The role of Glutathione-S-transferases in phoxim and chlorfenapyr tolerance in a major mulberry pest, Glyphodes pyloalis walker (Lepidoptera: Pyralidae).

Glyphodes pyloalis Walker is a destructive pest on mulberry trees and poses a significant threat to the sericultural industry in China. Phoxim and chlorfenapyr are two commonly used insecticides in mulberry fields. Glutathione-S-transferases (GSTs) comprise a multifunctional protein superfamily that plays important roles in the detoxification of insecticides and xenobiotic compounds in insects. However, whether GSTs participate in the tolerance of phoxim and chlorfenapyr in G. pyloalis is still unknown. To better understand the mechanism of insecticide tolerance in G. pyloalis, the enzymatic activity of GSTs was evaluated under phoxim and chlorfenapyr exposure, respectively. GST enzyme activity was significantly increased after 12, 36 and 48 h of phoxim treatment and 12, 24, 36 and 48 h of chlorfenapyr treatment. Subsequently, eighteen GST genes were identified from the larvae transcriptome of G. pyloalis. Among these, ten GpGSTs had GSH-binding sites and fifteen GpGSTs had variable hydrophobic substrate-binding sites. The expression levels of Delta-GpGST and Epsilon-GpGST genes were significantly influenced by phoxim and chlorfenapyr treatment, and by the time post insecticide application. Furthermore, after silencing GpGST-E4, the mortality rate of G. pyloalis larvae was increased when they were exposed to chlorfenapyr, but it did not significantly alter when the larvae were exposed to phoxim. Our results indicated the vital roles of GpGSTs in the tolerance of insecticides and this action depends on the categories of insecticides. The present study provides a theoretical basis for elucidating insecticide susceptibility and promotes functional research on GST genes in G. pyloalis.

A role of peptidoglycan recognition protein in mediating insecticide detoxification in Glyphodes pyloalis.

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

Fatty acid synthases and desaturases are essential for the biosynthesis of α-linolenic acid and metamorphosis in a major mulberry pest, Glyphodes pyloalis walker (Lepidoptera: Pyralidae).

BACKGROUND: α-linolenic acid is an essential unsaturated fatty acid in organisms. However, there is a large gap between α-linolenic acid accumulation and its synthesis mechanism in insects. Fatty acid synthases (FASs) and desaturases (Desats) are vital enzymes required for the synthesis of unsaturated fatty acids. RESULTS: The pupae of Glyphodes pyloalis (Lepidoptera: Pyralidae), which is a destructive pest of mulberry trees, contain the highest level of α-linolenic acid compared to other life-history stages. To further explore the synthesis mechanism of α-linolenic acid in G. pyloalis pupae, we constructed a pupal transcriptome dataset and identified 106 genes related to fatty acid metabolism from it. Among these, two fatty acid synthases (GpylFAS) and five desaturases (GpylDesat) were identified. A qRT-PCR validation revealed that GpylFAS1 and GpylDesat1, 2, 3, 5 were expressed highest at pupal stages. Furthermore, the content of α-linolenic acid decreased significantly after silencing GpylFAS1 and GpylDesat5, respectively. Besides, knocking down GpylFAS1 or GpylDesat5 resulted in more malformed pupae and adults, as well as lower emergence rates. Meanwhile, silencing GpylFAS1 or GpylDesat5 affected the expressions of the other GpylFASs and GpylDesats. CONCLUSION: The present results illustrate the pivotal function of FASs and Desats in α-linolenic acid biosynthesis and metamorphosis in insects. Our research also broadens the sources of unsaturated fatty acids, especially for α-linolenic acid from insects, and provides novel insights for the management of mulberry insect pests from the perspective of utilization rather than control. © 2022 Society of Chemical Industry.

Characterization, and Functional Analysis of Hsp70 and Hsp90 Gene Families in Glyphodes pyloalis Walker (Lepidoptera: Pyralidae).

Glyphodes pyloalis Walker (Lepidoptera: Pyralidae) is a destructive mulberry pest, causing great damage to mulberry in China. Heat shock proteins (Hsps) are involved in various signal pathways and regulate lots of physiological processes in insects. The function of Hsps in G. pyloalis, however, has still received less attention. Here, we identified five Hsp genes from G. pyloalis transcriptome dataset including two Hsp70 family genes (GpHsp71.3 and GpHsp74.9) and three Hsp90 family genes (GpHsp82.4, GpHsp89, and GpHsp93.4). Quantitative Real-time PCR validation revealed that all Hsps of G. pyloalis have significant expression in pupal and diapause stage, at which the larvae arrest the development. Expressions of GpHsp71.3 and GpHsp82.4 were increased significantly after thermal treatment at 40°C, and this upregulation depended on heat treatment duration. Furthermore, silencing GpHsp82.4 by RNA interference led to a significant increase in mortality of G. pyloalis larvae under the heat stress compared to the control group. After starvation stress, the expression levels of GpHsp82.4 and GpHsp93.4 were significantly increased. At last, after being parasitized by the parasitoid wasp Aulacocentrum confusum, Hsp70 and Hsp90 genes of G. pyloalis were decreased significantly in the early stage of parasitization and this moderation was affected by time post-parasitization. This study highlights the function of G. pyloalis Hsps in response to environmental stress and provides a perspective for the control of this pest.