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1.
Pest Manag Sci ; 78(1): 296-303, 2022 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-34487617

RESUMO

BACKGROUND: Hyblaea puera, commonly known as the teak defoliator, is a serious pest in teak plantations. Despite the availability of control measures, this pest causes losses in yield and quality of timber through voracious feeding. RNA interference (RNAi) is a promising strategy for the control of this pest. Chitin metabolism, which is vital for the growth and development of arthropods, is a potential target for developing RNAi-based insecticides. RESULTS: To assess the effects of chitin metabolism inhibition, H. puera larvae were treated with a chitin synthesis inhibitor, diflubenzuron (DFB). DFB treatment caused pupal deformities and disrupted eclosion. Partial gene sequences for three key genes of H. puera chitin metabolism were cloned and sequenced: chitin synthase 1 (HpCHS1), chitinase-h (HpChi-h) and ecdysone receptor (HpEcR). Feeding dsRNA cognate for these three target genes to the first instar of H. puera resulted in mortality and reduction in the corresponding transcript levels as assessed through qRT-PCR. This is the first report of RNAi in this forestry pest. The highest mortality was 45.9%, in response to dsHpEcR treatment; HpChi-h transcripts were the most down-regulated in response to dsHpEcR feeding. DsHpEcR RNAi resulted in growth inhibition and molting arrest. The mortalities were 29.7% and 32.4% for dsHpCHS1 and dsHpChi-h feeding, respectively. CONCLUSION: Chitin metabolism could be a potential target for RNAi-based control of H. puera, and HpCHS1, HpChi-h and HpEcR could be suitable target genes. However, the RNAi efficacy needs to be improved through formulations that improve stability and uptake, and employing better delivery strategies. © 2021 Society of Chemical Industry.


Assuntos
Quitina/metabolismo , Controle de Insetos , Mariposas , Animais , Quitina Sintase/genética , Agricultura Florestal , Larva/genética , Larva/metabolismo , Mariposas/genética , Mariposas/metabolismo , Interferência de RNA
2.
Gene ; 809: 146004, 2022 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-34648918

RESUMO

Recognition of invading foreign exogenous pathogen is the first step to initiate the innate immune response of insects, which accomplished by the pattern recognition receptors (PRRs). Peptidoglycan recognition proteins (PGRPs) serve as an important type of PRRs, which activate immune response by detecting peptidoglycan of microbial cell wall. In this study, we have cloned the full-length cDNA of PGRP gene called PGRP-S1 from the Diaphania pyloalis (Walker). The open reading frame (ORF) of D. pyloalis PGRP-S1 encodes 211 amino acids which containing a secretion signal peptide and a canonical PGRP domain. Multisequence alignment revealed that PGRP-S1 possess the amino acid residues responsible for zinc binding and amidase activity. D. pyloalis PGRP-S1 exhibited the highest transcript level in fat body and followed in head. The mRNA concentration dramatically increased after an injection of Escherichia coli or Micrococcus luteus. Purified recombinant PGRP-S1 exhibit binding ability to peptidoglycans from Staphylococcus aureus or Bacillus subtilis and cause intensive agglutination of E. coli, M. luteus or S. aureus in the presence of zinc ions. Furthermore, phenoloxidase activity significantly increased when the plasma from larvae was incubated with recombinant PGPR-S1 and peptidoglycans from B. subtilis or M. luteus simultaneously. These results implied that PGRP-S1 was a member involving the prophenoloxidase activation pathway. Overall, our results indicated that D. pyloalis PGRP-S1 serve as a PRR to participate in the recognition of foreign pathogen and prophenoloxidase pathway stimulation.


Assuntos
Proteínas de Transporte/metabolismo , Catecol Oxidase/metabolismo , Precursores Enzimáticos/metabolismo , Proteínas de Insetos/metabolismo , Mariposas/metabolismo , Peptidoglicano/metabolismo , Aglutinação/efeitos dos fármacos , Animais , Bacillus subtilis/química , Proteínas de Transporte/química , Proteínas de Transporte/genética , Parede Celular/metabolismo , Regulação da Expressão Gênica , Proteínas de Insetos/química , Proteínas de Insetos/genética , Lipopolissacarídeos/metabolismo , Mariposas/genética , Mariposas/microbiologia , Filogenia , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/farmacologia , Staphylococcus aureus/química
3.
Int J Mol Sci ; 22(24)2021 Dec 16.
Artigo em Inglês | MEDLINE | ID: mdl-34948319

RESUMO

Filippi's glands (FGs), formerly also called Lyonet's glands, are accessory secretory structures of the labial (silk) glands of lepidopteran caterpillars, which were implicated to play an important role in the maturation of the silk material and the construction of the cocoon. In our previous study, we have identified several species of giant silk moths that completely lack the FGs. Interestingly, the absence of FGs in these species correlates with the construction of a loose cocoon architecture. We investigated the functions of FGs by their surgical extirpation in the last instar larvae of the silkworm, Bombyx mori. We found that the absence of FGs altered the structure of the resulting cocoon, in which the different layers of silk were separated. In further experiments, we found no effects of the absence of FGs on larval cocoon formation behavior or on changes in cocoon mass or lipid content. Differential proteomic analysis revealed no significant contribution of structural proteins from FGs to silk cocoon material, but we identified several low abundance proteins that may play a role in posttranslational modifications of some silk proteins. Proteomic analysis also revealed a difference in phosphorylation of the N-terminal sequence of fibroin-heavy chain molecule. Thus, FGs appear to affect silk stickiness during spinning by regulating posttranslational modifications. This could also explain the link that exists between the absence of these glands and the formation of loose cocoons in some giant silk moth species.


Assuntos
Bombyx/metabolismo , Mariposas/metabolismo , Animais , Fibroínas/metabolismo , Larva/metabolismo , Proteômica/métodos , Seda/metabolismo
4.
Pestic Biochem Physiol ; 179: 104969, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34802519

RESUMO

Carboxylesterases (CarEs) usually play critical roles in the detoxification of toxic chemicals and therefore may be involved in insecticide resistance in agricultural pests. Previous work has shown that CarE 001C from Helicoverpa armigera was able to metabolize the isomers of cypermethrin and fenvalerate. In this study, seven mutants of CarE 001C with single amino acid substitution were produced and expressed in the Escherichia coli. Enzyme kinetic analysis indicated that all seven mutations dramatically reduced enzymatic activities toward the generic substrate α-naphthyl acetate, but in vitro metabolism assay showed that two of the mutations, H423I and R322L, significantly improved hydrolase activities toward fenvalerate, with their recorded specific activities being 3.5 and 5.1 nM·s-1·mg -1 proteins, respectively. Further, thermostability assay showed that the stability of one mutant enzyme was enhanced. This study will help us better understand the potential of CarEs in insecticide detoxification and resistance in H. armigera.


Assuntos
Inseticidas , Mariposas , Piretrinas , Animais , Carboxilesterase/genética , Carboxilesterase/metabolismo , Hidrolases de Éster Carboxílico/genética , Resistência a Inseticidas/genética , Inseticidas/farmacologia , Cinética , Mariposas/genética , Mariposas/metabolismo , Mutação , Nitrilas
5.
Arch Insect Biochem Physiol ; 108(3): e21845, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34605064

RESUMO

With the wide cultivation of transgenic plants throughout the world and the rising risk of resistance to Bacillus thuringiensis crystal (Cry) toxins, it is essential to design an adaptive resistance management strategy for continued use. Neuropeptide F (NPF) of insects has proven to be valuable for the production of novel-type transgenic plants via its important role in the control of feeding behavior. In this study, the gene encoding NPF was cloned from the diamondback moth, Plutella xylostella, an important agricultural pest. Real-time quantitative reverse transcription-polymerase chain reaction and in situ hybridization showed a relatively high expression of P. xylostella-npf (P. x-npf) in endocrine cells of the midgut of fourth instar larvae, and it was found to participate in P. xylostella feeding behavior and Cry1Ac-induced feeding inhibition. Prokaryotic expression and purification provided structure unfolded P. x-npf from inclusion bodies for diet surface overlay bioassays and the results demonstrated a significant synergistic effect of P. x-npf on Cry1Ac toxicity by increasing intake of noxious food which contains Cry toxins, especially quick death at an early stage of feeding. Our findings provided a potential new way to efficiently control pests by increasing intake of lower dose Cry toxins and a novel hint for the complex Cry toxin mechanism.


Assuntos
Toxinas de Bacillus thuringiensis , Endotoxinas , Proteínas Hemolisinas , Mariposas , Neuropeptídeos , Animais , Toxinas de Bacillus thuringiensis/farmacologia , Endotoxinas/farmacologia , Comportamento Alimentar/fisiologia , Expressão Gênica , Genes de Insetos , Proteínas Hemolisinas/farmacologia , Proteínas de Insetos/genética , Proteínas de Insetos/metabolismo , Proteínas de Insetos/farmacologia , Mariposas/efeitos dos fármacos , Mariposas/genética , Mariposas/metabolismo , Mariposas/fisiologia , Neuropeptídeos/genética , Neuropeptídeos/metabolismo , Neuropeptídeos/farmacologia , Controle de Pragas/métodos
6.
Arch Insect Biochem Physiol ; 108(3): e21843, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34490676

RESUMO

Gossypol is a toxic sesquiterpene dimer produced by cotton plants which deters herbivory by insects and vertebrates. Two highly reactive aldehyde groups contribute to gossypol toxicity by cross-linking herbivore proteins. We identified another consequence of consuming gossypol in two insect pests of cotton: increased amounts of fatty acid-amino acid conjugates (FACs). Eight different FACs in the feces of larval Helicoverpa armigera and Heliothis virescens increased when larvae consumed artificial diet containing gossypol, but not a gossypol derivative lacking free aldehyde groups (SB-gossypol). FACs are produced by joining plant-derived fatty acids with amino acids of insect origin in the larval midgut tissue by an unknown conjugase, and translocated into the gut lumen by an unknown transporter. FACs are hydrolyzed back into fatty acids and amino acids by an aminoacylase (L-ACY-1) in the gut lumen. The equilibrium level of FACs in the lumen is determined by a balance between conjugation and hydrolysis, which may differ among species. When heterologously expressed, L-ACY-1 of H. armigera but not H. virescens was inhibited by gossypol; consistent with the excretion of more FACs in the feces by H. armigera. FACs are known to benefit the plant host by inducing anti-herbivore defensive responses, and have been hypothesized to benefit the herbivore by acting as a surfactant and increasing nitrogen uptake efficiency. Thus in addition to its direct toxic effects, gossypol may negatively impact insect nitrogen uptake efficiency and amplify the signal used by the plant to elicit release of volatile compounds that attract parasitoids.


Assuntos
Amidoidrolases/metabolismo , Ácidos Graxos/metabolismo , Gossipol/farmacologia , Mariposas , Defesa das Plantas contra Herbivoria , Amidoidrolases/efeitos dos fármacos , Aminoácidos/metabolismo , Animais , Proteínas de Insetos/efeitos dos fármacos , Proteínas de Insetos/metabolismo , Larva/efeitos dos fármacos , Larva/metabolismo , Mariposas/efeitos dos fármacos , Mariposas/metabolismo
7.
Arch Insect Biochem Physiol ; 108(3): e21842, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34499777

RESUMO

Glyphodes pyloalis Walker has become one of the most significant mulberry pests, and it has caused serious economic losses in major mulberry growing regions in China. Peptidoglycan recognition proteins (PGRPs) are responsible for initiating and regulating immune signalling pathways in insects. However, their roles responding to chemical pesticides is still less known. This study aimed to investigate the possible detoxication function of GpPGRP-S2 and GpPGRP-S3 in G. pyloalis in response to chlorfenapyr and phoxim. The chlorfenapyr and phoxim treatment significantly induced the expression level of GpPGRP-S3 at 48 h. In addition, the expression levels of GpPGRP-S2 and GpPGRP-S3 in the chlorfenapyr/phoxim treatment group were significantly higher in midgut than those in the control group at 48 h. The results of the survival experiment showed that silencing either GpPGRP-S2 or GpPGRP-S3 would not influence the survival rate of G. pyloalis which treated with phoxim, however, silencing GpPGRP-S2 or GpPGRP-S3 would cause G. pyloalis to be more easily killed by chlorfenapyr. The expression of carboxylesterase GpCXE1 was significantly induced by chlorfenapyr/phoxim treatment, while it was suppressed once silenced GpPGRP-S2 followed with chlorfenapyr treatment or silenced GpPGRP-S3 followed with phoxim treatment. These results might suggest that under the chlorfenapyr/phoxim treatment condition, the connection between GpPGRPs and detoxification genes in insect was induced to maintain physiological homeostasis; and these results may further enrich the mechanisms of insects challenged by insecticides.


Assuntos
Proteínas de Transporte , Inseticidas , Mariposas , Animais , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Inseticidas/metabolismo , Inseticidas/farmacologia , Mariposas/efeitos dos fármacos , Mariposas/genética , Mariposas/metabolismo , Compostos Organotiofosforados/metabolismo , Compostos Organotiofosforados/farmacologia , Controle de Pragas/métodos , Piretrinas/metabolismo , Piretrinas/farmacologia
8.
J Agric Food Chem ; 69(39): 11582-11591, 2021 Oct 06.
Artigo em Inglês | MEDLINE | ID: mdl-34555899

RESUMO

The ionotropic γ-aminobutyric acid (iGABA) receptor is commonly considered as a fast inhibitory channel and is an important insecticide target. Since 1990, RDL, LCCH3, and GRD have been successively isolated and found to be potential subunits of the insect iGABA receptor. More recently, one orphan gene named 8916 was found and considered to be another potential iGABA receptor subunit according to its amino acid sequence. However, little information about 8916 has been reported. Here, the 8916 subunit from Chilo suppressalis was studied to determine whether it can form part of a functional iGABA receptor by co-expressing this subunit with CsRDL1 or CsLCCH3 in the Xenopus oocyte system. Cs8916 or CsLCCH3 did not form functional ion channels when expressed alone. However, Cs8916 was able to form heteromeric ion channels when expressed with either CsLCCH3 or CsRDL1. The recombinant heteromeric Cs8916/LCCH3 channel was a cation-selective channel, which was sensitive to GABA or ß-alanine. The current of the Cs8916/LCCH3 channel was inhibited by dieldrin, endosulfan, fipronil, or ethiprole. In contrast, fluralaner, broflanilide, and avermectin showed little effect on the Cs8916/LCCH3 channel (IC50s > 10 000 nM). The Cs8916/RDL1 channel was sensitive to GABA, but was significantly different in EC50 and Imax for GABA to those of homomeric CsRDL1. Fluralaner, fipronil, or dieldrin showed antagonistic actions on Cs8916/RDL1. In conclusion, Cs8916 is a potential iGABA receptor subunit, which can interact with CsLCCH3 to generate a cation-selective channel that is sensitive to several insecticides. Also, as Cs8916/RDL1 has a higher EC50 than homomeric CsRDL1, Cs8916 may affect the physiological functions of CsRDL1 and therefore play a role in fine-tuning GABAergic signaling.


Assuntos
Inseticidas , Mariposas , Sequência de Aminoácidos , Animais , Inseticidas/farmacologia , Mariposas/metabolismo , Receptores de GABA/genética , Receptores de GABA/metabolismo , Receptores de GABA-A , Ácido gama-Aminobutírico
9.
PLoS Pathog ; 17(9): e1009917, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34495986

RESUMO

Host-pathogen interactions are central components of ecological networks where the MAPK signaling pathways act as central hubs of these complex interactions. We have previously shown that an insect hormone modulated MAPK signaling cascade participates as a general switch to trans-regulate differential expression of diverse midgut genes in the diamondback moth, Plutella xylostella (L.) to cope with the insecticidal action of Cry1Ac toxin, produced by the entomopathogenic bacterium Bacillus thuringiensis (Bt). The relationship between topology and functions of this four-tiered phosphorylation signaling cascade, however, is an uncharted territory. Here, we carried out a genome-wide characterization of all the MAPK orthologs in P. xylostella to define their phylogenetic relationships and to confirm their evolutionary conserved modules. Results from quantitative phosphoproteomic analyses, combined with functional validations studies using specific inhibitors and dsRNAs lead us to establish a MAPK "road map", where p38 and ERK MAPK signaling pathways, in large part, mount a resistance response against Bt toxins through regulating the differential expression of multiple Cry toxin receptors and their non-receptor paralogs in P. xylostella midgut. These data not only advance our understanding of host-pathogen interactions in agricultural pests, but also inform the future development of biopesticides that could suppress Cry resistance phenotypes.


Assuntos
Infecções por Bactérias Gram-Positivas/metabolismo , Interações Hospedeiro-Patógeno/fisiologia , Sistema de Sinalização das MAP Quinases/fisiologia , Mariposas/metabolismo , Mariposas/microbiologia , Animais , Bacillus thuringiensis , Toxinas de Bacillus thuringiensis/metabolismo , Endotoxinas/metabolismo , Infecções por Bactérias Gram-Positivas/veterinária , Proteínas Hemolisinas/metabolismo , Resistência a Inseticidas
10.
Insect Biochem Mol Biol ; 138: 103636, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34478812

RESUMO

There are several known non-molting mutations of the silkworm, Bombyx mori, including non-molting dwarf (nm-d). Larvae with this mutation hatch normally and start eating leaves, but die before the completion of the first ecdysis. Genetic analysis of the nm-d mutation would contribute to the isolation of essential genes for the larval development of lepidopteran insects. To identify the causative gene of the nm-d locus, we conducted RNA-seq based rough mapping. Using two sets of RNA-seq data, one from a pooled sample of normal larvae, and one from a pooled sample of nm-d larvae, the nm-d locus was narrowed to a 500 kb region. Among the genes located in this region, a nm-d-specific exon loss was identified in the Bombyx homolog of the ATIC (5-aminoimidazole-4-carboxamide ribonucleotide transformylase/Inosine 5'-monophosphate cyclohydrolase) (BmATIC) gene, which catalyzes the final two steps of the de novo purine biosynthetic pathway in mammals. PCR and subsequent sequencing analysis revealed that a region containing exon 9 of the BmATIC gene is deleted in the nm-d larvae. A knockout allele of the BmATIC gene (BmATICKO), that was generated using the CRISPR/Cas9 system, revealed that first instar knockout larvae died while exhibiting the dark brown larval body that is a typical feature of mutants that lack uric acid in the integument. Lethal larvae resulted from crosses between +/BmATICKO moths. The uric acid content in the whole-body of the first instar was drastically reduced in the nm-d larvae compared to normal larvae. These results indicated that the BmATIC gene is responsible for the nm-d phenotype, and that nm-d larvae have a defect in purine biosynthesis, including uric acid. We also discuss the possibility that the BmATIC mRNA is maternally transmitted to eggs. Our results indicated that RNA-seq based mapping using pooled samples is a practical method for the identification of the causative genes of lethal mutations.


Assuntos
Proteínas de Insetos/genética , Mariposas/metabolismo , Mutação , Purinas/biossíntese , Animais , Proteínas de Insetos/metabolismo , Larva/genética , Larva/crescimento & desenvolvimento , Larva/metabolismo , Mariposas/genética , Mariposas/crescimento & desenvolvimento
11.
Molecules ; 26(15)2021 Jul 28.
Artigo em Inglês | MEDLINE | ID: mdl-34361721

RESUMO

To improve the proinsecticidal activity and phloem mobility of amino acid-tralopyril conjugates further, nine conjugates were designed and synthesized by introducing glutamic acid to tralopyril, and the length of the linker between glutamic acid and tralopyril ranged from 2 atoms to 10 atoms. The results of insecticidal activity against the third-instar larvae of P. xylostella showed that conjugates 42, 43, 44,and 45 (straight-chain containing 2-5 atoms) exhibited good insecticidal activity, and their LC50 values were 0.2397 ± 0.0366, 0.4413 ± 0.0647, 0.4400 ± 0.0624, and 0.4602 ± 0.0655 mM, respectively. The concentrations of conjugates 43-45 were higher than that of conjugate 42 in the phloem sap at 2 h, and conjugate 43 showed the highest concentration. The introduction of glutamic acid can improve phloem mobility. The in vivo metabolism of conjugates 42 and 43 was investigated in P. xylostella, and the parent compound tralopyril was detected at concentrations of 0.5950 and 0.3172 nmol/kg, respectively. According to the above results, conjugates 42 and 43 were potential phloem mobile pro-insecticide candidates.


Assuntos
Ácido Glutâmico/química , Inseticidas/farmacologia , Larva/efeitos dos fármacos , Mariposas/efeitos dos fármacos , Pró-Fármacos/farmacologia , Pirróis/farmacologia , Animais , Transporte Biológico , Biotransformação , Concentração Inibidora 50 , Inseticidas/síntese química , Inseticidas/metabolismo , Larva/crescimento & desenvolvimento , Larva/metabolismo , Longevidade/efeitos dos fármacos , Mariposas/crescimento & desenvolvimento , Mariposas/metabolismo , Floema/metabolismo , Floema/parasitologia , Plantas/metabolismo , Plantas/parasitologia , Pró-Fármacos/síntese química , Pró-Fármacos/metabolismo , Piretrinas/metabolismo , Piretrinas/farmacologia , Pirróis/síntese química , Pirróis/metabolismo , Relação Estrutura-Atividade
12.
Sci Rep ; 11(1): 15963, 2021 08 05.
Artigo em Inglês | MEDLINE | ID: mdl-34354188

RESUMO

One group of promising pest control agents are the entomopathogenic fungi; one such example is Conidiobolus coronatus, which produces a range of metabolites. Our present findings reveal for the first time that C. coronatus also produces dodecanol, a compound widely used to make surfactants and pharmaceuticals, and enhance flavors in food. The main aim of the study was to determine the influence of dodecanol on insect defense systems, i.e. cuticular lipid composition and the condition of insect immunocompetent cells; hence, its effect was examined in detail on two species differing in susceptibility to fungal infection: Galleria mellonella and Calliphora vicina. Dodecanol treatment elicited significant quantitative and qualitative differences in cuticular free fatty acid (FFA) profiles between the species, based on gas chromatography analysis with mass spectrometry (GC/MS), and had a negative effect on G. mellonella and C. vicina hemocytes and a Sf9 cell line in vitro: after 48 h, almost all the cells were completely disintegrated. The metabolite had a negative effect on the insect defense system, suggesting that it could play an important role during C. coronatus infection. Its high insecticidal activity and lack of toxicity towards vertebrates suggest it could be an effective insecticide.


Assuntos
Conidiobolus/metabolismo , Dodecanol/metabolismo , Dodecanol/farmacologia , Animais , Calliphoridae , Conidiobolus/química , Conidiobolus/patogenicidade , Ácidos Graxos/química , Ácidos Graxos/metabolismo , Fungos/química , Fungos/metabolismo , Cromatografia Gasosa-Espectrometria de Massas/métodos , Hemócitos/metabolismo , Imunidade Celular/efeitos dos fármacos , Imunidade Celular/imunologia , Insetos/metabolismo , Inseticidas , Larva/metabolismo , Mariposas/metabolismo
13.
Cell Mol Life Sci ; 78(19-20): 6593-6603, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34448011

RESUMO

The concept of reverse chemical ecology (exploitation of molecular knowledge for chemical ecology) has recently emerged in conservation biology and human health. Here, we extend this concept to crop protection. Targeting odorant receptors from a crop pest insect, the noctuid moth Spodoptera littoralis, we demonstrate that reverse chemical ecology has the potential to accelerate the discovery of novel crop pest insect attractants and repellents. Using machine learning, we first predicted novel natural ligands for two odorant receptors, SlitOR24 and 25. Then, electrophysiological validation proved in silico predictions to be highly sensitive, as 93% and 67% of predicted agonists triggered a response in Drosophila olfactory neurons expressing SlitOR24 and SlitOR25, respectively, despite a lack of specificity. Last, when tested in Y-maze behavioral assays, the most active novel ligands of the receptors were attractive to caterpillars. This work provides a template for rational design of new eco-friendly semiochemicals to manage crop pest populations.


Assuntos
Mariposas/efeitos dos fármacos , Mariposas/metabolismo , Receptores Odorantes/metabolismo , Animais , Drosophila/efeitos dos fármacos , Drosophila/metabolismo , Proteínas de Insetos/metabolismo , Repelentes de Insetos/farmacologia , Aprendizado de Máquina , Odorantes , Feromônios/farmacologia , Olfato/efeitos dos fármacos , Spodoptera/efeitos dos fármacos , Spodoptera/metabolismo
14.
Arch Insect Biochem Physiol ; 108(2): e21837, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34293199

RESUMO

The olfactory system must detect and discriminate various semiochemicals in the environment. In response to such diversity, insects have evolved a family of odorant-gated ion channels composed of a common receptor (coreceptor, Orco) and a ligand-binding tuning odorant receptor (OR) that confers odour specificity. This study aims to examine the expression pattern of Orco gene of Grapholita molesta (GmolOrco) and to elucidate the role of GmolOrco in detecting G. molesta sex pheromone and green leaf volatiles by using gene silencing via RNA interference (RNAi) coupled antennal electrophysiological (EAG). Multiple sequence alignment showed that GmolOrco shared high sequence similarities with the Orco ortholog of lepidopterans. The results of real-time quantitative PCR detection demonstrated that GmolOrco was predominantly expressed in adult antennae and had the highest expression quantity in adult period among the different developmental stages. Compared with the noninjected controls, GmolOrco expression in GmolOrcodouble-stranded RNA (dsRNA)-injected males was reduced to 39.92% and that in females was reduced to 40.43%. EAG assays showed that the responses of GmolOrco-dsRNA injected males to sex pheromones (Z)-8-dodecenyl acetate (Z8-12:OAc) and (Z)-8-dodecenyl alcohol (Z8-12:OH) were significantly reduced, and the GmolOrco-dsRNA-injected female to green leaf volatile (Z)-3-hexenyl acetate also significantly declined. We inferred that Orco-mediated olfaction was different in male and female G. molesta adults and was mainly involved in recognizing the sex pheromones released by female moths.


Assuntos
Mariposas , Receptores Odorantes , Animais , Antenas de Artrópodes/metabolismo , Feminino , Expressão Gênica , Genes de Insetos , Proteínas de Insetos/genética , Proteínas de Insetos/metabolismo , Masculino , Mariposas/genética , Mariposas/metabolismo , Mariposas/fisiologia , Odorantes , Plantas/metabolismo , Interferência de RNA , Receptores Odorantes/genética , Receptores Odorantes/metabolismo , Atrativos Sexuais/metabolismo , Caracteres Sexuais , Compostos Orgânicos Voláteis/metabolismo
15.
Int J Mol Sci ; 22(13)2021 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-34281211

RESUMO

To reveal the mechanism of temperature preference in Tuta absoluta, one of the top 20 plant pests in the world, we cloned and identified TaTRPA1, TaPain, and TaPyx genes by RACE and bioinformatic analysis, and clarified their expression profiles during different development stages using real-time PCR, and revealed their function in preference temperature by RNAi. The full-length cDNA of TaPain was 3136 bp, with a 2865-bp open reading frame encoding a 259.89-kDa protein; and the partial length cDNA of TaPyx was 2326-bp, with a 2025-bp open reading frame encoding a 193.16-kDa protein. In addition, the expression of TaTRPA1 and TaPyx was significantly lower in larvae than other stages, and it was significantly higher in pupae and newly emerging males for TaPain. After feeding target double-stranded RNA (dsRNA), the preferred temperature decreased 2 °C more than the control group. In conclusion, the results firstly indicated the molecular characterization of TRPA subfamily genes and their key role in temperature perception in T. absoluta, and the study will help us to understand the temperature-sensing mechanism in the pest, and will provide some basis for study of other Lepidoptera insects' temperature preference. Moreover, it is of great significance in enriching the research progress of "thermos TRP".


Assuntos
Mariposas/genética , Canal de Cátion TRPA1/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Comportamento Animal , Feminino , Larva/metabolismo , Masculino , Mariposas/metabolismo , Pupa/metabolismo , Temperatura
16.
Sci Rep ; 11(1): 14027, 2021 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-34234208

RESUMO

Insect olfaction system plays a key role in the foraging food, pollination, mating, oviposition, reproduction and other insect physiological behavior. Odorant binding protein are widely found in the various olfactory sensilla of different insect antennae and involved in chemical signals discrimination from natural environment. In this study, a novel OBP gene, MvitOBP3 is identified from the legume pod borer, Maruca vitrata, which it mainly harms important legume vegetables including cowpea, soybean and lablab bean. Real-time PCR results demonstrated that MvitOBP3 gene was abundantly expressed in the antennal tissue of M. vitrata, while low levels were distributed in the head, thorax, abdomen, leg and wing of adult moths. The recombinant OBP3 protein was purified using the prokaryotic expression and affinity chromatography system. Fluorescence competitive binding experiments indicated that that MvitOBP3 protein exhibited greater binding affinities with host-plant flower volatiles including Butanoic acid butyl ester, Limonene, 1H-indol-4-ol and 2-methyl-3-phenylpropanal, highlighting they may have attractant activities for the oviposition of female moths on the legume vegetables. Moreover, protein homology modeling and molecular docking analysis revealed that there are six amino acid sites of MvitOBP3 involved in the binding of the host-plant volatiles. These findings will further promote to understand the key role of odorant binding protein during host perception and oviposition of M. vitrata moths, which improve the efficiency of semiochemical-based prevention and monitoring for this pest in the legume vegetables field.


Assuntos
Fabaceae/parasitologia , Mariposas/genética , Mariposas/metabolismo , Receptores Odorantes/genética , Receptores Odorantes/metabolismo , Animais , Expressão Gênica , Proteínas de Insetos/química , Proteínas de Insetos/genética , Proteínas de Insetos/metabolismo , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Estrutura Molecular , Mariposas/classificação , Odorantes , Filogenia , Ligação Proteica , Receptores Odorantes/química , Proteínas Recombinantes , Análise de Sequência de DNA , Relação Estrutura-Atividade , Compostos Orgânicos Voláteis/metabolismo
17.
Pest Manag Sci ; 77(10): 4593-4606, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34092019

RESUMO

BACKGROUND: Broad use of insecticidal Cry proteins from Bacillus thuringiensis in biopesticides and transgenic crops has resulted in cases of practical field resistance, highlighting the need for novel approaches to insect control. Previously we described an anti-Cry1Ab idiotypic-antibody (B12-scFv) displaying toxicity against rice leafroller (Cnaphalocrocis medinalis) larvae, supporting the potential of antibodies for pest control. The goal of the present study was to generate insecticidal antibodies against diamondback moth (Plutella xylostella) larvae. RESULTS: Four genetically engineered antibodies (GEAbs) were designed in silico from B12-scFv using three-dimensional (3D) structure and docking predictions to alkaline phosphatase (ALP) as a Cry1Ac receptor in P. xylostella. Among these GEAbs, the GEAb-dVL antibody consisting of two light chains had overlapping binding sites with Cry1A and Cry1B proteins and displayed high binding affinity to P. xylostella midgut brush border membrane (BBM) proteins. Proteins in BBM identified by pull-down assays as binding to GEAb-dVL included an ABC transporter and V-ATPase subunit A protein. Despite lacking the α-helical structures in Cry1A that are responsible for pore formation, ingestion of GEAb-dVL disrupted the P. xylostella larval midgut epithelium and resulted in toxicity. Apoptotic genes were activated in gut cells upon treatment with GEAb-dVL . CONCLUSION: This study describes the first insecticidal GEAb targeting P. xylostella by mimicking Cry proteins. Data support that GEAb-dVL toxicity is associated to activation of intracellular cell death pathways, in contrast to pore-formation associated toxicity of Cry proteins. This work provides a foundation for the design of novel insecticidal antibodies for insect control. © 2021 Society of Chemical Industry.


Assuntos
Bacillus thuringiensis , Inseticidas , Mariposas , Animais , Bacillus thuringiensis/genética , Bacillus thuringiensis/metabolismo , Toxinas de Bacillus thuringiensis , Proteínas de Bactérias/metabolismo , Sítios de Ligação , Endotoxinas/metabolismo , Proteínas Hemolisinas/metabolismo , Proteínas de Insetos/genética , Proteínas de Insetos/metabolismo , Inseticidas/farmacologia , Larva/metabolismo , Mariposas/metabolismo , Ligação Proteica
18.
Artigo em Inglês | MEDLINE | ID: mdl-34082360

RESUMO

Chemosensory proteins (CSPs) are important for insect chemoreception, which bind, solubilize and transport hydrophobic chemical molecules from external environment to dendrite membrane of chemosensory neurons. Moreover, CSPs are also involved in non-sensory physiological activities. The peach fruit borers Carposina sasakii Matsumura (Lepidoptera: Carposinidae) seriously damage fruit trees and their chemoreception mainly occurs in the adult stage. We identified 10 putative CSPs (CsasCSP1 ~ CsasCSP10) from head transcriptomes of C. sasakii adult males and females, all of which are classic CSPs that have 4 conserved cysteines with a spacing pattern C1-X6-C2-X17-18-C3-X2-C4. Their phylogenetic characteristics were also described. An analysis using fluorescence quantitative PCR showed CsasCSP2 has the highest level of expression in the heads, so it is more likely to be involved in C. sasakii chemoreception than the other C. sasakii CSPs. CsasCSP1, CsasCSP3, CsasCSP4, CsasCSP6, CsasCSP7 and CsasCSP8 are expressed dominantly in the wings; CsasCSP5 and CsasCSP10 have the highest expression level in the thoraxes; CsasCSP9 is dominantly and equally expressed in the thoraxes and abdomens. This study contributes to understanding physiological functions of C. sasakii CSPs and chemosensory mechanism at C. sasakii molecular level.


Assuntos
Regulação da Expressão Gênica , Proteínas de Insetos/metabolismo , Larva/metabolismo , Mariposas/metabolismo , Transcriptoma , Animais , Frutas/parasitologia , Proteínas de Insetos/genética , Larva/genética , Larva/crescimento & desenvolvimento , Mariposas/genética , Mariposas/crescimento & desenvolvimento , Filogenia , Prunus persica/parasitologia
19.
Insect Biochem Mol Biol ; 135: 103597, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34089822

RESUMO

Lepidopteran P450s of the CYP6B and CYP9A subfamilies are thought to play important roles in host plant adaptation and insecticide resistance. An increasing number of paralogs and orthologs with high levels of sequence identity have been found in these subfamilies by mining recent genome projects. However, the biochemical function of most of them remains unknown. A better understanding of the evolution of P450 genes and of the catalytic competence of the enzymes they encode is needed to facilitate studies of host plant use and insecticide resistance. Here, we focused on the full complement of CYP6B (4 genes) and CYP9A (7 genes) in the generalist herbivore, Helicoverpa armigera. These P450s were heterologously expressed in Sf9 cells and compared functionally. In vitro assays showed that all CYP6B and CYP9A P450s can metabolize esfenvalerate efficiently, except for the evolutionarily divergent CYP6B43. A new 2'-hydroxy-metabolite of esfenvalerate was identified and found to be the main metabolite produced by CYP9A12. All tested P450s showed only low induction responses to esfenvalerate. To put these results from H. armigera P450s in perspective, 158 complete CYP6B and 100 complete CYP9A genes from 34 ditrysian species were manually curated. The CYP9A subfamily was more widespread than the CYP6B subfamily and the latter showed dramatic gains and losses, with ten species lacking CYP6B genes. Two adjacent CYP6B loci were found on chromosome 21, with different fates during the evolution of Lepidoptera. The diversity and functional redundancy of CYP6B and CYP9A genes challenge resistance management and pest control strategies as many P450s are available to insects to cope with chemical stresses they encounter.


Assuntos
Sistema Enzimático do Citocromo P-450 , Resistência a Inseticidas/genética , Mariposas , Piretrinas , Animais , Sistema Enzimático do Citocromo P-450/efeitos dos fármacos , Sistema Enzimático do Citocromo P-450/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Genes de Insetos , Genoma de Inseto , Controle de Insetos/métodos , Proteínas de Insetos/efeitos dos fármacos , Proteínas de Insetos/genética , Proteínas de Insetos/metabolismo , Inseticidas/metabolismo , Inseticidas/farmacologia , Mariposas/efeitos dos fármacos , Mariposas/genética , Mariposas/metabolismo , Filogenia , Piretrinas/metabolismo , Piretrinas/farmacologia
20.
Artigo em Inglês | MEDLINE | ID: mdl-34171685

RESUMO

The diamondback moth, Plutella xylostella (Lepidoptera: Plutellidae) is a holometabolous insect that its cuticles must undergo the significant changes during the larval-pupal metamorphosis development. To elucidate these changes at molecular levels, RNA-seq analysis of cuticles from LLS (later fourth instar larval stage), PPS (prepupal stage) and PS (pupal stage) were performed in P. xylostella. In this paper, a total of 17,710 transcripts were obtained in the larval-pupal transition of P. xylostella, and out of which 2293 (881 up-regulated and 1412 down-regulated) and 2989 transcripts (2062 up-regulated and 927 down-regulated) were identified to be differentially expressed between LLS and PPS, as well as PPS and PS, respectively. The further GO and KEGG analysis of differentially expressed genes (DEGs) revealed that the 'structural constituent of cuticle', 'chitin metabolic process', 'chitin binding', 'tyrosine metabolism' and 'insect hormone biosynthesis' pathways were significantly enriched, indicating these pathways might be involved in the process of larval pupation in P. xylostella. Then, we found some genes that encoded cuticular proteins, chitinolytic enzymes, chitin synthesis enzymes, and cuticle tanning proteins changed their expression levels remarkably, indicating these genes might play important roles in the restruction (degradation and biosynthesis) of insect cuticles during the larval metamorphosis. Additionally, the significant changes in the mRNA levels of 20-hydroxyecdysone (20E) and juvenile hormone (JH) related genes suggested their crucial roles in regulating cuticle remodeling during the larval metamorphosis of P. xylostella. In conclusion, the present study provide us the comprehensive gene expression profiles to explore the molecular mechanisms of cuticle metamorphosis in P. xylostella, which laid a molecular basis to study roles of specific pathways and genes in insect development.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Insetos/metabolismo , Larva/crescimento & desenvolvimento , Metamorfose Biológica , Mariposas/crescimento & desenvolvimento , Pupa/crescimento & desenvolvimento , RNA-Seq/métodos , Animais , Proteínas de Insetos/genética , Larva/genética , Larva/metabolismo , Mariposas/genética , Mariposas/metabolismo , Pupa/genética , Pupa/metabolismo , Transcriptoma
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