Rice volatile compound (E)-ß-caryophyllene induced by rice dwarf virus (RDV) attracts the natural enemy Cyrtorhinus lividipennis to prey on RDV insect vectors.

BACKGROUND: Rice dwarf virus (RDV)-induced rice plant volatiles (E)-ß-caryophyllene and 2-heptanol modulate the olfactory behavior of RDV insect vectors that promote viral acquisition and transmission. However, it remains elusive whether these two volatiles could influence the behaviors of the natural enemies of RDV insect vectors. Herein, we determined the effects of these two volatiles on the olfactory and predatory behaviors of Cyrtorhinus lividipennis (Hemiptera: Miridae), an important predator of RDV insect vectors in rice paddies. RESULTS: The results showed that C. lividipennis preferred RDV-infected rice plant odors over RDV-free rice plant odors. C. lividipennis was attracted by (E)-ß-caryophyllene, but showed no behavioral responses to 2-heptanol. The attraction of (E)-ß-caryophyllene towards C. lividipennis was further confirmed using oscas1 rice plants, which do not release (E)-ß-caryophyllene in response to RDV infection, through a series of complementary assays. The oviposition preference of the RDV vector insect Nephotettix cincticeps (Hemiptera: Cicadellidae) showed no significant difference between RDV-infected and RDV-free wild-type plants, nor between oscas1-RDV and oscas1 plants. However, the predation rate of C. lividipennis for N. cincticeps eggs on RDV-infected plants was higher than that on RDV-free plants, whereas there was no significant difference between oscas1-RDV and oscas1 plants. CONCLUSION: (E)-ß-caryophyllene induced by RDV attracted more C. lividipennis to prey on N. cincticeps eggs and played a crucial role in plant-virus-vector-enemy interactions. These novel findings will promote the design of new strategies for disease control by controlling the populations of insect vectors, for example recruiting more natural enemies by virus-induced plant volatiles. © 2023 Society of Chemical Industry.

Olfactory behavioral responses of two Drosophila species and their pupal parasitoid to volatiles from bananas.

BACKGROUND: Behavior-based manipulation is an essential part of Drosophila integrated pest management (IPM). Effective compounds are useful for improving the efficiency of baits and the development of attract-and-kill or push-pull strategies to manage Drosophila populations. Here, we investigated the olfactory behavior of two Drosophila species, as well as their pupal parasitoid, to volatiles from bananas, for the identification of effective compounds to control fly populations. RESULTS: The results showed that overripe bananas were most attractive to both flies, with a higher attraction index (AI) in Drosophila melanogaster than in Drosophila suzukii. The profiles of volatile organic compounds (VOCs) from bananas with three different ripening stages were compared, and six VOCs were selected for behavioral tests. D. suzukii showed significantly different responses to isoamyl alcohol with D. melanogaster. The effects of ethyl butyrate and ethyl isovalerate on the two flies were dose-dependent, with lower concentrations acting as repellent and higher concentrations acting as attractant. Isoamyl acetate, isoamyl butyrate and isoamyl isovalerate (0.005-0.05% v/v) were attractive to both flies. The parasitoid wasp Pachycrepoidus vindemmiae was repelled by isoamyl alcohol and ethyl butyrate, and attracted by ethyl isovalerate, but showed no significant response to isoamyl acetate, isoamyl butyrate and isoamyl isovalerate at the tested concentration. CONCLUSION: This study indicated that the behavioral response of Drosophila to the odor of chemical compounds is dose-dependent. Isoamyl alcohol and isoamyl acetate at certain concentrations could be used as repellents, whereas ethyl isovalerate as an attractant in control strategy design for D. suzukii. © 2023 Society of Chemical Industry.

Virus-Induced Plant Volatiles Promote Virus Acquisition and Transmission by Insect Vectors.

Rice dwarf virus (RDV) is transmitted by insect vectors Nephotettix virescens and Nephotettix cincticeps (Hemiptera: Cicadellidae) that threatens rice yield and results in substantial economic losses. RDV induces two volatiles ((E)-ß-caryophyllene (EBC) and 2-heptanol) to emit from RDV-infected rice plants. However, the effects of the two volatiles on the olfactory behavior of both non-viruliferous and viruliferous N. virescens are unknown, and whether the two volatiles could facilitate the spread and dispersal of RDV remains elusive. Combining the methods of insect behavior, chemical ecology, and molecular biology, we found that EBC and 2-heptanol influenced the olfactory behavior of non-viruliferous and viruliferous N. virescens, independently. EBC attracted non-viruliferous N. virescens towards RDV-infected rice plants, promoting virus acquisition by non-viruliferous vectors. The effect was confirmed by using oscas1 mutant rice plants (repressed EBC synthesis), but EBC had no effects on viruliferous N. virescens. 2-heptanol did not attract or repel non-viruliferous N. virescens. However, spraying experiments showed that 2-heptanol repelled viruliferous N. virescens to prefer RDV-free rice plants, which would be conducive to the transmission of the virus. These novel results reveal that rice plant volatiles modify the behavior of N. virescens vectors to promote RDV acquisition and transmission. They will provide new insights into virus-vector-plant interactions, and promote the development of new prevention and control strategies for disease management.

Identification and Expression Analysis of Odorant-Binding and Chemosensory Protein Genes in Virus Vector Nephotettix cincticeps.

The insect odorant binding proteins (OBPs) and chemosensory proteins (CSPs) are involved in the perception and discrimination of insects to host odor cues. Nephotettix cincticeps, one of the destructive pests of rice plants, not only directly damages hosts by sucking, but also indirectly transmits plant viruses in the field. Previous study found that two rice volatiles ((E)-ß-caryophyllene and 2-heptanol) induced by rice dwarf virus (RDV) mediated the olfactory behavior of N. cincticeps, which may promote virus dispersal. However, the OBPs and CSPs in N. cincticeps are still unknown. In this study, to identify the OBP and CSP genes in N. cincticeps, transcriptomic analyses were performed. In total, 46,623 unigenes were obtained. Twenty putative OBP and 13 CSP genes were discovered and identified. Phylogenetic analyses revealed that five putative OBPs belonged to the plus-C OBP family, and the other classic OBPs and CSPs were distributed among other orthologous groups. A total of 12 OBP and 10 CSP genes were detected, and nine OBP and three CSP genes were highly expressed in N. cincticeps antennae compared with other tissues. This study, for the first time, provides a valuable resource to well understand the molecular mechanism of N. cincticeps in the perception and discrimination of the two volatiles induced by RDV infection.

Identification and Characterization of Resistance of Three Aphid Species on Contrasting Alfalfa Cultivars.

Aphids on alfalfa (Medicago sativa) including Aphis craccivora Koch (cowpea aphid, CPA), Acyrthosiphon pisum Harris (pea aphid, PA) and Therioaphis trifolii Buckton (spotted alfalfa aphid, SAA) cause significant yield losses worldwide. In this experiment, the development of these three species of aphids on 16 alfalfa cultivars was compared. The results showed that the plant cultivar had a significant influence on the development of aphids as there are significant differences in the body weight of aphids reared on different alfalfa cultivars. In addition, antibiosis between the alfalfa cultivars Pegasis and Gannong NO.9 and the three species of aphids was evaluated by measuring aphid body weight and fecundity. Antixenosis was measured using choice tests, and feeding behavior was quantified using electrical penetration graphs (EPG). The Pegasis cultivar was observed to have both antibiosis and antixenosis effects with CPA, but was susceptible to PA and SAA compared with the Gannong NO.9 cultivar. CPA had less mean body weight, less fecundity, and shorter feeding time on the Pegasis cultivar, and preferred to settle on Gannong NO.9 cultivar. In contrast, Gannong NO.9 exhibits antibiosis and antixenosis to PA and SAA compared with Pegasis, as shown by lower body weight, lower fecundity and chose to settle less often, but EPG data showed that PA and SAA showed no different significance in feeding behavior between Pegasis and Gannong NO.9.

Anti-photoaging effects of flexible nanoliposomes encapsulated Moringa oleifera Lam. isothiocyanate in UVB-induced cell damage in HaCaT cells.

Skin photoaging is premature skin aging damage that occurs after repeated exposure to ultraviolet (UV) radiation. Although isothiocyanates extracted from the moringa tree (Moringa oleifera Lam.) (MITC) exhibit excellent effects against skin photoaging, its application is restricted because of its characteristics, such as extremely low water solubility, bioavailability, and easy degradation. Currently, flexible nanoliposomes have gained increasing interest as a biocompatible polymer for applications such as transdermal drug delivery. We prepare amphiphilic hyaluronic acid (HA) conjugated with ceramide (CE) to modify nanoliposomes for MITC (HACE/MITC NPs) delivery. The HACE/MITC nanoparticles (NPs) are prepared and characterized for entrapment efficiency, particle size, polydispersity index, zeta potential, in vitro release, in vivo skin permeation, and in vitro protective effect of photoaging. The zeta potential of MITC NPs and HACE/MITC NPs is -24.46 mV and -24.93 mV, respectively. After modification of HACE, the entrapment efficient of MITC liposome increased from 62.54% to 70.67%, and the particle size decreased from 266.1 nm to 192.8 nm. In vivo skin permeation, permeated drug increased from 49.42 to 71.40%. Moreover, the results showed that the entrapment of MITC in nanoliposomes improves its stability, efficacy, and skin permeation. Further, HACE/MITC NPs are favorable for uptake by HaCaT cells without requiring changes in cell morphology, which significantly improves the activities of antioxidant enzymes, scavenges UVB-induced reactive oxygen species, protects skin from damage, and reduces MMP-1, MMP-3, and MMP-9 expression caused by radiation-induced photoaging. Our results strongly suggest that flexible nanoliposomes successfully improved the cell membrane permeation of MITC, and that anti-photoaging and HACE/MITC NPs can potentially be used as candidates for photoaging therapy.

Virus-induced plant volatiles mediate the olfactory behaviour of its insect vectors.

Plant viruses can manipulate their hosts to release odours that are attractive or repellent to their insect vectors. However, the volatile organic compounds (VOCs), either individually or as mixtures, which play a key role in the olfactory behaviour of insect vectors remains largely unknown. Our study focused on green rice leafhoppers (GRLHs) vectoring rice dwarf virus (RDV) revealed that RDV infection significantly induced the emission of (E)-ß-caryophyllene and 2-heptanol by rice plants, which influenced the olfactory behaviour of both non-viruliferous and viruliferous GRLHs. (E)-ß-caryophyllene attracted non-viruliferous GRLHs to settle on RDV-infected plants, but neither attracted nor repelled viruliferous GRLHs. In contrast, 2-heptanol repelled viruliferous GRLHs to settle on RDV-infected plants, but neither repelled nor attracted non-viruliferous GRLHs. Suppression of (E)-ß-caryophyllene synthase OsCAS via CRISPR-Cas9 to generate oscas-1 plants enabled us to confirm the important role played by (E)-ß-caryophyllene in modulating the virus-vector-host plant interaction. These novel results reveal the role of these virus-induced VOCs in modulating the behaviour of its GRLH insect vector and may facilitate the design of new strategies for disease control through manipulation of plant volatile emissions.

Metabolic Analysis Reveals Cry1C Gene Transformation Does Not Affect the Sensitivity of Rice to Rice Dwarf Virus.

Metabolomics is beginning to be used for assessing unintended changes in genetically modified (GM) crops. To investigate whether Cry1C gene transformation would induce metabolic changes in rice plants, and whether the metabolic changes would pose potential risks when Cry1C rice plants are exposed to rice dwarf virus (RDV), the metabolic profiles of Cry1C rice T1C-19 and its non-Bt parental rice MH63 under RDV-free and RDV-infected status were analyzed using gas chromatography-mass spectrometry (GC-MS). Compared to MH63 rice, slice difference was detected in T1C-19 under RDV-free conditions (less than 3%), while much more metabolites showed significant response to RDV infection in T1C-19 (15.6%) and in MH63 (5.0%). Pathway analysis showed biosynthesis of lysine, valine, leucine, and isoleucine may be affected by RDV infection in T1C-19. No significant difference in the contents of free amino acids (AAs) was found between T1C-19 and MH63 rice, and the free AA contents of the two rice plants showed similar responses to RDV infection. Furthermore, no significant differences of the RDV infection rates between T1C-19 and MH63 were detected. Our results showed the Cry1C gene transformation did not affect the sensitivity of rice to RDV, indicating Cry1C rice would not aggravate the epidemic and dispersal of RDV.

Cry1C rice doesn't affect the ecological fitness of rice brown planthopper, Nilaparvata lugens either under RDV stress or not.

The potential risks of Bt rice on non-target arthropods (NTAs) should be evaluated and defined before commercial production. Recently, effects of Bt rice on NTAs under abiotic and biotic stress conditions attracted much attention. Here we reported the effects of Bt rice T1C-19 (Cry1C rice) on the non-target herbivore, Nilaparvata lugens (rice brown planthopper, BPH) with or without RDV (rice dwarf virus) infection conditions. BPH showed no feeding and oviposition preference between Bt rice T1C-19 and its non-Bt parental rice Minghui 63 (MH63), as well as between RDV-infected and RDV-free rice plants. Meanwhile, rice type, RDV infection status, and their interaction had little impacts on the survival, development and fecundity of BPH. By comparison with non-Bt control, Bt rice T1C-19 with or without RDV infection had no significant effects on the life-table parameters of BPH including rm, R0, T, DT and λ. Thus, it could be concluded that Bt rice T1C-19 doesn't affect the ecological fitness of BPH either under RDV stress or not.

Molecular and pharmacological characterization of a ß-adrenergic-like octopamine receptor from the green rice leafhopper Nephotettix cincticeps.

As the counterparts of noradrenaline and adrenaline in vertebrates, octopamine (OA) regulates multiple physiological and behavioral processes in invertebrate. OA mediates its effects via binding to specific octopamine receptors (OARs). Functional and pharmacological characterization of OARs have been reported in several insects. However, little work was documented in hemipteran insects. We cloned a ß-adrenergic-like OAR (NcOA2B2) from Nephotettix cincticeps. NcOA2B2 shares high similarity with members of the OA2B2 receptor class. Transcript level of NcOA2B2 varied in various tissues and was highly expressed in the leg. After heterologous expression in CHO-K1 cells, NcOA2B2 was dose-dependently activated by OA (EC50 = 2.56 nM) and tyramine (TA) (EC50 = 149 nM). Besides putative octopaminergic agonists, dopaminergic agonists and amitraz and DPMF potently activated NcOA2B2 in a dose-dependent manner. Receptor activity was blocked by potential antagonists and was most efficiently antagonized by asenapine. Phentolamine showed both antagonist and agonist effects on NcOA2B2. Our results offer the important information about molecular and pharmacological characterization of an OAR from N. cincticeps that will provide the basis for forthcoming studies on its roles in physiological processes and behaviors, and facilitate the design of novel insecticides for pest control.

A Novel Iflavirus Was Discovered in Green Rice Leafhopper Nephotettix cincticeps and Its Proliferation Was Inhibited by Infection of Rice Dwarf Virus.

The green rice leafhopper, Nephotettix cincticeps (Hemiptera: Cicadellidae), is a key insect vector transmitting rice dwarf virus (RDV) that causes rice dwarf disease. We discovered a novel iflavirus from the transcriptomes of N. cincticeps and named it as Nephotettix cincticeps positive-stranded RNA virus-1 (NcPSRV-1). The viral genome consists of 10,524 nucleotides excluding the poly(A) tail and contains one predicted open reading frame encoding a polyprotein of 3,192 amino acids, flanked by 5' and 3' untranslated regions. NcPSRV-1 has a typical iflavirus genome arrangement and is clustered with the members of the family Iflaviridae in the phylogenetic analysis. NcPSRV-1 was detected in all tested tissues and life stages of N. cincticeps and could be transmitted horizontally and vertically. Moreover, NcPSRV-1 had high prevalence in the laboratory populations and was widely spread in field populations of N. cincticeps. NcPSRV-1 could also infect the two-striped leafhopper, Nephotettix apicalis, at a 3.33% infection rate, but was absent in the zigzag leafhopper, Recilia dorsalis, and rice Oryza sativa variety TN1. The infection of RDV altered the viral load and infection rate of NcPSRV-1 in N. cincticeps, for which it seems that RDV has an antagonistic effect on NcPSRV-1 infection in the host.

Rice dwarf virus infection alters green rice leafhopper host preference and feeding behavior.

Host plants, pathogens and their herbivore vectors systems have complex relationships via direct and indirect interactions. Although there are substantial gaps in understanding these systems, the dynamics of the relationships may influence the processes of virus transmission and plant disease epidemics. Rice dwarf virus (RDV) is mainly vectored by green rice leafhoppers (GRLHs), Nephotettix cincticeps (Uhler) (Hemiptera: Cicadellidae) in a persistently circulative manner. In this study, host plant selection preferences of non-viruliferous and viruliferous (carrying RDV) GRLHs between RDV-free and RDV-infected plants were tested. Non-viruliferous GRLHs preferred RDV-infected rice plants over RDV-free rice plants, and viruliferous GRLHs preferred RDV-free rice plants over RDV-infected rice plants. In odor selection preference bioassay using a four-field olfactometer, non-viruliferous GRLHs preferred odors of RDV-infected rice plants over healthy rice and viruliferous GRLHs preferred odors of RDV-free rice plants over RDV-infected ones. In 6 h plant penetration behavior bioassay using electrical penetration graphs, non-viruliferous GRLHs spent shorter time in non-penetration and much longer time in xylem feeding on RDV-infected, compared to RDV-free rice plants. Viruliferous GRLHs exhibited more salivation and stylet movement on RDV-free rice plants than on RDV-infected rice plants. We infer from these findings that RDV influences these vector behaviors by altering host plant physiology to promote viral transmission.

Variation among conventional cultivars could be used as a criterion for environmental safety assessment of Bt rice on nontarget arthropods.

The current difficulty facing risk evaluations of Bacillus thuringiensis (Bt) crops on nontarget arthropods (NTAs) is the lack of criteria for determining what represents unacceptable risk. In this study, we investigated the biological parameters in the laboratory and field population abundance of Nilaparvata lugens (Hemiptera: Delphacidae) on two Bt rice lines and the non-Bt parent, together with 14 other conventional rice cultivars. Significant difference were found in nymphal duration and fecundity of N. lugens fed on Bt rice KMD2, as well as field population density on 12 October, compared with non-Bt parent. However, compared with the variation among conventional rice cultivars, the variation of each parameter between Bt rice and the non-Bt parent was much smaller, which can be easily seen from low-high bar graphs and also the coefficient of variation value (C.V). The variation among conventional cultivars is proposed to be used as a criterion for the safety assessment of Bt rice on NTAs, particularly when statistically significant differences in several parameters are found between Bt rice and its non-Bt parent. Coefficient of variation is suggested as a promising parameter for ecological risk judgement of IRGM rice on NTAs.

Does Bt rice pose risks to non-target arthropods? Results of a meta-analysis in China.

Transgenic Bt rice expressing the insecticidal proteins derived from Bacillus thuringiensis Berliner (Bt) has been developed since 1989. Their ecological risks towards non-target organisms have been investigated; however, these studies were conducted individually, yielding uncertainty regarding potential agroecological risks associated with large-scale deployment of Bt rice lines. Here, we developed a meta-analysis of the existing literature to synthesize current knowledge of the impacts of Bt rice on functional arthropod guilds, including herbivores, predators, parasitoids and detritivores in laboratory and field studies. Laboratory results indicate Bt rice did not influence survival rate and developmental duration of herbivores, although exposure to Bt rice led to reduced egg laying, which correctly predicted their reduced abundance in Bt rice agroecosystems. Similarly, consuming prey exposed to Bt protein did not influence survival, development or fecundity of predators, indicating constant abundances of predators in Bt rice fields. Compared to control agroecosystems, parasitoid populations decreased slightly in Bt rice cropping systems, while detritivores increased. We draw two inferences. One, laboratory studies of Bt rice showing effects on ecological functional groups are mainly either consistent with or more conservative than results of field studies, and two, Bt rice will pose negligible risks to the non-target functional guilds in future large-scale Bt rice agroecosystems in China.