Functional types of long trichoid sensilla responding to sex pheromone components in Plutella xylostella.

Sex pheromones, which consist of multiple components in specific ratios promote intraspecific sexual communications of insects. Plutella xylostella (L.) is a worldwide pest of cruciferous vegetables, the mating behavior of which is highly dependent on its olfactory system. Long trichoid sensilla on male antennae are the main olfactory sensilla that can sense sex pheromones. However, the underlying mechanisms remain unclear. In this study, 3 sex pheromone components from sex pheromone gland secretions of P. xylostella female adults were identified as Z11-16:Ald, Z11-16:Ac, and Z11-16:OH in a ratio of 9.4 : 100 : 17 using gas chromatography - mass spectrometry and gas chromatography with electroantennographic detection. Electrophysiological responses of 581 and 385 long trichoid sensilla of male adults and female adults, respectively, to the 3 components were measured by single sensillum recording. Hierarchical clustering analysis showed that the long trichoid sensilla were of 6 different types. In the male antennae, 52.32%, 5.51%, and 1.89% of the sensilla responded to Z11-16:Ald, Z11-16:Ac, and Z11-16:OH, which are named as A type, B type, and C type sensilla, respectively; 2.93% named as D type sensilla responded to both Z11-16:Ald and Z11-16:Ac, and 0.34% named as E type sensilla were sensitive to both Z11-16:Ald and Z11-16:OH. In the female antennae, only 7.53% of long trichoid sensilla responded to the sex pheromone components, A type sensilla were 3.64%, B type and C type sensilla were both 0.52%, D type sensilla were 1.30%, and 1.56% of the sensilla responded to all 3 components, which were named as F type sensilla. The responding long trichoid sensilla were located from the base to the terminal of the male antennae and from the base to the middle of the female antennae. The pheromone mixture (Z11-16:Ald : Z11-16:Ac : Z11-16:OH = 9.4 : 100 : 17) had a weakly repellent effect on female adults of P. xylostella. Our results lay the foundation for further studies on sex pheromone communications in P. xylostella.

Identification, expression profiles, and binding properties of chemosensory protein 18 in Plutella xylostella (Lepidoptera: Plutellidae).

Chemosensory proteins (CSPs) are highly efficient carry tools to bind and deliver hydrophobic compounds, which play an important role in the chemosensory process in insects. The diamondback moth, Plutella xylostella L. (Lepidoptera: Plutellidae), is a cosmopolitan pest that attacks cruciferous crops. However, the detailed physiological functions of CSPs in P. xylostella remain limited to date. Here, we identified a typical CSP, named PxylCSP18, in P. xylostella and investigated its expression patterns and binding properties of volatiles. PxylCSP18 was highly expressed in antennae and head (without antennae), and the expression level in the male antennae of P. xylostella was obviously higher than that in the female antennae. Moreover, PxylCSP18 has a relatively broad binding spectrum. Fluorescence competitive binding assays showed that PxylCSP18 had strong binding abilities with 14 plant volatiles (Ki < 10 µM) that were repellent or attractive to P. xylostella. Notably, PxylCSP18 had no significant binding affinity to (Z)-11-hexadecenal, (Z)-11-hexadecenyl acetate, and (Z)-11-hexadecenyl alcolol, which are the pheromone components of P. xylostella. The attractive effects of trans-2-hexen-1-ol and isopropyl isothiocyanate to male adults and the attractive effects of isopropyl isothiocyanate and the repellent effects of linalool to female adults were significantly decreased after knocked down the expression of PxylCSP18. Our results revealed that PxylCSP18 might play an important role in host plant detection, avoidance of unsuitable hosts, and selection of oviposition sites; however, it does not participate in mating behavior. Overall, these results extended our knowledge on the CSP-related functions, which provided insightful information about CSP-targeted insecticides.

Electrophysiological and Behavioral Responses of Plodia interpunctella (Hübner) Females to Aldehyde Volatiles from Dried Fruits.

Plodia interpunctella (Lepidoptera: Pyraloidea) is a notorious pest of stored grain globally. The dried fruits (Ziziphus jujuba, Malus pumila, and Fragaria ananassa) can strongly attract P. interpunctella. However, specific volatile compounds responsible for such effects have not been identified. Volatiles were analyzed by using headspace solid-phase microextraction (HS-SPME) and chromatography-mass spectrometry (GCMS) techniques. Five aldehyde compounds were selected for electroantennogram (EAG), single sensillum recording (SSR), and behavioral response assays. The three chemicals that elicited the strongest EAG responses to mated females at 100 µg/µL include hexanal (1.13 mV), heptanal (0.92 mV), and octanal (0.73 mV). In SSR experiments, the basiconic sensilla of the antennae responded to these aldehyde compounds. The results of behavioral responses showed that all aldehydes exhibited dose-dependent responses, with hexanal having the highest attractant rate of 74.56%. These compounds have the potential to be used for monitoring P. interpunctella and its integrated management program.

Identification and evaluation of cruciferous plant volatiles attractive to Plutella xylostella L. (Lepidoptera: Plutellidae).

BACKGROUND: The diamondback moth, Plutella xylostella, has developed resistance to almost all insecticides used for its control. The 'push-pull' method has been shown as an effective control strategy to address this resistance challenge of P. xylostella. The key focus of the strategy is the identification of attractive or repellent volatile components. The aim of this study was to identify attractive volatile compounds released from host plants. Identified compounds were applied in the biological control of this pest. RESULTS: Nine active compounds released into the headspace of seven cruciferous plant species were identified using gas chromatography-electroantennographic detection and gas chromatography-mass spectrometry. Electroantennographic detection-active compounds included five green leaf volatiles (hexanal, trans-2-hexen-1-ol, cis-3-hexen-1-ol, cis-3-hexenyl acetate, and 1-penten-3-ol), three isothiocyanates (isopropyl isothiocyanate, allyl isothiocyanate, and butyl isothiocyanate), and nonanal. Except for nonanal, all the identified green leaf volatiles and isothiocyanates elicited strong electrophysiological and behavioral responses in P. xylostella. The strongest attractive compounds, trans-2-hexen-1-ol and isopropyl isothiocyanate, were further evaluated in oviposition and field-trapping assays. Results showed that they both lured female moths to lay eggs, and were highly attractive to P. xylostella adults in field, especially when used in combination with yellow and green sticky boards. However, a blend of the two compounds showed no synergistic effect, but rather an antagonistic effect. CONCLUSIONS: Green leaf volatiles and isothiocyanates were identified as key olfactory cues for host selection of P. xylostella. Trans-2- hexen-1-ol and isopropyl isothiocyanate were identified as candidate attractive compounds to serve in a 'push-pull' strategy for P. xylostella control. © 2023 Society of Chemical Industry.

Behavioral, Electrophysiological, and Toxicological Responses of Plutella xylostella to Extracts from Angelica pubescens.

Plutella xylostella L. is a destructive pest affecting cruciferous vegetables, causing massive economic losses worldwide. Plant-based insecticides are considered promising insect control agents. The Angelica pubescens extract inhibited female oviposition, with an oviposition deterrence index (ODI) of 61.65% at 12.5 mg/mL. We aimed to identify the bioactive compounds in A. pubescens extract. The compounds from A. pubescens extract were analyzed using LC-MS techniques. The toxicity and behavioral responses of larvae and adults of P. xylostella to ten compounds were investigated. We found that the caryophyllene oxide and 3,4-dimethoxycinnamic acid inhibited female oviposition; the ODIs were 98.31% and 97.59% at 1.25 mg/mL, respectively. The A. pubescens extract, caryophyllene oxide, and 3,4-dimethoxycinnamic acid caused larval mortality, with LC50 values of 21.31, 4.56, and 5.52 mg/mL, respectively. The EAG response of females was higher than that of males under A. pubescens extract conditions, while the EAG response of males was higher than that of females in caryophyllene oxide and 3,4-dimethoxycinnamic acid conditions. The A. pubescens extract and caryophyllene oxide showed repellent activity against both female and male adults, while the 3,4-dimethoxycinnamic acid did not elicit any notable behavioral responses from P. xylostella adults. A. pubescens extract and caryophyllene oxide are potential insecticides, oviposition deterrents, and behavioral regulators against P. xylostella, and they could be potential candidates for the development of biological insecticides to control P. xylostella.

Role of DSC1 in Drosophila melanogaster synaptic activities in response to haedoxan A.

Drosophila sodium channel 1 (DSC1) encodes a voltage-gated divalent cation channel that mediates neuronal excitability in insects. Previous research revealed that DSC1 knockout Drosophila melanogaster conferred different susceptibility to insecticides, which indicated the vital regulation role of DSC1 under insecticide stress. Haedoxan A (HA) is a lignan compound isolated from Phryma leptostachya, and we found that HA has excellent insecticidal activity and is worthy of further study as a botanical insecticide. Herein, we performed bioassay and electrophysiological experiments to test the biological and neural changes in the larval Drosophila with/without DSC1 knockout in response to HA. Bioassay results showed that knockout of DSC1 reduced the sensitivity to HA in both w1118 (a common wild-type strain in the laboratory) and parats1 (a pyrethroid-resistant strain) larvae. Except for parats1 /DSC1-/- , electrophysiology results implicated that HA delayed the decay rate and increased the frequency of miniature excitatory junctional potentials of Drosophila from w1118 , parats1 , and DSC1-/- strains. Moreover, the neuromuscular synapse excitatory activities of parats1 /DSC1-/- larvae were more sensitive to HA than DSC1-/- larvae, which further confirmed the functional contribution of DSC1 to neuronal excitability. Collectively, these results indicated that the DSC1 channel not only regulated the insecticidal activity of HA, but also maintained the stability of neural circuits through functional interaction with voltage-gated sodium channels. Therefore, our study provides useful information for elucidating the regulatory mechanism of DSC1 in the neural system of insects involving the action of HA derived from P. leptostachya.

Nitric oxide synthase is required for the pea aphid's defence against bacterial infection.

Compared to other insects, the pea aphid Acyrthosiphon pisum has a reduced immune system with an absence of genes coding for a lot of immunity-related molecules. Notably, nitric oxide synthase (NOS), which catalyses the synthesis of nitric oxide (NO), is present in the pea aphid. However, the role of NO in the immune system of pea aphid remains unclear. In this study, we explored the role of NO in the defence of the pea aphid against bacterial infections and found that the NOS gene of the pea aphid responded to an immune challenge, with the expression of ApNOS observably upregulated after bacterial infections. Knockdown of ApNOS using RNA interference or inhibition of NOS activity increased the number of live bacterial cells in aphids and the mortality of aphids after bacterial infection. Conversely, the increase in NO level in aphids using NO donor inhibited the bacterial growth, increased the survival of bacteria-infected aphids, and upregulated immune genes, such as Toll pathway and phagocytosis related genes. Thus, NO promotes immune responses and plays an important role in the immune system of pea aphid.

Serpin-4 Negatively Regulates Prophenoloxidase Activation and Antimicrobial Peptide Synthesis in the Silkworm, Bombyx mori.

The prophenoloxidase (PPO) activation and Toll antimicrobial peptide synthesis pathways are two critical immune responses in the insect immune system. The activation of these pathways is mediated by the cascade of serine proteases, which is negatively regulated by serpins. In this study, we identified a typical serpin, BmSerpin-4, in silkworms, whose expression was dramatically up-regulated in the fat body and hemocytes after bacterial infections. The pre-injection of recombinant BmSerpin-4 remarkably decreased the antibacterial activity of the hemolymph and the expression of the antimicrobial peptides (AMPs) gloverin-3, cecropin-D, cecropin-E, and moricin in the fat body under Micrococcus luteus and Yersinia pseudotuberculosis serotype O: 3 (YP III) infection. Meanwhile, the inhibition of systemic melanization, PO activity, and PPO activation by BmSerpin-4 was also observed. Hemolymph proteinase 1 (HP1), serine protease 2 (SP2), HP6, and SP21 were predicted as the candidate target serine proteases for BmSerpin-4 through the analysis of residues adjacent to the scissile bond and comparisons of orthologous genes in Manduca sexta. This suggests that HP1, SP2, HP6, and SP21 might be essential in the activation of the serine protease cascade in both the Toll and PPO pathways in silkworms. Our study provided a comprehensive characterization of BmSerpin-4 and clues for the further dissection of silkworm PPO and Toll activation signaling.

The IMD pathway in Hemipteran: A comparative analysis and discussion.

The evolutionary patterns of the genes in the IMD pathway in hemipterans were characterized and compared. The hemipteran insects were clustered into two groups. One group that encompasses whitefly, plant lice, and scale insect partially lacks the IMD pathway and all antimicrobial peptide (AMP) genes, with the vast majority of IMD pathway and all AMP genes being absent in aphids. The reasons for the absence of the IMD pathway and AMP genes in hemipterans were analyzed based on aphids, in terms of fitness costs. In case of limited resources, aphids have to make a trade-off between the necessary costs such as clean food sources, the essential amino acids supplied by primary bacterial symbionts for survival, nutrients and/or protections against stress provided by secondary symbionts, and the high reproductive capacity, and the costs that do not increase the fitness. Obviously, aphids have to abandon the strong immune system, especially the AMPs and IMD pathway which is mainly against Gram-negative bacteria. The common ground shared with aphids may be the reason for the absence of the IMD pathway and AMP genes in other hemipteran insects.

Negative cross-resistance of a pyrethroid-resistant Drosophila mutant to Phryma leptostachya-derived haedoxan A.

Voltage-gated sodium channels are the primary target of pyrethroid insecticides. Mutations in sodium channel confer knockdown resistance (kdr) to pyrethroids in various arthropod pests. Haedoxan A (HA) is the major insecticidal component from Phryma leptostachya. It has been shown that HA alters electrical responses at the Drosophila neuromuscular junction and modifies the gating properties of cockroach sodium channels expressed in Xenopus oocytes. However, whether sodium channel mutations that confer pyrethroid resistance also affect the action of HA is unknown. In this study, we conducted bioassays using HA and permethrin in two Drosophila melanogaster strains: w1118 , an insecticide-susceptible strain, and parats1 , a pyrethroid-resistant strain due to a I265N mutation in the sodium channel, and identified a new case of negative cross-resistance (NCR) between permethrin and HA. Both parats1 larvae and adults were more resistant to permethrin, as expected. However, both parats1 larvae and adults were more sensitive to HA compared to w1118 . We confirmed that the I265N mutation reduced the sensitivity to permethrin of a Drosophila sodium channel variant, DmNav 22, expressed in Xenopus oocytes. Interestingly, the I265N mutation also abolished the effect of HA on sodium channels. Further characterization showed that I265 on the sodium channels is critical for the action of both pyrethroids and HA on sodium channels, pointing to an overlapping mode of action between pyrethroids and HA on the sodium channel. Overall, our results suggest an I265N-independnt mechanism(s) in parats1 flies that is responsible for the NCR between permethrin and HA at the whole insect level.

Transcriptome analysis of Plantago major as a phytoremediator to identify some genes related to cypermethrin detoxification.

Cypermethrin (CYP) is a toxic manmade chemical compound belonging to pyrethroid insecticides contaminating the environment. Plantago major (PM) has numerous excellent advantages like high biomass yield and great stress tolerance, which make it able to increase the efficacy of phytoremediation. So far, no study has directly or indirectly made a transcriptome analysis (RNA-seq) of PM under CYP stress. The aim of this study is to identify the genes in PM related to CYP detoxification (10 µg mL-1) and compared with control. In this study, BGISEQ-500 high-throughput sequencing technology independently developed by BGI was used to sequence the transcriptome of P. major. Six libraries were constructed including (CK_1, CK_2, and CK_3) and (CYP_1, CYP_2, and CYP_3) were sequenced for transcripts involved in CYP detoxification. Our data showed that de novo assembly generated 138,806 unigenes with an average length of 1129 bp. Analyzing the annotation results of the KEGG database between the samples revealed 37,177 differentially expressed genes (DEGs), 18,062 down- and 19,115 upregulated under CYP treatment compared with control. A set of 107 genes of cytochrome P450 (Cyt P450), 43 genes of glutathione S-transferases (GST), 25 genes of glycosyltransferases (GTs), 113 genes from ABC transporters, 21 genes from multidrug and toxin efflux (MATE), 11 genes from oligopeptide transporter (OPT), and 3 genes of metallothioneins (MT) were upregulated notably. By using quantitative real-time PCR (qRT-PCR), the results of gene expression for 12 randomly selected DEGs were confirmed, showing the different patterns of response to CYP in PM tissues. Furthermore, the enzyme activity of Cyt P450 and GST in PM under CYP stress was significantly increased in roots and leaves than in control. This study introduces a clue to understand the metabolic pathways of plants used in phytoremediation by identifying the highly expressed genes related to phytoremediation which would be utilized to enhance pesticide detoxification and reduce pollution problem.