1a. Objectives (from AD-416):
1. To explore improvements to formulation of biocontrol yeast for field application by growers. Types of formulations are wet formulations with additives (preservatives, nutritional supplements, surfactants for spreading) and dry formulations for delivery of viable yeast as a dry powder for tank mixing. 2. Evaluation of yeast longevity under different storage and application conditions: time and temperature. 3. Application for controlling l of other microbial pathogens: plant pathogenic fungi, microbial contaminants of crops (Salmonella, E. Coli)
1b. Approach (from AD-416):
Objective 1. Protective additives will be added to wet and dry yeast formulations and evaluated for improved shelf-life under lab conditions. Spray trials in the field may be used to evaluate effects on durability on plant leaves and fruits. Surfactants will be in the tank mix for even spreading. Objective 2. Yeast durability in a fully diluted mixture will be evaluated through time and temperatures series in lab so the upper threshold that might be expected under grower conditions. Objective 3. Ability to control fungi on agricultural crops. evaluations will be done using greenhouse tests on plants and pathogens of significance in tree fruit-nut and vegetable crops. Salmonella is a particular problem on almond. Initial experiments may involve field sprays on almond followed by collection of nuts and inoculation with Salmonella in lab-greenhouse and monitor Salmonella population Non-pathogenic E. Coli strains applied to lettuce and spinach in the greenhouse followed by yeast spray applications to determine whether potential control is possible. The effectiveness of the yeast against additional fungi of agricultural importance will initially be evaluated on tomato and potato. Documents SCA with UC-Davis.
3. Progress Report:
Saprophytic yeast, Pichia anomala has been demonstrated to inhibit the growth of Aspergillus flavus as well as aflatoxin production by this fungus. It was also shown to inhibit Alternaria spp, Penicillium spp, Botrytis spp in lab assays. A stable liquid formulation of the biocontrol yeast product has been developed and used by spraying onto horticultural crops. Yeast cells can be stored at 40C for up to six month with very little change in viability. Field trials to control Alternaria disease in pistachio orchards have been conducted over the past four years. Significant control of A. alternata were observed in 2009. Nut cluster damage was also evaluated and treatment effects for this measure of A. alternata infection was highly significant (P=0.001). Cluster damage was also moderately correlated (r=0.47, P=0.002) with yield differences. Pichia anomala was tested as a biocontrol agent to reduce the survival and growth of attenuated E. Coli 0157:H7 strain on spinach. The experiments were carried out in a quarantined green house. A water suspension of P. anomala was applied to spinach 24 hr before inoculating E. coli. Spinach plants were harvested at day 4 and day 7 after the treatment. The population of E. coli on spinach was reduced by five fold in comparison to plants sprayed with the bacteria without yeasts. For the harvest mold experiment, in-hull almonds were used. The results indicated the nut samples contain high incidence of harvest molds and the yeast does have an inhibitory effect on the growth of harvest molds for in hull almond samples. It was also observed that the nut control samples contain fungal growth with sporulation while the fungal growth in the yeast control and the yeast treated samples has no visual evidence of sporulation. In the experiment with E. coli, we tested five replicates of almonds in dried hulls. The results indicates the dried in shell almonds had a lower infection rate of bacteria by the yeast treatment. The goal of the specific cooperative agreement is to develop practical applications of P. anomala to control pathogenic fungi and bacteria, which contributes directly to Objective 4 of the in-house project.