1. Field evaluation of endophytes for management of Witches’ broom on cacao
Rachel Melnick Anissa poleatewich Department of Plant pathology Penn State university
Department of Plant Pathology

2. Previous work INIAP station in Pichilingue Ecuador
Isolated endophytic bacteria from elite cacao trees escaping disease
Screened isolates at Penn State to determine which bacterial endophytes to study further

3. Growth chamber studies
Plants maintained in growth chambers
Detached leaf assays used to screen isolates
Colonized leaves challenged with pathogen

4. Ecuador field trials
Selected 4 bacterial isolates based on performance in growth chamber studies
Investigated foliar colonization of these bacteria on 5 genotypes of cacao in the field
Susceptible (EE19 and IMC67)
Tolerant (A2634 and A2126)
High yielding and Resistant (CCN51)
Colonized foliage of small potted plants and placed under larger trees in May 2007

5. Bacterial Application
Experimental System
Disease Challenge
TIME
Bacterial Application
Colonization

6. Ecuador field trials
Colonizing experiment

7. Disease challenge in Ecuador
Susceptible tree full of witches’ broom
Spores raining from infected trees onto seedlings
Seedlings undergoing disease screening

8. Preliminary observations
Endophytic colonization occurred on all genotypes
Disease developed on seedlings
Disease ratings are being taken monthly
Preliminary data has been analyzed and published in PDMR
Bacteria were re-applied to plants in November 2007, March 2008, and May 2008

9. Disease severity
One bacterial treatment has completely suppressed disease....as of March 2008

10. Disease severity
Tolerant genotypes have significantly less disease than susceptible genotypes
A2634 statistically significantly reduced disease. A2634 & A2126 are nacional cocoa and tolerant. EET19 susceptible control. CCN51 widely planted genotypes due to high yield, but did no SIGNIFICANT disease reduction

11. Coupling tolerant genotypes with bacteria can reduce disease

12. Future
Continuing INIAP trial…..Rachel is currently in Ecuador collecting data
Replicating field trial on the Nestle farm in Valencia, Los Rios, Ecuador
Inoculation of bacteria on established tree branches
ARISA – automatic ribosomal intergenic spacer analysis
qPCR – Quantitative PCR. Real time PCR used to quantify mRNA to determine the expression leaves of genes in the plant

13. Future
Molecular analysis (ARISA) of microbial populations to determine how application of biocontrol agents impacts native microbes
Molecular analysis (qPCR) of leaves to determine how endophytes affect plant defense pathways
Real time PCR used to quantify mRNA to determine the expression of genes in the plant

14. Related Projects Management of postharvest disease on apple
Achieved 90% reduction in bitter rot severity in preliminary tests  Adams county, PA
Experiment repeated in Bolivia
Giovanna Plata at PRO-IMPA

15. Acknowledgements PSU Plant Path USDA-ARS SPCL
Paul Backman
USDA-ARS SPCL
Bryan Bailey
Eric Rosenquist
INIAP
Carmen Suarez
Danilo Vera
Karina Solis
USDA-ARS SPCL
USDA-ARS International Programs
PSU CAS Tag- Along Program
PSU Plant Path. Travel Award
PSU Department of Plant Pathology
USAID
SANREM CRSP
IPM CRSP