Prevention and Detection of Fungicide Resistance Development in Rhizoctonia zeae from Soybean and Corn in Nebraska

The goal of this research was to advance the foundational knowledge required to quantify and mitigate fungicide resistance in Rhizoctonia zeae, the seedling disease pathogen of soybean and corn. In vitro sensitivity to azoxystrobin, fludioxonil, sedaxane, and/or prothioconazole was determined for 91 R. zeae isolates obtained mostly from soybean and corn fields in Nebraska. Isolates were sensitive to fludioxonil, sedaxane, and prothioconazole (EC50 < 3 µg/ml) and had a positively skewed EC50 distribution. Isolates were not sensitive to azoxystrobin in vitro (EC50 > 100 µg/ml) or in planta. Application of azoxystrobin did not significantly decrease disease severity or improve total dry weight of the soybean plants (P > 0.05). The risk of resistance development in R. zeae was estimated by characterizing its population structure. Eighty-one R. zeae isolates were genotyped using six microsatellite markers. Results showed that the population has a mixed mode of reproduction and is structured according to geographic region, suggesting limited dispersal. These population characteristics suggest that R. zeae has an intermediate risk of resistance development. Overall, this research established the current status of fungicide sensitivity in R. zeae in Nebraska and estimated its risk of resistance development, which can inform fungicide resistance management for R. zeae.

Illumina sequencing of 18S/16S rRNA reveals microbial community composition, diversity, and potential pathogens in turfgrass seeds

The increasing need for turfgrass seeds is coupled with the high risk of dangerous microbial pathogens being transmitted through the domestic and international trade of seeds. Concerns continue to be raised about seed safety and quality. Here, we show that the next-generation sequencing (NGS) of DNA represents an effective and reliable tactic to monitor the microbial communities within turfgrass seeds. A comparison of DNA sequence data with reference databases revealed the presence of 26 different fungal orders. Among them, serious plant disease pathogens such as Bipolaris sorokiniana, Boeremia exigua, Claviceps purpurea, and Rhizoctonia zeae were detected. Seedborne bacteria, including Erwinia persicina and Acidovorax avenae, were identified from different bacterial orders. Our study indicated that the traditional culturing method and the NGS approach are complementary to each other for pathogen identification. The reliability of culturing and NGS methods was further validated by polymerase chain reaction (PCR) with specific primers. The combination of these different techniques ensures maximum sensitivity and specificity for turfgrass seed pathogen testing assay.

Screening Tall Fescue for Resistance to Rhizoctonia solani and Rhizoctonia zeae Using Digital Image Analysis

Brown patch, caused by Rhizoctonia solani, is a destructive disease on tall fescue. Compared with R. solani, Rhizoctonia zeae causes indistinguishable symptoms in the field but varies in geographic distribution. This may contribute to geographic variability observed in the resistance response of improved brown patch–resistant cultivars. This study examined R. solani and R. zeae susceptibility of four cultivars, selected based on brown patch performance in the National Turfgrass Evaluation Program (NTEP), and nine plant introductions (PIs). Twenty genotypes per PI/cultivar were evaluated by using four clonal replicates in a randomized complete block design. Plants were inoculated under controlled conditions with two repetitions per pathogen. Disease severity was assessed digitally in APS Assess, and analysis of variance and correlations were performed in SAS 9.3. Mean disease severity was higher for R. solani (65%) than for R. zeae (49%) (P = 0.0137). Interaction effects with pathogen were not significant for PI (P = 0.0562) but were for genotype (P < 0.001). Moderately to highly resistant NTEP cultivars compared with remaining PIs exhibited lower susceptibility to R. zeae (P < 0.0001) but did not differ in susceptibility to R. solani (P = 0.7458). Correlations between R. solani and R. zeae disease severity were not significant for either PI (R = 0.06, P = 0.8436) or genotype (R = 0.11, P = 0.09). Breeding for resistance to both pathogens could contribute to a more geographically stable resistance response. Genotypes were identified with improved resistance to R. solani (40), R. zeae (122), and both pathogens (26).

Evaluation of Soil-Applied Chemical Fungicide and Biofungicide for Control of the Fusarium Wilt of Chrysanthemum and Their Effects on Rhizosphere Soil Microbiota

Chemical fungicides are a frequently used intervention for the control of the Fusarium wilt of chrysanthemum, but are no longer considered environmentally friendly. However, the biofungicides offer one of the best alternatives to reduce the use of chemical fungicides. In this study, a series of two-year greenhouse experiments were conducted to evaluate the soil-applied chemical fungicide (dazomet, DZ) and biofungicide (biocontrol agent combined with B. subtilis NCD-2, BF) for controlling the Fusarium wilt of chrysanthemum and its effects on rhizosphere soil microbiota. The results indicated that DZ and BF showed good control efficacy of Fusarium wilt of chrysanthemum in the two-year application evaluation. However, the DZ treatment significantly decreased the soil catalase and urease activities compared with the control, while BF showed a significant increase in bacterium/fungus ratios (B/F), soil urease and acid phosphatase activities. Abundances of potential plant pathogens F. oxysporum, Rhizoctonia zeae and Rhizoctonia solani were also lower, while potential plant-growth-promoting micro-organisms like the Rhizobiales bacterium and Mariniflexile sp. were higher in the BF treatment than in the control. Our findings suggested that the overall effect of the soil biota on chrysanthemum growth was more positive and stronger in the BF treatment than in the DZ treatment.

Diversity and Pathogenicity of Rhizoctonia Species from the Brazilian Cerrado

Eighty-one Rhizoctonia-like isolates were identified based on morphology and nuclei-staining methods from natural and agricultural soils of the Cerrado (Brazilian savanna). The nucleotide similarity analysis of ITS1-5.8S-ITS2 regions identified 14 different taxa, with 39.5% of isolates assigned to Waitea circinata (zeae, oryzae, and circinata varieties), while 37.0% belonged to Thanatephorus cucumeris anastomosis groups (AGs) AG1-IB, AG1-ID, AG1-IE, AG4-HGI, and AG4-HGIII. Ceratobasidium spp. AG-A, AG-F, AG-Fa, AG-P, and AG-R comprised 23.5%. Rhizoctonia zeae (19.8%), R. solani AG1-IE (18.6%), and binucleate Rhizoctonia AG-A (8.6%) were the most frequent anamorphic states found. Root rot severity caused by the different taxa varied from low to high on common beans, and tended to be low to average in maize. Twenty-two isolates were pathogenic to both hosts, suggesting difficulties in managing Rhizoctonia root rots with crop rotation. These results suggest that cropping history affects the geographical arrangement of AGs, with a prevalence of AG1 in the tropical zone from central to north Brazil while the AG4 group was most prevalent from central to subtropical south. W. circinata var. zeae was predominant in soils under maize production. To our knowledge, this is the first report on the occurrence of W. circinata var. circinata in Brazil.

Waitea circinata. [Distribution map].

A new distribution map is provided for Waitea circinata [Rhizoctonia zeae] (Warcup) & Talbot. Fungi: Basidiomycota: Corticiales. Hosts: creeping bentgrass (Agrostis stolonifera), bluegrass (Poa spp.), rice (Oryza sativa), maize (Zea mays), wheat (Triticum spp.). Information is given on the geographical distribution in Europe (Hungary), Asia (Bangladesh, Cambodia, China, Hubei, Sichuan, India, Andhra Pradesh, Tamil Nadu, Iran, Japan, Honshu, Kyushu, Shikoku, Korea Republic, Malaysia, Sarawak, Myanmar, Nepal, Taiwan, Thailand, Turkey, Vietnam), Africa (Malawi, South Africa), North America (Canada, British Columbia, Ontario, USA, Alaska, Arizona, Arkansas, California, Colorado, Connecticut, Florida, Georgia, Idaho, Illinois, Indiana, Kentucky, Louisiana, Massachusetts, Minnesota, Mississippi, Nevada, New Jersey, New York, North Carolina, North Dakota, Ohio, Oregon, Pennsylvania, Rhode Island, South Carolina, Texas, Virginia, Washington, West Virginia, Wisconsin), Central America & Caribbean (Cuba, El Salvador, Panama), South America (Brazil, Para, Tocantins, Colombia, Venezuela), Oceania (Australia, New South Wales, Queensland, Western Australia, Fiji, New Zealand).

Antimicrobial Activity of Pyrrocidines from Acremonium zeae Against Endophytes and Pathogens of Maize

Acremonium zeae produces pyrrocidines A and B, which are polyketide-amino acid-derived antibiotics, and is recognized as a seedborne protective endophyte of maize which augments host defenses against microbial pathogens causing seedling blights and stalk rots. Pyrrocidine A displayed significant in vitro activity against Aspergillus flavus and Fusarium verticillioides in assays performed using conidia as inoculum, with pyrrocidine A being more active than B. In equivalent assays performed with conidia or hyphal cells as inoculum, pyrrocidine A revealed potent activity against major stalk and ear rot pathogens of maize, including F. graminearum, Nigrospora oryzae, Stenocarpella (Diplodia) maydis, and Rhizoctonia zeae. Pyrrocidine A displayed significant activity against seed-rotting saprophytes A. flavus and Eupenicillium ochrosalmoneum, as well as seed-infecting colonists of the phylloplane Alternaria alternata, Cladosporium cladosporioides, and Curvularia lunata, which produces a damaging leaf spot disease. Protective endophytes, including mycoparasites which grow asymptomatically within healthy maize tissues, show little sensitivity to pyrrocidines. Pyrrocidine A also exhibited potent activity against Clavibacter michiganense subsp. nebraskense, causal agent of Goss's bacterial wilt of maize, and Bacillus mojaviense and Pseudomonas fluorescens, maize endophytes applied as biocontrol agents, but were ineffective against the wilt-producing bacterium Pantoea stewartii.

Discovery and Characterization of Waitea circinata var. circinata Affecting Annual Bluegrass from the Western United States

Waitea circinata var. circinata was identified as the causal agent of a new disease of annual bluegrass (Poa annua) in the United States. This pathogen is also known to cause brown ring patch on creeping bentgrass (Agrostis stolonifera) in Japan, but it had not been reported on any turf species outside of Japan. Symptoms on annual bluegrass caused by this fungus included regular to irregular yellow rings several centimeters to 1 m in diameter, typically at maximum daytime temperatures of 15 to 35°C. A total of 26 isolates were collected from diseased annual bluegrass. Twenty-two of these isolates were multinucleate, grew optimally at 25 to 30°C, and in culture formed irregular sclerotia approximately 2 to 5 mm in size that were white to orange and remained orange or turned brown to dark brown over a 28-day period. The remaining four isolates were characterized as being W. circinata var. zeae (Rhizoctonia zeae), which is a known pathogen of annual bluegrass in the United States. Sequence analysis of the internal transcribed spacer (ITS) regions of rDNA on a subset of isolates confirmed the identifications of W. circinata var. circinata (n = 8) and W. circinata var. zeae (n = 1) based on deposited sequences in GenBank. The identity of the remaining 14 isolates suspected to be W. circinata var. circinata was confirmed by HapII digestion of the amplified rDNA ITS region. Pathogenicity of four W. circinata var. circinata isolates was confirmed on both annual bluegrass and creeping bentgrass. This study is the first morphological and molecular confirmation of the presence of W. circinata var. circinata as a pathogen of turfgrass in the United States.