**1. Introduction**

World population is expected to rise beyond 9 billion by 2050 [1]. Rice (*Oryza sativa*) is a staple food crop world-wide, providing about one fifth of the calories consumed by humans [2]. In particular, rice accounts for 35–75% of the calories consumed by more than 3 billion in Asian countries alone and planted on approximately 154 million hectares land annually [3]. Crop protection and food security go hand in hand, and breeding for resistance against crop diseases remains the essential ingredient for food security. Due to the labor-intensive nature of breeding, integrated disease control is often reduced to mere chemical control, leaving the very purpose of this environment-friendly approach in limbo. Advances in molecular tools in crop breeding, however, makes breeding an increasing sustainable effort in staying ahead of pathogen adaptation [4]. Bacterial blight (BB) of rice is a widespread vascular disease caused by *Xanthomonas oryzae* pv. *oryzae*

(*Xoo*). Epidemics can severely reduce grain yield due to collapse of the entire crop [5]. BB was first characterized in the late nineteenth century [6]. Introduction of resistance (*R*) genes into rice cultivars is considered as the best option for *Xoo management*. A total of 42 *R* genes have been identified in rice against *Xoo, and the number continues to grow* [7–9]. Due to co-evolution and selection pressure between *Xoo* and rice, these *R* genes are selective in their efficiency against specific *Xoo* strains or races, which are sets of strains that share incompatibility on defined sets of *R* genes [10].

## **2. Post genomic era and rice grain protection**

Advancements in genomics, referring here to DNA and RNA analyses, is as beneficial to crop protection as is to other discipline of biology. Rice MetaSysB, an open source which provides detailed information about BB-responsive genes, is based on the global expression analysis. The database provided 7475 unique genes and 5375 simple sequence repeats, which were responsive to *Xoo* in rice [11]. Such information is based on the compatible and incompatible rice-*Xoo* interactions. In another example, 454 and 498 differentially expressed genes were reported as exemplified by the incompatible and compatible rice-*Xoo* interactions, respectively, using cDNA microarray [12]. Genomics also provides functional information of genes up- and downstream of candidate resistance genes in the defense signal pathway, as is done in near-isogenic rice lines introgressed with *Xa39*, an as yet uncharacterized BB resistance gene [13].

Multiple rice and *Xoo* genomes have been sequenced, either in draft or complete form [14–23], paving the way to identify functional connections between host and pathogen genes. The functional validation of the candidate genes is helping develop new rice varieties by introduction of the gene of interest through traditional breeding, marker assisted breeding, or genetic engineering approaches [3]. BB disease resistance is overcome by the emergence of more virulent strains of *Xoo*. Whole genome sequencing of 100 *Xoo* strains from India revealed that these strains were distinct from African and US *Xoo* strains [24]. Based on the reaction towards ten major resistance genes of rice, 46 out of the 100 strains were grouped into 11 pathotypes [24].

## **3. The genetic context of rice-***Xoo* **interaction**

Many BB-resistance genes in modern rice germplasm were selected long before the concepts of modern plant breeding were established, and a rich assortment of major dominant and recessive *R* genes has been identified by genetic and molecular studies (**Table 1**).

Perhaps the best known of these genes, *Xa21* represents the receptor kinase (RLK) class of *R* genes. *Xa21* was originally introgressed into rice from the related species *O*. *longistaminata* and confers resistance to a broad range of *Xoo* strains [25]. *Xa26*, another cloned member of RLK gene family, also confers broad resistance with a somewhat different strain profile [26]. The cognate elicitor for Xa21 has been reported [27]. However, for *Xa26* has not been identified.

RLKs play a central role in disease immunity pathways in plants, largely via the characterization of the bacterial flagellin receptor FLS2 and the related receptor EFR in *Arabidopsis* [28, 29]. A typical RLK consists of an extracellular receptor domain comprising of leucine-rich repeats (LRRs), a transmembrane domain, and an intracellular kinase domain [30]. As a class, RLKs have great potential for

**111**

*Disease Resistance and Susceptibility Genes to Bacterial Blight of Rice*

*Xa21* RLK1 extracellular,

*Xa26* RLK similar to *Xa21*; same

*Xa1*, *Xo1* NBS-LRR2 cytoplasm; narrow

*Xa27*, *Xa23*, *Xa10* TAL effector inducible membrane and cell

of *TFIIAγ5*; small subunit of TFIIA transcription factor

*OsSWEET11*; nodulin

promoter mutant of OsSWEET13, nodulin

complex

3 family

3 family

*NBS-LRR, nucleotide binding site, leucine-rich repeat.*

*xa5* Missense mutant

*xa13* promoter mutants of

*xa25*, *OsSWEET13Kit*

*RLK, receptor linked kinase.*

*WAK, wall-associated kinase.*

*Cloned R genes to bacterial blight of rice.*

*1*

*2*

*3*

**Table 1.**

**Gene Class Comments Cognate** 

membrane and intracellular domains; kinase; broad resistance

locus as *Xa3*; broad resistance

wall; novel protein; broad resistance

nuclear; broad resistance

membrane;

TATA box polymorphisms; unresponsive to PthXo2

unresponsive to PthXo1

resistance

*Xa4* WAK3 narrow unknown [40]

**elicitor/ effector**

Multiple TALes

AvrXa27, AvrXa23, AvrXa10

TALe interference **Ref**

[31–33]

[37–39]

[51, 53, 54]

RaxX [25, 27]

Unknown [26]

PthXo1 [42, 47]

PthXo2 [44, 52]

enhancing resistance to BB in rice and in other disease complexes of crop plants *Xa21*, *Xa26*, and other RLKs represent genetic components of the pathogenassociated molecular patterns (PAMPs)-triggered immunity (PTI) surveillance pathway in rice. Improvements in the rationale design of RLK receptor specificities, and screening for novel genes in germplasm or wild relatives could lead to general

The nucleotide binding site-LRR (NBS-LRR) is another large class of *R* gene, represented in rice toward *Xoo* by *Xa1* and *Xo1* [31–33]. XA1 and XO1 recognize multiple TALe, and *Xoo* strains have adapted TALes, the so-called iTALes, that are

Specific TALe-dependent *R* genes governing dominant resistance in rice against *Xoo* are known as executor (*E*) genes. *E* genes are distinct from classical *R* genes, whose transcriptional activation by TALes of *Xoo* trigger immunity, leading to dominant resistance [35]. *Xa27* represents the E genes class of dominant *R* genes and confers broad resistance to BB in rice [36]. Although not expressed in susceptible host, *Xa27* is expressed only upon inoculation with *Xoo* strains harboring the TALe gene *avrXa27* [37]. The protein is localized to apoplastic space, cell membrane and cell wall, and when expressed under a pathogen-nonspecific inducible rice *OsPR1* promoter, conferred constitutive resistance to both compatible and incompatible

application for broad and durable resistance.

truncated and inhibit the function of XA1 and XO1 [32, 34].

*DOI: http://dx.doi.org/10.5772/intechopen.86126*


*Disease Resistance and Susceptibility Genes to Bacterial Blight of Rice DOI: http://dx.doi.org/10.5772/intechopen.86126*

*2 NBS-LRR, nucleotide binding site, leucine-rich repeat.*

*3 WAK, wall-associated kinase.*

### **Table 1.**

*Protecting Rice Grains in the Post-Genomic Era*

**2. Post genomic era and rice grain protection**

**3. The genetic context of rice-***Xoo* **interaction**

reported [27]. However, for *Xa26* has not been identified.

of *R* genes [10].

resistance gene [13].

pathotypes [24].

studies (**Table 1**).

(*Xoo*). Epidemics can severely reduce grain yield due to collapse of the entire crop [5]. BB was first characterized in the late nineteenth century [6]. Introduction of resistance (*R*) genes into rice cultivars is considered as the best option for *Xoo management*. A total of 42 *R* genes have been identified in rice against *Xoo, and the number continues to grow* [7–9]. Due to co-evolution and selection pressure between *Xoo* and rice, these *R* genes are selective in their efficiency against specific *Xoo* strains or races, which are sets of strains that share incompatibility on defined sets

Advancements in genomics, referring here to DNA and RNA analyses, is as beneficial to crop protection as is to other discipline of biology. Rice MetaSysB, an open source which provides detailed information about BB-responsive genes, is based on the global expression analysis. The database provided 7475 unique genes and 5375 simple sequence repeats, which were responsive to *Xoo* in rice [11]. Such information is based on the compatible and incompatible rice-*Xoo* interactions. In another example, 454 and 498 differentially expressed genes were reported as exemplified by the incompatible and compatible rice-*Xoo* interactions, respectively, using cDNA microarray [12]. Genomics also provides functional information of genes up- and downstream of candidate resistance genes in the defense signal pathway, as is done in near-isogenic rice lines introgressed with *Xa39*, an as yet uncharacterized BB

Multiple rice and *Xoo* genomes have been sequenced, either in draft or complete

Many BB-resistance genes in modern rice germplasm were selected long before the concepts of modern plant breeding were established, and a rich assortment of major dominant and recessive *R* genes has been identified by genetic and molecular

Perhaps the best known of these genes, *Xa21* represents the receptor kinase (RLK) class of *R* genes. *Xa21* was originally introgressed into rice from the related species *O*. *longistaminata* and confers resistance to a broad range of *Xoo* strains [25]. *Xa26*, another cloned member of RLK gene family, also confers broad resistance with a somewhat different strain profile [26]. The cognate elicitor for Xa21 has been

RLKs play a central role in disease immunity pathways in plants, largely via the characterization of the bacterial flagellin receptor FLS2 and the related receptor EFR in *Arabidopsis* [28, 29]. A typical RLK consists of an extracellular receptor domain comprising of leucine-rich repeats (LRRs), a transmembrane domain, and an intracellular kinase domain [30]. As a class, RLKs have great potential for

form [14–23], paving the way to identify functional connections between host and pathogen genes. The functional validation of the candidate genes is helping develop new rice varieties by introduction of the gene of interest through traditional breeding, marker assisted breeding, or genetic engineering approaches [3]. BB disease resistance is overcome by the emergence of more virulent strains of *Xoo*. Whole genome sequencing of 100 *Xoo* strains from India revealed that these strains were distinct from African and US *Xoo* strains [24]. Based on the reaction towards ten major resistance genes of rice, 46 out of the 100 strains were grouped into 11

**110**

*Cloned R genes to bacterial blight of rice.*

enhancing resistance to BB in rice and in other disease complexes of crop plants *Xa21*, *Xa26*, and other RLKs represent genetic components of the pathogenassociated molecular patterns (PAMPs)-triggered immunity (PTI) surveillance pathway in rice. Improvements in the rationale design of RLK receptor specificities, and screening for novel genes in germplasm or wild relatives could lead to general application for broad and durable resistance.

The nucleotide binding site-LRR (NBS-LRR) is another large class of *R* gene, represented in rice toward *Xoo* by *Xa1* and *Xo1* [31–33]. XA1 and XO1 recognize multiple TALe, and *Xoo* strains have adapted TALes, the so-called iTALes, that are truncated and inhibit the function of XA1 and XO1 [32, 34].

Specific TALe-dependent *R* genes governing dominant resistance in rice against *Xoo* are known as executor (*E*) genes. *E* genes are distinct from classical *R* genes, whose transcriptional activation by TALes of *Xoo* trigger immunity, leading to dominant resistance [35]. *Xa27* represents the E genes class of dominant *R* genes and confers broad resistance to BB in rice [36]. Although not expressed in susceptible host, *Xa27* is expressed only upon inoculation with *Xoo* strains harboring the TALe gene *avrXa27* [37]. The protein is localized to apoplastic space, cell membrane and cell wall, and when expressed under a pathogen-nonspecific inducible rice *OsPR1* promoter, conferred constitutive resistance to both compatible and incompatible

strains alike [37]. The rice *R* genes *Xa10* and *Xa23* have similar requirements for the transcription activation domain and nuclear localization sequence (NLS) motifs of the corresponding TALes for their induction [38, 39].

*Xa4* is the latest and, again, an unusual *R* gene of rice to be characterized. The protein is a wall-associated kinase (WAK) and provides attributes other than enhanced resistance. Rice plants with XA4 are shorter and stiffer in comparison to plants lacking the gene [40]. Xa4 is race-specific, meaning many strains of *Xoo* are compatible on plants with Xa4. How Xa4 functions in resistance is unknown at present.

### **3.1** *SWEET* **genes and recessive resistance**

A class of major TALe-dependent susceptibility (*S*) genes for BB in rice encodes sugar transporters, thereby named as SWEET gene family [41]. Specific TALes, referred to as major TALes, transcriptionally activate the corresponding SWEET genes in rice during infection to promote the disease in a gene-for-gene susceptibility manner [42]. Although at least five SWEET genes of the clade III members can function as an *S* gene in BB, only three members are known to be targeted by extant strains of *Xoo* [42–47]. A member of the SWEET gene family, *OsSWEET14*, is induced by multiple distinct TALes, which include AvrXa7, PthXo3, Tal5 and TalC and are present in strains of different geographic origins and genetic lineages [43, 45, 46]. Similarly, PthXo2 drives *OsSWEET13* expression in the susceptible rice variety IR24 [44], and *OsSWEET11* is induced by the cognate PthXo1 [42]. The typical TALe possesses a central repetitive domain, a nuclear localization signal domain, and a transcription activation domain. The repetitive domain is responsible for binding of the TALe to a sequence motif called the effector binding element (EBE), which is commonly located in the promoter region of the respective *S* gene.

Mutated *S* gene alleles are proposed to be potentially more durable than dominant *R* genes [48, 49]. Identifying the promoter variant alleles of major *S* genes has been proposed in breeding for BB resistance [42, 47, 50–53]. Recessive resistance is due to the cognate TALe cannot bind to the promoter variants of the *S* gene. The gene *xa13*, for example, is a recessive resistance insertion allele of 14.8 kb DNA fragment in the promoter of *OsSWEET11* [42, 47]. *OsSWEET11* encodes a protein related to *MtN3* encoding nodulin 3 (N3) protein of *Medicago truncatula*. The gene was originally named *Os8N3* due to its location on rice chromosome 8 and the similarity to *MtN3* [42]. The critical difference between resistant (*xa13*/*xa13*) and susceptible plants is the elevated expression of *OsSWEET11* during infection in otherwise susceptible plant genotypes [42]. RNAi-mediated silencing of *OsSWEET11* plants was similarly resistant to *Xoo* strains that are solely dependent on PthXo1 for SWEET induction. Silenced plants, but not promoter variants, showed low pollen viability, corroborating the fact that *Xoo* hijacked otherwise developmentally important genes in rice for pathogenicity [42, 47]. Similarly, the TALe PthXo2 cannot bind to the EBE of *xa25*, a recessive allele of *OsSWEET13*, or the EBE region of *OsSWEET13* in japonica rice cultivars, owing to single nucleotide polymorphisms in the respective EBEs [51, 52].

The gene *xa13* is a naturally occurring allele, actually a series of alleles that protects the plant from a genetic disease vulnerability in the plant developmental pathways [42, 47]. However, *xa13* is not a broad resistance provided in comparison to *Xa21*, *Xa27* and *xa5*), and many strains from China, Philippines, Japan and Korea are compatible on *xa13* lines [51]. Compatibility is derived by acquisition of major TALes that target alternative SWEET promoters [43]. As yet, not major TALe has been identified that replaces PthXo1 for *OsSWEET11* expression.

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*Disease Resistance and Susceptibility Genes to Bacterial Blight of Rice*

The gene *xa5* also affects TALe-dependent function but does not act at a specific SWEET gene. The recessive allele encodes a variant of the γ or small subunit of the transcription factor TFIIA [54, 55], which confers broad resistance. The gene differs from the susceptible allele by a single codon substitution of valine at position 39 to glutamic acid. TFIIA, consists of α, β, and γ subunits, and is involved in stabilizing the binding of the TATA box binding protein complex (TFIID) to the TATA box of gene promoters. The TFIIA components are highly conserved across the eukaryotes. Rice has two loci for *TFIIAγ*-one gene is on chromosome 5 (*TFIIAγ5, xa5*) and another on chromosome 1 (*TFIIAγ1*) [54]. The proteins are closely related but not identical. *xa5* provides broad BB resistance and functions in inhibiting TALe function [51, 56]. However, *xa5* is not effective against strains with the TALe PthXo1 [51]. Perhaps not all *SWEET S* genes are known or are not always induced in disease by *Xoo*. The Indian strain IX-80 was virulent but did not induce any known *SWEET* gene [57], suggesting an adaptation by the *Xoo* to relieve dependency on *SWEET* gene family. On the other hand, IX-80 remains TALe-dependent as the strain was not compatible on IR53 (*xa13*/*xa13*, *xa5*/*xa5*), a gene combination that blocks the xa5 compatible PthXo1 and all other major TALes at *OsSWEET14* and *OsSWEET14* [51].

**4. Implication of interactions between TALes and the corresponding** 

populations should provide a degree of broad and durable resistance.

of XA7 mediated resistance is as yet unknown.

In the case of *xa13*, induction of the dominant allele *SWEET11* is mediated by the TALe PthXo1 [42]. However, strains of *Xoo* that solely rely on PthXo1 cannot induce *xa13* allele, and rice homozygous for *xa13* is symptomless. *xa13*-dependent recessive resistance is phenotypically and qualitatively different from resistance provided by the dominant *R* gene *Xa7* [42, 63]. Quantitatively, however, resistance mediated by *xa13* and *Xa7* are approximately equal with respect to bacterial growth and lesion length [42, 58, 64]. *Xa7* resistance is the result of the presence of the appropriate AvrXa7 in the pathogen and dominant, while *xa13* resistance is dependent on the absence of an effective virulence factor and recessive. The mechanism

Due to the large reservoir of TALes in each strain of *Xoo* and the diverse roles of TALes in pathogenesis, the BB of rice represents an excellent plant/pathogen system for studying the biology of TALes. The apparent reason for the broad activity of *Xa27* and *Xa23* is the presence of the cognate TALes *avrXa27* and *avrXa23* in a large number of strains from southeast Asia, including Korea, China, Japan and the Philippines [37, 39]. On the other hand, the loss of *avrXa27*, *avrXa23*, or *avrXa10*, for that matter, does not appear to have an apparent fitness cost to the pathogen, and populations of *Xoo* may lose *avrXa27* if *Xa27* is widely deployed [37–39]. AvrXa7 is an important virulence factor for some strains of *Xoo*, and strains with AvrXa7 are incompatible on rice lines harboring the *Xa7*. In this case, loss of *avrXa7*, which is a major TALe for *OsSWEET14*, may result in strains that are weakly virulent or, essentially, nonpathogenic, if no other SWEET inducing TALes are present [43, 58]. A variety of other TALe genes are present in *Xoo* populations that can restore full virulence to strains missing *avrXa7* [59]. Evasion of *Xa7*-mediated resistance is possible by loss of the gene, rearrangement of the central repeats or recombination among different TALe genes [60, 61]. However, despite rapid adaptation of bacteria by genetic changes and gene flow, field studies in the Philippines indicated that deployment of *Xa7* was durable in test plots for more than 10 years [62]. Therefore, strains may have other limitations due to geographical location or rice genotype. Nevertheless, pyramiding broadly effective *R* genes with cognate TALes that are wide-spread in the pathogen

*DOI: http://dx.doi.org/10.5772/intechopen.86126*

**host genes**

*Disease Resistance and Susceptibility Genes to Bacterial Blight of Rice DOI: http://dx.doi.org/10.5772/intechopen.86126*

*Protecting Rice Grains in the Post-Genomic Era*

present.

the corresponding TALes for their induction [38, 39].

**3.1** *SWEET* **genes and recessive resistance**

region of the respective *S* gene.

in the respective EBEs [51, 52].

strains alike [37]. The rice *R* genes *Xa10* and *Xa23* have similar requirements for the transcription activation domain and nuclear localization sequence (NLS) motifs of

*Xa4* is the latest and, again, an unusual *R* gene of rice to be characterized. The protein is a wall-associated kinase (WAK) and provides attributes other than enhanced resistance. Rice plants with XA4 are shorter and stiffer in comparison to plants lacking the gene [40]. Xa4 is race-specific, meaning many strains of *Xoo* are compatible on plants with Xa4. How Xa4 functions in resistance is unknown at

A class of major TALe-dependent susceptibility (*S*) genes for BB in rice encodes sugar transporters, thereby named as SWEET gene family [41]. Specific TALes, referred to as major TALes, transcriptionally activate the corresponding SWEET genes in rice during infection to promote the disease in a gene-for-gene susceptibility manner [42]. Although at least five SWEET genes of the clade III members can function as an *S* gene in BB, only three members are known to be targeted by extant strains of *Xoo* [42–47]. A member of the SWEET gene family, *OsSWEET14*, is induced by multiple distinct TALes, which include AvrXa7, PthXo3, Tal5 and TalC and are present in strains of different geographic origins and genetic lineages [43, 45, 46]. Similarly, PthXo2 drives *OsSWEET13* expression in the susceptible rice variety IR24 [44], and *OsSWEET11* is induced by the cognate PthXo1 [42]. The typical TALe possesses a central repetitive domain, a nuclear localization signal domain, and a transcription activation domain. The repetitive domain is responsible for binding of the TALe to a sequence motif called the effector binding element (EBE), which is commonly located in the promoter

Mutated *S* gene alleles are proposed to be potentially more durable than dominant *R* genes [48, 49]. Identifying the promoter variant alleles of major *S* genes has been proposed in breeding for BB resistance [42, 47, 50–53]. Recessive resistance is due to the cognate TALe cannot bind to the promoter variants of the *S* gene. The gene *xa13*, for example, is a recessive resistance insertion allele of 14.8 kb DNA fragment in the promoter of *OsSWEET11* [42, 47]. *OsSWEET11* encodes a protein related to *MtN3* encoding nodulin 3 (N3) protein of *Medicago truncatula*. The gene was originally named *Os8N3* due to its location on rice chromosome 8 and the similarity to *MtN3* [42]. The critical difference between resistant (*xa13*/*xa13*) and susceptible plants is the elevated expression of *OsSWEET11* during infection in otherwise susceptible plant genotypes [42]. RNAi-mediated silencing of *OsSWEET11* plants was similarly resistant to *Xoo* strains that are solely dependent on PthXo1 for SWEET induction. Silenced plants, but not promoter variants, showed low pollen viability, corroborating the fact that *Xoo* hijacked otherwise developmentally important genes in rice for pathogenicity [42, 47]. Similarly, the TALe PthXo2 cannot bind to the EBE of *xa25*, a recessive allele of *OsSWEET13*, or the EBE region of *OsSWEET13* in japonica rice cultivars, owing to single nucleotide polymorphisms

The gene *xa13* is a naturally occurring allele, actually a series of alleles that protects the plant from a genetic disease vulnerability in the plant developmental pathways [42, 47]. However, *xa13* is not a broad resistance provided in comparison to *Xa21*, *Xa27* and *xa5*), and many strains from China, Philippines, Japan and Korea are compatible on *xa13* lines [51]. Compatibility is derived by acquisition of major TALes that target alternative SWEET promoters [43]. As yet, not major TALe has

been identified that replaces PthXo1 for *OsSWEET11* expression.

**112**

The gene *xa5* also affects TALe-dependent function but does not act at a specific SWEET gene. The recessive allele encodes a variant of the γ or small subunit of the transcription factor TFIIA [54, 55], which confers broad resistance. The gene differs from the susceptible allele by a single codon substitution of valine at position 39 to glutamic acid. TFIIA, consists of α, β, and γ subunits, and is involved in stabilizing the binding of the TATA box binding protein complex (TFIID) to the TATA box of gene promoters. The TFIIA components are highly conserved across the eukaryotes. Rice has two loci for *TFIIAγ*-one gene is on chromosome 5 (*TFIIAγ5, xa5*) and another on chromosome 1 (*TFIIAγ1*) [54]. The proteins are closely related but not identical. *xa5* provides broad BB resistance and functions in inhibiting TALe function [51, 56]. However, *xa5* is not effective against strains with the TALe PthXo1 [51].

Perhaps not all *SWEET S* genes are known or are not always induced in disease by *Xoo*. The Indian strain IX-80 was virulent but did not induce any known *SWEET* gene [57], suggesting an adaptation by the *Xoo* to relieve dependency on *SWEET* gene family. On the other hand, IX-80 remains TALe-dependent as the strain was not compatible on IR53 (*xa13*/*xa13*, *xa5*/*xa5*), a gene combination that blocks the xa5 compatible PthXo1 and all other major TALes at *OsSWEET14* and *OsSWEET14* [51].
