Lugens Feeding Research Articles

Cross-kingdom RNA interference mediated by insect salivary microRNAs may suppress plant immunity

Communication between insects and plants relies on the exchange of bioactive molecules that traverse the species interface. Although proteinic effectors have been extensively studied, our knowledge of other molecules involved in this process remains limited. In this study, we investigate the role of salivary microRNAs (miRNAs) from the rice planthopper Nilaparvata lugens in suppressing plant immunity. A total of three miRNAs were confirmed to be secreted into host plants during insect feeding. Notably, the sequence-conserved miR-7-5P is specifically expressed in the salivary glands of N. lugens and is secreted into saliva, distinguishing it significantly from homologues found in other insects. Silencing miR-7-5P negatively affects N. lugens feeding on rice plants, but not on artificial diets. The impaired feeding performance of miR-7-5P-silenced insects can be rescued by transgenic plants overexpressing miR-7-5P. Through target prediction and experimental testing, we demonstrate that miR-7-5P targets multiple plant genes, including the immune-associated bZIP transcription factor 43 (OsbZIP43). Infestation of rice plants by miR-7-5P-silenced insects leads to the increased expression of OsbZIP43, while the presence of miR-7-5P counteracts this upregulation effect. Furthermore, overexpressing OsbZIP43 confers plant resistance against insects which can be subverted by miR-7-5P. Our findings suggest a mechanism by which herbivorous insects have evolved salivary miRNAs to suppress plant immunity, expanding our understanding of cross-kingdom RNA interference between interacting organisms.

Effect of Silicon Amendment on Herbivore Induced Plant Volatiles of Rice Plant Infested by Brown Planthopper Nilaparvata Lugens (Stål)

Silicon (Si) is known to play a very important role in a plant’s direct and indirect defense. In rice plants, its impact on induced volatiles released upon Brown planthopper (BPH), Nilaparvata lugens feeding is less understood. The BPH-induced volatile compounds from Si amended rice plants were analyzed using Gas Chromatography-Mass Spectrometry (GC-MS). The alteration in Herbivore Induced Plant Volatiles (HIPV’s) blend was observed, wherein, total 38 HIPVs were found to be differentially released. The HIPV mainly belongs to alkane, alkene, alcohol and terpene groups. Overall, Si amendment caused a significant effect on the composition of HIPVs in rice, some of which are involved in tri-trophic interaction.

No impact of transgenic cry2Aa rice on Anagrus nilaparvatae, an egg parasitoid of Nilaparvata lugens, in laboratory tests

Anagrus nilaparvatae (Hymenoptera: Mymanidae), is an egg parasitoid of the rice planthopper, Nilaparvata lugens. In the current study, a seven generation bioassay was used to test for host-mediated effects of transgenic cry2Aa rice (T2A-1) on this parasitoid. When A. nilaparvatae parasitized the eggs of N.lugens reared on either Bt or non-Bt rice, there were no differences in survival, development time, longevity, sex ratio or fecundity over seven generations. In addition, direct exposure to a high dosage of Cry2Aa had no adverse effects on the survival or fecundity of A. nilaparvatae adults. Cry2Aa protein could not be detected in the eggs of N. lugens or in the adults of A. nilaparvatae parasitizing eggs of N. lugens feeding on Bt rice. These results indicate that A. nilaparvatae are not sensitive to Cry2Aa protein at the tested concentration and there were no detrimental effects on any life-table parameters in our study.

Selection of Reference Genes for Gene Expression Analysis in Nilaparvata lugens with Different Levels of Virulence on Rice by Quantitative Real-Time PCR

The brown planthopper Nilaparvata lugens Stål (Homoptera: Delphacidae) can cause hopperburn by feeding on rice and also can transmit the grassy stunt disease. Resistant rice varieties have been developed, but several N. lugens strains can recover their virulence to these resistant rice varieties. In the present study, reference genes with stable expression levels in N. lugens populations showed different levels of virulence to susceptible and resistant rice varieties. The expression of six candidate reference genes in N. lugens feeding on susceptible and resistant rice varieties was analyzed. These genes were evaluated for their potential use in the analysis of differential gene expression. Polymerase chain reaction data was generated from N. lugens, including two different treatments (resistant or susceptible rice) and three virulent N. lugens populations. Three software programs (BestKeeper, Normfinder and geNorm) were used to assess the candidate reference genes. Both geNorm and Normfinder identified the genes 18S, β-ACT, β-TUB and α-TUB as the most stable reference genes. BestKeeper identified ETIF1 as the optimal reference gene with the least overall variation, whereas 18S and α-TUB were the second and third most stably expressed genes, respectively. Therefore, we concluded that the genes 18S and α-TUB were the most suitable reference genes in N. lugens. These results will facilitate future transcript profiling studies on N. lugens populations that show variation in virulence levels on different rice varieties.

Feeding‐induced changes in defence enzymes and PR proteins and their implications in host resistance to Nilaparvata lugens

AbstractSix rice genotypes showing susceptible and resistant reactions to brown planthopper (BPH), Nilaparvata lugens were studied for feeding‐induced changes in defence enzymes and pathogenesis‐related (PR) proteins. The high resistant genotypes PTB 33, ADT 45 and ASD 7 and moderately resistant genotypes CO 43 and KAU 1661 recorded the greater expression of defence enzymes peroxidase, polyphenol oxidase, phenylalanine ammonia lyase, total phenol and β‐1,3 glucanase in response to N. lugens feeding at 1 day after infestation (DAI) compared with susceptible genotype TN1. The greater activity of chitinase was observed in resistant cultivars at 3 DAI and the activity was sustained for more than 1 week compared with susceptible TN1. In conclusion, the current study revealed that these defence enzymes and PR proteins might attribute to the resistance mechanisms in rice plants against BPH infestation.

Cultivar and Insecticide Applications Affect the Physiological Development of the Brown Planthopper, <I>Nilaparvata lugens</I> (Stål) (Hemiptera: Delphacidae)

Nilaparvata lugens (Stål) (Hemiptera:Delphacidae) is a serious migratory pest of rice in Asia. Pest resurgence often occurs because of insecticide overuse. Using both susceptible (TN1) and moderately resistant (Xieyou 963) cultivars, we studied the effect of foliar insecticides on the percentage of brachypterous adults, female reproductive rate, and crude fat and soluble sugar contents in third- and fifth-instar nymphs and adults. The percentage of brachypterous adults and reproductive rate of adult females developed from nymphs that fed on insecticide-treated plants varied significantly with rice cultivar, type of insecticide, and its concentration. Feeding on susceptible plants increased the percentage of brachypterous adults and reproductive rate of adult females. Also, deltamethrin increased brachypterous production relative to imidacloprid and triazophos. The highest reproductive rate was on plants treated with triazophos. All insecticide treatments in both cultivars resulted in increase of soluble sugar contents in third- and fifth-instar nymphs and adults developed from nymphs feeding on insecticide-treated rice plants. This effect was stronger on the susceptible cultivar. Changes of crude fat content after N. lugens feeding on insecticide-treated plants were related to its feeding duration. Crude fat content in adult developed from nymphs feeding on treated plants was significantly higher that on control plants. These studies showed that plant and insecticide influences on physiological ecology of this planthopper will influence its population dynamics under commercial production of rice in Asia.

Effects of Cry1Ab Toxin onPropylea japonica(Thunberg) (Coleoptera: Coccinellidae) Through Its Prey,Nilaparvata lugensStål (Homoptera: Delphacidae), Feeding on TransgenicBtRice

Laboratory feeding experiments using rice plants containing a synthetic cry1Ab gene derived from Bacillus thuringiensis Berliner (Bt) were carried out to study the effects of Bt rice-fed prey on the predator Propylea japonica (Thunberg). Plants were obtained from two homozygous transgenic Bt-cry1Ab expressing rice lines, Kemingdao 1 (KMD1) and Kemingdao 2 (KMD2), and their untransformed parental variety Xiushui 11 (XS11). The herbivorous prey species tested was the brown planthopper, Nilaparvata lugens Stål, one of most serious insect pests of rice and not targeted by KMD1 or KMD2. The concentrations of Cry1Ab toxin expressed in KMD1 and KMD2 plants and that of the toxin transferred to N. lugens feeding on these plants were determined by enzyme immunosorbent assay technique. Development parameters of P. japonica reared on KMD1- or KMD2-fed N. lugens were assessed in the laboratory. The results showed that the concentration of Cry1Ab in rice leaves and stems significantly increased from the booting to grain filling stage and subsequently decreased as the plants matured. Cry1Ab could be detected in nymphs and adults of N. lugens feeding on the Bt rice plants. Development time, pupation, adult eclosion, pupal and adult weight, and male-adult locomotive activity of P. japonica that had preyed on KMD1- or KMD2-fed N. lugens nymphs as larvae were not significantly different from those that preyed on XS11-fed nymphs. In short, our results indicate that the nontarget insect N. lugens and its predator P. japonica are exposed to Cry1Ab toxin from transgenic cry1Ab rice, but development of this predator was not affected by the toxin through tritrophic interactions.

Responses in nutrient uptake in rice roots to infestation of brown planthopper, Nilaparvata lugens (Stål) (Homoptera: Delphacidae)

This study investigates the uptake of nitrogen (N), phosphorus (P), and potassium (K) by rice roots as a response to infestation by the rice brown planthopper, Nilaparvata lugens (Stål) (Homoptera: Delphacidae) at two developmental stages (i.e. beginning-heading and grain-filling) of two rice varieties, Zhendao 2 (japonics rice) and Xieyou 63 (hybrid indica rice), respectively. The results show that the impact of N. lugens infestation at the beginning-heading stage on N, P, and K uptake was not only more serious, but also that the recovery rate of root uptake after the removal of N. lugens was slower, as compared to the grain-filling stage. The percent reduction of N uptake following N. lugens infestations at the beginning-heading stage of Xieyou 63 was significantly greater in comparison to Zhendao 2 at the same stage. There was no significant difference in N uptake between two developmental stages, but P and K uptake by roots following N. lugens infestation were significantly different between the two developmental stages. The maximum reduction of K uptake was shown at the beginning-heading stage in both rice varieties. The recovery rate of N, P, and K uptake by roots after the removal of N. lugens from rice plants varied with the rice variety, developmental stage of rice and infestation duration. In Xieyou 63, the recovery rate of K uptake by roots of the plants infested at the beginning-heading stage was slower than that at the grain-filling stage. Plants of Zhendao 2 infested at the two developmental stages had showed slower recovery rate of K uptake compared with N and P uptakes. These findings partially explain the virulence of N. lugens feeding on rice plants, and thus may have important implications for integrated management of N. lugens.

Identification of differentially expressed genes in brown planthopper Nilaparvata lugens (Hemiptera: Delphacidae) responding to host plant resistance

The brown planthopper Nilaparvata lugens Stål is one of the major insect pests of rice Oryza sativa L. The host resistance exhibits profound effects on growth, development and propagation of N. lugens. To investigate the molecular response of N. lugens to host resistance, a cDNA-amplified fragment length polymorphism (cDNA-AFLP) technique was employed to identify the differentially expressed genes in the nymphs feeding on three rice varieties. Of the 2,800 cDNA bands analysed, 54 were up-regulated and seven down-regulated qualitatively in N. lugens when the ingestion sources were changed from susceptible rice plants to resistant ones. Sequence analysis of the differential transcript-derived fragments showed that the genes involved in signalling, stress response, gene expression regulation, detoxification and metabolism were regulated by host resistance. Four of the transcript-derived fragments corresponding to genes encoding for a putative B subunit of phosphatase PP2A, a nemo kinase, a cytochrome P450 monooxygenase and a prolyl endopeptidase were further characterized in detail. Northern blot analysis confirmed that the expression of the four genes was enhanced in N. lugens feeding on resistant rice plants. The roles of these genes in the defensive response of N. lugens to host plant resistance were discussed.

Identification of genes responsive to brown planthopper Nilaparvata lugens St�l (Homoptera: Delphacidae) feeding in rice

The brown planthopper (Nilaparvata lugens Stal) is a rice (Oryza sativa L.)-specific herbivore that ingests assimilates from the phloem, which leads to stunted growth and therefore poor yield of rice. In this study, 27 planthopper-responsive genes in rice plants were isolated by suppression subtractive hybridization (SSH), and confirmed by filter cDNA array hybridization and northern blot analysis. Among these transcripts, 25 were induced and 2 were suppressed by N. lugens feeding. The responsive genes are involved in photosynthesis, macromolecule degradation, signal transduction and defenses against stress and pathogens. Overall, genes involved in macromolecule degradation and plant defenses were up-regulated, whereas those involved in photosynthesis and cell growth were down-regulated. Our data suggest that leaf senescence is most likely activated in the susceptible rice plants damaged by N. lugens insects.

Comparing Nilaparvata lugens (Stal) and Sogatella furcifera (Horvath) (Homoptera: Delphacidae) feeding effects on rice plant growth processes at the vegetative stage

The brown planthopper, Nilaparvata lugens, and the white-backed planthopper, Sogatella furcifera, are phloem feeders. Experiments were conducted to compare plant response of japonica and indica rice cultivars to N. lugens and S. furcifera feeding, and quantify the amount of feeding by these hopper species on various rice cultivars. Plant height, tiller number, leaf area and organ dry weights, and leaf photosynthetic rates and organ nitrogen contents were measured to determine plant response to hopper feeding. Hopper dry weights and honeydew weights were measured to quantify the amount of feeding by the hoppers and determine the effects of cultivar on hoppers. Reduction in plant height caused by S. furcifera feeding was greater than that of N. lugens. The effects of N. lugens feeding on roots were greater than those of S. furcifera. These results suggest a difference in the mechanism of damage caused by S. furcifera and N. lugens even though both hoppers are phloem feeders. Regardless of rice type, the reaction of different cultivars to S. furcifera and N. lugens feeding varied. Hinohikari was most susceptible to S. furcifera and N. lugens feeding, whereas Daili Zaoxian and Nipponbare were more susceptible to N. lugens and S. furcifera feeding, respectively. S. furcifera were smaller and excreted less honeydew than N. lugens, but their effects on tillers, leaf area and plant weight were similar to those of N. lugens suggesting that S. furcifera feeding effects at vegetative stage was greater than those of N. lugens. The demand for nitrogen was probably greater for S. furcifera than N. lugens, because the former contains greater nitrogen and thrives on the upper portion of the rice plant; however, this needs further investigation.

Probing behavior of the brown planthopper, Nilaparvata lugens Stal(Homoptera: Delphacidae) on a non-host barnyard grass, and resistant and susceptible varieties of rice.

The feeding activity of Nilaparvata lugens (Stål) (Homoptera: Delphacidae) was compared on the barnyard grass, Echinochloa Crus-galli var. oryzicola, which contains an antifeedant, (E)-aconitic acid, and on resistant (bph4) and susceptible rice varieties using an AC electronic monitoring system (EMS). N. lugens made more frequent probes on the barnyard grass and the resistant rice than on the susceptible rice. Mean duration of probes tended to be the shortest and the percentage of probes that consisted of the primarily salivation phase only was the highest on the barnyard grass, followed by the resistant rice. The most distinct difference in N. lugens feeding activities between the barnyard grass and the resistant rice was the frequency of the primarily ingestion phase. On the barnyard grass, the insect was rarely successful in achieving the primarily ingestion phase, indicating that probing was interrupted before the arrival of the stylets at the sieve elements. On the other hand, on the resistant rice the primarily ingestion phase was observed as frequently as it was on the susceptible rice. However, it stopped within 5 min on the resistant rice in most cases. Probing activities were strongly inhibited by 1% (E)-aconitic acid diluted with distilled water. The inhibitory effect was considerably mitigated when diluted with 15% sucrose. These results suggest that on the barnyard grass, interruption of probing activity by (E)-aconitic acid can occur in stylet pathway tissues rather than in ingestion tissues (e.g. the phloem where sucrose is located).

Levels of Tolerance and Antibiosis in Rice Varieties Having Moderate Resistance to the Brown Planthopper, Nilaparvata lugens (Stål) (Hemiptera: Delphacidae)

Rice varieties having a moderate level of resistance to the brown planthopper, Nilaparvata lugens, were evaluated to determine the mechanisms of resistance. The host plant-insect interactions were studied in the greenhouse and insectary to determine the levels of antibiosis and tolerance in the moderately resistant varieties. Parameters measured as the tolerance indicator were plant damage, plant weight loss, and yield reduction due to insect feeding. The insect biomass production, growth index, and population growth rate on the test varieties were used as antibiosis indicators. A regression of plant weight loss, due to N. lugens feeding, on N. lugens weight for five varieties indicated that the variety ‘Utri Rajapan’ has a high level of tolerance and no antibiosis. The resistance in ‘IR46,’ ‘Kencana,’ and ‘Triveni’ is due to a combination of antibiosis and tolerance. The relative amount of each of the two resistance components was measured by using the tolerance-antibiosis index. Field studies confirmed results obtained in the greenhouse and insectary studies. Although ‘Utri Rajapan’ had field populations of N. lugens equal to that of the susceptible check, yield reduction was only 25%, compared with 100% in the susceptible check.