Effects of cadmium stress at different concentrations on the reproductive behaviors of beet armyworm Spodoptera exigua (Hübner).

Insects are exposed to cadmium stress since cadmium pollution has increasingly become a serious global environmental issue. However, until now few studies have paid attention to the effect of heavy metals on insect reproductive behaviors. In our study, the courtship behaviors, mating behaviors and fecundity of beet armyworm Spodoptera exigua (Lepidoptera: Noctuidae) exposed to different concentrations of cadmium in artificial diets at larval stage were studied. The results showed that cadmium stress changed the courtship rhythm by significantly advancing or delaying the courtship starting time. Low dose of cadmium (0.2 mg/kg) increased the courtship frequency in the first two scotophases, but in the fourth phase, the two cadmium treatments reduced the frequency. The total courtship duration was significantly shortened in the first six scotophases except high dose of cadmium treatment (51.2 mg/kg) in the sixth dark phase. Paired adults did not mate after the seventh scotophase under low cadmium exposure, while high cadmium stress made the paired adults just copulate in the first four scotophases. The daily mating rate and total mating rate decreased with the increase in cadmium concentration. The number of eggs of low cadmium treatment was higher than that of control, but the difference was not significant; the number of eggs in high cadmium treatment was lower than that of control and low cadmium treatment. Our results indicate that cadmium exposure can disrupt the courtship rhythm for females and has negative influences on copulation behavior and high cadmium stress can reduce fecundity. Hence, the insect population increase will be affected by heavy metal pollution. Our study will provide scientific reference for environmental risk assessment of heavy metal pollution.

Better cold tolerance of Bt-resistant Spodoptera exigua strain and the corresponding cold-tolerant mechanism.

Spodoptera exigua is a secondary target pest of Bt cotton commercialized in China. With the continuous adoption of Bt cotton, populations of S. exigua have gradually increased. However, the cold tolerance ability of Bt-resistant S. exigua and the effect of continuous Bt diet on anti-cold materials are unknown. In our study, it was found that Bt-resistant S. exigua (Bt10) developed better with shorter larval and pupal duration and higher pupation rate compared to CK at the suboptimal low temperature. The supercooling points and freezing points of the Bt-resistant S. exigua strain were determined, and body water content and anti-cold materials such as total sugar, trehalose and glycogen, glycerol and fat were examined to explore the effect of Bt toxin on overwintering and on population increase. The results showed that the supercooling point and the freezing point of the Bt-resistant S. exigua pupae were both significantly lower than that of the Bt-susceptible strain. No difference was found in the body water content of pupae and adults between the two strains. Total sugar content of the Bt-resistant strain at both the pupal and adult stages was higher than that of the susceptible strain at the corresponding stages, and glycogen content of the Bt-resistant strain at the larval stage was higher than that of the susceptible larval S. exigua. Fat content of the Bt-resistant larvae, pupae and adults was for each higher than that of the susceptible strain, but the difference was not significant except for that of the 3rd instar larvae. Glycerol content of the Bt-resistant strain at larval, pupal and adult stages was for each higher than that of the corresponding life stages of the susceptible strain. It can be seen that more glycerol was accumulated in Bt-resistant S. exigua. The results indicate that Bt-resistant S. exigua has better cold tolerance. The contents of the anti-freeze substances of progeny, especially glycerol, were increased after previous generations were continuously fed on Bt protein, which means that the Bt-resistant secondary target pests could more easily overcome the overwinter season and become a source of crop damage the following year.

Does expression of Bt toxin matter in farmer's pesticide use?

Despite the widespread adoption of Bt cotton, farmers still spray excessive pesticides in their cotton fields. In contrast to scientists who always use high quality seeds in the laboratory and/or experimental fields, farmers may plant low quality seeds with a low expression of Bt toxin. How does the expression of Bt toxin influence farmers' pesticide use? On the basis of a plot-level survey and laboratory test data, this study shows that pesticide use on one cotton plot is influenced not only by the expression of Bt crops in this plot, but also by the average expression in the village in the early stage of the cotton growing season. In other words, high expression of Bt toxin benefits not only the farmers who plant the varieties but also all the other villagers.

Effect of farm management practices in the Bt toxin production by Bt cotton: evidence from farm fields in China.

Based on farm field plot level survey data and laboratory test, we examine the determinants of the expression of Bt toxin in China's Bt cotton production. The results show that the expression of Bt toxin differs significantly among varieties. Even for the same variety the expression of Bt toxin also varies substantially among villages and among farmers in the same village. Econometric analyses show that after controlling for the effects of varieties and locations (or villages), farm management, particular applications of phosphate and potash fertilizers, and manure, has significant positive effects on Bt toxin expression in farmer's fields. In contrast to previous studies which showed that nitrogen fertilizer has a positive impact on expression of Bt toxin, this study shows that nitrogen fertilizer has no significant impact on expression of Bt toxin in farmer's fields. On the other hand, the expression of Bt toxin has a positive relationship with phosphate fertilizer, potash fertilizer and manure application.

Accumulation and excretion of cadmium in three successive generations of Spodoptera exigua (Lepidoptera: Noctuidae) and impact on the population increase.

Nowadays, heavy metal pollution has become more and more serious. Cadmium is one of the heavy metals that have caused serious pollution. Spodoptera exigua Hübner (Lepidoptera: Noctuidae), one of important agricultural pests, has broken out more frequently in recent years. In this study, accumulation at different developmental stages and excretion of Cd were detected, and effect of cadmium (Cd concentrations were 0.2, 0.8, 3.2, 12.8, and 51.2 mg/kg, respectively) on three successive generations of beet armyworm was also studied using life table in the laboratory. Results showed that Cd2+ contents were significantly different when S. exigua was reared under different Cd2+ concentrations in the same generation in a significant dose-dependent manner. The concentration of Cd2+ increased with extension of stress time. Cd2+ concentration at different developmental stages was different, that is, larvae > pupa > adult. After emergence, Cd2+ concentration in both females and males decreased gradually, but that in paired adults decreased to a greater extent than that in unpaired ones. Beet armyworm could excrete heavy metals by means of faeces, prepupa exuviate, and puparium. Life table of the laboratory population was constructed, which showed that low doses of cadmium promoted the increase of population, whereas high doses inhibited the increase, meanwhile, with the extension of exposure length, the index of population trend (I) declined more rapidly. The study might provide references for the long-term and objective assessment of heavy metal pollution and its effect on the populations of important agricultural pests.

Transcriptomic and Metabolomic Responses in Cotton Plant to Apolygus lucorum Infestation.

With the wide-scale adoption of transgenic Bacillus thuringiensis (Bt) cotton, Apolygus lucorum (Meyer-Dür) has become the most serious pest and has caused extensive yield loss in cotton production. However, little is known about the defense responses of cotton at the seedling stage to A. lucorum feeding. In this study, to elucidate the cotton defense mechanism, cotton leaves were damaged by A. lucorum for 0, 4, 12 and 24 h. The transcriptomic results showed that A. lucorum feeding elicits a rapid and strong defense response in gene expression during the whole infestation process in cotton plants. Further analysis revealed that at each assessment time, more differentially expressed genes were up-regulated than down-regulated. The integrated analysis of transcriptomic and metabolic data showed that most of the genes involved in jasmonic acid (JA) biosynthesis were initially up-regulated, and this trend continued during an infestation. Meanwhile, the content levels of JA and its intermediate products were also significantly increased throughout the whole infestation process. The similar trend was displayed in condensed tannins biosynthesis. This research proved that, after plants are damaged by A. lucorum, the JA pathway mediates the defense mechanisms in cotton plants by promoting the accumulation of condensed tannins as a defense mechanism against A. lucorum. These results will help us to discover unknown defensive genes and improve the integrated pest management of A. lucorum.

Behavioral Responses of Bemisia tabaci Mediterranean Cryptic Species to Three Host Plants and Their Volatiles.

Bemisia tabaci Gennadius (Hemiptera: Aleyrodidae) is a worldwide pest that damages over 900 host plant species. The volatile organic compounds (volatiles) of contrasting plants, as well as their growth stage, influence this pest's infestation behavior. The chemical contents of volatiles isolated from three plants (Gossypium hirsutum, Abutilon theophrasti, and Ricinus communis) during various growth phases (pre-flowering, fluorescence, and fruiting) were examined, as well as their influence on the behavior of adult B. tabaci. The olfactometer studies demonstrated that growth periods of the three plants affected the preference of B. tabaci. Volatiles of piemarker and cotton plants had dissimilar levels of attraction to adults during all stages. Volatile substances released by the castor at the stage of flowering had repellent effect on B. tabaci. In the plant versus plant combination, piemarker volatiles before and during anthesis were most preferred by adults, followed by cotton and then castor. A total of 24, 24, and 20 compounds were detected from volatiles of piemarker, cotton, and castor, respectively, and proportions among the compounds changed during different stages of plant development. The olfactory responses of B. tabaci to volatile compounds presented that linalool and high concentration of (Z)-3-hexenyl acetate had a strong trapping effect on this pest, while nonanal had a significant repellent effect at high concentration. This study indicates that distinct plants and their growth stage affect their attractiveness or repellency to B.tabaci adults, which are mediated by changing plant volatiles. These compounds obtained by analysis screening can be adopted as potential attractants or repellents to control Mediterranean (MED) B. tabaci.

Reproduction Evaluation and Transcription Analysis of Aphis gossypii under Various Photoperiods.

Aphis gossypii Glover (Hemiptera, Aphidoidae) is a polyphagous pest, whose complex phenotypic form, combined with its high fecundity and short reproductive cycle, has caused serious economic losses to agriculture worldwide. Photoperiod plays an important role in the reproduction of aphids. However, the molecular mechanisms underlying its response to seasonal photoperiodic changes are not known. In this study, the effects of different photoperiod treatments (8 L:16 D, 10 L:14 D, 12 L:12 D and 14 L:10 D) on A. gossypii reproduction in the first, third, and fifth generations at low temperatures were investigated. Then, transcriptome sequencing analysis was performed after the fifth generation of A. gossypii, exposed to different photoperiods (8 L:16 D and 12 L:12 D), using high-throughput sequencing technology. The results showed that (I) the effect of photoperiod on aphids was gradually evident with increasing exposure generations. In general, daylight extension help the A. gossypii to reproduce with the optimum photoperiod of L:D 12:12. (II) The transcriptome analysis results showed that 170 differentially expressed genes (DEGs) (123 downregulated and 47 upregulated genes) were identified between aphids under 8 h daylight and 12 h daylight. (III) Gene Ontology (GO) enrichment analysis showed that the DEGs involved in "proteolysis", "metabolic process", "peptidase activity" and "structural molecule activity" were significantly enriched; Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis showed that there were more DEGs in "Longevity regulating pathway-multiple species", "Lysosome", "Endocytosis", "Spliceosome" and "Protein processing in endoplasmic reticulum". (IV) Ten related genes were chosen for validation of statistical analysis based on RNA-Seq by the reverse transcription quantitative (RT-qPCR). The comparison was consistent with the expression pattern and supported the accuracy and reliability of RNA-Seq. In summary, the genes involved in these pathways play an important role in the reproduction of A. gossypii under photoperiodical changes. These will contribute to the sustainable management of cotton aphids through the disruption of their reproduction by the method of RNA interference in the future.

Chromosome-level genome assemblies of two cotton-melon aphid Aphis gossypii biotypes unveil mechanisms of host adaption.

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.

Life Table Construction under Different Temperatures and Insecticide Susceptibility Analysis of Uroleucon formosanum (Hemiptera: Aphididae).

Uroleucon formosanum is an important aphid pest of lettuce, but basic information on its biology is scarce. In this study, effects of three constant temperatures (17, 21, and 25 °C, simulating the mean temperature range in greenhouses) on the development and fecundity of U. formosanum were analyzed by constructing a life table. U. formosanum could develop and reproduce under all three temperatures, but the survival rate, development, and fecundity of U. formosanum were affected by temperature. The intrinsic rate of increase was lowest at 17 °C (0.17) and it was significantly less than at 21 °C (0.20) and 25 °C (0.23). Furthermore, U. formosanum had the lowest finite rate of increase (1.19) and the largest mean generation time (20.21) at 17 °C. These results mean that U. formosanum is less adapted to the lower temperatures (17 °C) among these three set temperatures. To screen insecticides for control, susceptibility of U. formosanum to six insecticides including chlorpyrifos, abamectin, beta-cypermethrin, imidacloprid, nitenpyram, and thiamethoxam was evaluated. U. formosanum was relatively sensitive to all six test insecticides. Chlorpyrifos had the highest toxicity to U. formosanum (LC50 = 3.08 mg/L). These data may help to develop integrated management strategies for better population control of U. formosanum.

The Effect of Torsional Vibration in Longitudinal-Torsional Coupled Ultrasonic Vibration-Assisted Grinding of Silicon Carbide Ceramics.

Rotary longitudinal-torsional coupled ultrasonic vibration-assisted grinding (LTUAG) is a new manufacturing method that can improve the grinding ability of silicon carbide ceramics. However, compared with longitudinal ultrasonic vibration-assisted grinding (LUAG), the role of torsional vibration in the grinding process is unclear. In this study, an effective method for measuring longitudinal-torsional coupled ultrasonic vibration amplitude and an experimental setup for measuring actual amplitude during grinding are proposed. The trajectory of the abrasive grains under the same grinding parameters and the same longitudinal amplitude during LTUAG and LUAG are analysed. Ultrasonic amplitude curves under the condition of tool rotation are then measured and analysed. Finally, the effect of torsional vibration on grinding force and surface roughness under the same grinding conditions is explained. Experimental analysis shows that the introduction of torsional vibration has little effect on the trajectory length and does not change the number of interference overlaps between abrasive grain tracks. Torsional vibration will only increase the cutting speed during grinding and reduce the undeformed chip thickness, which will reduce the grinding force and improve the surface roughness of LTUAG.

The Effect of Mirid Density on Volatile-Mediated Foraging Behaviour of Apolygus lucorum and Peristenus spretus.

Plants would release herbivore-induced plant volatiles (HIPVs) to repel herbivores and attract natural enemies after being damaged by herbivores. In this study, after cotton plants were damaged by different densities of Apolygus lucorum, the behavioral responses of A. lucorum and Peristenus spretus to cotton plants volatiles were evaluated, and the quality and quantity of volatiles from cotton plants were analyzed. Only when cotton plants were damaged by four bugs did both A. lucorum and P. spretus show an obvious response to damaged cotton plants, which indicates that cotton defense is correlated with pest density. The collection and analysis of volatiles reveals that the increase in pest density results in the emission of new compounds and an increase in the total number of volatiles with an alteration in proportions among the compounds in the blend. These changes in volatile profiles might provide wasps and mirids with specific information on host habitat quality and thus could explain the behavioral responses of parasitoids and pests.

Symbiotic microbial studies in diverse populations of Aphis gossypii, existing on altered host plants in different localities during different times.

Complex interactions between symbiotic bacteria and insects ultimately result in equilibrium in all aspects of life in natural insect populations. In this study, abundance of principal symbiotic bacteria was estimated using qPCR in 1553 individuals of aphids, Aphis gossypii. Aphids were sampled from primary and secondary host plants-hibiscus and cotton. Hibiscus aphids were collected from 24 different locations in April, September, and November, whereas cotton aphids were collected between 2015 and 2017 from areas with wide variations in climatic conditions. About 30%-45% aphids were recorded with the most dominant symbiont, Arsenophonus. The other symbionts were in low frequency, and about 7% of aphids were noted with Hamiltonella, Acinetobacter, and Microbacterium, and 3% of aphids were verified with Serratia and Pseudomonas. Aphids infected with Hamiltonella, Arsenophonus, and Serratia can influence Buchnera densities. Hamiltonella has positive interaction with densities of Arsenophonus and Serratia. Almost 100% coinfection of Hamiltonella and Arsenophonus was detected in Xinxiang aphids and 50% coinfection was reported in aphids from North China, while no coinfection was detected in Hainan aphids. These findings describe the prevalence pattern and richness of core community of symbiotic bacteria in naturally occurring populations of A. gossypii and provide new insights for the study of symbiotic bacteria.

Transcriptome analysis of the ovary of beet armyworm Spodoptera exigua under different exposures of cadmium stress.

Heavy metal pollution is becoming an increasingly serious problem globally, and cadmium pollution ranks first in the world. Reproduction in insects is affected by cadmium stress in a dose-dependent manner. However, no previous studies have examined the molecular mechanisms underlying the influence of cadmium exposure on insect reproduction. In this study, RNA-Seq was used to investigate changes in ovary gene expression in newly emerged female beet army worms. The beet armyworms were reared under 4 cadmium concentrations: 0 mg/kg (control), low 0.2 mg/kg (L), medium 12.8 mg/kg (M) and high 51.2 mg/kg (H). Compared with the control (CK), a total of 3453 differentially expressed genes (DEGs) were identified in L cadmium stress, including 1791 up-regulated and 1662 down-regulated candidates; in L versus M groups, 982 up-regulated and 658 down-regulated DEGs; and in M versus H groups, 6508 up-regulated and 2000 down-regulated DEGs were identified and the expression patterns of ten genes were verified by q PCR. Many of the identified DEGs were relevant to juvenile hormone and molting hormone biosynthesis, insulin secretion, estrogen signaling, amino acid metabolism and lipid biosynthesis. These data will provide a molecular prospective to understand the ecological risk of heavy metal pollution and are a resource for selecting key genes as targets in gene-editing/silencing technologies for sustainable pest management.

Identification and expression pattern of putative odorant-binding proteins and chemosensory proteins in antennae of the Microplitis mediator (Hymenoptera: Braconidae).

The parasitoids of Cotton Bollworm Microplitis mediator (Hymenoptera: Braconidae) find their hosts through the odor released by stressed plants. In this study, preliminary characterization and isolation of cDNAs from male M. mediator antennal libraries identified 8 putative odorant-binding proteins (OBPs). Real-time polymerase chain reaction method was used to study the expression pattern of these isolated genes. Their gene expression profiles under a wide range of conditions indicated that only 4 OBP genes in M. mediator were antenna specific. The remaining 4 genes are either expressed ubiquitously or strictly regulated in specialized tissues or during different developmental stages. Some OBP genes were gender specific. These findings support that OBPs play dynamic roles during development of M. mediator and are likely to be involved in broader physiological functions.

Silk Gland Gene Expression during Larval-Pupal Transition in the Cotton Leaf Roller Sylepta derogata (Lepidoptera: Pyralidae).

The cotton leaf roller, Sylepta derogata, is a silk-producing insect pest. While young larvae feed on the underside of leaves, the older ones roll cotton leaves and feed on the leaf edges, which defoliates cotton plants. The larvae produce silk to stabilize the rolled leaf and to balloon from used to new leaves. Despite the significance of silk in the biology of pest insect species, there is virtually no information on the genes involved in their silk production. This is a substantial knowledge gap because some of these genes may be valuable targets for developing molecular pest management technologies. We addressed the gap by posing the hypothesis that silk gland gene expression changes during the transition from larvae to pupae. We tested our hypothesis using RNA-seq to investigate changes in silk gland gene expression at three developmental stages, 5th instar larvae (silk producing; 15,445,926 clean reads), prepupae (reduced silk producing; 13,758,154) and pupae (beyond silk producing; 16,787,792). We recorded 60,298 unigenes and mapped 50,158 (larvae), 48,415 (prepupae) and 46,623 (pupae) of them to the NCBI database. Most differentially expressed genes in the 5th instar larvae/prepupae libraries were relevant to nucleotide synthesis and maintenance of silk gland function. We identified down-regulated transcriptional factors and several genes involved in silk formation in the three libraries and verified the expression pattern of eight genes by qPCR. The developmental- and tissue-specific expression patterns of the fibroin light chain gene showed it was highly expressed during the larval silk-producing stage. We recorded highest expression of this gene in the larval silk gland, compared to other tissues, including midgut, hindgut, epidermis, Malpighian tubes, hemolymph and fat body. These data are a genetic resource to guide selection of key genes that may be targeted for in planta and other gene-silencing technologies for sustainable cotton agriculture.

[Accumulation of lead in Spodoptera exigua (Hübner) and its impact on the population].

The heavy metals pollution is one of the ecological problems which have been paid close attention to Spodoptera exigua Hübner, one of the key pests on vegetables and cotton in the Yangtze River and its southern area in China, has broken out more frequently in recent years. In this study, accumulation of Pb2+ in S. exigua and its excretion were detected when fed with artificial diets with different Pb2+ concentrations (0.3, 1.2, 4.8, 19.2 and 76.8 mg x kg(-1), respectively). Life table was constructed according to the survival and growth of different stages of S. exigua. The effect of lead on three successive generations of beet armyworm was studied using the life table in the laboratory. It was found that the Pb2+ contents were significantly different when S. exigua was reared at different Pb2+ concentrations in the same generation in a significant dose-dependent manner. The concentration of Pb2+ increased with prolonging the stress time at the same time. The Pb2+ concentrations at the three developmental stages of beet armyworm followed the order of larvae > adult > pupa. The beet armyworm could excrete heavy metals by means of faeces, prepupa exuviate and puparium, and the concentrations of Pb2+ in faeces and prepupa exuviate were far higher than in puparium. According to the survival rates of different developmental stages, the female ratios and the egg numbers, the laboratory population life table was constructed. It showed that low doses of Pb2+ promoted the growth of the population, and high doses inhibited the growth, and the index of population trend (I) declined more rapidly with the increase of stress time. The study could provide references for long-term and objective assessment of heavy metal hazard and its effect on the populations of important agricultural pests.