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Pantoea ananatis is a phytopathogen infecting many economically important crops, including rice worldwide. Here, we report the complete genome of P. ananatis strain TZ39 identified as causative agent of a new bacterial blight of rice that emerged in China in 2020. The assembled genome consists of one circular chromosome of 4,483,976 bp and two plasmids of 135,135 and 276,579 bp. This complete genome of the first Chinese pathogenic P. ananatis strain will provide new insights into the traits of pathogenicity on genomic level from China and worldwide.

Sheath blight is a serious rice disease that causes great yield losses worldwide. Sheath blight resistance is controlled by genes with minor effect. It is an urgent need to identify the resistance genes in rice germplasm and utilize them in breeding. The purpose of this study is to identify the sheath blight-resistant alleles in different rice germplasm by association mapping and explore the potential of the resistance alleles in utilization. A total of 273 rice genotypes were inoculated and evaluated for sheath blight resistance in the field for 5 years, and 158 genotypes with relatively stable disease resistance phenotype were chosen for association mapping. A wide range of phenotypic variation for sheath blight resistance was observed in the 158-genotype population. Structure analyses divided the 158 genotypes into 4 subgroups, which was consistent with principal component analysis and cluster analysis. Association mapping performed using mixed linear model with 213 markers identified 14 resistance loci, locating on all chromosomes except chromosome 6, 10 and 12. Three novel resistance loci (D704, D855, and D905) which were different from the previously reported were detected. Thirty-eight alleles were detected at the 14 loci, among which favorable alleles at 11 loci showed consistent resistance across 5 years. It was found that the favorable resistance alleles can be pyramided to improve sheath blight resistance. These results enhance the understanding of how different resistance alleles at multiple loci in regulating sheath blight resistance, and provide markers for sheath blight resistance breeding in rice.

The seed setting rate (spikelet fertility) is an important determinant of rice yield. In the past few decades, genes that control rice seed set have been cloned, and many quantitative trait loci (QTL) have been identified. However, the epistasis influencing rice seed set remains largely unclear. In this study, a recombinant inbred line population, which consisted of 219 lines developed by crossing the Lemont and Yangdao4 rice cultivars, was grown in five environments to identify the QTL and epistatic loci related to seed set. A total of 26 minor-effect QTL were detected by multiple interval mapping, which explained less than 12.7% of the phenotypic variation individually. A pair of new epistatic loci were detected and confirmed by two-way analysis of variance; the homozygous Yangdao4 allele at the qSS6.1 locus interacted with the homozygous Lemont allele at the qSS8.1 locus and resulted in a low seed setting rate. A linear regression analysis and a multiple comparison test suggested that eight alleles at four QTL (qSS1.3, qSS6.3, qSS7.1, and qSS8.1) control seed set simultaneously. Marker-assisted selection using these four loci guaranteed a greater than 70% average seed setting rate in all five environments.

To identify stable quantitative trait loci (QTLs) responsible for sheath blight resistance, a recombinant inbred line mapping population consisting of 219 lines was developed by crossing Lemont and Yangdao 4. Average disease rating, average lesion length, maximum disease rating and maximum lesion length were assayed in six different environments. A total of 128 minor effect QTLs were detected by multiple interval mapping. These QTLs explained less than 11.2% of the phenotypic variations individually, and 106 QTLs were clustered in 20 QTL-rich regions/putative loci. Significant QTL × environment interactions were detected at three putative loci (qSBR11.1, qSBR11.2 and qSBR11.3), indicating that these three loci were not stable. The other 17 stable loci (qSBR1.1, qSBR1.2, qSBR2.1, qSBR2.3, qSBR3.1, qSBR3.2, qSBR3.5, qSBR3.6, qSBR5.1, qSBR7.1, qSBR8.1, qSBR9.1, qSBR9.2, qSBR9.3, qSBR12.1, qSBR12.2 and qSBR12.4) provided a foundation for marker-assisted selection in breeding. Analysis of allelic effect on the 20 putative loci identified 7 highly stable loci, including qSBR3.2, qSBR7.1, qSBR8.1, qSBR9.2, qSBR9.3, qSBR12.1 and qSBR12.2. Keywords: Rice, Sheath blight resistance, Quantitative trait locus, Recombinant inbred line

Tiller angle, controlled by quantitative trait loci (QTL), is crucial for achieving ideal plant architecture in rice, which affects its final yield values. It is important to understand the genetic mechanisms underlying tiller angle when breeding new plant-type varieties. To uncover the genetic regulation mechanisms in cultivated rice, we performed QTL analysis using a recombinant inbred line (RIL) mapping population consisting of 219 lines developed by crossing two rice cultivars, Lemont and Yangdao4. The angle between the side tiller (ST) and the horizontal ground was measured as a tiller angle-related trait in QTL analysis. Twenty-three QTLs responsible for ST-ground angle were detected using multiple interval mapping in four mapping environments. A major QTL, qTA9, was detected on chromosome 9, which explained 12.9, 39.8, 37.9, and 28.3% of the phenotypic variation in the four mapping environments, respectively. Another QTL, qTA8, with a relatively large effect, was detected on chromosome 8. The other 21 QTLs explained

Rice sheath blight (SB) disease is a global issue that causes great yield losses each year. To explore whether SB field resistance can be predicted, 273 rice genotypes were inoculated and evaluated for SB field resistance across nine environments from 2012 to 2019 to identify loci associated with SB resistance by association mapping. A total of 80 significant marker–trait associations were detected in nine environments, among which six loci (D130B, D230A, D304B, D309, D427A, and RM409) were repeatedly detected in at least two environments. A linear regression model for predicting SB lesion length was developed using genotypic data of these six loci and SB field resistance data of the 273 rice genotypes: y = 34.44 – 0.56x, where y is the predicted value of lesion length, and x is the total genotypic value of the six loci. A recombinant inbred line (RIL) population consisting of 219 lines that was grown in six environments (from 2013 to 2018) for evaluation of SB field resistance was used to check the prediction accuracy of the prediction model. The average absolute error between the predicted lesion length and real lesion length for the RIL population was 6.67 cm. The absolute errors between predicted and real lesion lengths were

In autumn 2020, leaf blight was observed on rice (Oryza sativa L., variety Zhongzao39, Yongyou9, Yongyou12, Yongyou15, Yongyou18, Yongyou1540, Zhongzheyou8, Jiafengyou2, Xiangliangyou900 and Jiyou351) in the fields of 17 towns in Zhejiang and Jiangxi Provinces, China. The disease incidence was 45%-60%. Initially, water-soaked, linear, light brown lesions emerged in the upper blades of the leaves, and then spread down to leaf margins, which ultimately caused leaf curling and blight during the booting-harvest stage (Fig. S1). The disease symptoms were assumed to be caused by Xanthomonas oryzae pv. oryzae (Xoo), the pathogen of rice bacterial blight. 63 isolates were obtained from the collected diseased leaves as previously described (Hou et al. 2020). All isolates showed circular, smooth-margined, yellow colonies when cultured on peptone sugar agar (PSA) medium for 24h at 28℃. The cells were all gram-negative and rod-shaped with three to six peritrichous flagella; positive for catalase, indole, glucose fermentation and citrate utilization, while negative for oxidase, alkaline, phenylalanine deaminase, urease, and nitrate reductase reactions. 16S rRNA gene sequence analysis from the 6 isolates (FY43, JH31, JH99, TZ20, TZ39 and TZ68) revealed that the amplified fragments shared 98% similarity with Pantoea ananatis type strain LMG 2665T (GenBank JFZU01) (Table S3). To further verify P. ananatis identity of these isolates, fragments of three housekeeping genes including gyrB, leuS and rpoB from the 6 isolates were amplified and sequenced, which showed highest homology to LMG 2665T with a sequence similarity of 95%-100% (Table S3). Primers (Brady et al. 2008) and GenBank accession numbers of gene sequences from the 6 isolates are listed in Table S1 and Table S2. Phylogenetic analysis of gyrB, leuS and rpoB concatenated sequences indicated that the 6 isolates were clustered in a stable branch with P. ananatis (Fig. S2). Based on the above morphological, physiological, biochemical and molecular data, the isolates are identified as P. ananatis. For pathogenicity tests, bacterial suspension at 108 CFU/mL was inoculated into flag leaves of rice (cv. Zhongzao39) at the late booting stage using clipping method. Water was used as a negative control. The clipped leaves displayed water-soaked lesions at 3 to 5 days after inoculation (DAI); then the lesion spread downward and turned light brown. At about 14 DAI, blight was shown with similar symptoms to those samples collected from the rice field of Zhejiang and Jiangxi provinces (Fig. S1). In contrast, the control plants remained healthy and symptomless. The same P. ananatis was re-isolated in the inoculated rice plants, fulfilling Koch's postulates. In the past decade, P. ananatis has been reported to cause grain discoloration in Hangzhou, China (Yan et al. 2010) and induce leaf blight as a companion of Enterobacter asburiae in Sichuan province, China (Xue et al. 2020). Nevertheless, to the best of our knowledge, this is the first report of P. ananatis as the causative agent of rice leaf blight in southeast China. This study raises the alarm that the emerging rice bacterial leaf blight in southeast China might be caused by a new pathogen P. ananatis, instead of Xoo as traditionally assumed. Further, the differences of occurrence, spread and control between two rice bacterial leaf blight diseases caused by P. ananatis and Xoo, respectively need to be determined in the future.

Molecular screening of major rice blast resistance genes was determined with molecular markers, which showed close-set linkage to 11 major rice blast resistance genes (Pi-d2, Pi-z, Piz-t, Pi-9, Pi-36, Pi-37, Pi5, Pi-b, Pik-p, Pik-h and Pi-ta2), in a collection of 32 accessions resistant to Magnaporthe oryzae. Out of the 32 accessions, the Pi-d2 and Pi-z appeared to be omnipresent and gave positive express. As the second dominant, Pi-b and Piz-t gene frequencies were 96.9% and 87.5%. And Pik-h and Pik-p gene frequencies were 43.8% and 28.1%, respectively. The molecular marker linkage to Pi-ta2 produced positive bands in eleven accessions, while the molecular marker linkage to Pi-36 and Pi-37 in only three and four accessions, respectively. The natural field evaluation analysis showed that 30 of the 32 accessions were resistant, one was moderately resistant and one was susceptible. Infection types were negatively correlated with the genotype scores of Pi-9, Pi5, Pi-b, Pi-ta2 and Pik-p, although the correlation coefficients were very little. These results are useful in identification and incorporation of functional resistance genes from these germplasms into elite cultivars through marker-assisted selection for improved blast resistance in China and worldwide.

Plant height (PH) is controlled by quantitative trait loci (QTLs) in rice. In the present study, a recombinant inbred line population developed by crossing two rice cultivars, Lemont and Yangdao4, was grown in eight environments for QTL analysis. Multiple interval mapping detected 53 PH-QTLs, 39 of which clustered at 12 chromosome regions/putative loci. An examination of the 12 putative loci identified 24 alleles that are simultaneously involved in controlling PH. Linear regression analyses suggested that these 24 alleles function additively across the 12 loci to control PH, and plants carrying more PH-increasing alleles at the 12 loci were taller than those carrying more PH-decreasing alleles. Multiple comparison tests indicated that the effect of a single allele at the 12 loci was small and that multiple alleles must be pyramided to attain a statistically significant effect. The closest markers to the 12 loci can be used directly in marker-assisted breeding to manipulate PH.

Rice sheath blight (SB) is the most serious rice disease in China. Resistance of rice to SB is a quantitative trait that is easily influenced by the environment; however, the extent of environmental influence on SB field resistance is still poorly understood. To identify rice genotype by environment interactions for SB resistance, 211 rice genotypes originating from 15 countries were planted and evaluated for SB field resistance in six different environments between 2012 and 2016 after inoculation with the SB pathogen isolate ZJ03. In addition, 65 rice genotypes were evaluated for SB field resistance in another four environments between 2013 and 2016 using ZJ03. Variations in SB field resistance were observed in different genotypes in different environments using objective and subjective rating methods. Two-way analysis of variance indicated that the interaction between the genotype and environment had a highly significant effect on SB field resistance. This analysis indicated that the environment had more of an influence than the genotype itself on SB field resistance, and the genotype by environment interaction was the greatest obstacle in obtaining a precise determination of SB field resistance in rice. The most resistant genotype, GD66, is a good candidate for genetic studies and breeding.

Rice bakanae disease, caused by the pathogen Fusarium fujikuroi, is becoming severely detrimental to rice production worldwide. To understand the interaction between rice and F. fujikuroi, a moderate resistant genotype, 93-11 (Oryza sativa indica), and a susceptible genotype, Nipponbare (Oryza sativa japonica), were used for transcriptome analysis. Several cDNA libraries were constructed using mRNA isolated from the leaves of non-stressed 93-11 and NIPPONBARE plants and from the leaves of both genotypes inoculated with F. fujikuroi. In total, 1152 and 1052 transcripts were differentially expressed between the controls and the treatments for 93-11 and NIPPONBARE, respectively. Comparative transcriptome analysis revealed different expression patterns for the two genotypes. Although some common defense-related enriched GO terms were shared in both genotypes, specific defense-related terms were enriched exclusively in 93-11. A detailed comparison of defense-related differentially expressed genes revealed that certain WRKYs, WAK and MAP3Ks were responsible for the bakanae disease resistance in 93-11. The OsWAK112d gene was up-regulated in the resistant genotype. The POEI gene response to abiotic stress was modulated in Nipponbare. Further analysis suggested that the defense-related genes WRKYs and MARKs on chromosome 1 that are modulated in 93-11 upon infection might play a crucial role in the rice–F. fujikuroi interaction. Further characterization of these resistance genes may provide candidate genes for the development of molecular markers for rice bakanae resistance breeding programs. Transcriptomic dissection of the rice–F. fujikuroi interaction provides valuable information for future studies on the molecular mechanisms of rice bakanae resistance.

Rice blast, bacterial blight (BB) and brown planthopper (BPH) are the three main pests of rice. This study investigated pyramiding genes resistant to blast, BB and BPH to develop restorer lines. Ten new lines with blast, BB and/or BPH resistance genes were developed using marker-assisted selection (MAS) technique and agronomic trait selection (ATS) method. Only HR13 with resistance genes to blast, BB and BPH was obtained. In addition to blast and BB resistance, four lines (HR39, HR41, HR42, HR43) demonstrated moderate resistance to BPH, but MAS for BPH resistance genes were not conducted in developing these four lines. These data suggested that there were unknown elite BPH resistance genes in the Zhongzu 14 donor parent. A more effective defense was demonstrated in the lines with Pi1 and Pi2 genes although the weather in 2012 was favorable to disease incidence. Blast resistance of the lines with a single resistance gene, Pita, was easily influenced by the weather. Overall, the information obtained through pyramiding multiple resistance genes on developing the restorer lines is helpful for rice resistance breeding.

Sheath blight (SB) disease, caused by Rhizoctonia solani Kuhn, is one of the most serious diseases causing rice (Oryza sativa L.) yield loss worldwide. A doubled haploid (DH) population was constructed from a cross between a japonica variety CJ06 and an indica variety TN1, and to analyze the quantitative trait loci (QTLs) for SB resistance under three different environments (environments 1–3). Two traits were recorded to evaluate the SB resistance, namely lesion height (LH) and disease rating (DR). Based on field evaluation of SB resistance and a genetic map constructed with 214 markers, a total of eight QTLs were identified for LH and eight QTLs for DR under three environments, respectively. The QTLs for LH were anchored on chromosomes 1, 3, 4, 5, 6, and 8, and explained 4.35–17.53% of the phenotypic variation. The SB resistance allele of qHNLH4 from TN1 decreased LH by 3.08 cm, and contributed to 17.53% of the variation at environment 1. The QTL for LH (qHZaLH8) detected on chromosome 8 in environment 2 explained 16.71% of the variation, and the resistance allele from CJ06 reduced LH by 4.4 cm. Eight QTLs for DR were identified on chromosomes 1, 5, 6, 8, 9, 11, and 12 under three conditions with the explained variation from 2.0 to 11.27%. The QTL for DR (qHZaDR8), which explained variation of 11.27%, was located in the same interval as that of qHZaLH8, both QTLs were detected in environment 2. A total of six pairs of digenic epistatic loci for DR were detected in three conditions, but no epistatic locus was observed for LH. In addition, we detected 12 QTLs for plant height (PH) in three environments. None of the PH-QTLs were co-located with the SB-QTLs. The results facilitate our understanding of the genetic basis for SB resistance in rice.

Rice sheath blight (ShB), caused by Rhizoctonia solani, leads to severe yield losses in many rice production areas worldwide. Resistance to rice ShB is a typical quantitative trait controlled by quantitative trait loci (QTLs). To identify the QTLs responsible for ShB resistance, phenotypes for ShB resistance have been surveyed in different individuals/lines in various mapping populations. The 0–9 rating system, based on the percentage of plant height (PH) above the water line with ShB symptoms, is extensively used in the evaluation of ShB in rice communities. By reviewing previous ShB-QTL-mapping studies, we found that the ShB disease score attained using the 0–9 system was inevitably affected by PH and resulted in the colocalization of QTLs for PH and ShB resistance, i.e., some of the PH-QTLs were mistaken as “ShB-QTLs” and had pleiotropic effects. These colocalizing “ShB-QTLs” are irrelevant to physiological ShB resistance and should not be targeted for utilization. We analyzed the ways in which PH affects ShB resistance through the 0–9 rating system and suggest solutions for improving the accuracy of QTL mapping for rice ShB resistance. We also developed the first physical map of the ShB-QTLs and PH-QTLs detected since 1995. This map will be useful in the marker-assisted selection of ShB-resistant QTLs. Under current circumstances, we believe that conventional breeding remains an effective approach for developing ShB-resistant varieties.

Five physiological and eleven yield traits of two pairs of sister lines generated from a high generation with similar genetic background (SLs) for purple pericarp were investigated to explore the reasons behind low-yield production of colored rice. Of the five physiological traits examined, except grain anthocyanin content, there were generally similar trends between the P (purple-pericarp) lines and the corresponding W (white-pericarp) lines over two seasons (in the year 2009 and 2010 separately). The results demonstrated that the chlorophyll content of flag leaves, the net photosynthetic rate of flag leaves, and the grain anthocyanin content could be easily influenced by the environment. The physiological functions of the traits for the P lines were more active than those of the corresponding W lines in the year 2010. The grain anthocyanin content of the P lines was much greater in the year 2010 than in the year 2009 during the growth period. The investigation of yield traits revealed that the P lines had reduced 1000-grain weight, yield per plot and grain/brown rice thickness compared to the W lines. A difference comparison of these traits and a source-sink and transportation relationship analysis for these SLs suggested that small sink size was a key reason behind yield reduction of purple pericarp rice.

Uniformity in the height of main stem and tillers is a key factor affecting ideal plant type, a key component in super high-yielding rice breeding. An understanding of the genetic basis of the panicle layer uniformity may thus contribute to breeding varieties with good plant type and high yield. In the present study, a doubled haploid (DH) population, derived from a cross between indica rice variety Zhai-Ye-Qing 8 (ZYQ8) and japonica rice variety Jing-Xi 17 (JX17) was used to analyze quantitative trait loci (QTL) for panicle layer uniformity related traits. Six, four and three QTL were detected for the highest panicle height (HPH), lowest panicle height (LPH) and panicle layer dis-uniformity (PLD), respectively. qHPH-1-1 and qPLD-1 were located at the same interval on chromosome 1. The JX17 allele(s) of these QTL increased HPH and PLD by 2.57 and 1.26 cm, respectively. Similarly, qPLD-7 and qHPH-7 were located at the same interval on chromosome 7, where the ZYQ8 allele(s) increased HPH and PLD by 3.74 and 1.96 cm, respectively. These four QTL were unfavourable for panicle layer uniformity improvement because a decrease of the PLD was accompanied by decrease of the plant height. qPLD-6 and qLPH-6-1 were located at the same interval on chromosome 6, however here the JX17 allele(s) increased LPH, but decreased PLD, suggesting that this QTL was favourable for improvement of panicle layer uniformity. The markers identified in this study are potential for marker assisted breeding for the improvement of the panicle layer uniformity and ideal plant type.

A total of 32 rice genotypes carrying different dwarf or semi-dwarf genes were inoculated with the fungus Fusarium moniliforme Sheldon or treated with 50 mg l-1 GA3 in order to select resources resistant to rice bakanae disease from the dwarf materials. The length of the elongated seedlings was measured, and the percentage of death of the seedlings after transplanting to field was also counted. A significant correlation was found between the length of the seedling treated with GA3 and disease injury by bakanae fungus. Rice materials carrying dwarf gene such as sd1 were not only sensitivity to GA3 but also susceptive to rice bakanae disease. Materials carrying dwarf gene d1 were insensitive to GA3 but susceptive to bakanae. On the other hand, all materials carrying d29, sd6 or sdq(t) genes showed resistance to bakanae. The present study indicated that dwarf and semi-dwarf rice materials might be useful resources for improvement of bakanae resistance in rice breeding programs.

Pre-harvest sprouting (PHS) in rice production is usually caused by high temperature and humidity or continuous rains. It frequently happens in F1 in hybrid rice seed production. The PHS or “Physiologically germinated” seeds are of lower quality, by which the hybrid rice seed production is badly affected every three years at a loss of 20% or even 50% yield in seed production over the vast Yangtse River Valley and Southwest China. It is estimated that PHS causes an average decrease of seed activity by 10%. A recombinant inbred line population including 304 lines, derived from a cross between Indica rice (Oryza sativa L.) cultivars Zhong-156 and Gumei-2, was used to study the PHS physiology. Based on the data of sprouting rate in panicles and sprouting rate in grains, two kinds of lines, namely easy-to-sprout lines and hard-to-sprout lines, were selected to investigate their physiological differences when PHS happened. The experiment was conducted in a special field with a microclimate of higher temperature and humidity. The results indicated that it was easier to produce PHS from the female parent GM-2 than the male parent ZH-156, besides, the GA1 content and amylase activity in GM-2 grains were higher than those in ZH-156. However, the abscisic acid (ABA) content in GM-2 grains was lower than that in ZH-156. Higher temperature and higher humidity facilitated the GA1 increment from milk ripe stage to yellow ripe stage. GM-2 and the easily-sprouting lines showed an even higher increase in GA1 than ZH-156 and the hardly-sprouting lines, which enhanced the amylase activity and induced pre-harvest sprouting. This may be the physiological basis for pre-harvest sprouting induced by higher temperature and higher humidity, and these special characteristics must be inherited from their parents.