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.

Codominance Functional Marker of Bacterial Blight Resistance Gene Xa7 in Rice

[Objectives]A codominance functional marker of the broad-spectrum bacterial blight resistance gene,Xa7,of rice was identified for accurate detection,generation tracking,and differentiation between homozygous and hemizygous genotypes of the gene.[Methods]A potential functional marker containing four primers was designed using Premier 5 software and based on the differences on the sequences of Xa7,xa7,and allele-free genomes.The molecular distinctness of the marker in different materials was verified by PCR.Three crossbreed lines of Xa7 and their parents were inoculated with seven bacterial blight strains at the booting stage to examine the affected agronomic traits at maturation.[Results]The homozygous R084 of Xa7 could be amplified into a 91 bp band and the Nip free of allele with a 153 bp band,while the heterozygote Nip/R084,91 bp and 153 bp bands.The candidate codominance marker,Xa7fun,amplified fragments that matched the predicted target bands.No 91 bp fragment was amplified from 18 germplasms of varied types,indicating a lack of Xa7 in them.Whereas Ry1,Ry2 and Ry3 had a 91 bp band,suggesting the inclusion of homozygous Xa7.Under an elevated temperature,Huazhan responded to the seven bacterial blight pathogens as highly susceptible(HS),intermediate susceptible(MS),or susceptible(S);R084 to six of the seven pathogens(HNA1-4,FuJ,GDA2,GD1358,PX086,and YN24)as highly resistant(HR),intermediate resistant(MR)or resistant(R);Ry-1 to five pathogens(GDA2,HNA1-4,FuJ,GD1358,and YN24)as HR or MR;Ry-2 to five pathogens(GDA2,GD1358,HNA1-4,PXO86,and YN24)as HR or R;and Ry-3 to 6 pathogens(HNA1-4,FuJ,GDA2,GD1358,PXO86,and YN24)as HR or MR.Therefore,the infiltration of Xa7 in the improved crossbred lines RY-1,RY-2,and RY-3 significantly accentuated the blight resistance of Huazhan.[Conclusions]Homozygous or hemizygous Xa7 could be accurately differentiated by the currently identified codominance functional marker Xa7 fun.The Xa7 introgression did not significantly alter the critical agronomic traits in the hybridization from generation to generation and could be safely applied in breeding rice varieties with bacterial blight resistance.

Molecular Screening of Rice Cultivated in Benin for the Identification of Xanthomonas oryzae Pv. oryzae and Bacterial Leaf Blight Resistance Genes

One of the most devastating diseases of rice worldwide is bacterial blight (BLB) caused by Xanthomonas oryzae pv. Oryzae (Xoo). In Benin, Xoo was first described in 2013 on wild rice Oryzae longistaminata. So far, no study has been done on Beninese Xoo strains. We do not know whether the pathogen has already passed into the rice varieties grown, or if they are exposed to other bacteria. Whereas the use of resistant varieties, carrying resistance genes, is the only highly effective and environmentally friendly way to control this disease, no information is available on these Xoo resistance genes in rice varieties grown in Benin apart from the one we recently. This study aims to identify Beninese Xoo strains, causing BLB and screen rice varieties grown in Benin for the main resistance genes. Diseased rice leaves showing typical symptoms of fire blight collected from different rice fields in the three phytogeographic areas of Benin were analyzed by PCR for Xoo-specific sequence identification. Furthermore, seventy-five collected rice accessions were screened to identify xa5, Xa7, xa13, and Xa21 resistance genes to Xoo. The results reveal that Xanthomonas oryzae was identified in two fields in Banikouara and one in Malanville. On the other hand, Sphingomonas sp. has been identified in several other rice fields in Benin. Forty-seven of seventy-five rice accessions examined (62.66%) carried Xoo resistance genes with 3 (4%) and 40 (53.33%) of xa5 and Xa21 respectively. None of the accessions had either Xa7 or xa13 resistance genes. Three accessions possess both xa5 and Xa21 genes. Isogenic lines IRBB60 and IRBB21, supposed to be a positive control, presented a Xoo sensitivity allele. These results indicate that Xoo has moved from the wild rice variety to the cultivated variety in northern Benin and varietal improvement programs must be implemented with varieties having several resistance genes for the efficient response against a possible BLB pandemic in Benin.

Resistance of Antimicrobial Peptide Gene Transgenic Rice to Bacterial Blight

Antimicrobial peptide is a polypeptide with antimicrobial activity. Antimicrobial peptide genes Np3 and Np5 from Chinese shrimp （Fenneropenaeus Chinensis） were integrated into Oryza sativa L. subsp, japonica cv. Aichi ashahi by Agrobacterium mediated transformation system. PCR analysis showed that the positive ratios of Np3 and Np5 were 36% and 45% in To generation, respectively. RT-PCR analysis showed that the antimicrobial peptide genes were expressed in T1 generation, and there was no obvious difference in agronomic traits between transgenic plants and non-transgenic plants. Four Np3 and Np5 transgenic lines in T1 generation were inoculated with Xanthomonas oryzae pv. oryzae strain CR4, and all the four transgenic lines had significantly enhanced resistance to bacterial blight caused by the strain CR4. The Np5 transgenic lines also showed higher resistance to bacterial blight caused by strains JS97-2, Zhe 173 and OS-225. It is suggested that transgenic lines with Np5 gene might possess broad spectrum resistance to rice bacterial blight.

Marker-Assisted Pyramiding of Genes Conferring ResistanceAgainst Bacterial Blight and Blast Diseases into Indian RiceVariety MTU1010

Two major bacterial blight （BB） resistance genes （Xa21 and xa13） and a major gene for blastresistance （Pi54） were introgressed into an Indian rice variety MTU1010 through marker-assistedbackcross breeding. Improved Samba Mahsuri （possessing Xa21 and xa13） and NLR145 （possessingPi54） were used as donor parents. Marker-assisted backcrossing was continued till BC2 generationwherein PCR based functional markers specific for the resistance genes were used for foregroundselection and a set of parental polymorphic microsatellite markers were used for background selectionat each stage of backcrossing. Selected BC2F1 plants from both crosses, having the highest recoveriesof MTU1010 genome （90% and 92%, respectively）, were intercrossed to obtain intercross F1 （ICF1） plants,which were then selfed to generate 880 ICF2 plants possessing different combinations of the BB andblast resistance genes. Among the ICF2 plants, seven triple homozygous plants （xa13xa13Xa21Xa21Pi54Pi54）with recurrent parent genome recovery ranging from 82% to 92% were identified. All the seven ICF2plants showed high resistance against the bacterial blight disease with a lesion lengths of only 0.53–2.28 cm, 1%–5% disease leaf areas and disease scoring values of ‘1’ or ‘3’. The seven ICF2 plants wereselfed to generate ICF3, which were then screened for blast resistance, and all were observed to behighly resistant to the diseases. Several ICF3 lines possessing high level of resistance against BB andblast, coupled with yield, grain quality and plant type on par with MTU1010 were identified and advanced forfurther selection and evaluation.

Marker-Assisted Selection of Xa21 Conferring Resistance to Bacterial Leaf Blight in indica Rice Cultivar LT2

Bacterial leaf blight of rice (BLB), caused by Xanthomonas oryzae pv. oryzae, is one of the most destructive diseases in Asian rice fields. A high-quality rice variety, LT2, was used as the recipient parent.IRBB21, which carries the Xa21 gene, was used as the donor parent. The resistance gene Xa21 was introduced into LT2 by marker-assisted backcrossing. Three Xoo races were used to inoculate the improved lines following the clipping method. Eleven BC_3F_3 lines carrying Xa21 were obtained based on molecular markers and agronomic performance. The 11 lines were then inoculated with the three Xoo races. All the 11 improved lines showed better resistance to BLB than the recipient parent LT2. Based on the level of resistance to BLB and their agronomic performance, five lines (BC_3F_3 5.1.5.1, BC_3F_3 5.1.5.12, BC_3F_3 8.5.6.44, BC_3F_3 9.5.4.1 and BC_3F_3 9.5.4.23) were selected as the most promising for commercial release. These improved lines could contribute to rice production in terms of food security.

Evaluation of Inherited Resistance Genes of Bacterial Leaf Blight, Blast and Drought Tolerance in Improved Rice Lines

Improved rice lines were developed frome three parents with the resistance or tolerance to bacterial leaf blight,blast and drought stress,respectively,using single-,double-and three-way crosses.The improved lines were assessed for agro-morphological and yield traits under non-drought stress(NS)and reproductive-stage drought stress(RS)treatments.The mean comparison of traits measured between parent plants and progenies(improved lines)were similar,and there were significant and non-significant differences among the parents and improved lines(genotypes)under NS and RS.Smilarly,there was significant and non-significant differences in the interaction among both parent varieties and improved lines for NS and RS.Cluster and 3D-model of principal component analysis did not generate categorical clusters according to crossing methods,and there were no exclusive crossing method inclined variations under the treatments.The improved lines were high-yielding,disease resistant,and drought-tolerant compared with their parents.All the crossing methods were good for this crop improvement program without preference to any,despite the number of genes introgressed.

In vitro and in silico studies of salicylic acid on systemic induced resistance against bacterial leaf blight disease and enhancement of crop yield

Salicylic acid(SA)is an effective elicitor to promote plant defenses and growth.This study aimed to investigate rice(Oryza sativa L.)cv.Khao Dawk Mali 105 treated with salicylic acid(SA)-Ricemate as an enhanced plant protection mechanism against bacterial leaf blight(BLB)disease caused by Xanthomonas oryzae pv.oryzae(Xoo).Results indicated that the use of SA-Ricemate as a foliar spray at concentrations of more than 100 mg L^(-1)can reduce the severity of BLB disease by 71%.SA-Ricemate treatment also increased the hydrogen peroxide(H_(2)O_(2))content of rice leaf tissues over untreated samples by 39–61%.Malondialdehyde(MDA)in rice leaves treated with SA-Ricemate also showed an increase of 50–65%when comparing to non-treated samples.The differential development of these defense compounds was faster and distinct when the SA-Ricemate-treated rice was infected with Xoo,indicating plant-induced resistance.Besides,SA-Ricemate elicitor at a concentration of 50–250 mg L^(-1)was correlated with a substantial increase in the accumulation of total chlorophyll content at 2.53–2.73 mg g^(-1)of fresh weight which suggests that plant growth is activated by SA-Ricemate.The catalase-and aldehyde dehydrogenase-binding sites were searched for using the CASTp server,and the findings were compared to the template.Chemsketch was used to design and optimize SA,which was then docked to the catalase and aldehyde dehydrogenase-binding domains of the enzymes using the GOLD 3.0.1 Software.SA is shown in several docked conformations with the enzymes catalase and aldehyde dehydrogenase.All three catalase amino acids(GLN7,VAL27,and GLU38)were discovered to be involved in the creation of a strong hydrogen bond with SA when SA was present.In this mechanism,the aldehyde dehydrogenase amino acids LYS5,HIS6,and ASP2 were all implicated,and these amino acids created strong hydrogen bonds with SA.In field conditions,SA-Ricemate significantly reduced disease severity by 78%and the total grain yield was significantly increased which was an increase of plant height,tiller per hill,and panicle in three field trials during Aug–Nov 2017 and 2018.Therefore,SA-Ricemate can be used as an alternative elicitor on replacing harmful pesticides to control BLB disease with a high potential of increasing rice defenses,growth,and yield components.

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-

Screening of Effective Resistance Genes and Resistant Rice Parents against Rice Bacterial Blight(Xanthomonas oryzae pv. Oryzae) in Guangxi Province

[ Objective ] The paper was to confirm the resistance genes and resistant parents of rice against bacterial blight that could be used in Guangxi Province. [ Method] The dominant pathogenic types Ⅳ of Xanthomonas Oryzae pv. Oryzae in Guangxi were inoculated on a set of monogenic rice lines, the main hybrid rice parents in Guangxi and some important rice germplasm resources, and its resistant and susceptible conditions were investigated. [ Result ] IRBBS, IRBB7 and CBB23 were the resistant rice parents with resistance against pathogenic type IV, which contained resistance genes xa5, Xa7 and Xa23, respectively, and were identified to be the effective resistance genes against pathogenic type Ⅳ of X. Oryzae in Guangxi. [ Conclusion] The results provided basis for resistance breeding against bacterial blight.

Pyramiding blast,bacterial blight and brown planthopper resistance genes in rice restorer lines

Rice blast, bacterial blight （BB） and brown planthopper （BPH） are the three main pests of rice. This study investigated pyr-amiding genes resistant to blast, BB and BPH to develop restorer lines. Ten new lines with blast, BB and/or BPH resistance genes were developed using marker-assisted selection （MAS） technique and agronomic trait selection （ATS） method. Only HR13 with resistance genes to blast, BB and BPH was obtained. In addition to blast and BB resistance, four lines （HR39, HR41, HR42, HR43） demonstrated moderate resistance to BPH, but MAS for BPH resistance genes were not conducted in developing these four lines. These data suggested that there were unknown elite BPH resistance genes in the Zhongzu 14 donor parent. A more effective defense was demonstrated in the lines withPi1 andPi2 genes although the weather in 2012 was favorable to disease incidence. Blast resistance of the lines with a single resistance gene,Pita, was easily inlfuenced by the weather. Overal, the information obtained through pyramiding multiple resistance genes on developing the restorer lines is helpful for rice resistance breeding.

Genetics and Improvement of Bacterial Blight Resistance of Hybrid Rice in China

Since 1980s, rice breeding for resistance to bacterial blight has been rapidly progressing in China. The gene Xa4 was mainly used in three-line indica hybrid and two-line hybrid rice. The disease has been ＇quiet＇ for 20 years in China, yet in recent years it has gradually emerged and been prevalent in fields planted with newly released rice varieties in the Changjiang River valley. Under the circumstances, scientists inevitably raised several questions： what causes the resurgence and what should we do next？ And/or is resistance breeding still one of the main objectives in rice improvement？ Which approach do we take on resistance breeding so that the resistance will be more durable, and the resistance gene will be used more efficiently？ A combined strategy involving traditional method, molecular marker-assisted selection, and transgenic technology should bring a new era to the bacterial blight resistance hybrid rice breeding program. This review also briefly discusses and deliberates on issues related to the broadening of bacterial blight resistance, and suitable utilization of resistance genes, alternate planting of available resistance genes; and understands the virulent populations of the bacterial pathogen in China even in Asia.

Genetic Improvement of Rice for Bacterial Blight Resistance:Present Status and Future Prospects

The production and productivity of rice has been challenged due to biotic and abiotic factors.Bacterial blight(BB)disease,caused by Xanthomonas oryzae pv.oryzae,is one of the important biotic stress factors,which reduces rice production by 20%-50%.The deployment of host plant resistance is the most preferred strategy for management of BB disease,and breeding disease resistant varieties remains a very economical and effective option.However,it is difficult to develop rice varieties with durable broad-spectrum resistance against BB using conventional approaches alone.Modern biotechnological tools,particularly the deployment of molecular markers,have facilitated the cloning,characterization and introgression of BB resistance genes into elite varieties.At least 46 BB resistance genes have been identified and mapped from diverse sources till date.Among these,11 genes have been cloned and characterized.Marker-assisted breeding remains the most efficient approach to improve BB resistance by introducing two or more resistance genes into target varieties.Among the identified genes,xa5,xa13 and Xa21 are being widely used in marker-assisted breeding and more than 70 rice varieties or hybrid rice parental lines have been improved for their BB resistance alone or in combination with genes/QTLs conferring tolerance to other stress.We review the developments related to identification and utilization of various resistance genes to develop BB resistant rice varieties through marker-assisted breeding.

Improving the resistance of the rice PTGMS line Feng39S by pyramiding blast,bacterial blight,and brown planthopper resistance genes

Knowledge of rice(Oryza sativa L.)genes and various DNA markers can be used in genomic breeding programs aimed at developing improved elite rice cultivars.We used an efficient genomic breeding approach to pyramid four resistance genes(Pi2,Xa23,Bph14,and Bph15)in the popular photoperiod-and thermosensitive genic male sterile(PTGMS)rice line Feng39S.We performed foreground selection for the target genes,followed by recombinant selection and background selection.This process reduced the sizes of the genomic segments harboring the target genes(566.8 kb for Pi2,1143.9 kb for Xa23,774.7 kb for Bph14,and 1574.9 kb for Bph15)and accelerated the recovery of the recurrent parent genome to proportions ranging from 98.77%to 99.16%,thus resulting in four near-isogenic lines.To assemble the four resistance genes in Feng39S,we performed a double-way cross combined with foreground and background selection to generate two improved lines of Feng39S(Pi2+Xa23+Bph14+Bph15)with a recurrent parent genome recovery of 98.98%.The two lines showed agronomic performance,grain quality,and fertility–sterility transition characteristics similar to those of the original Feng39S line.The newly developed PTGMS lines and corresponding hybrid combinations were resistant to various field blast isolates and seven representative isolates of bacterial blight.At the seedling stage,the lines also showed resistance against brown planthopper.This study provides an efficient and accurate genomic breeding approach for introducing desirable traits into PTGMS lines.

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.

Breeding Rice Restorer Lines with High Resistance to Bacterial Blight by Using Molecular Marker-Assisted Selection

Two bacterial blight （BB） resistance genes, Xa21 and Xa4, from IRBB24 were introduced into hybrid rice restorer line Mianhui 725, which is highly susceptible to BB, by using hybridization and molecular marker-assisted selection technology. Four homologous restorer lines were obtained through testing the R target genes with molecular markers and analyzing parental genetic background. Inoculation of the four lines and their hybrids with the specific strains of Xanthomonas oryzae pv. oryzae, P1, P6 and seven representative strains of Chinese pathotype, C Ⅰ -CⅦ, showed that all of the four lines and their hybrids were highly resistant and presented broad resistance-spectrum to BB. The hybrids of G46A / R207-2 displayed good agronomic characters and high yield potential, and R207-2 was named Shuhui 207.

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.

Application of Functional Markers to Identify Genes for Bacterial Blight Resistance in Oryza rufipogon

Field resistances of nine accessions of common wild rice （Oryza rufipogon Griff.） and one rice variety （IR24） were evaluated by using nine strains of bacterial blight pathogen （Xanthomonas oryzae pv. oryzae） from the Philippines. IR24 was highly susceptible to all the strains, and six common wild rice accessions resisted all the nine strains, with a resistance frequency of 67%. The accessions Yulin and Wanning were only susceptible to PXO280 and PXO71, respectively. The accession Gaozhou was susceptible to the three strains PXO79, PXO99 and PXO339, whereas resistant to the other six strains. It could be concluded that there is at least one resistance gene in each common wild rice accession. The functional markers of the genes xa5, xa13, Xa21 and Xa27 were used to detect the presence of these resistance genes in the nine tested wild rice accessions, and it was found that four wild rice accessions contained heterozygous xa13. Among the nine common wild rice accessions, five were homozygous for Xa27 and three homozygous for xa27, and the accession Laibin contained neither xa27 nor Xa27. In addition, there were no xa5 and Xa21 in all of these accessions.