Challenges and Insights into Pyramided R Genes for Durable Rice Blast Resistance in Southwest China

Challenges and Insights into Pyramided R Genes for Durable Rice Blast Resistance in Southwest China | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Challenges and Insights into Pyramided R Genes for Durable Rice Blast Resistance in Southwest China Junhua Liu, Zhongya Cai, Mei Yang, Peng Liu, Shufan Lei, Zhiwen Lv, and 4 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-5734717/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 5 You are reading this latest preprint version Abstract Rice blast, caused by Magnaporthe oryzae , threatens global rice productivity through severe yield losses. Developing cultivars with durable resistance via strategic pyramiding of resistance (R) genes is essential for sustainable disease management. This study aimed to develop elite rice germplasm harboring effective R-gene compositions and evaluate their resistance stability across geographically distinct blast hotspots in Southwest China. From disease nursery screenings in Wanzhou and Enshi, 136 resistant lines were selected from crosses of elite parents. Subsequent multilocation evaluations identified 20 lines demonstrating stable resistance in Meitan, a region with distinct M. oryzae pathogenic populations, highlighting significant geographical variation in pathogen virulence. These results emphasize the necessity of multilocation testing for breeding durably resistant varieties. Allele-specific marker analysis of 14 major R genes revealed genetic compositions across resistant lines. Over 50% of lines carried Pi5 , Pi54 , Pita , or Pia , with most genotypes pyramiding 3-5 genes, reflecting the widespread adoption of R-gene pyramiding in rice breeding programs, with elite parental lines harboring diverse resistance gene combinations. Notably, lines demonstrating blast resistance in Meitan shared identical R-gene profiles with susceptible materials, suggesting the involvement of uncharacterized major-effect genes in conferring resistance. This finding underscores the prioritization of functional major-effect genes over quantitative gene stacking in resistance breeding. Our study establishes a regionally optimized framework for blast resistance evaluation and provides validated genetic resources for developing adapted rice cultivars in Southwest China. rice M. oryzae resistance genes rice resistance breeding disease nursery Figures Figure 1 Figure 2 Figure 3 Figure 4 Introduction Rice is a staple food in China, nourishing over half of the country's population and playing a crucial role in both social and economic development (Zhou et al., 2006). However, Rice production is constantly threatened by various biotic and abiotic stresses. Among these, rice blast disease, caused by Magnaporthe oryzae ( M. oryzae ), is one of the most devastating, impacting rice yields in over 85 countries worldwide (Wang et al., 2014). In China, annual yield losses due to rice blast exceed 0.4 million tons, severely threatening food security (Wancai, Zhendong et al. 2016). Although fungicides are widely used to control rice blast, their overuse has led to the emergence of fungicide-resistant M. oryzae strains, posing significant risks to human health and the environment (Luo et al., 2005). Therefore, the resistance breeding is considered the most cost-effective and environmentally sustainable strategy for managing rice blast disease (Wu et al., 2021). This dilemma underscores the urgency to develop sustainable disease management strategies. Since the cloning of the first blast resistance gene Pib in 1996, over 50 dominant R genes have been identified at 17 chromosomal loci in rice. Most of them as allelic variants, particularly at the Pi2 and Pik loci localized on chromosomes 6 and 11, respectively (Devanna et al., 2022; Huang et al., 2023). These genes encode conserved nucleotide-binding-site leucine-rich-repeat (NBS-LRR) proteins, which recognize corresponding avirulence proteins secreted by M. oryzae , triggering strong defense responses, including ROS burst, hypersensitive response (HR) and transcriptional reprogramming (Liu et al., 2021; Singh et al., 2016). Resistance genes have been extensively applied in rice breeding and production, effectively safeguarding the safe cultivation of rice. However, the efficacy of these resistance genes demonstrates region-specific characteristics (Rathour et al., 2016; Liu et al., 2021; Zeng et al., 2018). For example, the Pikm , Pi54 , and Pib genes confer the strongest resistance in Huanghuai, Shandong and Jiangsu rice cultivation regions. In contrast, the Pi5 and Pita genes are particularly effective in the middle and lower reaches of the Yangtze River (Wang et al., 2016; Chen et al., 2018; Zhou et al., 2022), while the Pi2 , Pish , Pikh , Pi9 and Pi41 genes are most effective in Guangdong province (Yang et al., 2008; Wu et al., 2023). However, the resistance conferred by individual R gene has been increasingly overcome by M. oryzae . Nevertheless, sustaining durable disease resistance remains a pivotal challenge in rice breeding. Pathogen populations undergo rapid co-evolutionary adaptation, frequently acquiring effector gene mutations that evade detection by host R proteins, ultimately leading to resistance breakdown. Notably, emerging empirical evidence demonstrates that pyramiding multiple R genes enhances broad-spectrum and durable resistance against M. oryzae in rice breeding, providing a genetically reinforced defense mechanism against pathogen evolution. For instance, pyramiding two R genes, such as Pi-1 with Pi-2 , Pi-46 with Pi-ta, or Pi9 with Pi54 , significantly enhanced resistance compares to varieties carrying a single R gene (Chen et al., 2001; Xiao et al., 2016; Zhou et al., 2020). However, not all combination of R gene pyramiding enhances rice blast resistance. For example, although pyramided lines carrying Pi9+Pi54 and Pizt+Pi54 exhibited higher resistance to leaf blast than the lines carrying the corresponding single R gene, pyramiding of Pi9+Pi54 unexpectedly reduced neck blast resistance compared to Pi9 alone (Xiao et al., 2017). These limitations highlight the need for region-specific R gene deployment strategies based on pathogen population dynamics. Southwest China represents a high-risk zone for rice blast due to its humid subtropical climate and complex pathogen diversity (Hu, Huang et al. 2022). To develop cultivars with durable blast resistance, we implemented an integrated approach combining multi-location field evaluation and molecular marker-assisted selection. Progenies derived from elite crosses were systematically screened across three disease nurseries (Wanzhou, Enshi, Meitan). Molecular characterization of 136 resistant lines identified 14 functional resistance genes through allele-specific marker analysis, including Pi5 (Lee et al., 2009), Pi54 (Arora et al., 2015), Pi-ta (Lee et al., 2011), Pia (Zeng et al., 2011), Pib (Wang et al., 1999), Pi2 (Zhou et al., 2006), Pid3 (Xu et al., 2014), Pik-m (Ashikawa et al., 2008), Pit (Li et al., 2014), Pigm (Deng et al., 2017), Pi1 (Hua et al., 2012), Pi9 (Chung et al., 2015), Pizt (Ning et al., 2020;Yokoo, 1983), Pik ( Ariya-anandech et al., 2018). Twenty F4 breeding lines demonstrating broad-spectrum resistance were ultimately selected, providing critical genetic resources and strategies to breed durable blast-resistant cultivars for Southwest China’s high-risk regions. Results Development of F2 segregating populations for screening of resistant restorer lines To develop resistant restorer lines adapted to the unique climatic conditions of the mountainous rice-growing regions in southwest China, we selected thirteen elite rice germplasm lines, including Huazhan, Yuehesimiao, Qhui 28 and ten additional lines detailed in Table S1 all of which had demonstrated excellent agronomic performance and stable high yields at Wanzhou over several years. These elite lines were then crossed with 26 donor germplasm lines, including Yunhui 11, Luhui 80, Chenghui 727 and others listed in Table S1, previously characterized for durable resistant to M. oryzae through systematic phenotyping in disease nurseries at Wanzhou and Enshi (Fig. 1A). Fifty-four F1 hybrid plants were generated from these crosses. The F1 plants were cultivated at Wanzhou and subjected to self-pollination to derive F2 segregating populations for subsequent resistance screening. Screening the resistant lines at Wanzhou and Enshi disease nurseries To identify blast-resistant germplasm, we first cultivated the 54 F2 segregating populations in the rice blast disease nursery at Wanzhou (Fig. 1A and 1B), where 800 lines resistant to both leaf blast and neck blast, while exhibiting high yield and elite agronomic traits, were selected. To validate resistance stability across environments, the F3 progeny lines derived these selections were further evaluated in the distinct epidemiological conditions of the Enshi disease nursery. 136 lines were selected for exhibiting high or moderate resistance to neck blast (Table 1). Notably, the cross between Qhui28 and Luhui 80 exhibited superior breeding efficiency, generating 10 resistance lines, the highest among all combinations. In contrast, 27 crosses yielded 2-9 lines each, and 26 crosses yielded single resistant line, such as the cross between 2015-B 36 and Mianhui 523. The data reflects a significant variation in combinatory resistance potential. The selected lines exhibited stable resistance across generations and environments To evaluate the heritability of resistance traits, the 136 F4 progeny lines were re-evaluated in Wanzhou and Enshi nurseries. In Wanzhou, all lines demonstrated resistance to leaf blast disease without segregation. For neck blast resistance, 2.94% of lines exhibited stable resistance, 95.59% showed moderate resistance, and 1.47% displayed moderate susceptibility (Figure 2A). In Enshi, 2.21% of the lines exhibited medium resistant to leaf blast disease, while the remaining lines maintained resistance. Neck blast disease evaluations identified 2.21% resistant lines, 96.32% moderately resistant, and 1.47% moderately susceptible (Fig. 2B). The consistent phenotypic performance patterns observed across both geographical locations and generational propagation confirm the high heritability and environmental stability of these resistance traits. The broad-spectrum resistance evaluation in Meitan rice blast nursery To characterize broad-spectrum resistance adaptability across heterogeneous rice ecosystems, we tested the 136 F4 lines at the Meitan disease nursery in Zunyi city, Guizhou province, where shares climatic similarities with Chongqing. All lines retained leaf blast resistance or moderate resistance phenotypes (Fig. 2A and B). In contrast, neck blast resistance displayed progressive attenuation, with no lines demonstrating resistance. Specifically, 20 lines (15.44%) demonstrated moderate neck blast resistance, 114 lines (83.82%) exhibited moderate susceptibility, and a single line (0.74%) showed full susceptibility (Fig. 2C). Ultimately, twenty lines were selected for rice resistance breeding based on their consistent resistance across the three disease nurseries. These lines exhibited minimal variability in resistance scores over multiple seasons, demonstrating strong potential for broad-spectrum resistance breeding (Table 2). Notably, compared to the uniform resistance patterns observed in Wanzhou and Enshi, the Meitan trials revealed substantial phenotypic variation among lines (Fig. 2), indicating broader pathogenic diversity within the M. oryzae population at this site. This geographical divergence in pathogen virulence profiles underscores the necessity of multi-location screening for breeding durable blast resistance. R gene composition in the 136 resistance lines Resistance to M. oryzae is primarily conferred by R genes through effector-triggered immunity (ETI) in rice. To characterize resistance mechanisms underlying variation in blast resistance, we performed molecular profiling of 14 NLR genes, including Pi5 , Pi54 , Pita , Pia , Pib , Pi2 , Pid3 , Pikm , Pit , PigmR , Pi1 , Pi9 , Pizt and Pik, across 136 F4 lines using gene-specific molecular markers (Figure S1 and Table S3). Three genes, Pi9 , Pizt and Pik , were absent in all analyzed lines. Seven genes, Pi2 , Pib , Pid3 , Pit , Pikm , Pi1 and PigmR, was detected in 49.26%, 49.26%, 30.15%, 16.18%, 16.18%, 2.21%and 0.74% of the lines, respectively. The remaining four R genes, Pi5 , Pi54 , Pita and Pia , were detected in more than 50% of the lines, with distribution frequency of 82.35%, 62.50%, 54.41%, and 53.680%, respectively (Figure 3A). The number of R genes per line varied from one to seven, with a normal distribution observed (Fig. 3B). While only one line carried a single R gene, the majority of the lines contained three to five R genes. This distribution suggests a high level of genetic diversity in the resistance composition among the F4 lines. Undetected R genes may exist in the resistant lines to confer resistance in Meitan nursery In the Meitan disease nursery, 143 rice lines exhibited susceptible phenotypes to M. oryzae infection, indicating compromised functionality of their intrinsic resistance genotypes under prevailing field conditions. Notably, the characterized resistance gene compositions identified in 20 resistant lines were paradoxically present in susceptible lines. For instance, resistant lines, K52, K42, K176 and K53, shared same resistance genotypes ( Pi5 + Pi54 + Pita ) with susceptible lines, K55, K57, K33, K174, K44, K45 and K103 (Table S1). Transcriptomic analysis revealed comparable expression patterns of Pi5 , Pi54 , and Pita in both resistant line K52 and susceptible line K55, both pre- and post-pathogen inoculation (Figure 4). This phenotypic-genotypic paradox implies that unknown or undetected known R genes not captured by in this study likely govern field resistance against M. oryzae . The identified resistant lines constitute valuable germplasm resources for discovery of novel R gene alleles, and optimization of pyramiding breeding strategies targeting durable blast resistance. Discussion Resistance to M. oryzae in rice is primarily conferred by R genes through effector-triggered immunity (ETI), which involves the recognition of pathogen-derived Avr proteins by plant immune receptors, triggering strong immune responses such as ROS bursts, hypersensitive responses (HR), and transcriptional reprogramming. These responses inhibit disease development by limiting the growth of M. oryzae isolates within the cells surrounding the infection site (Zhang et al., 2020; Kim et al., 2004; Cesari et al., 2013; Singh et al., 2016; Yan et al., 2023; Korinsak et al., 2022). Over 50 R genes have been cloned in rice, many of which have been utilized in breeding programs to improve resistance to rice blast (Li et al., 2019). However, resistance breeding faces significant challenges, especially the rapid evolution of M. oryzae. Resistance (R) genes impose strong selective pressure on M. oryzae populations. Strains harboring recognized Avr genes are suppressed in the field, while those with Avr loss (via mutations, deletions, or silencing) evade R gene recognition (Hu et al., 2022). Over time, these escaped strains dominate the pathogen population, leading to resistance breakdown in previously resistant rice varieties. In this study, we selected 136 resistant rice lines from two disease nurseries (Wanzhou and Enshi), which exhibited stable resistance to M. oryzae across multiple years. The majority of these lines harbor three or more pyramided R genes, demonstrating that R gene pyramiding has become a common practice in modern rice germplasm and breeding programs. This strategy effectively enhances blast resistance and delays resistance breakdown. However, pyramiding resistance genes presents challenges, as not all combinations of pyramided genes necessarily improve resistance (Wang et al., 2020; Variar et al., 2009). Our findings align with these reports. Notably, the genotypes of resistant individuals identified in the Meitan disease nursery were also detected in susceptible lines (Table S1), suggesting that unidentified or uncharacterized known resistance genes may predominantly govern resistance in these lines. In contrast, susceptible lines carrying multiple pyramided R genes, including those with up to eight in our study, still displayed susceptibility. This highlights that effective resistance in pyramided lines primarily relies on the presence of major-effect R genes, while the additive contributions of other pyramided genes may be less pronounced. Previous studies further confirm a non-linear correlation between R gene number and resistance intensity (Zhou et al., 2022), indicating that the genetic architecture governing M. oryzae resistance is more intricate than previously assumed. Complex interactions among pyramided R genes likely contribute to this phenomenon, though their mechanisms remain poorly characterized. Further systematic investigations are required to unravel these genetic complexities. The resistant lines identified in Wanzhou and Enshi disease nurseries showed less than 15% disease resistance (20 lines) in the Meitan nursery (Table 2), indicating significant regional differences in dominant M. oryzae populations. This highlights distinct geographical differentiation of rice blast pathogen populations. In northeastern China, the monogenic lines carrying Pi2 , Piz-t , Pi50 , Pi5 , or Pii gene exhibited strong resistance in local rice blast disease nurseries (Zhang et al., 2022), whereas Pi-1 , Pi2 , and Pi-ta2 confer robust resistance against leaf and panicle neck blast in Hunan Province, southern China (Li et al., 2023). Furthermore, systematic analysis for monogenic lines across 21 rice blast disease nurseries in Sichuan and Chongqing (2012–2013) revealed localized pathogenicity differences of M. oryzae even at smaller geographical scales (Zhang et al., 2017). Such regional pathogen divergence likely results from differential use of resistance genes. Local cultivar preferences impose selection pressures favoring distinct pathogen populations adapted to region-specific resistance profiles (Fukuta et al., 2019). Our data from the Meitan nursery, where M. oryzae displayed higher virulence than in Wanzhou and Enshi (Figure 2C), indirectly reflect these population differences. These findings emphasize the necessity of region-specific resistance evaluations for breeding broad-spectrum blast-resistant lines. The loss of resistance in the selected lines across three disease nurseries further emphasizes the need for continuous breeding to combat the evolving threat of M. oryzae . Resistance in rice varieties often diminishes over time as the pathogen adapts (Mao et al., 2022). This rapid evolution of pathogen virulence necessitates the development of new varieties with enhanced and durable resistance. In this context, molecular marker-assisted selection (MAS) can play a key role in accelerating the identification of superior resistant genotypes. By incorporating molecular markers for R genes, we can more accurately select lines with stable genetic backgrounds and pyramided R genes (Hasan et al., 2021). In our study, the application of MAS allowed for precise identification of R gene compositions in the 136 selected lines, enabling us to select 20 lines with stable resistance across three nurseries (Table 2). These lines show promise for further breeding, but the current evidence suggests that the genetic resistance provided by the pyramided R genes may not be sufficient to withstand evolving pathogen pressures in the long term. Therefore, future breeding programs should focus on exploring new R genes with broad-spectrum resistance or designing novel gene pyramids that combine both known and undetected resistance factors. In summary, pyramiding resistance (R) genes remains a critical approach for combating M. oryzae in rice, yet its efficacy is constrained by inherent challenges. The dynamic evolution of pathogen virulence, regional variations in pathogen populations, and unpredictable epistatic interactions among pyramided R genes demand continuous innovation in breeding programs. Sustained resistance will rely on identifying novel R genes, optimizing their combinations through MAS, and developing varieties with balanced durability and agronomic performance. To address these challenges, future strategies should integrate CRISPR-based editing to optimize R gene expression while reducing fitness costs, regional pathogen surveillance to inform dynamic R gene deployment, and synergistic integration of genetic and agronomic measures to disrupt pathogen adaptation. Materials and Methods Rice germplasm lines Thirteen elite rice germplasm lines were selected as female parents, including Huazhan, Yuehesimiao, Qhui 28, Wushansimiao, Wanhui 88, Wanhui 56, Wanhui 481, Naire 1317, 2015-B 36, Wanhui 16, Qhui 28, Wanhui 96 and Wanhui 99. Twenty-six additional lines served as male parents: Luhui80, Chenghui 727, Wanhui 16, Yunhui 11, Luhui 80, Mianhui 523, CT 18597, CT 18272, Luhui 615, Luhui 37, Huayousizhan, R 24, Wanhui 96, Taiguoxiaoxiangzhan, Guangyou 8, Yuenongsimiao, Wanhui 99, Exiang 1, Huangguangyouzhan, zhonghui 2827, Guiyu 9, Wushansimiao, Huanglizhan, Huazhan, Mabayouzhan, and Kang 4. Localization of rice blast disease nurseries Three rice blast disease nurseries were established for screening the resistance lines against M.oryzae . The Wanzhou nursery is localized at the Ganning base of Three Gorges Academy of Agricultural Sciences in Wanzhou District, Chongqing City (108.24°E, 30.67°N). The Enshi nursery is localized in Lianghekou Village, Enshi Tujia and Miao Autonomous Prefecture, Hubei Province (109.22°E, 30.17°N). The Meitan nursery is localized in Miaotangba Village, Meitan County, Guizhou Province (107.30°E, 27.40°N) (Figure 1A). Classification of rice blast resistance The rice seeds were sown in mid-March annually. Leaf blast symptoms were assessed from May 11 th to June 10 th , and neck blast symptoms were evaluated between July 10 th and August 31 th . Each rice line was planted with 40 individual plants, and three replicates per line were distributed at distinct locations within the disease nursery. Rice cultivation and assessment of rice blast disease on both leaves (tiller stage) and panicle necks (maturation stage) followed the guidelines set by the "NY/T2646-2014 Rice Variety Test Technical Procedures for Identification and Evaluation of Rice Blast Resistance" (Gu et al., 2014). A simplified disease index was applied: Grade 0: No visible symptoms (healthy leaf). Grade 1: Tiny pinhead-sized brown specks only. Grade 2: Slightly larger brown specks. Grade 3: Small circular to slightly elongated brown necrotic gray lesions, 1-2 mm in diameter. Grade 4: Typical spindle-shaped or elliptical blast lesions, 1-2 cm in length, confined to interveinal areas, covering <2% of leaf area. Grade 5: Typical blast lesions covering <10% of leaf area. Grade 6: Typical blast lesions covering 10%-25% of leaf area. Grade 7: Typical blast lesions covering 26%-50% of leaf area. Grade 8: Typical blast lesions covering 51%-75% of leaf area. Grade 9: Complete leaf necrosis (entire leaf dead). Each rice line was planted in three replicates per disease nursery. Three disease indices were derived from these replicates, and the mean value was calculated as the disease index for the line in the nursery. The disease index for each tested line is listed in Table S1. Based on this index, lines were categorized into four resistance groups: Resistance (R), Moderately Resistant (MR), Moderately Susceptible (MS), and Susceptible (S). The resistance levels were defined as follows: R: Index range 0-3. MR: Index range 4-5. MS: Index range 6-7. S: Index range 8-9. Specific molecular markers The primers used in this study were designed based on polymorphic sites identified in previous research (Yang et al., 2024a; Yang et al., 2024b; Yang et al., 2023), ensuring high specificity. To enhance PCR specificity, a mismatch was introduced by mutating the base adjacent to single nucleotide polymorphisms (SNPs) at the 3' end of the primers, creating a two-base mismatch with non-target sites to reduce non-specific amplification. The unique product amplified by these primers was used as a molecular marker to identify the corresponding R genes in the tested lines (Table S3). Genotyping R gene profile in rice The PCR reaction condition was conducted with the following conditions: 2 μL DNA (10 ng/μL), 2.0 μL buffer, 0.5 μL dNTP (2.5 mM each), 0.5 μL forward primer (10 μmol/L), 0.5 μL reverse primer (10 μmol/L), 0.5 μL Taq polymerase (2.5 U/μL, catalog number: ZT101), 14.0 μL ddH 2 O. The cycling protocol was optimized through several interactions with both positive and negative controls to ensure the amplification specificity (Table S3). Declarations Conflict of interest statement On behalf of all authors, the corresponding author states that there is no conflict of interest. Author contributions Conceptualization: Junhua Liu, Chengzhi Huang, Yanyan Huang; Methodology: Junhua Liu, Zhongya Cai, Mei Yang; Formal analysis and investigation: Junhua Liu, Peng Liu, Shufan Lei, Zhiwen Lv; Writing - original draft preparation: Junhua Liu, Yanyan Huang; Writing - review and editing: Junhua Liu, Chengzhi Huang, Yanyan Huang, Shiyan Huang, Zhongxian Liu; Funding acquisition: Junhua Liu, Chengzhi Huang; Resources: Junhua Liu, Chengzhi Huang; Supervision: Chengzhi Huang, Yanyan Huang. Funding This work was supported by research grants from Chongqing technology innovation and application development special key project (CSTB2022TIAD-KPX0018), National modern agricultural industrial technology system construction special project (CARS-01-77), Chongqing modern agricultural industrial technology system (CQMAITS202301), the Science and Technology Research Program of Chongqing Municipal Education Commission (KJQN202201243). Data availability statements The data supporting the results and analysis presented in this article are available upon reasonable request to the corresponding author. Competing Interests All authors certify that they have no affiliations with or involvement in any organization or entity with any financial interest or non-financial interest in the subject matter or materials discussed in this manuscript. References Arora, K., Rai, A. K., Gupta, S. K., Singh, P. K., Narula, A., & Sharma, T. R. (2015). Phenotypic expression of blast resistance gene Pi54 is not affected by its chromosomal position. Plant cell reports, 34(1), 63–70. Ashikawa, I., Hayashi, N., Yamane, H., Kanamori, H., Wu, J., Matsumoto, T., Ono, K., & Yano, M. (2008). Two adjacent nucleotide-binding site-leucine-rich repeat class genes are required to confer Pikm-specific rice blast resistance. Genetics, 180(4), 2267–2276. Ariya-anandech, K., Chaipanya, C., Teerasan, W., Kate-ngam, S., & Jantasuriyarat, C. (2018). 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The Plant cell, 35(5), 1360–1385. Yang H., Huang Y., Yi C., Shi J., Tan C., Ren W., Wang W. (2023). Development and Application of Specific Molecular Markers for Six Homologous Rice Blast Resistance Genes in Pi9 Locus of Rice. Scientia Agricultura Sinica, 56(21):4219-4233. Yang, H., Huang Y., Yi C., Tan C., Ren W., Huang F., Shi J., Li X., Wang W. (2024a). Development and application of specific molecular markers for five allelic rice blast resistance genes in Pik gene site in rice. ACTA PHYTOPATHOLOGICA SINICA, 54(3): 571-581. Yang H., Huang Y., Wang J., Yi C., Shi J., Tan C., Ren W., Wang W. (2024b). Development and Application of Specific Molecular Markers for Eight Rice Blast Resistance Genes in Rice. Chin J Rice Sci, 38(5): 525-534. Zhang, S., Zhong, X. L., Qiao, G. Y., Shen, L., Zhou, T. Y. and Peng, Y. L. (2017). Regional differentiation of Magnaporthe oryzae virulence in Sichuan, Chongqing, and Guizhou. Southwest China Journal of Agricultural Sciences 30(2): 359-365. Zhang, Y. L., Gao, Q., Zhao, Y. H., Liu, R., Fu, Z. J., Li, X., Sun, Y. J. and Jin, X. H. (2022). Evaluation of rice blast resistance and analysis of resistance gene structure in rice germplasm from Heilongjiang Province. Scientia Agricultura Sinica 55(4): 625-645. Zhou, B., Qu, S., Liu, G., Dolan, M., Sakai, H., Lu, G., Bellizzi, M., & Wang, G. L. (2006). The eight amino-acid differences within three leucine-rich repeats between Pi2 and Piz-t resistance proteins determine the resistance specificity to Magnaporthe grisea . Molecular plant-microbe interactions: MPMI, 19(11), 1216–1228. Zeng, S., Li, C., Du, C., Sun, L., Jing, D., Lin, T., Yu, B., Qian, H., Yao, W., Zhou, Y., & Gong, H. (2018). Development of Specific Markers for Pigm in Marker- Assisted Breeding of Panicle Blast Resistant Japonica Rice. Chinese rice science , 05, 453-461. Zhou, K., Zhang, C., Xia, J.,Wang, Y., Yun, P.,Ma, T., Wu, D., & Li, Z. (2022).Associated Analysis of Rice Blast Genotypes and Seedling Blast Resistance of Japonica Rice Resources in the Middle and Lower Reaches of the Yangtze River. Journal of Nuclear Agricultural Sciences, 10,1920-1928. Zhou, Y., Lei, F., Wang, Q., He, W., Yuan, B., & Yuan, W. (2020). Identification of Novel Alleles of the Rice Blast-Resistance Gene Pi9 through Sequence-Based Allele Mining. Rice (New York, N.Y.), 13(1), 80. Zeng, X., Yang, X., Zhao, Z., Lin, F., Wang, L., & Pan, Q. (2011). Characterization and fine mapping of the rice blast resistance gene Pia . Science China. Life sciences, 54(4), 372–378. Zhang, Z., Jia, Y., Wang, Y., & Sun, G. (2020). A Rapid Survey of Avirulence Genes in Field Isolates of Magnaporthe oryzae . Plant disease, 104(3), 717–723 Tables Table 1. Crosses leading to 136 resistance lines selected from WZ and ES disease nurseries NO. Parent lines Number of lines NO. Parent lines Number of lines NO. Parent lines Number of lines 1 Qhui 28/Luhui 80 10 19 18-TL9 2 37 Wanhui 16/Huayousizhan 1 2 Huazhan/Chenghui 727 9 20 Qhui 28/Wanhui 16 2 38 Wanhui 481/CT18597 1 3 Yuehesimiao/Yunhui 11 9 21 Qhui 28/Zhonghui 2827 2 39 Wanhui 56/CT18272 1 4 Yuehesimiao/Wanhui 16 8 22 Huazhan/Wanhui 99 2 40 Wanhui 79 1 5 Huazhan/Luhui 80F4 6 23 Wanhui 99/Guiyu 9 2 41 Wanhui 88/Luhui 37 1 6 Qhui 28/Yunhui 11 5 24 Wanhui 99/Taiguoxiaoxiangzhan 2 42 Wanhui 88/Luhui 615 1 7 Huazhan/Wanhui 16 5 25 Yuehesimiao/Huanglizhan 2 43 Wanhui 90 1 8 Yuehesimiao/Chenghui 727 5 26 Yuehesimiao/Wushansimiao 2 44 Wanhui 96/Huangguangyouzhan 1 9 Qhui 28/Kang 4 4 27 Yuenongsimiao/Guangyou 8 2 45 Wanhui 96/Zhonghui 2827 1 10 Qhui 28/Yuenongsimiao 4 28 Yuenongsimiao/Huazhan 2 46 Wanhui 99/Guangyou 8 1 11 Huazhan/Yunhui 11 4 29 2015-B 36/Mianhui 523 1 47 Wanhui 99/Huangguangyouzhan 1 12 Huazhans/Wanhui 99 3 30 Qhui 28/R24 1 48 Wanhui 99/Wushansimiao 1 13 Wanhui 99/Yuenongsimiao 3 31 Qhui 28/Chenghui 727 1 49 Yuehesimiao/Huazhan 1 14 Wushansimiao/Zhonghui 2827 3 32 Qhui 28/Guangyou 8 1 50 Yuehesimiao/Yuenongsimiao 1 15 Yuehesimiao/Guangyou 8 3 33 Qhui 28/Guiyu 9 1 51 Yuehesimiao/Zhonghui 2827 1 16 Yuenongsimiao/Guiyu 9 3 34 Huazhan/Exiang 1 1 52 Yuenongsimiao/Huangguangyouzhan 1 17 Yuenongsimiao/Wushansimiao 3 35 Huazhans/Wanhui 96 1 53 Yuenongsimiao/Mabayouzhan 1 18 Yuenongsimiao/Zhonghui 2827 3 36 Naire 1317/Chenghui 727 1 54 Yuenongsimiao/Taiguoxiaoxiangzhan 1 Table 2. Twenty resistant lines selected from Meitan rice blase disease nursery NO. Parent lines Generation Meitan R gene composition Leaf blast Neck blast E94 Qhui 28/Chenghui 727 F4 2 5 Pi2+Pi5+Pi54+Pia E97 Yuenongsimiao/Yunhui 11 F4 2 5 Pi5+Pi54+Pia K1 Huazhan/Luhui80 F4 2 5 Pi2+Pi5+Pi54+Pib+Pid3+Pita+Pia K125 Wanhui 96/zhonghui 2827 F4 1 5 Pi5+Pi54+Pib+Pita+Pia K127 Wanhui 99/Taiguoxiaoxiangzhan F4 2 5 Pi2+Pi5+Pid3+Pita K135 Wanhui 99/Wushansimiao F4 2 5 Pi2+Pi5+Pid3+Pita K158 Yuenongsimiao/Guiyu 9 F4 3 5 Pi2+Pi5+Pi54+Pib+Pit K162 Yuenongsimiao/Wushansimiao F4 3 5 Pi2+Pi5+Pi54+Pib+Pit K167 Yuenongsimiao/Huazhan F4 2 5 Pi2+Pi5+Pi54+Pib+Pit+Pita K169 Wushansimiao/zhonghui 2827 F4 1 5 Pi2+Pi5+Pid3+Pita K171 Wushansimiao/zhonghui 2827 F4 2 5 Pi2+Pi5+Pid3+Pita K176 Qhui28/zhonghui 2827 F4 1 5 Pi5+Pi54+Pita K179 Qhui 28/Kang 4 F4 2 5 Pi2+Pi5+Pi54+Pid3+Pita K3 Huazhan/Luhui80 F4 2 5 Pi5+Pi54+Pib+Pid3+Pita+Pia K4 Huazhan/Luhui80 F4 3 5 Pi5+Pib+Pid3+Pita+Pia K42 Qhui 28/Yunhui 11 F4 3 5 Pi5+Pi54+Pita K47 Qhui 28/Luhui 80 F4 2 5 Pi5+Pi54+Pita K51 Qhui 28/Luhui 80 F4 2 5 Pi5+Pid3+Pita+Pia K52 Qhui 28/Luhui 80 F4 2 5 Pi5+Pi54+Pita K63 Yuenongsimiao/Yunhui 11 F4 2 5 Pi54+Pib+Pia Supplementary Files FigureS1.docx TableS3revised.xlsx Cite Share Download PDF Status: Under Review Version 1 posted Reviewers agreed at journal 29 Apr, 2025 Reviewers invited by journal 29 Apr, 2025 Editor assigned by journal 15 Apr, 2025 First submitted to journal 08 Apr, 2025 Editorial decision: Minor revisions 13 Mar, 2025 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-5734717","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":449445837,"identity":"b3f0f712-b952-41d1-a468-56abac592f37","order_by":0,"name":"Junhua Liu","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Junhua","middleName":"","lastName":"Liu","suffix":""},{"id":449445838,"identity":"b2c1a106-301c-44ab-b3ec-4d3bf2d47c90","order_by":1,"name":"Zhongya Cai","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Zhongya","middleName":"","lastName":"Cai","suffix":""},{"id":449445839,"identity":"b67027a3-508f-4b42-b867-475f916a8969","order_by":2,"name":"Mei Yang","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Mei","middleName":"","lastName":"Yang","suffix":""},{"id":449445840,"identity":"2e786019-86a0-4f64-945d-409e24847346","order_by":3,"name":"Peng Liu","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Peng","middleName":"","lastName":"Liu","suffix":""},{"id":449445841,"identity":"62e27928-5700-442c-a0ec-c31eec624082","order_by":4,"name":"Shufan Lei","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Shufan","middleName":"","lastName":"Lei","suffix":""},{"id":449445842,"identity":"32480acb-e830-421c-8efd-834deefb00d2","order_by":5,"name":"Zhiwen Lv","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Zhiwen","middleName":"","lastName":"Lv","suffix":""},{"id":449445843,"identity":"a02105bc-be37-4ddd-be17-a741facceb01","order_by":6,"name":"Shiyan Huang","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Shiyan","middleName":"","lastName":"Huang","suffix":""},{"id":449445844,"identity":"29aea04b-1886-4c25-bcd8-19f7ba2ea0bc","order_by":7,"name":"Zhongxian Liu","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Zhongxian","middleName":"","lastName":"Liu","suffix":""},{"id":449445845,"identity":"055006f7-6308-49e3-a535-a17df070941a","order_by":8,"name":"Yanyan Huang","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAv0lEQVRIiWNgGAWjYBACA2bmg49/MDDIsbG3HyBSCztbsjGQNubjOZNApBZ+HjNpIJ04T8LBgDgt5sw8xsaFbYfT2yQYEhh+VGwjrMWyma3w8cy2w7lt0o0HGHvO3CbCYYeZNxvwgrTIHEhgZmwjSguDmQRQSzqbRIIBsVpYzKSBWhJI0cKWbDjjXLphGzCQDxLnl/OHDz74UGYtL9/efvDBjwoitIABI1szmD5ApHoQ+FNHguJRMApGwSgYcQAAX8A9Qu2/jOYAAAAASUVORK5CYII=","orcid":"","institution":"Sichuan Agricultural University - Chengdu Campus","correspondingAuthor":true,"prefix":"","firstName":"Yanyan","middleName":"","lastName":"Huang","suffix":""},{"id":449445846,"identity":"c71110be-eeec-44be-b6b7-5067a7d0810a","order_by":9,"name":"Chengzhi Huang","email":"","orcid":"","institution":"","correspondingAuthor":false,"prefix":"","firstName":"Chengzhi","middleName":"","lastName":"Huang","suffix":""}],"badges":[],"createdAt":"2024-12-30 10:18:27","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-5734717/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-5734717/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":81671626,"identity":"1558715c-6fe3-4aea-9a61-902ab657c1aa","added_by":"auto","created_at":"2025-04-30 06:00:47","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":156027,"visible":true,"origin":"","legend":"\u003cp\u003eSelection process for rice resistance lines.A. Localization of rice blast disease nurseries. The red triangles indicate the locations of the rice blast disease nurseries, located in Wanzhou (WZ), Enshi (ES), and Meitan (MT), which are key sites for evaluating blast disease resistance in rice. B. Selection process for rice resistance lines. This section outlines the steps in selecting rice lines with resistance to blast disease\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-5734717/v1/d9ad87333c8c9206e88898e6.png"},{"id":81671625,"identity":"e44b68fe-7087-4c22-aea7-178a72b78599","added_by":"auto","created_at":"2025-04-30 06:00:47","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":70915,"visible":true,"origin":"","legend":"\u003cp\u003eResistance analysis of 136 rice lines in different disease nurseries. A-C. Resistance analysis of leaf blast and neck blast at various disease nurseries. Resistance levels of the 136 rice lines were evaluated for leaf blast and neck blast at three different disease nurseries: Wanzhou (A), Enshi (B), and Meitan (C).\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-5734717/v1/8f104b82a66c1f8d9ebb10d9.png"},{"id":81671631,"identity":"1bf9c6ca-2459-4d76-9f8a-11bc014952b8","added_by":"auto","created_at":"2025-04-30 06:00:59","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":74514,"visible":true,"origin":"","legend":"\u003cp\u003eGenotyping of R genes in 136 rice lines. A. Distribution frequency of each R gene in the rice lines. The data highlight the prevalence of individual R genes across the evaluated lines. B. Ratio of lines carrying different numbers of R genes.\u003c/p\u003e","description":"","filename":"3.png","url":"https://assets-eu.researchsquare.com/files/rs-5734717/v1/2c44968dd64c80abab8835a6.png"},{"id":81671630,"identity":"9ce24b66-9f11-43e6-b047-e9ee31d783ee","added_by":"auto","created_at":"2025-04-30 06:00:58","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":223558,"visible":true,"origin":"","legend":"\u003cp\u003eExpression dynamics of R genes in rice lines K55 and K52 following \u003cem\u003eM. oryzae\u003c/em\u003e inoculation. A. Disease symptom on leaves of K55 and K52 after spray inoculation with \u003cem\u003eM. oryzae\u003c/em\u003e isolate DZ98. Representative images were captured at 5 days post-inoculation. B. Semi-quantitative PCR analysis of \u003cem\u003ePita\u003c/em\u003e, \u003cem\u003ePi5\u003c/em\u003e, and\u003cem\u003e Pi54 \u003c/em\u003eexpression in leaves sampled at 0, 12, and 48 hours post-inoculation. The ubiquitin gene (\u003cem\u003eUBI\u003c/em\u003e) served as an internal control. Primer sequences are provided in Table S3.\u003c/p\u003e","description":"","filename":"4.png","url":"https://assets-eu.researchsquare.com/files/rs-5734717/v1/de1ad163baa5499afc1063fe.png"},{"id":81672044,"identity":"ace85fc5-de5a-4f7c-adc2-f04e4fba7f35","added_by":"auto","created_at":"2025-04-30 06:08:48","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1434435,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-5734717/v1/ec73f8d3-24de-47f9-9535-44c36171cdfe.pdf"},{"id":81671627,"identity":"d06d77d3-0123-4245-8c38-d5e4dc5a4e80","added_by":"auto","created_at":"2025-04-30 06:00:47","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":158932,"visible":true,"origin":"","legend":"","description":"","filename":"FigureS1.docx","url":"https://assets-eu.researchsquare.com/files/rs-5734717/v1/1479d6a1cb2e0b055eace19c.docx"},{"id":81671634,"identity":"60b26aa4-eb7e-4e6d-9fdf-74ff141d8e5b","added_by":"auto","created_at":"2025-04-30 06:01:26","extension":"xlsx","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":16593,"visible":true,"origin":"","legend":"","description":"","filename":"TableS3revised.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-5734717/v1/8b01b00049ec88415d034d34.xlsx"}],"financialInterests":"","formattedTitle":"Challenges and Insights into Pyramided R Genes for Durable Rice Blast Resistance in Southwest China","fulltext":[{"header":"Introduction","content":"\u003cp\u003eRice is a staple food in China, nourishing over half of the country\u0026apos;s population and playing a crucial role in both social and economic development\u0026nbsp;(Zhou et al., 2006). However, Rice production is constantly threatened by various biotic and abiotic stresses. Among these, rice blast disease, caused by \u003cem\u003eMagnaporthe oryzae\u003c/em\u003e (\u003cem\u003eM. oryzae\u003c/em\u003e), is one of the most devastating, impacting rice yields in over 85 countries worldwide (Wang et al., 2014). In China, annual yield losses due to rice blast exceed 0.4 million tons, severely threatening food security\u0026nbsp;(Wancai, Zhendong et al. 2016). Although fungicides are widely used to control rice blast, their overuse has led to the emergence of fungicide-resistant \u003cem\u003eM. oryzae\u0026nbsp;\u003c/em\u003estrains, posing significant risks to human health and the environment\u0026nbsp;(Luo et al., 2005). Therefore, the resistance breeding is considered the most cost-effective and environmentally sustainable strategy for managing rice blast disease (Wu et al., 2021). This dilemma underscores the urgency to develop sustainable disease management strategies.\u003c/p\u003e\n\u003cp\u003eSince the cloning of the first blast resistance gene \u003cem\u003ePib\u003c/em\u003e in 1996, over 50 dominant R genes have been identified at 17 chromosomal loci in rice. Most of them as allelic variants, particularly at the \u003cem\u003ePi2\u003c/em\u003e and \u003cem\u003ePik\u003c/em\u003e loci localized on chromosomes 6 and 11, respectively (Devanna et al., 2022; Huang et al., 2023). These genes encode conserved nucleotide-binding-site leucine-rich-repeat (NBS-LRR) proteins, which recognize corresponding avirulence proteins secreted by\u003cem\u003e\u0026nbsp;M. oryzae\u003c/em\u003e, triggering strong defense responses, including ROS burst,\u0026nbsp;hypersensitive response (HR) and transcriptional reprogramming (Liu et al., 2021; Singh et al., 2016).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eResistance genes have been extensively applied in rice breeding and production, effectively safeguarding the safe cultivation of rice. However, the efficacy of these resistance genes demonstrates region-specific characteristics (Rathour\u0026nbsp;et al.,\u0026nbsp;2016;\u0026nbsp;Liu\u0026nbsp;et al.,\u0026nbsp;2021; Zeng\u0026nbsp;et al.,\u0026nbsp;2018). For example, the \u003cem\u003ePikm\u003c/em\u003e,\u003cem\u003e\u0026nbsp;Pi54\u003c/em\u003e, and\u003cem\u003e\u0026nbsp;Pib\u003c/em\u003e genes confer the strongest resistance in Huanghuai, Shandong and Jiangsu rice cultivation regions. In contrast, the \u003cem\u003ePi5\u0026nbsp;\u003c/em\u003eand \u003cem\u003ePita\u0026nbsp;\u003c/em\u003egenes are particularly effective in the middle and lower reaches of the Yangtze River (Wang et al., 2016; Chen et al., 2018; Zhou et al., 2022), while the \u003cem\u003ePi2\u003c/em\u003e, \u003cem\u003ePish\u003c/em\u003e, \u003cem\u003ePikh\u003c/em\u003e, \u003cem\u003ePi9\u0026nbsp;\u003c/em\u003eand \u003cem\u003ePi41\u0026nbsp;\u003c/em\u003egenes are most effective\u0026nbsp;in Guangdong province\u0026nbsp;(Yang\u0026nbsp;et al., 2008; Wu et al., 2023). However, the resistance conferred by individual \u003cem\u003eR\u003c/em\u003e gene has been increasingly overcome by \u003cem\u003eM. oryzae\u003c/em\u003e.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eNevertheless, sustaining durable disease resistance remains a pivotal challenge in rice breeding. Pathogen populations undergo rapid co-evolutionary adaptation, frequently acquiring effector gene mutations that evade detection by host R proteins, ultimately leading to resistance breakdown. Notably, emerging empirical evidence demonstrates that pyramiding multiple R genes enhances broad-spectrum and durable resistance against \u003cem\u003eM. oryzae\u003c/em\u003e in rice breeding, providing a genetically reinforced defense mechanism against pathogen evolution. For instance, pyramiding two \u003cem\u003eR\u003c/em\u003e genes, such as\u0026nbsp;\u003cem\u003ePi-1\u0026nbsp;\u003c/em\u003ewith \u003cem\u003ePi-2\u003c/em\u003e, \u003cem\u003ePi-46\u0026nbsp;\u003c/em\u003ewith \u003cem\u003ePi-ta, or Pi9\u0026nbsp;\u003c/em\u003ewith \u003cem\u003ePi54\u003c/em\u003e,\u003cem\u003e\u0026nbsp;\u003c/em\u003esignificantly enhanced resistance compares to varieties carrying a single \u003cem\u003eR\u003c/em\u003e gene (Chen et al., 2001; Xiao et al., 2016; Zhou et al., 2020). However, not\u0026nbsp;all combination of \u003cem\u003eR\u003c/em\u003e gene pyramiding enhances\u0026nbsp;rice blast resistance. For example, although pyramided lines carrying\u003cem\u003e\u0026nbsp;Pi9+Pi54\u003c/em\u003e and \u003cem\u003ePizt+Pi54\u003c/em\u003e exhibited higher resistance to leaf blast than the lines carrying the corresponding single \u003cem\u003eR\u003c/em\u003e gene, pyramiding of\u003cem\u003e\u0026nbsp;Pi9+Pi54\u003c/em\u003e unexpectedly reduced neck blast resistance compared to \u003cem\u003ePi9\u0026nbsp;\u003c/em\u003ealone\u0026nbsp;(Xiao\u0026nbsp;et al., 2017).\u0026nbsp;These limitations highlight the need for region-specific R gene deployment strategies based on pathogen population dynamics.\u003c/p\u003e\n\u003cp\u003eSouthwest China represents a high-risk zone for rice blast due to its humid subtropical climate and complex pathogen diversity (Hu, Huang et al. 2022). To develop cultivars with durable blast resistance, we implemented an integrated approach combining multi-location field evaluation and molecular marker-assisted selection. Progenies derived from elite crosses were systematically screened across three disease nurseries (Wanzhou, Enshi, Meitan). Molecular characterization of 136 resistant lines identified 14 functional resistance genes through allele-specific marker analysis, including \u003cem\u003ePi5\u003c/em\u003e (Lee\u0026nbsp;et al., 2009), \u003cem\u003ePi54\u003c/em\u003e (Arora\u0026nbsp;et al., 2015), \u003cem\u003ePi-ta\u003c/em\u003e (Lee\u0026nbsp;et al., 2011), \u003cem\u003ePia\u003c/em\u003e (Zeng\u0026nbsp;et al., 2011), \u003cem\u003ePib\u003c/em\u003e (Wang\u0026nbsp;et al., 1999), \u003cem\u003ePi2\u003c/em\u003e (Zhou\u0026nbsp;et al., 2006), \u003cem\u003ePid3\u003c/em\u003e (Xu\u0026nbsp;et al., 2014), \u003cem\u003ePik-m\u0026nbsp;\u003c/em\u003e(Ashikawa\u0026nbsp;et al., 2008), \u003cem\u003ePit\u0026nbsp;\u003c/em\u003e(Li\u0026nbsp;et al., 2014), \u003cem\u003ePigm\u0026nbsp;\u003c/em\u003e(Deng\u0026nbsp;et al., 2017),\u003cem\u003e\u0026nbsp;Pi1\u0026nbsp;\u003c/em\u003e(Hua\u0026nbsp;et al., 2012), \u003cem\u003ePi9\u0026nbsp;\u003c/em\u003e(Chung\u0026nbsp;et al., 2015),\u003cem\u003e\u0026nbsp;Pizt\u0026nbsp;\u003c/em\u003e(Ning\u0026nbsp;et al., 2020;Yokoo, 1983), \u003cem\u003ePik\u0026nbsp;\u003c/em\u003e( Ariya-anandech et al., 2018). Twenty F4 breeding lines demonstrating broad-spectrum resistance were ultimately selected, providing critical genetic resources and strategies to breed durable blast-resistant cultivars for Southwest China\u0026rsquo;s high-risk regions.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003e\u003cstrong\u003eDevelopment of F2 segregating populations for screening of resistant restorer lines\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo develop resistant restorer lines adapted to the unique climatic conditions of the mountainous rice-growing regions in southwest China, we selected thirteen elite rice germplasm lines, including Huazhan, Yuehesimiao, Qhui 28 and ten additional lines detailed in Table S1 all of which had demonstrated excellent agronomic performance and stable high yields at Wanzhou over several years. These elite lines were then crossed with 26 donor germplasm lines, including Yunhui 11, Luhui 80, Chenghui 727 and\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003eothers listed in Table S1, previously characterized for durable resistant to \u003cem\u003eM. oryzae\u003c/em\u003e through systematic phenotyping in disease nurseries at Wanzhou and Enshi (Fig. 1A). Fifty-four F1 hybrid plants were generated from these crosses. The F1 plants were cultivated at Wanzhou and subjected to self-pollination to derive F2 segregating populations for subsequent resistance screening.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eScreening the resistant lines at Wanzhou and Enshi disease nurseries\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo identify blast-resistant germplasm, we first cultivated the 54 F2 segregating populations in the rice blast disease nursery at Wanzhou (Fig. 1A and 1B), where 800 lines resistant to both leaf blast and neck blast, while exhibiting high yield and elite agronomic traits, were selected. To validate resistance stability across environments, the F3 progeny lines derived these selections were further evaluated in the distinct epidemiological conditions of the Enshi disease nursery. 136 lines were selected for exhibiting high or moderate resistance to neck blast (Table 1). Notably, the cross between Qhui28 and Luhui 80 exhibited superior breeding efficiency, generating 10 resistance lines, the highest among all combinations. In contrast, 27 crosses yielded 2-9 lines each, and 26 crosses yielded single resistant line, such as the cross between 2015-B 36 and Mianhui 523. The data reflects a significant variation in combinatory resistance potential.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eThe selected lines exhibited stable resistance across generations and environments\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo evaluate the heritability of resistance traits, the 136 F4 progeny lines were re-evaluated in Wanzhou and Enshi nurseries. In Wanzhou, all lines demonstrated resistance to\u003cem\u003e\u0026nbsp;\u003c/em\u003eleaf blast disease without segregation. For neck blast resistance, 2.94% of lines exhibited stable resistance, 95.59% showed moderate resistance, and 1.47% displayed moderate susceptibility (Figure 2A). In Enshi, 2.21% of the lines exhibited medium resistant to leaf blast disease, while the remaining lines maintained resistance. Neck blast disease evaluations identified 2.21% resistant lines, 96.32% moderately resistant, and 1.47% moderately susceptible (Fig. 2B). The consistent phenotypic performance patterns observed across both geographical locations and generational propagation confirm the high heritability and environmental stability of these resistance traits.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eThe broad-spectrum resistance evaluation in Meitan rice blast nursery\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo characterize broad-spectrum resistance adaptability across heterogeneous rice ecosystems, we tested the 136 F4 lines at the Meitan disease nursery in Zunyi city, Guizhou province, where shares climatic similarities with Chongqing. All lines retained leaf blast resistance or moderate resistance phenotypes (Fig. 2A and B). In contrast, neck blast resistance displayed progressive attenuation, with no lines demonstrating resistance. Specifically, 20 lines (15.44%) demonstrated moderate neck blast resistance, 114 lines (83.82%) exhibited moderate susceptibility, and a single line (0.74%) showed full susceptibility\u003cem\u003e\u0026nbsp;\u003c/em\u003e(Fig. 2C). Ultimately, twenty lines were selected for rice resistance breeding based on their consistent resistance across the three disease nurseries. These lines exhibited minimal variability in resistance scores over multiple seasons, demonstrating strong potential for broad-spectrum resistance breeding (Table 2).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eNotably, compared to the uniform resistance patterns observed in Wanzhou and Enshi, the Meitan trials revealed substantial phenotypic variation among lines (Fig. 2), indicating broader pathogenic diversity within the \u003cem\u003eM. oryzae\u003c/em\u003e population at this site. This geographical divergence in pathogen virulence profiles underscores the necessity of multi-location screening for breeding durable blast resistance.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eR gene composition in the 136 resistance lines\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eResistance to \u003cem\u003eM. oryzae\u003c/em\u003e is primarily conferred by \u003cem\u003eR\u003c/em\u003e genes through effector-triggered immunity (ETI) in rice. To characterize resistance mechanisms underlying variation in blast resistance, we performed molecular profiling of 14 NLR genes, including\u0026nbsp;\u003cem\u003ePi5\u003c/em\u003e, \u003cem\u003ePi54\u003c/em\u003e, \u003cem\u003ePita\u003c/em\u003e, \u003cem\u003ePia\u003c/em\u003e, \u003cem\u003ePib\u003c/em\u003e, \u003cem\u003ePi2\u003c/em\u003e, \u003cem\u003ePid3\u003c/em\u003e, \u003cem\u003ePikm\u003c/em\u003e, \u003cem\u003ePit\u003c/em\u003e, \u003cem\u003ePigmR\u003c/em\u003e, \u003cem\u003ePi1\u003c/em\u003e, \u003cem\u003ePi9\u003c/em\u003e, \u003cem\u003ePizt\u0026nbsp;\u003c/em\u003eand \u003cem\u003ePik,\u003c/em\u003e across 136 F4 lines using gene-specific molecular markers (Figure S1 and Table S3). Three genes, \u003cem\u003ePi9\u003c/em\u003e, \u003cem\u003ePizt\u0026nbsp;\u003c/em\u003eand \u003cem\u003ePik\u003c/em\u003e, were absent in all analyzed lines. Seven\u003cem\u003e\u0026nbsp;\u003c/em\u003egenes, \u003cem\u003ePi2\u003c/em\u003e, \u003cem\u003ePib\u003c/em\u003e, \u003cem\u003ePid3\u003c/em\u003e, \u003cem\u003ePit\u003c/em\u003e, \u003cem\u003ePikm\u003c/em\u003e, \u003cem\u003ePi1\u003c/em\u003e and \u003cem\u003ePigmR,\u0026nbsp;\u003c/em\u003ewas detected in 49.26%, 49.26%, 30.15%, 16.18%, 16.18%, 2.21%and 0.74% of the lines, respectively. The remaining four \u003cem\u003eR\u003c/em\u003e genes, \u003cem\u003ePi5\u003c/em\u003e, \u003cem\u003ePi54\u003c/em\u003e, \u003cem\u003ePita\u003c/em\u003e and \u003cem\u003ePia\u003c/em\u003e, were detected in more than 50% of the lines, with distribution frequency of 82.35%, 62.50%, 54.41%, and 53.680%, respectively (Figure 3A).\u003c/p\u003e\n\u003cp\u003eThe number of R genes per line varied from one to seven, with a normal distribution observed (Fig. 3B). While only one line carried a single R gene, the majority of the lines contained three to five R genes. This distribution suggests a high level of genetic diversity in the resistance composition among the F4 lines.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eUndetected R genes\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003emay exist in the resistant lines to confer resistance in Meitan nursery\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eIn the Meitan disease nursery, 143 rice lines exhibited susceptible phenotypes to\u003cem\u003e\u0026nbsp;M. oryzae\u003c/em\u003e infection, indicating compromised functionality of their intrinsic resistance genotypes under prevailing field conditions. Notably, the characterized resistance gene compositions identified in 20 resistant lines were paradoxically present in susceptible lines. For instance, resistant lines, K52, K42, K176 and K53, shared same resistance genotypes (\u003cem\u003ePi5\u003c/em\u003e + \u003cem\u003ePi54\u003c/em\u003e + \u003cem\u003ePita\u003c/em\u003e) with susceptible lines, K55, K57, K33, K174, K44, K45 and K103 (Table S1). Transcriptomic analysis revealed comparable expression patterns of\u003cem\u003e\u0026nbsp;Pi5\u003c/em\u003e, \u003cem\u003ePi54\u003c/em\u003e, and \u003cem\u003ePita\u003c/em\u003e in both resistant line K52 and susceptible line K55, both pre- and post-pathogen inoculation (Figure 4). This phenotypic-genotypic paradox implies that unknown or undetected known R genes not captured by in this study likely govern field resistance against \u003cem\u003eM. oryzae\u003c/em\u003e. The identified resistant lines constitute valuable germplasm resources for discovery of novel R gene alleles, and optimization of pyramiding breeding strategies targeting durable blast resistance.\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eResistance to \u003cem\u003eM. oryzae\u003c/em\u003e in rice is primarily conferred by R genes through effector-triggered immunity (ETI), which involves the recognition of pathogen-derived Avr proteins by plant immune receptors, triggering strong immune responses such as ROS bursts, hypersensitive responses (HR), and transcriptional reprogramming. These responses inhibit disease development by limiting the growth of \u003cem\u003eM. oryzae\u003c/em\u003e isolates within the cells surrounding the infection site (Zhang et al., 2020; Kim et al., 2004; Cesari et al., 2013; Singh et al., 2016; Yan et al., 2023; Korinsak et al., 2022). Over 50 R genes have been cloned in rice, many of which have been utilized in breeding programs to improve resistance to rice blast (Li et al., 2019). However, resistance breeding faces significant challenges, especially the rapid evolution of \u003cem\u003eM. oryzae. Resistance (R) genes impose strong selective pressure on M. oryzae populations. Strains harboring recognized Avr genes are suppressed in the field, while those with Avr loss (via mutations, deletions, or silencing) evade R gene recognition (Hu et al., 2022). Over time, these escaped strains dominate the pathogen population, leading to resistance breakdown in previously resistant rice varieties.\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eIn this study, we selected 136 resistant rice lines from two disease nurseries (Wanzhou and Enshi), which exhibited stable resistance to \u003cem\u003eM. oryzae\u003c/em\u003e across multiple years. The majority of these lines harbor three or more pyramided R genes, demonstrating that R gene pyramiding has become a common practice in modern rice germplasm and breeding programs. This strategy effectively enhances blast resistance and delays resistance breakdown. However, pyramiding resistance genes presents challenges, as not all combinations of pyramided genes necessarily improve resistance (Wang et al., 2020; Variar et al., 2009). Our findings align with these reports. Notably, the genotypes of resistant individuals identified in the Meitan disease nursery were also detected in susceptible lines (Table S1), suggesting that unidentified or uncharacterized known resistance genes may predominantly govern resistance in these lines. In contrast, susceptible lines carrying multiple pyramided R genes, including those with up to eight in our study, still displayed susceptibility. This highlights that effective resistance in pyramided lines primarily relies on the presence of major-effect R genes, while the additive contributions of other pyramided genes may be less pronounced. Previous studies further confirm a non-linear correlation between R gene number and resistance intensity (Zhou et al., 2022), indicating that the genetic architecture governing \u003cem\u003eM. oryzae\u003c/em\u003e resistance is more intricate than previously assumed. Complex interactions among pyramided R genes likely contribute to this phenomenon, though their mechanisms remain poorly characterized. Further systematic investigations are required to unravel these genetic complexities.\u003c/p\u003e\n\u003cp\u003eThe resistant lines identified in Wanzhou and Enshi disease nurseries showed less than 15% disease resistance (20 lines) in the Meitan nursery (Table 2), indicating significant regional differences in dominant \u003cem\u003eM. oryzae\u003c/em\u003e populations. This highlights distinct geographical differentiation of rice blast pathogen populations. In northeastern China, the monogenic lines carrying \u003cem\u003ePi2\u003c/em\u003e, \u003cem\u003ePiz-t\u003c/em\u003e, \u003cem\u003ePi50\u003c/em\u003e, \u003cem\u003ePi5\u003c/em\u003e, or \u003cem\u003ePii\u0026nbsp;\u003c/em\u003egene exhibited strong resistance in local rice blast disease nurseries (Zhang et al., 2022), whereas \u003cem\u003ePi-1\u003c/em\u003e, \u003cem\u003ePi2\u003c/em\u003e, and \u003cem\u003ePi-ta2\u003c/em\u003e confer robust resistance against leaf and panicle neck blast in Hunan Province, southern China (Li et al., 2023). Furthermore, systematic analysis for monogenic lines across 21 rice blast disease nurseries in Sichuan and Chongqing (2012\u0026ndash;2013) revealed localized pathogenicity differences of\u003cem\u003e\u0026nbsp;M. oryzae\u003c/em\u003e even at smaller geographical scales (Zhang et al., 2017). Such regional pathogen divergence likely results from differential use of resistance genes. Local cultivar preferences impose selection pressures favoring distinct pathogen populations adapted to region-specific resistance profiles (Fukuta et al., 2019). Our data from the Meitan nursery, where\u003cem\u003e\u0026nbsp;M. oryzae\u0026nbsp;\u003c/em\u003edisplayed higher virulence than in Wanzhou and Enshi (Figure 2C), indirectly reflect these population differences. These findings emphasize the necessity of region-specific resistance evaluations for breeding broad-spectrum blast-resistant lines.\u003c/p\u003e\n\u003cp\u003eThe loss of resistance in the selected lines across three disease nurseries further emphasizes the need for continuous breeding to combat the evolving threat of \u003cem\u003eM. oryzae\u003c/em\u003e. Resistance in rice varieties often diminishes over time as the pathogen adapts (Mao et al., 2022). This rapid evolution of pathogen virulence necessitates the development of new varieties with enhanced and durable resistance. In this context, molecular marker-assisted selection (MAS) can play a key role in accelerating the identification of superior resistant genotypes. By incorporating molecular markers for R genes, we can more accurately select lines with stable genetic backgrounds and pyramided R genes (Hasan et al., 2021).\u003c/p\u003e\n\u003cp\u003eIn our study, the application of MAS allowed for precise identification of R gene compositions in the 136 selected lines, enabling us to select 20 lines with stable resistance across three nurseries (Table 2). These lines show promise for further breeding, but the current evidence suggests that the genetic resistance provided by the pyramided R genes may not be sufficient to withstand evolving pathogen pressures in the long term. Therefore, future breeding programs should focus on exploring new R genes with broad-spectrum resistance or designing novel gene pyramids that combine both known and undetected resistance factors.\u003c/p\u003e\n\u003cp\u003eIn summary, pyramiding resistance (R) genes remains a critical approach for combating \u003cem\u003eM. oryzae\u003c/em\u003e in rice, yet its efficacy is constrained by inherent challenges. The dynamic evolution of pathogen virulence, regional variations in pathogen populations, and unpredictable epistatic interactions among pyramided R genes demand continuous innovation in breeding programs. Sustained resistance will rely on identifying novel R genes, optimizing their combinations through MAS, and developing varieties with balanced durability and agronomic performance. To address these challenges, future strategies should integrate CRISPR-based editing to optimize R gene expression while reducing fitness costs, regional pathogen surveillance to inform dynamic R gene deployment, and synergistic integration of genetic and agronomic measures to disrupt pathogen adaptation.\u0026nbsp;\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cp\u003e\u003cstrong\u003eRice germplasm lines\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThirteen elite rice germplasm lines were selected as female parents, including Huazhan, Yuehesimiao, Qhui 28, Wushansimiao, Wanhui 88, Wanhui 56, Wanhui 481, Naire 1317, 2015-B 36, Wanhui 16, Qhui 28, Wanhui 96 and Wanhui 99. Twenty-six additional lines served as male parents: Luhui80, Chenghui 727, Wanhui 16, Yunhui 11, Luhui 80, Mianhui 523, CT 18597, CT 18272, Luhui 615, Luhui 37, Huayousizhan, R 24, Wanhui 96, Taiguoxiaoxiangzhan, Guangyou 8, Yuenongsimiao, Wanhui 99, Exiang 1, Huangguangyouzhan, zhonghui 2827, Guiyu 9, Wushansimiao, Huanglizhan, Huazhan, Mabayouzhan, and Kang 4.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eLocalization of rice blast disease nurseries\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThree rice blast disease nurseries were established for screening the resistance lines against \u003cem\u003eM.oryzae\u003c/em\u003e. The Wanzhou nursery is localized at the Ganning base of Three Gorges Academy of Agricultural Sciences in Wanzhou District, Chongqing City (108.24\u0026deg;E, 30.67\u0026deg;N). The Enshi nursery is localized in Lianghekou Village, Enshi Tujia and Miao Autonomous Prefecture, Hubei Province (109.22\u0026deg;E, 30.17\u0026deg;N). The Meitan nursery is localized in Miaotangba Village, Meitan County, Guizhou Province (107.30\u0026deg;E, 27.40\u0026deg;N) (Figure 1A).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eClassification of rice blast resistance\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe rice seeds were sown in mid-March annually. Leaf blast symptoms were assessed from May 11\u003csup\u003eth\u003c/sup\u003e to June 10\u003csup\u003eth\u003c/sup\u003e, and neck blast symptoms were evaluated between July 10\u003csup\u003eth\u003c/sup\u003e and August 31\u003csup\u003eth\u003c/sup\u003e. Each rice line was planted with 40 individual plants, and three replicates per line were distributed at distinct locations within the disease nursery.\u003c/p\u003e\n\u003cp\u003eRice cultivation and assessment of rice blast disease on both leaves (tiller stage) and panicle necks (maturation stage) followed the guidelines set by the \u0026quot;NY/T2646-2014 Rice Variety Test Technical Procedures for Identification and Evaluation of Rice Blast Resistance\u0026quot; (Gu et al., 2014). A simplified disease index was applied: Grade 0: No visible symptoms (healthy leaf). Grade 1: Tiny pinhead-sized brown specks only. Grade 2: Slightly larger brown specks. Grade 3: Small circular to slightly elongated brown necrotic gray lesions, 1-2 mm in diameter. Grade 4: Typical spindle-shaped or elliptical blast lesions, 1-2 cm in length, confined to interveinal areas, covering \u0026lt;2% of leaf area. Grade 5: Typical blast lesions covering \u0026lt;10% of leaf area. Grade 6: Typical blast lesions covering 10%-25% of leaf area. Grade 7: Typical blast lesions covering 26%-50% of leaf area. Grade 8: Typical blast lesions covering 51%-75% of leaf area. Grade 9: Complete leaf necrosis (entire leaf dead).\u003c/p\u003e\n\u003cp\u003eEach rice line was planted in three replicates per disease nursery. Three disease indices were derived from these replicates, and the mean value was calculated as the disease index for the line in the nursery. The disease index for each tested line is listed in Table S1. Based on this index, lines were categorized into four resistance groups: Resistance (R), Moderately Resistant (MR), Moderately Susceptible (MS), and Susceptible (S). The resistance levels were defined as follows: R: Index range 0-3. MR: Index range 4-5. MS: Index range 6-7. S: Index range 8-9.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eSpecific molecular markers\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe primers used in this study were designed based on polymorphic sites identified in previous research (Yang et al., 2024a; Yang et al., 2024b; Yang et al., 2023), ensuring high specificity. To enhance PCR specificity, a mismatch was introduced by mutating the base adjacent to single nucleotide polymorphisms (SNPs) at the 3\u0026apos; end of the primers, creating a two-base mismatch with non-target sites to reduce non-specific amplification. The unique product amplified by these primers was used as a molecular marker to identify the corresponding R genes in the tested lines (Table S3).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eGenotyping R gene profile in rice\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe\u0026nbsp;PCR reaction condition was conducted with the following conditions: 2 \u0026mu;L DNA (10 ng/\u0026mu;L), 2.0 \u0026mu;L buffer, 0.5 \u0026mu;L dNTP (2.5 mM each), 0.5 \u0026mu;L forward primer (10 \u0026mu;mol/L), 0.5 \u0026mu;L reverse primer (10 \u0026mu;mol/L), 0.5 \u0026mu;L Taq polymerase (2.5 U/\u0026mu;L, catalog number: ZT101), 14.0 \u0026mu;L ddH\u003csub\u003e2\u003c/sub\u003eO. The cycling protocol was optimized through several interactions with both positive and negative controls to ensure the amplification specificity (Table S3).\u0026nbsp;\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eConflict of interest statement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eOn behalf of all authors, the corresponding author states that there is no conflict of interest.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eConceptualization: Junhua Liu, Chengzhi Huang, Yanyan Huang; Methodology: Junhua Liu, Zhongya Cai, Mei Yang; Formal analysis and investigation: Junhua Liu, Peng Liu, Shufan Lei, Zhiwen Lv; Writing - original draft preparation: Junhua Liu, Yanyan Huang; Writing - review and editing: Junhua Liu, Chengzhi Huang, Yanyan Huang, Shiyan Huang, Zhongxian Liu; Funding acquisition: Junhua Liu, Chengzhi Huang; Resources: Junhua Liu, Chengzhi Huang; Supervision: Chengzhi Huang, Yanyan Huang.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis work was supported by research grants from Chongqing technology innovation and application development special key project (CSTB2022TIAD-KPX0018), National modern agricultural industrial technology system construction special project (CARS-01-77), Chongqing modern agricultural industrial technology system (CQMAITS202301), the Science and Technology Research Program of Chongqing Municipal Education Commission (KJQN202201243).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData availability statements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe data supporting the results and analysis presented in this article are available upon reasonable request to the corresponding author.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting Interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll authors certify that they have no affiliations with or involvement in any organization or entity with any financial interest or non-financial interest in the subject matter or materials discussed in this manuscript.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n \u003cli\u003eArora, K., Rai, A. 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Pyramiding of \u003cem\u003ePi46\u0026nbsp;\u003c/em\u003eand \u003cem\u003ePita\u003c/em\u003e to improve blast resistance and to evaluate the resistance effect of the two R genes.\u003cem\u003e\u0026nbsp;Journal of Integrative Agriculture,\u0026nbsp;\u003c/em\u003e15, 2290-2298.\u003c/li\u003e\n \u003cli\u003eXiao, N., Wu, Y., Pan, C., Yu, L., Chen, Y., Liu, G., Li, Y., Zhang, X., Wang, Z., Dai, Z., Liang, C., \u0026amp; Li, A. (2017). Improving of Rice Blast Resistances in Japonica by Pyramiding Major R Genes. \u003cem\u003eFrontiers in plant science,\u0026nbsp;\u003c/em\u003e7, 1918.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eXu, X., Lv, Q., Shang, J., Pang, Z., Zhou, Z., Wang, J., Jiang, G., Tao, Y., Xu, Q., Li, X., Zhao, X., Li, S., Xu, J., \u0026amp; Zhu, L. (2014). Excavation of \u003cem\u003ePid3\u003c/em\u003e orthologs with differential resistance spectra to \u003cem\u003eMagnaporthe oryzae\u0026nbsp;\u003c/em\u003ein rice resource. \u003cem\u003ePloS one,\u0026nbsp;\u003c/em\u003e9(3), e93275.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eYang, J.,Chen, S.,Zeng, L.,Li, Y.,Chen, Z.,Zhu, X. (2008). Evaluation on Resistance of Major Rice Blast Resistance Genes to\u003cem\u003e\u0026nbsp;Magnaporthe grisea\u003c/em\u003e Isolates Collected from indica Rice in Guangdong Province China. \u003cem\u003eChain J Rice Sci,\u0026nbsp;\u003c/em\u003e22( 2) : 190-196.\u003c/li\u003e\n \u003cli\u003eYokoo, M. (1983). Near-Isogenic Lines of Rice with Respect to a \u003cem\u003ePi-zt\u0026nbsp;\u003c/em\u003eGene for Resistance to Blast Disease.\u003cem\u003e\u0026nbsp;Breeding Science,\u0026nbsp;\u003c/em\u003e33, 341-345.\u003c/li\u003e\n \u003cli\u003eYan, X., Tang, B., Ryder, L. S., MacLean, D., Were, V. M., Eseola, A. B., Cruz-Mireles, N., Ma, W., Foster, A. J., Os\u0026eacute;s-Ruiz, M., \u0026amp; Talbot, N. J. (2023). The transcriptional landscape of plant infection by the rice blast fungus\u003cem\u003e\u0026nbsp;Magnaporthe oryzae\u003c/em\u003e reveals distinct families of temporally co-regulated and structurally conserved effectors. \u003cem\u003eThe Plant cell,\u0026nbsp;\u003c/em\u003e35(5), 1360\u0026ndash;1385.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eYang H., Huang Y., Yi C., Shi J., Tan C., Ren W., Wang W. (2023). Development and Application of Specific Molecular Markers for Six Homologous Rice Blast Resistance Genes in \u003cem\u003ePi9\u0026nbsp;\u003c/em\u003eLocus of Rice. Scientia Agricultura Sinica, 56(21):4219-4233.\u003c/li\u003e\n \u003cli\u003eYang, H., Huang Y., Yi C., Tan C., Ren W., Huang F., Shi J., Li X., Wang W. (2024a). Development and application of specific molecular markers for five allelic rice blast resistance genes in\u003cem\u003e\u0026nbsp;Pik\u0026nbsp;\u003c/em\u003egene site in rice. ACTA PHYTOPATHOLOGICA SINICA, 54(3): 571-581.\u003c/li\u003e\n \u003cli\u003eYang H., Huang Y., Wang J., Yi C., Shi J., Tan C., Ren W., Wang W. (2024b). Development and Application of Specific Molecular Markers for Eight Rice Blast Resistance Genes in Rice. Chin J Rice Sci, 38(5): 525-534.\u003c/li\u003e\n \u003cli\u003eZhang, S., Zhong, X. L., Qiao, G. Y., Shen, L., Zhou, T. Y. and Peng, Y. L. (2017). Regional differentiation of Magnaporthe oryzae virulence in Sichuan, Chongqing, and Guizhou. Southwest China Journal of Agricultural Sciences 30(2): 359-365.\u003c/li\u003e\n \u003cli\u003eZhang, Y. L., Gao, Q., Zhao, Y. H., Liu, R., Fu, Z. J., Li, X., Sun, Y. J. and Jin, X. H. (2022). Evaluation of rice blast resistance and analysis of resistance gene structure in rice germplasm from Heilongjiang Province. Scientia Agricultura Sinica 55(4): 625-645.\u003c/li\u003e\n \u003cli\u003eZhou, B., Qu, S., Liu, G., Dolan, M., Sakai, H., Lu, G., Bellizzi, M., \u0026amp; Wang, G. L. (2006). The eight amino-acid differences within three leucine-rich repeats between \u003cem\u003ePi2\u003c/em\u003e and \u003cem\u003ePiz-t\u003c/em\u003e resistance proteins determine the resistance specificity to \u003cem\u003eMagnaporthe grisea\u003c/em\u003e. \u003cem\u003eMolecular plant-microbe interactions: MPMI,\u0026nbsp;\u003c/em\u003e19(11), 1216\u0026ndash;1228.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eZeng, S., Li, C., Du, C., Sun, L., Jing, D., Lin, T., Yu, B., Qian, H., Yao, W., Zhou, Y., \u0026amp;\u0026nbsp;Gong, H. (2018). Development of Specific Markers for \u003cem\u003ePigm\u003c/em\u003e in Marker- Assisted Breeding of Panicle Blast Resistant Japonica Rice. \u003cem\u003eChinese rice science\u003c/em\u003e, 05, 453-461.\u003c/li\u003e\n \u003cli\u003eZhou, K., Zhang, C., Xia, J.,Wang, Y., Yun, P.,Ma, T., Wu, D., \u0026amp; Li, Z. (2022).Associated Analysis of Rice Blast Genotypes and Seedling Blast Resistance of Japonica Rice Resources in the Middle and Lower Reaches of the Yangtze River.\u0026nbsp;\u003cem\u003eJournal of Nuclear Agricultural Sciences,\u0026nbsp;\u003c/em\u003e10,1920-1928.\u003c/li\u003e\n \u003cli\u003eZhou, Y., Lei, F., Wang, Q., He, W., Yuan, B., \u0026amp; Yuan, W. (2020). Identification of Novel Alleles of the Rice Blast-Resistance Gene\u003cem\u003e\u0026nbsp;Pi9\u003c/em\u003e through Sequence-Based Allele Mining. \u003cem\u003eRice (New York, N.Y.),\u0026nbsp;\u003c/em\u003e13(1), 80.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eZeng, X., Yang, X., Zhao, Z., Lin, F., Wang, L., \u0026amp; Pan, Q. (2011). Characterization and fine mapping of the rice blast resistance gene \u003cem\u003ePia\u003c/em\u003e. \u003cem\u003eScience China. Life sciences,\u0026nbsp;\u003c/em\u003e54(4), 372\u0026ndash;378.\u0026nbsp;\u003c/li\u003e\n \u003cli\u003eZhang, Z., Jia, Y., Wang, Y., \u0026amp; Sun, G. (2020). A Rapid Survey of Avirulence Genes in Field Isolates of \u003cem\u003eMagnaporthe oryzae\u003c/em\u003e.\u003cem\u003e\u0026nbsp;Plant disease,\u0026nbsp;\u003c/em\u003e104(3), 717\u0026ndash;723\u003c/li\u003e\n\u003c/ol\u003e"},{"header":"Tables","content":"\u003cp\u003eTable 1. Crosses leading to 136 resistance lines selected from WZ and ES disease nurseries\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\" width=\"590\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003eNO.\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eParent lines\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003eNumber of lines\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003eNO.\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003eParent lines\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003eNumber of lines\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003eNO.\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eParent lines\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003eNumber of lines\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eQhui 28/Luhui 80\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e19\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003e18-TL9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e37\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eWanhui 16/Huayousizhan\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eHuazhan/Chenghui 727\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003eQhui 28/Wanhui 16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e38\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eWanhui 481/CT18597\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eYuehesimiao/Yunhui 11\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e21\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003eQhui 28/Zhonghui 2827\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e39\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eWanhui 56/CT18272\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eYuehesimiao/Wanhui 16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e22\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003eHuazhan/Wanhui 99\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eWanhui 79\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eHuazhan/Luhui 80F4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e23\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003eWanhui 99/Guiyu 9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e41\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eWanhui 88/Luhui 37\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e6\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eQhui 28/Yunhui 11\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e24\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003eWanhui 99/Taiguoxiaoxiangzhan\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e42\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eWanhui 88/Luhui 615\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eHuazhan/Wanhui 16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003eYuehesimiao/Huanglizhan\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e43\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eWanhui 90\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eYuehesimiao/Chenghui 727\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e26\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003eYuehesimiao/Wushansimiao\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e44\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eWanhui 96/Huangguangyouzhan\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eQhui 28/Kang 4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e27\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003eYuenongsimiao/Guangyou 8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e45\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eWanhui 96/Zhonghui 2827\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eQhui 28/Yuenongsimiao\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e28\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003eYuenongsimiao/Huazhan\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e46\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eWanhui 99/Guangyou 8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e11\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eHuazhan/Yunhui 11\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e29\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003e2015-B 36/Mianhui 523\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e47\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eWanhui 99/Huangguangyouzhan\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e12\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eHuazhans/Wanhui 99\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003eQhui 28/R24\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e48\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eWanhui 99/Wushansimiao\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e13\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eWanhui 99/Yuenongsimiao\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e31\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003eQhui 28/Chenghui 727\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e49\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eYuehesimiao/Huazhan\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e14\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eWushansimiao/Zhonghui 2827\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003eQhui 28/Guangyou 8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eYuehesimiao/Yuenongsimiao\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eYuehesimiao/Guangyou 8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e33\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003eQhui 28/Guiyu 9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e51\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eYuehesimiao/Zhonghui 2827\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eYuenongsimiao/Guiyu 9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e34\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003eHuazhan/Exiang 1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e52\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eYuenongsimiao/Huangguangyouzhan\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e17\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eYuenongsimiao/Wushansimiao\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e35\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003eHuazhans/Wanhui 96\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e53\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eYuenongsimiao/Mabayouzhan\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e18\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 116px;\"\u003e\n \u003cp\u003eYuenongsimiao/Zhonghui 2827\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e36\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 125px;\"\u003e\n \u003cp\u003eNaire 1317/Chenghui 727\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 29px;\"\u003e\n \u003cp\u003e54\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 140px;\"\u003e\n \u003cp\u003eYuenongsimiao/Taiguoxiaoxiangzhan\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 41px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eTable 2. Twenty resistant lines selected from Meitan rice blase disease nursery\u003c/p\u003e\n\u003cdiv align=\"\"\u003e\n \u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\" width=\"548\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"2\" style=\"width: 32px;\"\u003e\n \u003cp\u003eNO.\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 162px;\"\u003e\n \u003cp\u003eParent\u0026nbsp;lines\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 55px;\"\u003e\n \u003cp\u003eGeneration\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" style=\"width: 102px;\"\u003e\n \u003cp\u003eMeitan\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 198px;\"\u003e\n \u003cp\u003e\u003cem\u003eR\u0026nbsp;\u003c/em\u003egene\u0026nbsp;composition\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 49px;\"\u003e\n \u003cp\u003eLeaf\u0026nbsp;blast\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 53px;\"\u003e\n \u003cp\u003eNeck\u0026nbsp;blast\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 32px;\"\u003e\n \u003cp\u003eE94\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 162px;\"\u003e\n \u003cp\u003eQhui\u0026nbsp;28/Chenghui\u0026nbsp;727\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 55px;\"\u003e\n \u003cp\u003eF4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 49px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 53px;\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 198px;\"\u003e\n \u003cp\u003e\u003cem\u003ePi2+Pi5+Pi54+Pia\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 32px;\"\u003e\n \u003cp\u003eE97\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 162px;\"\u003e\n \u003cp\u003eYuenongsimiao/Yunhui\u0026nbsp;11\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 55px;\"\u003e\n \u003cp\u003eF4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 49px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 53px;\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 198px;\"\u003e\n \u003cp\u003e\u003cem\u003ePi5+Pi54+Pia\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 32px;\"\u003e\n \u003cp\u003eK1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 162px;\"\u003e\n \u003cp\u003eHuazhan/Luhui80\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 55px;\"\u003e\n \u003cp\u003eF4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 49px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 53px;\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 198px;\"\u003e\n \u003cp\u003e\u003cem\u003ePi2+Pi5+Pi54+Pib+Pid3+Pita+Pia\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 32px;\"\u003e\n \u003cp\u003eK125\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 162px;\"\u003e\n \u003cp\u003eWanhui\u0026nbsp;96/zhonghui\u0026nbsp;2827\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 55px;\"\u003e\n \u003cp\u003eF4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 49px;\"\u003e\n \u003cp\u003e1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 53px;\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 198px;\"\u003e\n \u003cp\u003e\u003cem\u003ePi5+Pi54+Pib+Pita+Pia\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 32px;\"\u003e\n \u003cp\u003eK127\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 162px;\"\u003e\n \u003cp\u003eWanhui\u0026nbsp;99/Taiguoxiaoxiangzhan\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 55px;\"\u003e\n \u003cp\u003eF4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 49px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 53px;\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 198px;\"\u003e\n \u003cp\u003e\u003cem\u003ePi2+Pi5+Pid3+Pita\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 32px;\"\u003e\n \u003cp\u003eK135\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 162px;\"\u003e\n \u003cp\u003eWanhui\u0026nbsp;99/Wushansimiao\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 55px;\"\u003e\n \u003cp\u003eF4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 49px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 53px;\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n 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style=\"width: 55px;\"\u003e\n \u003cp\u003eF4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 49px;\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 53px;\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 198px;\"\u003e\n \u003cp\u003e\u003cem\u003ePi54+Pib+Pia\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003c/table\u003e\n\u003c/div\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":true,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"tropical-plant-pathology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"tppa","sideBox":"Learn more about [Tropical Plant Pathology](https://www.springer.com/journal/40858)","snPcode":"40858","submissionUrl":"https://www.editorialmanager.com/tppa","title":"Tropical Plant Pathology","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"rice, M. oryzae, resistance genes, rice resistance breeding, disease nursery","lastPublishedDoi":"10.21203/rs.3.rs-5734717/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-5734717/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eRice blast, caused by \u003cem\u003eMagnaporthe oryzae\u003c/em\u003e, threatens global rice productivity through severe yield losses. Developing cultivars with durable resistance via strategic pyramiding of resistance (R) genes is essential for sustainable disease management. This study aimed to develop elite rice germplasm harboring effective R-gene compositions and evaluate their resistance stability across geographically distinct blast hotspots in Southwest China. From disease nursery screenings in Wanzhou and Enshi, 136 resistant lines were selected from crosses of elite parents. Subsequent multilocation evaluations identified 20 lines demonstrating stable resistance in Meitan, a region with distinct \u003cem\u003eM. oryzae\u003c/em\u003e pathogenic populations, highlighting significant geographical variation in pathogen virulence. These results emphasize the necessity of multilocation testing for breeding durably resistant varieties. Allele-specific marker analysis of 14 major R genes revealed genetic compositions across resistant lines. Over 50% of lines carried \u003cem\u003ePi5\u003c/em\u003e, \u003cem\u003ePi54\u003c/em\u003e, \u003cem\u003ePita\u003c/em\u003e, or \u003cem\u003ePia\u003c/em\u003e, with most genotypes pyramiding 3-5 genes, reflecting the widespread adoption of R-gene pyramiding in rice breeding programs, with elite parental lines harboring diverse resistance gene combinations. Notably, lines demonstrating blast resistance in Meitan shared identical R-gene profiles with susceptible materials, suggesting the involvement of uncharacterized major-effect genes in conferring resistance. This finding underscores the prioritization of functional major-effect genes over quantitative gene stacking in resistance breeding. Our study establishes a regionally optimized framework for blast resistance evaluation and provides validated genetic resources for developing adapted rice cultivars in Southwest China.\u003c/p\u003e","manuscriptTitle":"Challenges and Insights into Pyramided R Genes for Durable Rice Blast Resistance in Southwest China","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-04-30 05:52:43","doi":"10.21203/rs.3.rs-5734717/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"reviewerAgreed","content":"","date":"2025-04-29T16:39:44+00:00","index":0,"fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-04-29T05:01:41+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-04-15T13:41:52+00:00","index":"","fulltext":""},{"type":"submitted","content":"Tropical Plant Pathology","date":"2025-04-08T21:10:18+00:00","index":"","fulltext":""},{"type":"decision","content":"Minor revisions","date":"2025-03-13T22:51:27+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"tropical-plant-pathology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"tppa","sideBox":"Learn more about [Tropical Plant Pathology](https://www.springer.com/journal/40858)","snPcode":"40858","submissionUrl":"https://www.editorialmanager.com/tppa","title":"Tropical Plant Pathology","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"7f6187c7-1352-4425-92a2-2a4b75f783fa","owner":[],"postedDate":"April 30th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[],"tags":[],"updatedAt":"2026-01-29T14:22:20+00:00","versionOfRecord":[],"versionCreatedAt":"2025-04-30 05:52:43","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-5734717","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-5734717","identity":"rs-5734717","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}