Setting up a qPCR plate
Author: Haley Stanton | Major: Animal Science
My name is Haley Stanton, and I’m a junior Animal Science major in Bumpers College. Under the supervision of my research mentor, Dr. Dowling of the Department of Entomology and Plant Pathology, I began my research in the summer of 2021 and finished the last data collection in the fall of 2021. Although the data collection may be over, there is still plenty left to do, and I plan to conduct the data analysis and formulate the conclusions of my paper by the end of my junior year. I plan to graduate in the spring of 2023 and attend veterinary school afterward – it was my passion for the health of animals (and people) that inspired my research idea. I wanted to conduct a project that concerned the health of both animals and people, which led me to research tick-borne diseases. After coming up with my idea, I contacted Dr. Dowling due to his similar research interests, and he has been a mentor to me ever since.
My goal with this research is to identify the different tick species in four Arkansas state parks, analyze the proportion of ticks that carry Ehrlichia and Rickettsia species by tick species and life stage, and determine if there is a significant variation in these proportions from park to park. Ehrlichia and Rickettsia are pathogens vectored by ticks and can cause ehrlichiosis and spotted fever rickettsiosis, including Rocky Mountain spotted fever (RMSF), which are diseases that affect both humans and animals. My hope with this research is to raise awareness of tick-borne diseases and inform others of the risks that ticks can pose to health so that proper precautions are taken when hiking in state parks.
To test the ticks for pathogens, I first had to collect them via standardized methods, which I did in the summer of 2021. This involved both dragging a piece of cloth on the ground and picking ticks off as they crawled, along with putting dry ice traps out overnight, which attract ticks. By far, this was the most challenging part of my research. In field research, many things can go wrong since there are so many uncontrolled variables. However, this was mitigated by visiting each park four times over the course of four months in case there were outliers. Things still didn’t go perfectly, such as there being popup rain showers overnight that saturated the dry ice traps, but these will be considered when analyzing and discussing the project. Unexpectedly, there was already a noticeable difference between the parks at this stage. Three of the four parks had approximately the same number of ticks each visit, but one park had only a fraction of the ticks present at other parks. This was an interesting result that I hope will be explored further!
After all ticks were collected, I identified each tick’s species and life stage to account for possible variation in results due to these variables. Then, I extracted the ticks’ DNA using genomic kits so that they could be tested for Ehrlichia and Rickettsia pathogens. Due to the high number of nymphs collected, they were pooled in groups up to 5 nymphs per DNA extraction. The presence of Ehrlichia and Rickettsia pathogens was determined through quantitative polymerase chain reaction (qPCR) assays. Specifically, the ticks were tested for E. chaffensis, E. ewingii, and any Rickettsia species in general. For the adult ticks, approximately 61.5% were positive for Rickettsia, 6.8% were positive for E. ewingii, and 1.3% were positive for E. chaffensis. Further analysis needs to be done to determine the minimum infection prevalence for nymph pools – since positive nymph pools have at least one positive nymph but not necessarily more than that – and any significant variation in results between parks.
Ultimately, this project has furthered my passion for veterinary medicine, piqued my interest in research as a career field, and given me invaluable experience in a laboratory setting. Knowing how to perform DNA extractions and qPCR assays successfully and without contamination will be incredibly useful for veterinary diagnostics. I’m also extremely grateful to my research mentor, Dr. Dowling, for being supportive of my ideas and helping me throughout the way. I’ve even been looking into dual DVM/PhD degree programs due to how much I have enjoyed doing research.