Differential Antioxidant Enzyme Gene Expression and Functional Analysis of Pyridaben-Susceptible and -Resistant Strains of Tetranychus truncatus (Acari: Tetranychidae) under High Temperature Stress.

Tetranychus truncatus (Acari: Tetranychidae) has caused serious economic losses on some crops (soybean, corn, and cotton) in China, and has developed resistance to most acaricides. Our laboratory study found that T. truncatus was resistant to pyridaben and also adapted to high temperature (34-40 °C). High temperature stress may cause arthropods to produce a large amount of reactive oxygen species (ROS), causing oxidative damage. Antioxidant enzymes, as the main antioxidants, can reduce the damage caused by excessive ROS in arthropods. In order to study the adaptation mechanism of the pyridaben-resistant strain of T. truncatus to high temperature and the role of antioxidant enzyme genes under high temperature stress, four antioxidant enzyme genes, TtSOD, TtPOD3, TtPOD4, and TtGSTs2, were screened according to the transcriptome sequencing data of pyridaben-susceptible and -resistant strains in T. truncatus. Firstly, the phylogeny and structure analyses of these four genes were carried out. Then, real-time quantitative PCR (RT-qPCR) technology was used to analyze the gene expression patterns of antioxidant enzymes in two strains of T. truncatus at three different high temperature ranges (34 °C, 38 °C, and 42 °C). The results showed that the expression levels of four antioxidant enzyme genes of two strains of T. truncatus were induced by high temperature stress, and the expression levels of antioxidant enzyme genes were significantly different in each development state. The gene expression of antioxidant enzyme genes in resistant strains at the adult stage was significantly higher than that in susceptible strains. After the TtSOD and TtPOD4 genes of adult mites of the resistant strain were silenced by RNA interference (RNAi) technology, the mortality rate of mites with TtPOD4 gene silencing reached 41.11% after 96 h at 34 °C, which was significantly higher than that of the control and TtSOD gene silencing. It has been confirmed that the TtPOD4 gene plays a key role in the adaptation of pyridaben-resistant strain of T. truncatus to high temperature. It lays a theoretical foundation for revealing the thermal adaptation mechanism of T. truncatus.

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

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

Changes in the bacterial communities of Harmonia axyridis (Coleoptera: Coccinellidae) in response to long-term cold storage and progressive loss of egg viability in cold-stored beetles.

Bacteria have a profound influence on life history and reproduction of numerous insects, while the associations between hosts and bacteria are substantially influenced by environmental pressures. Cold storage is crucial for extending the shelf life of insects used as tools for biological control, but mostly causes detrimental effects. In this study, we observed a great decrease in egg hatch rate of cold-stored Harmonia axyridis during the later oviposition periods. Furthermore, most eggs produced by their F1 offspring exhibited complete loss of hatchability. We hypothesized that long-term exposure to cold may greatly alter the bacterial community within the reproductive tracts of H. axyridis, which may be an important factor contributing to the loss of egg viability. Through sequencing of the 16S rRNA gene, we discovered considerable changes in the bacterial structure within the reproductive tracts of female cold-stored beetles (LCS_F) compared to non-stored beetles (Control_F), with a notable increase in unclassified_f_Enterobacteriaceae in LCS_F. Furthermore, in accordance with the change of egg hatchability, we observed a slight variation in the microbial community of eggs produced by cold-stored beetles in early (Egg_E) and later (Egg_L) oviposition periods as well as in eggs produced by their F1 offspring (Egg_F1). Functional predictions of the microbial communities revealed a significant decrease in the relative abundance of substance dependence pathway in LCS_F. Moreover, this pathway exhibited relatively lower abundance levels in both Egg_L and Egg_F1 compared to Egg_E. These findings validate that long-term cold storage can greatly modify the bacterial composition within H. axyridis, thereby expanding our understanding of the intricate bacteria-insect host interactions.

Canola bee pollen is an effective artificial diet additive for improving larval development of predatory coccinellids: a lesson from Harmonia axyridis.

BACKGROUND: Pollen is a common plant-derived food source for predatory ladybird beetles under field conditions, yet the potential for pollen to improve the quality of artificial diets remains largely unexplored. In this study, we developed three pollen diets by incorporating varying proportions of canola bee pollen (7.5%, 15.0% and 22.5% with 2.5%, 5.0%, and 7.5% of water, respectively) into a conventional diet. The feeding efficiency of Harmonia axyridis, an omnivorous predator, was evaluated and compared on three pollen diets, a conventional nonpollen diet and pea aphids. RESULTS: The larvae fed a medium or high pollen diet exhibited significantly higher survival in the 4th instar, pupa and adult stages than those fed a nonpollen diet. These larvae also developed into significantly heavier adults, and their survival rates in adulthood were comparable to those fed pea aphids. Specifically, we revealed the underlying mechanisms through which a high pollen diet enhances pupal development. Consumption of high pollen diet versus nonpollen diet resulted not only in a significant decrease in pupal glycogen content, but also an increase in adult lipid content. Both diet treatments induced similar changes in carbohydrate and glycogen content compared to the aphid diet while exhibiting different alterations in pupal protein content and adult lipid content. Furthermore, the transcriptome analysis revealed that the nutrient metabolism, immune response, and cuticle development pathways were predominantly enriched among the differentially expressed genes (DEGs). CONCLUSION: Canola bee pollen offers diverse advantages in terms of rearing H. axyridis larvae with an artificial diet, which will advance the development of effective diets for predaceous coccinellids. © 2024 Society of Chemical Industry.

The influence of high-temperature frequency variation on the life-history traits of pyridaben-sensitive and -resistant strains of Tetranychus truncatus.

With a generally warming global climate, the number of Tetranychus truncatus specimens in the Hexi region in China has been increasing. As ectotherms, the growth and development of T. truncatus are greatly affected by changes in environmental temperature. The effect of heatwaves on organisms depends on a delicate balance between damage and repair periods. Therefore, we simulated nine patterns of periodically recurring changes in the frequency of high-temperature days using an intraday gradual temperature change model to study and compare the effects on the development and reproduction of pyridaben-sensitive and -resistant strains of T. truncatus. The results showed that the influence of the frequency of high-temperature days on developmental stages, longevity and fecundity was different between the two strains. The egg and immature stages of the sensitive strain were all affected by hot days, whereas the adult stage was less affected by the frequency. The egg stage of the resistant strain was less affected; it was mainly affected in the immature and adult stages. Under the moderate condition of increasing the proportion of days at normal temperature, the longevity of the resistant strain gradually increased and reached a maximum at a 1:3 frequency, and then it decreased with the increase in high-temperature days. The longevity of the sensitive strain was less affected by frequency, and there was no significant difference between most treatment and control groups. In addition, both sensitive and resistant strains were able to complete growth and development under all nine frequencies of high-temperature days, but the reproductive rate was lower than it was at normal temperatures, indicating that both strains of T. truncatus adapted to high temperatures at the expense of reduced reproduction rates. This lays a key theoretical foundation for predicting the occurrence of agricultural pest populations under the background of climate warming and developing appropriate control strategies.

Effects of Fusarium solani on the Growth and Development of Anoplophora glabripennis Larvae.

Anoplophora glabripennis (Asian longhorned beetle) is a wood-boring pest that can inhabit a wide range of healthy deciduous host trees in native and non-native habitats. Lignocellulose degradation plays a major role in the acquisition of nutrients during the growth and development of A. glabripennis larvae. In this study, the lignocellulose degradation capacity of Fusarium solani, a fungal symbiont of A. glabripennis, was investigated in fermentation culture and in four host tree species. The impact of F. solani on larval growth and survival parameters was assessed. Fermentation culture demonstrated continuous and stable production of lignocellulolytic enzymes over the cultivation period. Furthermore, F. solani was able to degrade host tree lignocellulose, as shown by decreased soluble sugar and cellulose contents and an increase in protein content. No significant differences in larval survival were observed in larvae fed with or without F. solani. However, weight and head capsule width were higher in larvae fed on F. solani, and gut lignocellulose activities were elevated in fed larvae. Our results indicate a role for F. solani in the predigestion of lignocellulose during the colonization and parasitic stages of A. glabripennis larval development, and also the F. solani an important symbiotic partner to A. glabripennis, lowering barriers to colonization and development in a range of habitats.

The role of cytochrome P450 3A2 and 4V2 in response to high-temperature stress in Tetranychus truncatus (Acari: Tetranychidae).

Temperature is an important factor influencing the physiological activities of agricultural pests. Therefore, understanding pest physiological activities and the molecular response to high-temperature stress is of paramount importance for pest management. Tetranychus truncatus Ehara (Acari: Tetranychidae) is a harmful organism, that may cause serious harm to crops such as corn and cotton in high-temperature environments. Cytochrome P450 (CYP450) is induced by high-temperature stress, and it plays an important role in the resistance of spider mites to high temperatures. Because of their role in high-temperature tolerance, the cytochrome P450 genes of the spider mite are attracting more and more attention. In this study, we identified and analyzed CYP450 genes in T. truncatus to investigate their potential roles in growth and development and the resistance to high-temperature stress. Based on phylogenetic and structural analyses, we identified 17 CYP450 genes in T. truncatus. RNA-seq and real-time quantitative PCR (RT-qPCR) revealed differential expression patterns of these genes at different developmental stages and levels of high-temperature stress resistance. The RNA interference results of selected CYP450 showed that when TtCYP3A2 and TtCYP4V2 were silenced by feeding on dsRNA, respectively, the high-temperature resistance of T. truncatus was decreased, which indicated that the expression of these two CYP450 genes in this species may be related to high-temperature tolerance. Our results provide potential evidence for the response of spider mites to high-temperature stress and help to improve the understanding of T. truncatus's ability to resist high-temperature stress.

Functional Analysis of Amino Acid Transporter Genes ACYPI000536 and ACYPI004320 in Acyrthosiphon pisum.

In recent years, pea aphids have become major pests of alfalfa. Our previous study found that "Gannong 5" is a highly aphid-resistant alfalfa variety and that "Lie Renhe" is a susceptible one. The average field susceptibility index of "Gannong 5" was 31.31, and the average field susceptibility index of "Lie Renhe" was 80.34. The uptake and balance of amino acids in insects are usually dependent on amino acid transporters. RT-qPCR was used to detect the relative expression levels of seven amino acid transporter differential genes in the different instar pea aphids fed on resistant and susceptible alfalfa varieties after 24 h, and two key genes were selected. When pea aphids fed on "Gannong 5", the expression of ACYPI004320 was significantly higher than that in pea aphids fed on "Lie Renhe"; however, the expression of ACYPI000536 was significantly lower than that in pea aphids fed on "Lie Renhe". Afterward, the RNA interference with pea aphid ACYPI000536 and ACYPI004320 genes was performed using a plant-mediated method, and gene function was verified via liquid chromatography-mass spectrometry and pea aphid sensitivity to aphid-resistant and susceptible alfalfa varieties. The results showed that the down-regulation of the ACYPI000536 gene expression led to an increase in the histidine and lysine contents in pea aphids, which, in turn, led to an increase in mortality when pea aphids fed on the susceptible variety "Lie Renhe". The down-regulation of the ACYPI004320 gene expression led to an increase in phenylalanine content in pea aphids, which, in turn, led to a decrease in mortality when pea aphids fed on the resistant variety "Gannong 5".

Genetic diversity of Venturia inaequalis isolates from the scabs in apple trees in Gansu Province, China, using AFLP markers.

Apple scab is a serious disease that restricts the growth of cultivated apples. The objective of this study is to investigate the genetic variations and genetic structure of Venturia inaequalis in Gansu Province, China. 108 isolates of the pathogen V. inaequalis from the Jingning, Lingtai, Jingchuan, Xifeng, Ning and Maiji regions were collected, and their genetic diversity was analyzed using AFLP molecular marker technique. The results showed that genetic diversity was present among the isolates but was not statistically significant. Genetic distance values ranged from 0.0095 to 0.0762. Cluster analysis results showed that the 108 isolates could be divided into two clusters using a similarity coefficient of 0.69. A total of 104 isolates were contained in cluster I while four isolates were contained in Cluster II. From the AMOVA analysis, 98% of variations were observed within the same region, while 2% were observed across different regions. The analysis of population structure showed that 108 isolates had two common ancestors, with the Jingning isolates mainly being derived from the red ancestor. PCoA analysis showed that the Jingning isolates were independent to a certain extent. The different geographical location caused the genetic difference of the isolates. The genetic diversity of apple scab in Gansu Province is greatly aided by this work, which also offers a theoretical foundation for the use of molecular markers in assisted breeding to create novel resistant types.

Transcriptional response and functional analysis of ATP-binding cassette transporters to tannic acid in pea aphid, Acyrthosiphon pisum (Harris).

Although tannins are widely distributed in broad beans and alfalfa, the pea aphid (Acyrthosiphon pisum) can still destroy them. The ATP binding cassette (ABC) transporters participate in the metabolism of plant secondary metabolites and pesticides in insects. However, whether ABC transporter genes play a role in the metabolism of tannins in the pea aphid is unclear. Here, we found that verapamil (an ABC transporter inhibitor) significantly increased the mortality of tannic acid to pea aphid, which indicated that ABC transporter gene was related to the metabolism of tannic acid by pea aphid. Then, we identified 54 putative ABC transporter genes from the genome database of A. pisum. These genes were divided into eight subfamilies, ApABCA to ApABCH, of which subfamily G has the largest number of genes with 19, followed by the subfamily C with 14. RT-qPCR results show that the expression levels of ApABCA2, ApABCC7, ApABCG2, and ApABCG3 were highly expressed in the first instar, while those of ApABCA3, ApABCG6, ApABCG7, ApABCH3, and ApABCH4 were highly expressed in adults. Furthermore, transcription levels of many ABC transporter genes were induced by tannic acid. Especially, ApABCG17 and ApABCH2 were obviously induced after being exposed to tannic acid. Meanwhile, knockdown of ApABCG17 by RNA interference resulted in increased sensitivity of pea aphid to tannic acid. These results suggest that ApABCG17 may be involved in tannic acid metabolism in pea aphid. This study will help us to understand the mechanism of tannic acid metabolism in pea aphid, and provides a basis for further research on the physiological function of ABC transporter genes in pea aphid.

Mongolian pine forest decline by the combinatory effect of European woodwasp and plant pathogenic fungi.

Interactions between the decline of Mongolian pine woodlands and fungal communities and invasive pests in northeastern China are poorly understood. In this study, we investigated the fungal communities occurring in three tree samples: the woodwasp Sirex noctilio infested, healthy uninfested and unhealthy uninfested Mongolian pine trees. We analyzed the relationships of the Mongolian pine decline with fungal infection and woodwasp infestation. Twenty-six fungal species were identified from the sampled trees. Each tree sample harbored a fungal endophyte community with a unique structure. Pathogenic fungi richness was four times higher in infested and unhealthy un-infested trees compared to that in healthy uninfested trees. Sphaeropsis sapinea was the most dominant pathogenic fungus in the sampled Mongolian pine trees. The number of S. noctilio was higher than native bark beetles in the declining Mongolian pine trees. The invasion of the woodwasp appeared to be promoted by the fungal infection in the Mongolian pine trees. The incidence of S. noctilio infestation was higher in the fungi infected trees (83.22%) than those without infection (38.72%). S. sapinea population exhibited positive associations with within-tree colonization of S. noctilio and bark beetle. Collectively, these data indicate that the fungal disease may have caused as the initial reason the decline of the Mongolian pine trees, and also provided convenient conditions for the successful colonization of the woodwasp. The woodwasps attack the Mongolian pine trees infected by fungi and accelerated its decline.

Evidence of the Involvement of a Plus-C Odorant-Binding Protein HparOBP14 in Host Plant Selection and Oviposition of the Scarab Beetle Holotrichia parallela.

Holotrichia parallela is one of the agriculturally important scarab beetle pests in China. In this study, HparOBP14 was cloned, which is the most abundantly expressed among the OBP genes in the legs of female H. parallela adults. Sequence comparison and phylogenetic analysis showed that HparOBP14 has a Plus-C structure motif. The expression profile analysis revealed that HparOBP14 expression was the highest in the female antennae and then in the legs. The fluorescence competitive binding experiment of the recombinant HparOBP14 protein showed that HparOBP14 had an affinity with 6-methyl-5-heptene-2-one (plant volatile), 3-methylindole, p-cymene, methanol, formaldehyde, α-pinene, and geraniol (organic fertilizer volatile). Knockdown HparOBP14 expression decreased significantly the EAG response of the injected female adults to p-cymene, methanol, formaldehyde, α-pinene, and geraniol. Similarly, the injected female adults were significantly less attracted to geraniol and methanol. Therefore, HparOBP14 might bind organic matter volatiles during oviposition. These results are not only helpful to analyze the olfactory recognition mechanism of female adult H. parallela when choosing suitable oviposition sites, but also to provide target genes for green prevention and control of H. parallela in the future.

Chemical Identity and Functional Characterization of Semiochemicals That Promote the Interactions between Rice Plant and Rice Major Pest Nilaparvata lugens.

The interaction between food crops and insect pests is mediated by semiochemicals emitted from host plants. These semiochemicals are natural behavioral modifiers and act on the insect olfactory system to locate hosts and preys. In this study, eight rice neuroactive semiochemicals were identified from rice varieties by GC-EAG and GC-MS. Their ability to modify rice pest behaviors was further studied as individual chemicals and physiologically relevant blend. The total amount of each semiochemical and the expression of their biosynthesis genes were significantly higher in pest susceptible variety than in pest-resistant variety and upregulated by the infestation of the pest Nilaparvata lugens (BPH). The semiochemicals emitted by uninfested plants (UIRVs) were more attractive to BPHs. Interestingly, the attractiveness of UIRVs was significantly reduced by the addition of the blend that mimics the natural composition of these semiochemicals emitted by infested plants (IRVs). Our study suggests a mechanism for the spread of pest infestation from infested plants to uninfested plants nearby. UIRVs initially serve as attractive signals to rice insect pests. The pest infestation changes the rice semiochemical profile to be less attractive or even repellent, which pushes further colonization to uninfested plants nearby. The identified semiochemicals can be used for crop protection based on a push-pull strategy.

Investigating reproductive success of the ladybird beetle Harmonia axyridis from the perspective of micropyle variation.

Micropyles in insects are small openings that allow sperm entry into, and the number was usually decreased on unfertilized and (or) undeveloped eggs. However, reports showed that Harmonia axyridis, a reproductive success model, deposited similar number of micropyles on undeveloped and developing eggs. Thus, it was confusing whether micropyles in H. axyridis were unaffected. To solve this confusion, two experiments were conducted here. Firstly, virgin female and four different days delayed mating (DDM) experiments were conducted to reveal the effects of fertilization stimulus and delayed-fertilization. Secondly, intercrosses between a light-colored mutant (HAM, an adaptive deficiency) and wild type (HAW) were conducted to further reveal whether there were female and male interactions. We found that (1) eggs produced by virgin and DDM females had significantly less micropyles than control. Even so, more than 18 micropyles were observed on eggs following fertilization and, consequently, egg production as well as hatch rate was not negatively affected by mating delay; (2) number of micropyles was significantly varied among the four reciprocal crosses and virgin HAW female. Specifically, the heterozygous eggs (Cross-D) and wild-type homozygous eggs (Cross-B) respectively had the least and maximum micropyles, and eggs from virgin HAW female had significantly less micropyles compared to those from HAW female (Cross-B or Cross-C), but the number was significantly higher than those from HAM female (Cross-A or Cross-D). These results informed us that the number of micropyles in H. axyridis is plastic but maintaining a high-quantity that offers many benefits, which should contribute to its reproduction success.

[Changes of phenols and lignin contents in alfalfa leaf damaged by Odontothrips loti].

In order to investigate the changes of secondary metabolites content of alfalfa induced by thrips Odontothrips loti damaging, two alfalfa strains, one resistant to thrips (R-1) and the other susceptible to thrips (I-1) , were chosen to measure the phenols and lignin contents of alfalfa leaves under infestation with thrips at different densities (0, 1, 3, 5, 7 thrips x branch(-1), and 0 thrip x branch(-1) as control). After infestation 7 days, the polyphenols, tannin and condensed tannin contents increased in both leaves of R-l and I-i with the increasing thrips density, the simple phenols content had no significant difference, while the lignin content increased significantly compared with the control. After infestation 14 days, the polyphenols, tannin, condensed tannin and lignin contents in both leaves of R-1 and I-1 increased obviously with the increasing thrips density, while the simple phenols content had no significant difference. The lignin content increased significantly, and was significantly higher under 7 thrips x branch(-1) than under the control. After infestation 21 days, the polyphenols, tannin, and lignin contents in both leaves of R-1 and I-1 increased obviously with the increasing thrips density, and were the highest under 7 thrips x branch(-1). Simple phenols content of I-1 strain was increased significantly, but that of R-1 strain had no significant change. The condensed tannin content in both leaves of R-1 and I-1 was not obvious compared with the control. Phenols and lignin contents in R-1 and I-1 leaves increased obviously after thrips infestation, and the polyphenols, tannin and lignin contents increased faster in R-1 strain than in I-1 strain. Thrips infestation had inductive effects on phenols and lignin contents of alfalfa, which could be used to evaluate the resistance of alfalfa.