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First Report of Rosellinia quercina Causing Postharvest Decay on ‘Conference’ Pears in the Netherlands

Wenneker, M., Pham, K. T. K., Teeuw, L., Harteveld, D. O. C.
Plant disease 2019 v.103 no.2 pp. 370
DNA, Pyrus communis, Quercus pubescens, Rosellinia necatrix, apples, asexual reproduction, avocados, cherries, cold, cold storage, cork, cultivars, culture media, ethanol, fruit crops, fruits, fungi, hyphae, internal transcribed spacers, lesions (plant), mycelium, olives, orchard soils, pathogens, peaches, pears, plastic bags, plums, postharvest diseases, rain, root rot, runoff, sequence analysis, trees, Massachusetts, Netherlands
Pear (Pyrus communis) is an important fruit crop in the Netherlands, with a total production of 374,000 metric tons in 2016. ‘Conference’ is the main pear cultivar, representing 75% of the total pear production area. In the Netherlands, pears are kept in controlled atmosphere cold storage up to 11 months after harvest. In 2017, symptoms of an unknown cause were observed in low incidence (<1%) on ‘Conference’ pears in storage from four different locations across the Netherlands. The symptoms appeared as yellow-brown circular lesions with distinct borders and a whitish center, and the lesions were slightly sunken. To isolate the causal agent, fruit were rinsed with sterile water, lesions were sprayed with 70% ethanol until runoff, the skin was removed aseptically, and tissue under the lesion was isolated and placed onto potato dextrose agar (PDA). The PDA plates were incubated at 20°C in the dark. Pure cultures were obtained by transfer of hyphal tips onto fresh PDA plates. These isolates produced fast-growing colonies with white mycelium. Cultures did not produce any specialized structures associated with sexual or asexual reproduction. However, they presented the typical pear-shaped swelling immediately above the septum of mycelia, which is characteristic of the genus Rosellinia (Castro et al. 2013). The identity of two representative isolates (KP00109 and KP00113) was confirmed by means of gene sequencing. Genomic DNA was extracted using the LGC Mag Plant Kit (Berlin) in combination with the Kingfisher method (Waltham, MA). Segments of the internal transcribed spacer region (ITS) were amplified using ITS1/ITS4 primers (White et al. 1990) and deposited in GenBank as accessions MG775691 and MG775692. MegaBLAST analysis revealed that the ITS sequences matched with 100% identity to reference Rosellinia quercina isolate ATCC36702 in GenBank (AB017661), 99% identity to Rosellinia desmazieri (AY805591), and only 89% with several isolates of Rosellinia necatrix (KF719201, AY909001, and EF592568). Subsequently, Koch’s postulates were performed on 15 detached Conference pears. Experiments were carried out on surface-sterilized fruit. One side of the fruit was wounded using a sterile cork borer before inserting a mycelial plug (5-mm diameter) of an actively growing 7-day-old culture of R. quercina on PDA. Control inoculations were performed using plugs without mycelia. The inoculated fruit were covered in plastic bags and incubated in darkness at 20°C. Symptoms appeared after 7 days on 100% of the fruits, whereas mock-inoculated controls remained symptomless. Fungal colonies isolated from the lesions on PDA morphologically resembled the original isolates. Moreover, symptoms observed on artificially inoculated fruit were identical to the decay observed on the fruit that were obtained from cold storage. The identity of the reisolations was confirmed as R. quercina by sequencing. Rosellinia is a large and complex genus (Peláez et al. 2008). Among the best-known root pathogens is the white root rot caused by R. necatrix, destructive to many fruit tree species including apple, cherry, peach, plum, pear, olive, and avocado (Pérez-Jiménez 2006; Sun et al. 2008). R. quercina is known to cause root damage in young oaks (Quercus pubescens) (Peláez et al. 2008). This is the first report of R. quercina causing storage rot of pears. Possibly, fruits got contaminated through rain splash via the orchard soil or contaminated bins at harvest. Currently, the occurrence of Rosellinia spp. in the Netherlands is largely unknown. However, owing to its ability to affect fruit crops, attention is needed.