Intron Retention Fine-Tunes the Resistance of the Rice Mutant pls4 to Rice Sheath Blight(Rhizotonia solani AG I.1a)

OsPLS4 encodes aβ-ketoacyl carrier protein reductase(KAR).The role of OsPLS4 in rice sheath blight(Rhizoctonia solani)remains unclear.Our preliminary studies showed that premature leaf senescence mutants(pls4)were highly susceptive to sheath blight in the early stage of rice development.To explore the role of this gene in the development of rice sheath blight,the transcriptome profiles of the rice pls4 mutant and wild type were compared by RNA-seq.The results revealed 2,569 differentially expressed genes(DEGs).The down-regulated genes were significantly enriched in the defense response-related biological processes.These down-regulated genes included the chitinase genes and WRKY genes,which were significantly changed in pls4 mutants.Furthermore,467 genes induced significant alternative splicing(AS)events.Among them,intron retention(IR)affected gene expression levels and functions of the vitamin B6(VB6)metabolism pathway related to sheath blight.This result suggests that IR plays an important role in the sheath blight resistance of mutant pls4.Together,these results indicate that pls4 could be involved in the biological process of sheath blight via DEGs and the fine-tuning of IR.The present study provides a molecular basis for further investigation of the resistance of rice to sheath blight.

The Effect of Three New Fungicides against Rice Sheath Blight in Field Experiment

[Objective] The aim was to study the effect of three new fungicides against rice sheath blight in field experiment. [Methods] The experiment set up 7 treatments with three times of repetition and designed by random grouping. By using 5 sampling points in each plot, and investigating continuous 4 holes of each point, total plants, diseased plants and disease degrees were recorded. Then disease index and control efficiency were calculated, and variance analysis was carried out. [Results] 300 or 450 ml/hm^2 azoxystrobin ＋ difenoconazole 325 g/L SC had better control efficiency to rice sheath blight and had no phytotoxicity effect, we should use it at the initial disease stage and continuously spray 2-3 times. [Conclusion] The experiment provided a theoretical basis for controlling rice sheath blight using fungicides.

Research on the Pathogen of Rice Sheath Blight(Rhizoctonia solani Kühn)and Its Pathogenicity in Sichuan Regions

[ Objective] The paper was to study the cultural characteristic of the pathogen of Rice Sheath Blight （ Rhizoctonia solani Kuhn） and its pathogenicity in Sichuan regions. [ Method] The samples of rice sheath blight collected from six main rice planting areas in Sichuan regions were separated. The separated pathogen of rice sheath blight was cultured on PDA medium, and its cultural characteristic was recorded. Meanwhile, the pathogenicity of the obtained 23 strains was determined. [ Result] The growth rates among different strains had significant difference. According to the growth rate, only one strain belonged to medium strain（colony diameter： 40 mm≤（Ф≤60 mm）, the rest were all slow-type strains （colony diam- eter Ф〈40 mm）, and there was no strain with fast growth rate （colony diameter Ф 〉 60 mm）. Pathogenicity test showed that the pathogenicity among strains was significantly different, only one strain had strong pathogenicity, and the others all had moderate or weak pathogenicity. [ Conclu- sion] The study confirmed the basic biological characteristics of the pathogen of rice sheath blight in Sichuan region, which would provide theoretical basis for effective control of rice sheath blight in the region.

Control Efficacy of Validamycin A·(0.2 Billion Spores/ml) Paenibacillus polymyxa DN-1 3% AS for Rice Sheath Blight

With validamycin A.（0.2 billion spores/ml） Paenibacillus polymyxa DN-1 3% AS as the test agent, the effects of different dosage and different application time on the control efficacy for＇ rich sheath blight were investigated. The results of two- year test showed that when the application amount was in the range of 45-90 g.a.i/ hm^2, the field efficacy of validamycin A-（0.2 billion spores/ml） P. polymyxa DN-1 3% AS in the initial infection stage of rich sheath blight （Le., the diseased plant rate was below 5%） reached 80.38%-89.06%, and that in the peak infection stage （i.e., the diseased plant rate was higher than 10%） reached only 41.12%-53.26%. The field efficacy of validamycin A.（0.2 billion spores/ml） P. polymyxa DN-1 3% AS at the early onset of rich sheath blight was significantly better than that at the onset, so that the application time of validamycin A .（0.2 billion spores/ml） P. polymyxa DN-1 3% AS should be appropriately brought forward in the prevention and control of rice sheath blight.

Occurrence Regulation and Control Measure Comparison of Rice Sheath Blight

Rice sheath blight is widely distributed throughout the world, it can cause damage on many kinds of crops. Pathogen characteristics, infection cycle, occurrence law, damage law and different control methods of rice sheath blight in China are summarized, which will provide foundation for further worldwide research on the disease.

Efficacy of Difenoconazole-azoxystrobin 32.5% SC in Treatment of Rice Sheath Blight and Its Application Technology

[Objective] The efficacy of difenoconazole-azoxystrobin 32.5% SC for rice sheath blight and its application technology were discussed in this research. [Method] Three surveys were carried out. There were 5 fixed test clumps in each plot. The number of total plants and disease plants and disease progression in each fixed clump were recorded. The correlation effectiveness was calculated based on the growth rate of disease index. Significance analysis was performed with Dun- can＇s new multiple range method （DMRT）. [Result] The difenoconazole-azoxystrobin 32.5% SC had a good efficacy for rice sheath blight, and its efficacy increased with the increase of dose. If sprayed according to the dose of 450 ml/hm2 5 days before the beginning of heading stage of rice, difenoconazole-azoxystrobin 32.5% SC would have better effects on controlling rice sheath blight with correlation effectiveness reaching up to 95.14%, which was 2.06% higher than that of control drug （trifloxys- trobin-tebuconazole 75% WG, 225 g/hm2）. [Conclusion] Treated with difenoconazole- azoxystrobin 32.5% SC with dose of 450 ml/hm2, rice would have green upper leaves and less yellow middle and lower leaves. Moreover, the yield was in- creased significantly. The difenoconazole-azoxystrobin 32.5% SC had good safety. Therefore, difenoconazole-azoxystrobin 32.5% SC had a good application prospect in production.

Influencing Factors on Rice Sheath Blight Epidemics in Integrated Rice-Duck System

Sheath blight, a disease caused by the fungus Rhizoctonia solani Kuhn （anamorph）, has been the most economically significant disease of rice. It was frequently reported that the disease was well-controlled in integrated rice-duck system without the employment of fungicides. However, the effecting factors behind this phenomenon were rarely reported. In this research, experiment was carried out between two treatments, rice combined with ducks （RD） and conventional rice field without ducks rearing （CK） in early season rice paddy, to investigate the variations of sclerotia in floodwater and on rice plant, microclimate 10 cm above the waterline in rice paddy and activity of protective enzymes in rice plants. The results showed that the floating sclerotia in floodwater in RD was 86-91% lower than that in CK, and adhering sclerotia in rice plant in RD was 67-78% lower than that in CK. The relative humidity tested significantly lower and light intensity tested significantly higher in RD. The temperature in the early rice growth stages in RD was slightly lower than that in CK, but it was significantly higher （32.3-36.5~C） in the middle stage rice growth stages. The polyphenoloxidase （PPO） activity in RD were lower than that in CK, but the enhanced activity of phenylalanine ammonia-lyase （PAL）, peroxidase （POD） and Chitinase was observed in different stages of rice growth in RD, especially the Chitinase which showed higher activity in all investigating days.

Secondary metabolites of rice sheath blight pathogen Rhizoctonia solani Kühn and their biological activities

Eight compounds were isolated from the fermentation cultures of rice sheath blight pathogen Rhizoctonia solani Kuhn. They were identified as ergosterol （1）, 6β-hydroxysitostenone （2）, sitostenone （3）, m-hydroxyphenylacetic acid （4）, methyl m-hydroxyphenylacetate （5）, m-hydroxymethylphenyl pentanoate （6）, （Z）-3-methylpent-2-en-1,5-dioic acid （7） and 3-methoxyfuran-2-carboxylic acid （8） by means of physicochemical and spectroscopic analysis. Among them, 2, 3, 5-8 were isolated from R. solani for the first time. All the compounds were evaluated for their biological activities. 4-6 and 8 showed their inhibitory activities on the radical and germ elongation of rice seeds. 1,4 and 7 showed moderate antibacterial activity to some bacteria. 4, 7 and 8 exhibited weak inhibitory activities on spore germination of Magnaporthe oryzae. 8 showed moderate antioxidant activity with the 1,1-diphenyl-2-picryhydrazyl （DPPH） andβ-carotene-linoleic acid assays. This is the first time to reveal compounds 5, 6 and 8 from rice sheath blight pathogen R. solani to have in vitro phytotoxic activity.

Establishment of Agrobacterium tumefaciens-Mediated Transformation System for Rice Sheath Blight Pathogen Rhizoctonia solani AG-1 IA

To construct the T-DNA insertional mutagenesis transformation system for rice sheath blight pathogen Rhizoctonia solani AG-1 IA,the virulent isolate GD118 of this pathogen was selected as an initial isolate for transformation.The conditions for transformation of isolate GD118 were optimized in five aspects,i.e.pre-induction time,co-culture time,acetosyringone(AS) concentration at the co-culture phase,co-culture temperature and pH value of induction solid medium(ISM) at the co-culture phase.Finally,a system of Agrobacterium tumefaciens-mediated transformation(ATMT) for R.solani AG-1 IA was established successfully.The optimal conditions for this ATMT system were as follows:the concentration of hygromycin B at 30 μg/mL for transformant screening,8 h of pre-induction,20 h of co-culture,200 μmol/L of AS in ISM,co-culture at 25 ℃ and pH 5.6 to 5.8 of ISM at the co-culture phase.The transformants still displayed high resistance to hygromycin B after subculture for five generations.A total of 10 randomly selected transformants were used for PCR verification using the specific primers designed for the hph gene,and the results revealed that an expected band of 500 bp was amplified from all of the 10 transformants.Moreover,PCR amplification for these 10 transformants was carried out using specific primers designed for the Vir gene of A.tumefaciens,with four strains of A.tumefaciens as positive controls for eliminating the false-positive caused by the contamination of A.tumefaciens.An expected band of 730 bp was amplified from the four strains of A.tumefaciens,whereas no corresponding DNA band could be amplified from the 10 transformants.The results of the two PCR amplifications clearly showed that T-DNA was indeed inserted into the genome of target isolate GD118.

Characteristics of Resistance to Rice Sheath Blight of Zhongda 2,a Transgenic Rice Line as Modified by Gene “RC24”

The transgenic rice, Zhongda 2, which was genetically modified from an indica rice line Zhuxian B by rice chitinase gene (RC24), had high resistance to rice sheath blight (Rhizoctonia solan!) in laboratory and a two-year field experiment. The pathogen could invade sheath of Zhongda 2 and induce symptoms of the disease. No difference was noted in time of penetration or incubation period between Zhongda 2 and non-transgenic rice control, Zhuxian B, but the hyphae lysate could be observed earlier than control. Its resistance expressed as to inhibit the growth of mycelium in host tissue. Fis from Zhongda 2(4) crossed with other five non-transgenic rice lines showed higher resistance than donor non-transgenic parents, but the resistance was different along with the different maternal parents.

Influence of different inoculation methods on the evaluation of the resistance to rice sheath blight(Rhizoctonia solani Kuhn)

To establish a standard system for geneticstudies on sheath blight resistance, a field testwas conducted at the experimental farm ofYangzhou University to compare several pro-cedures for inoculating rice plants with R.solani Kuhn (RH9). The varieties used wereJasmine 85, Teqing (resistant or moderatelyresistant), and Lemont (susceptible). They

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

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

Study on the antibiosis of exudate JK-91-b against rice sheath blight (SHB)

Antagonistic bacterial strain 91-b possessedstronger antagonistic ability against SHBinfection. The control effects of the bacterizationagainst SHB were 58.1% in greenhouse and 45.1%in paddy. The effect of antibiosis JK-91-b exudat-ed by strain 91-b was studied in the present re-search.Bacterial strain 91-b was inoculated into

Verification test on an innovated method for the studies on inheritance of resistance to rice sheath blight

We have recently developed a systematic method for the study on the inheritance of resistance to sheath blight. The key of the system is an innovated method of inoculation and investigation along with the employment of the permanent population. This paper reported the procedure of the system and the result of its verification.

Advances in Mapping Loci Conferring Resistance to Rice Sheath Blight and Mining Rhizoctonia solani Resistant Resources

Sheath blight （SB） caused by Rhizoctonia solani is one of the three major diseases of rice, and now has become the most severe disease causing rice yield loss in China. Breeding and use of varieties resistant to SB is crucial in controlling the disease, but the advances achieved have been limited due to the tack of highly SB-resistant rice germplasm. Genetic analysis revealed that the SB resistance in rice was a typical quantitative trait controlled by multi-genes. Although many QTLs conferring resistance to SB have been identified in recent years, most of the QTLs only showed small effects and few of them have been evaluated for utilization potential. Many R. solani-resistant resources have been found in wild rice species, microorganisms and other plant species. It is already known that the SB-resistance could be improved in transgenic rice plants by genetic transformation. This paper reviewed the genetic mapping of loci associated with resistance to rice SB, the evaluation of the potential of resistance QTLs, and the resistant resources found in various organisms besides rice. To develop SB-resistant rice varieties, it is important to develop and explore new resistant rice germplasms, fine map and evaluate resistance QTLs, and also to pay attention to various bio-resources showing resistance to R. solani.

Control Effects of Jinggangmycin and Its Compounded Reagents against Rice Sheath Blight

The control effects of six kinds of reagents against rice sheath blight were compared in the test. The results showed that 24% thifluzamide SC 300 mL/hm2 and 20% Jinggangmycin ~ prochloraz WP 750 g/hm2 had good control effect against rice sheath blight, and their control effects after spraying for 7 and 14 d were 72.16%, 77.91% and 66.52%, 71.08%, respectively. The control effects of both reagents were significantly better than other test reagents and control re- agent Jinggangmycin, and they had no adverse effect on growth and development of rice. Considering from the aspects of control effect and economy, 20% Jinggang- mycin ~ prochloraz WP was suggested to be used to control rice sheath blight, its recommended dosage was 600 -750 g/hm2, and the reagent volume was 750kg with coarse spraying.

Diversity of ds RNA Viruses Infecting Rice Sheath Blight Fungus Rhizoctonia solani AG-1 IA

Rice sheath blight,caused by Rhizoctonia solani(Kühn),is a notorious soil-borne disease prevalent in many rice-growing regions.Although several sporadic studies of mycoviruses in R.solani AG-1 IA have been reported for single strain of R.solani AG-1 IA,there have been no reports describing the distribution and diversity of mycoviruses in natural populations.In this study,43 R.solani AG-1 IA strains collected from different locations in China were examined for the presence of ds RNA elements to confirm the presence of viral infections.Electrophoretypes showed that 16 of the 43 fungal strains(37.2%)contained ds RNAs that can be characterized as viruses.Furthermore,the species-specific reverse transcription PCR(RT-PCR)showed ds RNA bands with similar sizes do not always contain the same virus but exist as mixed mycoviral infections.Thus,our findings indicate mycoviruses infecting R.solani AG-1 IA in China are diverse,widespread and universal.

Analyses of the Temporal Development and Yield Losses due to Sheath Blight of Rice (Rhizoctonia solani AG1.1a)

Sheath blight of rice, caused by Rhizoctonia solani AG 1. 1a, has become the most important disease and caused serious yield losses in some major rice-growing regions in China in recent years. In the present study, field plot experiment was conducted to examine the relationships between disease intensity and inoculum density （ID）, the seasonal disease epidemic dynamics, and yield reductions due to disease damages. Results from the experiment demonstrated that the areas under progress curves of disease severity and those of percent rice tillers diseased were positively and closely related to the relative initial ID of the pathogen. The inoculum density-disease （IDD） relationships were simulated and the impractical linear models were obtained. Both logistic and Gompertz functions could be used to simulate the disease progress dynamics in time, but the progress curves of the disease severity were modeled better by the Gompertz than by logistic function. However, the Richards function was found to be the best in simulating the disease progress curves when a most appropriate value was chosen for the shape parameter m by using the computer software Epitimulator. Sheath blight infection decreased rice yield very significantly and a yield reduction of 40% was recorded in rice crop with the highest inoculum density. Rice yield was linearly and negatively correlated with the disease severity and the percent tillers affected. The simulated models for all these relationships were computed through executing Epitimulator software and were presented in this paper.

Enrichment of bacteria involved in the nitrogen cycle and plant growth promotion in soil by sclerotia of rice sheath blight fungus

Rice sheath blight pathogen,Rhizoctonia solani,produces numerous sclerotia to overwinter.As a rich source of nutrients in the soil,sclerotia may lead to the change of soil microbiota.For this purpose,we amended the sclerotia of R.solani in soil and analyzed the changes in bacterial microbiota within the soil at different time points.At the phyla level,Proteobacteria,Acidobacteria,Bacteroidetes,Actinobacteria,Chloroflexi and Firmicutes showed varied abundance in the amended soil samples compared to those in the control.An increased abundance of ammonia-oxidizing bacterium(AOB)Nitrosospira and Nitrite oxidizing bacteria(NOB)i.e.,Nitrospira was observed,where the latter is reportedly involved in the nitrifier denitrification.Moreover,Thiobacillus,Gemmatimonas,Anaeromyxobacter and Geobacter,the vital players in denitrification,N2O reduction and reductive nitrogen transformation,respectively,depicted enhanced abundance in R.solani sclerotia-amended samples.Furthermore,asymbiotic nitrogen-fixing bacteria,notably,Azotobacter as well as Microvirga and Phenylobacterium with nitrogen-fixing potential also enriched in the amended samples compared to the control.Plant growth promoting bacteria,such as Kribbella,Chitinophaga and Flavisolibacter also enriched in the sclerotia-amended soil.As per our knowledge,this study is of its kind where pathogenic fungal sclerotia activated microbes with a potential role in N transformation and provided clues about the ecological functions of R.solani sclerotia on the stimulation of bacterial genera involved in different processes of N-cycle within the soil in the absence of host plants.

Application Technology of Tebuconazole WG against Sheath Blight 75% Trifloxystrobin · Rice Blast and Rice

[ Objective] The paper was to confirm the best application period and frequency of 75% trifloxystrobin ·tebuconazole WG against rice blast and rice sheath blight. [ Method] Influences of different dosages and different application periods of 75% trifloxystrobin · tebuconazole WG on control effects against rice blast and rice sheath blight, as well as their effects on rice yields were studied in the paper. [Result] The control effects of three different fungicides application treatments against rice sheath blight were 80.24%, 83.0% and 67.99%, and the control effects against rice blast were 56.4%, 49.11% and 61.1%, respective- ly. Advanced application of fungicide for two times had good prevention effect against rice sheath blight ; properly delayed application of fungicides for two times was conducive to improving the control effect against rice blast, and one time application of sufficient fungicide during middle booting stage had higher control effect than application for two times. Effective panicle number per unit area, total grain number, 1 000-grain weight and moisture content of various fungicide application treat- ments did not have significant difference with those of the treatments without fungicide application, but grain number per panicle in treatments applied with fungicide was higher than the treatment without application. [ Conclusion] 75% Trifloxystrobin ·tebuconazole WG has better control effect on rice blast and sheath blight, which helps to promote the formation of rice grain and increases yield significantly. The fungicide application against rice sheath blight should be appropriately ad- vanced, and application for one time against panicle blast after middle booting stage is helpful to improve the control effect.