First Report of Phytopythium vexans Causing Root Rot on Quercus bicolor in Tennessee and the United States.

Abstract

Swamp white oak (Quercus bicolor) is a North American species of medium-sized trees in the beech family. One-year-old swamp white oak seedlings grown in field conditions in a commercial nursery in Warren County, Tennessee exhibited severe root rot in July 2024. Dark brown lesions were observed in the affected roots (Fig. 1a). Disease severity was approximately 40% of root area affected, and disease incidence was approximately 20% of 100 plants. Symptomatic fine root tissues were surface sterilized with 70% ethanol and rinsed twice with distilled water. Then, symptomatic root parts (1-cm pieces) were plated on V8-PARPH (V8 juice agar amended with pimaricin, ampicillin, rifampicin, pentachloronitrobenzene, and hymexazol) and incubated at 24°C under an 8-hour photoperiod. The rosette pattern accompanied by whitish mycelium resembling Phytopythium species was consistently observed after three days of incubation. Sporangia were globose or subglobose (19.11±1.71 μm, n=50) with or without papilla (Fig. 1b). Oogonia were smooth, filamentous to globose (21.04±1.74 μm, n=50) (Fig. 1c). Representative isolates (FBG7779-1 and FBG7779-2) were identified as Phytopythium vexans based on morphological characterization (de Cock et al. 2015; Ghimire and Baysal-Gurel 2023). To confirm pathogen identity, total DNA was extracted using the DNeasy PowerLyzer Microbial Kit from 7-day-old cultures of the isolates grown on V8-PARPH. The primer pairs ITS1/ITS4 (White et al. 1990), NL1/NL4 (Baten et al. 2014), OomCoxI-Levup/Fm85mod (Robideau et al. 2011), and Cox2-F/Cox2-R (Hudspeth et al. 2000) were used to amplify and sequence the ribosomal internal transcribed spacer (ITS), the large subunit (LSU), and the mitochondrial cytochrome c oxidase subunits I (CoxI) and II (CoxII) genetic markers, respectively. The ITS, LSU, CoxI and CoxII sequences of the isolates FBG7779-1 and FBG7779-2 (ITS: PQ567140 and PQ567141; LSU: PQ567376 and PQ567377; CoxI: PQ570510 and PQ570511; CoxII: PQ570512, and PQ570513) were 100% identical to those of P. vexans isolates MK011115, OQ754108, GU133478, and AB468910, respectively. To complete Koch's postulates, pathogenicity test was performed on two-year-old swamp white oak seedlings (165 to 170 cm height) grown in 3-gal containers. The plants were drench inoculated with pathogen slurry (150 ml per plant - two 9-cm plates of 7-day-old culture/liter) of the isolates FBG7779-1 and FBG7779-2 (five plants per isolate) (Panth et al. 2021). Five plants were drenched with agar slurry without the pathogen and served as a non-inoculated control plant. The study was conducted in a greenhouse condition (21 to 23°C, 70% relative humidity) and irrigated twice a day for 2 min each time using an overhead irrigation system. Two weeks after inoculation, dark brown lesions developed in the roots of all inoculated plants (Fig. 1d), whereas controls remained healthy (Fig. 1e). The morphology of the pathogen isolated on the V8-PARPH medium was identical to the original isolate and confirmed by sequencing the ITS, LSU, CoxI and CoxII markers. Phytopythium vexans has been previously reported to cause root and crown rot in flowering cherry, ginkgo, red maple, and redbud in Tennessee (Baysal-Gurel et al. 2021; Liyanapathiranage et al. 2023; Panth et al. 2021). To our knowledge, this is the first report of P. vexans causing root rot of swamp white oak in Tennessee and the United States. Identifying the pathogen as the causal agent is crucial in the development of a successful disease management strategy of P. vexans on swamp white oak.