silver bullet for growers because it was very effective. However, within the last five years, we have seen that it is not as effective as before. 

The next part of my research was working with the new pathogenic species Neopestalotiopsis, which has been a very important research subject for the past few years. Being a new pathogenic species in Florida strawberry production, we did not know much about it when we started our research. I actually spent all the time during the pandemic developing a semi-selective medium so we could isolate the pathogen from the soil and the strawberry crop debris. It was a tedious process to narrow down the ingredients so we could be confident the recipe could only grow Neopestalotiopsis, inhibiting the growth of all the other pathogens harbored in the plants, the plant residue, and the soil. 

Then I used the media to determine the colony forming units and the survival of the pathogen in the soil and crop debris over time. We found that the pathogen does survive in the soil from one season to the next. It also survives in the strawberry leaves until leaf decomposition, where it goes into the ground and is considered soilborne. The pathogen can survive in the crown for 16 months, and possibly longer! 

I also tried a heat treatment, or thermotherapy, technique to kill the Neopestalotiopsis on the transplants prior to planting. A heat treatment protocol has already been established for Botrytis and other pathogens, but it has not been tested yet on Neopestalotiopsis. In vitro, we found that after four hours at 44 degrees Celsius, it completely killed the spores. For comparison, it only takes 30 minutes of thermotherapy to kill other pathogens. In plants, this heat treatment can reduce the level of inoculant by up to 50%.  

Finally, because strawberry plants are annual, at the end of the season growers get rid of the crop, usually with an herbicide, so we studied the effect of terminating strawberries at the end of the season on the inoculum of Neopestalotiopsis. Unfortunately, terminating strawberries this way can spike Neopestalotiopsis levels in the plant. Since Neopestalotiopsis spp. are necrotrophs and feed on the damaged strawberry tissue, damaging the strawberry plants with herbicides just makes food more available for the pathogen.