Global characterization of OsPIP aquaporins reveals that the H_(2)O_(2)transporter OsPIP2;6 increases resistance to rice blast

Plasma membrane intrinsic proteins(PIPs)are conserved plant aquaporins that transport small molecules across the plasma membrane to trigger instant stress responses and maintain cellular homeostasis under biotic and abiotic stress.To elucidate their roles in plant immunity to pathogen attack,we characterized the expression patterns,subcellular localizations,and H_(2)O_(2)-transport ability of 11 OsPIPs in rice(Oryza sativa),and identified OsPIP2;6 as necessary for rice disease resistance.OsPIP2;6 resides on the plasma membrane and facilitates cytoplasmic import of the immune signaling molecule H_(2)O_(2).Knockout of OsPIP2;6 increases rice susceptibility to Magnaporthe oryzae,indicating a positive function in plant immunity.OsPIP2;6 interacts with OsPIP2;2,which has been reported to increase rice resistance to pathogens via H_(2)O_(2)transport.Our findings suggest that OsPIP2;6 cooperates with OsPIP2;2 as a defense signal transporter complex during plant–pathogen interaction.

Application of UAV-Based Imaging and Deep Learning in Assessment of Rice Blast Resistance

Rice blast is regarded as one of the major diseases of rice.Screening rice genotypes with high resistance to rice blast is a key strategy for ensuring global food security.Unmanned aerial vehicles(UAV)-based imaging,coupled with deep learning,can acquire high-throughput imagery related to rice blast infection.In this study,we developed a segmented detection model(called RiceblastSegMask)for rice blast detection and resistance evaluation.The feasibility of different backbones and target detection models was further investigated.RiceblastSegMask is a two-stage instance segmentation model,comprising an image-denoising backbone network,a feature pyramid,a trinomial tree fine-grained feature extraction combination network,and an image pixel codec module.The results showed that the model combining the image-denoising and fine-grained feature extraction based on the Swin Transformer and the feature pixel matching feature labels with the trinomial tree recursive algorithm performed the best.The overall accuracy for instance segmentation of RiceblastSegMask reached 97.56%,and it demonstrated a satisfactory accuracy of 90.29%for grading unique resistance to rice blast.These results indicated that low-altitude remote sensing using UAV,in conjunction with the proposed RiceblastSegMask model,can efficiently calculate the extent of rice blast infection,offering a new phenotypic tool for evaluating rice blast resistance on a field scale in rice breeding programs.

Preventive and Therapeutic Effects of New Fungicides on Rice Blast

[Objectives]The paper was to identify the characteristics and the potency of new fungicides to rice blast disease,and study their preventive and therapeutic effects on the disease.[Methods]Six new and conventional fungicides with different mechanisms of action were selected to compare their preventive and therapeutic effects on rice seedling blast in greenhouse.[Results]The agents which showed to be highly efficacious in blast prevention(greater than 90%)were 25%pyraclostrobin EC,20%fenoxanil SC,75%tricyclazole WP and 50%prochloraz-manganese WP sprayed at the dosages of 112.5-150,180-360,270-450 and 450-600 g/ha,respectively.The therapeutic effect of new fungicides 25%pyraclostrobin EC and 50%prochloraz-manganese WP on rice blast were above 90%when the dosages were 90-150 and 360-600 g/ha,respectively.The therapeutic effect of the conventional fungicide 40%isoprothiolane EC on rice blast was 69.04%when the dosage was 480 g/ha,while those of other fungicides were worse.[Conclusions]Both 25%pyraclostrobin EC and 50%prochloraz-manganese WP have good preventive and therapeutic effects on rice blast,which will provide a reference for the use of these fungicides in the field.

Effects of β-Amino Butyric Acid Induced Rice Blast Resistance on Reactive Oxygen Metabolism

[Objective] This study was to understand the effects of β-Amino butyric acid(abbreviated as BABA) induced rice blast resistance on reactive oxygen metabolism. [Method] Using the cultivar Chaochan 2 that is highly susceptible to disease as experimental material, the changes of catalase(CAT), and superoxide dismutase(SOD) and MDA activities in rice treated by BABA were investigated. [Result] In rice plants treated by BABA, the activities of CAT and SOD increased, meanwhile the MDA content also rose to some extent, resulting in the disease resistance to rice blast. [Conclusion] By influencing reactive oxygen metabolism, BABA endows rice plants with resistance to rice blast. BABA is safe to environment and has highly resistance-inducing capacity, it could be generalized in production.

Genotypic Analysis of Rice Blast Resistance Genes Pi-ta and Pi-b in Japonica Rice Varieties and Lines in Jiangsu Province

Pi-ta and Pi-b, the first cloned rice blast resistant genes, have been wide- ly used in rice blast resistance breeding for their lasting and stable resistance. To define the distribution of Pi-ta and Pi-b in japonica rice in Jiangsu, the genotypes of resistance genes Pi-ta and Pi-b in 40 varieties and 665 new lines were detected using functional markers of Pi-ta/pi-ta and Pi-b^pi-b alleles. The results showed that the resistance alleles of Pi-ta and Pi-b were widely spread in japonica rice varieties, and the distribution frequency of Pi-b was higher than that of Pi-ta. Most of the Lianjing serial varieties didn＇t carry the two resistance genes, but the two resistance genes were widely distributed in Wujing serial varieties. There was no significant dif- ference in distribution frequency of Pi-ta between new lines and commercial vari- eties. However, the distribution frequency of Pi-b in new lines was higher than that in commercial varieties. It was indicated that artificial selection was conducive to the improvement of distribution frequency of Pi-b in rice varieties. Among the 4 genotypes, the distribution frequency of pi-taJPi-b was highest （60.0%）, followed by Pi-ta/ Pi-b （33.5%） and pi-ta/pi-b （3.9%）. The frequency of Pi-taJpi-b was lowest, account- ing for only 2.6%. In terms of source of resistance genes in the four combinations, the resistant allele Pi-ta might be from parents of Wuxiangjing14, Wujing15 or Nanjing44, and Pi-b might come from parents of Wujing13, Wuxiangjing14, Wujing15 or Nanjing44. The analysis on the genotypic frequencies in offspring of the rice vari- eties showed that the resistance genotype of Pi-ta/Pi-b had the highest frequency in the cross combination of Nanjing44//Wujing13/Kantou194.

Study on the Prediction of Rice Blast Based on the Unbiased GM (1,1) Model

To create a new prediction model, the unbiased GM （1,1） model is optimized by the five-point slide method in this paper. Then, based on the occurrence areas of dce blast in Enshi District during 1995 -2004, the new model and unbiased GM （1, 1 ） model are applied to predict the occurrence areas of rice blast during 2005 -2010. Predicting outcomes show that the prediction accuracy of five-point unbiased sliding optimized GM （1, 1 ） model is higher than the unbiased GM （1,1） model. Finally, combined with the prediction results, the author provides some suggestion for Enshi District in the prevention and control of rice blast in 2010.

Study on the Occurrence and Epidemic Regularity and Region Division of Rice Blast in Nanchong City

With the aim to improve the level of monitoring and warning as well as the comprehensive control of rice blast disease, and to feasibly reduce the disease threat in Nanchong City, the methods of GPS and GIS, systematical monitoring and field survey, rice blast resistance identification, physiologic races of rice blast monitoring, and meteorological data analysis were performed to study the occurrence and epidemic region division, precise demarcation and occurrence and epidemic regularity of rice blast in Nanchong City. This study first completed the epidemic region division and precise demarcation; first clarified the initial affection（beginning period） locations, occurrence characteristics, epidemic trends and characteristics; explicated the existence of four epidemic peak periods of rice blast in the field, where the damage areas of first peak period played a decisive role during the blast epidemic years; in late May, the cumulative occurrence areas and annual occurrence areas presented higher positive correlation with the correlation coefficient of 0.817;and established a prediction model of occurrence areas per year based on the disease field rate at the end of boot stages and the diseased plant rate at dough stages. The results of investigation on the impact factors investigation of blast disease in Nanchong in recent years suggested that the internal causes were the decrease or loss of blast resistance of the rice cultivars, as well as the increase of physiological races with strong resistance to rice blast and the emergence of new virulent physiologic varieties; the external causes were suitable temperature, too much rainy, and sunlight shortage. Between 2010 and 2015, the short-term forecast accuracy for rice blast in Nanchong was up to 100%, and medium-and long-term forecast accuracy was also up to 98% and 95%, respectively, which increased by 5-15% than that before 1997, thereby making the control effect of rice blast in Nanchong increased by 15-30%.

Comprehensive Prevention and Control Technologies of Rice Blast in Disease Nanchong City

The study was conducted to reduce blast damage, the use of pesticides residue, environment pollution and control costs, and to make a significant contribution to the improvement of grain production, quality and agriculture ecological environment. Over these years, by the methods of systematical monitoring, regular surveys, field investigation, rice blast resistance identification, experiments and meteorological data analysis, the study on comprehensive prevention and control of rice blast in Nanchong City was conducted. The results showed that the rice varieties more sensitive to blast had a higher incidence of severe blast disease. Replacing,varieties with different source of resistance every three to five years and reasonable variety distribution can effectively reduce the prevalence of rice blast. Appropriate treatment of infected rice straw and pathogen, seed disinfection, seedling disinfection, and pesticide application at transplanting and etc. can delay blast occurrence and reduce the damage caused by blast. By analyzing the blast control efficiency of pesticides applied at different growth stages, we found that best control efficiency against blast was achieved by spraying pesticide twice during the whole growth stage, once 3 d before transplanting or 10 d after transplanting; and once at initial heading stage. Spraying 525 g/hm^2 75% tricyclazole was proven to be the best dosage for blast control. However, 375-450 g/hm^2 75% tricyclazole is enough if the blast incidence is not severe, or the rice varieties are slightly susceptible to blast.The control efficiency against leaf blast between 4% kasugamycin and 20% tricyclazole had no significant difference, but was significantly higher than that of 100 billion spores/g of Bacillus subtilis. The control efficiency against neck blast had no significant difference among 4% kasugamycin, 20% tricyclazole and 100 billion spores/g of B. subtilis. 450 g/hm^2 75% tricyclazole had better control efficiency against neck blast than 2 250 g/hm^2 2% 800 million spores/g Jinggangmycin-wax bud bacteria SC, 1 050 g/hm^2 41% kasugamycin-isoprothiolane WP and 900 g/hm^2 41%kasugamycin-isoprothiolane WP. The frequency of severe blast incidence in Nanchong City has reached 50% since 1997. The rice blast disease has been effectively controlled by comprehensive prevention and control technology, reducing the production loss to less than 2%, and pesticides by 4 523 t in total. In 2014, 327000 t rice grains were approved as pollution-free, green and organic agricultural products, indicating that the blast control measures produced great economic, social and ecological values.

Incidence Law and Disease Symptom of Rice Blast and Its Control Technique

The article systematically summarizes the pathogen, incidence law and disease symptom of rice blast, and raises the integrated control technique for the disease, which will provide the certain theoretical basis for the control of the disease, being benefit for the actual production of rice in field.

Inhibition Effect of Antagonistic Fungal Strain BOS-013 against the Pathogen of Rice Blast (Pyricularia oryzae)

[ ObjeeUve ] To search for the beneficial microorganisms and their metabolic products with control effect against rice Mast, the paper studied the inhibi- tion effect of the fermentation broth of actinomycetes BOS-013 strain against the pathogen of rice blast （Pyricularia oryzae）. [ Method ] With P. oryzae as the target indicator fungus, the inhibition effect of the fermentation broth of BOS-013 strain and its extracts from different solvents against the pathogen were determined using cylinder plate method and filter paper method. [ Result] The results showed that the inhibition rate of the fermentation broth of BOS-013 strain with fungus amount of 4×10^8 cfu/ml against P. oryzae was 92.0% ±0.5% ; the fermentation broth still had inhibition effect against P. oryaae after inactivation under high temperature and high pressure, and the inhibition rate was 60.0%±0.5%. After absorbed by non-polar CAD-45 type macroperous adsorption resin and eluted by 60% etha- nol, the antifungal activity of the fermentation broth was still high, and the diameter of inhibition zone reached 33.0 mm. [ Conclusion] The fermentation broth of BOS-O13 strain had significant inhibition effect against P. oryzae. The results would provide foundation for further clarification of inhibition mechanism of BOS-013 strain fermentation broth against P. oryzae and its practical application.

Resistance Evaluation of Part of Leading Rice Varieties to Rice Blast in Guangxi Province

In order to determine the resistance of leading rice varieties to rice blast in Guangxi, three natural induction rice blast nurseries were established in Cenxi, Sanjiang and Yongfu. The resistance of the 45 leading rice varieties in Guangxi to rice blast was determined under conditions of spray inoculation and natural induction at seedling stage. The results showed that resistance frequencies of the 45 leading rice varieties ranged from 8.33% to 94.44%. Among the 45 tested varieties, there were 8 varieties of which the resistance frequencies were all higher than 80%, accounting for 17.78%; there were 11 varieties of which the resistance frequencies ranged from 50% to 80%, accounting for 24.44%; the resistance frequencies of the remaining varieties were all lower than 50%, accounting for 57.78% of the total tested varieties. The twoconsecutive-year determination results showed none of the 45 leading rice varieties showed resistance to rice blast. In 2010, there were 26, 12 and 7 varieties that were moderately sensitive, sensitive and greatly sensitive to rice blast respectively, accounting for 57.78%, 26.67% and 15.56% of the total tested varieties. In 2011, there were 16, 23 and 12 varieties that were moderately sensitive, sensitive and greatly sensitive to rice blast respectively, accounting for 35.56%, 51.11% and 13.33%. The overall resistance of the 45 leading rice varieties trended to be decreased year by year. So in rice production, they should be selected properly.

Molecular Cloning of Early-expressed cDNAs from Rice in Response to Infection by Rice Blast Fungus Magnaporthe grisea

Compatible and incompatible reactions in rice plants ( Oryza sativa L. cv. Shenxianggen No.4) were resulted from inoculation with two different virulent races of rice blast fungus ( Magnaporthe grisea (Hebert) Barr), and thus an effective infecting system was established between rice plants and the rice blast pathogen. Two cDNA clones that showed induced and temporal patterns in expression in the very early stage in response to infection of the fungus were obtained from the plants by use of differential display. Of the two cDNA clones, Fastresp_a was induced to express in both compatible and incompatible interactions although it was expressed earlier in the former reaction. The second one, Fastresp_b , was only expressed in incompatible interaction. Southern blot analysis of the rice genomic DNA indicated that both of the two clones were from genome of the plant. No significant homology to the two genes was found from the rice gene database. This suggested that they were novel genes in rice and may play important roles in rice resistant response to infection of rice blast fungus.

Strategy for Use of Rice Blast Resistance Genes in Rice Molecular Breeding

Rice blast is one of the most destructive diseases affecting rice production worldwide.The development and rational use of resistant varieties has been the most effective and economical measure to control blast.In this review,we summarized the cloning and utilization of rice blast resistance genes,such as Pi1,Pi2,Pi9,Pi54,Pigm and Piz-t.We concluded that three main problems in the current breeding of rice blast resistance are:availability of few R(resistance)genes that confer resistance to both seedling and panicle blast,the resistance effect of pyramided lines is not the result of a simple accumulation of resistance spectrum,and only a few R genes have been successfully used for molecular breeding.Therefore,novel utilization strategies for rice blast R genes in molecular breeding were proposed,such as accurately understanding the utilization of R genes in main modern rice varieties,creating a core resistant germplasm with excellent comprehensive traits,screening and utilizing broadspectrum and durable resistance gene combinations.Lastly,the trends and possible development direction of blast resistance improvement were also discussed,including new genes regulating resistance identified via GWAS(genome-wide association study)and improving rice blast resistance using genetic editing.

Race Specificity of Major Rice Blast Resistance Genes to Magnaporthe grisea Isolates Collected from indica Rice in Guangdong, China

Race-specific resistance and field resistance of 30 rice blast resistance monogenic lines derived from different resources were evaluated. The spectra of resistance to 163 Magnaporthe grisea isolates collected from indica rice in Guangdong Province, China ranged from 0.6% to 89.6%. Most of the monogenic lines showed a narrow resistance spectrum and high susceptibility in rice blast area, whereas the lines with Pikh and Pi1(t) had the broad resistance spectra of 89.6% and 82.2% respectively, showing a high and stable blast resistance in fields. According to the cluster analysis of specific resistance to 163 blast isolates tested, the 30 monogenic lines were divided into 15 groups, and based on the principal factor analysis, nine kinds of race-specific resistance were identified. Pik, Piz5, Pi9 and Pish can be used as candidate resistance genes for rice breeding since their specific resistance differed from those of the backbone parents in Guangdong, China. Gene pyramiding of Pikh [or Pi1(t)], Pi9 (or Piz5) and Pish (or Pita2) will be effective to obtain broad-spectrum blast resistance in rice breeding program in Guangdong, China. The strategies for studying and application of rice blast resistance genes were discussed.

Analysis of Resistant Spectrum to Rice Blast in Transgenic Rice Lines Introduced Lysozyme Gene from T4 Phage

The receptor cultivar Nan29 and thirty-six T5 rice lines derived from ten T0 generation transgen-ic plants harboring lysozyme gene were challenged in the greenhouse by inoculating 63 isolates belonging to 48 races of Magnaporthe grisea from Yunnan Province. The transgenic rice lines exhibited resistance to more than 72% of isolates inoculated in this experiment, and 38.1% (24 isolates) of them could infect the receptor cultivar Nan29. The results indicated that the transgenic rice lines possessed wide-spectrum resistance against various rice blast races and the resistant spectrum of rice lines were different although some lines derived from same T0 plant. The transgenic rice lines exhibited also high resistance to leaf and neck blast in the disease field evaluation, but not all of resistant lines against leaf blast were resistant to neck blast.

The Use of Rice Varietal Diversity for Rice Blast Control

Field experiments of mixed- (intercropping) and pure-planting (monoculture) of four rice varieties, representing improved hybrid varieties (Shanyou63 and Shanyou22) and high-quality traditional varieties (Huangkenuo and Zigu) from Yunnan Province, were conducted based on their differences in genetic background and agro-economical characteristics. The results demonstrated that the mixed-planting of the hybrid rice and high-quality traditional rice varieties had a significantly greater effect on controlling rice blast disease than the monoculture of these varieties, particularly the traditional ones. It is evident for the highly susceptible traditional varieties in mixed-planting to achieve disease control, with significant decreases in blast incidences and severity indexes. The blast control efficiency reached up to 83-98% under such planting model. This suggests that an appropriate mixed-planting of rice varieties with diverse genetic background and agro-economical characteristics is an effective approach for rice blast control. In addition, resistance of the traditional rice varieties to lodging was considerably increased in the plots with mixed-planting, compared with the plots with monoculture. The average rate of grain-yield increase ranged from 6. 5 to 9.7% in the plots with mixed-planting.

Rice Blast Resistance of Transgenic Rice Plants with Pi-d2 Gene

Resistance to rice blast of transgenic rice lines harboring rice blast resistance gene Pi-d2 transformed from three different expression vectors of pCB6.3kb, pCB5.3kb and pZH01-2.72kb were analyzed. Nine advanced-generation transgenic rice lines with Pi-d2 gene displayed various resistance to 39 rice blast strains, and the highest disease-resistant frequency reached 91.7%. Four early-generation homozygous transgenic lines with Pi-d2 gene exhibited resistance to more than 81.5% of 58 rice blast strains, showing the characteristic of wide-spectrum resistance. The transgenic embryonic calli selected by the crude toxin of rice blast fungus showed that the callus induction rate of immature embryo from transgenic rice plants decreased as the concentration of crude toxin in the culture medium increased. When the concentration of crude toxin reached 40%, the callus induction rate of immature embryo from transgenic lines was 49.3%, and that of the receptor control was 5%. The disease incidence of neck blast of the transgenic rice lines in fields under induction was 0% to 50%, indicating that the rice blast resistance of transgenic rice lines is much higher than that of the receptor control.

Functional genomics in the rice blast fungus to unravel the fungal pathogenicity

A rapidly growing number of successful genome sequencing projects in plant pathogenic fungi greatly increase the demands for tools and methodologies to study fungal pathogenicity at genomic scale. Magnaporthe oryzae is an economically important plant pathogenic fungus whose genome is fully sequenced. Recently we have reported the development and application of functional genomics platform technologies in M. oryzae. This model approach would have many practical ramifications in design and implementation of upcoming functional genomics studies of filamentous fungi aimed at understanding fungal pathogenicity.

Genetic Transformation of Rice with Pi-d2 Gene Enhances Resistance to Rice Blast Fungus Magnaporthe oryzae

The gene Pi-d2, conferring gene-for-gene resistance to the Chinese blast strain ZB15, was isolated from a rice variety （Digu） by the map-based cloning strategy. Here, we constructed a control plasmid pZH01-pi-d2tp309 （pZH01-tp309） and three different expression constructs, pCB-Pi-d25.3kb （pCB5.3kb）, pCB-Pi-d26.3kb （pCB6.3kb） and pZH01-Pi-d22.72kb （pZH01-2.72kb） of Pi-d2, driven by Pi-d2 gene’s own promoter or CaMV35S promoter. These constructs were separately introduced into japonica rice varieties Lijiangxintuanhegu, Taipei 309, Nipponbare and Zhonghua 9 through Agrobacterium- mediated transformation. A total of 150 transgenic rice plants were obtained from the regenerated calli selected on hygromycin. PCR, RT-PCR and Southern-blotting assay showed that the gene of interest had been integrated into rice genome and stably inherited. Thirty-five transgenic lines independently derived from T1 progeny were inoculated with the rice blast strain ZB15. Transformants exhibited resistance to rice blast at various levels. The lesions on the transgenic plant leaves were less severe than those on the controls and the resistance level of transgenic plants harboring the gene of interest from three vectors had no difference. The own promoter of Pi-d2, about 2.2 kb or 3.2 kb, had the similar promoter function as CaMV35S. Field evaluation for three successive years supported the results of artificial trial, and some lines with high resistance to rice leaf blast and neck blast were obtained.

Effects of Silicon Fertilizer on Rice Blast (Magnaporthe oryzae) Green Control, Rice Growth and Yield

Three doses of silicon fertilizer and seven dominant rice varieties were selected to study the effects of silicon fertilizer on rice blast(Magnaporthe oryzae)green control,rice growth and yield.The results showed application of silicon fertilizer had certain control effect on panicle blast,and was beneficial for rice growth and yield.However,application effect varied among different varieties,and application of silicon fertilizer at the dose of 225 kg/hm2 received the best effect.Applying silicon fertilizer to control rice blast is safe and environmental-friendly,which is beneficial to the production of green food production.