Jasmonate-mediated polyamine oxidase 6 drives herbivore-induced polyamine catabolism in rice.

Polyamines (PAs) along with their conjugated forms, are important mediators of plant defense mechanisms against both biotic and abiotic stresses. Flavin-containing polyamine oxidases (PAOs) regulate PA levels through terminal oxidation. To date, the role of PAOs in plant-herbivore interaction remains poorly understood. We discovered that infestation by the brown planthopper (BPH) disrupts PA homeostasis within the leaf sheaths of rice plants, which co-occurs with the upregulation of OsPAO6, a tissue-specific inducible, apoplast-localized enzyme that regulates the terminal catabolism of spermidine (Spd) and spermine. Functional analysis using CRISPR-Cas9 genome-edited plants revealed that pao6 mutants accumulated significantly higher levels of Spd and phenylpropanoid-conjugated Spd in response to BPH infestation compared to wild-type controls. In addition, BPH feeding on pao6 mutants led to increased honeydew excretion and plant damage by female adults, consistent with in vitro experiments in which Spd enhanced BPH feeding. Furthermore, OsPAO6 transcription is regulated by jasmonate (JA) signaling, and it is dependent on MYC2, which directly binds to the G-box-like motif in the OsPAO6 promoter. Our findings reveal an important role of OsPAO6 in regulating polyamine catabolism in JA-induced responses triggered by herbivore attacks in rice.

Molecular and biological characterization of a bunyavirus infecting the brown planthopper (Nilaparvata lugens).

A negative-strand symbiotic RNA virus, tentatively named Nilaparvata lugens Bunyavirus (NLBV), was identified in the brown planthopper (BPH, Nilaparvata lugens). Phylogenetic analysis indicated that NLBV is a member of the genus Mobuvirus (family Phenuiviridae, order Bunyavirales). Analysis of virus-derived small interfering RNA suggested that antiviral immunity of BPH was successfully activated by NLBV infection. Tissue-specific investigation showed that NLBV was mainly accumulated in the fat-body of BPH adults. Moreover, NLBV was detected in eggs of viruliferous female BPHs, suggesting the possibility of vertical transmission of NLBV in BPH. Additionally, no significant differences were observed for the biological properties between NLBV-infected and NLBV-free BPHs. Finally, analysis of geographic distribution indicated that NLBV may be prevalent in Southeast Asia. This study provided a comprehensive characterization on the molecular and biological properties of a symbiotic virus in BPH, which will contribute to our understanding of the increasingly discovered RNA viruses in insects.

Brown planthopper infestation on rice reduces plant susceptibility to Meloidogyne graminicola by reducing root sugar allocation.

Plants are simultaneously attacked by different pests that rely on sugars uptake from plants. An understanding of the role of plant sugar allocation in these multipartite interactions is limited. Here, we characterized the expression patterns of sucrose transporter genes and evaluated the impact of targeted transporter gene mutants and brown planthopper (BPH) phloem-feeding and oviposition on root sugar allocation and BPH-reduced rice susceptibility to Meloidogyne graminicola. We found that the sugar transporter genes OsSUT1 and OsSUT2 are induced at BPH oviposition sites. OsSUT2 mutants showed a higher resistance to gravid BPH than to nymph BPH, and this was correlated with callose deposition, as reflected in a different effect on M. graminicola infection. BPH phloem-feeding caused inhibition of callose deposition that was counteracted by BPH oviposition. Meanwhile, this pivotal role of sugar allocation in BPH-reduced rice susceptibility to M. graminicola was validated on rice cultivar RHT harbouring BPH resistance genes Bph3 and Bph17. In conclusion, we demonstrated that rice susceptibility to M. graminicola is regulated by BPH phloem-feeding and oviposition on rice through differences in plant sugar allocation.

The Threat of the Fall Armyworm to Asian Rice Production Is Amplified by the Brown Planthopper.

The recent invasion of the fall armyworm (FAW) into Asia not only has had a major impact on maize yield but is feared to also pose a risk to rice production. We hypothesized that the brown planthopper (BPH) may aggravate this risk based on a recently discovered mutualism between the planthopper and the rice striped stem borer. Here we show that BPH may indeed facilitate a shift of FAW to rice. FAW females were found to strongly prefer to oviposit on BPH-infested rice plants, which emitted significantly elevated levels of five volatile compounds. A synthetic mixture of these compounds had a potent stimulatory effect on ovipositing females. Although FAW caterpillars exhibited relatively poor growth on both uninfested and BPH-infested rice, a considerable portion completed their development on young plants. Moreover, FAW were found to readily pupate and survive in exceedingly moist soils typical for rice cultivation, further highlighting FAW's potential to switch to rice. We conclude that BPH, by changing the bouquet of volatiles emitted by rice plants, may greatly facilitate this switch. These findings, together with a current increase of nonflooded upland rice in Asia, warrant careful monitoring and specific control measures against FAW to safeguard Asian rice production.

Silencing NlFAR7 destroyed the pore canals and related structures of the brown planthopper.

Fatty acyl-CoA reductase (FAR) is one of the key enzymes, which catalyses the conversion of fatty acyl-CoA to the corresponding alcohols. Among the FAR family members in the brown planthopper (Nilaparvata lugens), NlFAR7 plays a pivotal role in both the synthesis of cuticular hydrocarbons and the waterproofing of the cuticle. However, the precise mechanism by which NlFAR7 influences the formation of the cuticle structure in N. lugens remains unclear. Therefore, this paper aims to investigate the impact of NlFAR7 through RNA interference, transmission electron microscope, focused ion beam scanning electron microscopy (FIB-SEM) and lipidomics analysis. FIB-SEM is employed to reconstruct the three-dimensional (3D) architecture of the pore canals and related cuticle structures in N. lugens subjected to dsNlFAR7 and dsGFP treatments, enabling a comprehensive assessment of changes in the cuticle structures. The results reveal a reduction in the thickness of the cuticle and disruptions in the spiral structure of pore canals, accompanied by widened base and middle diameters. Furthermore, the lipidomics comparison analysis between dsNlFAR7- and dsGFP-treated N. lugens demonstrated that there were 25 metabolites involved in cuticular lipid layer synthesis, including 7 triacylglycerols (TGs), 5 phosphatidylcholines (PCs), 3 phosphatidylethanolamines (PEs) and 2 diacylglycerols (DGs) decreased, and 4 triacylglycerols (TGs) and 4 PEs increased. In conclusion, silencing NlFAR7 disrupts the synthesis of overall lipids and destroys the cuticular pore canals and related structures, thereby disrupting the secretion of cuticular lipids, thus affecting the cuticular waterproofing of N. lugens. These findings give significant attention with reference to further biochemical researches on the substrate specificity of FAR protein, and the molecular regulation mechanisms during N. lugens life cycle.

Comparative analysis of the diversity of symbionts in fat body of long- and short-winged brown planthoppers.

The microbial community structure plays an important role in the internal environment of brown planthopper (BPH), Nilaparvata lugens (Hemiptera: Delphacidae), which is an indispensable part to reflect the internal environment of BPH. Wing dimorphism is a strategy for balancing flight and reproduction of insects. Here, quantitative fluorescence PCR was used to analyse the number and changes of the symbionts in the fat body of long- and short-winged BPHs at different developmental stages. A metagenomic library was constructed based on the 16 S rRNA sequence and internal transcribed spacer sequence for high-throughput sequencing, to analyze the community structure and population number of the symbionts of long- and short-winged BPHs, and to make functional prediction. This study enriches the connotation of BPH symbionts, and laid a theoretical foundation for the subsequent study of BPH-symbionts interaction and the function of symbionts in the host.

Gut bacterium Burkholderia cepacia (BsNLG8) and immune gene Defensin A contribute to the resistance against Nicotine-induced stress in Nilaparvata lugens (Stål).

Nicotine, a naturally occurring alkaloid found in tobacco, is a potent neurotoxin extensively used to control Nilaparvata lugens (Stål), a destructive insect pest of rice crops. The insect gut harbors a wide array of resident microorganisms that profoundly influence several biological processes, including host immunity. Maintaining an optimal gut microbiota and immune homeostasis requires a complex network of reciprocal regulatory interactions. However, the underlying molecular mechanisms driving these symbiotic exchanges, particularly between specific gut microbe and immunity, remain largely unknown in insects. Our previous investigations identified and isolated a nicotine-degrading Burkholderia cepacia strain (BsNLG8) with antifungal properties. Building on those findings, we found that nicotine intake significantly increased the abundance of a symbiotic bacteria BsNLG8, induced a stronger bacteriostatic effect in hemolymph, and enhanced the nicotine tolerance of N. lugens. Additionally, nicotine-induced antimicrobial peptides (AMPs) exhibited significant antibacterial effects against Staphylococcus aureus. We adopted RNA-seq to explore the underlying immunological mechanisms in nicotine-stressed N. lugens. Bioinformatic analyses identified numerous differentially expressed immune genes, including recognition/immune activation (GRPs and Toll) and AMPs (i.e., Defensin, Lugensin, lysozyme). Temporal expression profiling (12, 24, and 48 hours) of immune genes revealed pattern recognition proteins and immune effectors as primary responders to nicotine-induced stress. Defensin A, a broad-spectrum immunomodulatory cationic peptide, exhibited significantly high expression. RNA interference-mediated silencing of Defensin A reduced the survival, enhanced nicotine sensitivity of N. lugens to nicotine, and decreased the abundance of BsNLG8. The reintroduction of BsNLG8 improved the expression of immune genes, aiding nicotine resistance of N. lugens. Our findings indicate a potential reciprocal immunomodulatory interaction between Defensin A and BsNLG8 under nicotine stress. Moreover, this study offers novel and valuable insights for future research into enhancing nicotine-based pest management programs and developing alternative biocontrol methods involving the implication of insect symbionts.

The down-regulation of salivary protein gene expression by etofenprox partially contributed to reducing the risk of increased fecundity in the brown planthopper.

Etofenprox is a pyrethroid insecticide that acts on the nervous system of insects. Due to its low toxicity to aquatic animals, it is permitted for use in controlling insect pests in rice fields. The brown planthopper (BPH), Nilaparvata lugens, a significant piercing-sucking pest feeding on rice exclusively, secretes various salivary components when feeding. Salivary proteins are essential for BPH feeding, but their response to etofenprox is not well understood. The application of etofenprox down-regulated the expression of 21 salivary protein genes, among which 9 genes (NlShpa, Salivap 3, CA, NlSEF1, Nl12, NlHSC70-3, NlSP1, NlG14, and NlDNAJB9) showed significant differences. Most differentially expressed genes are found important for BPH physiological processes, except Nl12. Here we found that silencing Nl12 impeded ovary development, thereby inhibiting oocyte formation. The potential explanation was that Nl12 was highly expressed in both salivary gland and ovary, and the ovary development abnormality may be due to the direct effect from expression reduction in ovary and/or indirect influence from expression reduction in salivary gland. Altogether, our findings provide a new insight into the mechanism of action of etofenprox on insect pests and explain part of the reason why etofenprox does not stimulate reproduction in BPH.

Area-wide survey and monitoring of insecticide resistance in the brown planthopper, Nilaparvata lugens (Stål), from 2020 to 2023 in China.

The brown planthopper (BPH), Nilaparvata lugens (Stål), is a notorious pest affecting Asian rice crops. The evolution of insecticide resistance in BPH has emerged as a significant challenge in effectively managing this pest. This study revealed the resistance status of BPH to nine insecticides in ten provinces and Shanghai City in China from 2020 to 2023. Monitoring results showed that the resistance of BPH to triflumezopyrim, nitenpyram, and dinotefuran increased rapidly. The average resistance ratio of BPH to triflumezopyrim increased from 2.5 to 7.1 fold, nitenpyram from 18.3 to 37.7 fold, and dinotefuran from 119.5 to 268.1 fold. All populations remained extremely high resistance to imidacloprid, thiamethoxam, and buprofezin. Most field populations of BPH maintained moderate resistance to chlorpyrifos and sulfoxaflor, and high resistance to pymetrozine by rice stem dipping method. However, considering the reproduction-inhibiting character of pymetrozine, susceptible to low resistance levels to pymetrozine were monitored by Insecticide Resistance Action Committee (IRAC) NO.005 method. This result indicated that pymetrozine might lose efficacy in the control of application generation, but it could significantly inhibit the reproduction of field populations of BPH. Additionally, we compared the expression levels of 11 nicotinic acetylcholine receptor (nAChR) genes, the targets of nAChR competitive modulators, in four field populations (FY23, YH23, LJ23, LP23) and susceptible strain. The expression level of nAChR α4 was significantly reduced in all field populations, while α1, α2, α6, and α7 were significantly reduced in some field populations. Our findings provide valuable information for resistance management strategies in N. lugens and offer new insights into the resistance mechanisms of nAChR competitive modulators.

Harnessing Lecanicillium attenuatum: A novel strategy for combatting Nilaparvata lugens in rice fields.

Nilaparvata lugens is a notorious rice pest causing significant annual yield and economic losses. The use of entomopathogenic fungi offers a promising and eco-friendly approach to sustainable pest management programs. However, research in this area is currently limited to a few specific types of insects and other arthropods. This study aimed to analyze the biocontrol potential of Lecanicillium attenuatum against N. lugens. Bioassays showed that L. attenuatum 3166 induced >80% mortality in N. lugens following 7 d exposure. Greenhouse and field investigations demonstrated that L. attenuatum 3166 application leads to a substantial reduction in N. lugens populations. Under greenhouse conditions, fluorescence was detected in GFP-labeled L. attenuatum 3166 hyphae enveloping the bodies of N. lugens. In field trials, L. attenuatum 3166 treatment exhibited a control efficacy of up to 68.94% at 14 d post-application, which was comparable to that of the commercial entomopathogenic fungal agent. Genomic sequencing of L. attenuatum 3166 revealed a comprehensive array of genes implicated in its infestation and lethality. Further, the transcriptome sequencing analysis highlighted the elevated expression levels of genes encoding proteases, chitinases, cutinases, and phospholipases. Our findings highlight the potential of L. attenuatum 3166 as an effective biological control agent against N. lugens.

Effects of imidacloprid combined with validamycin on the population fitness and symbiotic of Nilaparvata lugens (Hemiptera: Delphacidae).

Using a high-efficiency insecticide in combination with fungicides that have different mechanisms of action is a conventional method in the current management of brown planthopper (BPH) resistance. In this study, we investigate the separate and combined effects of the low-toxicity fungicide validamycin and the non-cross-resistant insecticide imidacloprid on the fitness and symbiosis of BPH. These research results indicate that when the proportion of active ingredients in validamycin is combined with imidacloprid at a ratio of 1:30, the toxicity ratio and co-toxicity coefficient are 1.34 and 691.73, respectively, suggesting that the combination has a synergistic effect on the control of BPH. The number of yeast-like symbiotic (YLS) and dominant symbiotic (Noda) in the imidacloprid + validamycin groups were significantly lower than the other three treatment groups (validamycin, imidacloprid, and water). The results of the study on population fitness show that the lifespan of the BPH population in validamycin, imidacloprid, and imidacloprid + validamycin was shortened. Notably, the BPH populations in the imidacloprid + validamycin groups were significantly lower than other groups in terms of average generation cycle, intrinsic growth rate, net reproduction rate, finite rate of increase, and fitness. The Real-time quantitative PCR showed that validamycin and imidacloprid + validamycin can significantly inhibit the expression of the farnesyl diphosphate farnesyl transferase gene (EC2.5.1.21) and uricase gene (EC1.7.3.3), with imidacloprid + validamycin demonstrating the most pronounced inhibitory effect. Our research results can provide insights and approaches for delaying resistance and integrated management of BPH.

Various functions of detoxification enzymes against insecticides in Nilaparvata lugens selected by toxicity assays and RNAi methods.

The brown planthopper (BPH), Nilaparvata lugens is a devastating agricultural pest of rice, and they have developed resistance to many pesticides. In this study, we assessed the response of BPH nymphs to nitenpyram, imidacloprid, and etofenprox using contact and dietary bioassays, and investigated the underlying functional diversities of BPH glutathione-S-transferase (GST), carboxylesterase (CarE) and cytochrome P450 monooxygenase (P450) against these insecticides. Both contact and ingestion toxicity of nitenpyram to BPH were significantly higher than either imidacloprid or etofenprox. Under the LC50 concentration of each insecticide, they triggered a distinct response for GST, CarE, and P450 activities, and each insecticide induced at least one detoxification enzyme activity. These insecticides almost inhibited the expression of all tested GST, CarE, and P450 genes in contact bioassays but induced the transcriptional levels of these genes in dietary bioassays. Silencing of NlGSTD2 expression had the greatest effect on BPH sensitivity to nitenpyram in contact test and imidacloprid in dietary test. The sensitivities of BPH to insecticide increased the most in the contact test was etofenprox after silencing of NlCE, while the dietary test was nitenpyram. Knockdown of NlCYP408A1 resulted in BPH sensitivities to insecticide increasing the most in the contact test was nitenpyram, while the dietary test was imidacloprid. Taken together, these findings reveal that NlGSTD2, NlCE, and NlCYP408A1 play an indispensable role in the detoxification of the contact and ingestion toxicities of different types of insecticides to BPH, which is of great significance for the development of new strategies for the sucking pest control.

The G932C mutation of chitin synthase 1 gene (CHS1) mediates buprofezin resistance as confirmed by CRISPR/Cas9-mediated knock-in approach in the brown planthopper, Nilaparvata lugens.

The brown planthopper (Nilaparvata lugens) is a major destructive rice pest in Asia. High levels of insecticide resistance have been frequently reported, and the G932C mutation in the chitin synthase 1 (CHS1) gene has been found to mediate buprofezin resistance. However, there has been no direct evidence to confirm the functional significance of the single G932C substitution mutation leading to buprofezin resistance in N. lugens. Here, we successfully constructed a knock-in homozygous strain (Nl-G932C) of N. lugens using CRISPR/Cas9 coupled with homology-directed repair (HDR). Compared with the background strain susceptible to buprofezin (Nl-SS), the knock-in strain (Nl-G932C) showed a 94.9-fold resistance to buprofezin. Furthermore, resistant strains (Nl-932C) isolated from the field exhibited a 2078.8-fold resistance to buprofezin, indicating that there are other mechanisms contributing to buprofezin resistance in the field. Inheritance analysis showed that the resistance trait is incomplete dominance. In addition, the Nl-G932C strain had a relative fitness of 0.33 with a substantially decreased survival rate, emergence rate, and fecundity. This study provided in vivo functional evidence for the causality of G932C substitution mutation of CHS1 with buprofezin resistance and valuable information for facilitating the development of resistance management strategies in N. lugens. This is the first example of using CRISPR/Cas9 gene-editing technology in a hemipteran insect to directly confirm the role of a candidate target site mutation in insecticide resistance.

Resistance to Planthoppers and Southern Rice Black-Streaked Dwarf Virus in Rice Germplasms.

The damage caused by the white-back planthopper (WBPH, Sogatella furcifera) and brown planthopper (BPH, Nilaparvata lugens), as well as southern rice black-streaked dwarf virus (SRBSDV), considerably decreases the grain yield of rice. Identification of rice germplasms with sufficient resistance to planthoppers and SRBSDV is essential to the breeding and deployment of resistant varieties and, hence, the control of the pests and disease. In this study, 318 rice accessions were evaluated for their reactions to the infestation of both BPH and WBPH at the seedling stage using the standard seed-box screening test method; insect quantification was further conducted at the end of the tillering and grain-filling stages in field trials. Accessions HN12-239 and HN12-328 were resistant to both BPH and WBPH at all tested stages. Field trials were conducted to identify resistance in the collection to SRBSDV based on the virus infection rate under artificial inoculation. Rathu Heenati (RHT) and HN12-239 were moderately resistant to SRBSDV. In addition, we found that WBPH did not penetrate stems with stylets but did do more probing bouts and xylem sap ingestion when feeding on HN12-239 than the susceptible control rice Taichung Native 1. The resistance of rice accessions HN12-239, HN12-328, and RHT to BPH, WBPH, and/or SRBSDV should be valuable to the development of resistant rice varieties.

Silencing an ATP-Dependent Caseinolytic Protease Proteolytic Subunit Gene Enhances the Resistance of Rice to Nilaparvata lugens.

The ATP-dependent caseinolytic protease (Clp) system has been reported to play an important role in plant growth, development, and defense against pathogens. However, whether the Clp system is involved in plant defense against herbivores remains largely unclear. We explore the role of the Clp system in rice defenses against brown planthopper (BPH) Nilaparvata lugens by combining chemical analysis, transcriptome, and molecular analyses, as well as insect bioassays. We found the expression of a rice Clp proteolytic subunit gene, OsClpP6, was suppressed by infestation of BPH gravid females and mechanical wounding. Silencing OsClpP6 enhanced the level of BPH-induced jasmonic acid (JA), JA-isoleucine (JA-Ile), and ABA, which in turn promoted the production of BPH-elicited rice volatiles and increased the resistance of rice to BPH. Field trials showed that silencing OsClpP6 decreased the population densities of BPH and WBPH. We also observed that silencing OsClpP6 decreased chlorophyll content in rice leaves at early developmental stages and impaired rice root growth and seed setting rate. These findings demonstrate that an OsClpP6-mediated Clp system in rice was involved in plant growth-defense trade-offs by affecting the biosynthesis of defense-related signaling molecules in chloroplasts. Moreover, rice plants, after recognizing BPH infestation, can enhance rice resistance to BPH by decreasing the Clp system activity. The work might provide a new way to breed rice varieties that are resistant to herbivores.

Long-wave opsin involved in body color plastic development in Nilaparvata lugens.

BACKGROUND: As one of the components of visual photopigments in photoreceptor cells, opsin exhibits different spectral peaks and plays crucial roles in visual function. Besides, it is discovered to evolve other functions despite color vision. However, research on its unconventional function is limited nowadays. With the increase in genome database numbers, various numbers and types of opsins have been identified in insects due to gene duplications or losses. The Nilaparvata lugens (Hemiptera) is a rice pest known for its long-distance migration capability. In this study, opsins were identified in N. lugens and characterized by genome and transcriptome analyses. Meanwhile, RNA interference (RNAi) was carried out to investigate the functions of opsins, and then the Illumina Novaseq 6000 platform-based transcriptome sequencing was performed to reveal gene expression patterns. RESULTS: Four opsins belonging to G protein-coupled receptors were identified in the N. lugens genome, including one long-sensitive opsin (Nllw) together with two ultraviolet-sensitive opsins (NlUV1/2) and an additional new opsin with hypothesized UV peak sensitivity (NlUV3-like). A tandem array of NlUV1/2 on the chromosome suggested the presence of a gene duplication event, with similar exons distribution. Moreover, as revealed by spatiotemporal expression, the four opsins were highly expressed in eyes with age-different expression levels. Besides, RNAi targeting each of the four opsins did not significantly affect the survival of N. lugens in phytotron, but the silencing of Nllw resulted in the melanization of body color. Further transcriptome analysis revealed that silencing of Nllw resulted in up-regulation of a tyrosine hydroxylase gene (NlTH) and down-regulation of an arylalkylamine-N-acetyltransferases gene (NlaaNAT) in N. lugens, demonstrating that Nllw is involved in body color plastic development via the tyrosine-mediated melanism pathway. CONCLUSIONS: This study provides the first evidence in a Hemipteran insect that an opsin (Nllw) takes part in the regulation of cuticle melanization, confirming a cross-talk between the gene pathways underlying the visual system and the morphological differentiation in insects.

Comprehensive transcriptomic analysis of three varieties with different brown planthopper-resistance identifies leaf sheath lncRNAs in rice.

BACKGROUND: Long non-coding RNAs (lncRNAs) have been brought great attention for their crucial roles in diverse biological processes. However, systematic identification of lncRNAs associated with specialized rice pest, brown planthopper (BPH), defense in rice remains unexplored. RESULTS: In this study, a genome-wide high throughput sequencing analysis was performed using leaf sheaths of susceptible rice Taichung Native 1 (TN1) and resistant rice IR36 and R476 with and without BPH feeding. A total of 2283 lncRNAs were identified, of which 649 lncRNAs were differentially expressed. During BPH infestation, 84 (120 in total), 52 (70 in total) and 63 (94 in total) of differentially expressed lncRNAs were found only in TN1, IR36 and R476, respectively. Through analyzing their cis-, trans-, and target mimic-activities, not only the lncRNAs targeting resistance genes (NBS-LRR and RLKs) and transcription factors, but also the lncRNAs acting as the targets of the well-studied stress-related miRNAs (miR2118, miR528, and miR1320) in each variety were identified. Before the BPH feeding, 238 and 312 lncRNAs were found to be differentially expressed in TN1 vs. IR36 and TN1 vs. R476, respectively. Among their putative targets, the plant-pathogen interaction pathway was significantly enriched. It is speculated that the resistant rice was in a priming state by the regulation of lncRNAs. Furthermore, the lncRNAs extensively involved in response to BPH feeding were identified by Weighted Gene Co-expression Network Analysis (WGCNA), and the possible regulation networks of the key lncRNAs were constructed. These lncRNAs regulate different pathways that contribute to the basal defense and specific resistance of rice to the BPH. CONCLUSION: In summary, we identified the specific lncRNAs targeting the well-studied stress-related miRNAs, resistance genes, and transcription factors in each variety during BPH infestation. Additionally, the possible regulating network of the lncRNAs extensively responding to BPH feeding revealed by WGCNA were constructed. These findings will provide further understanding of the regulatory roles of lncRNAs in BPH defense, and lay a foundation for functional research on the candidate lncRNAs.

Silencing a dehydration-responsive element-binding gene enhances the resistance of plants to a phloem-feeding herbivore.

Herbivore-induced plant defence responses share common components with plant responses to abiotic stresses. However, whether abiotic stress-responsive factors influence the resistance of plants to herbivores by regulating these components remains largely unknown. Here, we cloned a dehydration-responsive element-binding gene in rice, OsDREB1A, and investigated its role in the resistance of rice to the phloem-feeding herbivore, brown planthopper (BPH, Nilaparvata lugens), under normal and low temperatures. We found that OsDREB1A localized to the nucleus, and its transcripts in rice were up-regulated in response to BPH infestation, low temperatures and treatment with methyl jasmonate or salicylic acid. Silencing OsDREB1A changed transcript levels of two defence-related WRKY and two PLD genes, enhanced levels of jasmonic acid (JA), JA-isoleucine and abscisic acid, and decreased the ethylene level in rice; these changes subsequently enhanced the resistance of plants to BPH, especially at 17°C, by decreasing the hatching rate and delaying the development of BPH eggs. Moreover, silencing OsDREB1A increased the growth of rice plants. These findings suggest that OsDREB1A, which positively regulates the resistance of rice to abiotic stresses, negatively regulates the resistance of rice to BPH.

The salivary chaperone protein NlDNAJB9 of Nilaparvata lugens activates plant immune responses.

The brown planthopper (BPH) Nilaparvata lugens (Stål) is a main pest on rice. It secretes saliva to regulate plant defense responses, when penetrating rice plant and sucking phloem sap through its stylet. However, the molecular mechanisms of BPH salivary proteins regulating plant defense responses remain poorly understood. A N. lugens DNAJ protein (NlDNAJB9) gene was highly expressed in salivary glands, and the knock down of NlDNAJB9 significantly enhanced honeydew excretion and fecundity of the BPH. NlDNAJB9 could induce plant cell death, and the overexpression of NlDNAJB9 gene in Nicotiana benthamiana induced calcium signaling, mitogen-activated protein kinase (MAPK) cascades, reactive oxygen species (ROS) accumulation, jasmonic acid (JA) hormone signaling and callose deposition. The results from different NlDNAJB9 deletion mutants indicated that the nuclear localization of NlDNAJB9 was not necessary to induce cell death. The DNAJ domain was the key region to induce cell death, and the overexpression of DNAJ domain in N. benthamiana significantly inhibited insect feeding and pathogenic infection. NlDNAJB9 might interact indirectly with NlHSC70-3 to regulate plant defense responses. NlDNAJB9 and its orthologs were highly conserved in three planthopper species, and could induce ROS burst and cell death in plants. Our study provides new insights into the molecular mechanisms of insect-plant interactions.

The microRNA pathway core genes are indispensable for development and reproduction in the brown planthopper, Nilaparvata lugens.

MicroRNAs (miRNAs) are small single-stranded non-coding RNAs involved in a variety of cellular events by regulating gene expression at the post-transcriptional level. Several core genes in miRNA biogenesis have been reported to participate in a wide range of physiological events, in some insect species. However, the functional significance of miRNA pathway core genes in Nilaparvata lugens remains unknown. In the present study, we conducted a systematic characterisation of five core genes involved in miRNA biogenesis. We first performed spatiotemporal expression analysis and found that miRNA core genes exhibited similar expression patterns, with high expression levels in eggs and relatively high transcriptional levels in the ovaries and fat bodies of females. RNA interference experiments showed that injecting third-instar nymphs with dsRNAs targeting the miRNA core genes, NlAgo1, NlDicer1, and NlDrosha resulted in high mortality rates and various degrees of body melanism, moulting defects, and wing deformities. Further investigations revealed that the suppression of miRNA core genes severely impaired ovarian development and oocyte maturation, resulting in significantly reduced fecundity and disruption of intercellular spaces between follicle cells. Moreover, the expression profiles of miR-34-5p, miR-275-3p, miR-317-3p, miR-14, Let-7-1, and miR-2a-3p were significantly altered in response to the knockdown of miRNA core genes mixture, suggesting that they play essential roles in regulating miRNA-mediated gene expression. Therefore, our results provide a solid theoretical basis for the miRNA pathway in N. lugens and suggest that the NlAgo1, NlDicer1, and NlDrosha-dependent miRNA core genes are essential for the development and reproduction of this agricultural pest.