Bacterial Blight Resistance in RiceYanjun Kou and Shiping Wang
Rice is one of the most important cultivated food crops. Bacterial blight (BB) caused by Xanthomonasoryzae pv. oryzae (Xoo) is one of the major constraints for sustainable production of rice. Researchershave made tremendous progress in trying to elucidate the interaction between rice and Xoo. Thegenomes of three Xoo strains have been sequenced. Some factors affecting pathogenicity of Xoo, suchas type III secretion system, effectors translocated by type II and III secretion systems, have beenidentied. In rice, a number of genes contributing to qualitative and quantitative resistance against Xoohave been characterized. At least 37 major disease (MR) genes have been identied and named, and7 (Xa1, Xa3/Xa26, xa5, xa13, Xa21, xa25, and Xa27) of them have been isolated. Importantly, somekey components functioning in Xa3/Xa26- and Xa21-mediated defense signaling pathways have beencharacterized, which is helpful to understand molecular mechanisms of qualitative resistance to BB.At least 74 resistance QTLs against Xoo have been identied in different rice cultivars interacting withdifferent Xoo strains. One major resistance QTL (WRKY45) and eight minor resistance QTLs (NRR,WRKY13, OsDR8, MPK6, GH3-1, GH3-2, GH3-8, and C3H12) have also been identied. The wealthof information about molecular components that function in rice defense response is now accessiblefor rice improvement in breeding programs.
Rice (Oryza sativa L.) is perhaps the most widely cultivated food crop worldwide; it is consumedby approximately 50% of the worlds population, and its consumption has been dramatically increasedin many parts of the world (White 1994). Various factors affect rice productivity, including diseases.Bacterial blight (BB) is the most devastating bacterial disease of rice. It occurs in epidemic areas ofthe world and can result in yield loss of up to 50% (Ou 1985). Traditional management methods,including cultivation strategies, chemical control, and biological control, are useful tools to combatBB. However, these methods can be labor intensive, expensive, and may cause environment pollution.The most economical and environmentally friendly way to control BB is to use resistant varietiescarrying major disease resistance (MR) genes and/or resistance quantitative trait loci (QTLs) incombination with agricultural management practices. Resistance genes and QTLs have been identiedand provide valuable resources for developing broad-spectrum and/or durable resistance against BBin rice breeding programs.
Translational Genomics for Crop Breeding, Volume I: Biotic Stress, First Edition. Edited by Rajeev K. Varshney and Roberto Tuberosa.C 2013 John Wiley & Sons, Inc. Published 2013 by John Wiley & Sons, Inc.
12 TRANSLATIONAL GENOMICS FOR CROP BREEDING
The Disease and Pathogen
BB, also called kersek at early growth stage ofthe plant, is caused by Xanthomonas oryzae pv.oryzae (Xoo) and is one of the oldest known cropdiseases. It was rst reported by the farmers ofFukuoka (Japan) in 1884 (Yamanuki et al. 1962).Subsequently, it was found in various parts ofAsian countries, Australia, African countries,and the United States. BB occurs in both temper-ate and tropical regions, but outbreaks are morefrequent in irrigated and rainfed lowland areas.Severe epidemics often occur with strong windsand continuous heavy rains (Ou 1985). Xoo maybe seed-borne and can be spread by irrigationwater, but this is disputed (Mizukami 1961;Premalatha and Devadath 1983). The pathogenmay survive on infected cultivated rice plants orother hosts (wild rice and gramineous weeds)over winter (Ou 1985). Under favorable condi-tions, Xoo invades rice leaves through hydath-odes or wounds, multiplies in the intercellularspace of the underlying epithem, and spreadsinto the plant through the xylem vessels, result-ing in yellow lesions with wavy margins alongthe veins that may systemically extend to thesheath (Figures 2.1A, 2.1B). BB is observed onboth seedlings and adult plants and peaks at theowering stage.
Fig. 2.1. Bacterial blight disease of rice. (A) Rice cultivar infected by Xoo. (B) Infected rice leaves afterarticial inoculation of Xoo. (C) Xoo colonies. For a color version of this gure, please refer to the color plate.
Xoo is a gram-negative bacterium that isrod-shaped, round-ended, motile, and slime-producing with a polar agellum. The length andwidth of individual cells are approximately 0.7to 2.0 m and 0.4 to 0.7 m, respectively. Bac-terial colonies on nutrient solid media are yel-low, round, and convex (Webster and Gunnell1992) (Figure 2.1C). Xoo is aerobic, catalase-positive, able to produce acids from carbohy-drates, and unable to use nitrate. The optimaltemperature range for Xoo growth is 25C to30C (Bradbury 1984). Identication and clas-sication of the bacterial pathotypes of Xoo arehelpful for resistance breeding and disease con-trol of BB. However, the morphological, phys-iological, and biochemical characters of differ-ent pathotypes are identical (Reddy and Reddy1990). Based on the infection responses elicitedin rice lines, JapaneseXoo strains have been clas-sied into 6 virulence groups (I to VI), Philip-pines Xoo strains have been classied into 10virulence groups (race 1 to 10), Chinese Xoostrains include 7 virulence groups (C1 to C7),and Indian Xoo strains can be classied into 13clusters and 5 broad groups (Ezuka and Horino1974; Vera Cruz 1984; Fang 1990; Nayak et al.2008).
The genomes of three Xoo strains, includ-ing Japanese strain MAFF311018, Korean strain
BACTERIAL BLIGHT RESISTANCE IN RICE 13
KACC10331, and Philippine strain PXO99A,have been sequenced (Lee et al. 2005; Ochiaiet al. 2005; Salzberg et al. 2008). The Xoogenome is a single circular chromosome ofabout 50 million bases (Mb), and it containsnearly 5,000 open reading frames (ORFs). It fea-tures remarkable plasticity and evolves rapidly.There are large numbers of major rearrange-ments and indels between the three strains, whichcontributes to the genomic variation in Xoo.This genomic variation explains the diversity ofXoo genotypes and pathotypes (Salzberg et al.2008).
Factors Affecting Pathogenicityof Xoo
Key to Xoo pathogenicity is the type III secre-tion system that is encoded by hypersensitiveresponse and pathogenicity (Hrp) genes (Bochand Bonas 2010). The Hrp gene cluster is nec-essary for pathogenicity in susceptible hostsand for a hypersensitive response in resistanceplants and nonhost plants. In the Xoo genome,the Hrp gene cluster includes 26 genes thathave a high sequence similarity (Ochiai et al.2005). These genes are regulated by two cru-cial components, HrpG and HrpX, in the Xan-thomonas genus. The expression of HrpX geneis upregulated by HrpG protein (Koebnik et al.2006).
The type III secretion system translocateseffector proteins into plant cells to support bac-terial virulence, proliferation, and dissemina-tion. The largest effector family of Xoo is thetranscription activator-like (TAL) effector family(also called the avrBs3/pthA family) (Boch andBonas 2010). A common feature shared by TALeffectors is the central repeat region that con-sists of 1.5 to 28.5 repeats, with each repeat con-taining 3334 amino acids, and contributes tobinding the cis-elements named UTP (upregu-lated by TAL effector) boxes of plant gene pro-moters, the amino-terminal translocation region,the carboxyl-terminal nuclear localization sig-nal, and carboxyl-terminal acidic transcription
activator-like domain (Boch et al. 2009; Kayand Bonas 2009; Yuan and Wang 2012). TALeffectors function as specic transcriptional acti-vators in the plant cell nucleus. The speci-city of DNA recognition by the TAL effec-tor is determined by the variable amino acidsat residues 12 and 13 of each repeat. How-ever, some TAL effectors have been identiedas avirulence (Avr) proteins in disease resistance(R) gene-mediated Xoo resistance (Boch andBonas 2010).
In addition to the effectors translocated bythe type III secretion system, the other impor-tant virulence factors of Xoo are extracellularenzymes and polysaccharide and a diffusiblesignal factor (Feng et al. 1996; Buttner andBonas, 2010; He et al. 2010). The extracellu-lar enzymes, such as endoglucanases, xylanase,cellobiosidase, and esterase, are secreted by thetype II secretion system of Xoo to degradethe plant cell wall (Buttner and Bonas 2010).The extracellular polysaccharide protects bac-teria against environmental stress. Null muta-tion of rpfC in Xoo strain T3000 substan-tially inuences the synthesis of extracellularpolysaccharide and virulence in rice (Feng et al.1996). The diffusible signal factor is a cell-cell communication signal, and it can affect theexpression of virulence genes (He et al. 2010).Repeats in the structural toxin (RTX toxin),which has functions in biolm development, cel-lular adherence, and eukaryotic cell targeting,represent another type of important virulencefactors among gram-negative bacteria (Coote1992; Satchell 2011). Several RTX toxins,including phenylacetic acid, trans-3-methylthio-acrylic acid, and 3-methylthio-propionic acid,have been identied in Xoo (Noda et al. 1989).Thus, RTX toxins may also be virulence fac-tors of Xoo. In addition, the rax genes (suchas raxA, raxB, raxC, and raxST) of Xoo areinvolved in secretion by the type I secre-tion system and sulfation of peptide Ax21(activator of Xa21-mediated immunity), whichelicit rice Xa21 protein-mediated resistance (Leeet al. 2009).
14 TRANSLATIONAL GENOMICS FOR CROP BREEDING
Xoo Resistance in Rice
Overview of Disease ResistanceMechanism in Plants
Physical and biochemical barriers provide a rstline of defense against potential pathogen attack.These constitutive defenses include the presenceof many preformed barriers such as waxy epi-dermal cuticles, cell wall, bark, antimicrobialenzymes, and secondary metabolites. However,pathogens have evolved strategies to breach thesepassive defense barriers. When Xoo enters aleaf apoplast through hydathodes or wounds, theplant relies on its innate immune system to detectthe invading organisms and activate inducibledefenses.
The current view of plant-pathogen interac-tions has revealed that the innate immune systemconsists of a two-branched defense response. Therst branch is pathogen (microbe)-associatedmolecular patterns (PAMPs/MAMPs)-triggeredimmunity (PTI) or basal resistance, which isinitiated by the direct recognition pathogenPAMPs through plant pattern-recognition recep-tors (PRRs) (Jones and Dangl 2006; Boller andFelix 2009). PRRs are plasma membrane pro-teins. PAMPs, which are essential for microbetness or survival, are relatively conservedmolecules within a class of microbes dur-ing evolution, such as agellin, peptidogly-can, and lipopolysaccharides. The other branchis effector-triggered immunity (ETI) or race-specic resistance that is activated on direct orindirect detection of pathogen effectors by plantproteins encoded by R genes (Jones and Dangl2006; Thomma et al. 2011). R proteins are eitherintracellular, plasma membrane, or extracellular,and each of these R proteins recognizes one ora few specic effectors. Pathogen effectors arerapid evolving, which results in loss of functionof R proteins.
After the presence of PAMPs or effectorsactivates PPRs or R protein, the plant recep-tors transfer the defense signal to downstreamcomponents encoded by defense-responsive ordefense-related genes, which leads to defense
responses. Defense-responsive genes are char-acterized by their response to a pathogen attackvia changed expression levels or posttransla-tional modications of their encoding proteins(Kou and Wang 2010). In general, PTI is a rel-ative weak defense response and ETI is a high-level defense response. However, strong PTI andweak ETI have also been reported (Thommaet al. 2011). Furthermore, PAMPs and effec-tors as well as PRRs and R proteins cannot bestrictly maintained, because there is a continuumbetween PTI and ETI (Thomma et al. 2011). Forexample, rice Xa21-mediated Xoo resistance istriggered by a narrowly conserved PAMP, Ax21,and Xa21 protein is considered to be both a PRRand an R protein (Lee et al. 2009). In addition,the defense signaling pathways initiated byPRRsandRproteins are partially overlapping (Kou andWang 2010).
According to the speed and strength of theplant response to pathogen invasion, plant resis-tance can be divided into two major categories:qualitative or complete resistance and quantita-tive or partial resistance. Qualitative resistance isa rapid and high level of defense response medi-ated by MR genes, including R and PRR genesthat confer a high level of resistance. More than30 MR genes that mediate qualitative resistanceand have different resistance spectra against Xoohave been named. Quantitative resistance is con-trolled by multiple genes or resistance QTLs andcan be broad spectrum and/or durable (Kou andWang 2010). A large number of resistance QTLshave been identied in the interactions of dif-ferent rice varieties and Xoo strains (Kou andWang 2012).
In addition to innate immunity, plants havedifferent types of induced resistance, includingsystemic acquired resistance (SAR) and inducedsystemic resistance (ISR). Genetic studies inArabidopsis revealed that NPR1 (non-expressorof pathogenesis-related genes 1) is important forSAR, and TGA transcription factors are repres-sors of SAR (Vlot et al. 2009). Some evidencesupports rice having a similar SAR pathwayfor Xoo resistance. Overexpression of rice NH1,
BACTERIAL BLIGHT RESISTANCE IN RICE 15
which is a sequence and functional ortholog ofArabidopsis NPR1, results in enhanced resis-tance to Xoo (Chern et al. 2005). In rice, NH1interacts with TGA2.1 transcription factor andnegative regulator of resistance (NRR). TGA2.1negatively regulates basal defense responses toXoo (Fitzerald et al. 2005). Rice NRR nega-tively regulates SAR in Arabidopsis and basalandXa21-mediatedXoo resistance in rice (Chernet al. 2005, 2008). It is also known that arice mitogen-activated protein kinase, MPK6,negatively regulates SAR in rice-Xoo interac-tion (Shen et al. 2010). ISR of plants againstpathogens is a widespread phenomenon that acti-vates multiple defense mechanisms includingincreased activity of pathogenesis-related gene(PR) proteins.AttenuatedUV-mutantXoo strainshave been documented to induce rice ISR againstBB (Thein and Prathuangwong 2010).
Qualitative Resistance to XooAsian-cultivated rice (AA genome) consists oftwo major subspecies, indica (O. sativa L. ssp.indica) and japonica (O. sativa L. ssp. japon-ica). At least 37MR genes against Xoo have beenidentied and designated in a series from Xa1 toXa36, with one symbol having been used for twodifferent genes (Table 2.1). Most of these geneswere identied in Asian-cultivated rice whileonly a fewwere identied fromwild rice species,whichwere then introgressed into cultivated rice.It is generally accepted that R proteins encodedby dominantR genes recognize specic pathogeneffec...