First Report of Phytophthora lacustris Causing a Fruit Rot of Bartlett Pear in North America

Abstract

A major portion of Bartlett pears (Pyrus communis ‘Williams’) in the United States are produced in California. A fruit rot with symptoms similar to sprinkler rot caused by several Phytophthora spp. (Grove and Boal 1991) was observed in August 2017 and 2018 in an orchard in Ukiah Valley, Mendocino Co., California (N39°05′, W123°10′), where trees were irrigated by over-canopy sprinklers with water from the Russian River. Symptomatic pears were present throughout the canopy of trees, with ∼5% of fruit in the orchard affected. Symptoms initially appeared as small (5 to 10 mm in diameter), firm, light brown lesions with a diffuse margin on the exocarp. Lesions gradually expanded, covering 80 to 100% of the fruit surface, and darkened to almost black in color but remained firm. The mesocarp of infected fruit subsequently turned brown. To isolate the causal pathogen, the exocarp from 10 randomly collected infected fruit/year was removed aseptically, and 3 × 3 × 2-mm pieces of the mesocarp from the edge of lesions were placed onto a Phytophthora-selective agar medium (PARHFB-5% V8C; Hao et al. 2018). Colonies resembling a Phytophthora species consistently developed from every fruit after 5 to 7 days of incubation at 25°C in the dark. Two and four isolates obtained in 2017 and 2018, respectively, were further characterized. All isolates developed a similar petaloid mycelial growth pattern; sporangia, chlamydospores, hyphal swellings, or oospores were not observed on 5% V8C agar after 2 months. Abundant sporangia and zoospores, however, were produced in liquid culture following the protocol of Adaskaveg et al. (2015). All isolates produced nonpapillate, noncaducous sporangia from long (usually >300 μm) unbranched sporangiophores. Sporangia were mostly ovoid, varying in size (n = 60) from 38.0 to 56.0 μm (length) × 22.5 to 38.0 μm (width) (mean 43.5 × 28.6 μm) with a length/breadth ratio ranging from 1.14 to 1.83 (mean 1.52). Internal proliferation of sporangia was observed in all isolates. The sequences of ribosomal DNA internal transcribed spacers (ITS) 1 and 2 amplified with primers ITS6/ITS4 (Cooke et al. 2000) and cytochrome oxidase subunit 1 (cox 1) amplified with primers COXF4N/COXR4N (Kroon et al. 2004) of the six isolates were compared with sequences in GenBank. All were identified as Phytophthora lacustris Brasier, Cacciola, Nechw., Jung & Bakonyi based on 99% identity to the type strain P245 (Nechwatal et al. 2013; accession nos. AF266793 and JF896561 for ITS and cox 1 regions, respectively). ITS and cox 1 sequences were deposited in GenBank with accession numbers MN464235 and MN464236, and MN477337 and MN477338, for isolates 5721 and 5723, respectively. In two repeated pathogenicity tests, 30-µl drops of a zoospore suspension (1 × 10⁴ to 4 × 10⁴ zoospores/ml) of each of the six isolates of P. lacustris were pipetted onto the nonwounded surface of Bartlett pears (three drops per pear ∼15 mm apart). Fruit were placed into commercial fruit trays in enclosed plastic boxes in a randomized complete block design with two replicate fruit for each isolate and incubated at 25 ± 2°C and >95% relative humidity in the dark for 7 to 14 days. All six isolates caused brown lesions similar to those observed in the field. The pathogen was reisolated from all inoculated fruit and confirmed to be P. lacustris by ITS sequencing, completing Koch’s postulates. P. lacustris was previously isolated from necrotic roots of Alnus glutinosa, Prunus spp., Salix matsudana, and other tree species growing in flooded conditions (Nechwatal et al. 2013). It was also reported to cause a fruit rot of Bosc pear in Argentina (Sosa et al. 2015). This is the first report of P. lacustris causing a fruit rot of Bartlett pear in California. Because P. lacustris may be an opportunistic pathogen in wet habitats (Nechwatal et al. 2013) and has also been isolated from river water (Brazee et al. 2016), proper irrigation methods using pathogen-free water may effectively manage this disease.