Histopathological Alterations in Nilaparvata lugens (Hemiptera: Delphacidae) after Exposure to Cordyceps javanica.

The brown planthopper (BPH), Nilaparvata lugens (Stål, 1854), is a pest of rice plants worldwide. Cordyceps javanica is a destructive entomopathogenic fungus known to attack leafhoppers or BPHs specifically. Live adult BPH samples were inoculated with isolated C. javanica PSUC002, and their interaction was morpho-histologically examined from 0 to 120 h post-inoculation (pi). We observed that the mortality of BPH continuously increased until 120 h pi (Day 5). Tissue alterations in the host were examined after infection using morphological and histological methods, including the Grocott Methenamine Silver stain test (GMS). Filamentous fungi were first found on the external integument at 12 h pi, and fungal conidia attached to the integument at 24 h pi. However, the initial degeneration of BPHs was identified by histology at 6 h pi especially in the integument and adipose tissue. We identified the degeneration and loss of integument and adipose tissue of infected BPHs at 12 h pi, and their necrosis was completed at 96 h pi. The enzymatic index of the sampled fungi (chitinase and protease) peaked at 7 days of incubation. This study demonstrated that C. javanica PSUC002 is useful to control the BPHs as an eco-friendly practice and will possibly be applied in agriculture.

Osa-miR162a Enhances the Resistance to the Brown Planthopper via α-Linolenic Acid Metabolism in Rice (Oryza sativa).

The brown planthopper (BPH) is the most serious pest causing yield losses in rice. MicroRNAs (miRNAs) are emerging as key modulators of plant-pest interactions. In the study, we found that osa-miR162a is induced in response to BPH attack in the seedling stage and tunes rice resistance to the BPH via the α-linolenic acid metabolism pathway as indicated by gas chromatography/liquid chromatography-mass spectrometry analysis. Overexpression of osa-miR162a inhibited the development and growth of the BPH and simultaneously reduced the release of 3-hexenal and 3-hexen-1-ol to block host recognition in the BPH. Moreover, knockdown of OsDCL1, which is targeted by osa-miR162a, inhibited α-linolenic acid metabolism to enhance the resistance to the BPH, which was similar to that in miR162a-overexpressing plants. Our study revealed a novel defense mechanism mediated by plant miRNAs developed during the long-term evolution of plant-host interaction, provided new ideas for the identification of rice resistance resources, and promoted a better understanding of pest control.

Proteomics and Metabolomics Studies on the Biotic Stress Responses of Rice: an Update.

Biotic stresses represent a serious threat to rice production to meet global food demand and thus pose a major challenge for scientists, who need to understand the intricate defense mechanisms. Proteomics and metabolomics studies have found global changes in proteins and metabolites during defense responses of rice exposed to biotic stressors, and also reported the production of specific secondary metabolites (SMs) in some cultivars that may vary depending on the type of biotic stress and the time at which the stress is imposed. The most common changes were seen in photosynthesis which is modified differently by rice plants to conserve energy, disrupt food supply for biotic stress agent, and initiate defense mechanisms or by biotic stressors to facilitate invasion and acquire nutrients, depending on their feeding style. Studies also provide evidence for the correlation between reactive oxygen species (ROS) and photorespiration and photosynthesis which can broaden our understanding on the balance of ROS production and scavenging in rice-pathogen interaction. Variation in the generation of phytohormones is also a key response exploited by rice and pathogens for their own benefit. Proteomics and metabolomics studies in resistant and susceptible rice cultivars upon pathogen attack have helped to identify the proteins and metabolites related to specific defense mechanisms, where choosing of an appropriate method to identify characterized or novel proteins and metabolites is essential, considering the outcomes of host-pathogen interactions. Despites the limitation in identifying the whole repertoire of responsive metabolites, some studies have shed light on functions of resistant-specific SMs. Lastly, we illustrate the potent metabolites responsible for resistance to different biotic stressors to provide valuable targets for further investigation and application.

Combining next-generation sequencing and single-molecule sequencing to explore brown plant hopper responses to contrasting genotypes of japonica rice.

BACKGROUND: The brown plant hopper (BPH), Nilaparvata lugens, is one of the major pest of rice (Oryza sativa). Plant defenses against insect herbivores have been extensively studied, but our understanding of insect responses to host plants' resistance mechanisms is still limited. The purpose of this study is to characterize transcripts of BPH and reveal the responses of BPH insects to resistant rice at transcription level by using the advanced molecular techniques, the next-generation sequencing (NGS) and the single-molecule, real-time (SMRT) sequencing. RESULTS: The current study obtained 24,891 collapsed isoforms of full-length transcripts, and 20,662 were mapped to known annotated genes, including 17,175 novel transcripts. The current study also identified 915 fusion genes, 1794 novel genes, 2435 long non-coding RNAs (lncRNAs), and 20,356 alternative splicing events. Moreover, analysis of differentially expressed genes (DEGs) revealed that genes involved in metabolic and cell proliferation processes were significantly enriched in up-regulated and down-regulated sets, respectively, in BPH fed on resistant rice relative to BPH fed on susceptible wild type rice. Furthermore, the FoxO signaling pathway was involved and genes related to BPH starvation response (Nlbmm), apoptosis and autophagy (caspase 8, ATG13, BNIP3 and IAP), active oxygen elimination (catalase, MSR, ferritin) and detoxification (GST, CarE) were up-regulated in BPH responses to resistant rice. CONCLUSIONS: The current study provides the first demonstrations of the full diversity and complexity of the BPH transcriptome, and indicates that BPH responses to rice resistance, might be related to starvation stress responses, nutrient transformation, oxidative decomposition, and detoxification. The current result findings will facilitate further exploration of molecular mechanisms of interaction between BPH insects and host rice.

Insights into the structure-function relationship of brown plant hopper resistance protein, Bph14 of rice plant: a computational structural biology approach.

Brown plant hopper (BPH) is one of the major destructive insect pests of rice, causing severe yield loss. Thirty-two BPH resistance genes have been identified in cultivated and wild species of rice Although, molecular mechanism of rice plant resistance against BPH studied through map-based cloning, due to non-existence of NMR/crystal structures of Bph14 protein, recognition of leucine-rich repeat (LRR) domain and its interaction with different ligands are poorly understood. Thus, in the present study, in silico approach was adopted to predict three-dimensional structure of LRR domain of Bph14 using comparative modelling approach followed by interaction study with jasmonic and salicylic acids. LRR domain along with LRR-jasmonic and salicylic acid complexes were subjected to dynamic simulation using GROMACS, individually, for energy minimisation and refinement of the structure. Final binding energy of jasmonic and salicylic acid with LRR domain was calculated using MM/PBSA. Free-energy landscape analysis revealed that overall stability of LRR domain of Bph14 is not much affected after forming complex with jasmonic and salicylic acid. MM/PBSA analysis revealed that binding affinities of LRR domain towards salicylic acid is higher as compared to jasmonic acid. Interaction study of LRR domain with salicylic acid and jasmonic acid reveals that THR987 of LRR form hydrogen bond with both complexes. Thus, THR987 plays active role in the Bph14 and phytochemical interaction for inducing resistance in rice plant against BPH. In future, Bph14 gene and phytochemicals could be used in BPH management and development of novel resistant varieties for increasing rice yield.

Characterization of the Distal-less gene homologue, NlDll, in the brown planthopper, Nilaparvata lugens (Stål).

The brown planthopper, Nilaparvata lugens (Stål), is a globally devastating insect pest of rice, particularly in eastern Asia. Distal-less or Dll is a highly conserved and well studied transcription factor required for limb formation in invertebrates and vertebrates. We have identified a homologue of this gene, NlDll, and demonstrated that it is expressed in all life stages of N. lugens, particularly in adult brachypterous females. When we compared between specific adult tissues it was expressed most strongly in wings. Using RNAi techniques we demonstrated that downregulation of NlDll in the 3rd instar larvae led to the disrupted development of the leg, while downregulation of NlDll in the 5th instar larvae led to abnormal wing formation. Ectopic over-expression of NlDll in Drosophila melanogaster using the GAL4-UAS system led to fatal or visible phenotypic changes such as the loss of normal wing structure and disrupted haltere structure. Our work suggests that NlDll is a conserved homologue of Distal-less and is required for both leg development and wing structure. Since researches have shown that Dll is required for wing morphogenesis, understanding the role of NlDll during the wing development will further provide a basis for revealing the molecular mechanism of the wing dimorphism in brown planthopper. In the future, NlDll could be used as a target gene for brown planthopper pest management in the field.

Evidence of sibling species between two host-associated populations of brown planthopper, N. lugens (stål) (Homoptera: Delphacidea) complex based on morphology and host-plant relationship studies.

Morphological and host-plant relationship studies were conducted to differentiate two sympatric populations of brown planthopper (BPH), Nilaparvata lugens, one from rice (Oryza sativa) and the other from Leersia hexandra, a weed grass. In morphometric studies based on esterase activities, an UPGMA dendrogram using 17 quantitative morphological characters, including stridulatory organs (courtship signal-producing organs) between two sympatric populations of N. lugens, one from rice and the other from L. hexandra, a weed grass revealed that both populations were separated from each other. An out-group, N. bakeri, was found to be completely different from the two sympatric populations of N. lugens. Rice plants were best suited for the establishment of the rice-infesting population, and L. hexandra was a favourable host for the Leersia-infesting population. The individuals derived from one host did not thrive on the other host, as shown by a significant reduction in survival and nymphal development, ovipositional preferences, ovipositional response, and egg hatchability. Therefore, morphological and host-plant relationship studies indicate that rice-associated population with high esterase activities and L. heaxandra-associated population with low esterase activities are two closely related sibling species.