Methods for Rapid Identification and Functional Analysis of Fungi Causing Postharvest Decay of Pome Fruit

Objective 1: Isolate and identify fungi causing postharvest decay of pome fruit from packinghouses that use different pest management programs and determine their levels of resistance to postharvest fungicides. [NP 303; C1, PS 1A] Sub-objective 1A: Determine the tolerance of Penicillium spp., causing blue mold of apples from conventional and organic packinghouses, to postharvest fungicides. Sub-objective 1B: Determine cultural, morphological, and molecular markers that distinguish species and strains of Penicillium that differ in virulence and their sensitivity to selected fungicides. Sub-objective 1C: Design a real-time PCR method for rapid and accurate detection of Penicillium spp. causing blue mold of apple. Objective 2: Design and construct a genetic transformation system for functional analysis of genes in fungi that cause postharvest decay of pome fruit. [NP 303; C2, PS 2A] Sub-objective 2A: Construct binary vectors containing selectable antibiotic resistance gene(s), and transform into Penicillium expansum using Agrobacterium tumefaciens. Sub-objective 2B: Determine the mitotic stability and cultural morphology of Penicillium expansum transformants carrying single and multiple antibiotic resistance genes in vitro. Sub-objective 2C: Assess virulence and morphology in apple fruit of Penicillium expansum transformant strains harboring selectable antibiotic resistance genes. Objective 3: Identify and characterize genes encoding components of pathways that effect pathogen growth or virulence in fungi causing postharvest decay of pome fruit. [NP 303; C2, PS 2A] Sub-objective 3A: Identify signaling pathways and characterize genes that potentially affect conidial germination in Penicillium expansum causing blue mold. Sub-objective 3B: Analyze the function of candidate genes involved in pathways that affect Penicillium expansum growth or virulence.