Exogenous Copper Application for the Elemental Defense of Rice Plants against Rice Leaffolder (Cnaphalocrocis medinalis).

Metals that accumulate in plants may confer protection against herbivorous insects, a phenomenon known as elemental defense. However, this strategy has not been widely explored in important crops such as rice (Oryza sativa L.), where it could help to reduce the use of chemical pesticides. Here, we investigated the potential of copper (Cu) and iron (Fe) micronutrient supplements for the protection of rice against a major insect pest, the rice leaffolder (Cnaphalocrocis medinalis). We found that intermediate levels of Cu (20 µM CuSO4) and high concentrations of Fe (742 µM Fe) did not inhibit the growth of C. medinalis larvae but did inhibit rice root growth and reduce grain yield at the reproductive stage. In contrast, high levels of Cu (80 µM CuSO4) inhibited C. medinalis larval growth and pupal development but also adversely affected rice growth at the vegetative stage. Interestingly, treatment with 10 µM CuSO4 had no adverse effects on rice growth or yield components at the reproductive stage. These data suggest that pest management based on the application of Cu may be possible, which would be achieved by a higher effective pesticide dose to prevent or minimize its phytotoxicity effects in plants.

Insight into the mechanism of indium toxicity in rice.

Indium is widely used in the technology industry and is an emerging form of environmental pollution. The presence of indium in soil and groundwater inhibits shoot and root growth in crops, thus reducing yields. However, the underlying mechanisms are unknown, making it difficult to design effective countermeasures. We explored the spatiotemporal effects of excess indium on the morphological, physiological and biochemical properties of rice (Oryza sativa L.). Indium accumulated mainly in the roots, severely restricting their growth and causing the acute perturbation of phosphorus, magnesium and iron homeostasis. Other effects included leaf necrosis and anatomical changes in the roots (thinned sclerenchyma and enlarged epidermal and exodermal layers). Whole-transcriptome sequencing revealed that rice immediately responded to indium stress by activating genes involved in heavy metal tolerance and phosphate starvation responses, including the expression of genes encoding phosphate-regulated transcription factors and transporters in the roots. Direct indium toxicity rather than phosphate deficiency was identified as the major factor affecting the growth of rice plants, resulting in the profound phenotypic changes we observed. The application of exogenous phosphate alleviated indium toxicity by reducing indium uptake. Our results suggest that indium immobilization could be used to prevent indium toxicity in the field.

Divalent nutrient cations: Friend and foe during zinc stress in rice.

Zn deficiency is the most common micronutrient deficit in rice but Zn is also a widespread industrial pollutant. Zn deficiency responses in rice are well documented, but comparative responses to Zn deficiency and excess have not been reported. Therefore, we compared the physiological, transcriptional and biochemical properties of rice subjected to Zn starvation or excess at early and later treatment stages. Both forms of Zn stress inhibited root and shoot growth. Gene ontology analysis of differentially expressed genes highlighted the overrepresentation of Zn transport and antioxidative defense for both Zn stresses, whereas diterpene biosynthesis was solely induced by excess Zn. Divalent cations (Fe, Cu, Ca, Mn and Mg) accumulated in Zn-deficient shoots but Mg and Mn were depleted in the Zn excess shoots, mirroring the gene expression of non-specific Zn transporters and chelators. Ascorbate peroxidase activity was induced after 14 days of Zn starvation, scavenging H2 O2 more effectively to prevent leaf chlorosis via the Fe-dependent Fenton reaction. Conversely, excess Zn triggered the expression of genes encoding Mg/Mn-binding proteins (OsCPS2/4 and OsKSL4/7) required for antimicrobial diterpenoid biosynthesis. Our study reveals the potential role of divalent cations in the shoot, driving the unique responses of rice to each form of Zn stress.

Transcriptomics identifies key defense mechanisms in rice resistant to both leaf-feeding and phloem feeding herbivores.

BACKGROUND: Outbreaks of insect pests in paddy fields cause heavy losses in global rice yield annually, a threat projected to be aggravated by ongoing climate warming. Although significant progress has been made in the screening and cloning of insect resistance genes in rice germplasm and their introgression into modern cultivars, improved rice resistance is only effective against either chewing or phloem-feeding insects. RESULTS: In this study, the results from standard and modified seedbox screening, settlement preference and honeydew excretion tests consistently showed that Qingliu, a previously known leaffolder-resistant rice variety, is also moderately resistant to brown planthopper (BPH). High-throughput RNA sequencing showed a higher number of differentially expressed genes (DEGs) at the infestation site, with 2720 DEGs in leaves vs 181 DEGs in sheaths for leaffolder herbivory and 450 DEGs in sheaths vs 212 DEGs in leaves for BPH infestation. The leaf-specific transcriptome revealed that Qingliu responds to leaffolder feeding by activating jasmonic acid biosynthesis genes and genes regulating the shikimate and phenylpropanoid pathways that are essential for the biosynthesis of salicylic acid, melatonin, flavonoids and lignin defensive compounds. The sheath-specific transcriptome revealed that Qingliu responds to BPH infestation by inducing salicylic acid-responsive genes and those controlling cellular signaling cascades. Taken together these genes could play a role in triggering defense mechanisms such as cell wall modifications and cuticular wax formation. CONCLUSIONS: This study highlighted the key defensive responses of a rarely observed rice variety Qingliu that has resistance to attacks by two different feeding guilds of herbivores. The leaffolders are leaf-feeder while the BPHs are phloem feeders, consequently Qingliu is considered to have dual resistance. Although the defense responses of Qingliu to both insect pest types appear largely dissimilar, the phenylpropanoid pathway (or more specifically phenylalanine ammonia-lyase genes) could be a convergent upstream pathway. However, this possibility requires further studies. This information is valuable for breeding programs aiming to generate broad spectrum insect resistance in rice cultivars.

SWATH-MS-based quantitative proteomics reveals a uniquely intricate defense response in Cnaphalocrocis medinalis-resistant rice.

Cnaphalocrocis medinalis is a major insect pest of rice in Asia. A few defensive enzymes were reported to show higher activities in a resistant rice line (Qingliu) than in a susceptible rice line (TN1) upon leaffolder infestation. However, the overall molecular regulation of the rice defense response against leaffolder herbivory is unknown. Here, differential proteomic analysis by SWATH-MS was performed to identify differentially expressed proteins between the two rice varieties, Qingliu and TN1, at four time points of leaffolder herbivory, 0, 6, 24, and 72 h. Gene Ontology (GO) enrichment of the differentially expressed proteins indicated overrepresentation of (1) photosynthesis, (2) amino acid and derivative metabolic process, and (3) secondary metabolic process. Phenylalanine ammonia lyase and chalcone synthase, which catalyze flavonoid biosynthesis, and lipoxygenase, which catalyzes jasmonic acid biosynthesis, exhibited higher expression in Qingliu than in TN1 even before insect herbivory. Momentary activation of the light reaction and Calvin cycle was detected in Qingliu at 6 h and 24 h of insect herbivory, respectively. At 72 h of insect herbivory, amino acid biosynthesis and glutathione-mediated antioxidation were activated in Qingliu. A defense response involving jasmonic acid signaling, carbon remobilization, and the production of flavonoids and glutathione could underlie the resistance of Qingliu to leaffolder.

The Role of a Glucosinolate-Derived Nitrile in Plant Immune Responses.

Glucosinolates are defense-related secondary metabolites found in Brassicaceae. When Brassicaceae come under attack, glucosinolates are hydrolyzed into different forms of glucosinolate hydrolysis products (GHPs). Among the GHPs, isothiocyanates are the most comprehensively characterized defensive compounds, whereas the functional study of nitriles, another group of GHP, is still limited. Therefore, this study investigates whether 3-butenenitrile (3BN), a nitrile, can trigger the signaling pathways involved in the regulation of defense responses in Arabidopsis thaliana against biotic stresses. Briefly, the methodology is divided into three stages, (i) evaluate the physiological and biochemical effects of exogenous 3BN treatment on Arabidopsis, (ii) determine the metabolites involved in 3BN-mediated defense responses in Arabidopsis, and (iii) assess whether a 3BN treatment can enhance the disease tolerance of Arabidopsis against necrotrophic pathogens. As a result, a 2.5 mM 3BN treatment caused lesion formation in Arabidopsis Columbia (Col-0) plants, a process found to be modulated by nitric oxide (NO). Metabolite profiling revealed an increased production of soluble sugars, Krebs cycle associated carboxylic acids and amino acids in Arabidopsis upon a 2.5 mM 3BN treatment, presumably via NO action. Primary metabolites such as sugars and amino acids are known to be crucial components in modulating plant defense responses. Furthermore, exposure to 2.0 mM 3BN treatment began to increase the production of salicylic acid (SA) and jasmonic acid (JA) phytohormones in Arabidopsis Col-0 plants in the absence of lesion formation. The production of SA and JA in nitrate reductase loss-of function mutant (nia1nia2) plants was also induced by the 3BN treatments, with a greater induction for JA. The SA concentration in nia1nia2 plants was lower than in Col-0 plants, confirming the previously reported role of NO in controlling SA production in Arabidopsis. A 2.0 mM 3BN treatment prior to pathogen assays effectively alleviated the leaf lesion symptom of Arabidopsis Col-0 plants caused by Pectobacterium carotovorum ssp. carotovorum and Botrytis cinerea and reduced the pathogen growth on leaves. The findings of this study demonstrate that 3BN can elicit defense response pathways in Arabidopsis, which potentially involves a coordinated crosstalk between NO and phytohormone signaling.

Transposable Elements Adaptive Role in Genome Plasticity, Pathogenicity and Evolution in Fungal Phytopathogens.

Transposable elements (TEs) are agents of genetic variability in phytopathogens as they are a source of adaptive evolution through genome diversification. Although many studies have uncovered information on TEs, the exact mechanism behind TE-induced changes within the genome remains poorly understood. Furthermore, convergent trends towards bigger genomes, emergence of novel genes and gain or loss of genes implicate a TE-regulated genome plasticity of fungal phytopathogens. TEs are able to alter gene expression by revamping the cis-regulatory elements or recruiting epigenetic control. Recent findings show that TEs recruit epigenetic control on the expression of effector genes as part of the coordinated infection strategy. In addition to genome plasticity and diversity, fungal pathogenicity is an area of economic concern. A survey of TE distribution suggests that their proximity to pathogenicity genes TEs may act as sites for emergence of novel pathogenicity factors via nucleotide changes and expansion or reduction of the gene family. Through a systematic survey of literature, we were able to conclude that the role of TEs in fungi is wide: ranging from genome plasticity, pathogenicity to adaptive behavior in evolution. This review also identifies the gaps in knowledge that requires further elucidation for a better understanding of TEs' contribution to genome architecture and versatility.

Draft Genome Sequence of Rhizoctonia solani Anastomosis Group 1 Subgroup 1A Strain 1802/KB Isolated from Rice.

Sheath blight, caused by Rhizoctonia solani anastomosis group 1 subgroup 1A (AG1-1A), is one of the most devastating rice diseases worldwide. Here, we report the draft genome sequence of R. solani AG1-1A strain 1802/KB isolated from a popular Malaysian rice variety. To the best of our knowledge, this is the second reported representative genome from AG1-1A.

Identification of functionally important microRNAs from rice inflorescence at heading stage of a qDTY4.1-QTL bearing Near Isogenic Line under drought conditions.

A cross between IR64 (high-yielding but drought-susceptible) and Aday Sel (drought-tolerant) rice cultivars yielded a stable line with enhanced grain yield under drought screening field trials at International Rice Research Institute. The major effect qDTY4.1 drought tolerance and yield QTL was detected in the IR77298-14-1-2-10 Backcrossed Inbred Line (BIL) and its IR87705-7-15-B Near Isogenic Line (NIL) with 93.9% genetic similarity to IR64. Although rice yield is extremely susceptible to water stress at reproductive stage, currently, there is only one report on the detection of drought-responsive microRNAs in inflorescence tissue of a Japonica rice line. In this study, more drought-responsive microRNAs were identified in the inflorescence tissues of IR64, IR77298-14-1-2-10 and IR87705-7-15-B via next-generation sequencing. Among the 32 families of inflorescence-specific non-conserved microRNAs that were identified, 22 families were up-regulated in IR87705-7-15-B. Overall 9 conserved and 34 non-conserved microRNA families were found as drought-responsive in rice inflorescence with 5 conserved and 30 non-conserved families induced in the IR87705-7-15-B. The observation of more drought-responsive non-conserved microRNAs may imply their prominence over conserved microRNAs in drought response mechanisms of rice inflorescence. Gene Ontology annotation analysis on the target genes of drought-responsive microRNAs identified in IR87705-7-15-B revealed over-representation of biological processes including development, signalling and response to stimulus. Particularly, four inflorescence-specific microRNAs viz. osa-miR5485, osa-miR5487, osa-miR5492 and osa-miR5517, and two non-inflorescence specific microRNAs viz. osa-miR169d and osa-miR169f.2 target genes that are involved in flower or embryonic development. Among them, osa-miR169d, osa-miR5492 and osa-miR5517 are related to flowering time control. It is also worth mentioning that osa-miR2118 and osa-miR2275, which are implicated in the biosynthesis of rice inflorescence-specific small interfering RNAs, were induced in IR87705-7-15-B but repressed in IR77298-14-1-2-10. Further, gene search within qDTY4.1 QTL region had identified multiple copies of NBS-LRR resistance genes (potential target of osa-miR2118), subtilisins and genes implicated in stomatal movement, ABA metabolism and cuticular wax biosynthesis.

Identification of four functionally important microRNA families with contrasting differential expression profiles between drought-tolerant and susceptible rice leaf at vegetative stage.

BACKGROUND: Developing drought-tolerant rice varieties with higher yield under water stressed conditions provides a viable solution to serious yield-reduction impact of drought. Understanding the molecular regulation of this polygenic trait is crucial for the eventual success of rice molecular breeding programmes. microRNAs have received tremendous attention recently due to its importance in negative regulation. In plants, apart from regulating developmental and physiological processes, microRNAs have also been associated with different biotic and abiotic stresses. Hence here we chose to analyze the differential expression profiles of microRNAs in three drought treated rice varieties: Vandana (drought-tolerant), Aday Sel (drought-tolerant) and IR64 (drought-susceptible) in greenhouse conditions via high-throughput sequencing. RESULTS: Twenty-six novel microRNA candidates involved in the regulation of diverse biological processes were identified based on the detection of miRNA*. Out of their 110 predicted targets, we confirmed 16 targets from 5 novel microRNA candidates. In the differential expression analysis, mature microRNA members from 49 families of known Oryza sativa microRNA were differentially expressed in leaf and stem respectively with over 28 families having at least a similar mature microRNA member commonly found to be differentially expressed between both tissues. Via the sequence profiling data of leaf samples, we identified osa-miR397a/b, osa-miR398b, osa-miR408-5p and osa-miR528-5p as being down-regulated in two drought-tolerant rice varieties and up-regulated in the drought-susceptible variety. These microRNAs are known to be involved in regulating starch metabolism, antioxidant defence, respiration and photosynthesis. A wide range of biological processes were found to be regulated by the target genes of all the identified differentially expressed microRNAs between both tissues, namely root development (5.3-5.7 %), cell transport (13.2-18.4 %), response to stress (10.5-11.3 %), lignin catabolic process (3.8-5.3 %), metabolic processes (32.1-39.5 %), oxidation-reduction process (9.4-13.2 %) and DNA replication (5.7-7.9 %). The predicted target genes of osa-miR166e-3p, osa-miR166h-5p*, osa-miR169r-3p* and osa-miR397a/b were found to be annotated to several of the aforementioned biological processes. CONCLUSIONS: The experimental design of this study, which features rice varieties with different drought tolerance and tissue specificity (leaf and stem), has provided new microRNA profiling information. The potentially regulatory importance of the microRNA genes mentioned above and their target genes would require further functional analyses.