High-resolution genetic mapping of bacterial blight resistance gene <Emphasis Type="Italic">Xa10</Emphasis>

Bacterial blight of rice, caused by Xanthomonas oryzae pv. oryzae (Xoo), is the most devastating disease of rice (Oryza sativa L). Rice lines that carry resistance (R) gene Xa10 confer race-specific resistance to Xoo strains harboring avirulence (Avr) gene avrXa10. Here we report on genetic study, disease evaluation and fine genetic mapping of the Xa10 gene. The inheritance of Xa10-mediated resistance to PXO99^A(pHM1avrXa10) did not follow typical Mendelian inheritance for single dominant gene in F₂ population derived from IR24 × IRBB10. A locus might be present in IRBB10 that caused distorted segregation in F₂ population. To eliminate this locus, an F₃ population (F₃-65) was identified, which showed normal Mendelian segregation ratio of 3:1 for resistance and susceptibility. A new near-isogenic line (F₃-65-1743) of Xa10 in IR24 genetic background was developed and designated as IRBB10A. IRBB10A retained similar resistance specificity as that of IRBB10 and provided complete resistance to PXO99^A(pHM1avrXa10) from seedling to adult stages. Linkage analysis using existing RFLP markers and F₂ mapping population mapped the Xa10 locus to the proximal side of E1981S with genetic distance at 0.93 cM. With five new RFLP markers developed from the genomic sequence of Nipponbare, Xa10 was finely mapped at genetic distance of 0.28 cM between proximal marker M491 and distal marker M419 and co-segregated with markers S723 and M604. The physical distance between M491 and M419 on Nipponbare genome is 74 kb. Seven genes have been annotated from this 74-kb region and six of them are possible Xa10 candidates. The results of this study will be useful in Xa10 cloning and marker-assisted breeding.     

RaxH/RaxR: a two-component regulatory system in Xanthomonas oryzae pv. oryzae required for AvrXa21 activity

Xanthomonas oryzae pv. oryzae is the causal agent of bacterial leaf blight, one of the most serious diseases in rice. X. oryzae pv. oryzae Philippine race 6 (PR6) strains are unable to establish infection in rice lines expressing the resistance gene Xa21. Although the pathogen-associated molecule that triggers the Xa21-mediated defense response (AvrXa21) is unknown, six rax (required for AvrXa21 activity) genes encoding proteins involved in sulfur metabolism and Type I secretion were recently identified. Here, we report on the identification of two additional rax genes, raxR and raxH, which encode a response regulator and a histidine protein kinase of two-component regulatory systems, respectively. Null mutants of PR6 strain PXO99 that are impaired in either raxR, raxH, or both cause lesions significantly longer and grow to significantly higher levels than does the wild-type strain in Xa21-rice leaves. Both raxR and raxH mutants are complemented to wild-type levels of AvrXa21 activity by introduction of expression vectors carrying raxR and raxH, respectively. These null mutants do not affect AvrXa7 and AvrXa10 activities, as observed in inoculation experiments with Xa7- and Xa10-rice lines. Western blot and raxR/gfp promoter-reporter analyses confirmed RaxR expression in X. oryzae pv. oryzae. The results of promoter-reporter studies also suggest that the previously identified raxSTAB operon is a target for RaxH/RaxR regulation. Characterization of the RaxH/RaxR system provides new opportunities for understanding the specificity of the X. oryzae pv. oryzae-Xa21 interaction and may contribute to the identification of AvrXa21.     

Pathotypic variation of Xanthomonas oryzae pv. oryzae in Bangladesh

Bacterial Blight (BB) caused by Xanthomonas oryzae pv. oryzae (Xoo), a destructive disease of rice. Altogether, 96 isolates of Xoo were collected from 19 rice growing districts of Bangladesh in irrigated and rainfed seasons during 2014 to assess pathotypic variation. Pathotypic analyses on a set of 12 Near Isogenic Lines (NILs) of rice containing resistance genes viz. Xa1, Xa2, Xa3, Xa4, Xa5, Xa7, Xa8, Xa10, Xa11, Xa13, Xa14 and Xa21 and two check varieties IR24 and TN1 by leaf clip-inoculation technique. A total of 24 pathotypes were identified based on their virulence patterns on NILs tested. Among these, pathotypes VII, XII, and XIV considered as major, containing maximum number of isolates, (9.38% each) frequently distributed in North to Mid-Eastern districts of Bangladesh. Most virulent pathotype I recorded from Habiganj and Brahmanbaria. This pathotypic variation explained the pathogenic relatedness of X. oryzae pv. oryzae populations from diverse geographic areas in Bangladesh.     

Constitutive heterologous expression of avrXa27 in rice containing the R gene Xa27 confers enhanced resistance to compatible Xanthomonas oryzae strains

The vascular pathogen Xanthomonas oryzae pv. oryzae ( Xoo ) and nonvascular pathogen Xanthomonas oryzae pv. oryzicola ( Xoc ) cause bacterial blight (BB) and bacterial leaf streak (BLS) diseases of rice, respectively. We have previously identified the avirulence gene avrXa27 from Xoo PXO99^A, which specifically induces the expression of the rice resistance gene Xa27 , ultimately leading to resistance against BB disease in rice. In this study, we have generated a transgenic rice line (L24) that expresses avrXa27 constitutively under the control of the PR1 promoter, and have examined its role in the host–pathogen interaction. L24 is not more susceptible to BB, indicating that avrXa27 does not contribute to virulence. AvrXa27 retains avirulence activity in L24 and, after crossing with a line containing Xa27 , progeny display phenotypic changes including inhibition of tillering, delay in flowering, stiff leaves, early leaf senescence and activation of pathogenesis-related ( PR ) genes. On challenge with a variety of compatible strains of Xoo and Xoc strain L8, lines with both avrXa27 and Xa27 also show enhanced resistance to bacterial infection. The induction of Xa27 and subsequent inhibition of Xoc growth in Xa27 plants are observed on inoculation with Xoc L8 harbouring avrXa27 . Our results indicate that the heterologous expression of avrXa27 in rice containing Xa27 triggers R gene-specific resistance and, at the same time, confers enhanced resistance to compatible strains of Xoo and Xoc . The expression of AvrXa27 and related proteins in plants has the potential to generate broad resistance in plants.     

Field performance of Xa21 transgenic indica rice (Oryza sativa L.), IR72

Based on the characterization of the resistance phenotype and molecular analysis, several homozygous lines carrying Xa21 against the bacterial blight (BB) pathogen were obtained from previously transformed indica rice, IR72. The homozygous line, T103-10, with the best phenotype and seed-setting, was repeatedly tested under normal field conditions to evaluate its levels of resistance to the BB pathogen in Wuhan, China, in 1998 and 1999. The isolates of Xanthomonas oryzae pv oryzae (Xoo) used in this experiments were PXO61, PXO79, PXO99 and PXO112 isolated from the Philippines, T2 isolated from Japan, and Zhe173 isolated from China. The results demonstrated that the transgenic homozygous line expressed the same resistance spectrum, but with a shorter lesion length to each inoculated isolates as the lesion length of the Xa21 donor line IRBB21. The non-transformed control IR72 carrying Xa4 was resistant to PXO61, PXO112, Zhe173 and T2, but susceptible to PXO99 and PXO79. The negative control variety IR24 was susceptible to all isolates under field conditions. The results demonstrated clearly that the Xa21 transgene led to an excellent field performance of the introduced bacterial blight resistance trait on the recipient plants. The yield performance of this transgenic homozygous line, T103-10, is comparable with that of the control under field conditions. Received: 2 August 1999 / Accepted: 3 November 1999     

Evaluation of resistance genes in rice against local isolates of Xanthomonas oryzae pv. oryzae in Punjab Province of Pakistan

Absence of resistance/tolerance against bacterial leaf blight (BLB), incited by Xanthomonas oryzae pv. oryzae, in famous basmati varieties is one of the main reason for BLB epidemic in Punjab in 2007–2008. For developing resistance against BLB, the response of 26 IRBB lines of IRRI including 10 near isogenic lines (NILs) and 16 gene pyramids carrying two to five resistance genes (Xa series) was evaluated against 61 indigenous Xoo isolates under artificial inoculation field conditions. None of the NILs or gene pyramid provides complete protection against all the isolates. However, Xa21 and xa13 were found resistant against the majority of Xoo isolates, followed by Xa14 and Xa7. Of the 16 gene pyramids used in this study, IRBB-54 (Xa5 + Xa21), IRBB-55 (Xa13 + Xa21) followed by IRBB-58 (Xa4 + Xa13 + Xa21) were found effective against the majority of the Xoo isolates. These resistance genes (individually and in combinations) can be incorporated for the improvement of basmati rice cultivars cultivated in Punjab province of Pakistan. Effectiveness of gene combination supports the strategy of pyramiding appropriate resistance genes. Newly identified resistant genes may also be evaluated for achieving broad spectrum resistance against more Xoo isolates of the area.     

Virulence Analysis and Race Classification of Xanthomonas oryzae pv. oryzae in China

Two hundred and eighty‐five isolates of Xanthomonas oryzae pv. oryzae were randomly collected from 22 rice‐growing provinces in China. Ninety‐one representative isolates were chosen to assess the differential characteristics of 24 near‐isogenic rice lines containing a single resistance gene or two to four genes. Most isolates were avirulent on pyramided lines, except IRBB51, and hence, the pyramided lines cannot be used as differentials for the virulence analysis of X. oryzae pv. oryzae in China. The 13 rice lines with a single gene were used further to establish a system of races classification of X. oryzae pv. oryzae in China. IR24 and IRBB10 were susceptible to the isolates with several exceptions, whereas IRBB5, IRBB7 and IRBB21 were resistant. Based on the interactions between the isolates of X. oryzae pv. oryzae and the 13 near‐isogenic rice lines, six single‐gene rice cultivars (IRBB5, IRBB13, IRBB3, IRBB14, IRBB2 and IR24) were chosen as differentials, and the 285 tested isolates were classified into nine races. The reaction patterns of the nine races in order were: RRRRRR, RRRRRS, RRRRSS, RRRSSS, RRSSSS, RSRRRS, RSSRRS, RSSSSS and SSSSSS. The race frequencies were 10.18%, 10.53%, 4.91%, 10.18%, 24.21%, 5.96%, 11.23%, 22.46% and 0.35% respectively. The virulence of representative strains of eight Philippine races on 13 rice lines with a single gene was determined and compared with the Chinese races. The frequency distributions of X. oryzae pv. oryzae races were primarily described for the different regions in China.     

High-resolution mapping and gene prediction of <Emphasis Type="Italic">Xanthomonas Oryzae</Emphasis> pv. <Emphasis Type="Italic">Oryzae</Emphasis> resistance gene <Emphasis Type="Italic">Xa7</Emphasis>

Bacterial blight (BB) caused by Xanthomonas oryzae pv. oryzae (Xoo) is a devastating disease in rice worldwide. The resistance gene Xa7, which provides dominant resistance against the pathogen with avirulence (Avr) gene AvrXa7, has proved to be durably resistant to BB. A set of SSR markers were selected from the “gramene” database based on the Xa7 gene initial mapping region on chromosome 6. These markers were used to construct a high-resolution genetic map of the chromosomal region surrounding the Xa7 gene. An F₂ mapping population with 721 highly susceptible individuals derived from a cross between the near isogenic lines (NILs) IRBB7 and IR24 were constructed to localize the Xa7 gene. In a primary analysis with eleven polymorphic SSR markers, Xa7 was located in approximately the 0.28-cM region. To walk closer to the target gene, recombinant F₂ individuals were tested using newly developed STMS (sequence tagged microsatellite) markers. Finally, the Xa7 gene was mapped to a 0.21-cM interval between the markers GDSSR02 and RM20593. The Xa7-linked markers were landed on the reference sequence of cv. Nipponbare through bioinformatics analysis. A contig map corresponding to the Xa7 gene was constructed. The target gene was assumed to span an interval of approximately 118.5-kb which contained a total of fourteen genes released by the TIGR Genome Annotation Version 5.0. Candidate-gene analysis of Xa7 revealed that the fourteen genes encode novel domains that have no amino acid sequence similar to other cloned Xa(xa) genes. Shen Chen and Zhanghui Huang are contributed equally to this work.     

Genotypic and Pathotypic Diversity of Xanthomonas oryzae pv. oryzae Strains in Taiwan

Rice bacterial leaf blight, caused by Xanthomonas oryzae pv. oryzae [(Ishiyama) Swings et al. 1990] (Xoo), is a major rice disease of the second crop season in Taiwan. A total of 88 Xoo strains collected from 10 major rice cultivating areas in Taiwan from 1986, 1997, 2000, 2004, and 2011 were characterized by repetitive‐element PCR (REP‐PCR) fingerprinting and virulence analyses. Among the five genetic clusters identified by the pJEL1/pJEL2 (IS1112‐based) and REP1R‐Dt/REP2‐D [repetitive extragenic palindromic (REP)‐based] primer sets, clusters A, C and D contained Xoo strains from geographically distant regions, which suggests a high frequency of Xoo dispersal in Taiwan. The 88 Xoo strains were evaluated by inoculations on IRBB near‐isogenic lines and five Taiwan rice cultivars. A subset of 45 moderately or highly virulent strains were classified into 15 pathotypes by their compatible or incompatible reactions on IR24 and 12 IRBB near‐isogenic lines, each containing a single resistance gene. Analysis of molecular haplotypes and pathotypes revealed a partial relationship. IRBB5, IRBB21 and IRBB4 were incompatible with 96%, 96% and 73% of the strains, so xa5, Xa21 and Xa4 can recognize most of the Xoo strains in Taiwan and elicit resistance. In contrast, IRBB3 (Xa3), IRBB8 (xa8), IRBB10 (Xa10), IRBB11 (Xa11), IRBB13 (xa13) and IRBB14 (Xa14) were susceptible to almost all of the 45 Xoo strains. Inoculation trials revealed significant differences in the susceptibility of five Taiwan cultivars to Xoo (from high to low susceptibility: Taichung Sen 10 >  IR24, Taichung Native 1 >  Taichung 192, Taikeng 9, Tainan 11). This study provides useful information for resistance breeding and the development of disease management strategies against bacterial blight disease of rice.     

Analysis of Pathogenic Diversity of the Rice Bacterial Blight Pathogen (Xanthomonas oryzae pv. oryzae) in the Andaman Islands and Identification of Effective Resistance Genes

Bacterial blight (BB) caused by Xanthomonas oryzae pv. oryzae (Xoo) is a major disease of rice in the tropics for which genetic resistance in the host plants is the only effective solution. This study aimed at identification of resistance gene combinations effective against Xoo isolates and fingerprinting of the Xoo isolates of Andaman Islands (India). Here, we report the reaction of 21 rice BB differentials possessing Xa1 to Xa21 genes individually and in different combinations to various isolates of pathogen collected from Andaman Islands. Pathological screening results of 14 isolates revealed that among individual genes tested across 2 years, Xa4, Xa7 and Xa21 conferred resistance reaction across all isolates, whereas among combinations, IRBB 50 (Xa4 + xa5), IRBB 52 (Xa4 + Xa21) and IRBB 60 (Xa4 + xa5 + xa13 + Xa21) conveyed effective resistance against tested isolates. The nature of genetic diversity among four isolates selected on the basis of geographical isolation in the islands was studied through DNA finger printing. The RAPD primers S111, S119, S1117, S1109, S1103, S109 and S105 were found to be better indicators of molecular diversity among isolates than JEL primers. The diversity analysis grouped 14 isolates into three major clusters based on disease reaction wherein isolate no. 8 was found the most divergent as well as highly virulent. The remaining isolates were classified into two distinct groups. The importance of the study in the context of transfer of resistance gene(s) in the local cultivars specifically for tropical island conditions is presented and discussed.