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Helicoverpa armigera and Aphis gossypii are two important insect species that feed on cotton plants. These insects have distinct abilities to induce plant resistance and tolerate plant toxins, which results in interspecific competition imbalance that may be fatal to the low-tolerance A. gossypii and force these insects to develop avoidance behaviors and subsequently separate from their niche. We implemented ecological experiments to test the effects of H. armigera-induced plant resistance and behavioral avoidance in A. gossypii, and employed transcriptomics and metabolomics analyses to reveal changes in resistance genes and metabolites in plants.Our results demonstrate that cotton plants induced by H. armigera cause significant inhibitory and avoidance effects on A. gossypii insect populations. Electrical penetration graph (EPG) analysis showed changes in plant resistance induced by H. armigera leading to a decreased feeding efficiency of A. gossypii. In addition, genes associated with jasmonic acid and ethylene signaling pathways were significantly up-regulated in cotton plants after H. armigera induction, which led to a significant up-regulation of metabolites inducing plant resistance. These observations were corroborated by bioactivity analysis on metabolites, which showed that jasmonic acid, gossypol and tannins have significant inhibitory effects on A. gossypii populations. In contrast, methylparaben is associated with avoidance behaviors on A. gossypii populations.Our study suggests that the differences in the ability to induce plant resistance and tolerance between two non-predatory insects were lethal to low-tolerance A. gossypii insects, which might be a major factor determining their niche differentiation. This was further demonstrated by screening anti-insect and bio-hormonal metabolites. Our study provides a reference for investigating the evolutionary relationship between non-predatory insects and insights to implement effective insect biocontrol. © 2022 Society of Chemical Industry.

Cytochrome P450 monooxygenase (P450 or CYP) plays an important role in the metabolism of insecticides and plant allelochemicals by insects. CYP321B1, a novel Spodoptera litura P450 gene, was identified and characterized. CYP321B1 contains a 1488 bp open reading frame (ORF) that encodes a 495 amino acid protein. In fourth instar larvae, the highest CYP321B1 expression levels were found in the midgut and fat body. In the tannin feeding test, tannin can significantly induce the expression of CYP321B1 in the midgut and fat body of 4th instar larvae. To verify the function of CYP321B1, RNA interference and metabolome analysis were performed. The results showed that silencing CYP321B1 significantly reduced the rate of weight gain under tannin induction. Metabolome analysis showed silencing affected 47 different metabolites, mainly involved in secondary metabolite biosynthesis and amino acid metabolism, including amino acids, lipid fatty acids, organic acids and their derivatives. Henoxyacetic acid and cysteamine are the most highly regulated metabolites, respectively. These findings demonstrate that CYP321B1 plays an important role in tannin detoxification and metabolism. Functional knowledge about metabolite detoxification genes in this major herbivorous insect pest can provide new insights into this biological process and provide new targets for agricultural pest control.

The cotton-melon aphid Aphis gossypii is a sap-sucking insect that is considered a serious global pest. The species is distributed over a large geographical range and uses a wide variety of hosts, with some populations being specialized to attack different plant species. Here, we provide de novo chromosome-level genome assemblies of a cotton specialist population (Hap1) and a cucurbit specialist population (Hap3). We achieved this by using a combination of third-generation sequencing platforms, namely Illumina and Hi-C sequencing technologies. We were able to anchor a total of 334.89 Mb (scaffold N50 of 89.13 Mb) and 359.95 Mb (scaffold N50 of 68.88 Mb) to four chromosomes for Hap1 and Hap3, respectively. Moreover, our results showed that the X-chromosome of Hap3 (113.01 Mb) was significantly longer than that of Hap1 (100.26 Mb), with a high level of sequence conservation between the aphid species. We also report variation in the number of protein-coding genes and repeat sequences between Hap1 and Hap3. In particular, olfactory and gustatory receptor genes underwent a high level of gene duplication and expansion events in A. gossypii, including between Hap1 and Hap3. Moreover, we identified two glutathione S-transferase genes which underwent single gene duplications in Hap3, and tandem duplication and inversion events affecting the cytochrome P450 monooxygenase between Hap1 and Hap3, all of which include the CYP3 family. Our results illustrate the variance in the genomic composition of two specialized A. gossypii populations and provide a helpful resource for the study of aphid population evolution, host adaption and insecticide resistance.

Insects maintain a vast number of symbiotic bacteria, and these symbionts play key roles in the hosts' life processes. Propylea japonica (Coleoptera: Coccinellidae) is an abundant and widespread ladybeetle in agricultural fields in Asia. Both larvae and adults of P. japonica are likely to be exposed to insecticide residue in the field during their predatory activity. Sulfoxaflor is a highly powerful insecticide that has strong efficacy in controlling sap-sucking pests. To date, there have been several studies on the acute and long-term toxicity of sulfoxaflor to insects, but few studies have reported the impact of sulfoxaflor on the predators' micro-ecosystems. This study was to determine the impact of sulfoxaflor on the symbiotic bacteria and developmental performance of P. japonica. In the present study, two concentrations (1 mg/L and 5 mg/L) and two exposure periods (1 day and 5 days) were set for P. japonica under sulfoxaflor exposure. The survival rate, developmental duration, pupation rate, emergence rate, and body weight of P. japonica were examined. Moreover, the bacterial community of P. japonica was investigated by high-throughput 16S ribosomal RNA gene sequencing. Our results indicated that bacterial community of P. japonica was mainly composed of Staphylococcus, Pantoea, Acinetobacter, Rhodococcus, and Ralstonia at the genus level. The bacterial community of P. japonica in 1 mg/L and 5 mg/L sulfoxaflor groups was significantly altered on day 1, compared with that in control group. The results also showed that the larval duration was significantly prolonged but the pupal duration was significantly shortened in both sulfoxaflor groups. Meanwhile, the pupation and emergence rate was not significantly changed, but the body weights of adults were significantly decreased in both sulfoxaflor groups. Our study will provide a new perspective for evaluating the safety of pesticides to beneficial arthropods.

Propylea japonica has been regarded as one of the most remarkable natural enemies against aphid in China. However, the mechanism of juvenile hormone (JH) regulation of reproduction in P. japonica is still unclear. In this study, we investigated the JH titers of P. japonica and the development of the ovaries. We selected the six different developmental stages of ladybeetle females for transcriptome sequencing. We identified 583 genes involved in insect reproduction regulation, including 107 insect hormone synthesis signaling pathway-related genes and 476 nutrition-sensing signaling pathway-related genes. Transcriptome analysis indicated that a large number JH synthesis- and metabolism-related enzyme genes and some potential nutrient signal sensing- and transduction-related genes were significantly differentially expressed during P. japonica development. We investigated the effects of Met gene silencing on the reproduction of female adults and found that the ovarian maturation, vitellogenesis, and follicular epithelium development in the dsMet treatment group were significantly inhibited.

Agricultural soils and microplastics (MPs) are hotspots for antibiotic resistance genes (ARGs). Plastic mulch is the most important source of MPs in agricultural soil. ARGs, mobile genetic elements (MGEs), and their host profiles in long-term mulch MP-exposed soils remain unclear. In the present study, metagenomics was used to investigate the distribution patterns of ARGs and MGEs in eight Chinese provinces with a long history of plastic mulch use. A total of 204 subtypes of ARGs and thousands of MGEs (14 integrons, 28 insertions, and 2993 plasmids) were identified. A similar diversity of ARGs was found among MPs film-contaminated sites. The types of ARGs with a high abundance were more concentrated, and multidrug resistance genes were the dominant ARGs. Soils from regions with a longer history of plastic film use (such as Xinjiang province) had a higher abundance of ARGs and MGEs. The distribution of ARGs and MGEs exhibited a modular network distribution pattern. A total of 27 ARG subtypes and 29 MGEs showed co-occurrence network relationships. More than 10 common hosts of ARGs and MGEs, such as Pseudomonas, were found, and their abundances were highest in three provinces, including Xinjiang. This study may help elucidate the impact mechanism of long-term MP residues on the occurrence and spread of ARGs in soil.

Insect endosymbionts are widespread in nature and known to play key roles in regulating host biology. As a secondary endosymbiont, bacteria in the genus Hamiltonella help cotton aphids (Aphis gossypii) defend against parasitism by parasitoid wasps, however, the potential negative impacts of these bacteria on cotton aphid biology remain largely unclear.This study aims to evaluate the potential impacts of Hamiltonella on the growth and development of cotton aphids based on life table parameters and RNA sequencing. The results showed that infection with Hamiltonella resulted in smaller body type and lower body weight in aphids. Compared to the control group, there were significant differences in the finite and intrinsic rates of increase and mean generation time. Furthermore, the RNA sequencing data revealed that the genes related to energy synthesis and nutrient metabolism pathways were significantly downregulated and genes related to molting and nervous system pathways were significantly upregulated in the Hamiltonella population.Our results confirm that Hamiltonella retarded the growth and development of cotton aphids accompanied by the downregulation of genes related to energy synthesis and nutrient metabolism, which provides new insights into aphid-symbiont interactions and may support the development of improved aphid management strategies. © 2022 Society of Chemical Industry.

Background Aphis gossypii, a polyphagous and recurrent pest induced by pesticides, causes tremendous loss in the yield of crops each year. Previous studies of pesticide-induced sublethal effects mechanism mainly focus on gene level. The symbiotic bacteria are also important participants of this mechanism, but their roles in hormesis are still unclear. Results In this study, life table parameters and 16S rRNA sequencing were applied to evaluate the sublethal and transgenerational effects of sulfoxaflor on adult A. gossypii after 24-h LC20 (6.96 mg L-1 ) concentration exposure. The results indicated that the LC20 of sulfoxaflor significantly reduced the finite rate of increase (λ) and net reproductive rate (R0 ) of parent generation (G0), and significantly increased mean generation time (T) of G1 and G2, but not of G3 and G4. Both reproductive period and fecundity of G1 and G2 were significantly higher than those of the control. Furthermore, our sequencing data revealed that more than 95% bacterial communities were dominated by the phylum Proteobacteria, in which the maximum proportion genus was the primary symbiont Buchnera and the facultative symbiont Arsenophonus. Compared to those of the control, the abundance and composition of symbiotic bacteria of A. gossypii for three successive generations (G0-G2) were changed after G0 A. gossypii was exposed to sulfoxaflor: the diversity of the bacterial community was decreased, but the abundance of Buchnera was increased (G0), while the abundance of Arsenophonus was decreased. Contrary to G0, G1 and G2 cotton aphid exhibited an increased relative abundance of Arsenophonus in sublethal treatment group. Conclusion Taken together, our results provide an insight into the interactions among pesticide resistance, aphids, and symbionts, which will eventually help to better manage the resurgence of A. gossypii. This article is protected by copyright. All rights reserved.

With the aging population worldwide, osteoporosis, as an age-related bone metabolic disease, is becoming a hot issue in public health. However, it is still a great challenge to realize osteoporotic bone healing due to the alteration of the bone microenvironment in osteoporosis patients. In this study, a nano-structured akermanite (nAK) coating was in situ constructed on Ti–6Al–4V implants to improve osteoporotic bone repair. In vitro studies indicated that both the surface nano-topography and bioactive ions released from the nAK coatings promoted the proliferation, osteogenesis, angiogenesis and inhibited osteoclastogenesis of ovariectomy rabbit-derived bone marrow mesenchymal stem cells (OVX-rBMSCs). Furthermore, the nAK-coated Ti–6Al–4V implants improved new bone formation and osseointegration in an osteoporosis rabbit model in vivo. These results indicated that the AK coating with a nano-structured surface on the Ti–6Al–4V implant could synergistically promote bone formation and osseointegration for osteoporosis patients. This may be a promising strategy to improve the bone regeneration and osseointegration capability of orthopedic implants under osteoporosis conditions.

Osteoporosis (OP) is a major systemic bone disease leading to an imbalance in bone homeostasis which remains a challenge in the current treatment of bone defects. Our previous studies on strontium (Sr) doping apparently stimulated osteogenesis of bioceramics, which suggested a promising strategy for the treatment of bone defects. However, the potential effects and the underlying mechanisms of Sr-doping on osteoporotic bone defects still remain unclear. Autophagy is a conventional self-degradation process of cells involved in bone homeostasis and regeneration under physiological and pathological conditions. Therefore, it is essential to design appropriate biomaterials and investigate the associated osteogenic mechanisms via autophagy. Based on this hypothesis, Sr-doped 45S5 bioglass (Sr/45S5) was fabricated, and ovariectomy bone marrow-derived mesenchymal stem cells (OVX-BMSCs) were applied as the in vitro cell culture model. First, the optimal Sr-doping concentration of 10 mol% was screened by cell proliferation, ALP staining, alizarin red S staining and the real-time PCR assay. Then, the results of western blot (WB) analysis showed that Sr-induced osteogenic differentiation of OVX-BMSCs was associated with time-dependent modulation of autophagy and related to the AKT/mTOR signaling pathway. Meanwhile, the autophagy in Sr-induced osteogenic differentiation of OVX-BMSCs was detected by WB, immunofluorescence staining and transmission electron microscopy. Furthermore, the effect of osteogenic differentiation of OVX-BMSCs has been significantly inhibited by the administration of autophagy inhibitors and the AKT/mTOR pathway inhibitors, respectively, in the early and late periods of osteogenic differentiation. Finally, the results of the model of femoral condyle defects in OVX-rats indicated that Sr10/45S5 granules remarkably enhanced bone regeneration which provided the evidences in vivo. Our research indicates that Sr-doping provides a promising strategy to promote osteogenic differentiation of OVX-BMSCs and bone regeneration in osteoporotic bone defects via early improvement of autophagy and late activation of the Akt/mTOR signaling pathway.

Angiogenesis is closely coupled with osteogenesis and has equal importance. Thus, promoting angiogenesis during the bone repair process is vital for ideal bone regeneration. As important mediators of cell-cell communication and biological homeostasis, mesenchymal stem cell-derived small extracellular vesicles (MSC-sEVs) have been proved to be highly involved in bone and vascular regeneration. Because hypoxia microenvironment promotes the proangiogenic activity of MSCs, in the present study, we investigate the effect and underlying molecular mechanisms of sEVs from hypoxia-preconditioned MSCs (hypo-sEVs) on angiogenesis and develop an effective strategy to promote vascularized bone regeneration. Compared to sEVs from normoxia MSCs (nor-sEVs), hypo-sEVs promoted the proliferation, migration, and angiogenesis of HUVECs and ultimately enhanced bone regeneration and new blood vessel reconstruction in a critical-size calvarial bone defect model. miRNA sequence and the verified results showed that miR-210-3p in hypo-sEVs was increased via HIF-1α under hypoxia. The upregulated miR-210-3p in hypo-sEVs promoted angiogenesis by downregulating EFNA3 expression and enhancing the phosphorylation of the PI3K/AKT pathway. Thus, this study suggests a successful strategy to enhance vascularized bone regeneration by utilizing hypo-sEVs via the miR-210-3p/EFNA3/PI3K/AKT pathway. STATEMENT OF SIGNIFICANCE: Considering the significance of vascularization in ideal bone regeneration, strategies to promote angiogenesis during bone repair are required. Hypoxia microenvironment can promote the proangiogenic potential of mesenchymal stem cells (MSCs). Nonetheless, the therapeutic effect of small extracellular vesicles (sEVs) from hypoxia-preconditioned MSCs on cranio-maxillofacial bone defect remains unknown, and the underlying mechanism is poorly understood. This study shows that hypo-sEVs significantly enhance the proliferation, migration, and angiogenesis of HUVECs as well as promote vascularized bone formation. Moreover, this work indicates that HIF-1α can induce overexpression of miR-210-3p under hypoxia, and miR-210-3p can hinder EFNA3 expression and subsequently activate the PI3K/AKT pathway. The application of hypo-sEVs provides a facile and promising strategy to promote vascularized bone regeneration in a critical-size bone defect model.

As microplastics became the focus of global attention, the hazards of plastic plasticizers (PAEs) have gradually attracted people's attention. Agricultural soil is one of its hardest hit areas. However, current research of its impact on soil ecology only stops at the microorganism itself, and there is still lack of conclusion on the impact of soil metabolism. To this end, three most common PAEs (Dimethyl phthalate: DMP, Dibutyl phthalate: DBP and Bis (2-ethylhexyl) phthalate: DEHP) were selected and based on high-throughput sequencing and metabolomics platforms, the influence of PAEs residues on soil metabolic functions were revealed for the first time. PAEs did not significantly changed the alpha diversity of soil bacteria in the short term, but changed their community structure and interfered with the complexity of community symbiosis network. Metabolomics indicated that exposure to DBP can significantly change the soil metabolite profile. A total of 172 differential metabolites were found, of which 100 were up-regulated and 72 were down-regulated. DBP treatment interfered with 43 metabolic pathways including basic metabolic processes. In particular, it interfered with the metabolism of residual steroids and promoted the metabolism of various plasticizers. In addition, through differential labeling and collinear analysis, some bacteria with the degradation potential of PAEs, such as Gordonia, were excavated.

The complete mitochondrial genome of Aphis gossypii Glover cucumber biotype was sequenced using traditional PCR amplification coupled with Sanger sequencing. The genome is 15,870 bp long, with 83.7...

Plastic pollution in the soil ecosystem is currently receiving worldwide attention. However, little is known whether the presence of microplastics (MPs) in soil will affect the environmental behavior of pesticide residues in soil. Here, the effect of the addition of new mulch MPs (New-MPs), aged mulch MPs (Aged-MPs) and biodegradable mulch MPs (BioD-MPs) on the adsorption and degradation behaviors of two pesticides (imidacloprid and flumioxazin) in soil was investigated. Three MPs slowed down rapid adsorption stage of pesticides in soil and delayed the time to reach adsorption equilibrium. Adsorption rates: Soil Soil + New-MPs Soil + Aged-MPs Soil + BioD-MPs. Three MPs enhanced the adsorption strength of the soil system for the two pesticides, and the aging treatment of the MPs enhanced this effect. Three MPs affected the degradation process of the two pesticides. New-MPs promoted the degradation of two pesticides imidacloprid and flumioxazin, and the degradation half-lives were shortened to 0.93 and 0.85 times, respectively; while Aged-Mps and BioD-MPs delayed the degradation process of two pesticides, and the degradation half-lives were extended to 1.64 times and 1.21 times, respectively. The effect was more significant with the increase of MPs and pesticides concentration. Pesticide polarity, surface structure and functional groups of MPs are potentially important reasons for the differences in adsorption and degradation of MPs-soil systems. Our findings provide a deep insight into understanding the mechanism of interaction between MPs and pesticide residues in soil environment.

In insects, cytochrome P450 monooxygenases (P450s or CYPs) play an important role in the detoxification and metabolism of exogenous plant allelochemicals. In this study, a P450 gene CYP6AB12 was identified and characterized from Spodoptera litura. The cDNA contains an open reading frame (ORF) encoding 511 amino acid residues. CYP6AB12 was expressed at different ages of S. litura, with the highest levels found in the third and fourth instar larvae. Its highest expression was found in the midgut and fat body of fourth instar larvae fed with gossypol. Moreover, these expression levels were substantially increased compared with those from larvae fed with control diet. Gene silencing was then conducted by smearing dsRNA mixed with nanomaterials onto the cuticle. CYP6AB12 expression was significantly decreased in the midgut and fat body, and the net weight increase was substantially lower than that of the control group, indicating that the treatment group had more sensitivity to gossypol than the control. These results reveal that CYP6AB12 plays an important role in the detoxification and metabolism of gossypol, thus further confirming that P450s have a broad ability to detoxify and metabolize plant allelochemicals. It provides an important molecular basis for the exploration of detoxification metabolism and pest control of S. litura.

Research on microplastic pollution of terrestrial soils is catching up with the aquatic environment, especially agricultural soil systems. Plastic residues have caused various environmental problems in mulch film extensively used agricultural areas. However, studies focusing specifically on the potential influence of mulch film residues on the metabolic cycle of soil systems have yet to be conducted. Here, high-throughput sequencing combined with metabolomics were first used to study the effects of residual mulch on soil microbial communities and related metabolic functions. Plastic film treatment did not significantly affect soil physicochemical properties including pH, organic matter and nitrogen, etc in short term. However, it did significantly changed overall community structure of soil bacteria, and interfered with complexity of soil bacterial symbiosis networks; exposure time and concentration of residues were particularly important factors affecting community structure. Furthermore, metabolomics analysis showed that film residue significantly changed soil metabolite spectrum, and interfered with basic carbon and lipid metabolism, and also affected basic cellular processes such as membrane transport and, in particular, interfered with the biosynthesis of secondary metabolites, as well as, biodegradation and metabolism of xenobiotics. Additionally, through linear discriminant and collinear analysis, some new potential microplastic degrading bacteria including Nitrospira, Nocardioidaceae and Pseudonocardiaceae have been excavated.

The ladybird beetle Propylea japonica is an important natural enemy in agro-ecological systems. Studies on the strong tolerance of P. japonica to high temperatures and insecticides, and its population and phenotype diversity have recently increased. However, abundant genome resources for obtaining insights into stress-resistance mechanisms and genetic intra-species diversity for P. japonica are lacking. Here, we constructed the P. japonica genome maps using Pacific Bioscience (PacBio) and Illumina sequencing technologies. The genome size was 850.90 Mb with a contig N50 of 813.13 kb. The Hi-C sequence data were used to upgrade draft genome assemblies; 4,777 contigs were assembled to 10 chromosomes; and the final draft genome assembly was 803.93 Mb with a contig N50 of 813.98 kb and a scaffold N50 of 100.34 Mb. Approximately 495.38 Mb of repeated sequences was annotated. The 18,018 protein-coding genes were predicted, of which 95.78% were functionally annotated, and 1,407 genes were species-specific. The phylogenetic analysis showed that P. japonica diverged from the ancestor of Anoplophora glabripennis and Tribolium castaneum ~ 236.21 million years ago. We detected that some important gene families involved in detoxification of pesticides and tolerance to heat stress were expanded in P. japonica, especially cytochrome P450 and Hsp70 genes. Overall, the high-quality draft genome sequence of P. japonica will provide invaluable resource for understanding the molecular mechanisms of stress resistance and will facilitate the research on population genetics, evolution and phylogeny of Coccinellidae. This genome will also provide new avenues for conserving the diversity of predator insects.

Parasitism usually causes considerable changes in lipids and fatty acids by redirecting the development of the host. In this study, changes in weight and in free fatty acid content of cotton aphids were recorded after aphids had been parasitized. Results showed that the weight of parasitized Aphis gossypii was increased compared to nonparasitized aphids, and significantly increased weights were detected at 1, 2, and 3 instars after parasitization by Lysiphlebia japonica. Free fatty acid test kits and GC-MS showed that the fatty acid content increased in the early stage of parasitization but decreased after 3 days of parasitization. Seven genes related to the fatty acid synthesis pathway were significantly upregulated in the parasitized aphids, where they were 1.96-10.97 times greater. Our data described the change that occurs in the fatty acid content of parasitized A. gossypii.

Elevated CO

The complete mitochondrial genome of Aphis gossypii Glover cucumber biotype was sequenced using traditional PCR amplification coupled with Sanger sequencing. The genome is 15,870 bp long, with 83.7% AT content (MW048625). The genome encodes 37 typical mitochondrial genes, including 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNAs, a repeat region of 784 bp, and a control region of 627 bp. The base composition of the genome is A (45.4%), T (38.3%), C (10.5%), and G (5.8%). An analysis of two biotypes A. gossypii mitogenomes identified 77 single nucleotide polymorphisms and 1 insertion and deletion.

The predatory ladybird beetle, Harmonia axyridis, is a predominant natural enemy of pest insects in cotton fields. Commercialization of genetically modified crops has promoted the increased use of the herbicide glyphosate. In this study, to assess potential negative effects of glyphosate on beneficial non-target organisms in cotton fields, we first examined how glyphosate exposure affected the development and endosymbiotic bacterial community of H. axyridis. The results showed that the survival rate, development duration, pupation rate and emergence rate of H. axyridis under low and high concentrations of glyphosate exposure were not significantly changed, but glyphosate did significantly reduce the body weight of H. axyridis. Based on 16S rRNA sequencing, there were no significant differences in the diversity or richness of the endosymbiotic bacteria of H. axyridis before and after glyphosate exposure. The dominant bacterial phyla Firmicutes and Proteobacteria and genera Staphylococcus and Enterobacter remained the same regardless of treatment with glyphosate, however the abundance and copy number of these bacteria were altered. Glyphosate treatment significantly reduced the abundance and gene copy number of Staphylococcus and increased the abundance and gene copy number of Enterobacter. This is the first report demonstrating that glyphosate can reduce the body weight H. axyridis and alter the bacterial endosymbiont community by affecting the abundance and gene copy number of dominant bacteria.

Background The worldwide pest Aphis gossypii has three-winged morphs in its life cycle, namely, winged parthenogenetic female (WPF), winged gynopara (GP), and winged male, which are all produced by a wingless parthenogenetic female (WLPF). Most studies on A. gossypii have focused on WPF, while few have investigated GP and male. The shared molecular mechanism underlying the wing differentiation in the three wing morphs of A. gossypii remains unknown. The wing differentiation of WPF was explored in a previous study. Herein, GP and male were induced indoors. The characters of the body, internal genitals, wing veins, and fecundity of GP and male were compared with those of WPF or WLPF. Compared with WLPF, the shared and separate differentially expressed genes (DEGs) were identified in these three-wing morphs. Results Newly-born nymphs reared in short photoperiod condition (8 L:16D, 18 °C) exclusively produced gynoparae (GPe) and males in adulthood successively, in which the sex ratio was GP biased. A total of 14 GPe and 9 males were produced by one mother aphid. Compared with WLPF, the three-wing morphs exhibited similar morphology and wing vein patterns but were obviously discriminated in the length of fore- and underwings, reproductive system, and fecundity. A total of 37 090 annotated unigenes were obtained from libraries constructed using the four morphs via RNA sequencing (RNA-Seq). In addition, 10 867 and 19 334 DEGs were identified in the pairwise comparison of GP versus WLPF and male versus WLPF, respectively. Compared with WLPF, the winged morphs demonstrated 2 335 shared DEGs (1 658 upregulated and 677 downregulated). The 1 658 shared upregulated DEGs were enriched in multiple signaling pathways, including insulin, FoxO, MAPK, starch and sucrose metabolism, fatty acid biosynthesis, and degradation, suggesting their key roles in the regulation of wing plasticity in the cotton aphid. Forty-four genes that spanned the range of differential expression were chosen to validate statistical analysis based on RNA-Seq through the reverse transcription quantitative real time polymerase chain reaction (RT-qPCR). The comparison concurred with the expression pattern (either up- or downregulated) and supported the accuracy and reliability of RNA-Seq. Finally, the potential roles of DEGs related to the insulin signaling pathway in wing dimorphism were discussed in the cotton aphid. Conclusions The present study established an efficiently standardized protocol for GP and male induction in cotton aphid by transferring newly-born nymphs to short photoperiod conditions (8 L:16D, 18 °C). The external morphological characters, especially wing vein patterns, were similar among WPFs, GPe, and males. However, their reproductive organs were strikingly different. Compared with WLPF, shared (2 335) and exclusively (1 470 in WLPF, 2 419 in GP, 10 774 male) expressed genes were identified in the three-wing morphs through RNA-Seq, and several signaling pathways that are potentially involved in their wing differentiation were obtained, including insulin signaling, starch and sucrose metabolism.

Lysozymes are used in sterilisation, antisepsis, dairy additives, inflammation, and cancer. One transgenic goat line expressing high levels of human lysozyme (hLZ) in goat milk has been developed in China. Herein, we established an event-specific real-time polymerase chain reaction (real-time PCR) method to detect the transgenic hLZ goat line. The developed method has high specificity, sensitivity and accuracy, and a wide quantitative dynamic range. The limit of detection and limit of quantification was 5 and 10 copies per reaction, respectively. The practical sample analysis results showed that the method could identify and quantify transgenic lysozyme content in trace samples in routine lab analyses. Furthermore, the potential applicability in risk assessment, such as molecular characterisation and gene horizontal transfer, was confirmed. We believe that this method is suitable for the detection of transgenic hLZ goat line and its derivate.

Multiplex and high-throughput assays are becoming the main trends in the development of new nucleic acid detection and quantification methods, such as those for genetically modified organism (GMO) analysis. Here, we report a novel universal LNA probe-mediated droplet digital polymerase chain reaction (PCR) method (ULNA-ddPCR) for multiple DNA target quantification in GMOs. In ULNA-ddPCR, only one universal LNA probe is used for multiple DNA targets instead of using one to one TaqMan probe. The specificity, sensitivity, dynamic range, and accuracy of the ULNA-ddPCR method are determined by employing GM rice analysis as an example. Simplex and triplex ULNA-ddPCR assays for three GM rice events, T2A-1, T1C-19, and G6H1, are established and evaluated. All results indicate that the developed simplex and triplex ULNA-ddPCR assays are suitable for quantitative analysis of GM rice events with high sensitivity, accuracy, and low cost. The ULNA-ddPCR method also has the potential for multiple DNA target quantification in other research fields.

Spodoptera litura is a major insect with a cosmopolitan distribution and strong resistance to multiple insecticides. Determining the molecular basis and key candidate genes of the insecticide resistance of S. litura may help in managing this insect. In this study, fifth-instar S. litura larvae were subjected to transcriptome analysis at 6, 12, 24, 48, and 72 h after feeding on an LC20 dose of avermectin. The result showed that genes responding to avermectin changed dynamically with different gene counts and resistance mechanisms at the fifth instar based on a metabolic pathway map. These responses included degrading the insecticide by a series of P450 and glutathione-S-transferase enzymes starting at the 12 h time point, with subsequent increases in the number of genes involved and shifts to TOLL and immune deficiency (IMD) pathways at 48 h after feeding the insecticide. Weighted correlation network analysis (WGCNA) determined a co-expression module related to the avermectin response at 12 and 24 h (r = 0.403, p = 0.0371; r = 0.436, p = 0.023), in which a hub gene (LOC111358940) related to metalloproteinase activity was identified. In addition, Analysis of the genes in the co-expression module further revealed that eight genes encoding UDP-glucuronosyltransferases were directly associated with insecticide response in S. litura. These results provide better understanding of the avermectin response mechanism of S. litura and may be useful in developing improved control strategies for this species. SUPPLEMENTARY INFORMATION: The online version of this article (10.1007/s13205-021-02651-9) contains supplementary material, which is available to authorized users.

Background Endosymbiotic bacteria have been reported to mediate interactions between parasitoids and their insect hosts. How parasitic wasps influence changes in host microbial communities and the relationship between them are of great importance to the study of host-parasitoid co-evolutionary and ecological interactions. However, these interactions remain largely unreported for interactions between in Aphis gossypii and Lysiphlebia japonica. Results In this study, we characterize the bacterial microbiota of L. japonica wasps at different developmental stages and monitor changes over time in the bacterial microbiota of their parasitized and non-parasitized aphid hosts, using metagenomic analysis of 16S rDNA sequencing data. Proteobacteria, Firmicutes, and Actinobacteria were the three most abundant bacterial phyla identified in L. japonica. We found that parasitism was associated with an increased abundance of Buchnera nutritional endosymbionts, but decreased abundance of Acinetobacter, Arsenophonus, Candidatus_Hamiltonella, and Pseudomonas facultative symbionts in aphid hosts. Functional analysis of enriched pathways of parasitized aphids showed significant differences in the 'transport and metabolism of carbohydrates' and 'amino acid, lipid, and coenzyme biosynthesis' pathways. Notably, the composition of symbiotic bacteria in wasp larvae was highly similar to that of their aphid hosts, especially the high abundance of Buchnera. Conclusion The results provide a conceptual framework for L. japonica interactions with A. gossypii in which the exchange of symbiotic microbes provides a means by which microbiota can potentially serve as evolutionary drivers of complex, multilevel interactions underlying the ecology and co-evolution of these hosts and parasites. This article is protected by copyright. All rights reserved.

Simple Summary The Aphis gossypii is a global problem for its pesticide resistance with substantial economic and ecological cost and a wide host range, including cotton and cucurbits. The development of insecticide resistance is rapid and widespread and threatens crop productivity. Biopesticides have emerged as a better alternative for pest control. Cucurbitacin B (CucB) and epigallocatechin gallate (EGCG) are the major secondary metabolites of host plants cucurbits and cotton. In this study, we used cotton- and cucurbit-specialized aphids (CO and CU) as a study system to better understand the effects of CucB and EGCG on cotton aphid. Our study showed that CucB and EGCG can significantly reduce the population-level fitness of A. gossypii, affect their ability to adapt to nonhost plants and alter the levels of some detoxifying enzymes, which showed a potential to be developed into new biopesticides against the notorious aphids. Abstract Aphis gossypii (Glover) is distributed worldwide and causes substantial economic and ecological problems owing to its rapid reproduction and high pesticide resistance. Plant-derived cucurbitacin B (CucB) and epigallocatechin gallate (EGCG) are known to have insecticidal and repellent activities. However, their insecticidal activity on cotton- and cucurbit-specialized aphids (CO and CU), the two important host biotypes of A. gossypii, remains to be investigated. In the present study, we characterized, for the first time, the effects of these two plant extracts on the two host biotypes of A. gossypii. CucB and EGCG significantly reduced the A. gossypii population-level fitness and affected their ability to adapt to nonhost plants. Activities of important detoxification enzymes were also altered, indicating that pesticide resistance is weakened in the tested aphids. Our results suggest that CucB and EGCG have unique properties and may be developed as potential biopesticides for aphid control in agriculture.

Implantable self-powered generators (ISPGs) have been extensively explored as energy supplies for driving electronics and electrically stimulated therapeutics in vivo. However, some drawbacks arise, such as complicated architectonics, inescapability of wire connection, energy instability, and consumption. In this study, a host-coupling bio-nanogenerator (HCBG) is developed to configure a self-powered regional electrical environment for powerful bone regeneration. An HCBG consists of a porous electret nanofiber mat coupled with interstitial fluid and stimulated objects of the host after implantation, forming a host coupling effect. This bio-nanogenerator not only overcomes the disadvantages of general ISPGs, but also accomplishes both biomechanical energy scavenging and electrical stimulation therapeutics. The enhancement of osteogenesis differentiation of bone marrow mesenchymal stem cells in vitro and bone regeneration in vivo are remarkably achieved. Moreover, osteogenic ability is systematically evaluated by regulating the electrical performance of HCBGs. Osteogenic differentiation is activated by upregulating more cytosolic calcium ion, following to activate the calcium ion-induced osteogenic signal pathway, while applying electrical stimulation. As an implantable medical technology, the HCBG provides an explorative insight to facilitate the development of ISPG-based electrical medical therapeutics.

Objectives The polyester/hydroxyapatite (polyester/HA) composites play an important role in bone tissue repairing, mostly because they mimic the composition and structure of naturally mineralized bone tissue. This review aimed to discuss commonly used geometries of polyester/HA composites, including microspheres, membranes, scaffolds and bulks, and their applications in bone tissue repairing and to discuss existed restrictions and developing trends of polyester/HA. Methods The current review was conducted by searching Web of Science, and Google Scholar for relevant studies published related with polyester/HA composites. Selected studies were analyzed with a focus on the fabrication techniques, properties (mechanical properties, biodegradable properties and biological properties) and applications of polyester/HA composites in bone repairing. Results A total of 111 articles were introduced to discuss the review. Different geometries of polyester/HA composites were discussed. In addition, properties and applications of polyester/HA composites were evaluated. The addition of HA into polyester can adjust the mechanical and biodegradability of composites. Besides, the addition of HA into polyester can improve its osteogenic abilities. The results showed that polyester/HA composites can ideal candidate for bone tissue repairing. Conclusion Polyester/HA composites have many remarkable properties, such as appropriate mechanical strength, biodegradability, favorable biological properties. Diverse geometries of polyester/HA composites have been used in bone repairing, drug delivery and implant fixation. Further work needs to be done to investigate existed restrictions, including the controlled degradation rate, controlled drug release performance, well-matched mechanical properties, and novel fabrication techniques. The translational potential of this article The present review reveals the current state of the polyester/HA composites used in bone tissue repairing, contributing to future trends of polyester/HA composites in the forthcoming future.

Using bioactive nanomaterials in clinical treatment has been widely aroused. Nanomaterials provide substantial improvements in the prevention and treatment of oral and maxillofacial diseases. This review aims to discuss new progresses in nanomaterials applied to oral and maxillofacial tissue regeneration and disease treatment, focusing on the use of nanomaterials in improving the quality of oral and maxillofacial healthcare, and discuss the perspectives of research in this arena. Details are provided on the tissue regeneration, wound healing, angiogenesis, remineralization, antitumor, and antibacterial regulation properties of nanomaterials including polymers, micelles, dendrimers, liposomes, nanocapsules, nanoparticles and nanostructured scaffolds, etc. Clinical applications of nanomaterials as nanocomposites, dental implants, mouthwashes, biomimetic dental materials, and factors that may interact with nanomaterials behaviors and bioactivities in oral cavity are addressed as well. In the last section, the clinical safety concerns of their usage as dental materials are updated, and the key knowledge gaps for future research with some recommendation are discussed. This article is categorized under: Implantable Materials and Surgical Technologies > Nanomaterials and Implants Implantable Materials and Surgical Technologies > Nanotechnology in Tissue Repair and Replacement.

Spodoptera litura is one of the most devastating agricultural pests with a wide range of host plants. To study larval performance on different diets and midgut adaptation at transcriptional levels, feeding assay and RNA-Seq experiments were conducted. RNA interference technology was used to explore the detoxification and metabolism of two cytochrome P450 genes. The bioassay data showed that S. litura larvae developed more quickly when fed on cabbage than when fed on soybean, corn and cotton, tannin can inhibit the growth of S. litura . The result of RNA-Seq indicated that S. litura midgut modified gene expression levels to accommodate different diets, and the most differentially expressed genes were detoxification-related and digestion-related genes . Further analysis showed that the glutathione metabolism pathway was the common detoxification pathway in S. litura. The expression of cytochrome P450 genes showed a clear response to different plant hosts, and these differences may play key functions in primary detoxification of secondary metabolites from host plants. Meanwhile, the digestive enzymes of proteinases, lipases, and carbohydrases in midgut showed special responses to different plant hosts. After injection of dsRNA of CYP321A19 and CYP6AB60 , the expression level of target gene were decreased, and the sensitivity of insect to plant allelochemicals increased and the weight increase significantly slowed. In this study, genes involved in detoxification were identified, and the results demonstrate the genes and pathways S. litura utlize to detoxify specific plant-host allelochemicals. These results may also provide a theoretical basis for S. litura management.

Lysiphlebia japonica Ashmead (Hymenoptera, Braconidae) is an endophagous parasitoid and Aphis gossypii Glover (Hemiptera, Aphididae) is a major pest in cotton. The relationship between insect host-parasitoids and their hosts involves complex physiological, biochemical and genetic interactions. This study examines changes in the development and physiological metabolism of A. gossypii regulated by L. japonica. Our results demonstrated that both the body length and width increased compared to non-parasitized aphids. We detected significantly increases in the developmental period as well as severe reproductive castration following parasitization by L. japonica. We then used proteomics to characterize these biological changes, and when combined with transcriptomes, this analysis demonstrated that the differential expression of mRNA (up or downregulation) captured a maximum of 48.7% of the variations of protein expression. We assigned these proteins to functional categories that included immunity, energy metabolism and transport, lipid metabolism, and reproduction. We then verified the contents of glycogen and 6-phosphate glucose, which demonstrated that these important energy sources were significantly altered following parasitization. These results uncover the effects on A. gossypii following parasitization by L. japonica, additional insight into the mechanisms behind insect-insect parasitism, and a better understanding of host-parasite interactions.

The widespread commercialization of genetically modified (GM) cotton makes it important to assess the potential impact of this recombinant crop on non-target organisms. As important natural enemies of cotton field predators, green lacewing Chrysoperla sinica larvae are exposed to Bt insecticidal proteins expressed by GM cotton by feeding on herbivorous pests, and adults are directly exposed to Bt proteins by cotton pollen consumption. However, potential impacts of transgenic Bt cotton on C. sinica remain unclear. In this study, we evaluated the effects of two transgenic cotton varieties, CCRI41 and CCRI45, which express Cry1Ac (Bt toxin) and CpTI (Cowpea Trypsin Inhibitor), on C. sinica larvae and adults. After being fed with cotton aphids Aphis gossypii reared on transgenic cotton, the survival rate, developmental duration, pupation rate, and emergence rate of larvae were not adversely affected. After being fed two types of transgenic cotton pollen, the 7-day weight of adults and the preoviposition period and the cumulative oviposition of females were not significantly different from control specimen. Taken together, these results indicate that the potential risks of the two tested GM cotton varieties for the predator C. sinica are negligible. Capsule: Our study indicated that GM cotton varieties CCRI41 and CCRI45 have no adverse effects on insect predator C. sinica.

Background: Three wing morphs exists in the life cycle of the worldwide pest Aphis gossypii, i.e., wing parthenogenetic female (WPF), gynopara (GP) and male, which were produced mostly by crowding and host quality, photoperiod, loss of X chromosome, respectively. However, the shared molecular mechanism underlying their wing differentiation remains an enigma. Here we firstly induced gynoparae and males indoors and compared the characters of these wing morphs in body, internal genitals and fecundity. Then we identified the shared and separate differentially expressed genes (DEGs) and signaling pathways potentially involved in the wing morphs regulation in WPF, GP and male compared to wingless parthenogenetic female (WLPF). Results: Newly-born nymphs reared in short photoperiod condition exclusively produce gynoparae and males in adulthood successively, in which the sex ratio is gynoparae biased. Compared with WLPF, three wing morphs have similar morphology in bodies but is obviously discriminated in the reproductive system and fecundity. Built upon our previous study, 37 090 annotated unigenes were obtained from libraries constructed by the four morphs above through RNA-sequencing, in which 10 867, 19 334 DEGs were identified in pairwise comparison of GP vs. WLPF, Male vs. WLPF, respectively. Furthermore, 2 335 shared DEGs including 1 658 up- and 677 downregulated were obtained in these wing morphs compared to WLPF. The 1 658 shared up-regulated DEGs were enriched in multiple signaling pathways including insulin, FoxO, MAPK, strarch and sucrose metabolism, fatty acid biosynthesis and degradation which hint their key roles in the regulation of wing plasticity in cotton aphid. Gene expression levels were validated by using Pearson’s correlation (r) and potential roles of 15 DEGs related to the insulin signaling pathway in cotton wing dimorphism were discussed. Conclusions: The results of this study establish a solid foundation for deciphering molecular mechanisms underlying the switch between wingless and wing morphs in the cotton aphid and provide valuable resources for future research on the host-alternating aphid species.

Elevated CO2 concentrations can change the multi-level nutritional relationship of the ecosystem through the cascading effect of the food chain. To date, few studies have investigated the effects of elevated CO2 concentration on the Araneae species through the tritrophic system. Hylyphantes graminicola (Araneae: Linyphiidae) is distributed widely in Asia and is a dominant predator in cotton fields. This study investigated chemical components in the food chain of cotton (Gossypium hirsutum)—cotton aphid (Aphis gossypii)—predator (H. graminicola) and compared the development, reproduction, and predation of H. graminicola under ambient (400 ppm) and elevated concentration of CO2 (800 ppm). The results showed that the elevated CO2 concentration increased the chemicals of cotton and cotton aphid, but it did not affect the nutrients, development, reproduction, and predation of the spider. However, the survival rate of the spider was significantly decreased in elevated CO2. The results will further our understanding of the role of natural enemies in an environment with elevated CO2 concentration.

Lysiphlebia japonica (Ashmead) is a predominant parasitoid of cotton-melon aphids in the fields of northern China with a proven ability to effectively control cotton aphid populations in early summer. For accurate normalization of gene expression in L. japonica using quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR), reference genes with stable gene expression patterns are essential. However, no appropriate reference genes is L. japonica have been investigated to date. In the present study, 12 selected housekeeping genes from L. japonica were cloned. We evaluated the stability of these genes under various experimental treatments by RT-qPCR using four independent (geNorm, NormFinder, BestKeeper and Delta Ct) and one comparative (RefFinder) algorithm. We identified genes showing the most stable levels of expression: DIMT, 18S rRNA, and RPL13 during different stages; AK, RPL13, and TBP among sexes; EF1A, PPI, and RPL27 in different tissues, and EF1A, RPL13, and PPI in adults fed on different diets. Moreover, the expression profile of a target gene (odorant receptor 1, OR1) studied during the developmental stages confirms the reliability of the chosen selected reference genes. This study provides for the first time a comprehensive list of suitable reference genes for gene expression studies in L. japonica and will benefit subsequent genomics and functional genomics research on this natural enemy.

Risk assessments of insecticidal Cry proteins from Bacillus thuringiensis (Bt) cotton in soils have been mainly based on microcosm and short-term experiments. There is a lack of longer-term field results, especially when Bt-cotton residues are incorporated into the soil. In this study, we used a six-year field trial to evaluate how cultivation duration and Cry1Ac-inputted modes of Bt-cotton affect the persistence of Cry1Ac proteins and soil microbe-mediated enzymatic properties. The results showed that the persistence of Cry1Ac proteins increased with cultivation duration and periodic residue incorporation of the transgenic Bt-cotton variety ZM41. Moreover, temporal residue incorporation had a relatively larger contribution to the persistence of Cry1Ac proteins in the soil than their release in the growth period. Regardless of Bt-cotton cultivation or residue incorporation, soil microbial biomass was significantly suppressed. However, the dehydrogenase activity was significantly stimulated in Bt-cotton cultivation but suppressed in residue incorporation. The activities of β -glucosidase, nitrate reductase, phosphomonoesterase and arylsulfatase were significantly stimulated in soils with Bt-cotton residue incorporation. Based on a structural equation model analysis, the change in enzymatic activity of these four enzymes was attributed to both a direct effect from Cry1Ac proteins and an indirect effect via dehydrogenase.