Utilization Situation and Prospect of Gene xa5 Against Pathotype Ⅴ of Rice Bacterial Blight

In recent years,the strong virulence pathotype Ⅴ of rice bacterial blight grew up quickly in Southern China,which has become a major population and spread to the rice regions of Jiangsu and Zhejiang provinces in Southern China.Since pathotype Ⅴ caused serious bacterial blight disease in rice production regions,it is urgent to breed and promote resistant varieties against pathotype Ⅴ.The most economic and effective measure to control rice bacterial blight is to breed resistant cultivars for widely planting using resistance genes.The Institute of Plant Protection,Guangdong Academy of Agricultural Sciences,and Agricultural Science Research Institute of Panyu District of Guangzhou used IRBB5 carrying the recessive xa5 gene from IRRI that is resistant to pathotype Ⅴ to breed resistant varieties with rice blast resistance source,through hybridization,multiple cross,pedigree selection and synchronous resistance evaluation.We successfully bred series of new resistant rice varieties such as Baijiangzhan,Baijingzhan and Baisizhan,which showed resistance to strong virulence pathotype Ⅴ(grade 1-3)of bacterial blight and rice blast(mediate to high resistance),good grain quality(level 3 of rice quality of Guangdong),and yield equivalent to major cultivars(compared with region trial control cultivars of Guangdong).These new resistant varieties were promoted and planted in the strong virulence pathotype Ⅴ region along the west coast of Guangdong,which showed favorable superiority and wide application prospect in controlling rice bacterial blight with resistance varieties.

Development of Spectral Features for Monitoring Rice Bacterial Leaf Blight Disease Using Broad-Band Remote Sensing Systems

As an important rice disease, rice bacterial leaf blight (RBLB, caused by the bacterium Xanthomonas oryzae pv.oryzae), has become widespread in east China in recent years. Significant losses in rice yield occurred as a result ofthe disease’s epidemic, making it imperative to monitor RBLB at a large scale. With the development of remotesensing technology, the broad-band sensors equipped with red-edge channels over multiple spatial resolutionsoffer numerous available data for large-scale monitoring of rice diseases. However, RBLB is characterized by rapiddispersal under suitable conditions, making it difficult to track the disease at a regional scale with a single sensorin practice. Therefore, it is necessary to identify or construct features that are effective across different sensors formonitoring RBLB. To achieve this goal, the spectral response of RBLB was first analyzed based on the canopyhyperspectral data. Using the relative spectral response (RSR) functions of four representative satellite or UAVsensors (i.e., Sentinel-2, GF-6, Planet, and Rededge-M) and the hyperspectral data, the corresponding broad-bandspectral data was simulated. According to a thorough band combination and sensitivity analysis, two novel spectralindices for monitoring RBLB that can be effective across multiple sensors (i.e., RBBRI and RBBDI) weredeveloped. An optimal feature set that includes the two novel indices and a classical vegetation index was formed.The capability of such a feature set in monitoring RBLB was assessed via FLDA and SVM algorithms. The resultdemonstrated that both constructed novel indices exhibited high sensitivity to the disease across multiple sensors.Meanwhile, the feature set yielded an overall accuracy above 90% for all sensors, which indicates its cross-sensorgenerality in monitoring RBLB. The outcome of this research permits disease monitoring with different remotesensing data over a large scale.

The Class Ⅲ peroxidase gene OsPrx30,transcriptionally modulated by the AT-hook protein OsATH1, mediates rice bacterial blight-induced ROS accumulation

Class Ⅲ peroxidases(CⅢ Prxs) play critical roles in plant immunity by scavenging reactive oxygen species(ROS). However, the functions of CⅢ Prxs in rice(Oryza sativa L.) immunity are largely unexplored. Here, we report a Prx precursor, OsPrx30,that is responsive to the bacterial blight Xanthomonas oryzae pv. oryzae(Xoo). OsPrx30 was primarily expressed in rice roots, leaves, and stems,and its protein product was mainly localized at the endoplasmic reticulum. Overexpression of OsPrx30 enhanced the plant’s susceptibility to Xoo by maintaining a high level of peroxidase(POD) activity and reducing the content of H2O2, whereas depletion of OsPrx30 had the opposite effects. Furthermore, we identified an AT-hook transcription factor, Os ATH1, that is specifically bound to the OsPrx30 promoter. As observed in plants overexpressing OsPrx30, depletion of Os ATH1 enhanced susceptibility to Xoo. Finally, we demonstrated that depletion of Os ATH1 increased histone H3 acetylation at the AT-rich region of the OsPrx30 promoter.Taken together, these results reveal a mechanism underlying the POD-induced natural resistance to bacterial diseases and suggest a model for transcription regulation of Prx genes in rice.

Achieving broad-spectrum resistance against rice bacterial blight through targeted promoter editing and pathogen population monitoring

Plant diseases severely reduce crop yields and threaten global food security.Broad-spectrum resistance(BSR)is a desirable trait because it confers resistance against more than one pathogen species or the majority of races/strains of the same pathogen.To control plant diseases,breeders have selected BSR to reduce disease occurrence and prolong the life-span of newly released cultivars in the last several decades(Mundt,Phytopathology 108(7):792–802,2018).Although effective,breeding of BSR cultivars in crop plants is still time-consuming and technically challenging.Recently,new gene-editing technologies such as CRISPR/Cas9 have dramatically accelerated the process of plant breeding and provided an approach for rapidly creating new varieties with BSR and other beneficial traits(Borrelli et al.,Front Plant Sci 9:1245,2018).In addition,close surveillance of pathogen populations in the field can provide useful information for the deployment of appropriate resistance genes in the target regions.In this mini-review,we focus on the significance and application of the exciting results from two recent companion papers published in Nature Biotechnology that provide new strategies to develop crop plants with BSR against pathogens through targeted promoter editing of susceptibility genes in plants as well as pathogen population monitoring.

Resistance Evaluation of Dominant Varieties against Rice Blast (Magnaporthe oryzae) and Bacterial Blight (Xanthomonas oryzae pv. oryzae) in Guangzhou, China

The resistance of dominant rice varieties in Guangzhou against rice blast(Magnaporthe oryzae)and bacterial blight(Xanthomonas oryzae pv.oryzae)were evaluated by artificial inoculation and field natural blast nursery identification.Among 11 varieties tested,seven varieties showed resistance to rice blast,accounting for 63.6%;two varieties were highly resistant,accounting for 18.2%;three varieties were resistant,accounting for 27.3%;two varieties were moderately resistant,accounting for 18.2%;four varieties were susceptible(moderately susceptible),accounting for 36.4%.Varieties showed different resistance against the prevailing pathotype IV of bacterial blight;one variety was resistant,accounting for 9.1%;three varieties were moderately resistant,accounting for 27.3%;five varieties were moderately susceptible,accounting for 45.5%;one variety was susceptible,accounting for 9.1%;one variety was highly susceptible,accounting for 9.1%;all varieties tested showed susceptible to the virulent pathotype V.Huanghuazhan,Yuejingsimiao 2 and Fengxiuzhan showed resistance against both bacterial blight(pathotype IV)and rice blast,accounting for 27.3%.It is helpful to reduce the production risk by extending such dominant rice varieties with good resistance to rice blast and bacterial blight.

Analyses of gene effects and inheritance of resistance to bacterial blight in rice

We analyzed the resistant inheritance of ajaponica variety,Jia23,to two bacterial blightpathogen strains,KS-6-6 and Zhe 173,rep-resenting respectively the two predominantpathogenic types(Ⅱ and Ⅳ)in the rice crop-ping area along the Yangtze River Valley.Jia23 was crossed with susceptible vari-eties,Ewan8 and 7416.Fplants were back-

Inheritance of resistance to bacterial streak and bacterial blight in rice

Bacterial streak (BS) caused by Xanthomonascampestris pv. oryzicola has become one of themajor diseases in southern rice areas. Up todate, there are few reports on the inheritanceof resistance to BS and its relationship with theresistance to bacterial blight (BB). Therefore,we analyzed the inheritance of resistant to BSin three resistant rice cultivars and the geneticrelationship of resistance between BS and BB.

Breeding of near-isogenic japonica rice lines with three major genes for resistance to bacterial blight

From 1986 to 1993, a set of near-isogenic japonicarice Iines with three major genes Xα-3, Xα - 4,and Xα-12 for resistance to bacterial blight(Xan-thomonas oryzae pv.oryzae)were developed anddesignated as CBB3, CBB4, and CBB12 respective-

Production of bacterial blight resistant lines from somatic hybridization between Oryza sativa L. and Oryza meyeriana L.

Novel bacterial blight (BB) resistance gene(s) for rice was (were) introduced into a cultivated japonica rice variety Oryza sativa (cv. 8411), via somatic hybridization using the wild rice Oryza meyeriana as the donor of the resistance gene(s). Twenty-nine progenies of somatically hybridized plants were obtained. Seven somatically hybridized plants and their parents were used for AFLP (amplified fragment length polymorphism) analysis using 8 primer pairs. Results confirmed that these plants were somatic hybrids containing the characteristic bands of both parents. The morphology of the regenerated rice showed characters of both O.sativa and O.meyeriana. Two somatic hybrids showed highest BB resistance and the other 8 plants showed moderate resistance. The new germplasms with highest resistance have been used in the rice breeding program for the improvement of bacterial blight resistance.

Evaluatien of Near-lsogenic Rice Lines with 8 Genes for Bacterial Blight Resistance to Strains in China

A set of near-isogenic rice lines withmonogenic resistance to bacterial blight weredeveloped by IRRI.The Cultivar IR24 wasused as the recurred parent.They wereevaluated with 6 races of Xanthomonascampestris pv.oryzae(Xco)in the Philip-pines at the maximum tillering and the bootingstages by ZHANG and MEW at IRRI in 1989.

RICE TRANSFORMATION WITH A PHYTOALEXIN GENE AND BIOASSAY OF THE TRANSGENIC PLANTS

Immature embryos, mature embryos and embryogenic calli of 6 rice (Oryza sativa L.) materials were transformed with particle bombardment. The plasmids pSSVstl and pVE5+ were used, both containing the phytoalexin gene from grapevine coding for stilbene synthase, but driven by 35S and its own promoter respectively. Through resistance selection for G418 (100 to 150 mg/L) or hygromycin (50 mg/L), 54 independent transgenic plants were isolated and further assessed by PCR, Southern blot and Dot blot analyses. The transgenic plants and their progenies were tested for resistance to blast ( Pyricularia oryzae ) and bacterial blight of rice ( Xanthomonas oryzae ). Preliminary results indicated that the stilbene synthase gene could enhance the resistance of transgenic plants and their progenies to both pathogens.

Establishment of a system for screening and identification of novel bactericide targets in the plant pathogenic bacterium Xanthomonas oryzae pv. oryzae using Tn-seq and SPR

Xanthomonas spp. cause severe bacterial diseases. However, effective strategies for prevention and management of these diseases are scarce. Thus, it is necessary to improve the efficiency of control of diseases caused by Xanthomonas. In this study, Xanthomonas oryzae pv. oryzae(Xoo), which causes rice bacterial leaf blight, has been studied as a representative. A transposon insertion library of Xoo, comprising approximately 200,000 individual insertion mutants, was generated. Transposon sequencing data indicated that the mariner C9 transposase mapped at 35.7–36.4% of all potential insertion sites, revealing 491 essential genes required for the growth of Xoo in rich media. The results show that, compared to the functions of essential genes of other bacteria, the functions of some essential genes of Xoo are unknown, 25 genes might be dangerous for the Xanthomonas group, and 3 are specific to Xanthomonas. High-priority candidates for developing broad-spectrum, Xanthomonas-specific, and environment-friendly bactericides were identified in this study. In addition, this study revealed the possible targets of dioctyldiethylenetriamine using surface plasmon resonance(SPR) in combination with high performance liquid chromatography–mass spectrometry(HPLC–MS). The study also provided references for the research of some certain bactericides with unknown anti-bacterial mode of action. In conclusion, this study urged a better understanding of Xanthomonas,provided meaningful data for the management of bacterial leaf blight, and disclosed selected targets of a novel bactericide.

Identification of an avirulence gene,avrxa5,from the rice pathogen Xanthomonas oryzae pv.oryzae

Xanthomonas oryzae pv.oryzae,the causal agent of bacterial blight in rice,interacts with rice plants in a gene-for-gene manner.The specificity of the interaction is dictated by avirulence(avr) genes in the pathogen and resistance(R) genes in the host.To date,no avr genes that correspond to recessive R genes have been isolated.We isolated an avrBs3/pthA family gene,avrxa5,from our previously isolated clone p58,which was originally from strain JXOIII.The avrxa5 gene converted the PXO99A strain from compatible to incompatible in rice cultivars containing the recessive xa5 gene,but not in those containing the dominant Xa5 gene.Sequencing indicated that avrxa5,which is highly similar to members of the avrBs3/pthA family,encodes a protein of 1238 amino acid residues with a conserved carboxy-terminal region containing three nuclear localization signals and a transcription activation domain.It has 19.5 34-amino-acid direct repeats,but the 13th amino acid is missing in the fifth and ninth repetitive units.Domain swapping of the repetitive regions between avrxa5 and avrXa7 changed the avirulence specificity of the genes in xa5 and Xa7 rice lines,respectively.This indicates that avrxa5 is distinct from previously characterized avrBs3/pthA members.The specificity of avrxa5 toward recessive xa5 in rice could help us better understand the molecular mechanisms of plant-pathogen specific interactions.

Dynamic and Coordinated Expression Changes of Rice Small RNAs in Response to Xanthomonas oryzae pv.oryzae

Endogenous small RNAs are newly identified players in plant immune responses, yet their roles in rice （Oryza sativa） responding to pathogens are still less understood, especially for pathogens that can cause severe yield losses. We examined the small RNA expression profiles of rice leaves at 2, 6, 12, and 24 hours post infection ofXanthomonas oryzae pv. oryzae （Xoo） virulent strain PXO99, the causal agent of rice bacterial blight disease. Dynamic expression changes of some miRNAs and trans-acting siRNAs were identified, together with a few novel miRNA targets, including an RLK gene targeted by osa-miR159a. 1. Coordinated expression changes were observed among some small RNAs in response to Xoo infection, with small RNAs exhibiting the same expression pattern tended to regulate genes in the same or related signaling pathways, including auxin and GA signaling pathways, nutrition and defense-related pathways. These findings reveal the dynamic and complex roles of small RNAs in rice-Xoo interactions, and identify new targets for regulating plant responses to Xoo.

Paladin, a tyrosine phosphatase-like protein, is required for XA21-mediated immunity in rice

XA21 encodes a rice immune receptor that confers robust resistance to most strains of the Gram-negative bacterium Xanthomonas oryzae pv.oryzae(Xoo).XA21-mediated immunity is triggered by recognition of a small protein called RaxX-sY(required for activation of XA21-mediated immunity X,tyrosine-sulfated)secreted by Xoo.To identify components regulating XA21-mediated immunity,we generated and screened a mutant population of fast-neutron-mutagenized rice expressing Ubi:Myc-XA21 for those susceptible to Xoo.Here,we report the characterization of one of these rice mutants,named sxi2(suppressor of XA21-mediated immunity-2).Whole-genome sequencing revealed that sxi2 carries a deletion of the PALADIN(PALD)gene encoding a protein with three putative protein tyrosine phosphatase-like domains(PTP-A,-B,and-C).Expression of PALD in the sxi2 genetic background was sufficient to complement the susceptible phenotype,which requires the catalytic cysteine of the PTP-A active site to restore resistance.PALD coimmunoprecipitated with the full-length XA21 protein,whose levels are positively regulated by the presence of the PALD transgene.Furthermore,we foundd that sxi2 retains many hallmarks of XA21-mediated immunity,similar to the wild type.These results reveal that PALD,a previously uncharacterized class of phosphatase,functions in rice innate immunity,and suggest that the conserved cysteine in the PTP-A domain of PALD is required for its immune function.