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United States Department of Agriculture

Agricultural Research Service

Research Project: Assessment of Genetic Diversity and Biocontrol Strategies in the International Cacao Genebank

Location: Sustainable Perennial Crops

2012 Annual Report


1a.Objectives (from AD-416):
Part.
1)Genetic diversity This project will address three practical problems related to the control of cacao diseases, germplasm conservation, and varietal deployment in the Americas. Firstly it will address the critical need to understand the origin, dispersal and geographical variation of Moniliophthora roreri in the Americas, through the analysis of population structure and genetic diversity in CATIE’s collection of Moniliophthora roreri. Secondly, it will assess on-farm genetic diversity in the main cacao producing regions in Bolivia, as well as in other selected countries in Central America, with the intent to develop a rational strategy for on-farm conservation. Thirdly, this project will assist CATIE to verify the genetic identity of the elite clones which have been released by CATIE and are being propagated for distribution. Results of this project will contribute to more efficient management of cacao germplasm, both ex situ and on farm, better use of cacao germplasm for varietal development through the identification of new sources of resistance to moniliasis, black pod and other diseases, and rational deployment of new varieties in farmer fields.

Part.
2)Biocontrol The objectives of this research are to test, in the laboratory and in the field, the efficacy of selected biological agents for the control of cacao diseases, specifically, Moniliophthora roreri. The research will focus on endophytic Trichoderma spp. previously identified as having biocontrol potential. The development of biocontrol formulations with enhanced disease control efficacy in the field is of special interest. Little is known about how biocontrol formulations influence the establishment and efficacy of endophytic agents in tropical ecosystems, especially as applied to trees such as Theobroma cacao (cacao). The economic sustainability of optimized biocontrol strategies will be determined in order to demonstrate biocontrol’s merit as a “best practice” for cacao farmers.


1b.Approach (from AD-416):
1. Analyze geographic variation in the CATIE Moniliophthora roreri collection: Lyophilized mycelia from different populations will be prepared in CATIE and sent to USDA/ARS/BARC/SPCL. Microsatellite primers markers will be designed based on the USDA sequence information of M. roreri. Markers will be used to genotype the collection. Inter-population relationships and allelic diversity will be analyzed using haplotypes and multi-locus data. Population analysis will be used to evaluate genotypic diversity, allelic frequencies, gene diversity and population differentiation. Spatial genetic variation will be analyzed using both molecular and GIS data. 2. Assess on farm genetic diversity: Landraces or farmer selections will be selected from traditional cacao farms in Bolivia, Guatemala, El Salvador, Belize and Costa Rica. Leaf samples of cacao trees representing each landrace will be collected. Samples will be chosen based on interviews with farmers. The samples will be dried in silicone gel and sent to the SPCL for DNA extraction and fingerprint analysis using 100 SNP markers. The genetic relationship of these landraces will be analyzed using SNP data of reference populations. The asserted change in genetic diversity can be assessed by comparing the “disturbed” versus “undisturbed” farms or populations. The resultant data will allow us to measure the population structure and level of genetic diversity by farm, by region and by GIS. 3. Verify genotype identity of elite CATIE cacao clones: Leaves of the elite clones will be sampled following the guidelines established by SPCL and used for DNA extraction. The samples will be sent to SPCL and fingerprinted using 80 SNP markers. Off-types will be identified based on the DNA profiles. Parentage analysis will be applied to confirm pedigree relationships in case where parental combinations are known. Standard SNP profiles will be established for each elite clone and the SNP-based genotype verification will be used to improve and monitor varietal dissemination. 4. Access biocontrol effectiveness in field trials: At present, isolates to be considered for further field studies include 15 candidate endophytes (Trichoderma species) previously screened for biocontrol and endophytic efficacy in cacao by ARS, and several isolates of specific interest to CATIE scientists. Lab-based research on formulations with Trichoderma isolates indicate a significant benefit to Trichoderma efficacy in response to added nutrients, humectants, and vegetable oils (corn oil) and these will be used to enhance Trichoderma biocontrol efficacy in small scale and short term field studies. Formulation concepts will be optimized for readily available nutrient sources, concentrations, and application timing. Standard protocols will be developed for rapid evaluation of biocontrol agents, formulations, and their interactions in the field. Large scale field trials of promising endophyte formulations with biocontrol potential will be conducted. The economic impact of biocontrol strategies for the control of disease will be determined.


3.Progress Report:

Field applications of specific biocontrol agents to control diseases on cacao are underway. Treatments with these biocontrol agents have resulted in increased yields that are associated with reduced frosty pod rot disease. These results are linked to improved formulations that appear to increase the germination of the biocontrol organisms. Additional studies to look at the genetic diversity of the cacao pathogen Moniliophthora roreri are under way and 14 isolates of this fungus from Central and South America have been sent for RNA sequencing. The information derived from the sequencing will be compared and used to develop genetic markers to study the various populations of this fungal pathogen.


Last Modified: 4/19/2014
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