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

Progress report for PDP20-001

Project Type: Professional Development Program
Funds awarded in 2020: $66,013.00
Projected End Date: 08/31/2022
Host Institution Award ID: G120-21-W7903
Grant Recipients: University of Guam; University of Georgia
Region: Western
State: Guam
Principal Investigator:
Dr. Robert Schlub
University of Guam
Co-Investigators:
Dr. Marin Brewer
University of Georgia
Dr. Robert Kemerait
University of Georgia
Dr. Kisha Shelton
University of Georgia
Dr. Leilan Sumabat-Dacones
University of Philippines
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Project Information

Abstract:

The cornerstone of a sustainable plant disease control program for any farm community hinges on well-trained agricultural professionals. Diagnosing diseases in Guam and the Northern Mariana Islands is a constant challenge because of the yearly movement of nearly 2 million tourists and the importation of large quantities of fresh produce. Compounding the problem is the islands’ remoteness from mainstream plant pathology research and teaching facilities. Project’s long-term goals are to increase economic viability and enhance environmental quality of small commercial and subsistence farming operations in the region through accurate and timely diagnoses of plant diseases. The project’s immediate goal is to provide agricultural professionals with the skills and knowledge useful in discerning leaf spots caused by fungi as opposed to those caused by bacteria or chemical sprays.  Trainees will be provided with the skills necessary to differentiate among similar and often confused symptoms caused by three common and virulent fungal genera: Colletotrichum, Cercospora, and Corynespora. Using trainers from Georgia and the Philippines, will bring a national and  an international perspective to the workshop. Travel cost for the trainers will be kept to a minimum by scheduling the workshop to coincide with their other travel plans. Activities include: 1) Guam assembling workshop materials; 2) Guam collecting images of field symptoms and micrographs of spores from 15 foliar fungi including Colletotrichum, Cercospora, and Corynespora; 3) Guam preparing single spore isolates; 4) Georgia identifying isolates and depositing sequences to GenBank; 5) Guam updating Guam’s Index of Plant Diseases; 6) Guam, Georgia and Philippines conducting a four-day plant disease diagnostic workshop for 12 trainees composed of field trips, lectures, hands-on examination of samples, laboratory exercises, and engagement opportunities with the local farmers and the public; and 7) Guam, Georgia and Philippines posting identification tutorial online; and 8) Georgia and Guam conducting a workshop evaluation.

Project Objectives:

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.

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.

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.

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.

Timeline:

Proposal’s principle assumption is the belief that improvements in the skills of agricultural professionals in discerning various fungal leaf spots will translate into more specific and targeted recommendations to their clientele.

Inputs include expert workshop trainers (Dr. Schlub, Dr. Kemerait, Dr. Brewer, and Dr. Sumabat-Dacones); a mycologist with expertise in molecular techniques (Brewer et al., 2014; Stewart et al., 2015; Sumabat et al., 2018); and a Co-PI with expertise in instructional evaluations. Direct financial contributions will come from WSARE, while salary support will come from the PI, Co-PI, and trainees’ respective institutions and governments. Farmer volunteers will host field trips and participate in a half-day clinic for growers and the public.

The workshop will follow the “Train the Trainer” model and will include classroom and field instructions. On the final day, trainees will have an opportunity to use their newly acquired knowledge during a half-day clinic for growers and the public.  Dr. Shelton will use the Don Dillman’s tailored Design method (Dillman et al. 2014) to formulate survey questions and the IBM SPSS Statistics software (IBM 2017) in data analysis.

Activity 1: September 2020-February 2021: Create an online workshop manual.

 Activity 2: September 2020-January 2022: Guam will collect images of field symptoms and micrographs of spores of 15 foliar fungi including species of Colletotrichum, Cercospora, and Corynespora.

 Activity 3:  September 2020-January 2022: Two sets of 15 single spore fungal isolates will be prepared. One will be sent to Georgia for ID the other maintained on Guam.

 Activity 4: January 2021-August 2021: Trainers will develop a tutorial on distinguishing foliar infections caused by Colletotrichum, Cercospora, Corynespora, and other fungi. A working model will be available at the time of the workshop for trainees’ testing and feedback. When finished the tutorial will be placed online.

 Activity 5: March 2021: Trainers will conduct a four-day plant disease diagnostic workshop, composed of field trips, lectures, and microscopic examination of samples. There will be 4 trainers and 12 trainees consisting of seven from Guam and four from Northern Mariana Islands.

 Activity 6: May 2021-January 2022 Guam and Georgia will provide a written evaluation of the workshop and pre and post-tests, along with solicited suggestions from trainees for future workshops.

 Activity 7: January 2022-July 2022: Georgia will identify fungal cultures and deposit DNA sequences to GenBank.                                                                                                

Activity 8: August 2022: Guam and Georgia will add new fungal identification to Guam’s Index of Plant Diseases. 

Cooperators

Click linked name(s) to expand
  • Bernard Watson - Producer
  • Dr. Marin Brewer (Researcher)
  • Dr. Robert Kemerait, Jr. (Educator)
  • Dr. Kisha Shelton (Educator)
  • Dr. Leilani Sumabat-Dacones (Educator)

Education

Educational approach:

No educational approach has been used during this initial five-month reporting period.

Education & Outreach Initiatives

WSARE PDP Training
Objective:

Train Agriculture Professionals

Description:

No educational and outreach initiatives has occurred during this initial five-month reporting period.

Outcomes and impacts:

No learning and action outcomes has occurred during this initial five-month reporting period.

Project Outcomes

4 New working collaborations
Project outcomes:

          A training manual for identifying leaf spot diseases is being crafted. This manual consists of fact sheets developed using pictures of leaf spots recently gathered for this grant. The fact sheets include the name of the pathogen, condia morphology including microscope digital photos of the condia taken in our lab, leaf spot photos using a camera and microscope, a description of the disease development on Guam, and a description of the symptoms. Also, a simplified key with other information is being developed to compliment the fact sheets.

          We collected 25 leaf samples from plants around Guam between the months of November and December 2020. A google drive account was created to share sample data in real time with the University of Georgia.  The majority of leaf samples collected were of economically important crops but some leaves displaying pathogen symptoms were collected on ornamentals and native plants. Locations included farms in Northern Guam, the University of Guam campus, the tourism district, and coastal areas. Date and GPS location of each leaf sample was recorded and cataloged in our plant clinic log book.

 

Additional Outcomes:

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.

Objective 1 Activities, Products, Progress.

Assembly and development of training materials:

(1) Photographic key of fungal leaf spot symptoms caused by Colletotrichum, Cercospora, and Corynespora and other fungi.

          A list of Guam foliar pathogen genera was compiled, each was rated for incidence on Guam (common, occasional, rare), importance to Guam’s economy (high, moderate, low), and frequency reported in Guam’s Disease Index.  Eighty-nine pathogens were identified. From this list of pathogens approximately 15 will be chosen for the photographic key (fact sheets). A draft copy of a fact sheet follows:

(2) A simplified key to asexual morphological structures to Guam’s common leaf spots including Colletotrichum, Cercospora, and Corynespora;

          A list of Guam foliar pathogen genera was compiled, each was rated for incidence on Guam (common, occasional, rare), importance to Guam’s economy (high, moderate, low), and frequency reported in Guam’s Disease Index.  Eighty-nine pathogens were identified. From this list pathogens approximately 30 will be chosen for inclusion in the simplified key. Three outlines of the simplified key have been evaluated.

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.

          We collected 25 leaf samples from plants around Guam between the months of November and December 2020 (Table 1).

Table 1: Sample information for fungal id

UOG plant clinic ID#

Sample

Plant

Location

GPS

2020-023

1

Banana

UOG, GU

13.43016, 144.80016

2020-024

2

Papaya

UOG, GU

13.43015, 144.80013

2020-025

3

Wing Bean

UOG, GU

13.43017, 144.79982

2020-026

4

Breadfruit

Yigo Experiment Station, GU

13.53266, 144.87315

2020-027

5

Casssava

Yigo Experiment Station, GU

13.53239, 144.87305

2020-028

6

Hot pepper

Yigo Experiment Station, GU

13.53235, 144.87346

2020-029

7

Okra

Yigo Experiment Station, GU

13.53237, 144.87331

2020-030

8

Mango

Chalan Pago, GU

13.42799, 144.78479

2020-031

9

Mango

Chalan Pago, GU

13.42799, 144.78479

2020-032

10

Hibiscus tiliaceaus

Yigo, Ming Chun farm, GU

13.33745, 144.53522

2020-033

11

Guava

Yigo, Ming Chun farm, GU

13.33824, 144.53568

2020-034

12

Cucumber

Yigo, Ming Chun farm, GU

13.33865, 144.54568

2020-035

13

Plumeria

UOG, GU

13.25822, 144.48069

2020-036

14

Fig

Chalan Pago, GU

13.42799, 144.78479

2020-037

15

Corn

Dededo, Ernie Wusstig farm, GU

13.25822, 144.48069

2020-038

16

Avocado

Dededo, John Mesa farm, GU

13.32311, 144.51585

2020-039

17

Avocado

Dededo, John Mesa farm, GU

13.32311, 144.51585

2020-040

18

Calamansi

Dededo, John Mesa farm, GU

13.32318, 144.51623

2020-041

19

Calamansi

Dededo, John Mesa farm, GU

13.32318, 144.51623

2020-042

20

Ipomea pes-caprae

Tumon, Ypao Beach, GU

13.25880, 144.48029

2020-043

21

Heliotropium foertherianum

Tumon, Ypao Beach, GU

13.30160, 144.4730

2020-044

22

Panicum maximum

Tumon, Hyatt Hotel, GU

13.30430, 144.48160

2020-045

23

Wedelia sp.

Tumon, Ypao Beach, GU

13.3018, 144.47340

2020-046

24

Wedelia sp.

Tumon, Ypao Beach, GU

13.3018, 144.47340

2020-047

25

Banana

UOG, GU

13.43016, 144.80016

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.

Figure 1: Example of target leaf spot on Papaya leaf sample (Photo 1) sent to UGA for molecular analysis. The pathogen is suspected to be Corynespora sp. Condidiophore of Corynespora isolated from Papaya leaf sample (Photo2). Note the distinctive attachment scar (Credit: B. Deloso).
Figure 2: Similar spores isolated at UGA from same sample. Credit; M. Samaco
Figure 3: Suspect downy mildew with conidia supporting structures. Credit: B. Deloso

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.

          To assist in morphological identification of leaf fungal pathogens, help from the lab of Dr. Marin Brewer at the University of Georgia was utilized. Ultimately the University of Georgia will identify 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 UG after arrival. Possible identities were listed by mycologists at UGA based on leaf spot symptoms and spore morphology (Table 2).

 Table 2: Collection dates and other information

date collected

collected by

suspected disease

date received in UGA

 cultured

Spores

observed

possible identity (GA)

11/5/2020

B. Deloso

Fusarium

11/10/2020

yes

yes

Curvularia, Colletotrichum

11/5/2020

B. Deloso

Corynespora

11/10/2020

yes

yes

Colletotrichum

11/5/2020

B. Deloso

powdery mildew

11/10/2020

no

   

11/13/2020

B. Deloso & J. Hudson

anthracnose

11/17/2020

yes

yes

Colletotrichum

11/13/2020

B. Deloso & J. Hudson

 

11/17/2020

yes

yes

Colletotrichum

11/13/2020

B. Deloso & J. Hudson

 

11/17/2020

yes

yes

Colletotrichum, Pestalotiopsis

11/13/2020

B. Deloso & J. Hudson

 

11/17/2020

yes

yes

Fusarium, Curvularia

11/13/2020

R. Schlub

orange algae?

11/17/2020

no

   

11/13/2020

R. Schlub

anthracnose

11/17/2020

yes

yes

Colletotrichum, Fusarium

11/27/2020

B. Deloso & J. Hudson

 

11/30/2020

yes

yes

unidentified

11/27/2020

B. Deloso & J. Hudson

 

11/30/2020

yes

yes

Pestalotiopsis

11/27/2020

B. Deloso & J. Hudson

Colletotrichum

11/30/2020

yes

no

 

11/27/2020

B. Deloso & J. Hudson

 

11/30/2020

yes

yes

Colletotrichum

11/27/2020

R. Schlub

 

11/30/2020

yes

yes

Colletotrichum, Curvularia

12/4/2020

B. Deloso & J. Hudson

Bipolaris

12/7/2020

yes

   

12/4/2020

B. Deloso & J. Hudson

 

12/7/2020

yes

   

12/4/2020

B. Deloso & J. Hudson

 

12/7/2020

yes

   

12/4/2020

B. Deloso & J. Hudson

citrus scab

12/7/2020

yes

   

12/4/2020

B. Deloso & J. Hudson

 

12/7/2020

yes

   

12/10/20

B. Deloso & J. Hudson

 

12/14/2020

     

12/10/20

B. Deloso & J. Hudson

 

12/14/2020

     

12/10/20

B. Deloso & J. Hudson

A &B, two suspects

12/14/2020

yes

yes

Bipolaris

12/10/20

B. Deloso & J. Hudson

Cercospora

12/14/2020

     

12/10/20

B. Deloso & J. Hudson

downy mildew

12/14/2020

     

12/10/20

B. Deloso

Mycosphaerella

12/14/2020

     

          Georgia cultured spores from the leaves (spots) we sent them. Some are still growing out. PCR will be performed on cultured spores for species identification.

Any opinions, findings, conclusions, or recommendations expressed in this publication are those of the author(s) and do not necessarily reflect the view of the U.S. Department of Agriculture or SARE.