Ethyl 5-cyano-1,6-dihydro-2-methyl-4-(2\'-thienyl)-6-thioxonicotinate (A) was synthesized and reacted with ethyl chloroacetate in the presence of sodium acetate or sodium carbonate to give ethyl 5-cyano-6-((2-ethoxy-2-oxoethyl)thio)-2-methyl-4-(2\'-thienyl)nicotinate (1a) or its isomeric thieno[2,3-b]pyridine 2a. 3-Aminothieno[2,3-b]pyridine-2-carboxamide 2b was also synthesized by the reaction of A with 2-chloroacetamide. The reaction of 1a with hydrazine hydrate in boiling ethanol gave acethydrazide 3. Heating ester 1a with hydrazine hydrate under neat conditions afforded 3-amino-1H-pyrazolo[3,4-b]pyridine 10. Compounds 2b, 3, and 10 were used as precursors for synthesizing other new thieno[2,3-b]pyridines and pyrazolo[3,4-b]pyridines containing mainly the ethyl nicotinate scaffold. Structures of all new compounds were confirmed by elemental and spectral analyses. Most of the obtained compounds were evaluated for their insecticidal activity toward the nymphs and adults of Aphis gossypii (Glover,1887). Some compounds such as 4, 9b, and 9c showed promising results. The effect of some sublethal concentrations, less than LC50, of compounds 4, 9b, and 9c on the examined Aphis was subjected to a further study. The results demonstrated that exposure of A. gossypii nymphs to sublethal concentrations of compounds 4, 9b, and 9c had noticeable effects on their biological parameters, i.e., nymphal instar duration, generation time, and adult longevity. The highest concentration C1 of all three compounds increased the nymphal instar duration and generation time and decreased adult longevity and vice versa.

Tropical and subtropical crops are being increasingly cultivated in South Korea, leading to an increase in damage by exotic insect pests. Consequently, ethyl formate (EF) is currently being considered for quarantine and pre-shipment fumigation. In this study, we evaluated the effectiveness of EF fumigation for controlling Aphis spiraecola Patch and Aphis gossypii Glover, two representative quarantine pests on passion fruit (\"Pink Bourbon\") during greenhouse cultivation and post-harvest storage. The efficacy of EF against both aphids in terms of the lethal concentration causing 50% mortality (LCt50%) and LCt99% was 1.36-2.61 g h/m3 and 3.73-7.55 g h/m3 under greenhouse conditions (23 °C), and 1.37-2.02 g h/m3 and 3.80-14.59 g h/m3 post-harvest (5 °C), respectively. EF at 4 g/m3 for 4 h resulted in 100% mortality of A. spiraecola, which was more resistant to EF, without causing phytotoxic damage to the trees in a 340 m3 greenhouse. Post-harvest fruit fumigation at 10 g/m3 for 4 h in a mid-size (0.8 m3) fumigation chamber resulted in complete disinfection. Moreover, the EF level decreased below the EF threshold within 10 min after natural ventilation in the greenhouse. Therefore, our results suggest EF fumigation as an effective method for controlling A. spiraecola and A. gossypii.

Negeviruses are insect-specific enveloped RNA viruses that exhibit a wide geographic distribution. A novel nege-like virus, tentatively named Aphis gossypii nege-like virus (AGNLV, GenBank: OR880429.1), was isolated from aphids (Aphis gossypii) in Lijiang City, Yunnan, China. AGNLV has a genome sequence of 9258 nt (excluding the polyA tail) encoding three open reading frames (ORFs). ORF1 (7149 nt) encodes a viral methyltransferase, a viral RNA helicase, and an RNA-dependent RNA polymerase. ORF2 (1422 nt) encodes a DiSB-ORF2_chro domain and ORF3 encodes an SP24 domain. The genome sequence of AGNLV shares the highest nucleotide identity of 60.0% and 59.5% with Wuhan house centipede virus 1 (WHCV1) and Astegopteryx formosana nege-like virus (AFNLV), respectively. Phylogenetic analysis based on the RNA-dependent RNA polymerase shows that AGNLV is clustered with other negeviruses and nege-like viruses discovered in aphids, forming a distinct \"unclassified clade\". Interestingly, AGNLV only encodes three ORFs, whereas AFNLV and WHCV1 have four ORFs. Structure and transmembrane domain predictions show the presence of eight alpha helices and five transmembrane helices in the AGNLV ORF3. Translational enhancement of the AGNLV 5\' UTR was similar to that of the 5\' UTR of plant viruses. Our findings provide evidence of the diversity and structure of nege-like viruses and are the first record of such a virus from a member of the genus Aphis.

The cotton aphid, Aphis gossypii, is a polyphagous pest that stunts host plant growth via direct feeding or transmitting plant virus. Due to the long-term application of insecticides, A. gossypii has developed different levels of resistance to numerous insecticides. We found that five field populations had evolved multiple resistances to neonicotinoids. To explore the resistance mechanism mediated by uridine diphosphate glycosyltransferases (UGTs), two upregulated UGT genes in these five strains, UGT350C3 and UGT344L7, were selected for functional analysis of their roles in neonicotinoid detoxification. Transgenic Drosophila bioassay results indicated that compared with the control lines, the UGT350C3 and UGT344L7 overexpression lines were more tolerant to thiamethoxam, imidacloprid, and dinotefuran. Knockdown of UGT350C3 and UGT344L7 significantly increased A. gossypii sensitivity to thiamethoxam, imidacloprid, and dinotefuran. Molecular docking analysis demonstrated that these neonicotinoids could bind to the active pockets of UGT350C3 and UGT344L7. This study provides functional evidence of neonicotinoid detoxification mediated by UGTs and will facilitate further work to identify strategies for preventing the development of neonicotinoid resistance in insects.

Aphids are sap-sucking insects responsible for crop losses and a severe threat to crop production. Proteins in the aphid saliva are integral in establishing an interaction between aphids and plants and are responsible for host plant adaptation. The cotton aphid, Aphis gossypii (Hemiptera: Aphididae) is a major pest of Gossypium hirsutum. Despite extensive studies of the salivary proteins of various aphid species, the components of A. gossypii salivary glands are unknown. In this study, we identified 123,008 transcripts from the salivary gland of A. gossypii. Among those, 2933 proteins have signal peptides with no transmembrane domain known to be secreted from the cell upon feeding. The transcriptome includes proteins with more comprehensive functions such as digestion, detoxification, regulating host defenses, regulation of salivary glands, and a large set of uncharacterized proteins. Comparative analysis of salivary proteins of different aphids and other insects with A. gossypii revealed that 183 and 88 orthologous clusters were common in the Aphididae and non-Aphididae groups, respectively. The structure prediction for highly expressed salivary proteins indicated that most possess an intrinsically disordered region. These results provide valuable reference data for exploring novel functions of salivary proteins in A. gossypii with their host interactions. The identified proteins may help develop a sustainable way to manage aphid pests.

Ageratina adenophora can enhance its invasive ability by using beneficial rhizosphere bacteria. Bacillus cereus is able to promote plant growth and provide a positive feedback effect to A. adenophora. However, the interaction between A. adenophora and B. cereus under the influence of native polyphagous insect feeding is still unclear. In this study, Eupatorium lindleyanum, a local species closely related to A. adenophora, was used as a control, aimed to compare the content of B. cereus in the roots of A. adenophora and rhizosphere soil after different densities of Aphis gossypii feeding, and then investigated the variations in the population of A. gossypii and soil characteristics after the addition of B. cereus. The result showed that B. cereus content in the rhizosphere soil and root of A. adenophora increased significantly under A. gossypii feeding compared with local plants, which also led to the change of α-diversity and β-diversity of the bacterial community, as well as the increase in nitrate nitrogen (NO3 -N) content. The addition of B.cereus in the soil could also inhibit the population growth of A. gossypii on A. adenophora and increase the content of ammonium nitrogen (NH4 +-N) in the soil. Our research demonstrated that B. cereus enhances the ability of A. adenophora to resist natural enemy by increasing soil ammonium nitrogen (NH4 +-N) and accumulating other beneficial bacteria, which means that rhizosphere microorganisms help invasive plants defend themselves against local natural enemies by regulating the soil environment.

The toxicity and sublethal effects of three insecticides (spirotetramat, cyantraniliprole, and pymetrozine) on Aphis gossypii, a major agricultural pest, were investigated. The nymphal stage showed greater susceptibility than the adult stage to all the insecticides, with a difference of up to 8.9 times at the LC50 of spirotetramat. The effects of sublethal concentrations (LC10, LC30, LC50, and LC70) of the insecticides on the on the developmental period, survival rate, adult longevity, fecundity, and deformity rate were compared with those of the control. Compared with the control, cyantraniliprole and pymetrozine did not significantly affect the developmental period in the parental or F1 generation when applied at the nymphal stage at any concentration. Nonviable nymphs occurred in the F1 generation when both nymphs and adults were treated with spirotetramat and cyantraniliprole but not in the F2 generation. The age-specific maternity (lxmx) of A. gossypii treated with sublethal concentrations (LC10, LC30) decreased with increasing concentration. Spirotetramat at the LC30 resulted in significant differences in all life table parameters (R0, rm, λ, T, DT) compared with those of the control. Similarly, compared with that of the control (43.8), the net reproductive rate (R0) significantly decreased for all the insecticides except cyantraniliprole at the LC10 (37.5). Therefore, this study indicated that sublethal concentrations (over the LC30) of spirotetramat, cyantraniliprole, or pymetrozine might be useful for the density management of A. gossypii.

Upon acquisition of persistent circulative viruses such as poleroviruses, the virus particles transcytose through membrane barriers of aphids at the midgut and salivary glands via hemolymph. Such intricate interactions can influence aphid behavior and fitness and induce associated gene expression in viruliferous aphids. Differential gene expression can be evaluated by omics approaches such as transcriptomics. Previously conducted aphid transcriptome studies used only one host species as the source of virus inoculum. Viruses typically have alternate hosts. Hence, it is not clear how alternate hosts infected with the same virus isolate alter gene expression in viruliferous vectors. To address the question, this study conducted a transcriptome analysis of viruliferous aphids that acquired the virus from different host species. A polerovirus, cotton leafroll dwarf virus (CLRDV), which induced gene expression in the cotton aphid, Aphis gossypii Glover, was assessed using four alternate hosts, viz., cotton, hibiscus, okra, and prickly sida. Among a total of 2,942 differentially expressed genes (DEGs), 750, 310, 1,193, and 689 genes were identified in A. gossypii that acquired CLRDV from infected cotton, hibiscus, okra, and prickly sida, respectively, compared with non-viruliferous aphids that developed on non-infected hosts. A higher proportion of aphid genes were overexpressed than underexpressed following CLRDV acquisition from cotton, hibiscus, and prickly sida. In contrast, more aphid genes were underexpressed than overexpressed following CLRDV acquisition from okra plants. Only four common DEGs (heat shock protein, juvenile hormone acid O-methyltransferase, and two unannotated genes) were identified among viruliferous aphids from four alternate hosts. Gene ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) annotations indicated that the acquisition of CLRDV induced DEGs in aphids associated with virus infection, signal transduction, immune systems, and fitness. However, these induced changes were not consistent across four alternate hosts. These data indicate that alternate hosts could differentially influence gene expression in aphids and presumably aphid behavior and fitness despite being infected with the same virus isolate.

Aphis gossypii, a globally distributed and economically significant pest of several crops, is known to infest a wide range of host plants. Heat shock proteins (Hsps), acting as molecular chaperones, are essential for the insect\'s environmental stress responses. The present study investigated the molecular characteristics and expression patterns of AgHsp70, a heat shock protein gene, in Aphis gossypii. Our phylogenetic analysis revealed that AgHsp70 shared high similarity with homologs from other insects, suggesting a conserved function across species. The developmental expression profiles of AgHsp70 in A. gossypii showed that the highest transcript levels were observed in the fourth instar nymphs, while the lowest levels were detected in the third instar nymphs. Heat stress and exposure to four different xenobiotics (2-tridecanone, tannic acid, gossypol, and flupyradifurone (4-[(2,2-difluoroethyl)amino]-2(5H)-furanone)) significantly up-regulated AgHsp70 expression. Knockdown of AgHsp70 using RNAi obviously increased the susceptibility of cotton aphids to 2-tridecanone, gossypol and flupyradifurone. Dual-luciferase reporter assays revealed that gossypol and flupyradifurone significantly enhanced the promoter activity of AgHsp70 at a concentration of 10 mg/L. Furthermore, we identified the transcription factor heat shock factor (HSF) as a regulator of AgHsp70, as silencing AgHSF reduced AgHsp70 expression. Our results shed light on the role of AgHsp70 in xenobiotic adaptation and thermo-tolerance.

Predation risk posed by natural enemies can alter pest performance. In our previous study, we found Menochilus sexmaculatus provides risk cues to melon aphids, resulting in increased numbers of winged aphids. However, the effects of predation risk on multiple traits including behavior, physiology, growth rate, and reproductive capacity of pests are not clear. This study examined the effects of predation risk on host preference, the activities of two important defense enzymes (CAT and SOD), longevity, and offspring production. The Y-tube trial results showed that the risk of M. sexmaculatus significantly altered the host preference of the aphids, leading to avoidance behavior. When exposed to M. sexmaculatus for a long period (24 h), the reproductive period and offspring production were significantly decreased, and adult longevity was significantly shortened. The defense enzyme activities of SOD and CAT, as well as the MDA content (which is considered a marker of oxidative stress and cellular damage) in the aphids, significantly increased under M. sexmaculatus risk. The compounds of M. sexmaculatus extracted with n-hexane and volatile compounds collected with HS-SPME were analyzed by GC-MS, and when combined with the behavior response experiment, the results showed that the alkane compounds n-henicosane, n-docosane, n-tricosane, n-pentacosane, and n-hentriacontane may contribute to the impact of predation risk. The results will be helpful in the comprehensive evaluation of the ability of lady beetles to affect the aphid population, and provide new ideas for using these compounds in aphid control.