Progress 06/01/19 to 05/31/21
Outputs
Target Audience:• Students in agriculture • Scientific community • Vegetable Extension Agents • Georgia Farmers Changes/Problems:Major problems that have occured in this project include a delay in obtaining the toxigenic E. coli O157:H7 strains from the CDC. It took an extended period of time to find a point of contact that could find and deliver the isolates. Overall. it took a total of nine months to obtain the strains, even with consistent follow-up emails and calls. Additionally, the COVID-19 pandemic put a hault on research for several months. The University of Georgia discontinued any non-essential research and I was unable to continue my research during the months of March, April, May, and June. Furthermore, once non-essential research is allowed to resume, there will be restrictions on the amount of time I can work on the project and work within the lab to reduce possible contamination to the virus. I graduated from UGA in December 2020 and was unable to work on the project for the remainder of the year. I moved to the UK for a post-doc position in March 2021. Therefore no work on the project was completed from January 2021 to Mayb 2021. What opportunities for training and professional development has the project provided?I attended a professional development workshop series as part of The Plant Center Retreat at Unicoi State Park, GA in October 2019. The workshop touched on the following topics: • Teaching tools in the classroom • Money management in graduate school • Create publish worthy figures • Academic writing The workshop gave me some guidance on how to improve my academic writing and figure making for furture publications in perr-reviewed journals. I used the grant to pay for the workshop fee. I also attended the American Phytopathological Society Leadership Institute Workshop during the American Phytopathological Society meeting in August 2019. This workshop used interactive group activities to practice leadership skills and define what makes a good leader. I learned a lot about myself in terms of my leadership style and how to improve upon those skills. Through the grant, I was able to attend and present at the virtual American Phytopathological Sociaty meeting in August 2020. I presented the findings from my research through this grant at an e-poster session. At this virtual conference, I was able to network with other peers in my field as well as chair the graduate committee meeting. I was able to serve on the APS Foundation Board which aims to provide financial aid to graduate students to attend the meeting. I also organized and led a round table discussion titled "Working in Other Countries" about navigating US immigration for international students and employees. During my project, I hired an undergraduate researcher through the grant. I learned how to write a job posting with clearly defined expectations and qualifications. I mentored the undergraduate student on basic lab safety, lab maintenance, basic microbiology, plant care and maintenance, bacterial inoculation assays and sampling, and data analysis. She maintained her own lab notebook that we went over weekly. By the end of the second semester, she was a self-sufficient employee that could work independently and with little supervision. How have the results been disseminated to communities of interest?Results have been disseminated to the scientific community through oral presentations at regional and national conferences. Results have also been published through pre-print journals and disseertations. What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
Under these goals, I have screened ten cultivars of kale and collards for variation to opportunistic colonization of two Non- toxigenic E. coli O157:H7 strains (E. coli O157:H7 511 and E. coli O157:H7 BDMS T4169). The ten cultivars I used were recommended by the Georgia vegetable extension agent, Tim Coolong. The ten cultivars I tested are: • Georgia Green • Georgia Southern • White Mountain • Top Bunch • Vates • Morris Heading • Alabama Blue • Evenin Star • Flash • Champion I have found that there is variation in E. coli colonization during plant disease by P. syringae between strains and between cultivars of collards/kale. Both strains of E. coli O157:H7 5-11 (511) and BDMS T4169 (ATCC) had no difference in colonization of cultivars Vates, and Top Bunch regardless of whether they were co-inoculated with DC3K and HrcC or inoculated by itself. In the cultivar Georgia green, there was no different in colonization of strain 511 but the ATCC strain exhibited significantly greater colonization when co-inoculated with the HrcC strain than pathogenic DC3K strain. These phenotypes were maintained for all repeated experiments which suggest that these cultivars are good candidates for resistance to improved colonization of E. coli O157:H7 even during plant disease caused by P. syringae. In contrast, both O157:H7 strains had significantly greater colonization when co- inoculated with T3SS+ than T3SS- or by itself in B. oleracea var. acephala cultivars Alabama Blue, Morris Heading and Evenin' Star.Therefore, association with the plant pathogen, DC3K, can promote the growth of both strains of O157:H7 and this is most likely due to thesuppression of PTI by DC3K effectors delivered by the T3SS. As these cultivars exhibited the most consistent phenotype between experimental repeats, we selected these cultivars as good susceptible candidates for improved colonization of E. coli O157:H7 even during plant disease caused by P. syringae. The fact that this observed phenotype was not consistent from experiment to experiment despite our best efforts to maintain continuity in all variables, suggests that there are numerous standards that need to be met in order to create a permissive apoplastic environment for O157:H7 colonization during plant disease. Previously, we had demonstrated that the initial inoculum of either the P. syringae or E. coli O157:H7 strain is an important factor for successful colonization during plant diseaseWe had previously demonstrated this variability in colonization of enterics in apoplastic fluid collected from the two model organisms, Arabisopsis thaliana and Nicotiana benthamiana, with Salmonella enterica strains exhibiting more pronounced growth in apoplastic fluid collected from N. benthamiana than A. thaliana. Future testing of O157:H7 growth in apoplastic fluid collected from the resistant and susceptible candidate collard cultivars would shed some light as to whether variations in apoplastic-derived carbon sources influence apoplastic colonization of O157:H7 during plant disease. Although both O157:H7 strains exhibited similar behavior in our candidate resistant and susceptible cultivars, they exhibited variable opportunistic colonization during infection in some cultivars. For instance, in the Flash cultivar, the O157:H7 strain, BDMS T4169, had significantly greater colonization when co-inoculated with DC3K than when co-inoculated with HrcC or by itself. In contrast, O157:H7 strain, 5-11, had no difference in colonization regardless of its co-inoculation partner or lack thereof in the same cultivar, Flash. The opposite phenotype was observed in the Georgia Southern cultivar in that 5-11 exhibited enhanced colonization during disease but BDMS T4169 did not. This suggests that there are strain factors that contribute to opportunistic colonization of O157:H7 during plant disease. Previously we investigate several potential strain factors and found that variations in regulation of the general stress response, via the RpoS sigma factor, contribute to differential colonization of S. enterica in model hosts. Natural variations in RpoS levels in E. coli have been found to attribute to differential abilities in nutrient acquisition during starvation and survival during exposure to oxidative stress and low pH. It is possible that RpoS levels could vary between our two strains of O157:H7. Further characterization of rpoS expression and production during in planta colonization will need to beevaluated in our two strains in order to determine if RpoS plays a role in the differential colonization observed in cultivars Flash and Georgia Southern.
Publications
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2020
Citation:
Lovelace AH, Lee S, Ayoub S, Downs DM, Kvitko BH. March 2020. Variation in opportunistic colonization of enteric bacteria during plant disease on model and crop plant hosts by Pseudomonas syringae. Georgia Association of Plant Pathologists Meeting. Augusta, GA.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2020
Citation:
Lovelace AH, Lee S, Ayoub S, Downs DM, Kvitko BH. August 2020. Host and strain factors contribute to variation in colonization potential of enteric bacteria during Pseudomonas syringae plant disease. American Phytopathological Society.
- Type:
Theses/Dissertations
Status:
Published
Year Published:
2020
Citation:
Lovelace AH. (2020) Genome-Wide Bacterial Response to Plant Innate Immunity. University of Georgia. ProQuest Dissertations Publishing. 28154583.
- Type:
Journal Articles
Status:
Other
Year Published:
2020
Citation:
Lovelace AH, Lee S, Down DM, Soufi Z, Bota P, Preston GM, Kvitko BH. (2020) RpoS Contributes to Successful Opportunistic Colonization by Human Enteric Pathogens during Plant Disease. bioRxiv. https://doi.org/10.1101/2020.11.24.397208
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2019
Citation:
Lee S, Lovelace AH, Kvitko BH. (2019) Salmonella enterica Response to Immune Active and Suppressed Environments Established by Plant Pathogen, Pseudomonas syringae pv. tomato, in Model and Crop Plant Hosts
Center for Undergraduate Research Opportunities Symposium. University of Georgia, Georgia.
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Progress 06/01/19 to 05/31/20
Outputs
Target Audience:Efforts/Target Audiences: Labratory instruction and mentoring of a female undergraduate employee who is pursuing a degree in Agricultural & AppliedEconomics at The University of Georgia. Gave oral presentation on research titled "Variation in opportunistic colonization of enteric bacteria during plant disease on model and crop plant hosts byPseudomonas syringae"at the Georgia Association of Plant Pathologist Meeting to industry representatives, students, and faculty from Univerity of Georgia Plant Pathology department. Changes/Problems:Major problems that have occured in this project include a delay in obtaining the toxigenicE. coliO157:H7 strains from the CDC. It took an extended period of time to find a point of contact that could find and deliver the isolates. Overall. it took a total of nine months to obtain the strains, even with consistent follow-up emails and calls. Additionally, the COVID-19 pandemic put a hault on research for several months. The University of Georgia discontinued any non-essential research and I was unable to continue my research during the months of March, April, May, and June. Furthermore, once non-essential research is allowed to resume, there will be restrictions on the amount of time I can work on the project and work within the lab to reduce possible contamination to the virus. What opportunities for training and professional development has the project provided?I attended a professional development workshop series as part of The Plant Center Retreat at Unicoi State Park, GA in October 2019. The workshop touched on the following topics: Teaching tools in the classroom Money management in graduate school Create publish worthy figures Academic writing The workshop gave me some guidance on how to imporve my academic writing and figure making for furture publications in perr-reviewed journals. I used the grant to pay for the workshop fee. I also attended the American Phytopathological Society Leadership Institute Workshop during theAmerican Phytopathological Society meeting in August 2019. This workshop used interactive group activities to practice leadership skills and define what makes a good leader. I learned a lot about myself in terms of my leadership style and how to improve upon those skills. During my project, I hired an undergraduate researcher through the grant. I learned how to write a job posting with clearly defined expectations and qualifications. I mentored the undergraduate student on basic lab safety, lab maintenance, basic microbiology, plant care and maintenance, bacterial inoculation assays and sampling, and data analysis. She maintained her own lab notebook that we went over weekly. By the end of the second semester, she was a self-sufficient employee that could work independently and with little supervision. How have the results been disseminated to communities of interest?Results have been disseminated to the scientific community through oral presentations at regional conferences. What do you plan to do during the next reporting period to accomplish the goals?In the next reporting period, I plan to finish screening the toxigenic panel of E. colistrains in a resistant cultivar of collard. I plan to sequence the E. coliO157:H7 511 strain during co-inoculation with P. syringaewith and without a functional Type III Sectretion System, to determine the differences in the apoplastic environment created by the pathogen. I plant to compare the genomes of these enteric pathogens to find genetic differences that may explain variation in colonization during disease. I plan to validate these differences through genetic manipulation of these strains. I plan to publish my findings in a peer-reviewed journal. I also plan to include this data in my dissertation which will be published upon graduation. I will continue to present my findings atnational and regional conferences suchas The American Phytopathological Society and The Plant Center Retreat.
Impacts
What was accomplished under these goals?
Under these goals, I have screened tencultivars of kale and collards for variation to opportunistic colonization of two Non-toxigenicE. coliO157:H7 strains (E. coliO157:H7 511 and E. coliO157:H7 BDMS T4169). The tencultivars I used were recommended by the Georgia vegetable extension agent, Tim Coolong. The tencultivars I tested are: Georgia Green Georgia Southern White Mountain Top Bunch Vates Morris Heading Alabama Blue Evenin Star Flash Champion I have found that there is variation in E. colicolonization during plant disease byP. syringaebetween strains and between cultivars of collards/kale. Top Bunch, Vates, and Georgia Green are consistently resistant to E. coligrowth of both strains during disease caused by P. syringae. Alabama Blue, Morris Heading, and evening star are consistently susceptible to one or both strains of E. coliduring disease caused byP. syringae.This suggests that there are strain and cultivar factors that contribute to this phenotype. We have obtained E. coliO157:H7 isolates from the CDC that were isolated from various sources during the romaine lettuce outbreak. This panel of seven toxigenic strains are to be used for screening for natural variation of opportunistic colonization between strains in a resistant host.
Publications
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2020
Citation:
Lovelace AH, Lee S, Ayoub S, Downs DM, Kvitko BH. March 2020. Variation in opportunistic colonization of enteric bacteria during plant disease on model and crop plant hosts by Pseudomonas syringae. Georgia Association of Plant Pathologists Meeting. Augusta, GA.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2020
Citation:
Lovelace AH, Lee S, Ayoub S, Downs DM, Kvitko BH. August 2020. Host and strain factors contribute to variation in colonization potential of enteric bacteria during Pseudomonas syringae plant disease. American Phytopathological Society. Denver, CO.
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