Solani AG Research Articles

Horizontal and Vertical Transmission of a Mycovirus Closely Related to the Partitivirus RhsV717 That Confers Hypovirulence in Rhizoctonia solani.

Rhizoctonia solani virus717 (RhsV717) was isolated from the Rhizoctonia solani (R. solani) AG-2 strain Rhs717. This study isolated a virus designated as Rhizoctonia solani partitivirus BS-5 (RsPV-BS5) from the R. solani AG-3 strain BS-5, the causal agent of tobacco target spot disease. The virus was identified as a strain of RhsV717. Transmission electron microscopy (TEM) images showed that RsPV-BS5 had virus particles with a diameter of approximately 40 nm. Importantly, it can be horizontally transmitted through hyphal anastomosis and vertically transmitted via sexual basidiospores. Furthermore, this study demonstrated that RsPV-BS5 infection significantly impedes mycelial growth and induces hypovirulence in tobacco leaves. Thus, RsPV-BS5 presents a promising avenue for biocontrolling tobacco target spot disease. Transcriptome analysis unveiled differential expression of four genes related to cell wall-degrading enzymes between two isogenic strains, 06-2-15V and 06-2-15. These findings shed light on the molecular mechanism through which RsPV-BS5 reduces host pathogenicity.

First report of Rhizoctonia solani AG-1 IA causing rice sheath blight of Oryza rufipogon in China.

Red rice (Oryza rufipogon Griff.) is a valuable source of important agronomic traits as well as genes for biotic and abiotic stress tolerance. In June 2020, rice sheath blight on O. rufipogon cv. Bin09 was observed in Zhanjiang (20.93N, 109.79E), China. Initial symptoms on sheaths were water-soaked and light green lesions. Then, the lesions gradually expanded into oval or cloud shaped lesions with a gray white center. The lesions coalesced, causing the entire sheath to become blighted. Disease incidence reached approximately 30% in the fields (10 ha) surveyed. Twenty sheaths with symptoms were collected and cut into pieces of 2 × 2 cm in size. They were surface-disinfected with 75% ethanol for 30 s and 2% sodium hypochlorite (NaOCl) for 60 s, rinsed three times with sterile water, blotted dry on sterile paper, plated on potato dextrose agar (PDA), and incubated at 28°C in the dark for 4 days. Thirty-six pure cultures were obtained by transferring hyphal tips to new PDA plates, and three isolates (ORRS-1, ORRS-2, and ORRS-3) with similar morphological characteristics on PDA were selected as the representative isolates for study. Colony of isolate ORRS-1 was white initially, then turned brown with brown sclerotia. Septate hyphae were hyaline, smooth, and branched at right angles with a septum near the point of branching. Based on these morphological characteristics, the fungus was identified asRhizoctonia solani Kuhn (Sneh et al. 1991). The isolates were deposited in the fungus collection of the Aquatic Organisms Museum of Guangdong Ocean University. For molecular identification, genomic DNA from each of the three isolates was extracted, and the internal transcribed spacer (ITS) region was amplified, and sequenced with the primer pair ITS5/ITS4 (White et al. 1990). The sequences were deposited in GenBank (accession nos. OP497977 to OP497979). The three isolates were 100% identical (716/716 bp; 716/716 bp; and 716/716 bp) with those of R. solani AG-1 IA (accession nos. KX674518, MK481078, and MK480532) through BLAST analysis.The phylogenetic tree grouped the three isolates within theR.solaniAG-1 IA clade with high bootstrap support (99%) by the maximum likelihood method. A pathogenicity test was performed with these three isolates in a greenhouse at 24 to 30°C. Approximately 50 seedling of red rice cv. Bin09 were grown in each cup ( 250 ml in size with sterilesoil 50 cm3). At the 3-leaf stage, plants in five cups were inoculated with each isolate by spraying a mycelial suspension (106 mycelial fragments/ml) until runoff. The mycelial suspension was prepared by adding sterile distilled water to the cultures and gently scraping the surface with a sterilized scalpel blade. Five plants sprayed with sterile water served as the controls. The test was conducted three times. Sheath blight was observed on the inoculated leaves after 15 days while no disease was observed in the control plants. Morphological characteristics and the ITS sequences of fungal isolates re-isolated from the diseased sheaths were identical to those of R.solaniAG-1 IA. R.solani AG-1 IA is one of the most important plant pathogens worldwide, causing foliar diseases on maize, rice (O. sativa L.), and soybean (Joana et al. 2009). To our knowledge, this is the first report of R.solani AG-1 IA causing rice sheath blight of O. rufipogon in China (Farr and Rossman, 2022). With the spread of the pathogen on weedy populations of red rice, resistant races or pathotypes may evolve that could spread to cultivated rice.

Van’ın Erciş, Gevaş ve Edremit ilçelerinde Biber, Domates ve Kavundan Fusarium spp. ve Rhizoctonia spp.’nin Teşhisi ve Patojeniteleri

Van’ın Erciş, Gevaş ve Edremit ilçelerinde yetiştirilen domates, biber ve kavun bitkilerinin köklerinden izole edilen Rhizoctonia ve Fusarium türlerini teşhis etmek ve patojenitelerini belirlemek bu çalışmanın amacını oluşturmuştur. Sürvey çalışmaları 2018 yılında yapılmış ve domates örneklerinden 6 Rhizoctonia, 31 Fusarium, biber örneklerinden 4 Rhizoctonia, 6 Fusarium, kavun örneklerinden 10 Fusarium olmak üzere toplam 57 fungal izolat elde edilmiştir. Fusarium ve Rhizoctonia izolatlarının tür teşhisleri klasik yöntemlerle yapılmış, Rhizoctonia anastomosis grupları (AG) test izolatları ile eşleştirilerek belirlenmiştir. Buna göre Rhizoctonia izolatları; binükleik AG-K (domatesten 2 izolat), Rhizoctonia solani AG-4 (domatesten 2 izolat), R. solani AG-2 (domatesten 2, biberden 1 izolat), R. solani AG-3 (biberden 2 izolat) ve R. solani AG-5 (biberden 1 izolat) olarak belirlenmiştir. Fusarium izolatları ise; F. oxyporum (kavundan 6, biberden 3, domatesten 18 izolat), F. solani (kavundan 2, biberden 3, domatesten 13 izolat) ve F. equiseti (kavundan 2 izolat) olarak tespit edilmiştir. Patojenite denemesinde sayılarının az olması nedeniyle izole edilen bütün Rhizoctonia izolatları ile bölgeleri temsil edecek şekilde seçilen 14 Fusarium izolatı kullanılmıştır. Deneme sonucunda Fusarium izolatlarında biberde F. solani' nin, domates ve kavunlarda F. oxysporum’ un daha patojen oldukları belirlenmiştir. Rhizoctonia türlerinde ise biberlerde R. solani AG-3' ün, domateslerde R. solani AG-4 ve binükleik Rhizoctonia AG K’ nın daha patojen olduğu bulunmuştur. Sonuç olarak biberden R. solani AG-5 ve R. solani AG-3, domates ve biberden R. solani AG-2, domatesten binükleik Rhizoctonia AG-K Van' da ilk defa bu çalışma ile izole edilmiş ve anastomosis grupları belirlenmiştir.

First Report of Target Spot Caused by Rhizoctonia solani AG-5 on Tobacco in China.

Tobacco is one of the vital economic crops in China. Nevertheless, tobacco diseases cause substantial economic losses each year. Tobacco target spot is a fungal disease which commonly found on the leaves. While both sexual and asexual reproduction can occur, asexual reproduction is much more common in tobacco. In June 2022, target spot was found on tobacco leaf samples from Yibin, Sichuan Province and Meitan, Guizhou Province, China. The typical symptoms were light brown tissue with concentric ring marks, and the necrotic part of the disease spot was fragile and forming perforation after falling off. The diseased tissue in the sample was cut off and sterilized in 75% ethanol for 1 min, and rinsed three times in sterilized distilled water. Finally, the tissues were placed on potato glucose agar (PDA) medium with kanamycin (0.1 mg/mL). After incubation at 28 °C in darkness for 3 days,the culture of the isolate grew in the form of radial mycelium on PDA dishes, the mycelium was white initially, turned brown generally at the later stage, and finally thickened and separated with the growth of the culture. Nine pathogenic strains were isolated, including four isolates from Yibin and five from Meitan. They were all used for pathogen identification. Genomic DNA of each isolate was extracted using the CATB method, and PCR analysis was performed with primers specifically designed to detect individual fusion groups or fusion subgroups of solani: AG-1 IA, IB, and IC; AG-3 PT; AG-4 HG-I, HG-II and HG-III; AGs-5-6 and P-21-22. Among the 11 specific primer pairs, only AG-5-specific primer amplified the fungal DNA, indicating that the nine isolates tested all belonged to the R. solani AG-5 fusion group. BLASTn search was performed on the gene sequences obtained from these strains and they deposited in GenBank under accession no. OP647851-OP647859. These gene sequences were aligned with the voucher specimen R. solani AG-5, with more than 99% similarity . The nine isolates were then tested for mycelial anastomosis reactions using the R. solani AG-5 standard strain following the method described by Ogoshi (1987). A decrease in the diameter of the mycelia at the anastomosis site and death of adjacent cells were observed, indicating their anastomosis response. Therefore, these nine strains were identified as R. solani AG-5 based on morphological and genetic analysis. Subsequently, one pathogenic strain from Meitan and another one from Yibin were selected for pathogenicity verification. Mycelial PDA blocks (6 mm in diameter) of the two isolates were inoculated on healthy tobacco plants, while leaves containing only PDA blocks were used as controls. A total of 6 replicates were conducted. After inoculation, they were incubated at 85% relative humidity and 15 to 25 °C. Koch's hypothesis was confirmed by reisolating pathogens from diseased leaves 5 days after inoculation. Typical symptoms were observed on tobacco plants inoculated with the pathogen strains but not on control tobacco plants. To the best of our knowledge, tobacco target spot has been reported caused by R. solani AG-3, AG-6 and AG-2.1 groups in the field in China and in Argentina. Up until now, this is the first report of R. solani AG-5 causing tobacco target spot on tobacco in the field in China. It was also found to be highly virulent to chickpea in Turkey. Due to serious damages caused by this disease in the last five years in China, more attention should be paid in disease control measures to avoid economic losses. In addition, it also provides some theoretical help for the damage caused by this pathogen on other hosts and helps people to better understand Rhizoctonia solani AG-5.

Molecular Characterization of Defense of Brassica napus (Oilseed Rape) to Rhizoctonia solani AG2-1 Confirmed by Functional Analysis in Arabidopsis thaliana.

Rhizoctonia solani is a necrotrophic, soilborne fungal pathogen associated with significant establishment losses in Brassica napus (oilseed rape; OSR). The anastomosis group (AG) 2-1 of R. solani is the most virulent to OSR, causing damping-off, root and hypocotyl rot, and seedling death. Resistance to R. solani AG2-1 in OSR has not been identified, and the regulation of OSR defense to its adapted pathogen, AG2-1, has not been investigated. In this work, we used confocal microscopy to visualize the progress of infection by sclerotia of AG2-1 on B. napus varieties with contrasting disease phenotypes. We defined their defense response using gene expression studies and functional analysis with Arabidopsis thaliana mutants. Our results showed existing variation in susceptibility to AG2-1 and plant growth between OSR varieties, and differential expression of genes of hormonal and defense pathways related to auxin, ethylene, jasmonic acid, abscisic acid, salicylic acid, and reactive oxygen species regulation. Auxin, abscisic acid signaling, and the MYC2 branch of jasmonate signaling contributed to the susceptibility to AG2-1, while induced systemic resistance was enhanced by NAPDH RBOHD, ethylene signaling, and the ERF/PDF branch of jasmonate signaling. These results pave the way for future research, which will lead to the development of Brassica crops that are more resistant to AG2-1 of R. solani and reduce dependence on chemical control options.

Rhizoctonia solani AG1 IA extracellular polysaccharides: Structural characterization and induced resistance to rice sheath blight

Sheath blight, caused by Rhizoctonia solani (R. solani), is one of the most serious diseases of rice. Extracellular polysaccharides (EPS) are complex polysaccharides secreted by microbes that have a pivotal role in the plant-microbe interaction. At present, many studies have been carried out on R. solani, but it is not very clear whether the EPS is secreted by R. solani exists. Therefore, we isolated and extracted the EPS from R. solani, two kinds of EPS (EW-I and ES-I) were obtained by DEAE-cellulose 52 and Sephacryl S-300HR column further purification, and their structures were characterized by FT-IR, GC-MS, and NMR analysis. The results showed that EW-I and ES-I had similar monosaccharide composition but different molar ratio, they were composed of fucose, arabinose, galactose, glucose, and mannose with a ratio of 7.49: 27.72: 2.98: 6.66: 55.15 and 3.81: 12.98: 6.15: 10.83: 66.23, and their backbone may be composed of →2)-α-Manp-(1→ residues, beside ES-I was highly branched compared to EW-I. The exogenous application of EW-I and ES-I had no effect on the growth of R. solani AG1 IA itself, but their pretreatment of rice induced plant defense through activation of the salicylic acid pathway, resulting in enhanced resistance to sheath blight.

Preservation of Food Sugar Beet via the Control of Rhizoctonia solani AG 2-2IIIB by Extreme Factors

Sugar beet (Beta vulgaris L.) is a valuable sugar-providing crop accounting for around 25% of global white sugar production. It is an integral part of the food industry. Several factors cause a decrease in the yield of this food crop. The infection of phytopathogenic fungi is one of the most hazardous. Though several methods to control Rhizoctonia solani have been developed, the problem is still not solved. Thus, the goal of this work was to investigate the effect of abiotic factors (pH, Eh, presence or absence of O2) and biotic ones (microbial community) on the fungal pathogen R. solani AG 2-2IIIB for its control and sugar beet protection. Microbiological methods were used to conduct the research. Thermodynamic calculations were applied to determine the conditions necessary for suppression of R. solani AG 2-2IIIB. As a result, a comparison of the effect of abiotic and biotic mediated factors on the efficiency of the growth of Rhizoctonia solani AG 2-2IIIB was carried out. Obtained data showed the following: mainly the abiotic factors provided via chemical buffers or removal of oxygen caused the inhibition of R. solani; the values of 5.5 ≤ pH ≤ 8.5, Eh ≤ −200 mV, and oxygen absence inhibited its growth. The maintenance of these factors via microbial communities enhanced the suppressive effect, bringing about the death of mycelium. The study of microbial strains revealed the inhibitory effect of microbial metabolites and provided more effective control of Rhizoctonia solani AG 2-2IIIB.

Endophytic fungi-based wettable powder formulation for Rhizoctonia solani AG-4 biocontrol

Endophytic fungi-based wettable powder formulation for Rhizoctonia solani AG-4 biocontrol

Cover: Journal of Basic Microbiology. 2/2023

Journal of Basic MicrobiologyVolume 63, Issue 2 p. 117-117 COVER PICTUREFree Access Cover: Journal of Basic Microbiology. 2/2023 First published: 01 February 2023 https://doi.org/10.1002/jobm.202370003AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat Graphical Abstract Cover illustration: Rhizoctonia solani AG-3 TB strain M2J-22 formed a complex and beautiful colony pattern when grown on a Potato Dextrose Agar plate at 28°C for 3 days. (Photo: Xinru Liu, State Key Laboratory for Protection and Utilization of Bio-Resources in Yunnan, Yunnan Agricultural University, Kunming, Yunnan, China) Volume63, Issue2February 2023Pages 117-117 RelatedInformation

Novel QTL associated with Rhizoctonia solani Kühn resistance identified in two table beet × sugar beet F2:3 populations using a new table beet reference genome

AbstractThe necrotrophic fungus Rhizoctonia solani Kühn is a major concern for table beet (Beta vulgaris L. ssp. vulgaris) producers across the United States causing upward of 75% losses in severe instances. Thus far, there have been minimal efforts to incorporate host resistance to R. solani in table beet germplasm. To investigate the genetic control of R. solani resistance in table beet, we developed two mapping populations. Parents of the two populations were a Rhizoctonia‐susceptible table beet inbred W357A and a resistant sugar beet germplasm FC709‐2 (sugar beet resistance population, SBRP) and a Rhizoctonia‐resistant table beet inbred W364B and a susceptible sugar beet inbred FC901/C817 (table beet resistance population, TBRP). In Spring 2020, F2:3 families were evaluated for response to artificial inoculation with R. solani AG 2‐2 IIIB isolate R1 in replicated greenhouse experiments. This work also represents the first use of the W357B table beet reference genome, utilized here to align genotyping‐by‐sequencing reads to identify polymorphic markers. Using interval linkage mapping, we identified one quantitative trait locus (QTL) in each of the two populations, each accounting for 30% of the phenotypic variation. The QTL in both the SBRP and TBRP were found on chromosome 2 and contained several putative resistance genes in annotations of the Beta vulgaris and Arabidopsis thaliana genomes. This is the first report of a QTL on chromosome 2 for resistance to R. solani in B. vulgaris ssp. vulgaris and the first identification of QTL for disease resistance in table beet. The newly developed table beet reference genome and markers identified in this study may be of value for marker‐assisted selection in breeding for resistance to R. solani in both sugar beet and table beet breeding programs.

Identification and characterization of pathogenicity-related genes of Rhizoctonia solani AG3 during tobacco infection.

Tobacco target spot disease is caused by a ubiquitous soil-borne phytopathogen Rhizoctonia solani; the pathogenic mechanisms underlying the effects of R. solani remain unclear. Deeper understanding of the functional responses to R. solani during host plant infection would help identify the molecular mechanisms essential for successful host invasion. In this study, we performed global transcriptional analysis of R. solani during various stages (12, 24, 48, 72, 96, and 120h) of tobacco infection via an RNA sequencing method, while utilizing the pathosystem model R. solani AG3-tobacco (Nicotiana tabacum L.). After R. solani inoculation, the number of differentially expressed genes of R. solani differed at the various time points. Moreover, several gene ontology and Kyoto encyclopedia of genes and genomes pathways were unique in different infection stages, especially with respect to the genes involved in plant cell wall degradation and catalysis of biotransformation reactions, such as the pectin metabolic process and pectin catabolic process. The overexpressing-PD8 N. benthamiana plants enhanced the susceptibility to R. solani. In addition, we found that large amounts of reactive oxygen species (ROS) were generated in tobacco after infected by R. solani. R. solani encoding FAD/NAD binding oxidoreductase and peroxidase gene family to eliminating ROS and counteract oxidative stress. Moreover, Perox3 was validated that can enhance the ability of scavenging ROS by co-injecting. Overall, our findings show that pectin-degrading enzymes and cytochrome P450 genes are critical for plant infection. These results provide comprehensive insights into R. solani AG3 transcriptome responses during tobacco invasion.

Identification of 3-Methoxyphenylacetic Acid as a Phytotoxin, Produced by Rhizoctonia solani AG-3 TB

Tobacco target spot disease is caused by Rhizoctonia solani AG-3 TB, which causes serious harm to the quality and yield of tobacco. In this study, thin layer chromatography (TLC), high performance liquid chromatography (HPLC), infrared absorption spectroscopy (IR), and nuclear magnetic resonance spectroscopy (NMR) were used to purify and identify the potential phytotoxin produced by R. solani AG-3 TB. The result indicated that the purified toxin compound was 3-methoxyphenylacetic acid (3-MOPAA) (molecular formula: C9H10O3). The exogenous purified compound 3-MOPAA was tested, and the results revealed that 3-MOPAA can cause necrosis in tobacco leaves. 3-MOPAA is a derivative of phenylacetic acid (PAA), which should be produced by specific enzymes, such as hydroxylase or methylase, in the presence of PAA. These results enrich the research on the pathogenic phytotoxins of R. solani and provide valuable insights into the pathogenic mechanism of AG-3 TB.

Biocontrol of the causal brown patch pathogen Rhizoctonia solani by Bacillus velezensis GH1-13 and development of a bacterial strain specific detection method.

Brown patch caused by the basidiomycete fungus Rhizoctonia solani is an economically important disease of cool-season turfgrasses. In order to manage the disease, different types of fungicides have been applied, but the negative impact of fungicides on the environment continues to rise. In this study, the beneficial bacteria Bacillus velezensis GH1-13 was characterized as a potential biocontrol agent to manage brown patch disease. The strain GH1-13 strongly inhibited the mycelial growth of turf pathogens including different anastomosis groups of R. solani causing brown patch and large patch. R. solani AG2-2(IIIB) hyphae were morphologically changed, and fungal cell death resulted from exposure to the strain GH1-13. In addition, the compatibility of fungicides with the bacterial strain, and the combined application of fungicide azoxystrobin and the strain in brown patch control on creeping bentgrass indicated that the strain could serve as a biocontrol agent. To develop strain-specific detection method, two unique genes from chromosome and plasmid of GH1-13 were found using pan-genome analysis of 364 Bacillus strains. The unique gene from chromosome was successfully detected using both SYBR Green and TaqMan qPCR methods in bacterial DNA or soil DNA samples. This study suggests that application of GH1-13 offers an environmentally friendly approach via reducing fungicide application rates. Furthermore, the developed pipeline of strain-specific detection method could be a useful tool for detecting and studying the dynamics of specific biocontrol agents.

First report of Rhizoctonia solani AG 4 causing brown bean of lima bean in Delaware.

Lima bean production has been an economically valuable staple in Delaware agriculture for almost a century, with annual revenue approaching 8 million dollars (USDA-NASS, 2019; Evans et al. 2007). From 2019-2021, lima beans displaying symptoms of brown discoloration, referred to as "brown bean" were observed in the green baby lima variety 'Cypress' across multiple commercial and research fields. Symptoms were present in approximately 1-5% of beans and not visible until pods were opened for harvest. Thirty-seven symptomatic beans were collected and surface disinfested in 0.85% sodium hypochlorite for 30 s, rinsed in sterile deionized water for 30 s, sectioned into four pieces and plated onto potato dextrose agar (PDA) amended with 50 µg/ml penicillin G and streptomycin sulfate. Petri dishes were incubated at 23ºC and observed for colony morphology. Pure cultures were obtained with tan colonies that had mycelia with right angle branching and septations near the branch, consistent with the description of Rhizoctonia solani Kuhn (Sneh et al. 1991). DNA extraction and pathogen identification was confirmed by sequencing of the internal transcribed spacer (ITS) region of nuclear ribosomal DNA using primers ITS4/ITS5 (White et al. 1990) for thirty-seven isolates collected in 2019 and 2020. Isolates were identified as Rhizoctonia solani AG 4 (99.9% sequence identity with GenBank Accession [MN106359.1].) A representative isolate was selected to complete Koch's postulates and the sequence was deposited in GenBank as accession number MW560551. To observe colonization ability, 10 detached pods were sterilized in 75% EtOH for 60 s, then rinsed in Milli-Q water. The detached pods were divided among two 150 mm Petri dishes containing a single 150 mm filter paper saturated with Milli-Q water. Five 1 mm2 agar plugs colonized with the representative R. solani isolate were placed 0.5 cm apart along the length of the pod. Plates were sealed with parafilm and left at room temperature. Control pods were kept in identical conditions but inoculated using clean agar plugs. The trial was repeated and a second trial was conducted on 12 attached asymptomatic pods from C-Elite Select lima bean plants at the succulent seed stage to complete Koch's Postulates. Pods were surface disinfested with 70% ethanol. Three attached pods were wounded with the tip of a sterile scalpel blade where a colonized agar plug was placed and loosely wrapped with a thin parafilm layer to maintain contact. Three attached pods not wounded were also inoculated with a colonized agar plug and wrapped by parafilm. Three wounded and non-wounded pods received clean agar plug controls. Both attached and detached pods were kept at room temperature for one week until symptoms began to manifest on the pod surfaces, at which point the beans from infected pods were removed and placed on PDA, three to a plate. In the attached assay, all beans of both wounded and non-wounded pods developed symptoms. The plates were stored in identical conditions and monitored for 5 days until tan colonies were observed. Culture morphology was consistent with the original isolate in all beans. Sequencing of the ITS region confirmed identity as R. solani AG-4. No symptoms were observed on control pods or seeds. Rhizoctonia solani is most frequently associated with symptoms of root rot (Sharma-Poudyal et al. 2015), but no stem symptoms are associated with the late season "brown bean" that has been observed throughout production in recent years. To our knowledge, this is the first report of Rhizoctonia solani AG 4 causing symptoms of brown bean of lima bean in Delaware. In preliminary observations, symptoms seem to be worse in pods that could have had contact with the soil directly or via rain splash. This disease cannot be detected until pods are split open, which has potential to reduce lima bean quality at harvest. Further monitoring should be conducted to quantify yield impacts and develop appropriate preventative and curative techniques.

Chemical seed treatments for the control of cotton seedling damping-off caused by Rhizoctonia solani under greenhouse conditions

Abstract: The aim of this study was to determine the effectiveness of cotton seed treatment with fungicides in the control of seedling damping-off caused by Rhizoctonia solani, under greenhouse conditions. Treated and untreated delinted seeds of the cultivar DeltaOpal were sowed in sand contained in plastic trays, placed in individual and equidistant wells, 3 cm deep. Into each plastic tray were put 3.0 g of the pathogen inoculum. The inoculation of R. solani AG-4 was done by the homogeneous distribution of the fungus inoculum onto the substrate. The fungus was grown for 35 days on autoclaved oat seeds and then ground to powder using a mill (1 mm). Fungicide seed treatments had a significant effect (P ≤ 0.05) on initial and final seedling emergence, pre and post-emergence damping-off, injured seedlings and disease index according to McKinney formula (lesion severity). The best results were obtained by fludioxonil + metalaxyl-M + azoxystrobin, fipronil + pyraclostrobin + methyl-thiophanate, carbendazim + thiram + pencycuron + baytan and carbendazim + thiram + pencycuron + baytan + procymidone. Pearson correlation analysis indicated that injured seedlings and disease index were positively correlated (R = 0.95). This means that the greater the number of injured seedlings, the greater the disease index and the less efficient the fungicide. The fungicides used in the present study did not have any phytotoxic effect on cotton seedlings.

Potential Biocontrol Activities of Populus Endophytes against Several Plant Pathogens Using Different Inhibitory Mechanisms

The plant microbiome can be used to bolster plant defense against abiotic and biotic stresses. Some strains of endophytes, the microorganisms within plants, can directly inhibit the growth of plant fungal pathogens. A previously isolated endophyte from wild Populus (poplar), WPB of the species Burkholderia vietnamiensis, had robust in vitro antifungal activity against pathogen strains that are highly virulent and of concern to Pacific Northwest agriculture: Rhizoctonia solani AG-8, Fusarium culmorum 70110023, and Gaemannomyces graminis var. tritici (Ggt) ARS-A1, as well as activity against the oomycete, Pythium ultimum 217. A direct screening method was developed for isolation of additional anti-fungal endophytes from wild poplar extracts. By challenging pathogens directly with dilute extracts, eleven isolates were found to be inhibitory to at least two plant pathogen strains and were therefore chosen for further characterization. Genomic analysis was conducted to determine if these endophyte strains harbored genes known to be involved in antimicrobial activities. The newly isolated Bacillus strains had gene clusters for production of bacillomycin, fengicyn, and bacillibactin, while the gene cluster for the synthesis of sessilin, viscosin and tolaasin were found in the Pseudomonas strains. The biosynthesis gene cluster for occidiofungin (ocf) was present in the Burkholderia vietnamiensis WPB genome, and an ocf deletion mutant lost inhibitory activity against 3 of the 4 pathogens. The new isolates lacked the gene cluster for occidiofungin implying they employ different modes of action. Other symbiotic traits including nitrogen fixation, phosphate solubilization, and the production of auxins and siderophores were investigated. Although it will be necessary to conduct in vivo tests of the candidates with pathogen-infected agricultural crops, the wild poplar tree microbiome may be a rich source of beneficial endophyte strains with potential for biocontrol applications against a variety of pathogens and utilizing varying modes of action.

First Report of Rhizoctonia solani AG-4 HG-III Causing Sheath Blight on Foxtail Millet in China.

Foxtail millet (Setaria italica) is an important grain and forage crop. This crop is widely grown in Northern China (Yang et al.2020). In Aug 2021, foxtail millet variety of Jigu42 showing lodging were found in Baoding China with the incidence of 30% and irregular brown lesions were found in sheaths and leaves of infected plants. The center of the lesions was kraurotic and pale, and the edges were gray-brown or dark brown. Twelve samples with typical lesions were collected from the surveyed field to isolate the pathogen. The infected samples were cut into square pieces of about 3 to 5 mm and were immersed into NaOCl (1%) for 1 min followed by washing with sterile water for three times. Then all sterilized tissues were inoculated on potato dextrose agar (PDA) plates and incubated at 25℃. After 3 days, fresh mycelial tips grown from the tissues were transferred to new plates for purification and incubated in the dark at 25°C for 4-5 days until the hyphae covered the whole plates. The colonies of 15 isolates on PDA medium showed similar colonial characteristics, which were fluffy and white initially, gradually turned light brown, and no sclerotia was observed even at 20 days later. Micro-examination revealed that all isolates showed the identical morphological features as Rhizoctonia sp. (Sneh et al. 1991), which contained the septate and right-angled branching hyphae with slight constriction at the base of mycelial branches, and three to seven nuclei per cell (Yang et al. 2013). Total genomic DNA was extracted from 5-day-old cultures, and the internal transcribed spacer (ITS) region of rDNA was amplified with ITS1 and ITS4 as the primers (Garibaldi et al. 2019). The sequencing results showed that the nucleotide sequences of 15 amplicons were identical and shared 100% identity with the corresponding fragments of R. solani AG-4 HG-III from sugar beet (GenBank accession No. MH172666 and MH172663) in Blastn search. The sequencing size of ITS in this study was 3 bp shorter than that of sugar beet, with a length of 722, because the base 'T' in the beginning and 'GA' in the end of the sequences did not detected in our study. Phylogenetic tree of 16 isolates of different AG4 subgroups was created by the software MEGA 7.0 through the NJ method, and the showed that the isolates were clustered to the clade of AG-4 HG-III group. The sequences of three isolates were deposited in GenBank under the accession No. ON810364, ON810365 and ON810366. For pathogenicity test, 5 mm diameters plate of the 5-day-old fungus which cultured on PDA were inoculated to the sheath of 10 foxtail millet plants grown in pots at 5- or 6-leaf stage. Then, the inoculated plants were placed into a growth chamber, and the inoculated sheaths were covered with wet cotton ball for 2 days to keep humidity, while sterile water was inoculated as the control. All plants were cultivated at 26°C with 14 h light and 10 h dark for 14 days. The experiment was repeated for three times. As the result, the same lesions observed in the field appeared on the inoculated plants at 10-14 days post inoculation, whereas the mock was healthy. The pathogen was re-isolated from the infected samples. The morphological characteristics and the nucleotide sequences of ITSs were same as that of the original isolates. All in above, the pathogen cusing sheath blight on foxtail millet was identified as R. solani AG-4 HG-III. To our knowledge, this is the first report of R. solani AG-4 HG-III causing sheath blight on S. italica in China. This finding expands the host range known for R. solani AG-4 HG-III and will be helpful for developing effective control strategies of foxtail millet sheath blight.

Metabolomic analysis of sheath blight disease of rice (Oryza sativa L.) induced by Rhizoctonia solani phytotoxin.

To understand the mechanism of necrosis incited by a host-selective phytotoxin designated as Rhizoctonia solani toxin (RST) identified to be a potential pathogenic factor of R. solani AG1 IA, causing sheath blight (ShB) of rice. The metabolomic changes induced by the phytotoxic metabolite in a ShB susceptible rice cultivar were elucidated by gas chromatography-mass spectrometry analysis and compared with that of the pathogen to identify rice metabolites targeted by the phytotoxin. The profiles of about 29 metabolites with various physiological roles in rice plants have been identified worldwide. Unsupervised and supervised multivariate chemometrics (principal component analysis and partial least squares-discriminant analysis) and cluster (Heat maps) analyses were used to compare the metabolites obtained from chemical profiles of the treatments with sterile distilled water (SDW) control. The results indicated that the rice plant expressed more metabolites in response to the pathogen than the phytotoxin and was lowest in SDW control. The key metabolites expressed in rice in response to the treatments were investigated by the variable importance in projection (VIP) analysis using p < 0.05 VIP >15. The analysis identified 7 and 11 upregulating metabolites in the phytotoxin and the pathogen treatments, respectively, compared to the untreated control. Among the phytotoxin-treated and the pathogen inoculated samples, the phytotoxin-treated sample recorded upregulation of six metabolites, whereas nine metabolites were upregulated in the pathogen-inoculated samples. These upregulating metabolites are speculated for the necrotic symptoms characteristic to both the phytotoxin and pathogen. In this analysis, hexadecanoic acid and dotriacontane were highly expressed metabolites specific to the phytotoxin and pathogen-treated samples, respectively. Besides upregulation, the metabolites also have a VIP score of >1.5 and hence fulfilled the criteria of classifying them as reliable potential biomarkers. In the pathway analysis, hexadecanoic acid and dotriacontane were identified to be involved in several important biosynthetic pathways of rice, such as the biosynthesis of saturated fatty acid and unsaturated fatty acids cutin, suberin and wax. The study concludes that though certain metabolites induced by the phytotoxin in the susceptible variety during necrosis shares with that of the pathogen, the identification of metabolites specific to the phytotoxin in comparison to the pathogenic and SDW controls indicated that the phytotoxin modulates the host metabolism differently and hence can be a potential pathogenicity factor of the ShB fungus. Due to lack of knowledge on the pathway genes of RST and in the absence of an ShB-resistant variety, understanding differentially expressed metabolic changes induced in the susceptible variety by the phytotoxin in comparison to that of the pathogenic and uninoculated controls enables us to identify the key metabolite changes during the ShB infection. Such metabolomic changes can further be used to infer gene functions for exploitation in ShB control.

Molecular characterization of a novel mitovirus from Rhizoctonia solani AG-4 HGIII strain XMC-IF.

The nucleotide sequence of a viral double-stranded RNA (dsRNA) from Rhizoctonia solani AG-4 HGIII strain XMC-IF (designated as "Rhizoctonia solani mitovirus 106", RsMV-106) was determined. The complete sequence was 2794 bp in length with a 57.50% A + U content and contained a large open reading frame (ORF) when the fungal mitochondrial genetic code was used. The ORF potentially encodes a 95.76-kDa protein containing a conserved domain of an RNA-dependent RNA polymerase (RdRp). BLASTp analysis revealed that the RdRp domain of RsMV-106 shared 47.52-73.24% sequence identity with those of viruses of the genus Duamitovirus and was most similar (73.24% identity) to that of Alternaria alternata mitovirus 1 (AaMV1). Phylogenetic analysis showed that RsMV-106 is a novel member of the genus Duamitovirus, family Mitoviridae. This is the first report of the full genome sequence of a mitovirus associated with R. solani AG-4 HGIII.

Integrative transcriptome analysis revealed the pathogenic molecular basis of Rhizoctonia solani AG-3 TB at three progressive stages of infection.

Rhizoctonia solani has a broad host range and results in significant losses in agricultural production. Here, an integrated transcriptomic analysis was performed to reveal the critical genes responsible for the pathogenesis of R. solani AG-3 TB on Nicotiana tabacum at different infection stages. The results showed that various differential expressed genes (DEGs) were enriched in fatty acid metabolism, amino sugar, carbon metabolism, and cellular carbohydrate biosynthetic process at the early (6–12 hpi), middle (24–36 hpi), and late stage (48–72 hpi) of infection. Specifically, several critical genes such as shikimate kinase that were involved in the biosynthesis of an important fungal toxin, phenylacetic acid (PAA) showed markedly increase at 24 hpi. Additionally, the genes expression levels of carbohydrate-active enzymes (CAZymes) and cell wall degrading enzymes (CWDEs) were significantly increased at the late infection stage. Furthermore, we identified 807 potential secreted proteins and 78 small cysteine-rich proteins, which may function as fungal effectors and involved in the pathogenicity. These results provide valuable insights into critical and potential genes as well as the pathways involved in the pathogenesis of R. solani AG-3 TB.