Characterization and Aggressiveness of Take-All Root Rot Pathogens Isolated from Symptomatic Bermudagrass Putting Greens.

Take-all root rot is a disease of ultradwarf bermudagrass putting greens caused by Gaeumannomyces graminis (Gg), Gaeumannomyces sp. (Gx), Gaeumannomyces graminicola (Ggram), Candidacolonium cynodontis (Cc), and Magnaporthiopsis cynodontis (Mc). Many etiological and epidemiological components of this disease remain unknown. Improving pathogen identification and our understanding of the aggressiveness of these pathogens along with growth at different temperatures will advance our knowledge of disease development to optimize management strategies. Take-all root rot pathogens were isolated from symptomatic bermudagrass root and stolon pieces from 16 different golf courses. Isolates of Gg, Gx, Ggram, Cc, and Mc were used to inoculate 'Champion' bermudagrass in an in planta aggressiveness assay. Each pathogen was also evaluated at 10, 15, 20, 25, 30, and 35°C to determine growth temperature optima. Infected plant tissue was used to develop a real-time PCR high-resolution melt assay for pathogen detection. This assay was able to differentiate each pathogen directly from infected plant tissue using a single primer pair. In general, Ggram, Gg, and Gx were the most aggressive while Cc and Mc exhibited moderate aggressiveness. Pathogens were more aggressive when incubated at 30°C compared with 20°C. While they grew optimally between 24.4 and 27.8°C, pathogens exhibited limited growth at 35°C and no growth at 10°C. These data provide important information on this disease and its causal agents that may improve take-all root rot management.

Pythium spp. Associated with Root Rot and Stunting of Winter Crops in North Carolina.

Annual double-crop rotation systems that incorporate winter wheat, clary sage, or a cover crop are common in eastern North Carolina. Stunting and root rot of clary sage (Salvia sclarea L.) reduce yields of this crop, especially in wet soils. Stunting and reduced stand establishment also afflict winter cover crops, including rye, rapeseed, and winter pea. Pythium spp. are causal agents of root rot of winter wheat in this region, but their role in root rot and stunting of other winter crops is not understood. During the growing seasons of 2018 to 2019 and 2019 to 2020, samples of clary sage, rye, rapeseed, and winter pea displaying symptoms of stunting were collected across eastern North Carolina, resulting in the recovery of 420 isolates of Pythium from the roots of all hosts. Pythium irregulare, Pythium spinosum, and the complex Pythium sp. cluster B2A were the species most frequently isolated from clary sage. P. irregulare and P. spinosum were aggressive pathogens of clary sage at 18°C and caused moderate root rot at 28°C. Koch's postulates confirmed that isolates belonging to Pythium sp. cluster B2A, Pythium sylvaticum, Pythium pachycaule, Pythium aphanidermatum, Pythium myriotylum, and Pythium oopapillum are pathogens of clary sage. P. irregulare (37% of all isolates) and members of the species complex Pythium sp. cluster B2A (28% of all isolates) constituted the majority of isolates collected from all hosts and were the species most frequently isolated from rye, rapeseed, and winter pea. In pathogenicity assays, isolates representing P. irregulare and P. spinosum caused slight to moderate root necrosis on rye, rapeseed, and winter pea. Isolates representing Pythium sp. cluster B2A caused slight to moderate root necrosis on rapeseed and clary sage, but no symptoms on rye or winter pea.

Influence of post-application irrigation and mowing timing on fungicide fate on a United States Golf Association golf course putting green.

Fungicides are routinely applied to golf course putting greens throughout the growing season. Gaining a better understanding of fungicide fate can improve fungicide use and stewardship. Therefore, optimizing fungicide applications with post-application management practices may enhance fungicide movement and limit potential off-target effects. Two field studies were initiated on a golf course putting green to evaluate the influence of post-fungicide application irrigation and mowing timing on fungicide movement into the soil profile and removal in turfgrass clippings. Plots were treated with a single application of either pyraclostrobin, triadimefon, or penthiopyrad and received 0.64 cm post-application irrigation immediately or 6 h after application or received no post-application irrigation. Clippings were collected 0, 1, and 3 d after treatment (DAT). Cores were harvested 0, 1, 3, 5, 7, and 14 DAT and dissected into the remaining aboveground vegetation (RAV; verdure/thatch; 0-to-2.5-, 2.5-to-5.1-, and 5.1-to-7.6-cm soil subsections). A small amount of fungicide (<3.6%) was removed with clippings regardless of mowing and irrigation treatment. Post-application irrigation treatment influenced fungicide movement; however, >50% of fungicide remained restricted to the RAV for the first 3 DAT. Less fungicide remained restricted to the RAV, and more fungicide was detected in deeper soil depths when plots were irrigated immediately after application. Fungicide was only detected at the 5.1-to-7.6-cm depth when plots were irrigated immediately. Applying post-application irrigation immediately may result in more fungicide moving down to soilborne targets. Irrigating 6 h after application facilitated moderate fungicide movement compared with irrigating immediately but was better than no post-application irrigation.

Genome Resources for Seven Fungal Isolates That Cause Dollar Spot Disease in Turfgrass, Including Clarireedia jacksonii and C. monteithiana.

Fungi in the genus Clarireedia are widespread and destructive pathogens of grasses worldwide, and are best known as the causal agents of dollar spot disease in turfgrass. Here, we report genome assemblies of seven Clarireedia isolates, including ex-types of the two most widespread species, Clarireedia jacksonii and C. monteithiana. These datasets provide a valuable resource for ongoing studies of the dollar spot pathogens that include population diversity, host-pathogen interactions, marker development, and disease control.

Pythium spp. Associated with Root Rot and Stunting of Winter Wheat in North Carolina.

In eastern North Carolina, mild to severe stunting and root rot have reduced yields of winter wheat, especially during years with abundant rainfall. Causal agents of root rot of wheat in this region were previously identified as Pythium irregulare, P. vanterpoolii, and P. spinosum. To investigate species prevalence, 114 isolates of Pythium were obtained from symptomatic wheat plants collected in eight counties. Twelve species were recovered, with P. irregulare (32%), P. vanterpoolii (17%), and P. spinosum (16%) the most common. Pathogenicity screens were performed with selected isolates of each species, and slight to severe necrosis of young roots was observed. The aggressiveness of five isolates each of P. irregulare, P. vanterpoolii, and P. spinosum was compared on a single cultivar of wheat at 14°C, and very aggressive isolates were found within all species. In vitro growth of these isolates was measured at 14 and 20°C, and all isolates grew faster at the warmer temperature. The effects of varying temperatures and rates of nitrogen on root rot caused by Pythium spp. alone or in combination were investigated. All inoculation treatments caused severe root rot under all conditions tested, and disease was more severe at 12 and 14°C compared with 18 and 20°C; however, there was no effect of nitrogen application.

Evaluating Fungicide Selections to Manage Pythium Root Rot on Poinsettia Cultivars with Varying Levels of Partial Resistance.

Pythium aphanidermatum is the predominant species causing Pythium root rot of commercially grown poinsettia (Euphorbia pulcherrima Willd. ex Kotzch) in North Carolina. Pythium root rot is managed primarily through a combination of sanitation practices and preventative fungicide applications of mefenoxam or etridiazole. Insensitivity to mefenoxam is common but growers continue to rely on it from lack of inexpensive and efficacious alternatives. This research was conducted to identify alternative fungicides for Pythium root rot control and to evaluate their efficacy on poinsettia cultivars with varying levels of partial resistance. Greenhouse studies were conducted to assess efficacy of fungicide treatments in seven poinsettia cultivars inoculated with a mefenoxam-sensitive isolate of P. aphanidermatum. One study examined control with a single fungicide drench made at transplant and a second study examined repeat fungicide applications made throughout the experiment. Treatments containing etridiazole, mefenoxam, fenamidone, and cyazofamid provided control of Pythium root rot across all cultivars in both experiments whereas Fosetyl-al, potassium phosphite, and Trichoderma spp. failed to offer satisfactory control. Azoxystrobin, pyraclostrobin, and propamocarb reduced disease on some cultivars but failed to control Pythium root rot on highly susceptible cultivars. Four isolates of P. aphanidermatum cultured from plants growing in commercial greenhouses were evaluated for in vitro sensitivity to fungicides labeled for Pythium root rot control at four rates. Etridiazole, fosetyl-al, and potassium phosphite completely inhibited mycelial growth, whereas isolates varied in response to mefenoxam, cyazofamid, propamocarb, fenamidone, azoxystrobin, and pyraclostrobin in vitro. Twenty-one additional isolates then were evaluated at label rates of these fungicides. Seven isolates were insensitive to label rates of all three quinone outside inhibitors and one isolate was insensitive to the quinone outside inhibitors and mefenoxam. These results provide guidelines for selecting fungicides to maximize control of Pythium root rot on poinsettia cultivars.

Oxalic Acid Production in Clarireedia jacksonii Is Dictated by pH, Host Tissue, and Xylan.

Dollar spot is caused by the fungus Clarireedia jacksonii and is the most common disease of golf course turfgrass in temperate climates. Oxalic acid (OA) is an important pathogenicity factor in other fungal plant pathogens, such as the dicot pathogen Sclerotinia sclerotiorum, but its role in C. jacksonii pathogenicity on monocot hosts remains unclear. Herein, we assess fungal growth, OA concentration, and pH change in potato dextrose broth (PDB) following incubation of C. jacksonii. In addition, OA production by C. jacksonii and S. sclerotiorum was compared in PDB amended with creeping bentgrass or common plant cell wall components (cellulose, lignin, pectin, or xylan). Our results show that OA production is highly dependent on the environmental pH, with twice as much OA produced at pH 7 than pH 4 and a corresponding decrease in PDB pH from 7 to 5 following 96 h of C. jacksonii incubation. In contrast, no OA was produced or changes in pH observed when C. jacksonii was incubated in PDB at a pH of 4. Interestingly, C. jacksonii increased OA production in response to PDB amended with creeping bentgrass tissue and the cell wall component xylan, a major component of grass cell walls. S. sclerotiorum produced large amounts of OA relative to C. jacksonii regardless of treatment, and no treatment increased OA production by this fungus, though pectin suppressed S. sclerotiorum's OA production. These results suggest that OA production by C. jacksonii is reliant on host specific components within the infection court, as well as the ambient pH of the foliar environment during its pathogenic development.

Identification and Pathogenicity of Bacteria Associated with Etiolation and Decline of Creeping Bentgrass Golf Course Putting Greens.

Bacterial etiolation and decline has developed into a widespread issue with creeping bentgrass (CBG) (Agrostis stolonifera) putting green turf. The condition is characterized by an abnormal elongation of turfgrass stems and leaves that in rare cases progresses into a rapid and widespread necrosis and decline. Recent reports have cited bacteria, Acidovorax avenae and Xanthomonas translucens, as causal agents; however, few cases exist where either bacterium were isolated in conjunction with turf exhibiting bacterial disease symptoms. From 2010 to 2014, turfgrass from 62 locations submitted to the NC State Turf Diagnostic Clinic exhibiting bacterial etiolation and/or decline symptoms were sampled for the presence of bacterial pathogens. Isolated bacteria were identified using rRNA sequencing of the 16S subunit and internal transcribed spacer region (16S-23S or ITS). Results showed diverse bacteria isolated from symptomatic turf and A. avenae and X. translucens were only isolated in 26% of samples. Frequently isolated bacterial species were examined for pathogenicity to 4-week-old 'G2' CBG seedlings and 8-week-old 'A-1' CBG turfgrass stands in the greenhouse. While results confirmed pathogenicity of A. avenae and X. translucens, Pantoea ananatis was also shown to infect CBG turf; although pathogenicity varied among isolated strains. These results illustrate that multiple bacteria are associated with bacterial disease and shed new light on culturable bacteria living in CBG turfgrass putting greens. Future research to evaluate additional microorganisms (i.e., bacteria and fungi) could provide new information on host-microbe interactions and possibly develop ideas for management tactics to reduce turfgrass pests.

Brachypodium: A Potential Model Host for Fungal Pathogens of Turfgrasses.

Brachypodium distachyon is a C3 grass that is an attractive model host system for studying pathogenicity of major turfgrass pathogens due to its genetic similarity to many cool-season turfgrasses. Infection assays with two or more isolates of the casual agents of dollar spot, brown patch, and Microdochium patch resulted in compatible interactions with B. distachyon inbred line Bd21-3. The symptoms produced by these pathogens on Bd21-3 closely resembled those observed on the natural turfgrass host (creeping bentgrass), demonstrating that B. distachyon is susceptible to the fungal pathogens that cause dollar spot, brown patch, and Microdochium patch on turfgrasses. The interaction between Sclerotinia homoeocarpa isolates and Brachypodium ecotypes was also investigated. Interestingly, differential responses of these ecotypes to S. homoeocarpa isolates was found, particularly when comparing B. distachyon to B. hybridum ecotypes. Taken together, these findings demonstrate that B. distachyon can be used as a model host system for these turfgrass diseases and leveraged for studies of molecular mechanisms contributing to host resistance.

Plant Growth Regulator Effects on Bacterial Etiolation of Creeping Bentgrass Putting Green Turf Caused by Acidovorax avenae.

Bacterial etiolation, caused by Acidovorax avenae, is a widespread problem in creeping bentgrass putting green turf. The symptoms normally appear as abnormally elongated turfgrass stems and leaves. Observations at multiple field sites suggest the involvement of plant growth regulators (i.e., GA-biosynthesis inhibitors) commonly applied to turf, alluding to a phytohormone imbalance caused by the bacterium. A 2-year field study examined the effects of trinexapac-ethyl, flurprimidol, and paclobutrazol on bacterial etiolation severity caused by A. avenae. Trinexapac-ethyl applied at 0.05 kg a.i. ha-1 every 7 days and 0.10 kg ha-1 every 14 days increased etiolation compared with all other treatments in both years. Flurprimidol and paclobutrazol were not different from the control but high-rate applications caused phytotoxicity that lowered turf quality early in 2014. When the etiolated turfgrass was removed with mowing, turfgrass treated with trinexapac-ethyl exhibited the highest turfgrass quality on most rating dates. Results from this work illustrate that using plant growth regulator materials with different modes of action is a solution to managing creeping bentgrass growth while limiting the potential for bacterial etiolation outbreaks.