1University of Pennsylvania, Horticulture Department, State College, PA 16802, USA; 2USDA-ARS, Biocontrol of Plant Diseases Laboratory, Beitsville, MD 20705, USA

Background and objectives
At least five major cacao (Theobroma cacao) diseases destroy 40% of the total crop annually in cocoa producing countries [1]. A large percentage of this loss are due to fungal diseases such as black pod caused by Phytophthora spp., which is of worldwide importance; and witches' broom caused by Crinipellis perniciosa, presently restricted to South America. The current strategies available to control these diseases, including cultural practices, phytosanitation, fungicidal sprays and breeding for resistance, are labour intensive or at an early stage of development. However, research on biological control has been limited to antagonistic fungi such as Cladobotryum amazonense and Trichoderma spp., which were shown to have some potential to control witches' broom [2]. The constraints that exist for the large-scale application of microbial agents are lack of knowledge on the ecology of cacao-associated microorganisms and lack of selection and application strategies of microbial strains efficient under various agroclimatic conditions. The objective of this investigation was to study the microbial populations associated with the various plant parts of cacao thought to be the 'sites of infection' for cacao fungal pathogens and to identify the 'major group' or 'groups' of microorganisms associated with cacao with a potential for biological control.

Materials and methods
Serial dilutions of plant samples such as macerated leaves and flowers, scrappings from pod surfaces, brooms, roots and rhizosphere soils from cacao plantations were plated on selective and non-selective media for yeasts, bacteria, actinomycetes and fungi. Cacao cultivars susceptible (UF613) and tolerant (Scavina 6) to witches' broom, were used for the study. Approximately 1300 strains were purified and stored under glycerol at -7OC. A subset of strains were identified using morphological characteristics and biochemical tests and also tested for their antifungal activity in vitro against Phytopthora spp. and on leaf discs against C. perniciosa.

Results and conclusions
The study indicated that bacteria, fungi and actinomycetes were found associated with both aerial parts and the rhizosphere of cacao. However, yeasts were detected only on the aerial parts of the plant. Bacteria associated with the aerial parts were highly pigmented. Although total populations of bacteria, fungi and actinomycetes from the resistant and tolerant cultivars were not different from each other on the various plant parts, total populations of yeasts were significantly higher in the resistant than in the tolerant cultivar. Some of the classical biocontrol agents such as fluorescent pseudomonads, Burkholderia cepacia and Trichoderma spp. were detected mainly in the soil and on the roots. A few of these (yeasts, bacteria, actinomycetes and fungi) exhibited antifungal activity against Phytophthora spp. and C. perniciosa. Greenhouse tests are in progress to study the effects of these microorganisms on disease suppression.

1. Van der Vossen HAM, 1996. Proceedings International Workshop on the Contribution of Disease resistance to Cocoa Variety Improvement, pp. 27-31.
2. Bastos CN, 1996. Fitopatologia Brasileira 21, 50-54.