Biochemical and Molecular Basis of Chemically Induced Defense Activation in Maize against Banded Leaf and Sheath Blight Disease.

Maize is the third most vital global cereal, playing a key role in the world economy and plant genetics research. Despite its leadership in production, maize faces a severe threat from banded leaf and sheath blight, necessitating the urgent development of eco-friendly management strategies. This study aimed to understand the resistance mechanisms against banded leaf and sheath blight (BLSB) in maize hybrid "Vivek QPM-9". Seven fungicides at recommended doses (1000 and 500 ppm) and two plant defense inducers, salicylic acid (SA) and jasmonic acid (JA) at concentrations of 50 and 100 ppm, were applied. Fungicides, notably Azoxystrobin and Trifloxystrobin + Tebuconazole, demonstrated superior efficacy against BLSB, while Pencycuron showed limited effectiveness. Field-sprayed Azoxystrobin exhibited the lowest BLSB infection, correlating with heightened antioxidant enzyme activity (SOD, CAT, POX, ß-1,3-glucanase, PPO, PAL), similar to the Validamycin-treated plants. The expression of defense-related genes after seed priming with SA and JA was assessed via qRT-PCR. Lower SA concentrations down-regulated SOD, PPO, and APX genes but up-regulated CAT and ß-1,3-glucanase genes. JA at lower doses up-regulated CAT and APX genes, while higher doses up-regulated PPO and ß-1,3-glucanase genes; SOD gene expression was suppressed at both JA doses. This investigation elucidates the effectiveness of certain fungicides and plant defense inducers in mitigating BLSB in maize hybrids and sheds light on the intricate gene expression mechanisms governing defense responses against this pathogen.

Sustainable management of peanut damping-off and root rot diseases caused by Rhizoctonia solani using environmentally friendly bio-formulations prepared from batch fermentation broth of chitinase-producing Streptomyces cellulosae.

BACKGROUND: Soil-borne plant diseases represent a severe problem that negatively impacts the production of food crops. Actinobacteria play a vital role in biocontrolling soil-borne fungi. AIM AND OBJECTIVES: The target of the present study is to test the antagonistic activity of chitinase-producing Streptomyces cellulosae Actino 48 (accession number, MT573878) against Rhizoctonia solani. Subsequently, maximization of Actino 48 production using different fermentation processes in a stirred tank bioreactor. Finally, preparation of bio-friendly formulations prepared from the culture broth of Actino 48 using talc powder (TP) and bentonite in a natural as well as nano forms as carriers. Meanwhile, investigating their activities in reducing the damping-off and root rot diseases of peanut plants, infected by R. solani under greenhouse conditions. RESULTS: Actino 48 was found to be the most significant antagonistic isolate strain at p ≤ 0.05 and showed the highest inhibition percentage of fungal mycelium growth, which reached 97%. The results of scanning electron microscope (SEM) images analysis showed a large reduction in R. solani mycelia mass. Additionally, many aberrations changes and fungal hypha damages were found. Batch fermentation No. 2, which was performed using agitation speed of 200 rpm, achieved high chitinase activity of 0.1163 U mL- 1 min- 1 with a yield coefficient of 0.004 U mL- 1 min- 1 chitinase activity/g chitin. Nano-talc formulation of Actino 48 had more a significant effect compared to the other formulations in reducing percentages of damping-off and root rot diseases that equal to 19.05% and 4.76% with reduction percentages of 60% and 80%, respectively. The healthy survival percentage of peanut plants recorded 76.19%. Furthermore, the nano-talc formulation of Actino 48 was sufficient in increasing the dry weight of the peanut plants shoot, root systems, and the total number of peanut pods with increasing percentages of 47.62%, 55.62%, and 38.07%, respectively. CONCLUSION: The bio-friendly formulations of actinobacteria resulting from this investigation may play an active role in managing soil-borne diseases.

Genome analysis and hyphal movement characterization of the hitchhiker endohyphal Enterobacter sp. from Rhizoctonia solani.

Bacterial-fungal interactions are pervasive in the rhizosphere. While an increasing number of endohyphal bacteria have been identified, little is known about their ecology and impact on the associated fungal hosts and the surrounding environment. In this study, we characterized the genome of an Enterobacter sp. Crenshaw (En-Cren), which was isolated from the generalist fungal pathogen Rhizoctonia solani, and examined the genetic potential of the bacterium with regard to the phenotypic traits associated with the fungus. Overall, the En-Cren genome size was typical for members of the genus and was capable of free-living growth. The genome was 4.6 MB in size, and no plasmids were detected. Several prophage regions and genomic islands were identified that harbor unique genes in comparison with phylogenetically closely related Enterobacter spp. Type VI secretion system and cyanate assimilation genes were identified from the bacterium, while some common heavy metal resistance genes were absent. En-Cren contains the key genes for indole-3-acetic acid (IAA) and phenylacetic acid (PAA) biosynthesis, and produces IAA and PAA in vitro, which may impact the ecology or pathogenicity of the fungal pathogen in vivo. En-Cren was observed to move along hyphae of R. solani and on other basidiomycetes and ascomycetes in culture. The bacterial flagellum is essential for hyphal movement, while other pathways and genes may also be involved.IMPORTANCEThe genome characterization and comparative genomics analysis of Enterobacter sp. Crenshaw provided the foundation and resources for a better understanding of the ecology and evolution of this endohyphal bacteria in the rhizosphere. The ability to produce indole-3-acetic acid and phenylacetic acid may provide new angles to study the impact of phytohormones during the plant-pathogen interactions. The hitchhiking behavior of the bacterium on a diverse group of fungi, while inhibiting the growth of some others, revealed new areas of bacterial-fungal signaling and interaction, which have yet to be explored.

Biological characteristics and metabolic phenotypes of different anastomosis groups of Rhizoctonia solani strains.

BACKGROUND: Rhizoctonia solani is an important plant pathogen worldwide, and causes serious tobacco target spot in tobacco in the last five years. This research studied the biological characteristics of four different anastomosis groups strains (AG-3, AG-5, AG-6, AG-1-IB) of R. solani from tobacco. Using metabolic phenotype technology analyzed the metabolic phenotype differences of these strains. RESULTS: The results showed that the suitable temperature for mycelial growth of four anastomosis group strains were from 20 to 30oC, and for sclerotia formation were from 20 to 25oC. Under different lighting conditions, R. solani AG-6 strains produced the most sclerotium, followed by R. solani AG-3, R. solani AG-5 and R. solani AG-1-IB. All strains had strong oligotrophic survivability, and can grow on water agar medium without any nitrutions. They exhibited three types of sclerotia distribution form, including dispersed type (R. solani AG-5 and AG-6), peripheral type (R. solani AG-1-IB), and central type (R. solani AG-3). They all presented different pathogenicities in tobacco leaves, with the most virulent was noted by R. solani AG-6, followed by R. solani AG-5 and AG-1-IB, finally was R. solani AG-3. R. solani AG-1-IB strains firstly present symptom after inoculation. Metabolic fingerprints of four anastomosis groups were different to each other. R. solani AG-3, AG-6, AG-5 and AG-1-IB strains efficiently metabolized 88, 94, 71 and 92 carbon substrates, respectively. Nitrogen substrates of amino acids and peptides were the significant utilization patterns for R. solani AG-3. R. solani AG-3 and AG-6 showed a large range of adaptabilities and were still able to metabolize substrates in the presence of the osmolytes, including up to 8% sodium lactate. Four anastomosis groups all showed active metabolism in environments with pH values from 4 to 6 and exhibited decarboxylase activities. CONCLUSIONS: The biological characteristics of different anastomosis group strains varies, and there were significant differences in the metabolic phenotype characteristics of different anastomosis group strains towards carbon source, nitrogen source, pH, and osmotic pressure.

OsEIL2 balances rice immune responses against (hemi)biotrophic and necrotrophic pathogens via the salicylic acid and jasmonic acid synergism.

Plants typically activate distinct defense pathways against various pathogens. Heightened resistance to one pathogen often coincides with increased susceptibility to another pathogen. However, the underlying molecular basis of this antagonistic response remains unclear. Here, we demonstrate that mutants defective in the transcription factor ETHYLENE-INSENSITIVE 3-LIKE 2 (OsEIL2) exhibited enhanced resistance to the biotrophic bacterial pathogen Xanthomonas oryzae pv oryzae and to the hemibiotrophic fungal pathogen Magnaporthe oryzae, but enhanced susceptibility to the necrotrophic fungal pathogen Rhizoctonia solani. Furthermore, necrotroph-induced OsEIL2 binds to the promoter of OsWRKY67 with high affinity, leading to the upregulation of salicylic acid (SA)/jasmonic acid (JA) pathway genes and increased SA/JA levels, ultimately resulting in enhanced resistance. However, biotroph- and hemibiotroph-induced OsEIL2 targets OsERF083, resulting in the inhibition of SA/JA pathway genes and decreased SA/JA levels, ultimately leading to reduced resistance. Our findings unveil a previously uncharacterized defense mechanism wherein two distinct transcriptional regulatory modules differentially mediate immunity against pathogens with different lifestyles through the transcriptional reprogramming of phytohormone pathway genes.

Advances in breeding, biotechnology, and nanotechnological approaches to combat sheath blight disease in rice.

Sheath blight, caused by the fungus Rhizoctonia solani, is a major problem that significantly impacts rice production and can lead to substantial yield losses. The disease has become increasingly problematic in recent years due to the widespread use of high-yielding semi-dwarf rice cultivars, dense planting, and heavy application of nitrogenous fertilizers. The disease has become more challenging to manage due to its diverse host range and the lack of resistant cultivars. Despite utilizing traditional methods, the problem persists without a satisfactory solution. Therefore, modern approaches, including advanced breeding, transgenic methods, genome editing using CRISPR/Cas9 technology, and nanotechnological interventions, are being explored to develop rice plants resistant to sheath blight disease. This review primarily focuses on these recent advancements in combating the sheath blight disease.

The F-box protein ZmFBL41 negatively regulates disease resistance to Rhizoctonia solani by degrading the abscisic acid synthase ZmNCED6 in maize.

KEY MESSAGE: The maize F-box protein ZmFBL41 targets abscisic acid synthase 9-cis-epoxycarotenoid dioxygenase 6 for degradation, and this regulatory module is exploited by Rhizoctonia solani to promote infection. F-box proteins are crucial regulators of plant growth, development, and responses to abiotic and biotic stresses. Previous research identified the F-box gene ZmFBL41 as a negative regulator of maize (Zea mays) defenses against Rhizoctonia solani. However, the precise mechanisms by which F-box proteins mediate resistance to R. solani remain poorly understood. In this study, we show that ZmFBL41 interacts with an abscisic acid (ABA) synthase, 9-cis-epoxycarotenoid dioxygenase 6 (ZmNCED6), promoting its degradation via the ubiquitination pathway. We discovered that the ectopic overexpression of ZmNCED6 in rice (Oryza sativa) inhibited R. solani infection by activating stomatal closure, callose deposition, and jasmonic acid (JA) biosynthesis, indicating that ZmNCED6 enhances plant immunity against R. solani. Natural variation at ZmFBL41 across different maize haplotypes did not affect the ZmFBL41-ZmNCED6 interaction. These findings suggest that ZmFBL41 targets ZmNCED6 for degradation, leading to a decrease in ABA levels in maize, in turn, inhibiting ABA-mediated disease resistance pathways, such as stomatal closure, callose deposition, and JA biosynthesis, ultimately facilitating R. solani infection.

Lipase domain effector AGLIP1 in Rhizoctonia solani triggers necrotic killing in plants.

KEY MESSAGE: We highlight the emerging role of the R. solani novel lipase domain effector AGLIP1 in suppressing pattern-triggered immunity and inducing plant cell death. The dynamic interplay between plants and Rhizoctonia solani constitutes a multifaceted struggle for survival and dominance. Within this complex dynamic, R. solani has evolved virulence mechanisms by secreting effectors that disrupt plants' first line of defense. A newly discovered effector, AGLIP1 in R. solani, plays a pivotal role in inducing plant cell death and subverting immune responses. AGLIP1, a protein containing a signal peptide and a lipase domain, involves complex formation in the intercellular space, followed by translocation to the plant cytoplasm, where it induces cell death (CD) and suppresses defense gene regulation. This study provides valuable insights into the intricate molecular interactions between plants and necrotrophic fungi, underscoring the imperative for further exploration in this field.

Genome and transcriptome sequencing of Trichoderma harzianum T4, an important biocontrol fungus of Rhizoctonia solani, reveals genes related to mycoparasitism.

Trichoderma harzianum is a well-known biological control strain and a mycoparasite of Rhizoctonia solani. To explore the mechanisms of mycoparasitism, the genome and transcriptome of T. harzianum T4 were both assembled and analyzed in this study. The genome of T. harzianum T4 was assembled into 106 scaffolds, sized 41.25 Mb, and annotated with a total of 8118 predicted genes. We analyzed the transcriptome of T. harzianum T4 against R. solani in a dual culture in three culture periods: before contact (BC), during contact (C), and after contact (AC). Transcriptome sequencing identified 1092, 1222, and 2046 differentially expressed genes (DEGs), respectively. These DEGs, which are involved in pathogen recognition and signal transduction, hydrolase, transporters, antibiosis, and defense-related functional genes, are significantly upregulated in the mycoparasitism process. The results of genome and transcriptome analysis indicated that the mycoparasitism process of T. harzianum T4 was very complex. T. harzianum successfully recognizes and invades host cells and kills plant pathogens by regulating various DEGs at different culture periods. The relative expression levels of the 26 upregulated DEGs were confirmed by RT-qPCR to validate the reliability of the transcriptome data. The results provide insight into the molecular mechanisms underlying T. harzianum T4's mycoparasitic processes, and they provide a potential molecular target for the biological control mechanism of T. harzianum T4.

A pooled mycoviral resource in a strain of Rhizoctonia solani are regulators of fungal virulence.

Rhizoctonia solani is a widespread and devastating soil-borne plant fungal pathogen that causes diseases, including rice sheath blight, which are difficult to control. Some mycoviruses are potential biocontrol agents for the control of fungal diseases. In order to investigate the factors that influence the virulence of R. solani and search for mycoviruses with the potential for biocontrol of R. solani, a rice-infecting R. solani strain, ZJXD1-1, was isolated and confirmed to contain eight mycoviruses via dsRNA extraction and high-throughput sequencing. The identified mycoviruses belong to families of Endornaviridae (RsEV11 and RsEV12) and Mitoviridae (RsMV125 to RsMV129), and an unclassified Toti-like clade (RsTLV1). The C39 domain in RsEV12, which shares a close evolutionary relationship with bacteria, is observed for the first time in a mycovirus. Strains with different virus combinations were obtained through viral horizontal transfer, and pathogenicity test deduced that the Endornaviruses RsEV11 and RsEV12, and Mitovirus RsMV129 might potentially enhance the pathogenicity of R. solani, while RsMV125 might reduce the virulence or interfere with the function of other Mitoviruses. Furthermore, virus curing via protoplast regeneration and viral horizontal transfer demonstrated that RsMV129 is the causal agent of R. solani hypervirulence. Overall, our study provided the resource pool of viruses that may contribute to the discovery of new biocontrol agents against R. solani and enhance our understanding of the pathogenesis of R. solani regulated by mycoviruses.

Inhibitory Activity Against Rhizoctonia Solani and Chemical Composition of Extract from Moutan Cortex.

Rice sheath blight (RSB), caused by Rhizoctonia solani, is a significant disease of rice. The negative effects of chemical fungicides have created an urgent need for low-toxicity botanical fungicides. Our previous research revealed that the ethanol crude extract of Moutan Cortex (MC) exhibited superior antifungal activity against R. solani at 1000 µg/mL, resulting in a 100 % inhibition rate. The antifungal properties were mainly found in the petroleum ether extract. However, the active ingredients of the extract are still unclear. In this study, gas chromatography-mass spectrometry (GC-MS) was utilised for the analysis of its chemical components. The mycelium growth rate method was utilized to detect the antifungal activity. The findings indicated that paeonol constituted the primary active component, with a content of more than 96 %. Meanwhile, paeonol was the most significant antifungal active ingredient, the antifungal activity of paeonol (EC50=44.83 µg/mL) was much higher than that of ß-sitosterol and ethyl propionate against R. solani. Observation under an optical microscope revealed that paeonol resulted in abnormal mycelial morphology. This study provided theoretical support for identifying monomer antifungal compounds and developing biological fungicides for R. solani.

Evolution of pathogenicity-associated genes in Rhizoctonia solani AG1-IA by genome duplication and transposon-mediated gene function alterations.

BACKGROUND: Rhizoctonia solani is a polyphagous fungal pathogen that causes diseases in crops. The fungal strains are classified into anastomosis groups (AGs); however, genomic complexity, diversification into the AGs and the evolution of pathogenicity-associated genes remain poorly understood. RESULTS: We report a recent whole-genome duplication and sequential segmental duplications in AG1-IA strains of R. solani. Transposable element (TE) clusters have caused loss of synteny in the duplicated blocks and introduced differential structural alterations in the functional domains of several pathogenicity-associated paralogous gene pairs. We demonstrate that the TE-mediated structural variations in a glycosyl hydrolase domain and a GMC oxidoreductase domain in two paralogous pairs affect the pathogenicity of R. solani. Furthermore, to investigate the association of TEs with the natural selection and evolution of pathogenicity, we sequenced the genomes of forty-two rice field isolates of R. solani AG1-IA. The genomic regions with high population mutation rates and with the lowest nucleotide diversity are enriched with TEs. Genetic diversity analysis predicted the genes that are most likely under diversifying and purifying selections. We present evidence that a smaller variant of a glucosamine phosphate N-acetyltransferase (GNAT) protein, predicted to be under purifying selection, and an LPMP_AA9 domain-containing protein, predicted to be under diversifying selection, are important for the successful pathogenesis of R. solani in rice as well as tomato. CONCLUSIONS: Our study has unravelled whole-genome duplication, TE-mediated neofunctionalization of genes and evolution of pathogenicity traits in R. solani AG1-IA. The pathogenicity-associated genes identified during the study can serve as novel targets for disease control.

First report of Rhizoctonia solani AG-2-2 IIIB causing Rice sheath blight in China.

Rice (Oryza sativa) is an important crop worldwide, rice is susceptible to many pathogens, one of the most significant being Rice Sheath Blight, caused by Rhizoctonia solani. This disease initially produces cloudy spots on the leaf sheaths and later affects grain filling, resulting in yield losses of over 45%（Chen et al. 2013） when severe. In many southern rice-growing areas of China, the impact of this disease has risen to become the most damaging of the three major rice diseases (Margani et al. 2018). In July 2023, In Yongfu County, Guangxi (110.022°E, 25.010°N), symptoms of rice sheath blight were observed. The leaf sheaths were affected, with small, water-soaked, dark green spots with indistinct edges appearing near the water surface. These spots gradually expanded into elliptical or cloud-like lesions. Eventually, the center of the lesions turned straw-yellow to grayish-white, while the edges turned brown to dark brown. Often, several lesions merged into large cloud-like patches. Fifteen symptomatic sheaths were collected disinfecting pieces of necrotic tissue with 3% NaClO for 1.5 minutes, followed by 75% alcohol for 1 minute. The pieces were then rinsed with sterile distilled water, subsequently plated on Potato Dextrose Agar in Petri dishes, and incubated at 28°C in the dark. One isolate was obtained from each diseased plant using the hyphal tip method. (Feng et al. 2008). Isolates were obtained and displayed initially white mycelium and gradually turned brown after three to four days. Septate hyphae were 4.27 to 10.73 µ m (average 6.41 µ m) in diameter and branched at Right angle or acute angle with a constriction at the origin of the branch point. Staining with 1% safranin O and 3% KOH solution (Bandoni 1979) revealed multinucleated cells (three to nine nuclei per cell, n = 144). In summary, these characteristics were consistent with the description of Rhizoctonia solani Kühn (Meyer et al. 1990). The anastomosis group (AG) was confirmed by selecting three representative isolates (GL-Q-10, GL-Q-13, GL-Q-15) for molecular identification. The target DNA was extracted using Chelex-100. The internal transcribed spacer (ITS) region was amplified and sequenced with primers ITS1 and ITS4. The sequences were deposited in GenBank (ITS, PQ047154, PQ047150, and PQ047151 The base pairs are respectively 713bp, 715bp and 776bp, respectively). Upon searching GenBank, accession number MT385836 was found (Zhou et al. 2021), which has a similarity of 99.15% with PQ047154, 98.87% with PQ047150, and 99.30% with PQ047151. Phylogenetic tree analysis based on ITS sequences showed that the isolates clustered monophyletically with strains of R. solani AG-2-2 IIIB. The fusion group of the strain is verified by the shape and color of its mycelial growth on PDA at 35°C, enabling the distinct differentiation of AG-2-2 IIIB from AG-2-2 IV in terms of both morphology and coloration.（Aktaruzzaman et al. 2019） Pathogenicity tests involved culturing the pathogenic bacteria on PDA for 7-10 days, Then, 10 healthy rice plants (greenhouse potted rice variety Dian Heyou 615) were selected at the heading stage, and 5 plants were inoculated on the leaf sheaths with 5 strains of 5 mm fungus cake with pathogenic bacteria and 5 plants without pathogenic bacteria (The rice soil was disinfected), wrapped in cotton for moisture retention. All plants were sealed in transparent plastic bags and incubated in a greenhouse at 30 °C for 7-15 days, with daily moisturizing using sterile distilled water (Humidity control at 70%). Seven days postinoculation, Those containing pathogenic bacteria have symptoms of rice sheath blight, No symptoms were detected on control plants. Rhizoctonia solani AG-2-2 IIIB was re-isolated from the inoculated plants as previously described, thus fulfilling Koch's postulates. The pathogenicity tests were repeated three times. At present, Rhizoctonia solani AG-2-2 IIIB is primarily pathogenic in plants such as sugar beet and beans. It has only been reported in Japan and other countries to cause rice disease (Engelkes et al. 1996; Kenji Inagaki et al. 2004), and Rhizoctonia solani AG-2-2 IIIB has never been reported in China to cause disease in rice. To our knowledge, this study is the first to identify Rhizoctonia solani AG-2-2 IIIB causing rice sheath blight in China. This finding will aid further research on rice sheath blight defense strategies and contribute to the development of better management practices for this disease.

Anaerobic Soil Disinfestation and Vermicompost to Manage Bottom Rot in Organic Lettuce.

Rhizoctonia solani Kühn (teleomorph: Thanatephorus cucumeris [Frank] Donk) is an aggressive soilborne pathogen with a wide host range that survives saprophytically between crops, presenting a challenge for organic vegetable farmers who lack effective management tools. A 2-year field experiment was conducted at two organic farms to compare anaerobic soil disinfestation (ASD) and worm-cured compost (vermicompost) to manage bottom rot caused by R. solani subspecies AG1-IB in field-grown organic lettuce (Lactuca sativa). At each farm, four replicate plots of seven treatments were arranged in a randomized complete block design. Randomization was restricted by grouping treatments to evaluate ASD, and treatments to evaluate vermicompost in starter plugs. ASD experiment treatments were three different ASD carbon sources that are commonly used and widely available to local farmers in Vermont: compost, cover crop residues, and poultry manure fertilizer, as well as a tarped control. Vermicompost experimental treatments were vermicompost compared with two types of controls: a commercial biocontrol product (RootShield PLUS + G), and unamended (untarped control). This study demonstrated that the ASD method is achievable in a field setting on Vermont farms. However, neither ASD nor vermicompost produced significant disease suppression or resulted in higher marketable yields than standard growing practices. Given the laborious nature of ASD, it is likely more appropriate in a greenhouse setting with high-value crops that could especially benefit from being grown in plastic tarped beds (e.g., tomatoes and strawberries). This study is the first known attempt of field-implemented ASD for soil pathogen control in the northeastern United States.

Evaluation of Anaerobic Soil Disinfestation to Reduce Soilborne Diseases in Soilless and Soil-Based Substrates for Specialty Cut Flower Production.

Anaerobic soil disinfestation (ASD) is a nonchemical soil treatment where an easily decomposable carbon source is incorporated into soil, which is then irrigated to saturation and tarped to create anaerobic conditions, which prompts shifts in the soil microbiota from aerobes to anaerobes. ASD has been tested successfully for soilborne disease management in a variety of cropping systems but has not been sufficiently investigated in ornamentals. In this study, ASD was evaluated in soil-based and soilless substrates commonly used in specialty cut flower production using two model pathosystems: Rhizoctonia solani-Zinnia elegans and Phytophthora drechsleri-Gerbera jamesonii. Each substrate was mixed with pathogen-infested vermiculite and amended with either wheat bran, tomato pomace, or soybean meal as the carbon source. Amended substrates were incubated at 25°C for 4 weeks and used as growing substrates for the two crops mentioned above, which were monitored weekly for disease development for up to 5 weeks posttransplant. Additional experiments tested the effect of plant age and inoculum concentration in the substrate on ASD efficacy. Results showed that ASD has the potential to be deployed successfully for the control of Rhizoctonia stem rot in both substrates. Conversely, ASD was not effective at controlling Phytophthora crown rot on gerbera daisy in any of the experiments conducted in this study. More research is needed to understand the influence of carbon amendments, inoculum thresholds, and environmental conditions on ASD efficacy.

First Report of Rhizoctonia solani AG1-IB Causing Tobacco Target Spot in China.

Tobacco target spot, caused by Rhizoctonia solani Kühn, induces shot-hole lesions on leaves that that significantly reduce yield and quality of tobacco. In July 2022, samples (n=5) with target spot were collected from three tobacco fields, one each in Puer (22.63°N, 100.72°E, cv. Yunyan87) and Mengzi (23.26°N, 103.36°E, cv. Yunyan87) of Yunnan province and one in Dandong (40.63°N, 124.18°E, cv. Liaoyan17) of Liaoning province, China; disease incidence in these fields was approximately 30%~40%. Initial symptoms (2- to 3-mm-diameter lesions) appeared on the middle to lower leaves, then expanded to 2 to 3 cm in diameter and developed the shot-hole appearance. Pieces of tissue (5×5 mm) were cut from the edge of lesions, surface sterilized, rinsed in sterile water, then placed on the surface of water agar (WA) and incubated at 25℃ for 2 days in the dark. Single hyphal tips were taken from fungal isolates identified as R. solani based on the morphological traits (Tsror 2010), then transferred onto potato dextrose agar (PDA) and cultured for 3 d as described above. A total of 15 pure cultures were obtained. With the exception of YN-3 (isolated from Puer), YN-62 (isolated from Mengzi) and LN-95(isolated from Dandong) strains, which exhibited hyphal fusion reaction with AG1-IB standard strain, all the other strains demonstrated hyphal fusion with AG-3 standard strain (Ogoshi 1987). Genomic DNA of these three strains were extracted by the CTAB method and ITS regions of rDNA were sequenced (White et al. 1990). The sequences were deposited in GenBank with accession No. OR770079, OR770080 and OR770082. All the three rDNA-ITS sequences exhibited 99.85% similar to AG1-IB found in GenBank, and a phylogenetic tree using a neighbor-joining method grouped the three strains within the R. solani AG-1 IB clade. Therefore, based on the hyphal fusion reaction and molecular methods, these isolates were identified as R. solani AG1-IB. To determine pathogenicity of the isolates, the healthy leaves of tobacco plants (cv. Yunyan 87) were used. Five-mm-diameter mycelial plugs of the strain on PDA were inoculated on leaves that had been previously wounded with a sterile needle, and cotton balls moistened with sterile water were used for moisturizing the inoculation sites. Ten leaves were inoculated for each strain and leaves inoculated with PDA plugs were as control. The experiment was conducted twice. All plants were incubated for 2 d at 15℃ to 25℃ and 90% relative humidity with a 12 h photoperiod/day. Irregularly shaped lesions appeared on the leaves around each of the inoculated sites, but not on control leaves. The pathogens were reisolated and confirmed be R. solani AG1-IB by hyphal fusion and molecular identification tests as previously described, thereby fulfilling Koch's postulates. It has been reported that AG-3, AG-2 (Mercado Cardenas et al. 2012), AG-5 (Wang et al. 2023) and AG-6 (Sun et al. 2022) of R. solani could cause tobacco target spot, but AG-3 is considered the main causal agent (Marleny Gonzalez et al. 2011). To our knowledge, this is the first report of AG1-IB causing tobacco target spot in China and worldwide. The AG1-IB strain has a wide host range including cabbage, mint, lettuce, beans, and rice (Gonzalez et al. 2006). The discovery poses a new challenge for the prevention and control of tobacco target spot, especially when contemplating disease management strategies such as crop rotation and fungicide treatments.

First Report of Root and Bulb Rot Caused by Rhizoctonia solani AG-6 on Lanzhou Lily (Lilium davidii var. unicolor) in China.

Lanzhou lily (Lilium davidii var. unicolor) is the only famous sweet lily variety that has high edible, medicinal and ornamental value in China, which is mostly planted in the middle areas of Gansu Province in China. In recent years, severe yellowing and wilting of leaves, stem wilt, root and bulb rot symptoms were observed on Lanzhou lily in Qilihe District, Lanzhou, which has resulted in serious loss of bulb production. From June to August 2022, a survey of Lanzhou lily disease was carried out in Xiguoyuan and Weiling township of Qilihe District, Lanzhou. Typical symptoms of root and bulb rot were observed in Lanzhou lily fields. The disease incidence was estimated up to 30%. Fragments of symptomatic roots and bulbs were surface sterilized with 75% ethanol for 10 s, 2% sodium hypochlorite for 2 min, washed three times with sterilized distilled water, and then blotted dry on sterile filter paper. Fragments were placed on PDA medium and incubated at 25 ± 1°C in darkness for 5 days and 2 isolates were purified by the single-tip culture. Colonies of the fungus were white initially, and then turned light brown to brown, raised, and with entire or undulate edges. Sclerotia were brown and produced on PDA after 25 days of incubation at 25 ± 1°C in the dark. Genomic DNA from each of the two 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 of strains QLH22LD01 and QLH22LD02 were deposited in GenBank (OR710804 and OR710805). Phylogenetic analyses were performed using the Maximum Likelihood method with ITS sequences for anastomosis groups (AG) of Rhizoctonia solani. The phylogenetic tree grouped the two isolates within the R. solani AG-6 clade with high bootstrap support (100%). PCR analysis was performed with 21 primers specifically designed to detect individual anastomosis groups or anastomosis subgroups of R. solani (Carling et al., 2002; Misawa and Kurose, 2019; Misawa et al., 2020; Okubara et al., 2008). Among the 21 specific primer pairs, only AG-6 specific primer amplified the fungal DNA, indicating that the two isolates tested belonged to the R. solani AG-6. Therefore, these two strains were identified as R. solani AG-6. For pathogenicity tests, two isolates were grown individually on sterile wheat kernels at 25 ± 1°C for 14 days. Certified pathogen-free Lanzhou lily bulbs were grown in the plastic pot filled with the sterilized soil. Fifteen 2- week-old plants were inoculated by digging the soil and burying ten infested wheat kernels in the soil adjacent to the roots. Control plants were inoculated with sterile wheat kernels using the same procedure. All plants were placed in a greenhouse with a 12h/12h light/dark photoperiod at 15 to 30°C. Fifty days after inoculation, typical root and bulb rot symptoms developed on all inoculated plants, similar to symptoms observed in the field, whereas control plants remained symptomless. Pathogenicity test was performed three times with similar to symptoms observed in the field. Finally, the fungi were reisolated from the symptomatic plants and identified by molecular analysis as the isolates used for inoculation, thus fulfilling Koch's postulates. To our knowledge, this is the first confirmed report of R. solani AG-6 causing root and bulb rot on Lanzhou lily in China. Our findings improve knowledge about R. solani AGs occurring in Lanzhou lily fields in China. Due to serious damages caused by this disease in recent years in China, further studies should be conducted to investigate the diversity, prevalence, disease control measures and fungicide sensitivity of AGs distributed in the main Lanzhou lily-producing states in China.

First report of Rhizoctonia blight caused by Rhizoctonia solani on Chinese cabbage (Brassica rapa subsp. chinensis) in India.

During January and February 2021, foliar blight symptoms were observed on the leaves of Chinese cabbage (Pak choi) at Lembucherra research farm, College of Agriculture, Tripura, India. The incidence of disease symptoms ranged from 5 to 10% of the plants observed in the field. The symptomatic leaves showed grayish colored water-soaked lesions with an irreguar shape and size. A total of 10 symptomatic leaves (1 leaf per plant) from Chinese cabbage infected plant were sampled, surface decontaminated with 1% NaOCl, washed twice in sterile water, plated on 2% water agar, and incubated at 25 ± 2°C. Hyphal tips from mycelium of 7-day old culture (2 isolates from two different plants) with right-angled branching were transferred to potato dextrose agar (PDA) media (SRL, India). Cream or light brown hyphae that branched at right angles, with septa near the point of the origin of hyphae, and a slight constriction at the base of the branch) were visible under a microscope. Olive-brown sclerotia were observed after 5 days of incubation. Multiple nuclei per cell were visible after staining with 4', 6-diamidino-2-phenylindole (Estandarte et al. 2016). Based on morphological characteristics (Parmeter et al. 1970) the isolates TP36 and TP37 were identified as Rhizoctonia solani. The internal transcribed spacer (ITS) region and glyceraldehyde-3-phosphate dehydrogenase gene (GAPDH) were amplified with ITS1& ITS4 (White et al. 1990) and (GAPDH F-5'- CAAGGAGAACCCAGGTGTTAAG-3' and GAPDH R- 5'-GGCGTCGAAGATAGAAGAGTGT-3') respectively for both isolates and sequenced (accession #. PP458158, PP458159, PP425343, PP425344). BLASTn analysis showed 99.26%( 668/673 nt) to 99.46% (659/664 nt) identity with R. solani sequences (GenBank MG397062.1 and KX674524.1) for ITS and 98.42% (552/562 nt) to 100% 540/540 nt)identity with R. solani sequences (GenBank HQ425709.1 and CP102644.1) for GAPDH. Isolates TP36 and TP37 were deposited in the Indian Type Culture Collection (ITCC), New Delhi as R. solani (nos. 9154 and 9319, respectively). Both isolates were amplified using (anastomosis group) AG1 subgroup specific primers (Matsumoto 2002; Prashantha et al. 2021) to identify their AG. The presence of a 265 bp amplicon for both isolates suggested that they belong to AG1-IA. A multilocus analysis of R. solani isolates from different host plants with concatenated sequences ITS and GAPH showed that TP36 and TP37 are closely related to rice isolate RS107. A pathogenicity test on five plants per treatment was conducted and repeated twice on one month old Chinese cabbage plants (hybrid, TOKITA, India) grown under glasshouse conditions in a sterilized mixture of soil and sand (3:1) at 27-28oC during January 2024 at ICAR-IARI, New Delhi. R. solani isolates TP36 and TP37 were grown on PDA and plants were inoculated by placing single sclerotia of 10-day old colony on different plant parts and covering it with moist cotton. After 7 day, typical lesions of R. solani infection were visible. No symptoms were observed on the control plants. The fungus was reisolated from the inoculated plants and identified as R. solani based on morphology. R. solani has previously been reported to cause disease on some members of Brassicaceae in different countries (Budge et al. 2009; Hua et al. 2014). Based on literature available this is the first report of R. solani infecting Chinese cabbage in India.

Mechanism Analysis of OsZF8-Mediated Regulation of Rice Resistance to Sheath Blight.

Transcription factors are key molecules involved in transcriptional and post-transcriptional regulation in plants and play an important regulatory role in resisting biological stress. In this study, we identified a regulatory factor, OsZF8, mediating rice response to Rhizoctonia solani (R. solani) AG1-IA infection. The expression of OsZF8 affects R. solani rice infection. OsZF8 knockout and overexpressed rice plants were constructed, and the phenotypes of mutant and wild-type (WT) plants showed that OsZF8 negatively regulated rice resistance to rice sheath blight. However, it was speculated that OsZF8 plays a regulatory role at the protein level. The interacting protein PRB1 of OsZF8 was screened using the yeast two-hybrid and bimolecular fluorescence complementation test. The results showed that OsZF8 effectively inhibited PRB1-induced cell death in tobacco cells, and molecular docking results showed that PRB1 had a strong binding effect with OsZF8. Further, the binding ability of OsZF8-PRB1 to ergosterol was significantly reduced when compared with the PRB1 protein. These findings provide new insights into elucidating the mechanism of rice resistance to rice sheath blight.

Enhancing Vicia faba's immunity against Rhizoctonia solani root rot diseases by arbuscular mycorrhizal fungi and nano chitosan.

BACKGROUND: The spreading of root rot disease of faba bean plant (Vichia faba L, VF) in Egypt is still of great challenge faced researchers since VF is an important legume in Egypt, because their seeds are used for human feeding. Fungicides are used for treatment of either seeds or soil; unfortunately they cause environmental pollution. Therefore, there is a need to continue research to find out safe natural solutions. In this regard, Arbuscular mycorrhizal fungi (AMF) and chitosan (micro or nanoform) were used as an inhibitory product against Rhizoctonia solani OM918223 (R.solani) either singly or in combinations. RESULTS: The results employed herein have exhibited that R.solani caused root rot disease of VF plants in more than 80% of the plants under investigation. Chitosan nanoparticles (Chitosan NPs) were prepared by ionic gelatin method and characterized by using dynamic light scattering (DLS), transmission electron microscopy (TEM) imaging and Fourier transform infra-red (FTIR). Chitosan NPs are spherical with a diameter of 78.5 nm and exhibited the presence of different functional groups. The inhibitory natural products against R.solani were arranged according to their ability to inhibit the pathogen used in the following descending manner; combination of AMF with Chitosan NPs, AMF with micro chitosan and single AMF, respectively. Where, Chitosan NPs showed a potent influence on R.solani pathogen and reduced the pre-and post-emergence of R. solani. In addition, Chitosan NPs reduced Disease Incidence (DI %) and Disease Severity (DS %) of root rot disease and are widely functional through mixing with AMF by about 88% and 89%. Further, Chitosan NPs and micro chitosan were proved to increase the growth parameters of VF plants such as nutritional status (mineral, soluble sugar, and pigment content), and defense mechanisms including total phenol, peroxidase, and polyphenol oxidase in mycorrhizal plants more than non-mycorrhizal one either in infected or healthy plants. Moreover, activity of AMF as an inhibitory against R.solani and improvement natural agent for VF growth parameters was enhanced through its fusing with Chitosan NPs. CONCLUSIONS: The use of AMF and Chitosan NPs increased faba bean plant resistance against the infection of root rot R. solani, with both prevention and cure together. Therefore, this research opens the door to choose natural and environmental friendly treatments with different mechanisms of plant resistance to disease.