Effects of X-ray irradiation on the fitness of the established invasive pest fall armyworm Spodoptera frugiperda.

BACKGROUND: Spodoptera frugiperda has spread to Africa, Asia, and Oceania, posing a serious threat to global agriculture. We estimated the appropriate dose of X-ray sterilization for S. frugiperda using an X-ray irradiation instrument to investigate environmentally acceptable control techniques, laying the framework for future applications of sterile insect technology (SIT) to manage the pest environmentally-friendly. RESULTS: This study is the first to investigate the effects of X-ray irradiation on the growth, development, survival, reproduction, and flight of S. frugiperda. The results showed that irradiation with 50-400 Gy had no significant effect on pupal eclosion, but females were more sensitive than males in terms of reproductive parameters, especially when doses of radiation were > 350 Gy. After irradiation with a sub-sterilizing dose of 250 Gy, the parental sterility rate was > 85%, and the sterility traits could be passed on to their offspring, resulting in a continuous decrease in the population of F1 and F2 generations. CONCLUSION: Our laboratory experiments theoretically confirmed the feasibility of SIT for controlling S. frugiperda in the field using X-ray radiation. This study provides a theoretical basis for future regional pest management strategies. © 2022 Society of Chemical Industry.

Pro-Apoptotic Function Analysis of the Reaper Homologue IBM1 in Spodoptera frugiperda.

As an important type of programmed cell death, apoptosis plays a critical role in lepidopteran insects in response to various internal and external stresses. It is controlled by a network of genes such as those encoding the inhibitor of apoptosis proteins. However, there are few studies on apoptosis-related genes in Spodoptera frugiperda. In this study, an orthologue to the Drosophila reaper gene, named Sf-IBM1, was identified from S. frugiperda, and a full-length sequence was obtained by reverse transcription polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends PCR (RACE-PCR). The expression pattern of Sf-IBM1 was determined in different developmental stages and various tissues. Apoptotic stimuli including azadirachtin, camptothecin, and ultraviolet radiation (UV) induced the expression of Sf-IBM1 at both transcript and protein levels. Overexpression of Sf-IBM1 induced apoptosis in Sf9 cells, and the Sf-IBM1 protein was localized in mitochondria. The apoptosis induced by Sf-IBM1 could be blocked by the caspase universal inhibitor carbobenzoxy-valyl-alanyl-aspartyl-[O-methyl]-fluoromethylketone (Z-VAD-FMK) and Sf-IAP1. Our results provide valuable information that should contribute to a better understanding of the molecular events that lead to apoptosis in lepidopterans.

Elucidation of molecular expression associated with abnormal development and sterility caused by electron beam irradiation in Spodoptera litura (F.) (Lepidoptera: Noctuidae).

PURPOSE: The objective of the present study was to elucidate the mode of indirect action of electron beam irradiation at the molecular level against a quarantine pest, Spodoptera litura (F.). MATERIAL AND METHODS: Electron beam irradiation (50-200 Gy) was applied to S. litura eggs, larvae, pupae, and adults, after which the feeding area, body weight, deformity of pupae and adults, ovarian development, expression levels of vitellogenin (Vg) and vitellogenin receptor (VgR) genes, and protein levels were analyzed. RESULTS: The amount of feeding by S. litura larvae and the synthesis level of 70 kDa storage protein significantly decreased as the electron beam dose increased. When larvae were treated with the electron beam, morphological deformities appeared in the pupae, and abnormal wing disc (AWD) expression significantly decreased. Ovarian development was completely inhibited in emerged adults that had undergone 200 Gy electron beam irradiation as pupae. Quantitative real-time PCR (qRT-PCR) assays showed significant downregulation of the Vg and VgR genes due to electron beam irradiation; whereas the synthesis level of Vg protein (190 kDa) did not decrease with time in eggs unlike in non-irradiated (control) S. litura eggs, exhibiting irradiation induced impairment of Vg functioning. CONCLUSIONS: These findings of radiation-induced abnormal development and sterility in S. litura together with the correlated changes at the molecular level may facilitate the development of a phytosanitary strategy against this quarantine pest using electron beam irradiation.

Identification and expression analysis of the Spodoptera exigua neuropeptidome under different physiological conditions.

Neuropeptides are small signalling molecules acting as neurohormones, neurotransmitters and neuromodulators. Being part of the chemical communication system between cells within an organism, they are involved in the regulation of different aspects of animal physiology and behaviour such as feeding, reproduction, development and locomotion. Transcriptomic data from larval and adult tissues have been obtained and mined to generate a comprehensive neuropeptidome for the polyphagous insect pest Spodoptera exigua. Sixty-three neuropeptides have been identified and described based on their tissue specificity and their regulation in response to different abiotic perturbations. Expression analyses have identified those neuropeptides involved in ingestive and digestive behaviour of S. exigua larvae and revealed a general pattern of upregulation in the midgut during larval starvation. Our results represent a comprehensive neuropeptidome of a lepidopteran species that will be highly relevant to future studies and provide novel information of the insect's perception of its environment.

Irradiation influence on the phenoloxidase pathway and an anti-oxidant defense mechanism in Spodoptera litura (Lepidoptera: Noctuidae) and its implication in radio-genetic 'F 1 sterility' and biorational pest suppression tactics.

The present study was conducted to appraise the ontogenic radio-sensitivity of a serious tropical pest, Spodoptera litura (Fabr.). The molecular responses pertaining to the phenoloxidase (PO) pathway and an anti-oxidant defense mechanism were evaluated in order to understand its implication in pest control at pre-harvest and post-harvest intervals. Irradiation exhibited an inverse relationship with age with respect to impact on developmental and transcriptional responses. Transcript abundance of PO cascade enzymes, prophenoloxidase (slppo-2), its activating enzyme (slppae-1) and free-radical scavenging enzymes, superoxide dismutase (slsod) and catalase (slcat) was evaluated upon gamma irradiation alone and the dual-stress of radiation plus microbial challenge. The slppo-2, slppae-1, slsod and slcat transcripts were significantly up-regulated in F 1 L6 larvae (6th-instar) resulting from 100 Gy sub-sterilized male adults and unirradiated female moths. The extent of upregulation was relatively higher in comparison with L6 survivors (6th-instar larvae) developed from irradiated neonates (L1) treated with 100 Gy. Upon Photorhabdus challenge, the transcripts were down-regulated in irradiated L1 suggesting increased larval susceptibility to bacterial infections. Radioresistance increased with the age of the insect, and molecular responses (transcript abundance) of insect defense mechanism were less influenced when older age (F 1 progeny) were irradiated. These findings will help to optimize the gamma dose to be employed in inherited sterility technique for (pre-harvest) pest suppression and (post-harvest) phytosanitation and quarantine, and suggest compatible integration of biorational tactics including nuclear technology.

Radioresistant Sf9 insect cells display efficient antioxidant defence against high dose γ-radiation.

PURPOSE: To investigate the effect of gamma radiation-induced alterations in antioxidant defence of radioresistant Sf9 insect cells. MATERIALS AND METHODS: Sf9 cells were irradiated at doses ranging from 0.5-200 Gy. Lipid peroxidation and protein carbonylation levels were observed at 4 h post-exposure along with reduced glutathione/oxidized glutathione (GSH/GSSG) profile as well as specific activities of redox active enzymes superoxide dismutase (SOD), catalase, ascorbate peroxidase (APOx), and glutathione reductase (GR). Human brain malignant glioma (BMG-1) cells were used for comparing radiation response of mammalian cells. RESULTS: Sf9 cells displayed significantly less radiation-induced reactive oxygen/nitrogen species (ROS/RNS) generation, protein carbonylation and growth inhibition as compared to mammalian cells. Sf9 cells have higher basal APOx (∼4-fold), catalase (∼1.7-fold), SOD (∼1.3-fold) activity and GSH level (∼2.2-fold) compared to mammalian cells. A radiation dose-dependent increase in SOD, Catalase and APOx activity was found in Sf9 cells at least up to 100 Gy dose, while maximum activity in mammalian cells was achieved by 10 Gy. CONCLUSION: The present study suggests that Lepidopteran insect cells carry a stronger antioxidant system that protects against radiation-induced macromolecular damage, growth inhibition and cell death.

Ultraviolet-B light induced oxidative stress: effects on antioxidant response of Spodoptera litura.

Ultraviolet light (UV-B), which emits radiation in the range of 280-315 nm, has been used worldwide in light trapping of insect pests. In this article, we test the hypothesis that one of the duration of UV-B exposure has a differential impact on oxidative stress marker enzymes in Spodoptera litura. Effect of UV-B exposure on total protein and antioxidant activities of superoxide dismutase (SOD), catalase (CAT), peroxidases (POX) and glutathione-S-transferase (GST) were investigated in S. litura. The adults were exposed to UV-B light for various time periods (0, 30, 60, 90 and 120 min). We found that exposure to UV-B light for 30 and 60 min resulted in increased activities of POX. When the exposure time lasted for 60 and 90 min, the activities of SOD remained significantly higher than the control. However, the POX, CAT and GST activity decreased to control levels at 90 and 120 min. whereas relatively long duration exposure activates the xenobiotics detoxifying enzymes like GST and POX and CAT enzymes. Longer UV-B exposure may interfere with pesticide detoxification mechanism in insects, making them more susceptible to insecticides.

Spodoptera frugiperda FKBP-46 is a consensus p53 motif binding protein.

p53 protein, the central molecule of the apoptosis pathway, is mutated in 50% of the human cancers. Of late, p53 homologues have been identified from different invertebrates including Drosophila melanogaster, Caenorhabditis elegans, Squid, and Clams. We report the identification of a p53-like protein in Spodoptera frugiperda (Sf9) insect cells, which is activated during oxidative stress, caused by exposure to UV-B or H(2) O(2) , and binds to p53 consensus DNA binding motifs as well as other p53 cognate motifs. Sf9 p53 motif-binding protein is similar to murine and Drosophila p53 in terms of molecular size, which is around 50-60 kDa, as evident from UV cross-linking, and displays DNA binding characteristics similar to both insect and vertebrate p53 as seen from electrophoretic mobility shift assays. The N-terminal sequencing of the purified Sf9 p53 motif-binding protein reveals extensive homology to the pro-apoptotic FK-506 binding protein (FKBP-46), earlier identified in Sf9 cells as a factor which interacts with murine casein kinase. FKBP, an evolutionarily conserved protein of mammalian origin functions as a pro-apoptotic factor. Identification of FKBP-46 as a novel p53 motif-binding protein in insect cells adds a new facet to our understanding of the mechanisms of apoptosis under oxidative stress in the absence of a typical p53 homologue.

The synthesis and photoactivated cytotoxicity of 2-methyl-4-oxo-3-prop-2-yn-1-ylcyclopent-2-en-1-yl-3-(2,2-dichlorovinyl)-2,2-dimethylcyclopropanecarboxylate conjugated with alpha-terthienyl derivatives.

The synthesis of one pyrethroid insecticide [2-methyl-4-oxo-3-prop-2-yn-1-ylcyclopent-2-en-1-yl-3-(2,2-dichlorovinyl)-2,2-dimethylcyclopropanecarboxylate (Abbrev. JZ) (Fig. 1)] conjugated with a series of alpha-terthienyl derivatives (2-8) (Fig. 1) by palladium/copper-catalyzed cross-coupling reaction is presented here for evaluating the photoactivated cytotoxicity. The photoactivated cytotoxicity on Spodoptera litura (SL) cell line was detected by MTT assay. The inhibitory activity of all the conjugates was enhanced in the irradiation condition, compared with that of JZ. The IC(50) values of the most effective compound 9 (Fig. 1) treated with irradiation were 11.60 microg mL(-1) at 24h and 8.93 microg mL(-1) at 48 h, respectively. Generation of intracellular reactive oxygen species (ROS) and change of mitochondrial transmembrane potential (MMP) on SL cells treated with compound 9 were used for the further photoactivated study. A summary of these experiments on compound 9 demonstrated the notable ROS generation and dramatic MMP decrease when irradiated with UVA light. The results also represented statistically significant difference between dark and irradiation treatment of compound 9. However, in control and JZ groups, the effects were not statistically different. It was proved that our prepared compounds were ideal candidates for new photoactivated pyrethroid insecticides.

Changes in the haemolymph proteome of Spodoptera littoralis induced by the parasitoid Chelonus inanitus or its polydnavirus and physiological implications.

The egg-larval parasitoid Chelonus inanitus induces in its host Spodoptera littoralis two major developmental effects, namely a precocious onset of metamorphosis followed by a developmental arrest in the prepupal stage. Along with each egg, the wasp injects polydnavirus and venom into the host egg. The polydnavirus has been shown to play a major role in inducing the developmental arrest while the parasitoid larva is instrumental in inducing the precocious onset of metamorphosis. Here we report that experimental dilution of haemolymph of polydnavirus-containing larvae can partially prevent the developmental arrest while injection of native, but not of heat-treated, haemolymph or plasma from polydnavirus-containing larvae into nonparasitized larvae could induce developmental arrest in 14-15% of the larvae. This illustrates that heat-labile factors present in haemolymph play a role in causing developmental arrest. Injection of parasitoid medium increased the proportion of larvae entering metamorphosis precociously while injection of antibodies against a parasitoid-released protein had the opposite effect; this indicates that this protein and possibly other parasitoid-released substances are involved in inducing the precocious onset of metamorphosis. Analysis of the plasma proteome of nonparasitized, parasitized and polydnavirus-containing larvae revealed that the developmental effects are associated with only minor differences: eleven low abundant viral or virus-induced proteins and five parasitoid-released proteins were seen at specific stages of the host.

Radiation responses of Sf9, a highly radioresistant lepidopteran insect cell line.

PURPOSE: Lepidopteran insect cells are known to exhibit very high radioresistance. Although very effective DNA excision-repair has been proposed as a contributing factor, a detailed understanding of insect cell radiation responses has not yet been obtained. Therefore, the study was carried out to understand the in vitro radiation responses of Sf9 lepidopteran cells. MATERIALS AND METHODS: Exponentially growing asynchronous Sf9 cells (derived from ovaries of Spodoptera frugiperda) were exposed to gamma-radiation doses of 2-200 Gy. Cell survival, growth inhibition, cell cycle progression delay, alterations in cell morphology as well as induction of DNA damage, micronuclei and apoptosis were studied at various post-irradiation time intervals. RESULTS: Biphasic survival response curves were obtained with D0 rising from 20 Gy (at doses < or = 60 Gy) to 85 Gy (between 60 and 200 Gy), corroborating earlier reports on lepidopteran cells. An additional downward deviation at 2 Gy indicated a hypersensitive response. Dose-dependent growth inhibition with a transient G2 delay starting 12 h and extending up to 48-96 h was observed at doses of 10-200 Gy, while a brief G1/S transition delay was observed only at higher doses (> or = 100 Gy). Significant DNA damage was detected only at 20 Gy and higher doses, in contrast with human cells that showed similar damage at 2 Gy. Interestingly, micronuclei were not induced at any of the doses tested, although spontaneous micronucleation was evident in <1% of cells. Lack of micronucleus induction even at doses that induced significant DNA damage and a transient G2 block (20-50 Gy) strongly indicated a role of holocentric lepidopteran chromosomes. Apoptosis was detected only in a small proportion of cells (3%) exposed to 200 Gy, and cell/nucleus size and granularity increased by 72-96 h post-irradiation in a dose-dependent manner. Sf9 nucleoids extracted at 2 M NaCl showed higher compactness than the nucleoids prepared from human cells. CONCLUSIONS: It is clearly shown that lepidopteran cells are highly resistant to the induction of DNA damage and micronuclei, and display very low induction of apoptosis at doses up to 200 Gy. While the lack of micronucleus induction seems to be primarily due to the holocentric nature of their chromosomes, certain unique signalling pathways might be responsible for the low induction of apoptosis. Factors causing protection of Sf9 cellular DNA from radiation-induced damage are presently being investigated.