RESEARCHFree Access icon

Yield Losses and Control by Sedaxane and Fludioxonil of Soilborne Rhizoctonia, Microdochium, and Fusarium Species in Winter Wheat

    Affiliations
    Authors and Affiliations
    • Matthew Brown1
    • Dasuni P. Jayaweera1
    • Annabel Hunt1
    • James W. Woodhall2
    • Rumiana V. Ray1
    1. 1Division of Plant and Crop Sciences, School of Biosciences, University of Nottingham, Sutton Bonington Campus, Loughborough, Leicestershire, LE12 5RD, UK
    2. 2University of Idaho, Parma Research and Extension Center, Moscow, ID, U.S.A.

    Published Online:https://doi.org/10.1094/PDIS-11-20-2401-RE

    Abstract

    Soilborne Rhizoctonia, Microdochium, and Fusarium species are major causal agents of seedling and stem-base diseases of wheat. Currently, seed treatments are considered the most effective solution for their control. Rhizoctonia solani anastomosis groups (AGs) 2-1 and 5, R. cerealis, Microdochium, and Fusarium spp., were used in series of field experiments to determine their capability to cause soilborne and stem-base disease and to quantify their comparative losses in the establishment and yield of wheat. The effectiveness and response to seed treatment formulated with 10 g sedaxane and 5 g fludioxonil 100 kg−1 against these soilborne pathogens were also determined. Our results showed that damping-off caused by soilborne R. cerealis was associated with significant reductions in the emergence and establishment, resulting in stunted growth and low plant numbers. The pathogen also caused sharp eyespot associated with reductions in the ear partitioning index. R. solani AG 2-1 and AG 5 were weakly pathogenic and failed to cause significant damping-off, root rot, and stem-base disease in wheat. Fusarium graminearum and F. culmorum applied as soilborne inoculum failed to cause severe disease. Microdochium spp. caused brown foot rot disease and soilborne M. nivale reduced wheat emergence. Applications of sedaxane and fludioxonil increased plant emergence and reduced damping-off, early stem-base disease, and brown foot rot, thus providing protection against multiple soilborne pathogens. R. cerealis reduced the thousand grain weight by 3.6%, whereas seed treatment including fludioxonil and sedaxane against soilborne R. cerealis or M. nivale resulted in a 4% yield increase.

    Wheat (Triticum aestivum) is the most widely grown crop in the United Kingdom; it is grown on approximately 1.7 million ha, with a total annual production of 18 Mt (Department of Environment, Food and Rural Affairs 2019). Intense wheat rotations lead to the build-up of soilborne diseases associated with reductions in yield caused by loss during early plant establishment and disease-imposed limitations on root and stem water/nutrient uptake (Oerke 2006). The soilborne pathogens that occur most commonly in short wheat rotations in the UK include Rhizoctonia solani anastomosis group (AG) 2-1, AG 5, Rhizoctonia cerealis BNR AG-D (Brown et al. 2021), Microdochium, and Fusarium species (Turner et al. 2002).

    In the UK, R. cerealis is predominantly associated with the stem-base disease sharp eyespot of wheat (Hardwick et al. 2001; Parry 1990), which results in premature ripening, shriveled grains, and lodging of wheat (Lemańczyk and Kwaśna 2013). The most recent yield losses caused by sharp eyespot were estimated at 18% and 8 to 10% in New Zealand (Cromey et al. 2002) and in Poland (Lemańczyk and Kwaśna 2013), respectively. There is generally insufficient awareness by growers/agronomists in the UK of the capability of R. cerealis to cause pre-emergence and postemergence damping-off (Parry 1990), and the effects of this pathogen on emergence and establishment losses have not been previously investigated. Similar to R. cerealis, Fusarium and Microdochium spp. are adapted to wheat and cause three diseases within the Fusarium complex in cereals. Seed or soilborne infections develop into Fusarium seedling blight (FSB), which can transition to brown foot rot (BFR) and then to Fusarium head blight (Glynn et al. 2007). FSB arising from seed infection is known to reduce the seed germination capacity, leading to poor crop establishment of wheat (Haigh et al. 2009). However, yield loss caused by FSB from soilborne infection remains unknown. AG 2-1 and AG 5 of R. solani have a diverse host range and cause predominantly pre-emergence and postemergence damping-off on seedlings (Hamada et al. 2011a). Published pathogenicity experiments using wheat seedlings grown under a controlled environment (Demirci 1998; Roberts and Sivasithamparam 1986; Rush et al. 1994; Sturrock et al. 2015) demonstrated significant variations in virulence of their isolates; however, evidence of their ability to cause significant disease in field-grown wheat that may result in yield loss is lacking. Some of these soilborne pathogens occur in complexes that are confounded to specific tissues of the wheat host; for example, BFR and sharp eyespot are part of the stem-base disease complex in cereals. Current knowledge of the symptoms and field yield losses caused by soilborne pathogens is essential for growers and agronomists to optimize disease control as part of crop management.

    Cultural control methods are not consistently effective for intensive wheat cropping; therefore, seed treatments are the most reliable method of protecting seed germination and plant seedling growth when plants are most susceptible to soilborne pathogens (Haigh et al. 2009; Zeun et al. 2013). The broad-spectrum fungicide sedaxane developed by Syngenta Crop Protection inhibits the succinate dehydrogenase enzyme in complex II of the mitochondrial respiration chain (Zeun et al. 2013). Glasshouse studies have shown that sedaxane has activity against R. solani on several crop species, and field plots artificially inoculated with R. cerealis showed a yield advantage as a result of treatments containing sedaxane (Zeun et al. 2013). Fludioxonil, applied as a seed treatment, has been shown to significantly reduce Microdochium and Fusarium DNA in seedlings achieving >90% control against F. culmorum, M. nivale, and M. majus in the field (Glynn et al. 2007). R. cerealis has also been proven sensitive to fludioxonil in vitro (Hamada et al. 2011b). Based on the activity profiles of sedaxane and fludioxonil, a seed treatment containing the active fungicides will be potentially effective against the main soilborne pathogen complex that may pose an early threat to wheat in the UK.

    Therefore, in this study, we aimed to determine the yield losses and effectiveness of seed treatments containing sedaxane and fludioxonil against the main soilborne pathogens found in English wheat fields. The main objectives were to quantify the disease effects of soilborne R. solani AG5, AG2-1, and R. cerealis on the host from emergence through harvest yield, to compare yield loss caused by the most pathogenic Rhizoctonia spp. with soilborne Fusarium graminearum, F. culmorum, or Microdochium nivale, and to determine the effectiveness of fungicide seed treatments on disease severity and yield response.

    Materials and Methods

    Experimental design.

    Two series of field experiments repeated over the course of 2 years were performed. The first series of field experiments were performed in 2012/2013 and in 2013/2014 to determine the effects of different Rhizoctonia spp. on early emergence, establishment, and yield. The second series of experiments in 2016/2017 and in 2017/2018 focused on comparative yield losses with R. cerealis, identified as the most aggressive Rhizoctonia spp. during the first field experiments, Fusarium, and Microdochium spp. The effectiveness of fludioxonil used alone was tested only during the first series of experiments focused on Rhizoctonia spp. because this fungicide is already known to be effective in the field as a seed treatment against Microdochium and Fusarium spp. in wheat. Furthermore, because sedaxane is commercially available in a formulation with fludioxonil, only the formulated seed treatment was included during the second series of experiments.

    The first field series of experiments with winter wheat cultivar Santiago were designed as a randomized block with two factors, pathogen inoculation (noninoculated control, AG 2-1, AG 5, or R. cerealis) and seed treatment (untreated, fludioxonil [5 g active ingredient (a.i.) 100 kg−1], or sedaxane [10 g a.i. 100 kg−1] plus fludioxonil [5 g a.i. 100 kg−1]), allowing 12 treatment combinations with four and three replications in 2012/2013 and in 2013/2014, respectively. The second series of field experiments with winter wheat cultivar Leeds were also designed as a randomized block with two factors, pathogen inoculation (R. cerealis, F. graminearum, and F. culmorium, or M. nivale) and seed treatment (untreated or sedaxane [10 g a.i. 100 kg−1] plus fludioxonil [5 g a.i. 100 kg−1]), allowing six treatment combinations with three replications in 2016/2017 and in 2017/2018.

    Agronomy.

    All field experiments were conducted at the University farm in Sutton Bonington, UK, where winter wheat was sown in October at a standard rate of 320 seeds⋅m2. Inoculum grown on millet seed was drilled with a wheat seed at a rate of 30 g⋅m−2 using a Wintersteiger plot drill. Plot sizes were 6 × 1.6 m in 2012/2013, 1 × 1 m in 2013/2014, and 12 × 1.6 m in 2016/2017 and 2017/2018. Crop protection followed standard agronomic practices, except for the fungicide program, which was designed to give robust protection against foliar diseases and true eyespot utilizing active substances that were not active for R. solani, R. cerealis, F. graminiarum, F. culmorum, and M. nivale (Supplementary Tables S1 and S2).

    Inoculum preparation.

    Inoculum was grown on millet seed following the method described by Zeun et al. (2013). Isolates R. solani AG 2-1 (isolate 1917), AG 5 (isolate 1906), R. cerealis (Rc isolate 1480), F. graminiarum (isolates 13, 15, and 16), F. culmorium (isolates 218 and 236), and M. nivale (isolates 251, 252, and 253) taken from the University of Nottingham isolate collection were raised on potato dextrose agar plates for inoculum production.

    Mixed inoculum for the plot application of Fusarium and Microdochium spp., where more than one isolate was included, was prepared by adding individual isolate-inoculated millet seed in equal ratios. All isolates were of known pathogenicity to wheat. Inoculum was dry and visually inspected for adequate fungal colonization and lack of contamination prior to drilling with the wheat seed.

    Plant sampling and crop assessments during the first series of experiments.

    Crop growth stages were assessed according to Zadoks et al. (1974). Individual crop growth and physiological measurements and disease assessments were performed at growth stage (GS) 15, GS 31, GS 75, and GS 85 during the first series of experiments. At GS 15 and GS 31, 15 plants were randomly collected in each plot for assessments. Later, at GS 39 and GS 75, all plants were removed from within an area of a 0.25-m2 quadrant, randomly placed in each plot, and subjected to detailed biomass assessment. At each sampled growth stage, plants were removed while retaining all aboveground biomass and as much of the top 15 cm of the root system as feasible.

    Seedling emergence and assessment of plant populations.

    Seedling emergence counts were performed periodically after sowing during the first series of experiments and plant numbers were recorded at 29 days postinoculation (dpi) of the soil during the second series of experiments. Emerged seedlings were counted within a 0.25-m2 quadrant (0.5 × 0.5 m) in three replicates per plot. Then, this value was converted to a percentage of the 320 seeds m2 sown. Plant numbers at GS 39 were counted for each m2 per plot during the first experiment.

    Visual disease assessments.

    During the first series of experiments, visual disease assessments of the roots and stems were conducted at GS 15, GS 31, GS 39, and GS 75 for 15 plants per plot following the classification of the root rot disease assessment key described by Strausbaugh et al. (2004) and the classification of eyespot, sharp eyespot, and BFR severity described by Scott and Hollins (1974). During the second series of experiments, assessments were made at GS 15. At GS 15 and GS 31, all tillers were assessed for disease; at GS 39 and GS 75, only the main stem was assessed for disease. Plants were assessed for root rot, sharp eyespot, eyespot, and brown foot root on the stem bases. At GS 15, it was not possible to distinguish between stem-base diseases; therefore, stems were assessed using the stem browning–based Scott and Hollins (1974) disease assessment key.

    Pathogen DNA extraction and quantification using real-time PCR.

    Pathogen DNA extraction and quantification using real-time PCR were performed for the first series of experiments. DNA extraction from soil was only conducted for samples (50 g) collected at GS 15. Soil DNA was extracted using the method developed by Woodhall et al. (2012). Stem samples collected at GS 15 were extracted for DNA using BIOREBA extraction bags because of the small sample size. Stems (5-cm stem basal region) from 15 plants were weighed and placed in a BIOREBA extraction bag. Samples were frozen in liquid nitrogen. Cetyltrimethylammonium bromide (CTAB) buffer (6 ml) was added to the bag before macerating using a BIOREBA Homex 6 flatbed grinder. The resulting supernatant was centrifuged at 2,000 × g for 2 minutes, and 700 µl of clear lysate was transferred to a 2-ml tube containing 200 µl chloroform and vortexed until the mixture turned turbid. Then, these were centrifuged at 13,000 × g for 5 minutes. The resulting supernatant (500 µl) was used for extraction according to instructions for the Wizard food kit (Promega) in combination with the Kingfisher ML magnetic particle processor (Thermo Electron Corporation). Extracted DNA was quantified using a NanoDrop ND-1000 spectrophotometer (Thermo Scientific, Waltham, MA). At GS 31 and GS 75, DNA was extracted from the roots and stems of 15 plants per plot. DNA was extracted from the 10-cm stem basal region at GS 31 (all tillers) and the 15-cm stem basal region at GS 75 (main stems only). The DNA was extracted as described by Ray et al. (2004).

    Real-time PCR assays were performed for AG 2-1, AG 5, and R. cerealis from DNA extracted from plants grown in plots inoculated with the aforementioned pathogens. Primers and probes used during this study are shown in Supplementary Table S3. The quantitative PCR conditions used were the same as those described by Woodhall et al. (2017) for Rhizoctonia spp. and Nielsen et al. (2013) for Microdochium spp.

    Plant height, green area index, and ear partitioning index.

    The plant height (mm) from the stem base to the top of the longest leaf and green area index at GS 15, dry weight, and the ear partitioning index were measured during the first series of experiments. At GS 75, all plants were removed from a 0.25-m2 quadrant for detailed laboratory-based assessments of aboveground plant biomass. Roots were removed from each plant and the sample was weighed to determine the fresh weight (FW). Then, a subsample of 10% (by FW) of plants was selected and partitioned into leaf lamina (flag leaf, second leaf, remaining laminar), true stems, and ears (GS 75 only). The green area for each component part was measured (cm2) using a LI-3100C Area Meter (LI-COR Lincoln, NE). Then, the FW of each component part was recorded before samples were dried in a ventilated/forced draft oven at 80°C until a constant weight was achieved (generally 72 h later). Then, the dry weight of each component part was recorded. These data allowed calculations of the green area index (GAI), aboveground dry weight, and ear partitioning index (EPI) as per Equations 1 and 2.

    The GAI, defined as the green canopy area per unit of ground area, is a precise way of estimating the light-capturing capacity of a canopy (Pask et al. 2012). The GAI was calculated using the following equation:

    GAI (GS39)=((F W/10% of F W)*(GAL+GAS))/ 0.25/ 10000 (1)

    The EPI, defined as the fraction of the aboveground dry weight at GS75 in the ear, was determined using the following equation:

    EPI(GS 75)=DWE/(DWL+DWS+DWE)(2)

    Yield components.

    At maturity (GS 93), plots were harvested with a Sampo plot combine equipped with a grain-weighing system to establish the total yield per plot before converting to tons per hectare (t⋅ha−1). Grain samples were used to quantify the thousand grain weight using a moisture analyzer.

    Statistical analysis.

    All data were analyzed using an analysis of variance (ANOVA) with Genstat version 17.1 for Windows (VSN International Ltd, UK). DNA data were log10-transformed and the disease index, when required, was angular-transformed to normalize residuals. Back-transformed means of transformed data are presented in parentheses. Relationships between the disease index and pathogen DNA were analyzed using a regression analysis. The season (year of experimentation) was included in the treatment structure of the ANOVA, and the results are presented for significant interactions or in their absence for the main effects of factors in the analysis.

    Results

    Effects of Rhizoctonia spp. and seed treatment on emergence and plant populations.

    There were no interactions between factors, including season, for emergence and plant numbers; therefore, the main significant effects are presented in Figure 1. A significant and consistent decrease in plant emergence was observed under inoculation with R. solani AG 2-1 and R. cerealis compared with R. solani AG 5 at 18 dpi (P = 0.019) (Fig. 1A). Reductions of 14% were observed in R. cerealis–inoculated plots compared with the noninoculated control by 26 dpi. Fludioxonil alone or applied with sedaxane increased the emergence (Fig. 1B) and plant numbers at GS39 (Fig. 1C) by 33 and 15%, respectively.

    Fig. 1.

    Fig. 1. Effects of Rhizoctonia solani anastomosis group (AG) A, 2-1, AG 5, and R. cerealis and B, seed treatment on emergence (%) of wheat seedlings and plant populations (m−2) of winter wheat (‘Santiago’) at growth stage (GS) 39 (C) in 2012/2013 and in 2013/2014. Dpi, days postinoculation; control, not inoculated; UT, untreated; F, fludioxonil; S + F, sedaxane and fludioxonil. Error bars indicate standard error (se). * P < 0.05; ***: P < 0.001. lsd, least significant difference at P < 0.05.

    Download as PowerPoint

    Rhizoctonia diseases and effects of seed treatments.

    Fludioxonil and fludioxonil plus sedaxane reduced root rot symptoms by 29.1 and 35.1%, respectively, compared with no treatment (Fig. 2A). At GS 31, there was an interaction between treatment and season (P = 0.046), indicating that the control of root rot was not consistent during the two seasons of experimentation and in 2013/2014, root rot disease was significantly higher in fludioxonil (20%)-treated plots than in the untreated plots (13.6%) (Fig. 2B). At GS 75, there was also an interaction between inoculation and treatment (P = 0.038), with higher root rot disease in AG 5–inoculated plots after treatment with fludioxonil compared with untreated plots (Fig. 2C). Fludioxonil reduced the root rot disease index of the noninoculated control and of R. cerealis–inoculated plots, whereas the addition of sedaxane contributed to a decrease in root rot with AG 2-1 inoculation compared with no treatment (Fig. 2C).

    Fig. 2.

    Fig. 2. Effects of seed treatment on root rot disease index (DI; %) (angular-transformed) by Rhizoctonia solani anastomosis group (AG) 2-1, AG 5, or R. cerealis at A, growth stage (GS) 15, B, GS 31, and C, GS 75 of winter wheat (‘Santiago’) in 2012/2013 and in 2013/14. Control, not inoculated; UT, untreated; F, fludioxonil, S + F, sedaxane and fludioxonil; lsd, least significant difference at P < 0.05.

    Download as PowerPoint

    Stem-base diseases and effects of seed treatments.

    Because of the difficulty identifying the early symptoms of individual stem-base diseases at GS 15, stems were assessed for general stem browning (Table 1). The interaction between inoculation and season was significant (P = 0.002), indicating that AG 5 caused greater stem browning compared with the noninoculated control during 2012/2013. During both seasons, the highest stem browning at GS 15 occurred in R. cerealis–inoculated plots. There was also stem browning in the noninoculated control in 2012/2013, suggesting that other pathogens were present. Real-time PCR revealed that these symptoms were associated with Microdochium spp. Seed treatments reduced stem browning (P < 0.001), and the interaction with season was significant (P < 0.001) because fludioxonil did not control stem browning in 2013/2014. In contrast, sedaxane plus fludioxonil significantly reduced stem browning compared with no treatment by 47 and 38% in 2012/2013 and 2013/2014, respectively (Table 1).

    Table 1. Effects of inoculation with Rhizoctonia solani anastomosis group (AG) 2-1, AG 5, or R. cerealis and seed treatment with sedaxane and/or fludioxonil on stem browning disease index (angular-transformed; %) at growth stage (GS) 15 of winter wheat (‘Santiago’) in 2012/2013 and 2013/2014a

    Sharp eyespot and BFR were assessed at GS 31, GS 39, and GS 75 (Fig. 3). The sharp eyespot index was highest for the R. cerealis–inoculated plots throughout the assessment period (P < 0.001) (Fig. 3A). Slight symptoms were observed in plots inoculated with AG 2-1 (2.1 to 5.1%) and AG 5 (3 to 5%) and the noninoculated control plots (3 to 5.7%). Seed treatments had no effects on sharp eyespot disease (data not presented).

    Fig. 3.

    Fig. 3. Effects of inoculation with Rhizoctonia solani anastomosis group (AG) A, 2-1, AG 5, and R. cerealis on sharp eyespot disease index (DI; %) (angular-transformed) B, brown foot rot DI (%) (angular-transformed), and C, effect of seed treatment on brown foot rot DI (%) (angular-transformed) of winter wheat (‘Santiago’) in 2012/2013 and in 2013/2014. Control, not inoculated; F, fludioxonil; S + F, sedaxane and fludioxonil. Error bars indicate standard error (SE). *P < 0.05; ***P < 0.001.

    Download as PowerPoint

    The brown foot rot index was significantly (P < 0.001) lower for R. cerealis–inoculated (8.4 to 15.6%) plots than for AG 2-1–inoculated (20.4 to 27.5%), AG 5–inoculated (21.1 to 27.5%), and noninoculated control plots (25.8 to 28.5%) (Fig. 3B). Sedaxane plus fludioxonil reduced BFR by 30.4 and 23% compared with no treatment at GS 31 and GS 39, respectively (Fig. 3C). No seed treatment effect was present at GS 75.

    Effects of inoculation and seed treatment on Rhizoctonia spp. DNA in soil and in plants.

    Pathogen DNA in soil samples was quantified at GS 15 (Table 2). The highest DNA concentrations were quantified in the inoculated untreated plots. DNA of R. cerealis was found at >4000 pg⋅g−1 of soil, followed by AG 2-1 DNA (>220 pg⋅ng−1 of soil) and AG 5 DNA, which differed significantly in 2012/2013 and 2013/2014 (141 and 0.10 pg⋅g−1 of soil, respectively) (Table 2). DNA of AG 2-1, AG 5, and R. cerealis were detected in the roots at GS 31 and GS 75. There was a significant interaction between inoculation and season for the amount of Rhizoctonia spp. in the roots of the wheat host at GS 31 (P = 0.049) and at GS 75 (P = 0.001), possibly because of inconsistencies in the DNA amounts of AG 5 quantified during the two seasons (Table 2). Treatment had no effect on pathogen DNA in soil and in roots at GS 31. At GS 75, DNA in the roots of AG 2-1 and R. cerealis were at lower concentrations than those at the previous growth stage (Table 2). There was a significant interaction between inoculation and treatment (P = 0.025) associated with inconsistencies in the effects of treatments on inoculated plots with AG 5. Overall, lower amounts of DNA of AG 2-1 and R. cerealis were found in roots during both seasons in plots treated with sedaxane plus fludioxonil (Table 2).

    Table 2. Effects of inoculation with Rhizoctonia solani anastomosis group (AG) 2-1, AG 5, or R. cerealis and seed treatment with sedaxane and/or fludioxonil on DNA of Rhizoctonia spp. (pg⋅g−1 of soil) in soil at growth stage (GS) 15 and in roots at GS 31 and GS 75 of winter wheat (‘Santiago’) in 2012/2013 and 2013/2014a

    At GS 15, there was a significant interaction among inoculation, season, and treatment with higher DNA concentrations of R. cerealis and AG 5 in stems during the first season in contrast to AG 2-1 DNA concentrations, which accumulated more during the second season (Table 3). Furthermore, the effectiveness of seed treatment to reduce pathogen DNA in stems was inconsistent between species and seasons; for example, fludioxonil reduced R. cerealis DNA during the first season, but not during the second season. At GS 31, the amount of DNA of all pathogens increased in untreated stems (Table 3). AG 2-1 and R. cerealis DNA in stems were 4-fold and 12.5-fold higher than that in AG 5, respectively (P = 0.002) (Table 3). However, there was no effect of seed treatment and there were no interactions at GS 31. Less DNA accumulated in stems at GS 75 than at GS 31, although AG 5 DNA during the first season and R. cerealis DNA during the second season were highest from the three pathogens (P < 0.001) (Table 3). Seed treatment had no significant effect on pathogen DNA in stems, and there were no interactions for this growth stage.

    Table 3. Effects of inoculation with Rhizoctonia solani anastomosis group (AG) 2-1, AG 5, or R. cerealis and seed treatment with sedaxane and/or fludioxonil on DNA of Rhizoctonia spp. (pg⋅ng−1 of total DNA) in wheat stems at growth stage (GS) 15, GS 31, and GS 75 of winter wheat (‘Santiago’) in 2012/2013 and 2013/2014a

    Effects of inoculation and seed treatment on Microdochium spp. DNA in stems.

    M. nivale and M. majus were detected predominantly in 2012/2013 at GS 15 (Table 4). There was no significant pathogen or treatment effect for M. nivale (Table 4). DNA of M. majus was lower than DNA of M. nivale in plots inoculated with AG 2-1 and R. cerealis, and it was significantly reduced by sedaxane plus fludioxonil compared with no treatment.

    Table 4. Effects of inoculation with Rhizoctonia solani anastomosis group (AG) 2-1, AG 5, or R. cerealis and seed treatment with sedaxane and/or fludioxonil on DNA of Microdochium majus and M. nivale (pg⋅ng−1 of total DNA) in stems at growth stage (GS) 15 of winter wheat (‘Santiago’) in 2012/2013 and 2013/2014a

    Yield components.

    Plant height was reduced significantly by R. cerealis and R. solani AG 2-1 compared with that of the noninoculated plots during both seasons (Table 5). This effect was negated by fludioxonil and sedaxane plus fludioxonil treatments in AG 2-1 and R. cerealis; however, the effects of seed treatments on plants in AG 5–inoculated plots were less consistent (Table 5).

    Table 5. Effects of inoculation with Rhizoctonia solani anastomosis group (AG) 2-1, AG 5, or R. cerealis and seed treatment with sedaxane and/or fludioxonil on plant height (mm) and green area (cm2) at growth stage (GS) 15 of winter wheat (‘Santiago’) in 2012/2013 and 2013/2014a

    In contrast to plant height, inoculation had no effect on GAI at GS 15, but fludioxonil increased the GAI of inoculated plots. Sedaxane plus fludioxonil treatment showed the same effects during the first season of experimentation; however, during the second experiment, the effects were not consistent in AG 5–inoculated and AG 2-1–inoculated plots.

    The EPI is the fraction of aboveground DM partitioned in the ear. A significant interaction was observed between inoculation and season (P = 0.048) (Supplementary Table S4). Therefore, in 2012/2013, there were only slight differences in the EPI of the inoculated and noninoculated control plots. However, in 2013/2014, R. cerealis significantly reduced EPI compared with the control (Table S4).

    The yield is presented only for 2012/2013 because the plots in 2013/2014 were too small (1 × 1 m) to accurately determine field harvest yield on a per-hectare basis. Differences based on inoculation were not significant (P < 0.05); however, yields of infected plots with R. cerealis, AG2-1, and AG5 were 0.83, 0.44, and 0.22 t⋅ha−1, respectively, lower than that of the control (Fig. 4A). R. cerealis reduced the thousand grain weight significantly (by 3.6%) during both seasons compared with the noninoculated control (Fig. 4B). There was no significant effect of seed treatment on the yield or thousand grain weight.

    Fig. 4.

    Fig. 4. Effects of inoculation with Rhizoctonia solani anastomosis group (AG) A, 2-1, AG 5, or R. cerealis on grain yield (t⋅ha−1) in 2012/2013 and B, thousand grain weight (TGW; g) of winter wheat (‘Santiago’) in 2012/2013 and in 2013/2014. Control, not inoculated; lsd, least significant difference at P < 0.05.

    Download as PowerPoint

    Relationships between disease assessments and pathogen DNA.

    Regression analyses (R2) using disease indexes and pathogen DNA revealed that there were no significant correlations between Rhizoctonia DNA in roots and root rot (data not shown). There were significant, but generally weak to moderate (R2 ≤ 0.40), relationships between Rhizoctonia spp. DNA in stems and symptoms on the stems (Supplementary Table S5). The strongest relationship (R2 = 0.60) was between Rhizoctonia spp. DNA in stems at GS 15 and the stem browning index at GS 15 and between Rhizoctonia spp. DNA in stems at GS 15 and the sharp eyespot index at GS 75 (R2 = 0.45). Sharp eyespot was negatively related to BFR at GS 31, GS 39, and GS 75. There was also a very weak but significant relationship between stem browning at GS 15 and Microdochium DNA in stems at GS 15 (Supplementary Table S5).

    Effects of seed treatments on early disease by Fusarium spp., M. nivale, and R. cerealis and yield.

    To determine the effects of sedaxane plus fludioxonil against other common soilborne pathogens and final yield, we performed wheat field experiments during two consecutive seasons using Fusarium spp., M. nivale, and R. cerealis inoculation. Plant populations (m−2) were lowest during the first season of experimentation in plots inoculated with M. nivale and lowest during the second season in plots inoculated with R. cerealis (Fig. 5A). Higher plant numbers were observed in sedaxane plus fludioxonil–treated plots during both seasons but not for Fusarium spp. in 2017/2018 (Fig. 5A). The greatest effect of sedaxane plus fludioxonil treatment was seen under R. cerealis inoculation in 2017, showing a 34.3% increase in plant numbers compared with the untreated plots. There was a 9% increase under M. nivale inoculation. R. cerealis caused more severe stem browning disease compared with Fusarium spp. or M. nivale (Fig. 5B), and treatment reduced disease symptoms in inoculated plots at GS 15 by 43% overall (Fig. 5C). There were no interactions between factors for yield, and the yield response to treatment was 0.27 t⋅ha−1 (P = 0.02) (Fig. 5D).

    Fig. 5.

    Fig. 5. A, Effects of inoculation with Fusarium graminearium, F. culmorum, M. nivale, or R. cerealis and treatment with sedaxane and fludioxonil on plant populations (m−2) at 29 dpi, B and C, stem browning disease index (growth stage [GS] 15), and D, yield of winter wheat (‘Leeds’) in 2016/2017 and 2017/2018. UT, untreated; S + F, sedaxane and fludioxonil; lsd, least significant difference at P < 0.05.

    Download as PowerPoint

    Discussion

    In these studies, we show that R. cerealis is capable of causing pre-emergence and postemergence damping-off before developing into sharp eyespot, thus causing losses in the establishment and yield of wheat from the combined effects of both seedling and stem-base disease. In contrast, R. solani AG 2-1 and AG 5 caused relatively small reductions in emergence, suggesting that these species or the isolates we used in this work are unlikely to cause pre-emergence or postemergence damping-off in field-grown wheat. R. cerealis has been previously reported to cause damping-off (Hamada et al. 2011a; Parry 1990), but it has not been generally associated with establishment losses. In the second series of field experiments in 2016/2017 and 2017/2018, we compared the effects of soilborne R. cerealis, identified as the more aggressive Rhizoctonia spp., with FSB pathogens such as Fusarium spp. or M. nivale that are commonly associated with reductions in emergence and establishment (Humphreys et al. 1995) when they are seed-borne. Results using soilborne inoculum showed that R. cerealis and M. nivale were the main pathogens causing low emergence associated with the greatest reductions in plots in 2016/2017. Microdochium nivale and M. majus confirmed in stems at GS 15 in the untreated plots in 2012/2013 were also implicated in reduced emergence, stem browning, and BFR during the first series of field experiments, although it is possible that M. nivale and M. majus infection may have been seed-borne. We did not confirm the seed load of these species before sowing; however, seed viability by the seed producer was confirmed. During the second series of experiments, F. graminearum and F. culmorum failed to cause significant disease with soilborne inoculum compared with M. nivale or R. cerealis. It is possible that environmental conditions or the inoculation method used here failed to favor Fusarium infection. Therefore, it is not surprising that the effects of disease or seed treatments were not detected in Fusarium-inoculated plots.

    There are contrasting results of the pathogenicity and aggressiveness of isolates of AG 2-1 and AG 5 in wheat roots (Roberts and Sivasithamparam 1986; Rush et al. 1994). Throughout the period of our studies, there was no significant difference in root rot disease in R. solani–inoculated and noninoculated plots, suggesting that the isolates of AG 2-1 and AG 5 used here were only weakly pathogenic to wheat roots under field conditions. Limitations in the accuracy of visual assessments of root rot symptoms can be overcome by quantifying pathogen DNA in roots. Real-time PCR assays detected AG 2-1, AG 5, and R. cerealis DNA in roots, indicating these pathogens were able to colonize wheat roots; however, no association was found between Rhizoctonia DNA in roots and root rot symptoms, indicating that symptoms may be caused by a complex of species in the rhizosphere that cause disease with very similar symptoms (Harris and Moen 1985) or additional damage caused by pests. Furthermore, the results here showed significant reductions of root rot symptoms with the application of seed treatments but no consistent effects on individual pathogen DNA in roots, corroborating that symptoms may have been also associated with organisms other than AG 2-1, AG 5, or R. cerealis.

    Stem-base diseases at early growth stages of the host are difficult to distinguish (Brown et al. 2021; Turner et al. 1999, 2001). Therefore, plants at GS 15 were assessed for indiscriminate stem browning. The most stem browning consistently occurred in R. cerealis–inoculated plots during both seasons. In addition, R. cerealis DNA was quantified at higher concentrations than AG 2-1 and AG 5 in stems, suggesting that wheat is more susceptible to infection by the adapted R. cerealis than by the generalists, AG 2-1 or AG 5. R. cerealis on plants at GS 15 significantly reduced plant height, resulting in the appearance of stunted plants. R. solani AG 5 also caused considerable stem browning in 2012/2013, when pathogen DNA in the soil of plots sown with untreated seed was >1,400-fold higher than that in 2013/2014. This inconsistency in DNA accumulation in soil suggests that differences in environmental conditions have a significant role in the occurrence and severity of stem disease with AG 5. The acute stem browning on seedlings in the noninoculated control plots in 2012/2013 indicated the presence of other pathogens confirmed as Microdochium spp. and quantified at relatively high amounts in stems at GS 15 in 2012/2013 compared with 2013/2014. Because DNA of Rhizoctonia spp., rather than DNA of Microdochium spp., accounted for 60% of the variation in stem browning at GS 15, we can be confident that Rhizoctonia spp. were responsible for the majority of stem symptoms at this early stage. In naturally infected fields, symptoms of sharp eyespot are typically observed after stem extension (Lemańczyk and Kwaśna 2013). However, the inoculation method here delivered inoculum directly next to the sown seed, allowing the pathogen to colonize the developing seedling before stem extension (GS 31), thus resulting in early sharp eyespot by R. cerealis. The BFR index of the naturally occurring Microdochium was significantly lower in R. cerealis–inoculated plots than in AG 2-1–inoculated, AG 5–inoculated, or noninoculated control plots throughout the growing season. There were also weak but significant negative relationships between the sharp eyespot and BFR index, suggesting competitive interactions between R. cerealis and the BFR-causing species of Microdochium. R. cerealis had a competitive advantage because of the higher inoculum density in soil, and it may have suppressed the naturally occurring Microdochium, in agreement with observations of a previous study by Pettitt et al. (2003).

    Reductions in plant height, EPI, and yield were mostly associated with the effects of R. cerealis. Our results suggest that R. cerealis causing sharp eyespot imposes significant limitations on source-to-sink partitioning via biomass reductions, resulting in a yield loss of 8%, similar to the estimated maximum yield losses in Poland of 8 to 10% in naturally infected commercial wheat fields (Lemańczyk and Kwaśna 2013).

    Fludioxonil and sedaxane plus fludioxonil increased plant emergence by 47.9 and 50.2%, respectively, in 2012/2013. Therefore, under high disease pressure caused by a mixture of Rhizoctonia and naturally occurring Microdochium spp., there was a significant response to both seed treatments. Fludioxonil (Glynn et al. 2007, 2008) and sedaxane (Zeun et al. 2013) have been shown to effectively control Microdochium spp. in vitro and in the field (Jonavičienė et al. 2016). Furthermore, in this study, sedaxane, as part of a mixture with fludioxonil, under high disease pressure significantly increased plant populations at GS 39 by 29% compared with no treatment. The effectiveness of the two actives of the seed treatment was confirmed in 2017/2018 against emergence losses caused by R. cerealis, when the observed increase above that of the control was 35%. This demonstrated an additional benefit of sedaxane controlling damping-off caused by R. cerealis, which is in agreement with studies that showed high effects of sedaxane on mycelial growth inhibition of Rhizoctonia spp. (Da Silva et al. 2017; Zeun et al. 2013). Sedaxane and fludioxonil were also able to reduce root rot on winter wheat at GS 15 in field conditions, in agreement with a previous report of a controlled environment showing similar results for rhizoctonia root rot in maize, corn, and cotton (Zeun et al. 2013). The main period of activity of a seed treatment is generally considered to last 4 to 6 weeks after sowing; therefore, the lack of consistency at the later growth stages of 31 and 75, was not surprising. Fludioxonil alone was less consistent because its effect was only significant during the 2012/2013 experiment. However, the mixture of sedaxane and fludioxonil reduced stem browning and the disease index in all experiments. Therefore, the addition of sedaxane extends the period of effectiveness of seed treatment when controlling early stem-base diseases caused by Rhizoctonia spp. and Microdochium spp. Quantification of Microdochium spp. DNA showed that fludioxonil consistently controlled M. nivale, but that DNA of M. majus was not detected with sedaxane plus fludioxonil treatments during both seasons, suggesting that sedaxane contributed to controlling this species. Seed-treated plots also had significantly lower BFR index values at GS 31 and GS 39 compared with untreated plots, indicating that fludioxonil controlled BFR up to GS 39. There was no significant effect of seed treatment on Rhizoctonia DNA concentrations or sharp eyespot after GS 31, GS 39, or GS 75, suggesting that sharp eyespot control required additional stem base fungicide applications that should be applied at the beginning of stem extension (GS 30 to GS 31) (Nicholson et al. 2002). During the second series of experiments, F. graminearum and F. culmorum failed to cause significant disease in soil inoculum compared with M. nivale or R. cerealis, and we were unable to detect the effects of seed treatments in Fusarium-inoculated plots. The inoculum method used here was developed for and favored Rhizoctonia, in contrast to Fusarium or even Microdochium spp., which have been shown to cause severe seedling blight/foot rot disease from infected seed (Haigh et al. 2009; Ren et al. 2016) or ground surface inoculation simulating leftover debris (Jones et al. 2018).

    Artificially inoculated experiments provide useful information regarding the worst-case scenarios for losses caused by pathogens applied at a high inoculum density, and this approach is appropriate for establishing comparative differences and the effectiveness of control methods for individual pathogens. Under natural infection, the inoculum density of these pathogens is likely to be lower; Fusarium and Microdochium spppose significant threats to the host from other sources of inoculum. However, this work focused on diseases initiated by soilborne inoculum and, as such, is the first report of their comparative effects on wheat. Based on our results, soilborne disease control in the UK should focus on R. cerealis and M. nivale, which reduced wheat emergence and establishment. Fludioxonil and sedaxane are effective seed treatments that reduce damping-off and foot rot, thus providing control against more than one pathogen and disease, resulting in a modest but significant response of 4% in the yield of wheat.

    Acknowledgments

    We thank Gina Swart, Christian Schlatter, Brigitte Slaats, Michael Tait, and Jon Ronskey from Syngenta for their support.

    The author(s) declare no conflict of interest.

    Literature Cited

    The author(s) declare no conflict of interest.

    Funding: Funding was received from Syngenta International (310004) and the PhD studentship of Matthew Brown “Elucidating crop loss and control of Rhizoctonia solani and Rhizoctonia cerealis in winter wheat (RG35DC).