Effects of Klebsiella michiganensis LDS17 on Codonopsis pilosula growth, rhizosphere soil enzyme activities, and microflora, and genome-wide analysis of plant growth-promoting genes.

Codonopsis pilosula is a perennial herbaceous liana with medicinal value. It is critical to promote Codonopsis pilosula growth through effective and sustainable methods, and the use of plant growth-promoting bacteria (PGPB) is a promising candidate. In this study, we isolated a PGPB, Klebsiella michiganensis LDS17, that produced a highly active 1-aminocyclopropane-1-carboxylate deaminase from the Codonopsis pilosula rhizosphere. The strain exhibited multiple plant growth-promoting properties. The antagonistic activity of strain LDS17 against eight phytopathogenic fungi was investigated, and the results showed that strain LDS17 had obvious antagonistic effects on Rhizoctonia solani, Colletotrichum camelliae, Cytospora chrysosperma, and Phomopsis macrospore with growth inhibition rates of 54.22%, 49.41%, 48.89%, and 41.11%, respectively. Inoculation of strain LDS17 not only significantly increased the growth of Codonopsis pilosula seedlings but also increased the invertase and urease activities, the number of culturable bacteria, actinomycetes, and fungi, as well as the functional diversity of microbial communities in the rhizosphere soil of the seedlings. Heavy metal (HM) resistance tests showed that LDS17 is resistant to copper, zinc, and nickel. Whole-genome analysis of strain LDS17 revealed the genes involved in IAA production, siderophore synthesis, nitrogen fixation, P solubilization, and HM resistance. We further identified a gene (koyR) encoding a plant-responsive LuxR solo in the LDS17 genome. Klebsiella michiganensis LDS17 may therefore be useful in microbial fertilizers for Codonopsis pilosula. The identification of genes related to plant growth and HM resistance provides an important foundation for future analyses of the molecular mechanisms underlying the plant growth promotion and HM resistance of LDS17. IMPORTANCE: We comprehensively evaluated the plant growth-promoting characteristics and heavy metal (HM) resistance ability of the LDS17 strain, as well as the effects of strain LDS17 inoculation on the Codonopsis pilosula seedling growth and the soil qualities in the Codonopsis pilosula rhizosphere. We conducted whole-genome analysis and identified lots of genes and gene clusters contributing to plant-beneficial functions and HM resistance, which is critical for further elucidating the plant growth-promoting mechanism of strain LDS17 and expanding its application in the development of plant growth-promoting agents used in the environment under HM stress.

Development of a Greenhouse Assay to Screen Soybean Varieties for Resistance to Aerial Blight Caused by Rhizoctonia solani Anastomosis Group 1-IA.

Aerial blight, caused by the fungus Rhizoctonia solani anastomosis group (AG) 1-IA, is an economically important soybean disease in the mid-Southern United States. Management has relied on fungicide applications during the season, but there is an increasing prevalence of resistance to commonly used strobilurin fungicides and an urgent need to identify soybean varieties resistant to aerial blight. Because the patchy distribution of the pathogen complicates field variety screening, the present study aimed to develop a greenhouse screening protocol to identify soybean varieties resistant to aerial blight. For this, 88 pathogen isolates were collected from commercial fields and research farms across five Louisiana parishes, and 77% were confirmed to be R. solani AG1-IA. Three polymorphic codominant microsatellite markers were used to explore the genetic diversity of 43 R. solani AG1-IA isolates, which showed high genetic diversity, with 35 haplotypes in total and only two haplotypes common to two other locations. Six genetically diverse isolates were chosen and characterized for their virulence and fungicide sensitivity. The isolate AC2 was identified as the most virulent and was resistant to both active ingredients, azoxystrobin and pyraclostrobin, tested. The six isolates were used in greenhouse variety screening trials using a millet inoculation protocol. Of the 31 varieties screened, only Armor 48-D25 was classified as moderately resistant, and plant height to the first node influenced final disease severity. The study provides short-term solutions for growers to choose less susceptible varieties for planting and lays the foundation to characterize host resistance against this important soybean pathogen.

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.

Establishment and application of quantitative detection of Bacillus velezensis HMB26553, a biocontrol agent against cotton damping-off caused by Rhizoctonia.

A highly sensitive quantitative PCR (qPCR) method was developed for detection and quantification of Bacillus velezensis HMB26553 in cotton rhizosphere. The study aimed to develop a quantitative detection method for the strain HMB26553, and explore the relationship between its colonization of the cotton rhizosphere and its control effect. The whole genome sequence of strain HMB26553 was obtained by genome sequencing and a unique specific sequence pB-gene0026 on plasmid plaBV2 was identified by using high-throughput alignment against NCBI. Plasmid plaBV2 could be stably genetically inherited. Based on this sequence, specific primers for amplifying 106 bp and a minor groove binder (MGB) TaqMan probe for enhancing sensitivity were designed. The copy number of plaBV2 in strain HMB26553, which was 2, was confirmed by internal reference primers and the MGB TaqMan probe based on housekeeping gene gyrB. The established detection technique based on these primers and probes had high specificity and sensitivity compared to traditional plate counting method, with a detection limit of 1.5 copy genome. Using this method, the study discovered a likely correlation between the quantity of colonization in cotton rhizosphere and efficacy against cotton damping-off caused by Rhizoctonia after seed soaking and irrigation with strain HMB26553. Thus, this method provides scientific support for the rational application of strain HMB26553 in the future.

Characterization of a novel endornavirus isolated from the phytopathogenic fungus Rhizoctonia solani.

Rhizoctonia solani endornavirus 8 (RsEV8) was isolated from strain XY175 of Rhizoctonia solani AG-1 IA. The full-length genome of RsEV8 is 16,147 nucleotides (nt) in length and contains a single open reading frame that encodes a large polyprotein of 5227 amino acids. The polyprotein contains four conserved domains: viral methyltransferase, putative DEAH box helicase, viral helicase, and RNA-dependent RNA polymerase (RdRp). RsEV8 has a shorter 3'-UTR (58 nt) and a longer 5'-UTR (404 nt). A multiple sequence alignment indicated that the RdRp of RsEV8 possesses eight typical RdRp motifs. According to a BLASTp analysis, RsEV8 shares 39.31% sequence identity with Rhizoctonia cerealis endornavirus-1084-7. Phylogenetic analysis demonstrated that RsEV8 clusters with members of the genus Betaendornavirus.

Mitochondrial-targeting effector RsIA_CtaG/Cox11 in Rhizoctonia solani AG-1 IA has two functions: plant immunity suppression and cell death induction mediated by a rice cytochrome c oxidase subunit.

Rhizoctonia solani AG-1 IA causes a necrotrophic rice disease and is a serious threat to rice production. To date, only a few effectors have been characterized in AG-1 IA. We previously identified RsIA_CtaG/Cox11 and showed that infiltration of the recombinant protein into rice leaves caused disease-like symptoms. In the present study, we further characterized the functionality of RsIA_CtaG/Cox11. RsIA_CtaG/Cox11 is an alternative transcript of cytochrome c oxidase copper chaperone Cox11 that starts from the second AUG codon, but contains a functional secretion signal peptide. RNA interference with RsIA_CtaG/Cox11 reduced the pathogenicity of AG-1 IA towards rice and Nicotiana benthamiana without affecting its fitness or mycelial morphology. Transient expression of the RsIA_CtaG/Cox11-GFP fusion protein demonstrated the localization of RsIA_CtaG/Cox11 to mitochondria. Agro-infiltration of RsIA_CtaG/Cox11 into N. benthamiana leaves inhibited cell death by BAX and INF1. In contrast to rice, agro-infiltration of RsIA_CtaG/Cox11 did not induce cell death in N. benthamiana. However, cell death was observed when it was coinfiltrated with Os_CoxVIIa, which encodes a subunit of cytochrome c oxidase. Os_CoxVIIa appeared to interact with RsIA_CtaG/Cox11. The cell death triggered by coexpression of RsIA_CtaG/Cox11 and Os_CoxVIIa is independent of the leucine-rich repeat receptor kinases BAK1/SOBIR1 and enhanced the susceptibility of N. benthamiana to AG-1 IA. Two of the three evolutionarily conserved cysteine residues at positions 25 and 126 of RsIA_CtaG/Cox11 were essential for its immunosuppressive activity, but not for cell death induction. This report suggests that RsIA_CtaG/Cox11 appears to have a dual role in immunosuppression and cell death induction during pathogenesis.

Characterization of three-nucleate Rhizoctonia AG-E based on their morphology and phylogeny.

The genus Rhizoctonia has been classified into two main groups according to the number of nuclei. Binucleate Rhizoctonia strains have two nuclei in each cell, whereas multinucleate Rhizoctonia fungi were observed to have a variable number of nuclei ranging from 4 to 16 in each cell. In the study, twelve Polish isolates were tested. According to ITS1-5,8S-ITS2 rDNA sequences, the isolates were classified in the AG-E. Their affiliation to AG was confirmed by anastomosis reactions with tester isolates. The number of nuclei was counted with DAPI staining under a fluorescent microscope, and the diameter of the hyphae was also measured. Not all AG-E isolates had the same number of nuclei in their cells: one group among these fungi produced cells with a diverse number of nuclei, usually 3; however, this number ranged from 2 to 4, making the average number of nuclei close to 3. It can be assumed that all isolates with three nuclei belong to this group, which may greatly facilitate the preliminary identification of trinucleate isolates of Rhizoctonia spp. belonging to AG-E. Based on these characters, we call these isolates AG-E-3n isolates. The thiamine requirement is not helpful in classifying and describing the AG-E strains.

Functional Analyses of the Bacillus velezensis HMB26553 Genome Provide Evidence That Its Genes Are Potentially Related to the Promotion of Plant Growth and Prevention of Cotton Rhizoctonia Damping-Off.

Bacillus spp. is one kind of the important representative biocontrol agents against plant diseases and promoting plant growth. In this study, the whole genomic sequence of bacterial strain HMB26553 was obtained. A phylogenetic tree based on the genome and ANI (average nucleotide identity), as well as dDDH (digital DNA-DNA hybridization), was constructed, and strain HMB26553 was identified as Bacillus velezensis. Fourteen biosynthetic gene clusters responsible for secondary metabolite were predicted via anti-SMASH, and six secondary metabolites were identified by UHPLC-QTOF-MS/MS (ultra-high-performance liquid chromatography coupled to quadrupole-time-of-flight tandem mass spectrometry). When the phytopathogen Rhizoctonia solani was treated with B. velezensis HMB26553, the mycelial structure changed, ROS (reactive oxygen species) accumulated, and the mitochondrial membrane potential decreased. Characteristics of strain HMB26553 were predicted and confirmed by genomic information and experiments, such as producing IAA, siderophore, extracellular enzymes and biofilm, as well as moving and promoting cotton growth. All these results suggested the mechanisms by which B. velezensis HMB26553 inhibits pathogen growth and promotes cotton growth, which likely provided the potential biocontrol agent to control cotton Rhizoctonia damping-off.

Multiomics analysis reveals the molecular mechanisms underlying virulence in Rhizoctonia and jasmonic acid-mediated resistance in Tartary buckwheat (Fagopyrum tataricum).

Rhizoctonia solani is a devastating soil-borne pathogen that seriously threatens the cultivation of economically important crops. Multiple strains with a very broad host range have been identified, but only 1 (AG1-IA, which causes rice sheath blight disease) has been examined in detail. Here, we analyzed AG4-HGI 3 originally isolated from Tartary buckwheat (Fagopyrum tataricum), but with a host range comparable to AG1-IA. Genome comparison reveals abundant pathogenicity genes in this strain. We used multiomic approaches to improve the efficiency of screening for disease resistance genes. Transcriptomes of the plant-fungi interaction identified differentially expressed genes associated with virulence in Rhizoctonia and resistance in Tartary buckwheat. Integration with jasmonate-mediated transcriptome and metabolome changes revealed a negative regulator of jasmonate signaling, cytochrome P450 (FtCYP94C1), as increasing disease resistance probably via accumulation of resistance-related flavonoids. The integration of resistance data for 320 Tartary buckwheat accessions identified a gene homolog to aspartic proteinase (FtASP), with peak expression following R. solani inoculation. FtASP exhibits no proteinase activity but functions as an antibacterial peptide that slows fungal growth. This work reveals a potential mechanism behind pathogen virulence and host resistance, which should accelerate the molecular breeding of resistant varieties in economically essential crops.

Selection of reference genes for RT-qPCR analysis of rice with Rhizoctonia solani infection and biocontrol PGPR/KSi application.

BACKGROUND: Rhizoctonia solani (AG1 IA) is an important pathogen of rice (Oryza sativa L.) that causes rice sheath blight (RSB). Since control of RSB by breeding and fungicides have had limited success, novel strategies like biocontrol with plant growth-promoting rhizobacteria (PGPR) can be an effective alternative. METHOD AND RESULTS: Seven commonly used reference genes (RGs), 18SrRNA, ACT1, GAPDH2, UBC5, RPS27, eIF4a and CYP28, were evaluated for their stability in rice-R. solani-PGPR interaction for real-time quantitative PCR (RT-qPCR) analysis. Different algorithms were examined, Delta Ct, geNorm, NormFinder, BestKeeper, and comprehensive ranking by RefFinder, to evaluate RT-qPCR of rice in tissues infected with R. solani and treated with the PGPR strains, Pseudomonas saponiphilia and Pseudomonas protegens, with potassium silicate (KSi) alone or in combination with each PGPR strain. RG stability was affected for each treatment and treatment-specific RG selection was suggested. Validation analysis was done for nonexpressor of PR-1(NPR1) for each treatment. CONCLUSION: Overall, ACT1 was the most stable RG with R. solani infection alone, GAPDH2 with R. solani infection plus KSi, UBC5 with R. solani infection plus P. saponiphilia, and eIF4a with R. solani infection plus P. protegens. Both ACT1 and RPS27 were the most stable with the combination of KSi and P. saponiphilia, while RPS27 was the most stable with the combination of KSi and P. protegens.

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.

Complete genome sequence of a novel fusarivirus from Rhizoctonia solani AG-3 PT strain 3P-2-2.

Here, we describe a novel mycovirus, tentatively designated as "Rhizoctonia solani fusarivirus 6" (RsFV6), which was discovered in Rhizoctonia solani AG-3 PT strain 3P-2-2. The virus has a single-stranded positive-sense RNA (+ssRNA) genome of 6141 nucleotides containing two open reading frames (ORFs) and a poly(A) tail. ORF1 encodes a large polypeptide of 1,862 amino acids (aa) with conserved RNA-dependent RNA polymerase (RdRp) and helicase (Hel) domains. ORF2 encodes a putative 167-aa protein of unknown function. BLASTp searches revealed that the ORF1-encoded polypeptide showed the highest sequence similarity (70.67% identity) to that of Rhizoctonia solani fusarivirus 3 (RsFV3), which was isolated from Rhizoctonia solani AG-2-2LP. Multiple sequence alignments and phylogenetic analysis based on RdRp and Hel sequences indicated that RsFV6 could be a novel member of the genus Alphafusarivirus family Fusariviridae.

Biocontrol of sheath blight of rice (Oryza sativa L.) through alteration in expression dynamics of candidate effector genes of Rhizoctonia solani AG1-IA during pathogenesis.

In genome analyses of Rhizoctonia solani AG1-IA causing sheath blight (ShB) of rice, many genes were identified to have a hypothetical role in pathogenesis. To understand their roles in pathogenesis, their expressions during fungal infection were studied. An aggressive R. solani strain, RIRS-K, was first identified among six isolates, RIRS-K, RIRS-17, RIRS-S, RIRS-T, RIRS-MU and RIRS-FD, for inducing a maximum relative lesion height (RLH) of 32.7% on a ShB susceptible cultivar, Pusa Basmati-1. Hypothetical pathogenicity genes (52 nos) identified by in silico analyses of the publicly available genomic database of the pathogen strain were evaluated in Pathogen-Host Interaction (PHI) blast and RIRS-K. Though PHI blast identified 26 genes as potential ones, only 8 were constitutively expressive in RIRS-K cultured in a minimal broth. Among them, only expressions of AG1IA_06195, AG02692, AG04508, and AG05730 were induced in the rice plant inoculated with RIRS-K and, hence, were identified as the candidate ones. The candidate genes were highly expressed in the aggressive strain (RIRS-K) in comparison to the less aggressive one (RIRS-17). In further testing of their expressions in the highly aggressive fungal strain, RIRS-K infecting PB-1 pre-colonized by a potent biocontrol consortium comprising of Bacillus subtilis (S17TH), Pseudomonas putida (TEPF-Sungal-1), and Trichoderma harzianum (S17TH), the disease scoring and gene expression studies indicated that the candidate genes were downregulated. The studies, therefore, speculated that these genes might play a role in pathogen aggressiveness and ShB development.

Genome Resource of Rhizoctonia solani Anastomosis Group 4 Strain AG4-JY, a Pathomycete of Sheath Blight of Foxtail Millet.

The basidiomycetous fungus Rhizoctonia solani Kühn (teleomorph Thanatephorus cucumeris [Frank] Donk) is a fungal pathogen that causes various diseases on economically important crops, such as foxtail millet, maize, and rice. Using the PacBio Sequel platform, we assembled a draft genome of an R. solani strain AG4-JY that was isolated from foxtail millet with sheath blight at the stem. The genome was approximately 43.43 Mb on 53 scaffolds, with a scaffold N50 length of 2.10 Mb. In all, 10,545 genes and 179 noncoding RNAs were predicted, and 10,488 genes had at least one database annotation. In addition, the proteins encoded by 709 genes were predicted as secretory proteins. The AG4-JY genome sequence provides a valuable resource for understanding the interactions between R. solani and foxtail millet and controls sheath blight in the world.

A small secreted protein, RsMf8HN, in Rhizoctonia solani triggers plant immune response, which interacts with rice OsHIPP28.

The necrotrophic phytopathogen Rhizoctonia solani (R. solani) causes disease in many plant species. This fungal genome encodes abundant small cysteine-rich (SCR)-secreted proteins in R. solani that may induce pathogenesis. To test their molecular functions, we introduced 10 SCR-secreted protein genes from R. solani into tobacco leaves via agroinfiltration. Consequently, we identified RsMf8HN, a novel SCR protein that triggers cell death and an oxidative burst in tobacco. RsMf8HN comprises 182 amino acids (aa), including a signal peptide (SP) of 17aa, and the protein has unique features: it is orthologous to an allergen protein Mal f 8 occurring in Malassezia species, and possesses a high glycine and serine content. RsMf8HN is coded in a genomic location along with its paralogues and a few other effector candidates. The elicitation of plant immunity by RsMf8HN was dependent on HSP90 and SGT1. RsMf8HN was translocated to multiple locations within the host cells: i.e., nuclei, chloroplasts, and plasma membranes. We confirmed the occurrence of in vivo cross-interactions of RsMf8HN with a rice molecule, the heavy metal-associated isoprenylated plant protein OsHIPP28, which is a protein related to the disease susceptibility factor Pi21. In summary, our results suggest that RsMf8HN is a potential effector that enables necrotrophic phytopathogens to interfere with host plant immunity.

Nanosheet-Facilitated Spray Delivery of dsRNAs Represents a Potential Tool to Control Rhizoctonia solani Infection.

Rhizoctonia solani is one of the important pathogenic fungi causing several serious crop diseases, such as maize and rice sheath blight. Current methods used to control the disease mainly depend on spraying fungicides because there is no immunity or high resistance available in crops. Spraying double-strand RNA (dsRNA) for induced-gene silencing (SIGS) is a new potentially sustainable and environmentally friendly tool to control plant diseases. Here, we found that fluorescein-labelled EGFP-dsRNA could be absorbed by R. solani in co-incubation. Furthermore, three dsRNAs, each targeting one of pathogenicity-related genes, RsPG1, RsCATA, and RsCRZ1, significantly downregulated the transcript levels of the target genes after co-incubation, leading to a significant reduction in the pathogenicity of the fungus. Only the spray of RsCRZ1 dsRNA, but not RsPG1 or RsCATA dsRNA, affected fungal sclerotium formation. dsRNA stability on leaf surfaces and its efficiency in entering leaf cells were significantly improved when dsRNAs were loaded on layered double hydroxide (LDH) nanosheets. Notably, the RsCRZ1-dsRNA-LDH approach showed stronger and more lasting effects than using RsCRZ1-dsRNA alone in controlling pathogen development. Together, this study provides a new potential method to control crop diseases caused by R. solani.

Transcriptome analysis reveals genes potentially related to maize resistance to Rhizoctonia solani.

Banded leaf and sheath blight (BLSB) is a devasting disease caused by the necrotrophic fungus Rhizoctonia solani that affects maize (Zea mays L.) fields worldwide, especially in China and Southeast Asia. Understanding how maize plants respond to R. solani infection is a key step towards controlling the spread of this fungal pathogen. In this study, we determined the transcriptome of maize plants infected by a low-virulence strain (LVS) and a high-virulence strain (HVS) of R. solani for 3 and 5 days by transcriptome deep-sequencing (RNA-seq). We identified 3,015 (for LVS infection) and 1,628 (for HVS infection) differentially expressed genes (DEGs). We confirmed the expression profiles of 10 randomly selected DEGs by quantitative reverse transcription PCR. We also performed a Gene Ontology (GO) enrichment analysis to establish which biological processes are associated with these DEGs, which revealed the enrichment of defense-related GO terms in LVS- and HVS-regulated genes. We selected 388 DEGs upregulated upon fungal infection as possible candidate genes. Among them, the overexpression of ZmNAC41 (encoding NAC transcription factor 41) or ZmBAK1 (encoding BRASSINOSTEROID INSENSITIVE 1-associated receptor kinase 1) in rice enhanced resistance to R. solani. In addition, overexpressing ZmBAK1 in rice also increased plant height, plant weight, thousand-grain weight, and grain length. The identification of 388 potential key maize genes related to resistance to R. solani provides significant insights into improving BLSB resistance.

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.

Reduction of Rhizoctonia cerealis Infection on Wheat Through Host- and Spray-Induced Gene Silencing of an Orphan Secreted Gene.

Rhizoctonia cerealis is a soilborne fungus that can cause sharp eyespot in wheat, resulting in massive yield losses found in many countries. Due to the lack of resistant cultivars, fungicides have been widely used to control this pathogen. However, chemical control is not environmentally friendly and is costly. Meanwhile, the lack of genetic transformation tools has hindered the functional characterization of virulence genes. In this study, we attempted to characterize the function of virulence genes by two transient methods, host-induced gene silencing (HIGS) and spray-induced gene silencing (SIGS), which use RNA interference to suppress the pathogenic development. We identified ten secretory orphan genes from the genome. After silencing these ten genes, only the RcOSP1 knocked-down plant significantly inhibited the growth of R. cerealis. We then described RcOSP1 as an effector that could impair wheat biological processes and suppress pathogen-associated molecular pattern-triggered immunity in the infection process. These findings confirm that HIGS and SIGS can be practical tools for researching R. cerealis virulence genes. [Formula: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.