Odorant Receptor BdorOR49b Mediates Oviposition and Attraction Behavior of Bactrocera dorsalis to Benzothiazole.

The ability to recognize a host plant is crucial for insects to meet their nutritional needs and locate suitable sites for laying eggs. Bactrocera dorsalis is a highly destructive pest in fruit crops. Benzothiazole has been found to induce oviposition behavior in the gravid B. dorsalis. However, the ecological roles and the olfactory receptor responsible for benzothiazole are not yet fully understood. In this study, we found that adults were attracted to benzothiazole, which was an effective oviposition stimulant. In vitro experiments showed that BdorOR49b was narrowly tuned to benzothiazole. The electroantennogram results showed that knocking out BdorOR49b significantly reduced the antennal electrophysiological response to benzothiazole. Compared with wild-type flies, the attractiveness of benzothiazole to BdorOR49b knockout adult was significantly attenuated, and mutant females exhibited a severe decrease in oviposition behavior. Altogether, our work provides valuable insights into chemical communications and potential strategies for the control of this pest.

Cd pollution in agroforestry ecosystems: An abiotic factor affecting the passive and active lethal efficiency of Beauveria bassiana to Lymantria dispar larvae.

This study evaluated the effects of cadmium (Cd) exposure on the passive and active lethal efficiency of Beauveria bassiana (Bb) to Lymantria dispar larvae and analyzed the corresponding mechanism from mycelial vegetative growth, fungal and host nutrient competition, and fungal spore performance. The results showed that the passive lethal efficiency of Bb to Cd-exposed L. dispar larvae was significantly higher than that of larvae not exposed to Cd. After Bb infection, the fungal biomass in living larvae and the mycelium encapsulation index of dead larvae were significantly decreased under Cd exposure. Cd exposure damaged the mycelial structure, as well as inhibited the mycelial growth and sporulation quantity. A total of 15 and 39 differentially accumulated mycotoxin metabolites were identified in Bb mycelia treated with low Cd and high Cd, respectively, and the contents of these differentially accumulated mycotoxins in the low Cd and high Cd treatment groups were overall lower than those in the control group. Nutrient content and energy metabolism-related gene expression were significantly decreased in Cd-exposed larvae, both before and after Bb infection. Trehalose supplementation alleviated the nutritional deficiency of larvae under the combined treatment of Cd and Bb and decreased the larval susceptibility to Bb. Compared with untreated Bb, the lethal efficiency of low Cd-exposed Bb to larvae increased significantly, while high Cd-exposed Bb was significantly less lethal to larvae. Cd exposure promoted at low concentrations but inhibited the hydrophobicity and adhesion of spores at higher concentrations. Spore germination rate and stress resistance of Bb decreased significantly under the treatment of both Cd concentrations. Taken together, heavy metals can be regarded as an abiotic environmental factor that directly affects the lethal efficiency of Bb to insect pests.

Identification and potential application of key insecticidal metabolites in Tilia amurensis, a low-preference host of Hyphantria cunea.

Developing effective insecticidal strategies is an important means of reducing the spread and host plant damage by Hyphantria cunea. In this study, key metabolites with insecticidal activity against H. cunea were screened by targeted metabolomics in Tilia amurensis, a low-preference host plant. Subsequently, the potential of key metabolites that could be used as botanical pesticides was evaluated. The results showed that coumarin was the key insecticidal metabolite of T. amurensis and had a significant insecticidal effect and weight inhibition effect on H. cunea larvae. Coumarin treatment significantly decreased the larval nutrient content and the gene expression of rate-limiting enzymes in the glycolytic pathway and tricarboxylic acid cycle. A significantly enhanced detoxification enzyme activity (CarE and GST), antioxidant oxidase activity (SOD and CAT), non-enzymatic antioxidant levels (GSH), and total antioxidant capacity were observed in coumarin-treated larvae. Coumarin treatment resulted in a significant increase in the expression levels of detoxification enzyme genes (CarE1, CarE2, CarE3, GST2, and GST3) and antioxidant oxidase genes (SOD1, CAT1, and CAT2) in H. cunea larvae. Coumarin treatment significantly increased the levels of MDA and H2O2 in larvae but did not cause pathological changes in the ultrastructure of the larval midgut. Coumarin solution sprayed directly or as a microcapsule suspension formulation with coumarin as the active ingredient had significant insecticidal activity against the H. cunea larvae. Overall, coumarin, a key anti-insect metabolite identified from T. amurensis, can significantly inhibit the growth and survival of H. cunea larvae and has the potential to be developed as a botanical pesticide.

A new insight into Cd exposure-induced hemocyte reduction in Lymantria dispar larvae: Involvement of the ROS-ATF6-ER stress-apoptosis pathway.

Hemocytes are important targets for heavy metal-induced immunotoxicity in insects. This study aimed to investigate the mechanism by which cadmium (Cd) exposure affects the hemocyte count in Lymantria dispar larvae. The results showed that the number of larval hemocytes was significantly decreased under Cd exposure, accompanied by a significant increase in the apoptosis rate and the expression of Caspase-3. The endoplasmic reticulum (ER) of hemocytes in the Cd-treated group showed irregular swelling. Expression levels of ER stress indicator genes (CHOP, Bip1, Bip2, Bip3, and Bip4) were significantly higher in the Cd-treated group. Among the three pathways that potentially mediate ER stress, only the key genes in the ATF6 pathway (ATF6, S1P-1, S1P-2, and WFS1) exhibited differential responses to Cd exposure. Cd exposure significantly increased the levels of reactive oxygen species (ROS) and the expression of oxidative stress-related genes (CNCC, P38, and ATF2) in hemocytes. Studies using inhibitors confirmed that apoptosis mediated the decrease in hemocyte count, ER stress mediated apoptosis, ATF6 pathway mediated ER stress, and ROS or oxidative stress mediated ER stress through the activation of the ATF6 pathway. Taken together, the ROS-ATF6-ER stress-apoptosis pathway is responsible for the reduction in the hemocyte count of Cd-treated L. dispar larvae.

The exposure risk of heavy metals to insect pests and their impact on pests occurrence and cross-tolerance to insecticides: A review.

Heavy metal (HM) pollution is a severe global environmental issue. HMs in the environment can transfer along the food chain, which aggravates their ecotoxicological effect and exposes the insects to heavy metal stress. In addition to their growth-toxic effects, HMs have been reported as abiotic environmental factors that influence the implementation of integrated pest management strategies, including microbial control, enemy insect control, and chemical control. This will bring new challenges to pest control and further highlight the ecotoxicological impact of HM pollution. In this review, the relationship between HM pollution and insecticide tolerance in pests was analyzed. Our focus is on the risks of HM exposure to pests, pests tolerance to insecticides under HM exposure, and the mechanisms underlying the effect of HM exposure on pests tolerance to insecticides. We infer that HM exposure, as an initial stressor, induces cross-tolerance in pests to subsequent insecticide stress. Additionally, the priming effect of HM exposure on enzymes associated with insecticide metabolism underlies cross-tolerance formation. This is a new interdisciplinary field between pollution ecology and pest control, with an important guidance value for optimizing pest control strategies in HM polluted areas.

A classic screening marker does not affect antennal electrophysiology but strongly regulates reproductive behaviours in Bactrocera dorsalis.

The key phenotype white eye (white) has been used for decades to selectively remove females before release in sterile insect technique programs and as an effective screening marker in genetic engineering. Bactrocera dorsalis is a representative tephritid pest causing damage to more than 150 fruit crops. Yet, the function of white in important biological processes remains unclear in B. dorsalis. In this study, the impacts of the white gene on electrophysiology and reproductive behaviour in B. dorsalis were tested. The results indicated that knocking out Bdwhite disrupted eye pigmentation in adults, consistent with previous reports. Bdwhite did not affect the antennal electrophysiology response to 63 chemical components with various structures. However, reproductive behaviours in both males and females were significantly reduced in Bdwhite-/- . Both pre-copulatory and copulation behaviours were significantly reduced in Bdwhite-/- , and the effect was male-specific. Mutant females significantly delayed their oviposition towards γ-octalactone, and the peak of oviposition behaviour towards orange juice was lost. These results show that Bdwhite might not be an ideal screening marker in functional gene research aiming to identify molecular targets for behaviour-modifying chemicals. Instead, owing to its strong effect on B. dorsalis sexual behaviours, the downstream genes regulated by Bdwhite or the genes from white-linked areas could be alternate molecular targets that promote the development of better behavioural modifying chemical-based pest management techniques.

MFS Transporter Bdorwp Does Not Affect Antennal Electrophysiology but Regulates Reproductive Behaviors in Bactrocera dorsalis.

Developing behavioral modifying chemicals through molecular targets is a promising way to improve semiochemical-based technology for pest management. Identifying molecular targets that affect insect behavior largely relies on functional genetic techniques such as deletions, insertions, and substitutions. Selectable markers have thus been developed to increase the efficiency of screening for successful editing events. However, the effect of selectable markers on relevant phenotypic traits needs to be considered. In this study, we cloned the wp gene ofBactrocera dorsalis. Knocking out Bdorwp causes white pupae phenotypes. Reproductive behaviors in both males and females were strongly regulated by Bdorwp. Remarkably, Bdorwp did not affect the antennal electrophysiology response to 63 chemical components with various structures. It is recommended to indirectly apply Bdorwp as a selectable marker in functional gene research on behavioral modifying chemicals. Moreover, Bdorwp could also be a potential molecular target for developing new insecticides for tephritid species control.

Assessment of cytisine as an insecticide candidate for Hyphantria cunea management: Toxicological, biochemical, and control potential insights.

In the present study, the toxicological effects of cytisine on the H. cunea larvae were investigated, and the potential of cytisine as a botanical insecticide through field simulation experiments was evaluated. The results showed that cytisine treatment (0.25-2.5%) exerted significant biotoxic effects on the H. cunea larvae, including diminished weight, disruption of both positive (HcCKS1, HcPLK, HcCCNA) and negative (HcGADD and HcCDKN) regulatory genes associated with larval growth, increased mortality, and heightened oxidative damage (H2O2 and MDA). Cytisine treatment significantly reduced glucose content and inhibited the expression of key rate-limiting enzyme genes (HcPFK, HcPK, HcHK1, HcCS, and HcIDH2) within glycolysis and the tricarboxylic acid cycle pathways. Under cytisine treatment, detoxification enzyme activities (CarE and GST) and expression of detoxification genes (HcCarE1, HcCarE2, HcCarE3, HcGST1, and HcGST3) were inhibited in H. cunea larvae. An increased contents of SOD, CAT, ASA and T-AOC, as well as expression of antioxidant enzyme genes HcSOD1 and HcCAT2, was found in cytisine-treated H. cunea larvae. Simultaneously, this is accompanied by a significant reduction in the expression of four antioxidant enzyme genes (e.g., HcPOD1 and HcPOD2). In the field experiment, a cytisine aqueous solution (25 g/L) with pre-sprayed and directly sprayed ways demonstrated potent insecticidal activity against H. cunea larvae, achieving a mortality rate of 53.75% and 100% at 24 h, respectively. Taken together, cytisine has significantly weight inhibition and lethal toxicity on the H. cunea larvae, and can be developed as a botanical insecticide for H. cunea control.

Ultrastructural and Descriptive Study on the Adult Body Surface of Heortia vitessoides (Lepidoptera: Crambidae).

Heortia vitessoides Moore, 1885 (Lepidoptera: Crambidae) is an economically important lepidopteran pest that caused severe damage to the plantation area of Aquilaria sinensis (Lour.) Gilg, 1825 (Thymelaeaceae), resulting in extensive defoliation of the trees during an epidemic. In this study, we used scanning electron microscopy (SEM) to analyze the external morphology and ultrastructure of sensilla on various body parts of H. vitessoides. Specifically, seven, four, four, and five types of sensilla were found, respectively, on the antennae, proboscis, labial palps, and legs. We described the types, distributions, and sexual dimorphism of these sensilla on antennae, and found that the number and size of sensilla differed significantly between males and females. This study provides crucial information for future investigations into the function of these sensilla in H. vitessoides.

Digestive Characteristics of Hyphantria cunea Larvae on Different Host Plants.

Digestive physiology mediates the adaptation of phytophagous insects to host plants. In this study, the digestive characteristics of Hyphantria cunea larvae feeding preferences on different host plants were investigated. The results showed that the body weight, food utilization, and nutrient contents of H. cunea larvae feeding on the high-preference host plants were significantly higher than those feeding on the low-preference host plants. However, the activity of larval digestive enzymes in different host plants presented an opposite trend, as higher α-amylase or trypsin activity was observed in the group feeding on the low-preference host plants than that feeding on the high-preference host plants. Upon treatment of leaves with α-amylase and trypsin inhibitors, the body weight, food intake, food utilization rate, and food conversion rate of H. cunea larvae significantly decreased in all host plant groups. Furthermore, the H. cunea comprised highly adaptable compensatory mechanisms of digestion involving digestive enzymes and nutrient metabolism in response to digestive enzyme inhibitors. Taken together, digestive physiology mediates the adaptation of H. cunea to multiple host plants, and the compensatory effect of digestive physiology is an important counter-defense strategy implemented by H. cunea to resist plant defense factors, especially the insect digestive enzyme inhibitors.

Susceptibility of Hyphantria cunea larvae to Beauveria bassiana under Cd Stress: An integrated study of innate immunity and energy metabolism.

Biological control is widely used for integrated pest management. However, there are many abiotic factors that can affect the biocontrol efficiency. In this study, we investigated the susceptibility of Hyphantria cunea larvae to Beauveria bassiana under Cd stress, and the corresponding mechanism was analyzed around innate immunity and energy metabolism. The results showed that mortality of H. cunea larvae treated with Cd and B. bassiana was significantly higher than those treated with B. bassiana alone, and the combined lethal effect exhibited a synergistic effect. Compared with the single fungal treatment group, the total hemocyte count in the combined Cd and fungal treatment group decreased significantly, accompanied by a decrease in phagocytosis, encapsulation, and melanization activity. The expression levels of three phagocytosis-related genes, one encapsulation-promoting gene, and one melanization-regulating gene were significantly lower in the combined treatment group than those in the single fungal treatment group. Furthermore, pathogen recognition ability, signal transduction level, and immune effector expression level were weaker in the combined treatment group than those in the single fungal treatment group. The expression levels of 14 key metabolites and 7 key regulatory genes in glycolysis and tricarboxylic acid cycle pathways were significantly lower in the combined treatment group than those in the single fungal treatment group. Taken together, the weakness of innate immunity and energy metabolism in response to pathogen infection resulted in an increased susceptibility of H. cunea larvae to B. bassiana under Cd pre-exposure. Microbial insecticide is a preferred strategy for pest control in heavy metal-polluted areas. AVAILABILITY OF DATA AND MATERIAL: All the data that support the findings of this study are available in the manuscript.

Cadmium exposure through the food chain reduces the parasitic fitness of Chouioia cunea to Hyphantria cunea pupae: An ecotoxicological risk to pest control.

The toxicity of heavy metals on various trophic levels along the food chain has been extensively investigated, but no studies have focused on parasitic natural enemy insects. Herein, we constructed a food chain consisting of soil-Fraxinus mandshurica seedlings-Hyphantria cunea pupae-Chouioia cunea to analyze the effects of Cd exposure through food chain on the fitness of parasitic natural enemy insects and its corresponding mechanism. The results showed that the transfer of Cd between F. mandshurica leaves and H. cunea pupae and between H. cunea pupae and C. cunea was a bio-minimization effect. After parasitizing Cd-accumulated pupa, the number of offspring larvae, and the number, individual size (body weight, body length, abdomen length) and life span of offspring adults decreased significantly, while the duration of embryo development extended significantly. The contents of malondialdehyde and H2O2 in Cd-exposed offspring wasps increased significantly, accompanied by a significantly decrease in antioxidant capacity. The cellular immunity parameters significantly decreased in Cd-accumulated pupae, including the number of hemocytes, melanization activity and the expression level of cellular immunity genes (e.g. Hemolin-1 and PPO1). The humoral immunity disorder was found in the Cd-accumulated pupae, as evidenced by that the expression level of immune recognition gene (PGRP-SA), signal transduction genes (IMD, Dorsal, and Tube), as well as all antimicrobial peptide genes (e.g. Lysozym and Attacin) decreased significantly. Cd exposure decreased the content of glucose, trehalose, amino acid, and free fatty acid in H. cunea pupae. The expression of Hk2 in glycolysis pathway and the expression of Idh2, Idh3, Cs, and OGDH in TCA cycle pathway were significantly down-regulated in Cd-accumulated pupae. Taken together, exposure to Cd through the food chain causes oxidative damage on the offspring wasps and disrupts energy metabolism of the host insect, ultimately reducing the parasitic fitness of C. cunea to H. cunea pupae.

Transfer of Cd along the food chain: The susceptibility of Hyphantria cunea larvae to Beauveria bassiana under Cd stress.

Heavy metal can affect the bio-control efficiency of entomopathogenic fungi on pests, but this has not been studied in the food chain. Here, the food chain of soil-Fraxinus mandshurica-Hyphantria cunea was constructed to investigate the effect of cadmium (Cd) exposure on the susceptibility of H. cunea larvae to Beauveria bassiana (Bb) and to analyze the corresponding mechanism through larval innate immunity and energy metabolism. Cd through the food chain synergistically enhanced the susceptibility of H. cunea larvae to Bb. Cellular immunity-related parameters decreased when the Cd treatment group was compared with the control group and when the combined treatment group of Cd and Bb was compared with the Bb treatment group. Cd exposure induced hormesis on pathogen recognition and signal transduction genes of humoral immunity, but reduced the expression of effector genes. The expression of the 13 humoral immunity-related genes in the combined treatment group was lower than in the Bb treatment group. Cd exposure decreased the energy storage of H. cunea larvae before Bb infection and aggravated the disorder level of energy metabolism after Bb infection. Taken together, disturbance of innate immunity and energy metabolism improves the susceptibility of H. cunea larvae to Bb in the Cd-polluted food chain.

Cadmium exposure-triggered growth retardation in Hyphantria cunea larvae involves disturbances in food utilization and energy metabolism.

Serious environmental pollution in the ecosystem makes phytophagous insects face a great risk of exposure to pollutants, especially heavy metals. This study aims to understand the effects of Cd exposure on the growth and development of Hyphantria cunea and to elucidate the mechanism of growth toxicity induced by Cd from the perspective of food utilization and energy metabolism. Our results showed that the larval basal growth data, growth index, fitness index, and standard growth index were significantly decreased after feeding on Cd-containing artificial diets. The Cd-treated larvae had significantly higher digestibility than the untreated larvae. However, the food consumption, efficiency of conversion of digested food, and efficiency of conversion of ingested food were significantly lower than those of untreated larvae. Eight key metabolites in the glycolysis pathway and six key metabolites in the tricarboxylic acid cycle pathway were significantly reduced in Cd-treated larvae. The mRNA expression levels of two regulatory genes (6-phosphofructokinase 1 and hexokinase-1) belonging to two key enzymes in the glycolysis pathway and four regulatory genes (isocitrate dehydrogenase-1, isocitrate dehydrogenase-3, citrate synthase, and oxoglutarate dehydrogenase) belonging to three key enzymes in the tricarboxylic acid cycle pathway were significantly lower in the Cd-treated group than in the control group. Furthermore, most fitness-related traits were significantly and positively correlated with food utilization (except approximate digestibility) or energy metabolism parameters. Taken together, Cd exposure-triggered growth retardation of H. cunea larvae is a consequence of disturbances in food utilization and energy metabolism, thereby emphasizing the toxicity of heavy metals.

The susceptibility of Hyphantria cunea larvae to microbial pesticides Bacillus thuringiensis and Mamestra brassicae nuclear polyhedrosis virus under Cd stress.

Heavy metal pollution is an abiotic factor that can affect the efficiency of pest control. In this study, two microbial pesticides, Bacillus thuringiensis and Mamestra brassicae nuclear polyhedrosis virus (MbNPV), were used to treat Hyphantria cunea larvae with Cd pre-exposure, and the humoral and cellular immunity of H. cunea larvae with Cd exposure were evaluated. The results showed that Cd exposure increased the susceptibility of H. cunea larvae to microbial pesticides B. thuringiensis and MbNPV, and the lethal effect of Cd exposure and microbial pesticides on H. cunea larvae was synergistic. Cd exposure significantly decreased the expression of pathogen recognition genes (GNBP1 and GNBP3), signal transduction genes (Relish, Myd88, Tube, and Imd), and antimicrobial peptide gene (Lebocin) in the humoral immunity of H. cunea larvae compared with the untreated larvae. Parameters of cellular immunity, including the number of hemocytes, phagocytic activity, melanization activity, encapsulation activity, and the expression of three phagocytic regulatory genes (HEM1, GALE1, GALE2), were also found to decrease significantly in Cd-treated larvae. TOPSIS analysis showed that humoral immunity, cellular immunity, and total immunity levels of H. cunea larvae with Cd exposure were weaker than those in untreated larvae. Correlation analysis showed that the mortality of two microbial pesticides investigated in H. cunea larvae was negatively correlated with the humoral and cellular immunity of larvae. Taken togther, Cd exposure results in immunotoxic effects on H. cunea larvae and the use of microbial pesticides are an effective strategy for pest control in heavy metal-polluted areas.

Sex Pheromone Receptor-Derived Peptide Biosensor for Efficient Monitoring of the Cotton Bollworm Helicoverpa armigera.

The cotton bollworm, Helicoverpa armigera (H. armigera), causes damage to a wide range of cultivated crops and is one of the pests with the greatest economic importance for global agriculture. Currently, the detection of H. armigera is based on manual sampling. A low limit of detection (LOD), convenient, and real-time monitoring method is urgently needed for its early warning and efficient management. Here, we characterized the amino acid sequence in the sex pheromone receptors (SPRs) recognizing the pheromone components of H. armigera by three-dimensional (3D) modeling and molecular docking. Next, sex pheromone receptor-derived peptides (SPRPs) were synthesized and conjugated to nanotubes by chemical connection. The modified nanotubes were used to fabricate a sensor capable of real-time monitoring of gaseous sex pheromone compounds with a low LOD (∼10 ppb for Z11-16:Ald) and selectivity, and the sensor was able to detect a single live H. armigera. Furthermore, the developed biosensor allowed direct monitoring of the pheromone release dynamics by female H. armigera and showed that the release was instantly reduced in response to light. Here, we report the first demonstration of a biosensing method for detecting gaseous sex pheromones and live H. armigera. The findings show the great potential of the SPRP sensor for broad applications in insect biology study and infestation monitoring.

Defense response of Fraxinus mandshurica seedlings to Hyphantria cunea larvae under Cd stress: A contradiction between attraction and resistance.

Heavy metal pollution, as a common and serious environmental problem worldwide, has been regarded as an abiotic stimulus that can affect plant insect resistance and pest occurrence. This study evaluated the defense response of Fraxinus mandshurica seedlings to Hyphantria cunea larvae under Cd stress, with consideration given to chemical defense, physical defense, and elemental defense. Our results showed that the H. cunea larvae had a strong preference for Cd-treated F. mandshurica seedlings, but there was a significant reduction in body weight and survival rate in larvae that fed on leaves of Cd-treated seedlings. Under Cd treatment, the increase in attractant metabolites (e.g., styrene, dibutyl phthalate, and d-limonene) and the decrease in repellent metabolites (e.g., aromadendrene, heptadecane, and camphene) in leaf volatiles were responsible for the high attractant activity to H. cunea larvae. Based on leaf physicochemical properties, tissue structure, and phenolic acid content, an overall reduction in physical defense, chemical defense and their combination in F. mandshurica seedlings exposed to Cd stress was identified by Technique for Order of Preference by Similarity to Ideal Solution (TOPSIS) analysis. Elemental defense occurred in Cd-treated F. mandshurica seedlings, as evidenced by the high concentration of Cd in leaves and H. cunea larvae under Cd treatment. Taken together, these findings demonstrate that under Cd stress, elemental defense replaces the dominant role of basic defense in F. mandshurica seedlings and accounts for the enhanced ability to defend against H. cunea larvae.

Ultrasensitive Flexible Olfactory Receptor-Derived Peptide Sensor for Trimethylamine Detection by the Bending Connection Method.

Trimethylamine (TMA) is a harmful gas that exists ubiquitously in the environment; therefore, the sensitive and specific monitoring of TMA is necessary. In this work, we prepared ultrasensitive flexible sensors for TMA detection based on single-walled carbon nanotubes (SWCNTs) and olfactory receptor-derived peptides (ORPs) on low-cost plastic substrates. A novel bending connection method was developed by intentionally bending the interdigitated electrodes with SWCNTs to form a three-dimensional structure during the ORP-connection process, leading to the exposure of more modification sites. The method showed ∼4.7-fold more effective connection amount of the ORPs to SWCNTs compared to the conventional flat-condition connection method. The flexible ORP-SWCNT sensors could significantly improve the limit of detection for gaseous TMA from the reported lowest limit of 10 parts per quadrillion (ppq) to 0.1 ppq. The flexible ORP sensors also exhibited excellent sensitivity to vaporized TMA standards and TMA generated by different kinds of foods under different bending conditions. The results showed that the bending connection method in this work was effective for ultrasensitive flexible ORP sensors and their associated applications.

Effects of Arbuscular Mycorrhizal Fungi-Colonized Populus alba × P. berolinensis Seedlings on the Microbial and Metabolic Status of Gypsy Moth Larvae.

Arbuscular mycorrhizal fungi (AMF) are considered as important biological factors that can affect insect resistance of plants. Herein, we used AMF-poplar seedlings that could either increase or decrease the resistance to gypsy moth larvae, to elucidate the mechanism of mycorrhizal-induced insect resistance/susceptibility at the larval microbial and metabolic levels. Our results found that larval plant consumption and growth were significantly inhibited in the Glomus mossae (GM)-colonized seedlings, whereas they were enhanced in the Glomus intraradices (GI)-colonized seedlings. GM inoculation reduced the beneficial bacteria abundance in the larval gut and inhibited the detoxification and metabolic functions of gut microbiota. However, GI inoculation improved the larval gut environment by decreasing the pathogenic bacteria and activating specific metabolic pathways. Furthermore, GM inoculation triggers a metabolic disorder in the larval fat body, accompanied by the suppression of detoxification and energy production pathways. The levels of differentially accumulated metabolites related to amino acid synthesis and metabolism and exogenous toxin metabolism pathways were significantly increased in the GI group. Taken together, the disadaptation of gypsy moth larvae to leaves of GM-colonized seedlings led to the GM-induced insect resistance in poplar, and to the GI-induced insect susceptibility involved in the improvement of larval gut environment and fat body energy metabolism.

Protocols for CRISPR/Cas9 Mutagenesis of the Oriental Fruit Fly Bactrocera dorsalis.

The Oriental fruit fly, Bactrocera dorsalis, is a highly invasive and adaptive pest species that causes damage to citrus and over 150 other fruit crops worldwide. Since adult fruit flies have great flight capacity and females lay their eggs under the skins of fruit, insecticides requiring direct contact with the pest usually perform poorly in the field. With the development of molecular biological tools and high-throughput sequencing technology, many scientists are attempting to develop environmentally friendly pest management strategies. These include RNAi or gene editing-based pesticides that downregulate or silence genes (molecular targets), such as olfactory genes involved in searching behavior, in various insect pests. To adapt these strategies for Oriental fruit fly control, effective methods for functional gene research are needed. Genes with critical functions in the survival and reproduction of B. dorsalis serve as good molecular targets for gene knockdown and/or silencing. The CRISPR/Cas9 system is a reliable technique used for gene editing, especially in insects. This paper presents a systematic method for CRISPR/Cas9 mutagenesis of B. dorsalis, including the design and synthesis of guide RNAs, collecting embryos, embryo injection, insect rearing, and mutant screening. These protocols will serve as a useful guide for generating mutant flies for researchers interested in functional gene studies in B. dorsalis.