First Report of Phytopythium litorale Causing Root Rot on Kousa Dogwood in Tennessee and the United States.

Kousa dogwood (Cornus kousa) is an economically important woody ornamental crop that exhibits creamy, white, pointed bracts in late spring, and reddish to pink drupe fruits in late summer and fall. It bears shiny dark green leaves that become reddish-purple to scarlet in the fall. In August of 2023, 3-year-old container grown C. kousa var. chinensis plants in a commercial nursery in Warren Co., Tennessee, exhibited severe yellowing, dieback and root rot symptoms (Fig. 1a and 1b). Dark brown to black lesions were observed in the root and crown region of the plants. Disease severity was 40% to 60% of root area affected, and disease incidence was approximately 40% of 1,000 plants. Surface-sterilized (10% NaOCl: 1 min) symptomatic root tissues were plated on V8-PARPH and incubated at 25°C. Sparse aerial mycelium, showing a distinct rosette or faint radiate to chrysanthemum colony pattern, was observed within four days of incubation (Fig. 2). All isolates produced ovoid or subglose, papillate, and proliferating sporangia in grass blade water cultures (Dervis et al. 2020). Sporangia measured as 19.18 to 24.80 µm X 18.08 to 22.16 µm (n = 50) with a length/width ratio of 1.06 to 1.11. Zoospores observed were between 7.07 to 9.98 µm in diameter (n = 50). Oogonia and oospores were not produced. The ribosomal internal transcribed spacer (ITS) and large subunit (LSU), as well as mitochondrial cytochrome oxidase subunit II (COX-II) genetic markers were amplified and sequenced using primer pairs ITS1/ITS4 (White et al. 1990), NL1/NL4 (Baten et al. 2014), and cox2-F/cox2-RC4 (Choi et al. 2015), respectively. The ITS, LSU, and COX-II sequences of isolates FBG6343, FBG6344 (ITS: PP458373 and PP461387; LSU: PP461390 and PP461391; COXII: PP477112 and PP477113) were 100% identical to those of MN306118, HQ643386, and MN206732, respectively. Based on the morphology (Nechwatal and Mendgen 2006) and sequence data, the isolates were identified as Phytopythium litorale (Nechw.) Abad, De Cock, Bala, Robideau, Lodhi & Lévesque. The pathogenicity test was performed on 3-year-old C. kousa var. chinensis plants grown in a 3-gal container to fulfill Koch's postulates. Kousa dogwood plants were drench inoculated (800 ml/plant) with a pathogen slurry (two plates of 7-day-old culture/liter) of isolates FBG6343 and FBG6364 (five plants per isolate). Control plants were drenched with agar slurry without the pathogen. The study was conducted in a greenhouse maintained at 21 to 23°C and 70% relative humidity with a 16-h photoperiod and irrigated twice a day for 2 min using an overhead irrigation system. Forty-five days after inoculation, plants showed dieback symptoms, and dark brown lesions developed in the roots of all inoculated plants. Isolates with morphology and sequences identical to those of FBG6343 and FBG6364 were recovered from root tissues of all inoculated plants. All control plants remained symptom-free, and P. litorale was not isolated from the root tissue. Previously, P. litorale was reported to cause disease on apple, kiwi, planatus, and rhododendron (Dervis et al. 2020; Li et al. 2021; Mert et al. 2020; Polat et al. 2023). To our knowledge, this is the first report of P. litorale causing root rot of kousa dogwood in Tennessee and the United States. Identification of this pathogen as the causal agent is crucial to developing timely management practices.

Evaluating UV-C Sensitivity of Calonectria pseudonaviculata in Model Buffer Solution Using UV-C Light-Emitting-Diode System.

Calonectria pseudonaviculata, responsible for boxwood blight, produces sticky conidia that pose a contamination risk in boxwood production, cross-contamination tools and equipment and other resources. This study evaluated UV-C LED irradiation (263-287 nm) as a disinfection method by examining its effectiveness in inactivating conidia and determining the UV-C sensitivity. Conidial suspensions were exposed to quantifiable UV-C doses under dynamic stirring condition. Average volumetric intensity was quantified by accounting for UV gradients and UV dose was calculated as a product of average fluence rate (mW/cm2) and exposure time (s). UV-C irradiation effectively inactivated the tested pathogen following log-linear + shoulder kinetics as identified by parameters of goodness of model fit (i.e. high R2 and low RMSE values). The model predicted the UV sensitivity of C. pseudonaviculata conidia as 46.6 mJ/cm2/log. A total of 2.04 log reductions of the population could be obtained by an exposure of 60 mJ.cm-2 of UV-C dose. The calculated D10 was 13.53 ± 0.98 mJ.cm-2 (R2 = 0.97, RMSE = 0.14), Kmax = 0.17 ± 0.01, and shoulder length (Sl) = 33.06 ± 1.81 mJ.cm-2. These findings indicate that UV-C irradiation could be a viable option for disinfecting tools, equipment, and possibly propagation cuttings in nurseries.

Real-Time and Rapid Detection of Phytopythium vexans Using Loop-Mediated Isothermal Amplification Assay.

Phytopythium vexans (de Bary) Abad, de Cock, Bala, Robideau, A. M. Lodhi & Levesque is an important waterborne and soil-inhabiting oomycete pathogen causing root and crown rot of various plants including certain woody ornamentals, fruit, and forest trees. Early and accurate detection of Phytopythium in the nursery production system is critical, as this pathogen is quickly transported to neighboring healthy plants through the irrigation system. Conventional methods for the detection of this pathogen are tedious, frequently inconclusive, and costly. Hence, a specific, sensitive, and rapid molecular diagnostic method is required to overcome the limitations of traditional identification. In the current study, loop-mediated isothermal amplification (LAMP) for DNA amplification was developed for the identification of P. vexans. It was evaluated using real-time and colorimetric assays. Several sets of LAMP primers were designed and screened, but PVLSU2 was found to be specific to P. vexans as it did not amplify other closely related oomycetes, fungi, and bacteria. Moreover, the developed assays were sensitive enough to amplify DNA up to 102 fg per reaction. The real-time LAMP assay was more sensitive than traditional PCR and culture-based methods to detect infected plant samples. In addition, both LAMP assays detected as few as 100 zoospores suspended in 100 ml water. These LAMP assays are anticipated to save time in P. vexans detection by disease diagnostic laboratories and research institutions and enable early preparedness in the event of disease outbreaks.

Efficacy and Timing of Application of Fungicides, Biofungicides, Host-Plant Defense Inducers, and Fertilizer to Control Phytophthora Root Rot of Flowering Dogwood in Simulated Flooding Conditions in Container Production.

Phytophthora root rot, caused by Phytophthora cinnamomi Rands, is one of the major diseases of flowering dogwood (Cornus florida L.). The severity of root rot disease increases when the plants are exposed to flooding conditions. A study was conducted to determine the efficacy and timing of application of different fungicides, biofungicides, host-plant defense inducers, and fertilizer to manage Phytophthora root rot in month-old seedlings in simulated flooding events for 1, 3, and 7 days. Preventative treatments were drench applied 3 weeks and 1 week before flooding whereas curative treatments were applied 24 h after flooding. Dogwood seedlings were inoculated with P. cinnamomi 3 days before the flooding. Plant height and width were recorded at the beginning and end of the study. At the end of the study, plant total weight and root weight were recorded and disease severity in the root was assessed using a scale of 0 to 100%. Root samples were plated using PARPH-V8 medium to determine the percent recovery of the pathogen. Empress Intrinsic, Pageant Intrinsic, Segovis, and Subdue MAXX, as preventative and curative applications, were able to suppress the disease severity compared with the inoculated control in all flooding durations. All treatments, with the exception of Stargus as a preventative application 3 weeks before flooding and Orkestra Intrinsic as a curative application, were able to suppress the disease severity compared with the inoculated control for a 1-day flooding event. Aliette and ON-Gard were effective in the first trial when applied preventatively at both 1 week and 3 weeks before flooding but not in the second trial. Signature Xtra was effective as a preventative application but not as a curative application. Interface was effective as a curative application but not as a preventative application. The findings of this study will help nursery growers to understand the performance of fungicides, biofungicides, host-plant defense inducers, and fertilizer at different time intervals and repeated applications to manage Phytophthora root rot in flooding conditions.

First Report of Fusarium poae Causing Fusarium Head Blight of Wheat in Georgia, USA.

Fusarium head blight (FHB) is one of the most troublesome fungal diseases challenging US wheat (Triticum aestivum L.) production (Savary et al. 2019). Harmful mycotoxin contamination, primarily due to deoxynivalenol (DON) in the Fusarium-damaged kernels (FDK), can negatively impact human and livestock health (McMullen et al. 1997). Although Fusarium graminearum is the primary causal agent of FHB, several other species including F. poae could pose a risk by producing dangerous mycotoxins such as nivalenol, DON, HT-2, and T-2 (Stenglein 2009). Severe FHB epidemics on wheat have occurred in recent years along with increased corn acreage across the southeast US specifically in Georgia (Ghimire et al. 2020). Five symptomatic wheat heads displaying bleaching symptoms were randomly collected from 19 different fields across 13 counties of Georgia in late spring of 2018. Infected kernels were dipped in 6% sodium hypochlorite for 10 min and rinsed three times with sterilized water. Blot dried kernels were placed on potato dextrose agar (PDA) and incubated for 7 days at 25°C under 12-h photoperiod. Three isolates (GA18W-2.1.6, GA18W-6.1.4, and GA18W-10.2.3) from Terrell, Peach, and Sumter counties exhibited dense, whitish mycelium colony typical of F. poae (Leslie and Summerell 2006). When grown in carboxymethylcellulose broth, isolates produced globose to piriform microconidia (5.1 to 12.4 µm by 4.4 to 11.2 µm) that were aseptate or had a single septation. The morphological identification was further confirmed by DNA sequencing. Single hyphal tip isolates were grown on cellophane overlain on PDA for 10 days. Fungal DNA was extracted using a Qiagen DNeasy Plant Mini Kit. Genomic DNA was sequenced using TEF1 and TEF2 primer pairs that target the translation elongation factor 1-α (EF1-α) locus (O'Donnell et al. 1998). BLASTn query of the obtained sequences of GA18W-2.1.6 (accession no. MT856907) and GA18W-10.2.3 (accession no. MT856909) were identified as F. poae with a 99% sequence homology with GenBank reference accession MK629641, while GA18W-6.1.4 (accession no. MT856908) displayed 100% similarity with F. poae accession KJ947343. Koch's postulates were performed under greenhouse conditions. Three seeds of the FHB susceptible wheat cultivar 'SS8641' were planted in individual cone-tainers with three replications (two cone-tainers/replicate). Wheat plants were vernalized for six weeks and then moved back to the greenhouse. Each F. poae isolate was spray inoculated (50,000 spores/ml) at the flowering stage onto 18-24 wheat heads. A field isolate of F. graminearum was included as a positive control whereas heads mock-inoculated with water were used as a negative control. Inoculated wheat heads were incubated in black plastic bags for 48 hours. Disease severity and FDK were recorded three weeks post inoculation. Disease severities were 6.7% (GA18W-2.1.6), 8.3% (GA18W-10.2.3), and 15.2% (GA18W-6.1.4) compared to 90.0% in the positive control similar to Arrúa et al (2019). No symptoms were observed in the negative control. FDK was 18% (GA18W-2.1.6), 28% (GA18W-10.2.3) and 44% (GA18W-6.1.4). F. poae was re-isolated from the infected heads and found to be morphologically identical to the isolates used for inoculation. To our knowledge, this is the first report of F. poae associated with FHB of wheat in the state of Georgia, USA. F. poae isolates from Georgia might produce mycotoxins in addition to reducing grain yield which needs further study.