First Report of Colletotrichum fioriniae Causing Postharvest Decay on 'Nittany' Apple Fruit in the United States.

Abstract

Bitter rot of apple is caused by Colletotrichum acutatum and C. gleosporioides and is an economically important disease in the mid-Atlantic and southern regions of the United States (1). However, other Colletotrichum spp. have been found to infect apple and pear fruit in Croatia that include C. fioriniae and C. clavatum (3). The disease is favorable under wet, humid conditions and can occur in the field or during storage causing postharvest decay (2). In February 2013, 'Nittany' apples with round, brown, dry, firm lesions having acervuli in concentric rings were observed at a commercial cold storage facility in Pennsylvania. Samples were placed on a paper tray in an 80-count apple box and immediately transported to the lab. Fruit were rinsed with sterile water, and lesions were sprayed with 70% ethanol until runoff. The skin was aseptically removed with a scalpel, and tissue under the lesion was placed onto potato dextrose agar (PDA) petri dishes. Dishes were incubated at 25°C with constant light, and a single-spore isolate was propagated on PDA. Permanent cultures were maintained as PDA slants stored at 4°C in darkness. The isolate was identified as a Colletotrichum sp. based on culture morphology, having light gray mycelium with a pinkish reverse and abundant pin-shaped melanized acervuli oozing pink conidia on PDA. Conidia were fusiform, pointed at one or both ends, one-celled, thin-walled, aseptate, hyaline, and averaged 10.5 μm (7.5 to 20 μm) long and 5.1 μm (5 to 10 μm) wide (n = 50). Genomic DNA was extracted from mycelia and amplified using conventional PCR and gene specific primers for 313 bp of the Histone 3 gene and with ITS4/5 primers for the internal transcribed spacer (ITS) rDNA region. MegaBLAST analysis of both gene sequences showed that our isolate was identical to other Colletotrichum fioriniae sequences in GenBank and was 100% identical to culture-collection C. fioriniae isolate CBS:128517, thus confirming the morphological identification. To prove pathogenicity, Koch's postulates were conducted using organic 'Gala' apple fruit that were washed with soap and water, sprayed with 70% ethanol, and wiped dry. The fruit were wounded with a sterile nail to a 3-mm depth, inoculated with 50 μl of a conidial suspension (1 × 104 conidia/ml), and stored at 25°C in 80-count boxes on paper trays for 14 days. Lesion diameter was measured from 10 replicate fruit with a digital micrometer and averaged 31.2 mm (±2.5 mm) over two experiments (n = 20). Water-only controls were symptomless. Artificially inoculated 'Gala' apples had identical external and internal symptoms (v-shaped decay pattern when the fruit were cut in half) to those observed on 'Nittany' apples that were originally obtained from cold storage. Bitter rot caused by C. fioriniae may become an emerging problem for the pome fruit growing industry in the near future, and may require investigation of new disease management practices to control this fungus. This is the first report of postharvest decay caused by C. fioriniae on apple fruit from cold storage in the United States.