Fungal leaf spots: field, lab, and online tutorial for professionals in Guam and the Northern Mariana Islands

Objective 1) Increase the technical capacity of the University of Guam to train agricultural professionals about foliar fungal leaf spots through the assembly and development of training materials.

Activity: Foliar, filamentous pathogens were identified for use in fact sheets and as type specimens for the Simplified Key. Using the Guam Plant Disease Index and other sources, 89 pathogen candidates were identified. The candidates were ranked according to their incidence on Guam (common, occasional, rare), and importance to Guam’s economy (high, moderate, low).

Product: Fact sheets were developed for 16 important pathogens (Table 1). Drafts of the fact sheets were distributed to trainees at the foliar fungal workshop held in August 2021. All 16 fact-sheets are now finalized and posted on the UOG Cooperative Extension & Outreach website here: https://www.uog.edu/extension/agriculture-and-natural-resources#Resources under the “Plant Disease Diagnostics 2021” tab. They are also attached to this report.

Table 1. Compiled fact sheets of filamentous, foliar pathogens in Guam and the diseases they cause.

Product:  A simplified key to asexual morphological structures to Guam’s common leaf spots including Colletotrichum, Cercospora, and Corynespora and others was developed. Based on the pathogen ranking, 18 were chosen as type specimens (Table 2). A draft key was distributed to trainees who attended the August 2021 foliar fungal workshop. After the workshop, the simplified key was finalized and posted to the UOG Cooperative Extension and Outreach website here:  https://www.uog.edu/_resources/files/extension/publications/Pathogen_Simplified_Key.pdf. It is also included as an attachment on this report. 

Table 2. Guam foliar pathogens chosen for inclusion in the simplified key.

Objective 2) Increase diagnostic collaboration between the Universities of Guam, Georgia, and the Philippines by identifying unknown species of Colletotrichum, Cercospora, and Corynespora and other fungal leaf spots that occur on Guam and enter such information into the Guam Index of Plant Diseases and other databases.

Activity: Twenty-five leaf samples were collected between the months of November and December 2020 (Table 3).

Table 3: Sample information for fungal id

Activity: Field symptoms and microscopic images were photographed (Figure 1, 2, 3). Leica MZ95 dissecting scope was used for the photomicrographs. If spores were found they were scraped off the leaf sample and wet mounted on a glass microscope slide and examined under a Leica DM1000 compound light microscope.

Duplicates of each leaf sample were dried over two-day periods and pressed between cardboard sheets for later use in PDP training (Figure 4).

Figure 4: Examples of duplicate samples dried on herbarium sheets for future use in diagnostic educational workshops. Credit: B. Deloso

Activity: To assist in morphological identification of leaf fungal pathogens, help from the lab of Dr. Marin Brewer at the University of Georgia (UGA) was utilized. Ultimately, the University of Georgia identified these samples molecularly. Leaf samples were sent on the same day of field collection to UGA from Guam. After receiving samples, graduate student Manuela Samaco assessed the condition upon arrival, took photographs, and attempted to identify presence or absence of spores. If spores were found photographs were taken using a light and dissecting scope. Samaco attempted to get subsets of each sample into tissue culture, some were successful. Some samples had spores observed while others did not. Spores were later observed on 11 out of 25 samples by UGA after arrival. Possible identities were listed by mycologists at UGA based on leaf spot symptoms and spore morphology (Table 4).

 Table 4: Sample information and tentative identification.

Activity: Specimens were cultured if spores were present on the tissue sample and the pathogen was one that was known to be culturable. Using spores from the culture grow-outs, the University of Georgia was able to molecularly identify 14 fungal pathogens from 9 different host plants using Fungal ITS Sequencing (Table 5).

Table 5. Fungal leaf pathogens from different host plants collected from Guam (2021) identified by the University of Georgia using ITS Sequencing. (*unidentified: based on culture and spore morphology in PDA)

Product: The University of Georgia tissue sample analyses were added to the Index of Plant Diseases in Guam. The updated Index can be found on the University of Guam Cooperative Extension and Outreach website here: https://www.uog.edu/_resources/files/extension/publications/Plant_Diseases_Guam_Index_10_25_2022.pdf

Objective 3) Increase the skills of agricultural professionals in Guam and the Northern Mariana Islands to diagnose fungal leaf spots and to be able to pass this information on to their constituents, through the production of a diagnostic workshop composed of field trips, lectures, and microscopic examination of plant samples.

Activity: A three and one-half day workshop/training was held August 11-14, 2021 on identification of prevalent fungal leaf pathogens and their diseases. Fourteen professionals attended: four from Northern Marianas College, four from Guam Department of Agriculture, one from Guam Landscape Management Systems, and five from the University of Guam. Topics included differentiating among similar and often confused symptoms caused by three common and virulent fungal genera: Colletotrichum, Cercospora, and Corynespora, how to collect and process fungal samples, and how to use dissecting and compound microscopes. The workshop was hosted by the University of Guam PDP Project Coordinator Dr. Robert Schlub, University of Georgia Professor of Plant Pathology Dr. Robert Kemerait, University of Georgia Associate Professor of Mycology Dr. Marin Brewer, University of Georgia Plant Pathology Academic Professional Associate Dr. Kisha Shelton, and University of the Philippines Assistant Professor Dr. Leilani Sumabat-Dacones.  

            Prior to the start of the workshop, trainees were given hard copies of all training materials and print-outs of PowerPoint presentations. Training materials included final drafts of 14 foliar fungal factsheets, final draft of the simplified table key, a copy of the Index of Plant Diseases in Guam, excerpts from The Diseases of Cultivated Crops in Pacific Island Countries, and numerous other excerpts from publications, PowerPoints, and online materials which contained pictures and descriptions of foliar fungal diseases. Trainees were also given a tote bag which contained sample collecting and processing supplies, including a clipboard, a notepad, pen, sharpie, Ziploc bags, flagging tape, a hand lens, and a basic dissection kit. All training materials were posted on the University of Guam website, and can be found here: https://www.uog.edu/extension/agriculture-and-natural-resources#Resources under the “Plant Disease Diagnostics 2021” tab.

The following was the agenda for the workshop: [WSARE PDP Workshop Final Agenda]

            During the first two days of the workshop, classroom training was held in the morning hours and lab/field training was held in the afternoon. On the third day, the field training was held in the morning to accommodate the farmer’s schedule, and classroom training was held in the afternoon. Trainees were given instruction on identifying foliar pathogens and how to collect samples. During the fourth and final day, the Guam public was invited to bring in suspected plant disease samples for a trainee-hosted half-day diagnostic clinic at the University of Guam Agriculture and Life Science building laboratory. Over the course of the workshop, trainees were instructed on how to create their own YouTube tutorial which could be used teach others how to diagnose foliar fungal pathogens and specifically how to differentiate between Colletotrichum, Cercospora, and Corynespora. 

            During the first day of training, trainees received an introduction to foliar pathogens, how to ID foliar pathogens, and how to collect and process foliar fungal samples (Figure 5). Instructors were Drs. Schlub, Kemerait, and Brewer. In the afternoon, trainees moved into the lab where Extension Associate Ben Deloso gave a quick review of how to use the dissecting and compound microscopes which were set up for trainees to use (Figure 6). This was followed by a quick walking tour of the campus demonstration garden, where trainees were encouraged to flag and collect samples. Trainees then headed back to the lab where they examined and processed any samples collected (Figure 7).

Figure 5. Dr. Marin Brewer gives a presentation on foliar pathogens. 

Figure 6. Extension Associate Ben Deloso reviews how to use a microscope. 

Figure 7. Trainees returned to the lab and examined samples under the microscope. 

             During the second day of training, instruction began with a zoom presentation on how to create a YouTube tutorial by Dr. Shelton (Figure 8). Next, instruction focused on the life cycle and ID of Colletotrichum, Cercospora, and Corynespora. Instructors were Drs. Brewer and Sumabat-Dacones. In the afternoon, trainees and instructors made a field trip to Bernard Watson’s farm in Yigo, where a brief introduction was conducted by Dr. Robert Schlub and Mr. Frank Cruz, who has assisted Watson’s farm for many years (Figure 9). Mr. Watson was unable to be present due to health concerns. The Watson farm mainly grows banana plants and noted their two most concerning diseases are banana bunchy top virus and black sigatoka. Participants were encouraged to collect as many leaf samples as they could (Figure 10). Next, the field trip moved on to John Mesa farm in Dededo. Upon arrival, the farmer gave a brief introduction to the farm, and discussed his struggles with pests, certain pesticides, and fertilizers (Figure 11). John Mesa farm grows a wide variety of vegetable and fruit crops. Trainees were allowed to speak to the farmers and ask questions, collect samples, and take notes (Figure 12).

Figure 8. Dr. Kisha Shelton gives a presentation via Zoom. 

Figure 9. A brief introduction of Bernard Watson's farm was conducted by Dr. Robert Schlub and Frank Cruz. 

Figure 10. Trainees collect leaf samples from a banana tree plot at Bernard Watson's farm. 

Figure 11. One of the workers at John Mesa farm presented a short history of the farm. 

Figure 12. Trainees collect samples at John Mesa's farm. 

             During the third day of training, trainees and instructors began with a field trip to Ernie Wusstig’s farm and the University of Guam Triton Farm. Mr. Ernie Wusstig introduced his farm (Figure 13), which manly focuses on corn and field crops. Trainees were allowed to explore the fields, examining and collecting samples (Figure 14, 15). Next, the field trip moved on to the University of Guam Triton Farm. Trainees continued collecting samples, and Dr. Robert Schlub pointed out specific diseases found at the farm (Figure 16). In the afternoon, Dr. Leilani Sumabat-Dacones gave a virtual lecture on Molecular tools in fungal detection (Figure 17). This was followed by a two-hour open lab where trainees were given the opportunity to examine any samples collected over the past two days (Figure 18).

Figure 13. Mr. Ernie Wusstig gives an introduction to his farm to trainees. 

Figure 14. Trainees walk through corn fields. Mr. Wusstig is available to answer any questions. 

Figure 15. A trainee uses a hand lens to examine a disease symptom on a corn leaf. 

Figure 16. At the UOG Triton Farm, Dr. Robert Schlub points out damaged roots caused by a fungal pathogen. 

Figure 17. Dr. Leilani Sumabat-Dacones gives a virtual presentation. 

Figure 18. Dr. Robert Kemerait (red hat) gives instruction to a trainee during the open lab period. 

            During the fourth day, the Guam public was invited to bring in suspected leaf disease samples for a free trainee-hosted diagnostic clinic at the University of Guam Agriculture and Life Sciences building laboratory. The free clinic began with a short presentation by Dr. Robert Schlub on how to identify Guam’s common fungal diseases. The group was then taken on a short walking tour of the campus demonstration garden, where Dr. Schlub pointed out some fungal pathogen symptoms on a breadfruit leaf (Figure 19). The public was allowed to collect samples as they walked. At the end of the tour, the agricultural trainees assisted the public in diagnosing foliar plant disease samples using what they had learned over the previous three-day workshop (Figure 20).

Figure 19. Dr. Robert Schlub points out fungal symptoms on a breadfruit tree leaf. 

Figure 20. Trainees assisted in diagnosing disease samples from the public and used field trip samples to provide additional samples to share with island residents. 

Product: A YouTube tutorial was developed. On the last day of the workshop, trainees and instructors worked together to develop ideas and a working model for a YouTube tutorial to distinguish between these foliar infections. With assistance from Dr. Kisha Shelton at the University of Georgia, Extension Associates at the University of Guam assembled the YouTube tutorial and placed it online. It can be found on the UOG Cooperative Extension and Outreach YouTube page here: https://www.youtube.com/watch?v=QQiSizXChtw&ab_channel=UOGCNAS

Objective 4) Use workshop evaluations consisting of surveys and pre and post-tests to measure knowledge gained and identify opportunities for future plant pathology related activities and the means through which delivery of diagnostic skills and knowledge to trainees or stakeholders in future workshops can be improved.

Activity: Participant change in knowledge was measured using pre and post-tests. Pre-tests were given to each participant prior to the workshop, which was followed by post-tests at the conclusion of the workshop. Three participants were not present to take the post-test. The average gain in knowledge from the 11 agricultural professionals who completed both pre- and post-tests was 27.5 percent.

A post-workshop evaluation was also given to the 14 participants and consisted of 21 questions:

1) Pre-workshop communication was well organized; 2) Was it clear that the workshop objective was to provide you with an opportunity to gain skills and knowledge useful in discerning leaf spots caused by fungi as opposed to those caused by other pathogens or pesticides?; 3) After completing the workshop, do you feel like you were well informed about the objectives of this workshop?; 4) I found the content of this workshop to be relevant to my job; 5) I found the classroom materials to be useful; 6) The workshop activities stimulated my learning; 7) The activities in this workshop gave me sufficient practice and feedback; 8) The difficulty level of this workshop was appropriate; 9) The pace of this workshop was appropriate; 10) The field trips were a good addition to the classroom instruction; 11) The lecture room was clean and well set-up; 12) The laboratory was clean and well set-up; 13) I was satisfied with the snacks, lunch, and beverages provided; 14) The instructor was well prepared; 15) I learned something from this instructor; 16) The instructor was helpful; 17) I learned how to diagnose common foliar fungal genera found on Guam; 18) My skills/ knowledge have improved by attending this workshop; 19) I learned valuable information/ skills that can be applied to my work; 20) I am confident that I can teach others what I learned; 21) Rate your overall workshop experience.

Score was based on a scale of 1 (strongly disagree) to 5 (strongly agree). The average overall score was 4.7. The lowest average score for a question was 4.00 while the highest score for a question was 5.00.

            Due to the increase in knowledge gained and the overall positive workshop evaluation score, the participants in the training will have a positive impact on diagnosing foliar fungal diseases for their respective agencies.