First Report of <i>Pythium dissotocum</i> Causing Root Rot on Nandina in Tennessee and the United States

Nandina (<i>Nandina domestica</i>) is a popular landscape plant found extensively beyond the southeastern and southwestern parts of the United States (Wilson et al. 2021). In April 2024, one-year-old nandina 'Fire Power' plants grown in containers at a commercial nursery in Warren County, Tennessee, exhibited severe leaf yellowing, stunted growth, and brown to black root rot lesions. Disease severity affected 65% of the root area, with an incidence of 80% of 450 plants. Symptomatic eight plants' root tissues were surface-sterilized with 70% ethanol and washed twice with distilled water. Small sections of root tissues were placed in Petri dishes containing V8 agar (V8A) and incubated at 24°C in an 8-hour photoperiod cycle. White, radial mycelial growth were observed within 3-4 days of incubation. The aseptate hyphae ranged from 1.81 to 7.21 μm in diameter. The filamentous sporangia formed dendroid-like branches. Oogonia were subglobose, measured 15.06 ± 0.65 μm (<i>n</i>=50) in diameter. Oospores were globose and measured 4.58 ± 0.44 μm (<i>n</i>=50). The isolates (FBG7211 and FBG7216) were identified as <i>Pythium dissotocum</i> based on morphological characterization (Van der Plaats-Niterink 1981). For molecular identification, total DNA was extracted using the DNeasy PowerLyzer Microbial Kit (Qiagen, MA) from 7-day-old cultures of the isolates FBG7211 and FBG7216 grown on V8A. The primer pairs ITS1/ITS4 (White et al. 1990), OomCoxI-Levup/Fm85mod (Robideau et al. 2011), Cox2F/Cox2RC4 (Choi et al. 2015) and Oom-Btub-up415/Oom-Btub-lo1401 (Bilodeau et al., 2007) were used to amplify and sequence isolates using four genetic markers, including the ribosomal internal transcribed spacer (ITS), the mitochondrial cytochrome c oxidase subunit I (COX1), cytochrome c oxidase subunit II (COX2), and nuclear beta-tubulin (BT), respectively. The 100% coverage of sequences (GenBank accession nos. PQ329213 and PQ284875 for ITS; PQ349316 and PQ349317 for COX1; PQ349318 and PQ349319 for COX2; PQ349314 and PQ349315 for BT) were 100%, 99.43%, 100%, and 99.56% Identical to <i>P. dissotocum</i> sequences of ITS, COX1, COX2, and BT genetic markers in the GenBank using core nt BLAST search (ITS: MT348554; COX1: MZ027313; COX2: MG719859; BT: KJ595479). Pathogenicity tests were performed on 1-year-old nandina 'Fire Power' seedlings grown in 1-gal containers to fulfill Koch's postulates. Each seedling was drench-inoculated with a 150 mL/plant of 2 x 105 zoospore suspension of the isolates FBG7211 and FBG7216 (five plants per isolate) (Fu and Kim 2024). Control plants were drenched with sterile distilled water. The plants were maintained in a greenhouse at 21 to 23°C and 70% relative humidity with a 16-h photoperiod and were irrigated twice daily for 2 min using an overhead irrigation system. Fourteen days after inoculation, dark brown necrotic lesions developed on the root of all inoculated plants, similar to those observed in the original samples. In contrast, no symptoms were observed in the control plants. Isolates resembling the morphological characteristics of <i>P. dissotocum</i> were recovered from inoculated plants, and their identity was confirmed as <i>P. dissotocum</i> using sequencing. <i>P. dissotocum</i> has been reported to cause root rot in nursery plants (Weiland et al. 2013). However, to our knowledge, this is the first report of root rot on nandina caused by <i>P. dissotocum</i> in Tennessee and the United States. This finding is crucial to develop an efficient disease management strategy of Pythium root rot on nandina.