Cuticle Protein Genes Research Articles

RNAi of nuclear receptor E78 inhibits the cuticle formation in the molting process of spider mite, Tetranychus urticae.

The two-spotted spider mite, Tetranychus urticae, is an important pest mite in agriculture worldwide. E78, as a member of the nuclear receptor superfamily and a downstream responsive gene of ecdysteroids, plays a crucial role in regulating physiological behaviors such as development and reproduction in insects. However, its function in mites remains unclear. The aim of this study was to explore how E78 functions in the molting process of spider mites. In this study, reverse transcription quantitative polymerase chain reaction (RT-qPCR) experiments to analyze the expression pattern of TuE78 during the development of Tetranychus urticae, demonstrated that the expression level of TuE78 was higher during the molting state than that after the completion of molting, and it reached a peak expression level when the deutonymph mites entered the molting stage. RNA interference (RNAi)-mediated gene-silencing of TuE78 resulted in 95% deutonymph mite molt failure. A series of analysis under a light microscope, and scanning and transmission electron microscopy revealed that RNAi mites died within the exuvium without ecdysis, and that apolysis had started but the new cuticle was thin and the typical cuticular lamellae were absent, indicating blockage of the post-apolysial processes and explaining molt failure. Hence, transcriptome sequencing confirmed that the expression of cuticle protein and lipid metabolism-related genes was significantly affected after TuE78 silencing. This study demonstrated that TuE78 participates in the molting process of Tetranychus urticae by regulating the post-apolysial processes with the formation of new cuticle and successful ecdysis. This in turn suggests the potential of TuE78 as a target for pest mite control and provides a theoretical basis for further exploration of the molecular regulatory mechanism of spider mite molting. © 2024 Society of Chemical Industry. Published by John Wiley & Sons Ltd.

The transcription factor RUNT-like regulates pupal cuticle development via promoting a pupal cuticle protein transcription.

Holometabolous insects undergo morphological remodeling from larvae to pupae and to adults with typical changes in the cuticle; however, the mechanism is unclear. Using the lepidopteran agricultural insect Helicoverpa armigera, cotton bollworm, as a model, we revealed that the transcription factor RUNT-like (encoded by Runt-like) regulates the development of the pupal cuticle via promoting a pupal cuticle protein gene (HaPcp) expression. The HaPcp was highly expressed in the epidermis and wing during metamorphosis and was found being involved in pupal cuticle development by RNA interference (RNAi) analysis in larvae. Runt-like was also strongly upregulated in the epidermis and wing during metamorphosis. Knockdown of Runt-like produced similar phenomena, a failure of abdomen yellow envelope and wing formation, to those following HaPcp knockdown. The insect molting hormone 20-hydroxyecdysonen (20E) upregulated HaPcp transcription via RUNT-like. 20E upregulated Runt-like transcription via nuclear receptor EcR and the transcription factor FOXO. Together, RUNT-like and HaPCP are involved in pupal cuticle development during metamorphosis under 20E regulation.

Imidazole-modified graphene quantum dots can effectively promote the efficient silencing of the larval cuticle protein gene HaLCP17 in Helicoverpa armigera

Imidazole-modified graphene quantum dots can effectively promote the efficient silencing of the larval cuticle protein gene HaLCP17 in Helicoverpa armigera

The cuticular protein gene ApCP7 and ApCP62 are essential for reproduction in Acyrthosiphon pisum, affecting ecdysis and survival

Cuticular proteins, in conjunction with chitin, compose the insect exoskeleton, and play a key role in the growth, development, and molting of insects. However, the specific functions of most cuticular protein genes in the growth, development, and reproductive processes of the pea aphid (Acyrthosiphon pisum) remain unclear. In this study, we have identified six cuticular protein genes in the pea aphid, namely ApCP7, ApCP10, ApCP19, ApCP19.8-like, ApCP35 and ApCP62. We found that the expression levels of six genes were highly expressed during the adult stage, and except for ApCP10, which is highly expressed in the pea aphid cuticle, other genes were highly expressed in the ovaries. Subsequently, we observed that the survival rate and fecundity of pea aphid were significantly lower than those of the control group after silencing ApCP7 and ApCP62 through RNA interference. Furthermore, when ApCP7 transcript levels were reduced, aphid encountered difficulties in molting, were smaller in body size, and exhibited a darker body color. These results indicate that ApCP7 and ApCP62 are involved in the development and reproduction of pea aphid, and could be used as RNAi targets for controlling pea aphid.

The cuticle proteome of a planktonic crustacean.

The cuticles of arthropods provide an interface between the organism and its environment. Thus, the cuticle's structure influences how the organism responds to and interacts with its surroundings. Here, we used label-free quantification proteomics to provide a proteome of the moulted cuticle of the aquatic crustacean Daphnia magna, which has long been a prominent subject of studies on ecology, evolution, and developmental biology. We detected a total of 278 high-confidence proteins. Using protein sequence domain and functional enrichment analyses, we identified chitin-binding structural proteins and chitin-modifying enzymes as the most abundant protein groups in the cuticle proteome. Structural cuticular protein families showed a similar distribution to those found in other arthropods and indicated proteins responsible for the soft and flexible structure of the Daphnia cuticle. Finally, cuticle protein genes were also clustered as tandem gene arrays in the D. magna genome. The cuticle proteome presented here will be a valuable resource to the Daphnia research community, informing genome annotations and investigations on diverse topics such as the genetic basis of interactions with predators and parasites.

Genome-Wide Exploration of Long Non-Coding RNAs of Helicoverpa armigera in Response to Pyrethroid Insecticide Resistance.

Genome-wide long non-coding RNAs (lncRNAs) in low, moderate, and high pyrethroid insecticide-resistant and -susceptible strains of Helicoverpa armigera were identified in this study. Using 45 illumina-based RNA-sequencing datasets, 8394 lncRNAs were identified. In addition, a sublethal dose of deltamethrin was administered to a Korean-resistant strain (Kor-T). The average length of lncRNAs was approximately 531 bp, and the expression ratio of lncRNAs was 28% of the total RNA. The identified lncRNAs were divided into six categories-intronic, intergenic, sense, antisense, cis-RNA, and trans-RNA-based on their location and mechanism of action. Intergenic and intronic lncRNA transcripts were the most abundant (38% and 33%, respectively). Further, 828 detoxification-related lncRNAs were selected using the Gene Ontology analysis. The cytochrome P450-related lncRNA expression levels were significantly higher in susceptible strains than in resistant strains. In contrast, cuticle protein-related lncRNA expression levels were significantly higher in all resistant strains than in susceptible strains. Our findings suggest that certain lncRNAs contribute to the downregulation of insecticide resistance-related P450 genes in susceptible strains, whereas other lncRNAs may be involved in the overexpression of cuticle protein genes, potentially affecting the pyrethroid resistance mechanism.

Transcriptomic profiling of different developmental stages reveals parasitic strategies of Wohlfahrtia magnifica, a myiasis-causing flesh fly

BackgroundWohlfahrtia magnifica is an obligatory parasite that causes myiasis in several warm-blooded vertebrates. Adult females deposit the first-stage larvae directly onto wounds or natural body orifices (e.g., genitalia) of the host, from where they quickly colonize the host tissue and feed on it for development. The infestation of W. magnifica can lead to health issues, welfare concerns, and substantial economic losses. To date, little is known about the molecular mechanisms of the W. magnifica-causing myiasis.ResultsIn this study, we collected parasitic-stage larvae of W. magnifica from wounds of naturally infested Bactrian camels, as well as pupae and adult flies reared in vitro from the wound-collected larvae, for investigating the gene expression profiles of the different developmental stages of W. magnifica, with a particular focus on examining gene families closely related to the parasitism of the wound-collected larvae. As key proteins related to the parasite-host interaction, 2049 excretory/secretory (ES) proteins were identified in W. magnifica through the integration of multiple bioinformatics approaches. Functional analysis indicates that these ES proteins are primarily involved in cuticle development, peptidase activity, immune response, and metabolic processes. The global investigation of gene expression at different developmental stages using pairwise comparisons and weighted correlation network analysis (WGCNA) showed that the upregulated genes during second-stage larvae were related to cuticle development, peptidase activity, and RNA transcription and translation; during third-stage larvae to peptidase inhibitor activity and nutrient reservoir activity; during pupae to cell and tissue morphogenesis and cell and tissue development; and during adult flies to signal perception, many of them involved in light perception, and adult behavior, e.g., feeding, mating, and locomotion. Specifically, the expression level analysis of the likely parasitism-related genes in parasitic wound-collected larvae revealed a significant upregulation of 88 peptidase genes (including 47 serine peptidase genes), 110 cuticle protein genes, and 21 heat shock protein (hsp) genes. Interestingly, the expression of 2 antimicrobial peptide (AMP) genes, including 1 defensin and 1 diptericin, was also upregulated in the parasitic larvae.ConclusionsWe identified ES proteins in W. magnifica and investigated their functional distribution. In addition, gene expression profiles at different developmental stages of W. magnifica were examined. Specifically, we focused on gene families closely related to parasitism of wound-collected larvae. These findings shed light on the molecular mechanisms underlying the life cycle of the myiasis-causing fly, especially during the parasitic larval stages, and provide guidance for the development of control measures against W. magnifica.

Chromosome-level genome of spider Pardosa pseudoannulata and cuticle protein genes in environmental stresses

Spiders are representative arthropods of adaptive radiation. The high-quality genomes have only been reported in several web weaver spider species, leaving the wandering spiders’ genomic information scarce. The pond wolf spider, Pardosa pseudoannulata, is a representative species in the retrolateral titial apophysis (RTA) clade. We present a chromosome-level P. pseusoannulata genome assembly of 2.42 Gb in size with a scaffold N50 of 169.99 Mb. Hi-C scaffolding assigns 94.83% of the bases to 15 pseudo-chromosomes. The repeats account for 52.79% of the assembly. The assembly includes 96.2% of the complete arthropod universal single-copy orthologs. Gene annotation predicted 24,530 protein-coding genes with a BUSCO score of 95.8% complete. We identified duplicate clusters of Hox genes and an expanded cuticle protein gene family with 243 genes. The expression patterns of CPR genes change in response to environmental stresses such as coldness and insecticide exposure. The high-quality P. pseudoannulata genome provides valuable information for functional and comparative studies in spiders.

Identifying the genes involved in the egg-carrying ovigerous hair development of the female blue crab Callinectes sapidus: transcriptomic and genomic expression analyses

BackgroundCrustacean female sex hormone (CFSH) controls gradually developing adult female-specific morphological features essential for mating and brood care. Specifically, ovigerous hairs are developed during the prepuberty molt cycle of the blue crab Callinectes sapidus that are essential for carrying the eggs until they finish development. Reduced CFSH transcripts by CFSH-dsRNA injections result in fewer and shorter ovigerous hairs than the control. This study aimed to identify the specific genes responsible for ovigerous hair formation using transcriptomic, genomic and expression analyses of the ovigerous setae at three stages: prepuberty at early (OE) and late premolt (OL), and adult (AO) stages.ResultsThe de novo Trinity assembly on filtered sequence reads produced 96,684 Trinity genes and 124,128 transcripts with an N50 of 1,615 bp. About 27.3% of the assembled Trinity genes are annotated to the public protein sequence databases (i.e., NR, Swiss-Prot, COG, KEGG, and GO databases). The OE vs. OL, OL vs. AO, and OE vs. AO comparisons resulted in 6,547, 7,793, and 7,481 differentially expressed genes, respectively, at a log2-fold difference. Specifically, the genes involved in the Wnt signaling and cell cycle pathways are positively associated with ovigerous hair development. Moreover, the transcripts of ten cuticle protein genes containing chitin-binding domains are most significantly changed by transcriptomic analysis and RT-qPCR assays, which shows a molt-stage specific, down-up-down mode across the OE-OL-AO stages. Furthermore, the expression of the cuticle genes with the chitin-binding domain, Rebers and Riddiford domain (RR)-1 appears at early premolt, followed by RR-2 at late premolt stage. Mapping these 10 cuticle protein sequences to the C. sapidus genome reveals that two scaffolds with a 549.5Kb region and 35 with a 1.19 Mb region harbor 21 RR1 and 20 RR2 cuticle protein genes, respectively. With these findings, a putative mode of CFSH action in decapod crustaceans is proposed.ConclusionsThe present study describes a first step in understanding the mechanism underlying ovigerous hair formation in C. sapidus at the molecular level. Overall, demonstrating the first transcriptome analysis of crustacean ovigerous setae, our results may facilitate future studies into the decapod female reproduction belonging to the suborder Pleocyemata.

Screening of morphology-related genes based on predator-induced transcriptome sequencing and the functional analysis of Dagcut gene in Daphnia galeata.

High fish predation pressure can trigger "induced defense" in Daphnia species, resulting in phenotypic plasticity in morphology, behavior, or life-history traits. The molecular mechanisms of defense morphogenesis (e.g., the tail spine and helmet) in Daphnia remain unclear. In the present study, the tail spine, helmet, and body of Daphnia galeata under fish and non-fish kairomones conditions were collected for transcriptome analysis. A total of 24 candidate genes related to the morphological defense of D. galeata were identified, including 2 trypsin, one cuticle protein, 1 C1qDC protein, and 2 ferritin genes. The function of the Dagcut gene (D. galeata cuticle protein gene) in relation to tail spine morphology was assessed using RNA interference (RNAi). Compared with the EGFP (Enhanced green fluorescent protein) treatment, after RNAi, the expression levels of the Dagcut gene (D. galeata cuticle protein gene) showed a significant decrease. Correspondingly, the tail spines of the offspring produced by D. galeata after RNAi of the Dagcut gene appeared curved during the experiment. In whole-mount in situ hybridization, a clear signal site was detected on the tail spine of D. galeata before RNAi which disappeared after RNAi. Our results suggest that the Dagcut gene may play an important role in tail spine formation of D. galeata, and will provide a theoretical basis for studying the molecular mechanisms of the morphological plasticity in cladocera in the future.

Insights into the Effects of Insecticides on Aphids (Hemiptera: Aphididae): Resistance Mechanisms and Molecular Basis

With the passage of time and indiscreet usage of insecticides on crops, aphids are becoming resistant to their effect. The different classes of insecticides, including organophosphates, carbamates, pyrethroids and neonicotinoids, have varied effects on insects. Furthermore, the molecular effects of these insecticides in aphids, including effects on the enzymatic machinery and gene mutation, are resulting in aphid resistance to the insecticides. In this review, we will discuss how aphids are affected by the overuse of pesticides, how resistance appears, and which mechanisms participate in the resistance mechanisms in various aphid species as significant crop pests. Gene expression studies were analyzed using the RNA-Seq technique. The stress-responsive genes were analyzed, and their expression in response to insecticide administration was determined. Putative insecticide resistance-related genes, cytochrome P450, glutathione S-transferase, carboxylesterase CarEs, ABC transporters, cuticle protein genes, and trypsin-related genes were studied. The review concluded that if insecticide-susceptible aphids interact with ample dosages of insecticides with sublethal effects, this will result in the upregulation of genes whose primary role is to detoxify insecticides. In the past decade, certain advancements have been observed regarding insecticide resistance on a molecular basis. Even so, not much is known about how aphids detoxify the insecticides at molecular level. Thus, to attain equilibrium, it is important to observe the manipulation of pest and insect species with the aim of restoring susceptibility to insecticides. For this purpose, this review has included critical insights into insecticide resistance in aphids.

The tissue-specific expression of silkworm cuticle protein gene ASSCP2 is mediated by the Sox-2 transcription factor

The silk gland of silkworm is a unique organ in which silk proteins are synthesized, secreted, and transformed into fibers. The anterior silk gland (ASG) is located at the end of the silk gland, and is thought to be involved in silk fibrosis. In our previous study, a cuticle protein, ASSCP2, was identified. This protein is specifically and highly expressed in the ASG. In this work, the transcriptional regulation mechanism of ASSCP2 gene was studied by a transgenic route. The ASSCP2 promoter was analyzed, truncated sequentially, and used to initiate the expression of EGFP gene in silkworm larvae. After egg injection, seven transgenic silkworm lines were isolated. Molecular analysis revealed that the green fluorescent signal could not be detected when the promoter was truncated to −257 bp, suggesting that the −357 to −257 sequence is the key region responsible for the transcriptional regulation of the ASSCP2 gene. Furthermore, an ASG specific transcription factor Sox-2 was identified. EMSA assays showed that Sox-2 binds with the −357 to −257 sequence, and thus regulates the tissue-specific expression of ASSCP2. This study on the transcriptional regulation of ASSCP2 gene provides theoretical and experimental basis for further studies of the regulatory mechanism of tissue-specific genes.

Food quality and exposure time affect induced defences in Daphnia: Based on key morphology, life history traits, and expression of related genes

Abstract Kairomone‐induced anti‐predation defences in zooplankton mediate trophic interactions in aquatic ecosystems. Changes in food quality greatly affect induced defences because of the potential influence on resource allocation. Understanding how zooplankton respond to predation risk under poor nutrition conditions is critical for predicting ecosystem function. The present study focused on the fish kairomone‐induced defences in Daphnia pulex fed unicellular non‐toxic Microcystis aeruginosa. Transcriptions of key genes were detected to interpret changes in morphological and life history traits. Additionally, the degree of responsiveness of traits to increasing proportions of M. aeruginosa in the diet was compared at different endpoints with varied exposure duration. Daphnia pulex decreased body size but showed an elongated tail spine as morphological adaptations to the presence of fish kairomones, supported by enhanced transcriptions of chitin deacetylase and cuticle protein genes. However, with suppressed expression of the related genes, tail spine length was linearly reduced by increasing proportions of M. aeruginosa in the diet, accompanied by decreased somatic growth and reproduction. Presence of fish kairomones interactively affected the inhibitive effects of increasing proportions of M. aeruginosa in the diet on Daphnia growth, reproduction, and tail spine defence but not body size. In addition, the magnitude of effects of M. aeruginosa on Daphnia growth and morphological responses under fish kairomone exposure was increased with the increasing length of exposure time from 3–7 days (reaching maturity) to 14 days (by the end of tests), indicating an aggravated inhibitive effect of M. aeruginosa on induced defences over time. The present study provides references for evaluating zooplankton‐fish interactions under poor nutrition conditions. Since a cyanobacterial bloom may last from a few weeks to several months, experiments involving the whole life span of zooplankton are thought to fully reveal the actual impact of cyanobacteria on interspecific interactions related to zooplankton. In addition, unifying the exposure time to cyanobacteria for comparative studies is recommended in future research.

Cuticle protein gene LmCP8 is involved in the structural development of the ovipositor in the migratory locust Locusta migratoria.

The ovipositor comprises the external genitalia of female insects, which plays an important role in the mating and ovipositing process of insects. However, it remains rudimentary of regional gene expression and physiological function in the ovipositor during structural development. Here, we analysed the basic structure and characteristics of the ovipositor in the migratory locust Locusta migratoria. RNA-seq analysis revealed the specialization of chitin metabolism, lipids synthesis and transport, tanning and cuticular protein genes in the ovipositor. Among them, two cuticle protein genes, LmCP8 and LmACP79, were identified, which are specifically expressed in the ovipositor. Functional analysis based on RNA interference showed that deficiency of LmCP8 affected the structural development of the ovipositor resulting in the retention of a large number of remaining unproduced oocysts in the ovary of the locusts. Our results provide a fundamental resource to investigate the structural development and physiological function of the ovipositor in L. migratoria.

Knockdown of UDP-N-acetylglucosamine pyrophosphorylase and chitin synthase A increases the insecticidal efficiency of Lufenuron to Spodoptera exigua

Spodoptera exigua (Lepidoptera, Noctuidae) has been responsible for causing considerable and widespread agricultural losses worldwide. Owing to strong selective pressure, S. exigua showed increased resistance to Lufenuron (LUF). Consequently, RNA interference (RNAi)-based insecticides had more benefits than chemical insecticides. Therefore, to enhance the insecticidal activity of LUF to S. exigua, in the present study, we aimed to elucidate the impact of double-stranded RNAs (dsRNAs) on S. exigua larval susceptibility to LUF. First, the transcriptome of S. exigua was sequenced following the treatment with LUF. By comparing the upregulated and downregulated GO enrichment, chitin binding and chitin metabolic processes were the significantly enriched pathways. According to transcriptome sequencing, 8 genes associated with chitin biosynthesis, 8 chitin degradation genes, and 17 cuticle protein genes were obtained. UDP-N-acetylglucosamine pyrophosphorylase (UAP) and Chitin synthase A (CHSA) showed significantly downregulated expression after treatment with different sublethal doses of LUF. Downregulation of UAP increased mortality from 31.97% to 47.91% when the larvae were exposed to LUF. A significant increase in the mortality of S. exigua from 30.63% to 50.19% was observed following LUF administration after dsCHSA. In addition, the expression analysis of genes associated with chitin biosynthesis was significantly changed after LUF treatment, dsRNAs-RNAi, and their combination (LUF-dsRNAs). Significant differences were observed in the chitin content between the control group at 72 h after treatments. Results of the present study can help further elucidate the understanding of the combined effects of RNAi and LUF on S. exigua. Additionally, this research provides a suitable foundation for future studies with the aim to develop an efficient method of delivery for large-scale pest control in the fields.

Cuticle Protein LmACP19 Is Required for the Stability of Epidermal Cells in Wing Development and Morphogenesis of Locusta migratoria.

Insect wing consists of a double layer of epidermal cells that produce and secrete the dorsal and ventral cuticular components. It is important for the stability of epidermal cells during wing development and morphogenesis, but its specific gene expression and physiological function during this process remain unclear. In our previous work, a wing cuticle protein gene LmACP19 was identified in Locusta migratoria based on transcriptomic data. Here, we report on its roles in wing development and morphogenesis. LmACP19 encodes a chitin-binding protein belonging to RR-2 subfamily of CPR family, which is highly homologous to CP19-like proteins in other insect species. RT-qPCR analysis revealed that LmACP19 is highly expressed in wing pads of fifth-instar nymphs, and its encoded protein is located in two layers of epidermal cells but not in the cuticle. Suppression of LmACP19 by RNA interference led to abnormal wing pad and wing morphogenesis with curved, unclosed, and wrinkled phenotypes during nymph-to-nymph and nymph-to-adult transition, respectively. Furthermore, deficiency of LmACP19 affected arrangement of epidermal cells, resulting in apoptosis. Our results indicate that LmACP19 is indispensable for wing development and normal morphological structure by maintaining the stability of epidermal cells during L. migratoria molting.

Toxicity of Ribavirin to Spodoptera litura by Inhibiting the Juvenile Hormone.

Ribavirin is an antiviral drug showing high and delayed toxicity to the destructive agricultural pest Spodoptera litura. Larvae fed with artificial diets containing ribavirin could not molt successfully and showed abnormal phenotypes, including cuticle melanization and heavy wrinkle of the newly formed procuticle. RNA-Seq analysis suggested that ribavirin has great negative influence on cuticle. Quantitative real-time-polymerase chain reaction results indicated that ribavirin treatment decreased the expression of key genes in juvenile hormone (JH) biosynthesis (CYP15C1 and JH acid methyltransferase) and most cuticle protein genes, whereas the genes in melanin biosynthesis and bursicon genes were upregulated by ribavirin treatment. These results coincided with the decreased titer of JH I, JH II, and JH III determined by enzyme-linked immunosorbent assay, the much thinner procuticle layer exhibited by histopathological examination, and the cuticle melanization after ribavirin treatment. These results provided a valuable theoretical basis for the creation of green insecticides targeting JH and the development of new insecticide derivatives from 1,2,4-triazole.

Systematic identification and functional analysis of long noncoding RNAs involved in indoxacarb resistance in Spodoptera litura

Long noncoding RNAs (lncRNAs) are noncoding transcripts that are more than 200 nucleotides long. They play essential roles in regulating a variety of biological processes in many species, including insects, and some lncRNAs have been found to be associated with insecticide resistance. However, the characteristics and biological functions of lncRNAs involved in indoxacarb resistance are unknown in Spodoptera litura. We performed RNA sequencing in the SS, InRS, and FInRS of S. litura and identified 11978 lncRNAs, including 3136 intergenic lncRNAs, 7393 intronic lncRNAs, and 1449 anti-sense lncRNAs. Compared with the SS, 51 lncRNAs were upregulated and 134 lncRNAs were downregulated in the two resistant strains, and 908 differentially expressed mRNAs were predicted as the target genes of the 185 differentially expressed lncRNAs. Further analysis showed that 112 of differentially expressed lncRNAs may be associated with indoxacarb resistance by regulating the expression of 14 P450s, seven CCEs, one GST, six UGTs, five ABC transporters, and 24 cuticle protein genes, and 79 of differentially expressed lncRNAs may regulate the expression of 14 detoxification genes and 19 cuticle protein genes to participate in indoxacarb resistance by sponging 10 microRNAs. Interestingly, 47 of differentially expressed lncRNAs may mediate indoxacarb resistance through both lncRNA-mRNA and lncRNA-miRNA-mRNA regulatory pathways. Furthermore, quantitative PCR, RNA interference, and indoxacarb bioassay analyses indicated that overexpressed LNC_004867 and LNC_006576 were involved in indoxacarb resistance. This study provides comprehensive information for lncRNAs of S. litura, and presents evidence that lncRNAs have key roles in conferring insecticide resistance in S. litura.

Transcriptional analysis of Bemisia tabaci MEAM1 cryptic species under the selection pressure of neonicotinoids imidacloprid, acetamiprid and thiamethoxam

BackgroundNeonicotinoids are widely applied in the control of the destructive agricultural pest Bemisia tabaci, and resistance against these chemicals has become a common, severe problem in the control of whiteflies. To investigate the molecular mechanism underlying resistance against nenonicotinoids in whiteflies, RNA-seq technology was applied, and the variation in the transcriptomic profiles of susceptible whiteflies and whiteflies selected by imidacloprid, acetamiprid and thiamethoxam treatment was characterized.ResultsA total of 90.86 GB of clean sequence data were obtained from the 4 transcriptomes. Among the 16,069 assembled genes, 584, 110 and 147 genes were upregulated in the imidacloprid-selected strain (IMI), acetamiprid-selected strain (ACE), and thiamethoxam (THI)-selected strain, respectively, relative to the susceptible strain. Detoxification-related genes including P450s, cuticle protein genes, GSTs, UGTs and molecular chaperone HSP70s were overexpressed in the selected resistant strains, especially in the IMI strain. Five genes were downregulated in all three selected resistant strains, including 2 UDP-glucuronosyltransferase 2B18-like genes (LOC 109030370 and LOC 109032577).ConclusionsTen generations of selection with the three neonicotinoids induced different resistance levels and gene expression profiles, mainly involving cuticle protein and P450 genes, in the three selected resistant whitefly strains. The results provide a reference for research on resistance and cross-resistance against neonicotinoids in B. tabaci.

Chitin synthase 1 and five cuticle protein genes are involved in serosal cuticle formation during early embryogenesis to enhance eggshells in Nilaparvata lugens.

Many holo- and hemimetabolous insects enhance their eggshells during embryogenesis by forming a serosal cuticle (SC). To date, scholarly understanding of the SC composition and SC-related gene functions has been limited, especially for hemimetabolous insects. In this study, we initially performed transmission electron microscopic (TEM) observation and chitin staining of the SC in Nilaparvata lugens, a hemimetabolous rice pest known as the brown planthopper (BPH). We confirmed that the SC was a chitin-rich lamellar structure deposited gradually during the early embryogenesis. Parental RNA interference (RNAi) against Nilaparvata lugens chitin synthase 1 (NlCHS1) in newly emerged and matured females resulted in decreases of egg hatchability by 100% and 76%, respectively. Ultrastructural analyses revealed loss of the lamellar structure of the SC in dsNlCHS1-treated eggs. According to temporal expression profiles, five cuticle protein coding genes, NlugCpr1/2/3/8/90, were specifically or highly expressed during the SC formation period, and NlugCpr1/2/3/90 were further detected in 72 h eggshells extract by ultra-performance liquid chromatography-tandem mass spectrometry/mass spectrometry. NlugCpr2/3/90 were likely three SC-specific cuticle proteins. TEM observations of the SC following parental RNAi against NlugCpr1/2/3/8/90 demonstrated that NlugCpr3/8/90 were essential for SC formation. The study provided an understanding of the SC formation process and SC-related cuticle proteins in BPHs, which offer potential targets for pest control in the egg stage as well.