Genetic analysis and quantitative trait locus mapping of Bacterial Panicle Blight (BPB) disease resistance in rice (Oryza sativa L.)

Rice (Oryza sativa L.) is an important staple food for more than half of the world’s population, providing up to 23% of the total calories consumed by humans. Rice crop is prone to many diseases. Among the diseases, Bacterial Panicle Blight (BPB) disease has been reported to cause enormous yield loss. In 2018, over 12,000 hectares of paddy fields were infested with Bacterial Panicle Blight (BPB) disease in Malaysia. BPB disease caused by Burkholderia glumae is a devastating bacterial disease. There is no report of single genes identified for complete resistance to BPB. Although, previous studies have successfully identified a few BPB resistance QTLs, but could not be narrowed down to the candidate genes. Therefore, it is imperative to search for more QTLs resistance by screening the available rice germplasm worldwide. This study was undertaken to characterize and map out QTLs for resistance to BPB using the F2 mapping population. To achieve the objective, the study was portioned into five parts. The first part of the study is about evaluating 52 rice accessions for genetic diversity and phylogenetic relationships. Data was taken on vegetative, yield, and yield components, while phenotypic and genotypic coefficients, variance components, expected genetic advance, and heritability was calculated. Significant differences were observed among the accessions for all the characters studied. The evaluated genotypes were grouped into 13 major clusters based on the assessed traits with the aid of the UPGMA dendrogram. The highest numbers of accessions were grouped in cluster IV, while the least number was found in clusters II and IX, with one accession each. From the principal component analysis, the first four principal components explained 97.3 percent of the overall variance in the data set. The second part of the study seeks to characterize the causal pathogen of BPB. Burkholderia glumae was isolated from rice panicles, showing typical disease symptoms from Ladang 15, Faculty of Agriculture Universiti Putra Malaysia. Cultural, morphological, biochemical, and polymerase chain reaction (PCR) methods revealed that the isolated strain was Burkholderia glumae. The amplified isolate showed 100% identity when the sequence was aligned with other reported B. glumae strains. Following that, a pathogenicity test was conducted on three susceptible commercial rice varieties using the syringe inoculation method in the glasshouse to affirm the virulence of the isolated pathogen. All the inoculated rice accessions showed typical BPB symptoms similar to those obtained from the naturally infected rice panicles. Prior to the characterization of Burkholderia glumae, selected rice accessions were screened in search of novel resistant sources to BPB. Twenty eight rice accessions were selected based on yield and days to maturity as revealed in the cluster analysis conducted in the first study. The selected 28 accessions were inoculated with the B.glumae to assess external symptoms at 7, 14, and 21 days after inoculation. Based on the disease severity index (DSI), rice accession MRC 12 was the most susceptible with a DSI mean of 85.2% and AUDPC value of 114.8 unit2 in the third week after inoculation. About 14 accessions were classified as moderately susceptible (MS) with mean DSI value ranging from 37.04% (MRC24) to 53.09% (MRC11) and AUDPC range of 90.77 unit2 and 114.81 unit2 . Also, 13 accessions were found highly susceptible. Their mean DSI value ranged from 58.02% (MRC22) to 85.19% (MRC12) with the corresponding AUDPC of 103.70 unit2 and 114.81 unit2 , respectively. Only one accession (MRC 23) was found resistant (R) with a disease incidence score between 0-3 and a mean DSI value of 14.81% with the corresponding AUDPC of 38.89 unit2 in the third week after inoculation. Based on the cluster analysis and pathogenicity assay from the first and third studies, MRC 10 (high yielding susceptible) and MRC 23 (resistant) accessions were selected and used as parents to develop the mapping population for the QTL analysis. Hand emasculation and pollination were used to develop F1 from a cross between MRC 10 (female/recipient) and MRC 23 (male/donor). 201 rice SSR markers were screened to select polymorphic markers between MRC 10 and MRC 23. It is revealed that 39.98% polymorphism spread across the 12 chromosomes. For the confirmation of F1 hybridity, a total of 66 F1 plants were grown from the F1 seeds and tested for heterozygosity using the polymorphic background makers. Out of the 66 grown F1 plants, 44 were confirmed heterozygous true F1 plants. The true confirmed F1 were advanced to the F2 generation. In the fifth part of the study, a population of 62 F2 progenies was used to map QTL for BPB resistance. Progeny families were evaluated for disease reactions and genotyped with 46 SSR markers. Two significant (logarithm of odds ratio > 2.5) quantitative trait loci (QTLs) were identified for BPB resistance on chromosomes 9 and 10, with individual effects explaining 7.9 and 40.3% of the total phenotypic variance respectively. Furthermore, the bulk segregant analysis (BSA) method BSA was used to confirm the identified QTLs. BSA was done by bulking DNA of 10 BPB resistant and 10 BPB susceptible F2 progenies. Out of the 201 rice SSR markers genotyped, RM 228 co-segregate among the F2 progenies constituting the respective bulks. RM228 was mapped at the QTL region on chromosome 10. From this study, it is interesting to show that the region RM590 – RM228 on chromosome 10 harbors a large effect QTL for BPB resistance across several genetic backgrounds in rice. Altogether, findings from this study can be utilized to further investigate BPB resistance improvement in rice.

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