1a.Objectives (from AD-416)
Objective 1: Elucidate the role of fungal extracellular enzymes as virulence factors important in postharvest decay of fresh fruit including the effect of pathogen modification of the host environment on virulence factors.

Objective 2: Evaluate potential control strategies targeting fungal extracellular enzymes to reduce postharvest decay of fresh fruit. Sub-objective 2.A. Optimize the application of recombinant antibodies targeting extracellular enzymes of P. expansum and P. solitum to reduce postharvest decay of pome fruits. Sub-objective 2.B. Determine the effect of antioxidants and their analogs on decay development.

1b.Approach (from AD-416)
To elucidate the role of fungal extracellular enzymes as virulence factors in postharvest decay of apple and pear fruit, we will first compare the ability of several strains of a highly virulent and a weakly virulent pathogen of the same genus to produce extracellular enzymes (such as polygalacturonases and cellulases) during fruit infection and in response to fungal modification of the host/pathogen microenvironment. 'Golden Delicious' apple and 'Anjou' pear fruit will be infected with P. expansum or P. solitum and we will extract, purify, and characterize the extracellular enzymes produced by these organisms using standard procedures. We will develop and utilize recombinant antibodies specific for binding to and inactivating enzymes produced by the pathogens in vivo and in vitro. The antibodies will allow us to determine which specific enzymes play a major role in virulence of Penicillium species on pome fruit. Strategies incorporating recombinant antibodies against virulence factors, antioxidants known to have anti-fungal activity, and mild stress treatments will be tested for their ability to control postharvest decays of pome fruits caused by Penicillium species.

3.Progress Report
Production of polyclonal antisera and a recombinant phage library against polygalacturonase produced by P. expansum. It is necessary and ecologically responsible to develop alternatives to chemical control for reducing postharvest losses. In cooperation with and ARS scientist at the Appalachian Fruit Research Station, Kearneysville, West Virginia, and ARS scientists in Beltsville, MD, have made progress toward the production of polyclonal antisera and a phage library against polygalacturonase from P. expansum, the causal agent of blue mold of apples. Polygalacturonase from P. expansum-infected apple fruit was purified using gel filtration chromatography and injected in to mice. Spleens from immunized mice were harvested and polyclonal antisera were produced. Current progress provides a foundation for the construction of a recombinant phage library specific to polygalacturonase produced by P. expansum which will be used to produce recombinant antibodies. Blocking polygalacturonase activity on the fruit surface using recombinant antibodies against polygalacturonase may lead to a novel control strategy to halt postharvest decay mediated by P. expansum on apple fruit. This control strategy could potentially benefit the postharvest industry by reducing dependency on chemical fungicides. National Program 303, component IIa.

4.Accomplishments
1. Virulence factor associated with Pencicillium solitum during decay of apple fruit.

Penicillium solitum is a blue mold fungus that can cause losses of apples in storage due to decay. We have purified and characterized a polygalacturonase produced by P. solitum in infected apple fruit. The role for this polygalacturonase in the disease process is yet to be determined and is one of the objectives of the current project. This information will be of use to researchers as well as members of the apple storage industry that are exploring alternative methods of blue mold decay control. National Program 303, component IIa.

5.Significant Activities that Support Special Target Populations
None.

6.Technology Transfer

Number of Non-Peer Reviewed Presentations and Proceedings

1

Review Publications
Janisiewicz, W.J., Saftner, R.A., Conway, W.S., Forsline, P.L. 2008. Preliminary evaluation of apple germplasm from Kazakhstan for resistance to blue mold decay caused by Penicillium expansum after harvest. HortScience.43(2):420-426.