The genome- and transcriptome-wide analysis of innate immunity in the brown planthopper, Nilaparvata lugens.

BACKGROUND: The brown planthopper (Nilaparvata lugens) is one of the most serious rice plant pests in Asia. N. lugens causes extensive rice damage by sucking rice phloem sap, which results in stunted plant growth and the transmission of plant viruses. Despite the importance of this insect pest, little is known about the immunological mechanisms occurring in this hemimetabolous insect species. RESULTS: In this study, we performed a genome- and transcriptome-wide analysis aiming at the immune-related genes. The transcriptome datasets include the N. lugens intestine, the developmental stage, wing formation, and sex-specific expression information that provided useful gene expression sequence data for the genome-wide analysis. As a result, we identified a large number of genes encoding N. lugens pattern recognition proteins, modulation proteins in the prophenoloxidase (proPO) activating cascade, immune effectors, and the signal transduction molecules involved in the immune pathways, including the Toll, Immune deficiency (Imd) and Janus kinase signal transducers and activators of transcription (JAK-STAT) pathways. The genome scale analysis revealed detailed information of the gene structure, distribution and transcription orientations in scaffolds. A comparison of the genome-available hemimetabolous and metabolous insect species indicate the differences in the immune-related gene constitution. We investigated the gene expression profiles with regards to how they responded to bacterial infections and tissue, as well as development and sex expression specificity. CONCLUSIONS: The genome- and transcriptome-wide analysis of immune-related genes including pattern recognition and modulation molecules, immune effectors, and the signal transduction molecules involved in the immune pathways is an important step in determining the overall architecture and functional network of the immune components in N. lugens. Our findings provide the comprehensive gene sequence resource and expression profiles of the immune-related genes of N. lugens, which could facilitate the understanding of the innate immune mechanisms in the hemimetabolous insect species. These data give insight into clarifying the potential functional roles of the immune-related genes involved in the biological processes of development, reproduction, and virus transmission in N. lugens.

Field response of aboveground non-target arthropod community to transgenic Bt-Cry1Ab rice plant residues in postharvest seasons.

Risk assessments of ecological effects of transgenic rice expressing lepidoptera-Cry proteins from Bacillus thuringiensis (Bt) on non-target arthropods have primarily focused on rice plants during cropping season, whereas few studies have investigated the effects in postharvest periods. Harvested rice fallow fields provide a critical over-wintering habitat for arthropods in the Chinese rice ecosystems, particularly in the southern region of the country. During 2006-08, two independent field trials were conducted in Chongqing, China to investigate the effects of transgenic Cry1Ab rice residues on non-target arthropod communities. In each trial, pitfall traps were used to sample arthropods in field plots planted with one non-Bt variety and two Bt rice lines expressing the Cry1Ab protein. Aboveground arthropods in the trial plots during the postharvest season were abundant, while community densities varied significantly between the two trials. A total of 52,386 individual insects and spiders, representing 93 families, was captured in the two trials. Predominant arthropods sampled were detritivores, which accounted for 91.9% of the total captures. Other arthropods sampled included predators (4.2%), herbivores (3.2%), and parasitoids (0.7%). In general, there were no significant differences among non-Bt and Bt rice plots in all arthropod community-specific parameters for both trials, suggesting no adverse impact of the Bt rice plant residues on the aboveground non-target arthropod communities during the postharvest season. The results of this study provide additional evidence that Bt rice is safe to non-target arthropod communities in the Chinese rice ecosystems.

Interspecific competition between two endoparasitoids Cotesia vestalis (Hymenoptera: Braconidae) and Oomyzus sokolowskii (Hymenoptera: Eulophidae).

Two endoparasitoids, Cotesia vestalis and Oomyzus sokolowskii, parasitize the same host, larvae of Plutella xylostella. These two species have evolved different parasitization strategies. O. sokolowskii expresses a single factor, venom, and exerts virtually no detrimental effects on the development of its host. C. vestalis, on the other hand, injects polydnavirus (PDV) and venom during oviposition, and teratocytes are released into the host's hemolymph after egg hatching. Parasitization suppresses host immune reactions and redirects its developmental program. Because both these species parasitize the same stage of their hosts, there is the possibility of multiparasitism in nature. Only one species survives multiparasitism and because of its parasitic strategy, we hypothesized that C. vestalis would invariably be the stronger competitor. We designed competition experiments which revealed that C. vestalis is a stronger competitor than O. sokolowskii. We also show that C. vestalis survives intrinsic competition with O. sokolowskii through two mechanisms: physical attack and physiological suppression. We discovered melanized wounds on O. sokolowskii eggs and larvae, which is strong evidence of physical attacks. The physiological suppression is due to PDV and venom injected by C. vestalis. To test this idea more rigorously, we designed a pseudoparasitization experiment which revealed that no O. sokolowskii emerged from multiparasitized hosts when infertile C. vestlais eggs and normal O. sokolowskii larvae are both present inside the same host. These results support our hypothesis that C. vestalis is the stronger competitor and demonstrate two mechanisms that account for the outcome of intrinsic competition between these two endoparasitoids.

Cloning and expression analysis of a G-protein α subunit--Gαo in the rice water weevil Lissorhoptrus oryzophilus Kuschel.

The open reading frame (ORF) encoding a novel G protein α subunit, Lo Gα(o), was cloned from the parthenogenetic rice water weevil, Lissorhoptrus oryzophilus Kuschel (Coleoptera: Curculionidae). The Lo Gα(o) ORF encodes a protein of 354 amino acid residues. The deduced protein sequence shares high homology with Gα(o) from other species. The expression patterns of Lo Gα(o) in various adult tissues were indicated by real-time quantitative PCR and Western blot. The results showed that Lo Gα(o) mRNA was expressed at similar levels in tissues except relative high levels in the antennae of adult, and Lo Gα(o) protein of an apparent molecular mass of about 40 kDa was expressed in various tissues of the adult. Immunocytochemical localization showed that Lo Gα(o) was mainly expressed in the dendrites of the trichoid sensilla in the antenna of the weevil. The tissue and cellular localization of Lo Gα(o) suggests that Lo Gα(o) may take a part in signal transduction of olfactory/gustatory.

Identification, Characterization, and Distribution of Southern rice black-streaked dwarf virus in Vietnam.

A novel dwarf and twisting syndrome first observed on rice in Nghe An Province, Vietnam, in 2009 has spread rapidly to the other 19 provinces of North and Central Vietnam. Infected rice plants showed stunting, darkening of leaves, twisting of leaf tips, and splitting of leaf margins. At a later stage, white waxy enations that eventually turned black were observed on the underside of leaf blades, leaf sheaths, and culms. The disease also infected maize after rice was harvested. Infected maize plants were stunted and dark green with small enations along the minor veins on the back of leaves. The disease agent has now been identified as Southern rice black-streaked dwarf virus (SRBSDV) recently reported from Southern China. Typical fijivirus viroplasms containing crystalline arrayed spherical virions approximately 70 to 75 nm in diameter were observed under the electron microscope in ultrathin sections of infected rice leaves. The virus was transmitted to rice and maize seedlings by the white-backed planthopper (Sogatella furcimera). A one-step reverse transcription-polymerase chain reaction (RT-PCR) protocol was used to confirm the presence of SRBSDV in 477 samples of rice or maize from 29 provinces among 5 agroecological regions in North and Central Vietnam. Rice black-streaked dwarf virus was not detected in these samples. Partial sequences of RNA segments 4 and 10 from several isolates showed very low genetic divergences between isolates from Vietnam and China, suggesting a common origin, and phylogenetic analysis confirmed the placement of SRBSDV as a distinct virus within subgroup 2 of the genus Fijivirus.

Ultrastructural alterations in midgut and Malpighian tubules of Boettcherisca peregrina exposure to cadmium and copper.

The effects of Cu and Cd at their at their low concentrations (80microg/g diet) on the morphology and ultrastructure of the midgut and Malpighian tubules of Boettcherisca peregrina larvae were observed by light and transmission electron microscopy. After exposure to both metals, the midgut got darker, shorter, and thicker than in control, and many strumae occurred on the surface of the midgut. Similarly, Malpighian tubules got shorter and thinner. Ultrastructural alterations in the midgut included mitochondrial condensation, swelling, and lysis. The rough endoplasmic reticulum (rER) showed dilation and vesiculation. The microvilli were shortened and disorganized. The stored glycogens increased and many mineral spherites appeared along with lipid droplets decreased. Ultrastructural alterations observed in the Malpighian tubules included rER vesiculation and mitochondria swelling with loss of cristae. Shortened and disordered microvilli, increased numbers of large hydropic vacuoles, and mineral spherites were also observed.

Organic Regime Promotes Evenness of Natural Enemies and Planthopper Control in Paddy Fields.

Planthoppers (Nilaparvata lugens, Sogatella furcifera, and Laodelphax striatellus) (Hemiptera: Delphacidae) are the most important pests affecting rice production. Pesticide spraying for its control may cause harmful effects on human health and the environment, especially the loss of biodiversity. The consequences of these changes on biodiversity and ecological services are well studied in tropical irrigated paddy fields, but are largely unknown in subtropical areas. Organic regime provides an environment-friendly method for biodiversity conservation; however, it is unclear whether this regime can suppress planthopper populations effectively in paddy fields. Consequently, we compared species richness, abundance, community structure, and evenness of natural enemies and planthoppers between organic and conventional rice fields (n = 35) distributed across four sites in China. The results showed that species richness was higher in organic fields than in conventional fields. Shannon index and evenness of predators and parasitoids were higher in most of the organic fields than their conventional counterparts. Furthermore, planthopper density showed a significant negative relationship with increased richness and evenness for both predators and parasitoids. These results underscore the notion that management regimes influence biodiversity in rice field. Most importantly, this has direct implications on the efficacy of natural pest control services rendered by predators and parasitoids associated with planthoppers in China and potentially other rice production regions in Asia.

Differential fipronil susceptibility and metabolism in two rice stem borers from China.

The susceptibilities of larvae of two rice stem borers, namely, Chilo suppressalis (Walker) (Lepidoptera: Crambidae) and Sesamia inferens (Walker) (Lepidoptera: Nocutidae) to fipronil and its metabolites were investigated, and then the activities of microsomal O-demethylase, and glutathione transferase (GST) in two species were measured. The metabolism of fipronil in both stem borers was determined in vivo and in vitro. The LD50 value of fipronil to S. inferens was 118.5-fold higher than that of C. suppressalis. The bioassay results offipronil metabolites showed that the toxicities of sulfone and sulfide were higher than fipronil for both species, and the differential toxicity between sulfone and fipronil was remarkable. Alternatively, the activities of microsomal O-demethylase and GST of C. suppressalis were 1.35- and 2.06-fold higher than S. inferens, respectively. The in vivo and in vitro studies on metabolism of fipronil showed that all of fipronil, sulfone, and sulfide were detected and the content of sulfone was higher than sulfide in both stem borers. The residue of sulfone in C. suppressalis was significantly higher than that in S. inferens. These results suggest that the higher activity of mixed function oxidases may cause the higher capacity of C. suppressalis to produce fipronil-sulfone, which is more toxic than fipronil leading to the higher susceptibility of this species.

Do rice water weevils and rice stem borers compete when sharing a host plant?

The rice water weevil (RWW) Lissorhoptrus oryzophilus Kuschel (Coleoptera: Curculionidae) is an invasive insect pest of rice Oryza sativa L. in China. Little is known about the interactions of this weevil with indigenous herbivores. In the present study, adult feeding and population density of the weevil, injury level of striped stem borer Chilo suppressalis (Walker) (Lepidoptera: Pyralidae) and pink stem borer Sesamia inferens (Walker) (Lepidoptera: Noctuidae) to rice, as well as growth status of their host plants were surveyed in a rice field located in Southeastern Zhejiang, China, in 2004 with the objective to discover interspecific interactions on the rice. At tillering stage, both adult feeding of the weevil and injury of the stem borers tended to occur on larger tillers (bearing 5 leaves) compared with small tillers (bearing 2~4 leaves), but the insects showed no evident competition with each other. At booting stage, the stem borers caused more withering/dead hearts and the weevil reached a higher density on the plants which had more productive tillers and larger root system; the number of weevils per tiller correlated negatively with the percentage of withering/dead hearts of plants in a hill. These observations indicate that interspecific interactions exist between the rice water weevil and the rice stem borers with negative relations occurring at booting or earlier developmental stages of rice.

Oogenesis in summer females of the rice water weevil, Lissorhoptrus oryzophilus Kuschel (Coleoptera: Curculionidae), in southern Zhejiang, China.

The rice water weevil, Lissorhoptrus oryzophilus Kuschel, has two generations in southern Zhejiang, China. To determine oogenesis in first-generation females (summer females) and its relations to temperature, females were collected from a rice field in early and mid-July and reared on young rice plants at 28, 31 and 34 degrees C in the laboratory. Percentage of females having oocytes, number of oocytes of different stages (stage-I, from early previtellogenesis to middle vitellogenesis; stage-II, late vitellogenesis; and mature-oocyte stage), and length of ovarioles were determined every 10 d of feeding. At each temperature, oogenesis took place in over 40% of females after 20~40 d of feeding, but only 0.0~3.3 stage-I, 0.0~0.8 stage-II and 0.0~1.1 mature oocytes were observed at each observation date. Temperature had significant effect on number of stage-I oocytes but not on number of stage-II and mature oocytes in early July females; temperature had no significant effect on number of oocytes of either stage in mid-July females. Conclusively, in southern Zhejiang, summer L. oryzophilus females have great potential to become reproductive on rice, but their oogenesis activity is very low, with the overall procedures little affected by temperature.

Spider (Araneae) predations on white-backed planthopper Sogatella furcifera in subtropical rice ecosystems, China.

BACKGROUND: Spiders are effective biological control agents in rice ecosystems, but the comparative study of predations among main spider species under field conditions has not been fully explored owing to a lack of practical methodology. In this study, more than 6000 spiders of dominant species were collected from subtropical rice ecosystems to compare their predations on Sogatella furcifera (Horváth) (white-backed planthopper, WBPH) using DNA-based gut content analysis. RESULTS: The positive rates for all spider taxa were closely related to prey densities, as well as their behaviors and niches. The relationships of positive rates to prey planthopper densities for Pardosa pseudoannulata (Böes. et Str.), Coleosoma octomaculata (Böes. et Str.), Tetragnatha maxillosa Thorell and Ummeliata insecticeps (Böes. et Str.) under field conditions could be described using saturated response curves. Quantitative comparisons of predations among the four spider species confirmed that P. pseudoannulata and C. octomaculata were more rapacious than U. insecticeps and T. maxillosa under field conditions. A comparison of ratio of spiders to WBPH and positive rates between fields revealed that biological control by spiders could be effectively integrated with variety resistance. CONCLUSION: Generalist spiders could follow up WBPH population timely, and assemblages of spiders coupled with variety resistance could effectively suppress WBPH population. © 2016 Society of Chemical Industry.

[Cloning and comparative analysis of the venom prepromelittin genes from four wasp species].

The total RNAs were isolated from the venom glands of female Polistes hebraeus, Vespula maculifrons, Vespa velutina nigrithorax and Vespa magnifica, respectively. Using the ReverAid First Strand cDNA Synthesis Kit, the first cDNAs were synthesized from the total RNA templates. PCR was carried out for 30 cycles (40s at 94 degrees C, 40s at 52 degrees C and 1 min at 72 degrees C), with a final extension of 10 min at 72 degrees C. RT-PCR products were examined by electrophoresis in 1.5% agarose gels with ethidium bromide staining. After purification using QIAquick PCR Purification Kit, the PCR products were ligated into pGEM*-T easy vector. Positive clones were identified by double digestion of Xho I and Hind III and PCR amplification and then were sequenced. The sequencing results showed that the amplified cDNAs containing the open reading frames of prepromelittin, and their lengths were all 213 bp. The ORFs were potential to encode polypeptides of 70 amino acid residues with predicted molecular weight of 7.7 kDa, including a signal peptide of 21 residues and a promelittin of 49 residues. Comparative analysis showed that the prepromelittins from different wasp species shared more than 93% identities in nucleotide sequences and more than 95% identities in amino acid sequences with each other, respectively. The sequences of prepromelittins of Polistes hebraeus, Vespula maculifrons, Vespa velutina nigrithorax and Vespa magnifica share 94%, 100%, 94%, 93% homologies in nucleotide sequences, and 95.8%, 100%, 97.2% and 97.2% identities in amino acid sequences with those of the European honey bee, Apis mellifera, respectively. There are 67 common amino acid residues among the prepromelittins of the four wasps and two honey bee species, Apis mellifera and Apis cerana. Phylogenetic relationships based on the prepromelitin nucleotide sequences were also examined using the software DNAStar5.01. The result showed that a closer relationship lied in between Polistes hebraeus, Vespa velutina nigrithorax and Vespa magnifica, while Vespula maculifrons had a closer relationship with the honey bees, Apis mellifera and Apis cerana. In conclusion, the prepromelittins were very conserved in the primary structure and the wasp insects also contain the melittins in their venoms, which are very similar to that of honey bee, although they belong to different superfamilies.

Cloning and characterization analysis of the genes encoding precursor of mast cell degranulating peptide from 2 honeybee and 3 wasp species.

The precursors of mast cell degranulating peptide (MCDP) genes were amplified by RT-PCR from the total RNA of venom gland of two honeybee species, Apis mellifera ligustica, Apis cerana cerana, and three wasp species, Vespa magnifica, Vespa velutina nigrothorax and Polistes hebraeus, respectively. Their PCR products were ligated into pGEM T-easy vector and the nucleotide sequences were analyzed. The length of five fragments was the same, it was 341 bp containing an ORF of 153 bp coding the precursor of MCDP and 188 bp 3' noncoding region. They have more than 90% homologues with each other in nucleotide sequences. The precursors of MCDP of A. cerana cerana, V. magnifica, V. velutina nigrothorax and P. hebraeus shared 96%, 100%, 94% and 98% homology with A. mellifera ligustica, respectively. The two species of wasps, V. magnifica and V. velutina nigrothorax, contained the same MCDP as A. mellifera ligustica, though they belong to different families with quite different biological properties, while A. cerana cerana contained the different MCDP in their venom as A. mellifera ligustica though they belong to the same genus. The fifth amino acid residue of MCDP in A. cerana cerana and P. hebraeus is arginine, replacing the cysteine, an important disulfide bridges element, in the position as in A. mellifera ligustica.

Heavy metal bioaccumulation and mobility from rice plants to Nilaparvata lugens (Homoptera: Delphacidae) in China.

Samples of soils, rice plants, and the adult, long-winged, brown planthoppers, Nilaparvata lugens (Stål) (Homoptera: Delphacidae), were collected from 18 sites of 9 regions in southern China. The concentrations of seven elements (Cu, Zn, As, Mo, Ag, Cd, and Pb) were measured using inductively coupled plasma mass spectrometry. Heavy metal mobility and bioaccumulation were analyzed in the rice plant-N. lugens system. The concentrations of Zn, As, Cd, and Pb in rice plants were positively correlated with their relevant concentrations in soil samples The bioconcentration factors of the seven elements in the rice plant-N. lugens system showed that the order of metal accumulation was Mo>Zn>Ag>Cd>Cu>Pb>As. In particular, Mo and Zn showed significantly high accumulation in N. lugens. A cluster analysis and factor analysis showed that the bioaccumulation of these seven elements in the rice plant-N. lugens system could be classified into two groups, closely related to their molar mass. The first group consisted of five elements with relatively light molar masses: Cu, Zn, As, Mo, and Ag. Cu and Zn, which have nearly equal molar masses, showed similar accumulation levels in N. lugens. The second group included two elements with relatively heavy molar masses: Cd and Pb. This study demonstrated that bioaccumulation of seven heavy metals was regular in the rice plant-N. lugens system. N. lugens could be used as bioindicators of the contaminated degree for Zn in rice paddy fields. This information may provide a basis for future ecological research on the bioaccumulation mechanism in N. lugens.

Identification and characterization of microRNAs in small brown planthopper (Laodephax striatellus) by next-generation sequencing.

MicroRNAs (miRNAs) are endogenous non-coding small RNAs that regulate gene expression at the post-transcriptional level and are thought to play critical roles in many metabolic activities in eukaryotes. The small brown planthopper (Laodephax striatellus Fallén), one of the most destructive agricultural pests, causes great damage to crops including rice, wheat, and maize. However, information about the genome of L. striatellus is limited. In this study, a small RNA library was constructed from a mixed L. striatellus population and sequenced by Solexa sequencing technology. A total of 501 mature miRNAs were identified, including 227 conserved and 274 novel miRNAs belonging to 125 and 250 families, respectively. Sixty-nine conserved miRNAs that are included in 38 families are predicted to have an RNA secondary structure typically found in miRNAs. Many miRNAs were validated by stem-loop RT-PCR. Comparison with the miRNAs in 84 animal species from miRBase showed that the conserved miRNA families we identified are highly conserved in the Arthropoda phylum. Furthermore, miRanda predicted 2701 target genes for 378 miRNAs, which could be categorized into 52 functional groups annotated by gene ontology. The function of miRNA target genes was found to be very similar between conserved and novel miRNAs. This study of miRNAs in L. striatellus will provide new information and enhance the understanding of the role of miRNAs in the regulation of L. striatellus metabolism and development.

Genomes of the rice pest brown planthopper and its endosymbionts reveal complex complementary contributions for host adaptation.

BACKGROUND: The brown planthopper, Nilaparvata lugens, the most destructive pest of rice, is a typical monophagous herbivore that feeds exclusively on rice sap, which migrates over long distances. Outbreaks of it have re-occurred approximately every three years in Asia. It has also been used as a model system for ecological studies and for developing effective pest management. To better understand how a monophagous sap-sucking arthropod herbivore has adapted to its exclusive host selection and to provide insights to improve pest control, we analyzed the genomes of the brown planthopper and its two endosymbionts. RESULTS: We describe the 1.14 gigabase planthopper draft genome and the genomes of two microbial endosymbionts that permit the planthopper to forage exclusively on rice fields. Only 40.8% of the 27,571 identified Nilaparvata protein coding genes have detectable shared homology with the proteomes of the other 14 arthropods included in this study, reflecting large-scale gene losses including in evolutionarily conserved gene families and biochemical pathways. These unique genomic features are functionally associated with the animal's exclusive plant host selection. Genes missing from the insect in conserved biochemical pathways that are essential for its survival on the nutritionally imbalanced sap diet are present in the genomes of its microbial endosymbionts, which have evolved to complement the mutualistic nutritional needs of the host. CONCLUSIONS: Our study reveals a series of complex adaptations of the brown planthopper involving a variety of biological processes, that result in its highly destructive impact on the exclusive host rice. All these findings highlight potential directions for effective pest control of the planthopper.

Development and preliminary application of a triplex real-time polymerase chain reaction assay for evaluating predation on three planthoppers in a rice ecosystem.

A multiplex real-time quantitative polymerase chain reaction (PCR) assay was developed to simultaneously detect the DNA of three rice planthoppers, that is, Sogatella furcifera (Horváth) (white-backed planthopper), Nilaparvata lugens (Stål) (brown planthopper) and Laodelphax striatellus (Fallén) (small brown planthopper), in the gut of their predators. The sets of primers and ALLGlo probes were targeted to the regions of internal transcribed spacer 2 (ITS2) genes in nuclear ribosomal DNA (rDNA). The sensitivity, specificity and interference test for the multiplex real-time quantitative PCR assay were analysed. The assay's detection limits were 100, 1000 and 100 copies for the white-backed planthopper, the brown planthopper and the small brown planthopper, respectively. The specificity tests showed no cross-reactivity with genomic DNA from 30 other dominant herbivores, saprophagous insects and predators from rice ecosystem for each planthopper species. The assay was used in a preliminary study of predation events on the three planthoppers by three major spiders viz., Pardosa pseudoannulata (Bösenberg et Strand), Ummeliata insecticeps (Bösenberg et Strand) and Tetragnatha maxillosa Thorell which each differ in their preferred microhabitat as well as their predatory habits in rice field, and the results showed their predation on each planthopper species could be well evaluated using this method. Therefore, the multiplex real-time quantitative PCR assay provides a new tool to study the mechanisms of prey shifting and natural regulation of the three rice planthoppers by generalist predators in rice ecosystem.

Feeding-induced interactions between two rice planthoppers, Nilaparvata lugens and Sogatella furcifera (Hemiptera: Delphacidae): effects on feeding and honeydew excretion.

A series of laboratory experiments was conducted using electrical penetration graph, salivary flange, and honeydew measurement to study the effects of feeding-induced intra- and interspecific interactions on feeding behavior and honeydew excretion between planthoppers Nilaparvata lugens (Stål) and Sogatela furcifera (Horváth). Feeding-induced intra- and interspecific interactions affected many measures of feeding behavior. The number of salivary flanges, mean duration of pathway activities per insect, and mean duration from first probe to first sustained phloem ingestion for both N. lugens and S. furcifera were significantly shorter on rice plants with feeding-induced conspecific and heterospecific effects than those for planthoppers fed on control plants. Feeding-induced intra- and interspecific interactions also affected the duration per insect of phloem ingestion for both N. lugens and S. furcifera. The durations per insect of phloem ingestion on host plants with feeding-induced conspecific and heterospecific effects were significantly longer than those on control plants. An asymmetric facilitative effect of induced interspecific interactions on the weight of honeydew excreted was detected, because only the honeydew weights of S. furcifera were significantly increased by the induced heterospecific effect on both varieties. The results demonstrated that the facilitative effects on honeydew excretion were consistent with previously documented effects on performance. Both facilitative effects on honeydew excretion and performance were asymmetrical, with more benefits to S. furcifera from N. lugens. Such facilitative effects might be mainly related to altered nutrient status and induced allelochemistry in rice.

Feeding-induced interactions between Nilaparvata lugens and Laodelphax striatellus (Hemiptera: Delphacidae): effects on feeding behavior and honeydew excretion.

Using electrical penetration graph, salivary flange, and honeydew measurement, this study investigated the effects of feeding-induced intra- and interspecific interactions on feeding behavior and honeydew excretion of brown planthopper (Nilaparvata lugens) compared with small brown planthopper (Laodelphax striatellus). Results showed that many measures of feeding behavior were affected by feeding-induced intra- and interspecific interactions on two different rice varieties. There were significantly fewer salivary flanges for both brown planthopper and small brown planthopper on rice plants with feeding-induced conspecific or heterospecific effects than on relevant control plants. In contrast, only small brown planthopper on rice plants with feeding-induced heterospecific effects had significantly fewer salivary flanges than those with feeding-induced conspecific effects. The mean durations of pathway activities per insect and mean durations from first probe to first sustained phloem ingestion for small brown planthopper were significantly shorter, whereas the mean duration per insect of phloem ingestion was significantly longer, on rice plants with feeding-induced heterospecific effects than those on relevant control plants, as well as rice plants with feeding-induced conspecific effects. Honeydew weights of small brown planthopper were significantly increased by the induced heterospecific effect. Thus, all results indicated indirect, asymmetrical, facilitative effects of induced interspecific interactions on the feeding behavior and honeydew weight for small brown planthopper on both varieties. These findings are consistent with the previously documented asymmetrical effects on performance, with more benefits to small brown planthopper from brown planthopper indirectly. The change of nutrient and induced allelochemistry in host plant probably underlies these facilitative effects.

Molecular characterization of the flightin gene in the wing-dimorphic planthopper, Nilaparvata lugens, and its evolution in Pancrustacea.

Flightin was initially identified in Drosophila melanogaster. Previous work has shown that Drosophila flightin plays a key role in indirect flight muscle (IFM) function and has limited expression in the IFM. In this study, we demonstrated that flightin is conserved across the Pancrustacea species, including winged insects, non-winged insects, non-insect hexapods and several crustaceans. The brown planthopper (BPH), Nilaparvata lugens (Stål) (Hemiptera: Delphacidae), a long-distance migration insect with wing dimorphism, is the most destructive rice pest in Asia. We showed that flightin was one of the most differentially expressed genes in macropterous and brachypterous BPH adults. In female BPHs, flightin was expressed in the IFM of macropterous adults, no expression was detected in brachypterous ones; while in male BPHs, flightin was not only expressed in the IFM of macropterous adults, but also in the dorsal longitudinal muscle (DLM) in the basal two abdominal segments of both macropterous and brachypterous ones. RNAi and transmission electron microscopy results showed that flightin played key roles in maintaining IFM and male DLM structure, which drive wing movements in macropterous adults and the vibration of the male-specific tymbal, respectively. Using Daphnia magna as an example of a crustacean species, we observed that flightin was expressed in juvenile instars and adults, and was localized in the antenna muscles. These results illustrate the functional variations of flightin in insects and other arthropod species and provide clues as to how insects with flight apparatuses evolved from ancient pancrustaceans.