First Report of Diaporthe sp. from the D. arctii Species Complex Causing Postharvest Decay of European Pear in West Virginia, United States

In November 2022, a European pear (Pyrus communis L.) ‘Shenandoah’ presenting brown discoloration and softening of the tissue over 75% of the fruit surface was found in cold storage at the USDA Appalachian Fruit Research Station in Kearneysville, West Virginia. Two of the 24 ‘Shenandoah’ pears displayed the disease symptoms described. Following surface sanitation with 70% ethanol, tissue was taken at the margin of the lesion area, transferred to potato dextrose agar (PDA), and incubated at 25°C under continuous light. The isolate was hyphal-tip purified and propagated on PDA at 25°C. A growth assay using 7 mm plugs on PDA showed average daily growth of 14.5 mm over 7 days. Colonies appeared cream to light tan darkening with age. Elliptical, guttiferous, and aseptate α-conidia with mean dimensions of 8.74 µm (± 0.13 µm) by 3.7 µm (± 0.05 µm), n = 60, were observed on oatmeal agar after 25 days. β-conidia were filiform with mean dimensions of 24.4 µm (± 0.42 µm) by 1.34 µm (± 0.03 µm), n = 60. Fungal DNA was extracted using a CTAB protocol from one isolate (WV22SR1P5), and five genomic loci were amplified: internal transcribed spacer (ITS), translation elongation factor-1 alpha (TEF1), beta-tubulin (TUB), histone H3 (HIS), and calmodulin (CAL) (Udayanga et al. 2014) (GenBank accession nos. OR504473, OR504505, OR504506, PP213454, and PP213453 respectively). Additionally, part of the MAT1-1-1 gene was amplified (GenBank PP806137), but the MAT1-2-1 gene could not be detected (Santos et al. 2010). Based on a maximum-likelihood phylogenetic tree of concatenated genes (ITS-TEF1-TUB-HIS-CAL) from published isolates (Dissanayake et al. 2024; Gomes et al. 2013; Gomzhina et al. 2021; Moodispaw et al. 2023; Udayanga et al. 2015), WV22SR1P5 was most closely related to a Diaporthe sp. in the D. arctii species complex, Section Sojae, reported to cause disease on cucumber that has not yet been given a Latin binomial (Moodispaw et al. 2023). The WV22SR1P5 isolate was deposited in the USDA-ARS Culture Collection (NRRL# 64834). Organic European pears ‘Bartlett’ were surface sanitized with 70% ethanol and dried in a laminar flow hood. Fruits were wounded with a cork borer (4 mm diameter x 4 mm depth) and inoculated with a 4-mm mycelial plug of 7- to 10-day-old culture of WV22SR1P5 grown on PDA. Inoculated fruit were placed in fruit trays in plastic bins and stored at 25°C for 7 days in the dark. As a control, fruits were wounded and sterile PDA plugs were placed in wounds under the same conditions. Five fruits were used per treatment group, and the entire test was repeated for a total of two replications. Lesion development was observed within 72 hours and expanded to an average of 44 mm by day 7. Lesions were not observed on control fruit. Lesion appearance on inoculated pears matched the decay symptoms on the original pear selected from cold storage. Fungal isolates obtained from inoculated pears were morphologically and molecularly identified as the same Diaporthe sp. by sequencing of TEF1 and TUB loci. The related species D. eres and D. rudis were recently reported to cause fruit rots on European pear in northern Italy (Bertetti et al. 2018) and in Oregon, United States (KC et al. 2019), respectively. D. sojae isolates have been associated with pear shoot canker in Asian pear trees (Pryus pyrifolia) in China (Guo et al. 2020). To our knowledge, this is the first report of a Diaporthe sp. in the D. arctii species complex causing postharvest decay of European pear in the United States and West Virginia, specifically.