Kristen Gates Senior Biology Major, 2021
Author: Kristen Gates | Major: Biology
My name is Kristen Gates and I am a biology major and African and African American studies minor in the Fulbright College of Arts and Sciences. In the fall semester of 2021, I conducted research in Dr. Clemencia Rojas’ lab in the department of entomology and plant pathology. The focus of my project is isolating and identifying antimicrobials secreted by beneficial plant bacteria in order to control Bacterial Panicle Blight (BPB) in rice. Over the course of the semester, our lab made progress in refining the isolation of antimicrobial compounds by using minimal media to grow beneficial plant bacteria. I refined the isolation process in order to obtain samples to submit for mass spectrometry and chemical characterization with our collaborators at the University of North Carolina at Chapel Hill.
Bacterial Panicle Blight is a disease in rice caused by the bacterium Burkholderia glumae. BPB causes grain yield and economic losses that are likely to get worse due to global warming. Antimicrobials secreted by the beneficial plant bacterium Pseudomonas protegens PBL3 can be lyophilized or freeze-dried for storage. The antimicrobial compounds present in the freeze-dried powder can be reconstituted and applied as a biopesticide to control BPB in rice. We are exploring the growth curves of different Pseudomonas species in order to identify possible negative control strains, optimizing the growth conditions for the strains, and optimizing lyophilization of P. protegens PBL3 in minimal media.
The opportunity to work in the lab of Dr. Clemencia Rojas was brought to my attention by Honors College Dean Lynda Coon. In the summer between my freshman and sophomore year, I worked in Dr. Rojas’ lab as a research assistant. I performed antimicrobial growth inhibition assays to determine the extent of the antimicrobial activity exhibited by P. protegens PBL3. Dr. Clemencia Rojas introduced the prospect of working on a cross-disciplinary research project that required biological and chemical separations. Prior to joining the lab, I had been introduced to separation methods such as SDS-PAGE and affinity chromatography in the biochemistry laboratory of Dr. T. K. S. Kumar. I was interested in the antimicrobial isolation project proposed by Dr. Rojas because it combined my interests in pharmaceutical science and sustainability.
This past semester, most of my time in the lab was spent measuring the optical density of Pseudomonas samples at different stages of bacterial growth and performing experiments to determine the effects of myo-inositol concentration on the quality of the final product. Comparing the results for the different strains of Pseudomonas, I could determine which bacterial strains exhibit the most antimicrobial activity. The bacterial strains that exhibited antimicrobial activity were grown in minimal media and lyophilized. We used minimal media because the samples that we previously submitted to mass spectrometry contained compounds that were too complex to produce a meaningful result. We also decided to scrap the media that we had been using to plate serial dilutions earlier this year because there were issues with the selectivity of the media.
Laura Ortega, a graduate student in the lab, determined that growing P. protegens PBL3 in a larger volume in an initial step would result in a lyophilized sample with greater antimicrobial activity. Laura lyophilized samples and familiarized me with new protocols. The initial experimentation with solid phase extraction in the Rojas lab was performed by Casey Patrick. Casey worked on the initial phases of the antimicrobial isolation project and showed me how to perform solid phase extraction using a vacuum manifold. Dr. Clemencia Rojas encouraged me to apply for the Honors College Research Grant, looked over my drafts numerous times, and made invaluable suggestions. Dr. Rojas gave me and every member of the lab helpful feedback at our weekly lab meetings. The weekly lab meetings provided us an opportunity to present our findings from the past week and our research plan for the upcoming weeks. Dr. Rojas would sometimes ask me to perform experiments that would lead me to revise the protocols. These revisions saved me from repeating the same mistakes and allowed me to make progress in isolating the antimicrobial compounds.
I submitted a presentation to the Arkansas Crop Protection Association (ACPA) Annual Research Conference that took place in early December, 2020. The ACPA Research Conference gives undergraduate, masters, and doctoral students in Arkansas a chance to present their research. The event was held virtually due to the pandemic and my presentation placed in the undergraduate category. Even though the research was challenging at times, the months spent learning the nature of Pseudomonas species growth and antimicrobial isolation was important to developing a more nuanced understanding of bacteria. This experience will stick with me as I learn more about bacterial lifestyles through clinical experience. My future plans consist of finishing out my undergraduate degree in biology, taking a gap year, and applying to medical school in 2022.