First Report of <i>Phytopythium vexans</i> Causing Root Rot on Tulip Poplar in Tennessee and the United States

Tulip poplar (<i>Liriodendron tulipifera</i>) is a member of the Magnolia family, is a large, fast-growing, long-lived, deciduous tree native to eastern North America. One-year-old tulip poplar seedlings grown under field conditions in a commercial nursery in Warren County, Tennessee, exhibited severe root rot in May 2024. Dark brown to black lesions were observed on the affected roots. Disease severity was 60% of the affected root area, and disease incidence was 45% of 300 plants. Symptomatic root tissues were surface-disinfected with 70% ethanol and washed twice with distilled water. Small sections of root tissue were placed in Petri dishes containing V8-PARPH (V8 juice agar amended with pimaricin, ampicillin, rifampicin, pentachloronitrobenzene, and hymexazol) agar and incubated at 24°C in an 8-hour photoperiod cycle. After 3 days of incubation, whitish cottony mycelia with radiate and chrysanthemum flower-like growth patterns were observed (Supplementary Fig. 1a). The hyphae were coenocytic and 5.5 μm wide. Sporangia were subglobose with papillate, filamentous to globose smooth oogonia (18.35 to 23.86 μm; <i>n</i> = 50), bell-shaped antheridia, and spherical zoospores that are characteristic of <i>Phytopythium vexans</i> (de Cock et al. 2015) (Supplementary Fig. 1b and c). Total DNA was extracted (DNeasy PowerLyzer Microbial Kit) from 7-day-old isolates FBG7781-1 and FBG7781-2 and amplified using the ITS1/ITS4 (White et al. 1990), NL1/NL4 (Baten et al. 2014), Levup/Fm85mod (Robideau et al. 2011) and Cox2F/Cox2RC4 (Hudspeth et al. 2000) primer pairs. Isolates were sequenced using for genetic markers, including the ribosomal internal transcribed spacer (ITS), the 28S large subunit (LSU) of ribosomal RNA, the mitochondrial cytochrome c oxidase subunit I (COXI) and cytochrome c oxidase subunit II (COXII), respectively. The sequences (GenBank accession nos. PQ555199 and PQ555200 for ITS, PQ555205 and PQ555206 for LSU, PQ562067 and PQ562068 for COXI, and PQ562065 and PQ562066 for COXII) were 100% identical to the ITS, LSU, COX1, and COXII genetic markers of <i>P. vexans</i> isolates in GenBank (ITS: PQ050141, LSU: AB468723, COXI: GU133478, COXII: AB468910). Pathogenicity tests were performed on 1-year-old tulip poplar seedlings grown in 1-gal containers (3.8 L) to fulfill Koch's postulates. Tulip poplar seedlings were drenched and inoculated (150 ml/plant) with a pathogen slurry (two plates of 7-day-old culture/liter) as described in Panth et al. 2021, using isolates FBG7781-1 and FBG7781-2 (five plants per isolate). Five plants were drenched with V8-PARPH agar slurry without the pathogen and served as controls. The study was conducted in a greenhouse maintained at 21 to 23°C and 70% relative humidity and watered twice daily for 2 min using an overhead irrigation system. Fifteen days after inoculation, dark brown lesions developed in the roots of all inoculated plants (Supplementary Fig. 2a). No symptoms were observed in the control plants (Supplementary Fig. 2b). Isolates resembling the morphological characteristics of <i>P. vexans</i> were recovered from inoculated plants using the method described above, and their identity as <i>P. vexans</i> was confirmed by DNA sequencing with the same four primer pairs previously described. <i>Phytopythium vexans</i> has been reported to cause root 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 <i>P. vexans</i> causing root rot of tulip poplar in Tennessee, the United States and worldwide. This finding is significantly important for the development of a successful disease management strategy for <i>P. vexans</i> in tulip poplar.