Rice Sheath Research Articles

Design, Synthesis, and Fungicidal Activity against Rice Sheath Blight of Novel N-Acyl-1,2,3,4-tetrahydroquinoline Derivatives.

To discover fungicides with novel targets, a series of N-acyl-1,2,3,4-tetrahydroquinoline (NATHQ) derivatives were designed and synthesized by linking the active substructure NATHQ moiety in aspernigerin with the O-benzyl oxime-ether scaffold in commercial agrochemicals. Target compound structures were identified using proton and carbon-13 nuclear magnetic resonance spectroscopies and high-resolution mass spectrometry. Preliminary bioassays indicated that at 40 mg/L, some target compounds exhibited moderate to considerable in vitro fungicidal activities against Rhizoctonia solani and Botrytis cinerea. In particular, compound 3j exhibited higher fungicidal activities both in vitro (EC50 = 0.733 mg/L) and in vivo (EC50 = 15.2 mg/L) against R. solani than the commercial fungicide prochloraz; therefore, it should be a promising fungicide candidate against rice sheath blight. Additionally, compound 3j exhibited good laccase inhibitory activity (73.2% at 200 mg/L). Molecular docking revealed that the bis-cyano-oxime-ether moiety of compound 3j exhibited an excellent binding mode with the laccase target protein and could be used as a lead compound for developing laccase inhibitors. The structural features of these NATHQ derivatives will provide inspiration for developing laccase inhibitors and discovering more effective fungicides to control agricultural diseases.

Aspergillus sclerotiorum Whole-Cell Biocatalysis: A Sustainable Approach to Produce 3-Hydroxy-phenazine 1-Carboxylic Acid from Phenazine 1-Carboxylic Acid

In green chemistry, filamentous fungi are regarded as a kind of robust microorganism for the biotransformation of natural products. Nonetheless, the screening of microorganisms is crucial for the effective biotransformation of natural products, such as phenazine compounds. The precursor metabolite of most phenazine derivatives in Pseudomonas spp. is phenazine-1-carboxylic acid (PCA), the key constituent of shenqinmycin, widely used to control rice sheath blight in southern China. In this study, a new fungus strain Aspergillus sclerotiorum was isolated, which can efficiently convert PCA into 3-hydroxy-phenazine 1-carboxylic acid (3-OH-PCA). Moreover, an effective whole cells biotransformation system was designed by screening optimal reaction conditions and carbon sources. Hence, Aspergillus sclerotiorum exhibited desirable adaptation by the consumption of different carbon sources and maximum whole-cell biomass (10.6 g/L DCW) was obtained as a biocatalyst from glucose. Optimal conditions for whole-cell biocatalysis of PCA were evaluated, including a PCA concentration of 1120 mg/L, a pH of 7.0, a temperature of 25 °C, a rotation rate of 200 rpm, and dry cell weight of 15 g/L for 60 h; thus, 1060 mg/L of 3-OH-PCA was obtained and the conversion efficiency of PCA was 94%. Hence, the results of the repeated batch mood revealed that the biotransformation efficiency of fungus pellets reduced with each subsequent cycle, but remained stable in all five cycles with the provision of a glucose supplement. These findings present the prospect of using filamentous fungi for the whole-cell biocatalysis of phenazine in enormous amounts and the efficient production of 3-OH-PCA. Moreover, these results laid the foundation for further research to disclose the genetic-based mechanism of the strain responsible for PCA biotransformation.

Induced resistance to rice sheath blight (Rhizoctonia solani Kühn) by β-amino-butyric acid conjugate of phenazine-1-carboxylic acid

Rice sheath blight caused by Rhizoctonia solani Kühn is a major fungal disease that plagues commercially grown rice. Occurring mainly in leaf sheaths and leaves, the disease leads to great losses in food production. β-amino-butyric acid (BABA) has been demonstrated to activate an induced resistance response and is a potent inducer of broad-spectrum disease resistance in different plant species. In this study, β-amino-butyric acid conjugate of phenazine-1-carboxylic acid (PCA) with prominent induced resistance to rice sheath blight was tested. The in vitro fungicidal activity, as well as in vivo efficacy, systemicity, induced resistance and defense enzyme activity of BABA conjugate of PCA against R. solani in rice seedlings was systematically evaluated. The results indicated that in vitro fungicidal activity of PCA-β-aminobutyric acid (4e) against R. solani was lower than that of PCA, but in vivo curative ability of 4e was the highest among all tested compounds. The systemicity tests in rice seedlings revealed that PCA did not possess phloem mobility, while 4e exhibited moderate phloem mobility but much lower thanα-amino-butyric acid conjugate of PCA (4d). In addition, Compound 4e showed the highest induced activity against rice sheath blight. The observed effects of defense enzymes help to explain this high level of induced activity. The current research results indicate that in rice seedlings, BABA conjugate of PCA induce observable resistance to rice sheath blight and exhibit moderate phloem mobility, which could be used as an induced resistance fungicide against rice sheath blight in commercial rice production. The BABA conjugate of PCA might provide a useful example of induced resistance to R. solani.

Antagonistic potential and growth promoting activities of novel indigenous strains of <i>Trichoderma</i>

Rice (Oryza sativa L.), as a cereal grain, is the most widely consumed staple food for a large part of the world’s human population, especially in Asia and Africa. Rice production is often subjected to several biotic and abiotic stresses. Rice sheath blight is one of the most destructive diseases causing economic losses in rice yields and affecting quality worldwide. Twenty soil samples were collected from the rhizosphere of rice crop from different regions of Northern Karnataka. Trichoderma spp. were isolated from the rhizospheric soil samples. The antagonistic potential of Trichoderma spp. was studied using dual culture technique. Among twenty strains of Trichoderma the highest inhibition (>50%) was recorded in nine strains which ranged from 54.20 to 65.10 per cent. Trichoderma harzianum recorded highest seed germination (100%), root length (13.73 cm), shoot length (8.64 cm) and seedling vigour index. In pot culture experiment, the Trichoderma strains significantly improved the growth parameters such as root length, shoot length and number of tillers per plant. Among the different treatments, seed treatment + root dipping + foliar spray with T. harzianum strain was highly effective in increasing the shoot length, root length and seedling vigour index at 30, 60 and 90 days after sowing. This strain was found highly effective in inhibiting the pathogen and promoting the growth of rice plants.

β-Glucanase Family Genes Promote Resistance to Sheath Blight in Rice by Inhibiting the Permeability of Plasmodesmata.

Rice sheath blight (ShB) caused by Rhizoctonia solani is one of the most serious diseases that threatens rice (Oryza sativa) production. However, the mechanisms of defense against ShB in rice remain largely unknown. In this study, we identified that the expression levels of β-glucanase (OsBGL) family genes sensitively respond to infection by R. solani, and OsBGLs positively regulate rice resistance to ShB. In addition, OsBGL2 colocalized with AtPDCB1 at the plasmodesmata (PD) and limited the PD permeability. The level of callose accumulation in osbgls mutants and overexpressors was examined, and OsBGLs were found contribute to callose accumulation. Taken together, these data suggest that OsBGLs can regulate the deposition of callose at the PD to reduce its permeability to defend itself against ShB. Through the identification of these genes and the elucidation of their functions, this research fills the gap in the mechanism of PD permeability in rice ShB resistance.

BZIP23 interacts with NAC028 to modulate rice resistance to sheath blight disease

Rice sheath blight disease (ShB) is a serious threat to rice production, and breeding ShB-resistance varieties is the most effective strategy for ShB control. However, the molecular mechanisms of rice resistance to ShB are largely unknown. In this study, the NAC transcription factor NAC028 was shown to be sensitive to ShB infection. ShB inoculation assays revealed that NAC028 is a positive regulator of ShB resistance. To elucidate the molecular basis of NAC028-mediated ShB resistance, another transcription factor (bZIP23) was identified as a NAC028-interacting protein. Results of the transcriptome and qRT-PCR analyses demonstrated that CAD8B, a key enzyme for lignin biosynthesis and ShB resistance, is regulated by both bZIP23 and NAC028. The combination of the yeast-one hybrid, ChIP-qPCR, and transactivation assays illustrated that both bZIP23 and NAC028 directly bind the CAD8B promoter and activate its expression. The transcriptional connection between bZIP23 and NAC028 was also investigated and the results of in vitro and in vivo assays demonstrated that NAC028 was one of the target genes of bZIP23, but not vice versa. The results presented here provide new insights into the molecular basis of ShB resistance and contribute to the potential targets for the ShB resistance breeding program.

Effect of enzyme-response Mag@CMCS@CPSG film on rice sheath blight disease (caused by Rhizoctonia solani) control and rice quality

To find a precise, efficient, and safe method to control rice sheath blight disease (caused by Rhizoctonia solani), this study used the natural product magnolol as the fungicide. Based on the characteristics of cell wall-degrading enzymes (CWDEs) released by R. solani during the early stages of infection, a Mag@CMCS@CPSG (magnolol@carboxymethyl chitosan@carboxymethyl cellulose, pectin, sodium alginate, and glycerol) film was constructed to control the release of magnolol in response to CWDEs. The Mag@CMCS@CPSG film increased magnolol toxicity against R. solani, decreased EC50 from 7.33 μg/mL to 4.37 μg/mL, and effectively blocked the invasion of mycelia. Under the stimulation of CWDEs released during R. solani infection, the cumulative release of magnolol by the Mag@CMCS@CPSG film reached 75.62% after 96 h, and the disease index of rice treated with Mag@CMCS@CPSG film significantly decreased. The maximum field control effect of Mag@CMCS@CPSG film on rice sheath blight disease was 72.47%, which was not significantly different from that of the commercial preparation jinggangmycin; however, the holding period was better. The Mag@CMCS@CPSG film reduced the loss of rice yield and quality caused by infection with R. solani and did not cause phytotoxicity or residue. In conclusion, the Mag@CMCS@CPSG film can achieve accurate and green control of rice sheath blight.

Jinggangmycin stimulates reproduction and increases CHCs-dependent desiccation tolerance in Drosophila melanogaster

Jinggangmycin (JGM), an agricultural antibiotic compound, is mainly used against the rice sheath blight (RSB) Rhizoctonia solani. However, its application may lead to unexpected consequences in insects. In this study, the effects of JGM on the physiological parameters of Drosophila melanogaster were investigated. The results showed that 0.005 g/ml JGM exposure increased female daily egg production and extended the oviposition period, while there was no significant effect on reproduction at 0.016 g/ml. At the same time, desiccation tolerance increased in flies fed 0.005 g/ml JGM. The RT-qPCR results revealed that FAS1 and FAS3 expression were upregulated in 0.005 g/ml JGM treated flies. Consistently, the amount of CHCs accumulated on the cuticle surface increased upon JGM treatment at 0.005 g/ml. Moreover, RNAi for FAS3 decreased desiccation tolerance of JGM-treated flies. These results suggest that JGM affects fatty acid biosynthesis, which in turn enhances reproduction and desiccation tolerance in Drosophila.

Effects of fungicides and antagonistic marine-derived fungi on rice seedling promotion and rice sheath blight control

Fourteen fungicides and three marine-derived fungi were tested for their activities in the promotion of rice seedling development and their efficacy in controlling rice sheath blight disease in two rice varieties, RD43 and KMDL105, under greenhouse conditions. The results showed that the rice seeds treated with carbendazim 50%SC and azoxystrobin 25%SC displayed the best promotion of rice growth in root length and shoot height at seven and 14 days in rice var. RD43. Rice seeds from var. KMDL105 treated with thiophanate-methyl 70%WP exhibited significant stimulation of rice seedling growth resulting in the greatest increase in root length and shoot height at seven and 14 days. In pot experiments with rice var. RD43, spraying with triforine 19%EC, propiconazole 25%EC, trifloxystrobin 50%WG, and validamycin 3%SL showed a high level of fungicidal activity in controlling sheath blight disease, with this evidenced by significant suppression of the disease severity by 71–75% and 61–73% at seven and 14 days after inoculation, respectively. The treatment of rice var. KMDL105 with validamycin 3%SL exhibited the best activity in controlling sheath blight disease by reducing disease severity by up to 62% and 64% at seven and 14 days, respectively. The antagonistic marine fungi tested in this study displayed low activity against sheath blight disease and low effects on rice seedling promotion compared to fungicides. Specifically, Trichoderma harzianum KUFA 0689 and T. asperellum KUFA 0677 reduced sheath blight disease severity by 42–46% and 27–37% at seven and 14 days, respectively.

Swietenia mahagoni Leaves Extract: Antifungal, Insecticidal, and Phytochemical Analysis

In this study, we investigated the antifungal properties of an acetone extract derived from the leaves of Swietenia mahagoni (SMAL) against two isolated fungi, Fusarium equiseti (OQ820153) and Rhizoctonia solani (OQ820152), from rice sheath. The extract was effective in inhibiting the growth of both fungi at the highest concentration tested, 3000 μg·mL−1. Laboratory tests on the LC20 of SMAL extract (49.86 mg·L−1) versus pyriproxyfen 10% EC (1.96 mg·L−1) were accomplished on Aphis gossypii Glover. The extract potently reduced the survival of the nymphs (49.58%) more than the other treatments. The longevity of nymphs treated with the extract had the highest prolongation at 9.67 days. The olfactory choice test exhibited the lowest aphid attraction percentage (23.33%). The HPLC of SMAL extract contained various phenolic compounds, and the most abundant found were catechin (752.64 µg·g−1), gallic acid, and chlorogenic acid, as well as flavonoids such as rutin (585.24 µg·g−1) and naringenin. A GC–MS analysis revealed n-hexadecanoic acid (37.1%) as the major compound, followed by oleic acid. These results suggest that SMAL extract has the potential to help plants fight against fungal and insect infections, making it a promising natural and renewable solution for long-term plant pest regulation.

Preparation of cinnamaldehyde nanoemulsions: Formula optimization, antifungal activity, leaf adhesion, and safety assessment

Plant essential oils have become an increasingly popular topic in agriculture due to their outstanding fungicidal activity, good biocompatibility and abundance of sources. Cinnamaldehyde is found in cinnamon essential oil and has good antifungal activity, but cinnamaldehyde also has negative characteristics such as high volatility, poor water solubility and easy oxidative degradation, which seriously hinder the potential of its antimicrobial properties and its application in agriculture. In this study, the effects of the surfactant ratio, concentration and emulsification method on the stability of cinnamaldehyde nanoemulsions were systematically investigated by observing the sample appearance, mean droplet size and differential scanning calorimetry (DSC) analysis. The results showed that the optimal formula of cinnamaldehyde nanoemulsion was a surfactant (R5:848) ratio of 1:1 and a concentration of 7%, and the emulsification method was E1: adding dropwise aqueous phase (deionized water) to the oil phase (cinnamaldehyde + surfactant). Cinnamaldehyde nanoemulsions showed good antifungal activity against rice sheath blight (Rhizoctonia solani), wheat sheath blight (Rhizoctonia cerealis) and wheat take-all (Gaeumannomyces graminis). The nanoemulsions also had excellent wetting properties on rice and wheat leaves, higher biosafety for nontarget environmental organisms such as zebrafish and earthworms, low cytotoxicity to human BASE-2B cells, and no inhibition of germination and growth of wheat and rice. The results of this research provide important ideas for the design of efficient, safe and environmentally friendly plant-derived fungicides.

Kurstakin Triggers Multicellular Behaviors in Bacillus cereus AR156 and Enhances Disease Control Efficacy Against Rice Sheath Blight.

Kurstakin is the latest discovered family of lipopeptides secreted by Bacillus spp. In this study, the effects of kurstakin on the direct antagonism, multicellularity, and disease control ability of Bacillus cereus AR156 were explored. An insertion mutation in the nonribosomal peptide synthase responsible for kurstakin synthesis led to a significant reduction of antagonistic ability of AR156 against the plant-pathogenic fungi Rhizoctonia solani, Ascochyta citrullina, Fusarium graminearum, and F. oxysporum f. sp. cubense. The loss of kurstakin synthesis ability significantly impaired the swarming motility of AR156 and reduced biofilm formation and amyloid protein accumulation. Although the loss of kurstakin synthesis ability did not reduce the competitiveness of AR156 under laboratory conditions, the colonization and environmental adaptability of the mutant was significantly weaker than that of wild-type AR156 on rice leaves. The cell surface of wild-type AR156 colonizing the leaf surface was covered by a thick biofilm matrix under a scanning electron microscope, but not the mutant. The colonization ability on rice roots and control efficacy against rice sheath blight disease of the mutant were also impaired. Thus, kurstakin participates in the control of plant diseases by B. cereus AR156 through directly inhibiting the growth of pathogenic fungi and improving long-term environmental adaptability and colonization of AR156 on the host surface by triggering multicellularity. This study explored the multiple functions of kurstakin in plant disease control by B. cereus.

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.

Nitrogen and Growth Stage Influence Epicuticular Wax Composition on Rice Leaf and Sheath

Epicuticular wax plays an important part in protecting plant aerial organs from biotic and abiotic stresses. Nitrogen is the key limiting nutrient for crops and influences metabolite synthesis. However, how nitrogen influences wax composition on rice leaf and sheath at different growth stages has not been systematically examined. In this study, a total of 19 wax compounds in rice leaf and sheath were detected by using gas chromatography–mass spectrometry. Permutational multivariate analysis of variance (PermANOVA) indicated a significant influence of both nitrogen and growth stage on rice leaf and sheath wax composition. Nitrogen influenced wax composition on leaf and sheath mainly at 70 and 110 d, while the growth stage significantly influenced wax composition under all nitrogen levels. Our study provides the first report that rice wax composition changes with nitrogen and growth stage. This may support exploring nitrogen application and the associated ecological effects.

Aspergillus sp. R3, a new producer for cyclopyazonic acid, inhibits rice sheath blight fungus Rhizoctonia solani Kühn

This study investigated the activity of the novel Aspergillus sp. R3 against rice sheath blight (RSB) fungus Rhizoctonia solani Kühn. R3 strain had strong antagonistic activity. R3 strain was Aspergillus sp. by morphological observation and molecular identification. After enrichment, separation, purification and nuclear magnetic resonance (NMR) identification of culture solution and mycelium of Aspergillus sp. R3, the main metabolite was identified as cyclopyazonic acid (CPA). We studied the effect of CPA on R. solani, including mycelial morphology, membrane permeability and lipid peroxidation. The control effect of CPA on rice seedlings infected with sheath blight was also tested. CPA inhibited the growth of R. solani in vitro and led to the atrophy of R. solani mycelia under microscopic observation. In addition, CPA exhibited antimicrobial activity against a variety of plant pathogens in vitro. With the treatment of CPA with R. solani, the malondialdehyde content and electrolyte leakage of R. solani mycelia increased compared with the control. Additionally, CPA could significantly reduced the severity of sheath blight disease of rice seedlings infected with R. solani. This is the first report of the CPA produced by Aspergillus sp. with antagonistic activity against R. solani and R. solani-induced RSB. This study indicates that Aspergillus sp. and its metabolites may serve as potential biocontrol agents against some plant pathogens.

In vitro/vivo antifungal activity study of novel mandelic acid derivatives as potential fungicides against Thanatephorus cucumeris

To discover highly efficient and novel lead compounds against Thanatephorus cucumeris, a series of novel mandelic acid derivatives containing 1,3,4-thiadiazole thioether was designed and synthesized. The bioassay results revealed that target compound F10 exhibited excellent antifungal activity against T. cucumeris with EC50 value of 9.7 μg/mL. Further studies found that F10 not only significantly inhibited the growth of T. cucumeris mycelia but also effectively inhibited the formation of sclerotia, and exhibited significant in vivo protective (61.1%) and curative (67.9%) activities at 200 μg/mL. Mechanism studies demonstrated that F10 can damage the integrity of the cell membrane structure, resulting in increased permeability of the cell membrane, releasing the intracellular electrolyte and inhibiting the growth of fungi. In general, this work is helpful for managing the formation and diffusion of the infection source and provides an effective method to control rice sheath blight disease infected with sclerotia of T. cucumeris.

Organic amendments and rice sheath blight management

Soil amended with rice straw compost significantly reduced rice sheath blight severity. The disease caused by the fungus Rhizoctonia solani Kühn is a severe disturbance to rice cultivation in Indonesia. R. solani is soil borne fungi and highly competitive as saprophyte. In mature compost, R. solani propagules in the form of sclerotia germination were suppressed by hyperparasites and other biological control agents. Compost maturity level affects the ability of sclerotia to germinate. Initial inoculum reduction as sclerotia in steril soil was studied in glass house which is treated with the addition of variations in the maturity of the compost. Result showed that mature compost added to sterile soil in the pot decreased sclerotia germination for about 14%, otherwise fresh organic matter added to steril soil increased sclerotia germination, this condition lead R. solani grew and new sclerotia formed in the soil surface. The decomposition levels of organic matter influences disease development, it significantly affected on disease severity. Disease severity of rice sheath blight decreased by about 32.6-39.9 % with mature compost application.

Rational Exploration of Novel SDHI Fungicide through an Amide-β-ketonitrile Bioisosteric Replacement Strategy

The identification of succinate dehydrogenase inhibitor (SDHI) fungicides bearing a novel scaffold is of great importance to control pathogenic fungi. Difluoromethyl-pyrazole β-ketonitrile derivatives were rationally designed through an innovative amide-β-ketonitrile bioisosteric replacement strategy and evaluated for their antifungal activities. In preliminary fungicidal screening, our new β-ketonitrile compounds showed outstanding in vitro activity. Compounds A7 and A14 exhibited EC50 values of 0.116 and 0.165 μg/mL against Sclerotinia sclerotiorum, respectively, and A14 also displayed an EC50 of 0.0774 μg/mL against Rhizoctonia solani. Furthermore, A14 exhibited moderate in vivo protective activity against rice sheath blight on rice plants. Results from SDH enzymatic assays demonstrated that A14 possesses significant inhibitory effect toward porcine heart SDH, with an IC50 value of 0.183 μM, which was 20-fold more potent than that of fluxapyroxad (IC50 = 3.76 μM). A docking study indicated that H-bonds, cation-π interactions, and edge-to-face π-π interactions play key roles in the binding of A14 with R. solani SDH. The CoMSIA model guided the approach to further structural optimizations and indicated that hydrophobic and steric substituents on the benzene ring have decisive effects on the fungicidal activity against R. solani. The present work describes for the first time the successful bioisosteric replacement of the common SDHI amide moiety by a β-ketonitrile group and highlights the potential of β-ketonitriles as an innovative novel SDHI subclass.

Progress in rice sheath blight resistance research.

Rice sheath blight (ShB) disease poses a major threat to rice yield throughout the world. However, the defense mechanisms against ShB in rice remain largely unknown. ShB resistance is a typical quantitative trait controlled by multiple genes. With the rapid development of molecular methods, many quantitative trait loci (QTLs) related to agronomic traits, biotic and abiotic stresses, and yield have been identified by genome-wide association studies. The interactions between plants and pathogens are controlled by various plant hormone signaling pathways, and the pathways synergistically or antagonistically interact with each other, regulating plant growth and development as well as the defense response. This review summarizes the regulatory effects of hormones including auxin, ethylene, salicylic acid, jasmonic acid, brassinosteroids, gibberellin, abscisic acid, strigolactone, and cytokinin on ShB and the crosstalk between the various hormones. Furthermore, the effects of sugar and nitrogen on rice ShB resistance, as well as information on genes related to ShB resistance in rice and their effects on ShB are also discussed. In summary, this review is a comprehensive description of the QTLs, hormones, nutrition, and other defense-related genes related to ShB in rice. The prospects of targeting the resistance mechanism as a strategy for controlling ShB in rice are also discussed.

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

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. 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. 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.