Solani AG Research Articles

Diversity and Aggressiveness of Rhizoctonia spp. from Nebraska on Soybean and Cross-Pathogenicity to Corn and Wheat.

Rhizoctonia and Rhizoctonia-like species of fungi that cause disease are known to have varying host ranges and aggressiveness. Accurate identification of these species causing disease is important for soybean disease management that relies upon crop rotation. The anamorphic genus Rhizoctonia contains several diverse species and anastomosis groups (AGs) including some known soybean pathogens, such as Rhizoctonia solani, whereas for others the ability to cause disease on soybean has not been well described. The present study was conducted to identify the predominant species and AG of Rhizoctonia from soybean, corn, and wheat fields that are pathogenic on soybean and characterize cross-pathogenicity to common rotational crops, corn and wheat. We surveyed for Rhizoctonia spp. in Nebraska; isolates were identified to species and AG, and aggressiveness was assessed. A total of 59 R.zeae isolates, 49 R.solani, nine binucleate Rhizoctonia, three R.circinata, and two R.oryzae isolates were collected in 2016 and 2017 from a total of 29 fields in 15 counties. The most abundant R.solani AGs were AG-4, AG-1 IB, AG-2-1, AG-3, and AG-5. R.solani AG-4 and R. zeae were found in all three regions of the state (west, central, and eastern). Some isolates that were most aggressive to soybean seedlings were cross-pathogenic on both wheat and corn. In addition, R. zeae was pathogenic on soybean when evaluated at 25°C, which is warmer than temperatures used previously, and isolates were identified that were aggressive on soybean and cross-pathogenic on both corn and wheat.

Secreted Glycosyltransferase RsIA_GT of Rhizoctonia solani AG-1 IA Inhibits Defense Responses in Nicotiana benthamiana

Anastomosis group AG-1 IA of Rhizoctonia solani Khün has a wide host range and threatens crop production. Various glycosyltransferases secreted by phytopathogenic fungi play an essential role in pathogenicity. Previously, we identified a glycosyltransferase RsIA_GT (AG11A_09161) as a secreted protein-encoding gene of R. solani AG-1 IA, whose expression levels increased during infection in rice. In this study, we further characterized the virulence function of RsIA_GT. It is conserved not only in Basidiomycota, including multiple anastomosis groups of R. solani, but also in other primary fungal taxonomic categories. RsIA_GT possesses a signal peptide (SP) for protein secretion, and its functionality was proven using yeast and Nicotiana benthamiana. The SP-truncated form of RsIA_GT (RsIA_GT(ΔS)) expressed in Escherichia coli-induced lesion-like phenotype in rice leaves when applied to punched leaves. However, Agrobacterium-mediated transient expressions of both the full-length RsIA_GT and RsIA_GT(ΔS) did not induce cell death in N. benthamiana leaves. Instead, only RsIA_GT(ΔS) suppressed the cell death induced by two reference cell death factors BAX and INF1 in N. benthamiana. RsIA_GT(ΔS)R154A D168A D170A, a mutant RsIA_GT(ΔS) for the glycosyltransferase catalytic domain, still suppressed the BAX- or INF1-induced cell death, suggesting that the cell death suppression activity of RsIA_GT(ΔS) would be independent from its enzymatic activity. RsIA_GT(ΔS) also suppressed the H2O2 production and callose deposition and showed an effect on the induction of defense genes associated with the expression of BAX and INF1. The transient expression of RsIA_GT(ΔS) in N. benthamiana enhanced the lesion area caused by R. solani AG-1 IA. The secreted glycosyltransferase, RsIA_GT, of R. solani AG-1 IA is likely to have a dual role in virulence inside and outside of host cells.

Adaptability of Rhizoctonia solani AG-1 IA for mancozeb sensitivity under temperature stress

The genetic architecture of quantitative characters in plants can be influenced by stress due to environmental changes, in combination with the decrease in the organism’s average performance, resulting in genetic and environmental variances. The main objective of this study was to determine how the high-temperature stress affects the sensitivity of three populations of the soybean foliar blight pathogen Rhizoctonia solani AG-1 IA from Mato Grosso, Maranhão, and Tocantins to a broad-spectrum fungicide. The specific objective was to determine the effect of environmental stress on evolvability components (i.e., the selection response measures such as genotypic, environmental, and phenotypic variances) associated with sensitivity to the broad spectrum dithiocarbamate fungicide mancozeb. The fungal isolates from the three pathogen populations were grown under two temperatures (25 °C and 33.5 ºC, optimum and stress, respectively) and three fungicide concentrations (0.0, 0.32, and 0.64 g·L-1 of active ingredient). The mycelial growth was measured, and evolvability components, such as the genotypic variance coefficient (IG), the environmental variance (IE), and the broad-sense heritability (h2), were determined. The results showed that high-temperature stress decreased (≈ 0.1 units, in a scale from 0 to 1.0) the genotypic variance and the heritability for mancozeb sensitivity in three populations of the soybean foliar blight pathogen R. solani AG-1 IA.

Occurrence of Root and Stem Rot Caused by Rhizoctonia solani AG-4 HGI on Torenia fournieri in China.

Torenia fournieri (wishbone flower) is widely cultivated for ornamental and medicinal use in southern China. In July 2021, severe root and stem rot of T. fournieri were observed in approximately 70% of the 3,000 plants grown in an ornamental plant nursery under natural conditions located in Guangzhou, Guangdong Province, China. Lesions on the roots and basal stems were water-soaked and dark brown. In late infection, the whole plant wilted and died. Eventually, infected plants lose their ornamental and economic value. Fifteen diseased samples were collected, and diseased tissues (4 × 4 mm) were surface sterilized in 1% NaOCl for 45 s, rinsed with sterile water, dried, and incubated for 3 d on potato dextrose agar (PDA) medium at 25°C in the dark. Pure cultures were obtained by transferring hyphal tips to new plates of PDA and incubated at 25°C for 5 d. A fungus was consistently isolated from symptomatic root and stem samples (100% isolation rate). The colonies on PDA medium were initially white, turning light brown and with age, no sclerotia was present after 15 days. Width of hyphae was 3.78 to 11.10 μm , which with a constriction at the base of hyphal branches, septa near the branch, and right-angled branching. Multinucleate hyphal cells (3 to 12 nuclei per cell) were determined by 4'-6-diamidino-2-phenylindole (DAPI) staining (Yang et al. 2013). These characteristics matched those described for Rhizoctonia solani (Sneh et al. 1991). A representative isolate LZ1 was chosen for molecular identification, the internal transcribed spacer (ITS) region was amplified using the primers ITS1/ ITS4, and the resulting sequence (633 bp) were deposited in GenBank (accession no. MZ798228). A BLASTn search of the ITS sequence exhibited 99% identity with that of R. solani AG-4 HGI (MK430998). The phylogenetic analysis (neighbor-joining method and 1,000 bootstrap values) showed LZ1 was clearly assigned to group of R. solani AG-4 HGI. For pathogenicity testing, ten healthy T. fournieri plants (approximately 18 cm high) were planted into plastic pot filled with sterilized soil. Inoculum of R. solani was produced by adding a PDA agar plug (5 mm in diameter) from a 6-day-old colony to a flask with 150 ml of potato dextrose broth (PDB), shaken at 25°C and 150 rpm for 7 days, and poured evenly into the sterilized soil (50ml per pot ). Another 10 plants were poured with sterile PDB as controls. All the inoculated plants placed in a greenhouse at 25°C and 70-90% humidity conditions. After 14 days, rot symptoms of the roots and basal part of stems similar to those in the nursery were observed on all inoculated plants, while no visible symptoms were observed on the controls. We reisolated the fungal pathogen from the inoculated plants and identified to be R. solani AG-4 HGI by microscopy and molecular characteristics mentioned above. R. solani AG-4 HGI was previously reported causing Impatiens walleriana Stem Rot and muskmelon fruit rot in Liaoning, China (Sun et al. 2015). To the best of our knowledge, this is the first report of Rhizoctonia solani AG-4 HGI causing root and stem rot on Torenia fournieri in China as well as worldwide. This finding contributes to expand the host range known for R. solani AG-4 HGI and will assist in developing effective control strategies of this disease to minimize the losses.

Bioprospecting Fluorescent Pseudomonas from the Brazilian Amazon for the Biocontrol of Signal Grass Foliar Blight

Biological control is considered the only viable integrated disease management practice for controlling the widely distributed and destructive foliar blight and sudden death disease caused by the fungus Rhizoctonia solani AG-1 IA on signal grass (Urochloa brizantha) pastures. Since major signal grass varieties are highly susceptible and fungicide sprays are not labeled for grass pasture cropping systems, biological control is sought as an alternative for managing this fungal disease. In this study, 24 fluorescent Pseudomonas isolates obtained from naturally suppressive soils from the Amazon biome were bio-prospected for their role as biocontrol agents against R. solani AG-1 IA. Based on in vitro antagonism, three isolates (Amana, Poti, and Yara) were selected for further in vivo assays. Multilocus phylogenetic analysis indicated that Amana and Yara were grouped into the Pseudomonas putida group while Poti was grouped into the Pseudomonas asplenii group, and could well constitute a new Pseudomonas species. For in vivo biocontrol assays, the biocontrol agents were applied either via seed-treatment or via foliar spray. All three isolates produced siderophores and solubilized phosphate, while Amana and Poti showed protease and chitinase in vitro activity. Foliar application of P. putida Amana from Amazonian suppressive soils resulted in a significant reduction of the foliar blight disease severity on signal grass. We discuss further steps for the development and labeling of Pseudomonas-based biofungicides for managing the foliar blight disease on signal grass pastures in Brazil.

Thiamine and Piriformospora indica induce bean resistance against Rhizoctonia solani: The role of polyamines in association with iron and reactive oxygen species

In the present study, the protective effects of thiamine and Piriformospora indica against the pathogenic fungus Rhizoctonia solani AG-4 HGII, which causes root rot, have been investigated in bean plants. Our results revealed that thiamine and P. indica significantly reduced the disease progress in the leaf discs. Disease suppression via application of thiamine and the endophytic fungus, P. indica, was linked to biochemical and metabolic changes in bean plants. Accumalation of hydrogen peroxide (H2O2), superoxide anion (O2–), and iron ions decreased in the leaf discs pretreated with thiamine and/or P. indica, while the formation of polyamines (PAs) such as putrescine (Put), spermidine (Spd), and spermine (Spm) dramatically increased. We observed that occurrence of cell death, the levels of ferric (Fe3+) and ferrous (Fe2+) ions were positively correlated with H2O2 accumulation. Simultaneous increase in the generation of H2O2, Fe3+ and Fe2+ may play a role in induction of cell death. A positive correlation was also observed between PAs and H2O2 levels. Decreased accumulation of H2O2 in different treatments may be a result of an increase in the PAs contents. These findings suggest that formation of PAs may decrease progress of the disease via decreasing H2O2 accumulation in the plant cells. This is the first report on the crucial role of PA contents and their interplay with iron ions and reactive oxygen species in thiamine and P. indica mediated induction of bean defense responses against a necrotrophic fungal pathogen.

Three Genes Related to Trehalose Metabolism Affect Sclerotial Development of Rhizoctonia solani AG-1 IA, Causal Agent of Rice Sheath Blight

Trehalose metabolism is related to the sclerotial development of Rhizoctonia solani AG-1 IA, the causal agent of rice sheath blight (RSB). Here, we further elucidated the functions of three genes Rstre, Rstps1 and Rstpp that encode three key enzymes trehalase (TRE), alpha, alpha-trehalose- phosphate synthase (TPS1) and trehalose 6-phosphate phosphatase (TPP) in the sclerotial development of R. solani AG-1 IA. Due to the lack of a stable genetic transformation system for R. solani, the heterologous expression of these three genes in Pichia pastoris GS115 was performed. The results showed that reactive oxygen species (ROS) contents and enzyme activities in R. solani decreased significantly in the treatments of the fermentation broths of Rstps1 and Rstpp transformants, and that in the treatment of the fermentation broth of Rstre transformant visibly increased. Furthermore, the fermentation broths of the transformants of all the three genes were added to potato dextrose agar (PDA) medium for the cultivation of R. solani, as a result, the dry weight of sclerotia in each PDA plate containing the fermentation broths of Rstps1 and Rstpp transformants significantly increased compared with the control, and that of Rstre transformant obviously decreased. Finally, 178 proteins were found to interact with RSTPS1, and 16 of them were associated with ROS. Taken together, the findings suggest that all these three genes related to trehalose metabolism play important roles in the sclerotial development of R. solani AG-1 IA, and can be used as new targets for the development of novel high-efficiency fungicides for the controlling of RSB.

Six Novel Mycoviruses Containing Positive Single-Stranded RNA and Double-Stranded RNA Genomes Co-Infect a Single Strain of the Rhizoctoniasolani AG-3 PT.

Six novel mycoviruses that collectively represent the mycovirome of Rhizoctonia solani anastomosis group (AG)-3 PT strain ZJ-2H, which causes potato black scurf, were identified through metatranscriptome sequencing and putatively designated as Rhizoctonia solani fusarivirus 4 [RsFV4, positive single-stranded RNA (+ssRNA)], Rhizoctonia solani fusarivirus 5 (RsFV5, +ssRNA), Rhizoctonia solani mitovirus 40 (RsMV40, +ssRNA), Rhizoctonia solani partitivirus 10 [RsPV10, double-stranded RNA (dsRNA)], Rhizoctonia solani partitivirus 11 (RsPV11, dsRNA), and Rhizoctonia solani RNA virus 11 (RsRV11, dsRNA). Whole genome sequences of RsFV4, RsMV40, RsPV10, RsPV11, and RsRV11, as well as a partial genome sequence of RsFV5, were obtained. The 3’- and 5’- untranslated regions of the five mycoviruses with complete genome sequences were folded into stable stem-loop or panhandle secondary structures. RsFV4 and RsFV5 are most closely related to Rhizoctonia solani fusarivirus 1 (RsFV1), however, the first open reading frame (ORF) of RsFV4 and RsFV5 encode a hypothetical protein that differs from the first ORF of RsFV1, which encodes a helicase. We confirmed that RsPV10 and RsPV11 assemble into the spherical virus particles (approximately 30 nm in diameter) that were extracted from strain ZJ-2H. This is the first report that +ssRNA and dsRNA viruses co-infect a single strain of R. solani AG-3 PT.

Rhizoctonia Resistance Is Negatively Correlated to Early Root Growth Rate in Synthetic Hexaploid Wheat Derivatives.

Resistance to the soilborne fungal pathogen Rhizoctonia solani AG-8 is desirable in adapted wheat and barley but remains an elusive trait for prebreeders and breeders. In a previous study, we observed that emergence and root growth was faster in the Rhizoctonia-susceptible 'Scarlet' than in its resistant counterpart, 'Scarlet-Rz1'. The objective of the current study was to quantify early root growth rate and total root length in resistant and susceptible synthetic hexaploid wheat lines, including parental lines and 22 recombinant inbred lines derived crosses between parental lines. In Petri dish assays, the susceptible lines displayed a faster rate of root growth during the first 40 h of root emergence compared with resistant lines. This growth differential was observed in 14-day and 48-h greenhouse assays, in which the total root length of susceptible parental lines was significantly (P < 0.05) greater than that of resistant parental lines. However, the resistant lines sustained less root loss compared with susceptible lines when R. solani AG-8 was present in the soil. Early root growth rate and total root length were not correlated to freezing tolerance in a set of wheat cultivars selected for cold tolerance. The findings indicated that early root growth was negatively correlated to R. solani AG-8 damage in resistant synthetic wheat lines developed for the Pacific Northwest, United States, and suggested that the dynamics of root emergence affect resistance to this soilborne pathogen.

First Report of Rhizoctonia solani AG-4 HG-I Causing Fruit Rot on Tomato in Mexico.

The culture of tomato (Solanum lycopersicum L.) is an important food, fruits are source of vitamins and minerals, they also prevent chronic diseases such as cancer and vascular diseases (Willcox, 2003); in Mexico 47,372 ha are destined to tomato cultivation (SIAP, 2019). In February 2021, in a tomato plantation located in Rancho el Progreso (17.743639N, 99.243250W, 1554 masl), municipality of Zitlala, Guerrero, Mexico, the presence of rot of tomato fruits hybrid DRD-8551 (seed company, Seminis) was detected in tomatoes grown in greenhouses. Disease incidence was 8% in a sample of 250 plants. The infected fruits initially exhibited circular to irregular lesions, with a soft rot of infected tissue, which in advanced stages infected more than 50% of the fruit surface and a white to light brown mycelium cover was observed. It was observed that fruits were those that were closer to the soil were heavily affected. From infected fruits, cuts of approximately 0.5 cm were made, disinfested with 1% NaOCl and transferred to PDA medium, using this technique for all samples that were processed. Eight representative isolates were purified; developed colonies were initially light-white, after seven days they turned light brown. Right-angle hyphae were observed, with a slight constriction at the base of the branches, and produced dark brown sclerotia; morphological characteristics were consistent with those described for Rhizoctonia solani Kühn (Parmeter 1970). To confirm the anastomosis group (AG), the ITS region was amplified with the ITS1 and ITS4 primers (White et al. 1990) from the ESCJZ9 and ESCJZ10 strains. A search with BLAST tool indicated that both strains were 99.4% identity with R. solani AG-4 HG-I strain ICMP 20043 (GenBank: KM013470) (Ireland et al. 2015). Phylogenetic analysis compared with different sequences of AGs, allowed to group ESCJZ9 and ESCJZ10 strains with the clade AG-4 HG-I. The sequences were deposited in GenBank (MZ274468 for ESCJZ9 and, MZ274469 for ESCJZ10). Pathogenicity was confirmed by inoculating 15 healthy tomato fruits (hybrid DRD-8551), in which a portion of mycelium from the ESCJZ9 strain was inserted with a sterile toothpick in two points / fruit. In eight fruits only a sterile toothpick (two points / fruit) was inserted. The fruits were housed in laboratory conditions at average temperature of 27 °C. After five days, fruit rot symptoms were observed in all inoculated fruits, similar to those observed in the field. Control fruits remained asymptomatic. Additional pathogenicity tests were performed on 50 tomato plants (hybrid DRD-8551) of 15-day-old. Two 5 mm diameter mycelial discs from strain ESCJZ9 were deposited at the base of the stem 10 mm below the soil surface. As control treatments, sterile PDA was deposited in 20 plants. The plants were housed in a greenhouse at an average temperature and relative humidity of 28 ° C and 85%, respectively. After four days, all the inoculated plants showed root and stem rot and leaf flaccidity. Control plants remained asymptomatic. R. solani was re-isolated from the infected fruits and plants, fulfilling Koch's postulates. Previously, in Iran and Brasil R. solani AG-4 HG-I has been reported that caused stem and fruit rot on tomato (Rahimian 1988; Kuramae et al. 2003). In Mexico R. solani AG-4 has been reported in other solanaceae species such as potato and chili (Virgen-Calleros et al. 2000; Montero-Tavera et al. 2013). To our knowledge, this is the first report of R. solani AG-4 HG-I causing tomato fruit rot in Mexico. This research provides information in order to establish disease management strategies. It is important to establish measures to minimize the spread of this pathogen to other tomato-producing areas of this country.

Identification of Leaf Spot and Stem Rot on Plectranthus verticillatus Caused by Rhizoctonia solani AG-4 HGI in China.

Plectranthus verticillatus (Swedish ivy) is a plant in the family Lamiaceae (Labiatae), native to South-east Africa, that is commonly grown as a houseplant in China. In June 2018, the initial symptoms water-soaked spots were found on the stem base of sixteen P. verticillatus plants(1-year-old plants) in a commercial greenhouse, situating at Daxing district, Beijing, China (116°22'E, 39°42'N), where there were 150 potted P. verticillatus plants obtained by cuttage propagation and grown in conventional nutrient soils. Almost amonthlater, the disease was more severe and leaves appeared symptoms. Infected leaves displayed water-soaked, darkbrown lesions and fell off easily. Symptoms on the stems were dark brown, rotted and lodging. In the survey, the incidence of affected plants was 70%. To isolate the causal agent, the infected leaf and stem tissues were cut into 3 mm segments and surface disinfected in 1% NaClO for 1 min, then rinsed three times with sterile distilled water and cultured on potato dextrose agar (PDA) at 25±1 °C with a 12-h photoperiod. After 2 days, hyphal tips were transferred to new PDA plates to obtain pure cultures. Six morphologically similar isolates (CC1 to CC6) were obtained from the symptomatic samples. All the fungal colonies were white initially, turned brown gradually, and formed irregular shaped sclerotia after 9 days of incubation on PDA. Observationofmicrostructures indicated that all the isolates resembled Rhizoctonia sp.(Sneh et al. 1991), including septate hyphal with right-angled branching, a slight constriction at the base of hyphal branches and 3 to 8 nuclei per cell. Total genomic DNA was extracted from pure isolates using a Fungal DNA Kit (GBCBIO, Guangzhou, China) and the internal transcribed spacer (ITS) region of rDNA was amplified with primers ITS1/ITS4 and sequenced (White et al. 1990 and Garibaldi et al.2019).The results showed that sequences of 6 isolates were identical and had a 100% identity with those of R. solani AG-4 HGI (GenBank accession no. MH172679 and MH172664). The phylogenetic analysis performed by the neighbour-joining method (MEGA 7 software) showed that the representative isolate (accession no. MZ723940) was assigned to group ofR. solaniAG-4 HG-I. The pathogenicity of these six isolates to the leaves and stems of P. verticillatus were conducted. Healthy P. verticillatus plants(1-year-old) were grown in 2-liter pots with sterile substrate in a greenhouse at 14-19°C (night)/23-28°C (day). After 7 days, five P. verticillatus plantsfor each isolate were inoculated with a 6-mm-diameter PDA plug taken from a 10-day-old PDA culture, which was placed onto leaf blade and the base of the stem. Five control pots were inoculated with sterile PDA plugs. All the inoculated plants were incubated in a glass cabinet under 90% humidity and 25°C for 2 days and were then moved back to the greenhouse with natural daylight conditions. The test was repeated three times. Ten days later, all inoculated leaves and stems appeared dark brown and water-soaked lesions similar to those observed on naturally infected plants, whereas control plants remained asymptomatic. The fungus that was reisolated from diseased tissues had the same morphological characters as the original isolates and confirmed to beR. solaniAG-4 HG-I by molecular identification. R. solani AG-4 HGI has an extensive host range, which had previously been recorded to cause leaf, root and stem rot on Plectranthus sp. in Florida in 1984 (Alfieri Jr. et al. 1984). To our knowledge, this is the first report ofR. solani AG-4 HGI infecting P. verticillatus in China. Its confirmation is a significant step toward management recommendations for growers.

Molecular Mechanisms Associated with the Resistance of Rhizoctonia solani AG-4 Isolates to the Succinate Dehydrogenase Inhibitor Thifluzamide

Thifluzamide, a succinate dehydrogenase (SDH) inhibitor, possesses high activity against Rhizoctonia. In this study, 144 Rhizoctonia solani AG-4 (4HGI, 4HGII, and 4HGIII) isolates, the predominate pathogen associated with sugar beet seedling damping-off, were demonstrated to be sensitive to thifluzamide with a calculated mean median effective concentration of 0.0682 ± 0.0025 μg/ml. Thifluzamide-resistant isolates were generated using fungicide-amended media, resulting in four AG-4HGI isolates and eight AG-4HGII isolates with stable resistance and almost no loss in fitness. Evaluation of cross-resistance of the 12 thifluzamide-resistant isolates and their corresponding parental-sensitive isolates revealed a moderately positive correlation between thifluzamide resistance and the level of resistance to eight other fungicides from three groups, the exception being fludioxonil. An active efflux of fungicide through ATP-binding cassette and major facilitator superfamily transporters was found to be correlated to the resistance of R. solani AG-4HGII isolates to thifluzamide based on RNA-sequencing and quantitative reverse transcription-PCR analyses. Sequence analysis of sdhA, sdhB, sdhC, and sdhD revealed replacement of isoleucine by phenylalanine at position 61 in SDHC in 9 of the 12 generated thifluzamide-resistant isolates. No other mutations were found in any of the other genes. Collectively, the data indicate that the active efflux of fungicide and a point mutation in sdhC may contribute to the resistance of R. solani AG-4HGI and AG-4HGII isolates to thifluzamide invitro. This is the first characterization of the potential molecular mechanism associated with the resistance of R. solani AG-4 isolates to thifluzamide and provides practical guidance for the use of this fungicide.

First Report of Multinucleate Rhizoctonia solani AG4 HG-I Causing Crown and Root Rot on Strawberry in Italy

First Report of Multinucleate <i>Rhizoctonia solani</i> AG4 HG-I Causing Crown and Root Rot on Strawberry in Italy

Biocontrol of Large Patch Disease in Zoysiagrass (Zoysia japonica) by Bacillus subtilis SA-15: Identification of Active Compounds and Synergism with a Fungicide

Bacillus subtilis SA-15 is a plant growth-promoting bacterium isolated from non-farming soil. We aimed to identify lipopeptides produced by B. subtilis SA-15 and evaluate the control efficacy of B. subtilis SA-15 against large patch disease caused by Rhizoctonia solani AG 2-2 (IV) in zoysiagrass (Zoysia japonica). Bacillus subtilis SA-15 inhibited mycelial growth of R. solani AG 2-2 (IV) in vitro and produced fengycin A and dehydroxyfengycin A, which are antifungal compounds. Fengycin A and deghydroxyfengycin A inhibited R. solani mycelial growth by 30.4 and 63.2%, respectively. We formulated B. subtilis SA-15 into a wettable powder and determined its control efficiency against large patch in a field trial. The control efficacy was 51.2–92.0%. Moreover, when B. subtilis SA-15 powder was applied together with half the regular dose of the fungicide pecycuron, the control efficacy was 88.5–100.0%. These results indicate that B. subtilis SA-15 can be used to control soil-borne diseases, including large patch caused by R. solani, because of lipopeptide production. The use of this bacterium can also reduce the amount of fungicide needed, providing an eco-friendly management option for turfgrass.

Cloning and sequence analysis of a serine protease gene from Rhizoctonia solani Kühn AG5.

The present study aimed to identify the subtilisin-like proteases (SLPs) of Rhizoctonia solani Kühn potentially involved in the virulence of this phytopathogenic fungus, which has 14 anastomosis groups (AGs) responsible for many crop diseases. Through mycelial microscope observation and strain identification of pathogenic fungus MS-3, it was determined to be R. solani AG-5. Both 5' and 3' rapid amplification of cDNA ends were used to clone the serine protease gene RsSLP from R. solani AG-5. The full-length obtained for RsSLP was 1714bp with an open reading frame of 1587bp, encoding a protein of 528 amino acids with a molecular mass of 55.8kDa. This protein contained a predicted signal peptide for secretion but lacked a transmembrane domain or membrane anchor site. Bioinformatics analysis identified this protein as a serine protease with the Peptidase_S8 and Inhibitor_I9 characteristic domains of SLPs. Phylogenetic analysis suggested that frequent gene duplications of the SLPs occurred in R. solani (RsSLP), and RsSLP shares characteristic sequence features with virulence factors of other phytopathogenic fungi. Because the secretory serine protease RsSLP from R. solani AG5 is similar to the virulence factors of other phytopathogenic fungi, its identification will be helpful in studies considering the roles of these proteases in pathogen virulence.

Improving the Genome Annotation of Rhizoctonia solani Using Proteogenomics

Background Rhizoctonia solani is a pathogenic fungus that causes serious diseases in many crops, including rice, wheat, and soybeans. In crop production, it is very important to understand the pathogenicity of this fungus, which is still elusive. It might be helpful to comprehensively understand its genomic information using different genome annotation strategies.MethodsAiming to improve the genome annotation of R. solani, we performed a proteogenomic study based on the existing data. Based on our study, a total of 1060 newly identified genes, 36 revised genes, 139 single amino acid variants (SAAVs), 8 alternative splicing genes, and diverse post-translational modifications (PTMs) events were identified in R. solani AG3. Further functional annotation on these 1060 newly identified genes was performed through homology analysis with its 5 closest relative fungi.ResultsBased on this, 2 novel candidate pathogenic genes, which might be associated with pathogen-host interaction, were discovered. In addition, in order to increase the reliability and novelty of the newly identified genes in R. solani AG3, 1060 newly identified genes were compared with the newly published available R. solani genome sequences of AG1, AG2, AG4, AG5, AG6, and AG8. There are 490 homologous sequences. We combined the proteogenomic results with the genome alignment results and finally identified 570 novel genes in R. solani.ConclusionThese findings extended R. solani genome annotation and provided a wealth of resources for research on R. solani.

Genome Organizations and Functional Analyses of a Novel Gammapartitivirus from Rhizoctonia solani AG-1 IA Strain D122.

Here, we describe a novel double-stranded (ds) RNA mycovirus designated Rhizoctonia solani dsRNA virus 5 (RsRV5) from strain D122 of Rhizoctonia solani AG-1 IA, the causal agent of rice sheath blight. The RsRV5 genome consists of two segments of dsRNA (dsRNA-1, 1894 bp and dsRNA-2, 1755 bp), each possessing a single open reading frame (ORF). Sequence alignments and phylogenetic analyses showed that RsRV5 is a new member of the genus Gammapartitivirus in the family Partitiviridae. Transmission electron microscope (TEM) images revealed that RsRV5 has isometric viral particles with a diameter of approximately 20 nm. The mycovirus RsRV5 was successfully removed from strain D122 by using the protoplast regeneration technique, thus resulting in derivative isogenic RsRV5-cured strain D122-P being obtained. RsRV5-cured strain D122-P possessed the traits of accelerated mycelial growth rate, increased sclerotia production and enhanced pathogenicity to rice leaves compared with wild type RsRV5-infection strain D122. Transcriptome analysis showed that three genes were differentially expressed between two isogenic strains, D122 and D122-P. These findings provided new insights into the molecular mechanism of the interaction between RsRV5 and its host, D122 of R. solani AG-1 IA.

First Report of Rhizoctonia solani AG2-1 on roots of wheat in Kazakhstan.

In June 2019, approximately 20 tillers of wheat (Triticum aestivum L.) were sampled at the ripening stage (Feekes scale 11) from four different fields in Almaty, Kazakhstan. Brown lesions (3-5 mm in length) were present on the roots of sampled plants, with 20% incidence. To determine the causal agent, diseased roots were surface disinfected in sodium hypochlorite solution (1%) for 3 min, rinsed triple with sterile distilled water, air-dried in a laminar flow hood, and plated onto one-fifth strength potato dextrose agar (PDA) supplemented with 50 ppm chloramphenicol. After three days, the hyphal fragments that developed from the sections were transferred to fresh PDA and incubated at 23°C with 12-h photoperiod for 7 days to obtain pure cultures. Brown pigmented fungal colonies with a constriction at the base of hyphal branches, septa near the branching point, and right-angled branching resembling Rhizoctonia solani were observed. The identification anastomosis group (AG) of a representative isolate for each field was conducted by sequencing the internal transcribed spacer (ITS) region of rDNA with the universal primers ITS4 and ITS5 (White et al. 1990). The resulting sequences of 693 bp length were deposited in GenBank (accession nos. MW898143:MW898146). These sequences were 100% identical to the isolate 8Rs of R. solani AG2-1 (accession no. AF354063). To confirm the pathogenicity of the four isolates, the colonized wheat kernels method described by Demirci (1998) was used to inoculate a sterile potting mix containing peat, vermiculite, and soil (1:1:1 by v/v/v) into which wheat (cv. Seri) was planted. Control pots were inoculated with sterile wheat kernels using the same procedure. Wheat plants were left to grow for four weeks under controlled environmental conditions with a 23°C temperature regime. During the period that the plants remained in the glasshouse, the typical light regime was 16 h. Brown lesions were observed on the roots of plants in the inoculated pots whereas no symptoms were observed on plants grown in the control pots. R. solani was consistently reisolated from symptomatic plants, thereby confirming Koch's postulates. To our knowledge, this is the first report of R. solani AG2-1 on roots of wheat in Kazakhstan. R. solani AG2-1 isolates have been previously reported to be a weak pathogen to wheat (Roberts and Sivasithamparam 1986; Sturrock et al. 2015; Jaaffar et al. 2016; Özer et al. 2019). We suggest further studies are required to characterize the impact of R. solani AG2-1 in wheat. Considering crop rotation, the selection of non-host crops to this AG group is important to pathogen management, by reducing the amount of inoculum in the soil.

Effect of Rhizoctonia spp. on Polyphenol Oxidase Enzyme Activity in Alfalfa Seedling

Soil borne diseases cause significant losses on quantity and quality of many crop species annually. Rhizoctonia is a widespread and ecologically diverse soilborne fungus causing different types of diseases in many plant species including alfalfa (Medicago sativa). Rhizoctonia species have been traditionally identified based on the cell nuclear condition. Polyphenol oxidases (PPOs) are ubiquitous copper-containing enzymes that are widely occurring enzymes among plants. PPOs are involved in the oxidation of polyphenols into quinones (antimicrobial compounds) and lignification of plant cells that contribute to the formation of defense barriers against pathogens. The study was conducted with the aim to determine the effect of indigenous isolates of a multinucleate (Rhizoctonia solani AG-4) and nineteen isolates of binucleate Rhizoctonia on PPO activity in alfalfa (cv. Gea) seedling under in vitro conditions. The activity of PPO enzyme was determined in inoculated and uninoculated control alfalfa plants after ten days from inoculation. There was a significant increase in the activity of PPO after treatment of alfalfa seedling with isolates of Rhizoctonia. Among Rhizoctonia isolates, highest induction of PPO activity was recorded with pathogenic R. solani AG-4. In present study, increased amounts of PPO were also observed in plants that were challenged with Rhizoctonia spp. PPO has a role in catalyzing phenolic oxidation in limiting disease development. PPO may therefore be involved in induction of defense resistance against plant diseases.

The role of biological and chemical inducers in activating bean defense responses against Rhizoctonia solani

Rhizoctonia root rot is an important bean disease that causes significant yield losses. Increasing the levels of plant resistance using biological and chemical inducers is a sustainable strategy for plant disease management. In this study, the efficiency of thiamine, riboflavin, pyridoxine, folic acid, menadione, quercetin, naringenin, humic acid, fulvic acid, boric acid, zinc sulfate, manganese sulfate, Piriformospora indica and binucleate Rhizoctonia were evaluated to improve bean resistance to root rot pathogen Rhizoctonia solani AG-4 HGII. Under laboratory conditions, effect of various biological agents and chemical compounds on mycelial growth of the pathogen was evaluated. Results showed that applying boric acid, zinc sulfate and manganese sulfate at 1000 and 2000 mg L−1 concentrations and menadione at 2000, 5000, 10000 and 20000 μM completely inhibited the pathogen growth and 10000 and 20000 μM concentrations of thiamine reduced linear growth of R. solani. Under greenhouse conditions, application of the inducers significantly reduced the disease progress on treated plants compared to the non-treated plants. The highest level of disease reduction was observed using zinc sulfate and thiamine applied at 3 days before the pathogen inoculation, and also by using P. indica 14 days before inoculation with the pathogen. Thiamine, zinc sulfate and P. indica significantly reduced the disease index and improved plant growth parameters compared to the controls. Induction of resistance against R. solani by individual and combined treatments using thiamine, zinc sulfate and P. indica was associated with increased activity of various antioxidants and increased levels of membrane stability index (MSI), relative water content (RWC), lignin accumulation and total soluble phenolics in bean seedlings. Our findings revealed a practical possibility of inducing resistance and protection of bean plants against Rhizoctonia root rot with the help of biological and chemical resistance inducers.