A General Protocol for Gene Expression Analysis in Chemically Mutant Coffee Plants in Response to Rust (Hemileia vastatrix Berk & Broome) Infection.

Coffea spp. is the source of one of the most widely consumed beverages in the world. However, the cultivation of this crop is threatened by Hemileia vastatrix Berk & Broome, a fungal disease, which reduces the productivity and can cause significant economic losses. In this protocol, coffee leaf segment derived from a chemical mutagenesis process are inoculated with uredospores of the pathogen. Subsequently, the gene expression changes are analyzed over the time (0, 5, 24, 48, and 120 h) using quantitative real-time polymerase chain reaction (RT-qPCR). The procedures and example data are presented for expression analysis in the CaWRKY1 gene. This procedure can be applied for quantitative analysis of other genes of interest to coffee breeders and scientists for elucidating the molecular mechanisms involved in the interaction between the plant and pathogen, potentially leading to the development of more efficient approaches for managing this disease.

Identification of Hsp20 gene family in Malus domestica and functional characterization of Hsp20 class I gene MdHsp18.2b.

Heat shock protein 20 (Hsp20) is a small molecule heat shock protein that plays an important role in plant growth, development, and stress resistance. Little is known about the function of Hsp20 family genes in apple (Malus domestica). Here, we performed a genome-wide analysis of the apple Hsp20 gene family, and a total of 49 Hsp20s genes were identified from the apple genome. Phylogenetic analysis revealed that the 49 genes were divided into 11 subfamilies, and MdHsp18.2b, a member located in the CI branch, was selected as a representative member for functional characterization. Treatment with NaCl and Botryosphaeria dothidea (B. dothidea), the causal agent of apple ring rot disease, significantly induced MdHsp18.2b transcription level. Further analysis revealed that overexpressing MdHsp18.2b reduced the resistance to salt stress but enhanced the resistance to B. dothidea infection in apple calli. Moreover, MdHsp18.2b positively regulated anthocyanin accumulation in apple calli. Physiology assays revealed that MdHsp18.2b promoted H2O2 production, even in the absence of stress factors, which might contribute to its functions in response to NaCl and B. dothidea infection. Hsps usually function as homo- or heterooligomers, and we found that MdHsp18.2b could form a heterodimer with MdHsp17.9a and MdHsp17.5, two members from the same branch with MdHsp18.2b in the phylogenetic tree. Therefore, we identified 49 Hsp20s genes from the apple genome and found that MdHsp18.2b was involved in regulating plant resistance to salt stress and B. dothidea infection, as well as in regulating anthocyanin accumulation in apple calli.

Characterization of the olive endophytic community in genotypes displaying a contrasting response to Xylella fastidiosa.

BACKGROUND: Endophytes mediate the interactions between plants and other microorganisms, and the functional aspects of interactions between endophytes and their host that support plant-growth promotion and tolerance to stresses signify the ecological relevance of the endosphere microbiome. In this work, we studied the bacterial and fungal endophytic communities of olive tree (Olea europaea L.) asymptomatic or low symptomatic genotypes sampled in groves heavily compromised by Xylella fastidiosa subsp. pauca, aiming to characterize microbiota in genotypes displaying differential response to the pathogen. RESULTS: The relationships between bacterial and fungal genera were analyzed both separately and together, in order to investigate the intricate correlations between the identified Operational Taxonomic Units (OTUs). Results suggested a dominant role of the fungal endophytic community compared to the bacterial one, and highlighted specific microbial taxa only associated with asymptomatic or low symptomatic genotypes. In addition, they indicated the occurrence of well-adapted genetic resources surviving after years of pathogen pressure in association with microorganisms such as Burkholderia, Quambalaria, Phaffia and Rhodotorula. CONCLUSIONS: This is the first study to overview endophytic communities associated with several putatively resistant olive genotypes in areas under high X. fastidiosa inoculum pressure. Identifying these negatively correlated genera can offer valuable insights into the potential antagonistic microbial resources and their possible development as biocontrol agents.

Comparative transcriptome revealed the molecular responses of Aconitum carmichaelii Debx. to downy mildew at different stages of disease development.

BACKGROUND: Aconitum carmichaelii Debx. has been widely used as a traditional medicinal herb for a long history in China. It is highly susceptible to various dangerous diseases during the cultivation process. Downy mildew is the most serious leaf disease of A. carmichaelii, affecting plant growth and ultimately leading to a reduction in yield. To better understand the response mechanism of A. carmichaelii leaves subjected to downy mildew, the contents of endogenous plant hormones as well as transcriptome sequencing were analyzed at five different infected stages. RESULTS: The content of 3-indoleacetic acid, abscisic acid, salicylic acid and jasmonic acid has changed significantly in A. carmichaelii leaves with the development of downy mildew, and related synthetic genes such as 9-cis-epoxycarotenoid dioxygenase and phenylalanine ammonia lyase were also significant for disease responses. The transcriptomic data indicated that the differentially expressed genes were primarily associated with plant hormone signal transduction, plant-pathogen interaction, the mitogen-activated protein kinase signaling pathway in plants, and phenylpropanoid biosynthesis. Many of these genes also showed potential functions for resisting downy mildew. Through weighted gene co-expression network analysis, the hub genes and genes that have high connectivity to them were identified, which could participate in plant immune responses. CONCLUSIONS: In this study, we elucidated the response and potential genes of A. carmichaelii to downy mildew, and observed the changes of endogenous hormones content at different infection stages, so as to contribute to the further screening and identification of genes involved in the defense of downy mildew.

Risk assessment and molecular mechanism of Sclerotium rolfsii resistance to boscalid.

Boscalid, a widely used SDHI fungicide, has been employed in plant disease control for over two decades. However, there is currently no available information regarding its antifungal activity against Sclerotium rolfsii and the potential risk of resistance development in this pathogen. In this study, we evaluated the sensitivity of 100 S. rolfsii strains collected from five different regions in China during 2018-2019 to boscalid using mycelial growth inhibition method and assessed the risk of resistance development. The EC50 values for boscalid ranged from 0.2994 µg/mL to 1.0766 µg/mL against the tested strains, with an average EC50 value of 0.7052 ± 0.1473 µg/mL. Notably, a single peak sensitivity baseline was curved, indicating the absence of any detected resistant strains. Furtherly, 10 randomly selected strains of S. rolfsii were subjected to chemical taming to evaluate its resistance risk to boscalid, resulting in the successful generation of six stable and inheritable resistant mutants. These mutants exhibited significantly reduced mycelial growth, sclerotia production, and virulence compared to their respective parental strains. Cross-resistance tests revealed a correlation between boscalid and flutolanil, benzovindiflupyr, pydiflumetofen, fluindapyr, and thifluzamide; however, no cross-resistance was observed between boscalid and azoxystrobin. Thus, we conclude that the development risk of resistance in S. rolfsii to boscalid is low. Boscalid can be used as an alternative fungicide for controlling peanut sclerotium blight when combined with other fungicides that have different mechanisms of action. Finally, the target genes SDHB, SDHC, and SDHD in S. rolfsii were initially identified, cloned and sequenced to elucidate the mechanism of S. rolfsii resistance to boscalid. Two mutation genotypes were found in the mutants: SDHD-D111H and SDHD-H121Y. The mutants carrying SDHD-H121Y exhibited moderate resistance, while the mutants with SDHD-D111H showed low resistance. These findings contribute to our comprehensive understanding of molecular mechanisms underlying plant pathogens resistance to SDHI fungicides.

High-throughput diagnostic markers for foliar fungal disease resistance and high oleic acid content in groundnut.

BACKGROUND: Foliar diseases namely late leaf spot (LLS) and leaf rust (LR) reduce yield and deteriorate fodder quality in groundnut. Also the high oleic acid content has emerged as one of the most important traits for industries and consumers due to its increased shelf life and health benefits. RESULTS: Genetic mapping combined with pooled sequencing approaches identified candidate resistance genes (LLSR1 and LLSR2 for LLS and LR1 for LR) for both foliar fungal diseases. The LLS-A02 locus housed LLSR1 gene for LLS resistance, while, LLS-A03 housed LLSR2 and LR1 genes for LLS and LR resistance, respectively. A total of 49 KASPs markers were developed from the genomic regions of important disease resistance genes, such as NBS-LRR, purple acid phosphatase, pentatricopeptide repeat-containing protein, and serine/threonine-protein phosphatase. Among the 49 KASP markers, 41 KASPs were validated successfully on a validation panel of contrasting germplasm and breeding lines. Of the 41 validated KASPs, 39 KASPs were designed for rust and LLS resistance, while two KASPs were developed using fatty acid desaturase (FAD) genes to control high oleic acid levels. These validated KASP markers have been extensively used by various groundnut breeding programs across the world which led to development of thousands of advanced breeding lines and few of them also released for commercial cultivation. CONCLUSION: In this study, high-throughput and cost-effective KASP assays were developed, validated and successfully deployed to improve the resistance against foliar fungal diseases and oleic acid in groundnut. So far deployment of allele-specific and KASP diagnostic markers facilitated development and release of two rust- and LLS-resistant varieties and five high-oleic acid groundnut varieties in India. These validated markers provide opportunities for routine deployment in groundnut breeding programs.

Integrated genome-wide association and transcriptomic analysis to identify receptor kinase genes to stripe rust resistance in wheat germplasm from southwestern China.

Stripe rust of wheat, caused by Puccinia striiformis f. sp. tritici (Pst), is one of the most important diseases of wheat worldwide. Identification of new and elite Pst-resistance loci or genes has the potential to enhance overall resistance to this pathogen. Here, we conducted an integrated genome-wide association study (GWAS) and transcriptomic analysis to screen for loci associated with resistance to stripe rust in 335 accessions from Yunnan, including 311 landraces and 24 cultivars. Based on the environmental phenotype, we identified 113 protein kinases significantly associated with Pst resistance using mixed linear model (MLM) and generalized linear model (GLM) models. Transcriptomic analysis revealed that 52 of 113 protein kinases identified by GWAS were up and down regulated in response to Pst infection. Among these genes, a total of 15 receptor kinase genes were identified associated with Pst resistance. 11 candidate genes were newly discovered in Yunnan wheat germplasm. Our results revealed that resistance alleles to stripe rust were accumulated in Yunnan wheat germplasm, implying direct or indirect selection for improving stripe rust resistance in elite wheat breeding programs.

Characterization and identification of sources of rust resistance in Triticum militinae derivatives.

Triticum militinae (2n = 4X = 28, AtAtGG), belonging to the secondary gene pool of wheat, is known to carry resistance to many diseases. Though some disease resistance genes were reported from T. timopheevii, the closest wild relative of T. militinae, there are no reports from T. militinae. Twenty-one T. militinae Derivatives (TMD lines) developed at the Division of Genetics, IARI, New Delhi, were evaluated for leaf and stripe rusts at seedling and adult plant stages. Eight TMD lines (6-4, 6-5, 11-6, 12-4, 12-8, 12-12, 13-7 and 13-9) showed seedling resistance to both leaf and stripe rusts while six TMD lines (7-5, 7-6, 11-5, 13-1, 13-3 and 13-4) showed seedling resistance to leaf rust but adult plant resistance to stripe rust and three TMD lines (9-1, 9-2 and 15) showed seedling resistance to leaf rust but susceptibility to stripe rust. Three TMD lines (2-7, 2-8 and 6-1) with adult plant resistance to leaf and stripe rusts were found to carry the known gene Lr34/Yr18. Ten TMD lines (7-5, 7-6, 9-1, 9-2, 11-5, 11-6, 12-12, 12-4, 12-8, and 15) with seedling resistance to leaf rust, showing absence of known genes Lr18 and Lr50 with linked markers requires further confirmation by the test of allelism studies. As not a single stripe rust resistance gene has been reported from T. militinae or its close relative T. timpopheevii, all the 8 TMD lines (6-4, 6-5, 11-6,12-4, 12-8, 12-12, 13-7 and 13-9) identified of carrying seedling resistance to stripe rust and 3 TMD lines (13-1, 13-3 and 13-4) identified of carrying adult plant resistance to stripe rust are expected to carry unknown genes. Also, all the TMD lines were found to be cytologically stable and thus can be used in inheritance and mapping studies.

Analyzing genetic diversity in luffa and developing a Fusarium wilt-susceptible linked SNP marker through a single plant genome-wide association (sp-GWAS) study.

BACKGROUND: Luffa (Luffa spp.) is an economically important crop of the Cucurbitaceae family, commonly known as sponge gourd or vegetable gourd. It is an annual cross-pollinated crop primarily found in the subtropical and tropical regions of Asia, Australia, Africa, and the Americas. Luffa serves not only as a vegetable but also exhibits medicinal properties, including anti-inflammatory, antidiabetic, and anticancer effects. Moreover, the fiber derived from luffa finds extensive applications in various fields such as biotechnology and construction. However, luffa Fusarium wilt poses a severe threat to its production, and existing control methods have proven ineffective in terms of cost-effectiveness and environmental considerations. Therefore, there is an urgent need to develop luffa varieties resistant to Fusarium wilt. Single-plant GWAS (sp-GWAS) has been demonstrated as a promising tool for the rapid and efficient identification of quantitative trait loci (QTLs) associated with target traits, as well as closely linked molecular markers. RESULTS: In this study, a collection of 97 individuals from 73 luffa accessions including two major luffa species underwent single-plant GWAS to investigate luffa Fusarium wilt resistance. Utilizing the double digest restriction site associated DNA (ddRAD) method, a total of 8,919 high-quality single nucleotide polymorphisms (SNPs) were identified. The analysis revealed the potential for Fusarium wilt resistance in accessions from both luffa species. There are 6 QTLs identified from 3 traits, including the area under the disease progress curve (AUDPC), a putative disease-resistant QTL, was identified on the second chromosome of luffa. Within the region of linkage disequilibrium, a candidate gene homologous to LOC111009722, which encodes peroxidase 40 and is associated with disease resistance in Cucumis melo, was identified. Furthermore, to validate the applicability of the marker associated with resistance from sp-GWAS, an additional set of 21 individual luffa plants were tested, exhibiting 93.75% accuracy in detecting susceptible of luffa species L. aegyptiaca Mill. CONCLUSION: In summary, these findings give a hint of genome position that may contribute to luffa wild resistance to Fusarium and can be utilized in the future luffa wilt resistant breeding programs aimed at developing wilt-resistant varieties by using the susceptible-linked SNP marker.

Antifungal Activity of Ribosome-Inactivating Proteins.

The control of crop diseases caused by fungi remains a major problem and there is a need to find effective fungicides that are environmentally friendly. Plants are an excellent source for this purpose because they have developed defense mechanisms to cope with fungal infections. Among the plant proteins that play a role in defense are ribosome-inactivating proteins (RIPs), enzymes obtained mainly from angiosperms that, in addition to inactivating ribosomes, have been studied as antiviral, fungicidal, and insecticidal proteins. In this review, we summarize and discuss the potential use of RIPs (and other proteins with similar activity) as antifungal agents, with special emphasis on RIP/fungus specificity, possible mechanisms of antifungal action, and the use of RIP genes to obtain fungus-resistant transgenic plants. It also highlights the fact that these proteins also have antiviral and insecticidal activity, which makes them very versatile tools for crop protection.

Soil prokaryotic and fungal biome structures associated with crop disease status across the Japan Archipelago.

Archaea, bacteria, and fungi in the soil are increasingly recognized as determinants of agricultural productivity and sustainability. A crucial step for exploring soil microbiomes with important ecosystem functions is to perform statistical analyses on the potential relationship between microbiome structure and functions based on comparisons of hundreds or thousands of environmental samples collected across broad geographic ranges. In this study, we integrated agricultural field metadata with microbial community analyses by targeting 2,903 bulk soil samples collected along a latitudinal gradient from cool-temperate to subtropical regions in Japan (26.1-42.8 °N). The data involving 632 archaeal, 26,868 bacterial, and 4,889 fungal operational taxonomic units detected across the fields of 19 crop plant species allowed us to conduct statistical analyses (permutational analyses of variance, generalized linear mixed models, randomization analyses, and network analyses) on the relationship among edaphic factors, microbiome compositions, and crop disease prevalence. We then examined whether the diverse microbes form species sets varying in potential ecological impacts on crop plants. A network analysis suggested that the observed prokaryotes and fungi were classified into several species sets (network modules), which differed substantially in association with crop disease prevalence. Within the network of microbe-to-microbe coexistence, ecologically diverse microbes, such as an ammonium-oxidizing archaeon, an antibiotics-producing bacterium, and a potentially mycoparasitic fungus, were inferred to play key roles in shifts between crop-disease-promotive and crop-disease-suppressive states of soil microbiomes. The bird's-eye view of soil microbiome structure will provide a basis for designing and managing agroecosystems with high disease-suppressive functions.IMPORTANCEUnderstanding how microbiome structure and functions are organized in soil ecosystems is one of the major challenges in both basic ecology and applied microbiology. Given the ongoing worldwide degradation of agroecosystems, building frameworks for exploring structural diversity and functional profiles of soil microbiomes is an essential task. Our study provides an overview of cropland microbiome states in light of potential crop-disease-suppressive functions. The large data set allowed us to explore highly functional species sets that may be stably managed in agroecosystems. Furthermore, an analysis of network architecture highlighted species that are potentially used to cause shifts from disease-prevalent states of agroecosystems to disease-suppressive states. By extending the approach of comparative analyses toward broader geographic ranges and diverse agricultural practices, agroecosystem with maximized biological functions will be further explored.

Biological characterization of emerging fungal pathogen Colletotrichum associated with mango (Mangifera indica L.) post-harvest anthracnose from Vietnam.

BACKGROUND: Post-harvest anthracnose (PHA) of mango is a devastating disease, which results in huge loss to mango producers and importers. Various species of PHA, diverse pathogenicity, and different resistance towards fungicides make it essential to evaluate the pathogen taxonomic status and biological characterization. METHODS AND RESULTS: Two strains DM-1 and DM-2 isolated from the fruit of DaQing mango from Vietnam were identified as Colletotrichum fructicola and C. asianum respectively, based on the morphological features, along with the phylogenetic tree of ITS and ApMat combined sequences. The growth status of different Colletotrichum strains under different conditions was analyzed to reveal the biological characteristics. The optimum growth temperature of DM-1 and DM-2 was 28 °C and mycelia grew rapidly in the dark. Both strains could grow in media with pH 4-11, while the optimum pH value was 6. Maltose and soluble starch were the most suitable carbon source for DM-1 and DM-2 respectively, and the peptone was the most suitable nitrogen source for both strains. The lethal temperatures were recorded as 55 °C 5 min for DM-1, and 50 °C 10 min for DM-2. CONCLUSIONS: To the best of our knowledge, it is the first study reporting the identification of the pathogens: C. fructicola and C. asianum responsible for postharvest fruit anthracnose of mango in Vietnam.

Association mechanism between Arabidopsis immune coreceptor BAK1 and Pseudomonas syringae effector HopF2.

Brassinosteroid activated kinase 1 (BAK1) is a cell-surface coreceptor which plays multiple roles in innate immunity of plants. HopF2 is an effector secreted by the bacterial pathogen Pseudomonas syringae pv. tomato DC3000 into Arabidopsis and suppresses host immune system through interaction with BAK1 as well as its downstream kinase MKK5. The association mechanism of HopF2 to BAK1 remains unclear, which prohibits our understanding and subsequent interfering of their interaction for pathogen management. Herein, we found the kinase domain of BAK1 (BAK1-KD) is sufficient for HopF2 association. With a combination of hydrogen/deuterium exchange mass spectrometry and mutational assays, we found a region of BAK1-KD N-lobe and a region of HopF2 head subdomain are critical for intermolecular interaction, which is also supported by unbiased protein-protein docking with ClusPro and kinase activity assay. Collectively, this research presents the interaction mechanism between Arabidopsis BAK1 and P. syringae HopF2, which could pave the way for bactericide development that blocking the functioning of HopF2 toward BAK1.

Preventing multi-resistance: New insights for managing fungal adaptation.

Sustainable crop protection is vital for food security, yet it is under threat due to the adaptation of a diverse and evolving pathogen population. Resistance can be managed by maximising the diversity of selection pressure through dose variation and the spatial and temporal combination of active ingredients. This study explores the interplay between operational drivers for maximising the sustainability of management strategies in relation to the resistance status of fungal populations. We applied an experimental evolution approach to three artificial populations of Zymoseptoria tritici, an economically significant wheat pathogen, each differing in initial resistance status. Our findings reveal that diversified selection pressure curtails the selection of resistance in naïve populations and those with low frequencies of single resistance. Increasing the number of modes of action most effectively delays resistance development, surpassing the increase in the number of fungicides, fungicide choice based on resistance risk, and temporal variation in fungicide exposure. However, this approach favours generalism in the evolved populations. The prior presence of multiple resistant isolates and their subsequent selection in populations override the effects of diversity in management strategies, thereby invalidating any universal ranking. Therefore, the initial resistance composition must be specifically considered in sustainable resistance management to address real-world field situations.

Methionine biosynthetic genes and methionine sulfoxide reductase A are required for Rhizoctonia solani AG1-IA to cause sheath blight disease in rice.

Rhizoctonia solani is a polyphagous necrotrophic fungal pathogen that causes sheath blight disease in rice. It deploys effector molecules as well as carbohydrate-active enzymes and enhances the production of reactive oxygen species for killing host tissues. Understanding R. solani ability to sustain growth under an oxidative-stress-enriched environment is important for developing disease control strategies. Here, we demonstrate that R. solani upregulates methionine biosynthetic genes, including Rs_MET13 during infection in rice, and double-stranded RNA-mediated silencing of these genes impairs the pathogen's ability to cause disease. Exogenous treatment with methionine restores the disease-causing ability of Rs_MET13-silenced R. solani and facilitates its growth on 10 mM H2O2-containing minimal-media. Notably, the Rs_MsrA gene that encodes methionine sulfoxide reductase A, an antioxidant enzyme involved in the repair of oxidative damage of methionine, is upregulated upon H2O2 treatment and also during infection in rice. Rs_MsrA-silenced R. solani is unable to cause disease, suggesting that it is important for the repair of oxidative damage in methionine during host colonization. We propose that spray-induced gene silencing of Rs_MsrA and designing of antagonistic molecules that block MsrA activity can be exploited as a drug target for effective control of sheath blight disease in rice.

Effects of ferulic acid combined with light irradiation on deoxynivalenol and its production in Fusarium graminearum.

This study aimed to investigate the effects of ferulic acid (FA), a natural phenolic phytochemical, in combination with light irradiation at three wavelengths (365, 385 and 405 nm) on the concentration and toxicity of deoxynivalenol (DON), a mycotoxin produced by Fusarium graminearum. Moreover, this study examined the influence of the combination treatment on DON production in the cultured fungus. FA activated by light at a peak wavelength of 365 nm exhibited the most effective decrease in DON concentration of the tested wavelengths; a residual DON ratio of 0.23 at 24 h exposure was observed, compared with the initial concentration. The reduction in DON using 365-nm light was dependent on the concentration of FA, with a good correlation (r2 = 0.979) between the rate constants of DON decrease and FA concentration, which was confirmed by a pseudo-first-order kinetics analysis of the photoreaction with different FA concentrations (50-400 mg/L) for 3 h. The viability of HepG2 cells increased by 56.7% following in vitro treatment with a mixture containing the photoproducts obtained after treatment with 20 mg/L DON and 200 mg/L FA under 365-nm irradiation for 6 h. These results suggested that the photoreaction of FA under 365-nm irradiation induces the detoxification of DON through degradation or modification of DON. The antifungal effects of the combination (FA and 365-nm light) on F. graminearum were investigated. Conidia treated with the combination did not show additive or synergistic inhibition of fungal biomass and DON production in 7-day cultivated fungal samples compared with samples after single treatment. However, successive treatment, composed of 90 min irradiation at 365 nm and then treatment with 200 mg/L FA for 90 min in the dark, suppressed fungal growth and DON yield to 70% and 25% of the untreated sample level, respectively. This photo-technology involving the two treatment methods of 365-nm irradiation and FA addition as a food-grade phenolic acid in combination or successively, can aid in developing alternative approaches to eliminate fungal contaminants in the fields of environmental water and agriculture. However, further research is required to explore the underlying mechanisms of DON decontamination and its biosynthesis in F. graminearum.

[Genetic dissection of rice resistance to bacterial blight].

Bacterial blight, a major disease in rice, poses a serious impact on rice production. In this study, a doubled haploid (DH) population derived from a cross between the introduced japonica cultivar 'Maybelle' and the indica landrace 'Baiyeqiu' was used to investigate the pathogenicity of four pathogen races causing bacterial blight. The results showed that the pathogenicity of all the pathogen races exhibited continuous, transgressive distribution in the DH population. Moreover, strong correlations existed between every two pathogen races, with the correlation coefficients ranging from 0.3 to 0.6. A total of 12 quantitative trait loci (QTLs) distributed on chromosomes 1, 2, 3, 5, 6, 7, 9, and 12 were detected for rice bacterial blight, explaining 4.95% to 16.05% of the phenotype. Among these QTLs, a major QTL located in the interval RM6024-RM163 on chromosome 5 was detected in three pathogen races. In addition, the pyramiding of the positive alleles can apparently improve the rice resistance to bacterial blight. This study is of great significance for broadening the genetic resources with resistance to bacterial blight in China.

[Silencing GmATG5 genes accelerated senescence and enhanced disease resistance in soybean].

Autophagy plays an essential role in recycling/re-utilizing nutrients and in adaptions to numerous stresses. However, the roles of autophagy in soybean have not been investigated extensively. In this study, a virus-induced gene silencing approach mediated by bean pod mottle virus (BPMV) was used to silence autophagy-related gene 5 (ATG5) genes in soybean (referred to as GmATG5). Our results showed that ATG8 proteins were massively accumulated in the dark-treated leaves of the GmATG5-silenced plants relative to the vector control plants (BPMV-0), indicating that autophagy pathway is impaired in the GmATG5-silenced plants. Consistent with the impaired autophagy, an accelerated senescence phenotype was observed on the leaves of the dark-treated GmATG5-silenced plants, which was not shown on the leaves of the dark-treated BPMV-0 plants. In addition, the accumulation levels of both reactive oxygen species (ROS) and salicylic acid (SA) were significantly induced in the GmATG5-silenced plants compared with that of the vector control plants (BPMV-0), indicating an activated immunity. Accordingly, the GmATG5-silenced plants exhibited significantly enhanced resistance against Pseudomonas syringae pv. glycinea (Psg) in comparison with the BPMV-0 plants. Nevertheless, the activated immunity observed in the GmATG5-silenced plant was independent of the activation of mitogen-activated protein kinase (MAPK).

[GmAGB1 plays a positive regulatory role in soybean defense responses].

Heterotrimeric GTP-binding protein (G-proteins) complex, which consists of Gα, Gß and Gγ subunits, plays critical roles in defense signaling. Arabidopsis genome contains only a single Gß-encoding gene, AGB1. Loss function of AGB1 in Arabidopsis results in enhanced susceptibility to a wide range of pathogens. However, the function of soybean AGB1 in immunity has not been previously interrogated. Bioinformatic analysis indicated that there are four GmAGB1 homologous genes in soybean genome, sharing homology of 86%-97%. To overcome the functional redundancy of these GmAGB1 homologs, virus-induced gene silencing (VIGS) mediated by the bean pod mottle virus (BPMV) was used to silence these four genes simultaneously. As expected, these four GmAGB1 homologous genes were indeed silenced by a single BPMV-VIGS vector carrying a conserved fragments among these four genes. A dwarfed phenotype was observed in GmAGB1s-silenced soybean plants, suggesting that GmAGB1s play a crucial role in growth and development. Disease resistance analysis indicated that silencing GmAGB1s significantly compromised the resistance of soybean plants against Xanthomonas campestris pv. glycinea (Xag). This reduced resistance was correlated with the decreased accumulation of pathogen-induced reactive oxygen species (ROS) and the reduced activation of GmMPK3 in response to flg22, a conserved N-terminal peptide of flagellin protein. These results indicate that GmAGB1 functions as a positive regulator in disease resistance and GmAGB1 is indispensable for the ROS production and GmMPK3 activation induced by pathogen infection. Yeast two hybrid assay showed that GmAGB1 interacted with GmAGG1, suggesting that an evolutionary conserved heterotrimeric G protein complex similarly functions in soybean.

[Characterization of sequences, expression profiling, and natural allelic variation analysis of the MYC gene family in sorghum (Sorghum bicolor)].

Sorghum aphid (Melanaphis sacchari) and head smut fungi (Sporisorium reilianum) infesting sorghum cause delayed growth and development, and reduce yield and quality. This study use bioinformatics and molecular biological approaches to profile the gene expression pattern during sorghum development and under pest infestation, and analyzed the natural allelic DNA variation of sorghum MYC gene family. The findings provide insights for potential application in breeding the stress resistant and high productivity sorghum varieties. The results indicated that there are 28 MYC genes identified in sorghum genome, distributed on 10 chromosomes. The bHLH_MYC_N and HLH domains are the conserved domains of the MYC gene in sorghum. Gene expression analysis showed that SbbHLH35.7g exhibited high expression levels in leaves, SbAbaIn showed strong expression in early grains, and SbMYC2.1g showed high expression levels in mature pollen. In anti-aphid strains at the 5-leaf stage, SbAbaIn, SbLHW.4g and SbLHW.2g were significantly induced in leaves, while SbbHLH35.7g displayed the highest expression level in panicle tissue, which was significantly induced by the infection of head smut. Promoter cis-element analysis identified methyl jasmonate (MJ), abscisic acid (ABA), salicylic acid (SA) and MYB-binding sites related to drought-stress inducibility. Furthermore, genomic resequencing data analysis revealed natural allelic DNA variations such as single nucleotide polymorphism (SNP) and insertion-deletion (INDEL) for the key SbMYCs. Protein interaction network analysis using STRING indicated that SbAbaIn interacts with TIFYdomain protein, and SbbHLH35.7g interacts with MDR and imporin. SbMYCs exhibited temporal and spatial expression patterns and played vital roles during the sorghum development. Infestation by sugarcane aphids and head smut fungi induced the expression of SbAbaIn and SbbHLH35.7g, respectively. SbAbaIn modulated the jasmonic acid (JA) pathway to regulate the expression of defensive genes, conferring resistance to insects. On the other hand, SbbHLH35.7g participated in detoxification reactions to defend against pathogens.