BACKGROUND:Brassica oleracea includes many highly nutritious and widely consumed vegetable crops, yet it is severely threatened by Plutella xylostella, a notorious pest in agriculture. The insect-resistant traits developed by host plants over hundreds of millions of years provide one of the most promising methods for pest control, which is why we have screened germplasm for resistance to P. xylostella. Among 50 B. oleracea germplasm resources, we identified a key variety, DM187, that caused 53% mortality in larvae and inhibited the development and reproduction of this moth species. RESULTS:In this study, we collected and cultivated 50 B. oleracea germplasm resources and evaluated their resistance to P. xylostella. Among these germplasm resources, larvae feeding on DM187 caused the least leaf damage and exhibited the highest mortality rate. Moreover, DM187 significantly inhibited the larval length, larval weight, pupation rate, pupal weight, and adult egg production of surviving P. xylostella. Furthermore, ovarian anatomy showed that DM187 resulted in incomplete ovarian development and a reduced egg count in adult females. CONCLUSION:Our findings indicate that the B. oleracea germplasm DM187 significantly impairs the survival, development, and reproductive capacity of P. xylostella. These findings provide a theoretical basis for the identification and utilization of natural insect-resistant resources in vegetables, as well as for research on insect-resistant breeding, and offer a strategy for the effective management of pests in agricultural settings. © 2026 Society of Chemical Industry.

BACKGROUND:Unraveling the compensatory mechanisms that mitigate fitness costs associated with resistance to Bacillus thuringiensis (Bt) pesticidal proteins is essential for delaying the evolution of Bt resistance. The diamondback moth Plutella xylostella, the first insect reported to evolve field resistance to Bt biopesticides, serves as an ideal model for elucidating the compensatory mechanisms underlying Bt resistance, as its evolution of Bt Cry1Ac resistance is not always accompanied by significant fitness costs. In this study, we identified a novel juvenile hormone esterase (JHE) gene (PxJHE2) and explored its role in compensating for the fitness costs associated with Bt Cry1Ac resistance in P. xylostella. RESULTS:We found a noncanonical JHE gene through sequence alignment and phylogenetic analysis. Metabolic analysis based on ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) demonstrated that PxJHE2 possesses the ability to degrade juvenile hormone (JH) into JH acid. We cloned PxJHE2 gene, and found its expression was significantly reduced in the midgut of both Bt Cry1Ac-treated and Bt-resistant strains. Furthermore, small interfering RNA (siRNA)-mediated silencing of PxJHE2 gene significantly increased JH titers, accompanied by significant fitness costs such as reduced fecundity, decreased pupation rate, and lowered eclosion rate. CONCLUSION:Our findings identify PxJHE2 as a key regulator of JH homeostasis that mediates compensation of fitness costs in Bt Cry1Ac-resistant P. xylostella. Targeting PxJHE2 or its hormonal regulatory pathway may provide a promising strategy to disrupt resistance-associated compensatory mechanisms, thereby enhancing the sustainability of Bt-based pest management programs. These results further provide valuable molecular insights for the development of innovative bioinsecticide strategies. © 2026 Society of Chemical Industry.

RNAi-based biopesticides represent a transformative approach to sustainable pest control, offering enhanced specificity and environmental safety compared to conventional synthetic pesticides. This technology utilizes doublestranded RNA (dsRNA) to silence critical genes in target pests through a naturally occurring biological pathway. Despite its promise, the commercialization of RNAi-based biopesticides faces significant challenges, including the absence of standardized ecological risk assessment protocols, regulatory uncertainties, public skepticism, and ethical considerations. This review critically examines the development, mechanisms, and applications of RNAi-based biopesticides and their potential to reshape future pest management strategies. We analyze the molecular basis of RNAi-mediated gene silencing, recent advances in dsRNA delivery systems, and key environmental and regulatory hurdles. Furthermore, we propose a comprehensive ecological risk assessment framework to address concerns regarding non-target effects and environmental persistence. Strategies to facilitate commercialization including regulatory harmonization, stakeholder engagement, and integration into integrated pest management are also discussed. Critical research gaps are identified, highlighting the need for long-term ecological monitoring, resistance management protocols, and cost-effective production and delivery methods. Successful implementation will require coordinated collaboration among researchers, regulators, industry stakeholders, and the public to ensure the responsible integration of RNAi-based biopesticides into global agriculture.

The steroid hormone 20-hydroxyecdysone (20E) is the central regulator of insect molting, yet the upstream transcription factors controlling its pathway remain poorly characterized. Here, we investigate the role of the bHLH-PAS transcription factor Trachealess (HvTrh) in the 28-spotted ladybeetle, Henosepilachna vigintioctopunctata. HvTrh was ubiquitously expressed during larval development, with peak transcript levels in the pupal stage. RNAi-mediated silencing of HvTrh induced severe molting defects and high mortality across all larval instars. This was associated with a significant reduction in 20E titers and the suppression of key 20E pathway genes. The resulting molting defects were not rescued by exogenous 20E, indicating a disruption of tissue competence. Histological analysis confirmed that HvTrh knockdown impaired chitin deposition and new cuticle remodeling. Among the downregulated genes, HvE75 was the most strongly suppressed. Mechanistically, we demonstrated that HvTrh directly activates the HvE75 promoter by binding to a specific central nervous system midline element (CME), as validated by dual-luciferase reporter assays and electrophoretic mobility shift assays. Our findings establish HvTrh as a critical upstream regulator of insect molting, proposing a model where it governs the 20E signaling cascade, at least in part, through the direct transcriptional control of HvE75.

To identify potential strategies for increasing the efficiency of tomato leaf metabolism with a focus on the links between nitrogen/carbon metabolism, we explored a diel flux balance analysis (FBA) model of a source leaf in which the metabolic output was varied up to the theoretically achievable maximum. We noticed a potentially interesting switch in the use of glutamine synthetase (GS) isoforms-from the chloroplast isoform to the mitochondrial one-for nitrogen assimilation. To further explore this prediction, we characterized transgenic tomato plants as overexpressing 2 tomato GS genes, GS1 and GS2, targeted to mitochondria. Both sets of transgenic plants were characterized as displaying faster growth rate, early flowering, and increased fruit yield. In leaves, metabolomic profiling and enzyme activity analysis pointed out that GS activity in mitochondria plays a role in increasing the intracellular synthesis and subsequent export of sugar. Consistent with these changes, higher sucrose concentration in leaf exudates and reduced activities of enzymes involved in leaf starch synthesis were observed. Moreover, mitochondrial GS activity affected chloroplast redox status in a manner that modulated photorespiration and nitrogen metabolism. The combined data reveal the influence of mitochondrial GS activity on both foliar carbon/nitrogen balance and regulation of source-sink metabolism in tomato plants.

The widespread use of neonicotinoid insecticides has resulted in extensive resistance in field populations of the whitefly Bemisia tabaci, posing a major challenge to sustainable pest management. However, the molecular basis of cross-resistance to multiple neonicotinoids remains to be fully elucidated. In this study, resistance levels to thiamethoxam, thiacloprid, and nitenpyram were evaluated in eleven field populations of B. tabaci MED collected over a two-year period from vegetable fields in multiple provinces of China. Comparative transcriptomic analysis identified CYP402C5 as a candidate cytochrome P450 gene consistently overexpressed in resistant populations. RNA interference-mediated silencing of CYP402C5 significantly increased the susceptibility of resistant populations to all three neonicotinoids but had little effect on the susceptible strain, indicating its important role in neonicotinoid resistance. Structural prediction and molecular docking suggested that CYP402C5 possesses a conserved P450 catalytic pocket capable of accommodating multiple neonicotinoids. Functional characterization using a baculovirus expression system, coupled with UPLC-MS/MS analysis, demonstrated NADPH-dependent metabolism of thiamethoxam, thiacloprid, and nitenpyram by recombinant CYP402C5. Moreover, the identification of a nitenpyram-derived metabolite (NIT-IM) provided direct evidence for CYP402C5-mediated biotransformation. Collectively, these results demonstrate that CYP402C5 functions as a neonicotinoid-detoxifying P450 enzyme and contributes to cross-resistance in B. tabaci. This study expands the repertoire of resistance-associated P450 genes in B. tabaci and provides valuable insights for resistance monitoring and neonicotinoid resistance management.

RNA interference (RNAi), a highly conserved gene silencing mechanism triggered by sequence-specific doublestranded RNA (dsRNA), holds great promise for targeted pest control. The bean flower thrips, Megalurothrips usitatus, a devastating pest of cowpeas, has developed widespread resistance to conventional insecticides, necessitating alternative management strategies. While RNAi-based approaches offer an eco-friendly solution, our preliminary studies revealed remarkably low RNAi efficiency in this species. To investigate this limitation, we employed an artificial membrane feeding system and discovered that thrips salivary secretions rapidly degrade dsRNA, suggesting a potential mechanism for reduced RNAi efficacy. Further exploration identified five dsRNase genes (MudsRNase1-5) in M. usitatus, with high expression levels in both midgut and salivary glands. Exposure to exogenous dsGFP triggered upregulation of all five MudsRNase genes in the 2nd instar nymphs and specifically induced MudsRNase3 in adults, implicating their role in dsRNA catabolism. Functional analyses demonstrated that RNAi-mediated knockdown of MudsRNase1, MudsRNase3 and MudsRNase4 substantially enhanced RNAi efficiency in the 2nd instar nymphs. Notably, recombinant MudsRNase3 protein exhibited robust dsRNA-degrading activity in vitro, directly linking this enzyme to dsRNA clearance. These findings reveal that dsRNasemediated degradation represents a critical barrier to RNAi efficacy in M. usitatus. By characterizing the molecular basis of dsRNA instability in this pest, we identified MudsRNases as promising targets for RNAi optimization, and we established a framework for developing enhanced RNAi-based management strategies targeting thrips.

The whitefly, Bemisia tabaci, as a globally significant agricultural pest, poses a serious threat to various food crops and thus causes enormous economic losses to global agricultural production. In recent decades, with the long-term and extensive use of chemical pesticides, B. tabaci has developed high levels resistance to thiamethoxam. Therefore, an in-depth analysis of the molecular mechanisms involved in insecticide detoxification within B. tabaci is of great theoretical and practical significance. Neuropeptide FF receptor 2 (NPFF2), a G protein-coupled receptor (GPCR), plays a key role in the neonicotinoid resistance of B. tabaci. Whether NPFF2 is involved in the resistance to thiamethoxam remains unclear. In this study, we cloned the full-length NPFF2 gene of B. tabaci, and further confirmed its vital role in thiamethoxam resistance via Quantitative real-time PCR and RNA interference assays. To investigate the signaling pathway of NPFF2 involved in thiamethoxam resistance in B. tabaci. The expression of six published detoxification-related genes (UGT354A1, UGT352A3, CYP6CX1, CYP6CX2, CYP6CX3 and CYP6CX4) were determined in the resistant and susceptible strains. The results showed that the expression of CYP6CX4 was over-expressed in resistant, and decreased by knockdown of NPFF2. Additionally, in a metabolism assay in vitro, the CYP6CX4 protein could metabolize 52.64% of thiamethoxam with a clothianidin-guanidine hydrochloride metabolite via UPLC-QTOF/MS. Overall, our study reveals a novel NPFF2 - CYP6CX4 signaling pathway underlying thiamethoxam resistance in B.tabaci, providing a theoretical basis for its resistance monitoring and novel control target development.

BACKGROUND:Viruses and their insect vectors have evolved intricate co-evolutionary mechanisms to enhance transmission efficiency. This study aimed to investigate the impact of tomato spotted wilt virus (TSWV) infection on the development of Frankliniella occidentalis, with particular emphasis on the potential involvement of Halloween genes. RESULTS:Genomic analysis identified seven Halloween genes. Transcriptome data from revealed significant upregulation of FoCYP307A1 and FoCYP306A1 during the nymphal and pupal stages in TSWV-infected individuals. These findings were validated by quantitative real-time polymerase chain reaction, which demonstrated a 1.5- to 3-fold increase in transcript levels of these genes during the nymphal, propupal, and pupal developmental stages. TSWV infection significantly shortened the nymphal and pupal developmental durations by 1 day and 0.6 days, respectively. RNA interference (RNAi)-mediated silencing of FoCYP307A1 and FoCYP306A1 significantly prolonged the nymphal and pupal development time, accompanied by a 27.5% and 55% reduction in 20-hydroxyecdysone (20E) levels, respectively. In addition, the transcript levels of 20E key signaling pathway genes, including FoUSP, FoBr-C, FoDHR3, FoE74A, and FoE75, decreased by 16-67%. Importantly, supplementation with 20E effectively reversed the developmental delays caused by RNAi-induced silencing of FoCYP307A1 and FoCYP306A1. Collectively, these results suggest that FoCYP307A1 and FoCYP306A1 play a critical role in mediating TSWV-facilitated development in thrips. CONCLUSION:These findings provide valuable insights into how TSWV exploits ecdysone biosynthesis to accelerate development in insect vectors, offering important implications for integrated pest and disease management strategies. © 2026 Society of Chemical Industry.

Plant metabolism is increasingly being demonstrated to be partially controlled by dynamically assembled metabolons-multienzyme complexes that enable substrate channeling, insulate reactive intermediates, and permit rapid, low-energy flux control. Rigorous criteria are defined to distinguish true metabolons from generic assemblies, and evidence is synthesized across cyanogenic glucoside, phenylpropanoid/flavonoid, alkaloid, terpenoid, polyamine, sporopollenin, and auxin pathways. A practical workflow is presented in which AP-MS (Affinity purification mass spectrometry)/Co-IP (Co-immunoprecipitation), proximity labeling, BiFC (Bimolecular fluorescence complementation)/FRET (Förster resonance energy transfer)/Split-luciferase, and isotope-dilution metabolomics are integrated to resolve composition, dynamics, and direct channeling in vivo. In enzyme-based substrate channeling engineering, design rules are distilled for membrane anchoring, modular scaffolds, compartment targeting, and inducible/optogenetic control, and limitations such as metabolic burden, stoichiometry, and leakiness are noted. An AI-assisted loop is outlined in which structure-aware generative models produce binders/interfaces that are coupled to spatial optimization of enzyme order, orientation, and distance. Together, these advances reposition metabolons as a deployable technology for programmable flux in plants, enabling safer handling of labile intermediates and higher titers of valuable natural products.

Plant growth and productivity are strongly constrained by herbivorous insects, which reduce both yield and quality. Over the past two centuries, extensive efforts have been devoted to identifying natural insect-resistant traits and genes that have evolved in plants. Here, we first review key milestones and advances, including empirical observation, mechanism onset, theoretical emergence, gene identification, and breeding application. Then, we categorize research on plant defenses for insect resistance into four major types: physical, chemical, ecological modulation, and behavioral defenses, and integrate them to highlight the coordination and synergy among different defenses. Finally, we propose priority areas and future directions for research to advance studies on plant-insect interactions and support effective and sustainable pest management.

BACKGROUND:Conventional lambda-cyhalothrin formulations are limited by rapid photodegradation, short field persistence, high aquatic toxicity, and insufficient active ingredient loading for large-scale agricultural applications. Herein, a high-loading (23%) lambda-cyhalothrin microcapsule suspension (CS) with an organic-inorganic hybrid shell was developed by integrating interfacial polymerization with a sol-gel process using isophorone diisocyanate (IPDI) and tetraethyl orthosilicate (TEOS). This hybrid strategy was designed to simultaneously enhance sustained release, photostability, insecticidal efficacy and environmental safety. RESULTS:The optimized formulation produced spherical microcapsules with an average particle size of 0.832 μm, D50 = 0.762 μm, D90 = 1.204 μm, an encapsulation efficiency of 76.46% and a suspension rate of 95.26%. The high-performance liquid chromatography (HPLC) method exhibited excellent linearity (R2 = 0.99977). The microcapsules showed controlled release without burst release, reaching a cumulative release of 78.3% after 7 days, compared with 92.3% for the emulsion in water (EW) and almost complete release of the technical material within 24 h. After 4 h of UV irradiation (365 nm), the degradation rate of the microcapsules was only 52.25%, markedly lower than that of the EW (70.79%) and technical material (96.84%). Against 3rd-instar Hyblaea puera larvae, the CS achieved 69.52% corrected mortality at 0.5 mg/L after 12 h and 100% mortality across all tested concentrations within 36 h. Furthermore, the 96 h median lethal concentration toward juvenile crucian carp was 2.17-fold higher than that of the EW, indicating reduced acute aquatic toxicity. CONCLUSION:The organic-inorganic hybrid microcapsule integrates high active ingredient loading with sustained release, enhanced UV resistance, superior insecticidal performance, and improved ecological compatibility. This study provides a promising formulation strategy for stabilizing photolabile pyrethroids and offers a practical platform for developing next-generation controlled-release pesticide formulations suitable for sustainable crop protection. © 2026 Society of Chemical Industry.

The tomato leafminer, Tuta absoluta, poses a severe threat to tomato production in China, yet regional resistance patterns remain poorly characterized. This study investigates the insecticide resistance status and underlying molecular mechanisms in field populations of T. absoluta across major tomato-producing regions in China. Ten populations were evaluated for susceptibility to seven commonly used insecticides using standardized bioassays, alongside screening for resistance-associated target-site mutations in key genes. The results indicate that most populations remain susceptible to spinetoram, abamectin, and indoxacarb, supporting their continued effectiveness in field control. However, moderate resistance to emamectin benzoate was detected in several populations, with resistance ratios reaching up to 75.09 fold. Additionally, low to moderate resistance to Bacillus thuringiensis (Bt) and broflanilide was observed in certain regions. Although resistance ratios to bifenthrin were relatively low, consistently high LC50 values suggest a widespread reduction in baseline susceptibility. Molecular analyses revealed that the L1014F mutation in the voltage-gated sodium channel (VGSC) gene was fixed across all populations, while M918T and T929I mutations occurred at high frequencies. Notably, no significant correlation was found between these mutations and phenotypic resistance levels, and no resistance-associated mutations were detected in the nAChR gene. These findings highlight the complex and potentially multifactorial nature of resistance in Chinese T. absoluta populations and provide important guidance for optimizing insecticide rotation and resistance management strategies.

Abstract Horizontal gene transfer (HGT) has enabled insects to acquire novel genetic material that can fuel adaptation to environmental change. However, the role of HGT in the evolution of insecticide resistance remains poorly characterised. Here, we identify BtUCH19 , a fungal gene that has integrated into the genome of the global pest Bemisia tabaci and functions as a deubiquitinating enzyme (DUB). We show that compared to endogenous DUB, BtUCH19 specifically removes K63-linked ubiquitin chains from a cytochrome P450, CYP4C64, thereby stabilizing this key detoxification enzyme in vivo . Sustained CYP4C64 abundance enhances metabolic detoxification of two commonly used insecticides thiamethoxam (TMX) and clothianidin (CLO) to their low-toxicity products TMX-Urea and CLO-Urea. Consequently, BtUCH19 drives insecticide resistance through a novel "HGT-PTM-Metab" axis. Our work reveals the instrumental role of a HGT in orchestrating ubiquitin-proteasome system (UPS)-mediated protein level regulation of a key detoxification pathway. These findings offer new insights into the intricate mechanisms underlying co-evolutionary biology, and provide a new molecular target for Integrated Pest Management (IPM).

Megalurothrips usitatus (bean flower thrips) has transitioned from a tropical Asian pest into a global threat to legumes, particularly cowpea (Vigna unguiculata). Driven by climate warming and trade, its recent expansion across the Americas causes 20%-30% yield losses and sometimes leads to crop failure. Heavy reliance on chemical control has rapidly selected for multi-class insecticide resistance. This review synthesizes current research on the pest's biological traits, invasion ecology, cryptic diversity, and the molecular mechanisms driving resistance to spinosyns, neonicotinoids, and pyrethroids. Our review also evaluates integrated pest management strategies, ranging from cultural controls to RNAi biotechnologies, while addressing the critical challenges of laboratory-to-field translation. Finally, we propose key research priorities, including intelligent monitoring, localized resistance management, and synergistic control packages, to provide a sustainable framework for global legume protection and biosecurity policy.

Protein-Protein interactions (PPIs) wire plant cells, assembling metabolons, routing signals, and coordinating organelle crosstalk. We review experimental platforms and the computational signals long used to predict PPIs. While experimental platforms and traditional computational approaches have long been employed for PPIs prediction, recent advances in artificial intelligence offer unprecedented opportunities to map plant interactomes comprehensively. To provide a systematic overview, we categorize current methodologies into four thematic families: (i) sequence-centric predictors utilizing protein language models to extract evolutionary features; (ii) structure-based predictors integrating coevolutionary signals to reconstruct 3D complex arrangements; (iii) network-level learners employing graph architectures to capture global interactome topology; and (iv) geometric and generative methods leveraging symmetry-aware networks for specific site identification and de novo design. Despite rapid gains, plant applications are constrained by paralog expansion, compartmentalization, dynamic microenvironments, and the sparse availability of gold standards in the field. Next-generation plant AI PPI models should be organelle-aware, multimodal, rigorously benchmarked, structure-gated, and condition-validated.

Reducing dependence on synthetic nitrogen fertilizer requires biologically grounded alternatives. Symbiotic nitrogen fixation supplies fixed nitrogen but is restricted to a narrow angiosperm clade, limiting direct deployment in most major nonleguminous crops. We synthesize how telomere-to-telomere genomes and pangenomes expose structural and regulatory variants for nodulation; how single-cell and spatial transcriptomics resolve stage-specific cell states and division of labor; and how epigenomic and 3D genome maps reveal principles of regulatory control for infection, organogenesis, and fixation. Extending to actinorhizal symbioses tests single- versus multiple-origin models. We present an artificial intelligence-guided roadmap that integrates sequence, chromatin accessibility, and expression data to prioritize regulatory elements, propose compact edit sets, and guide cell type-specific deployment in nonleguminous crops, advancing from descriptive catalogs to testable models and iterative validation.

Abstract N6-methyladenosine (m 6 A) modification is the most predominant and ubiquitous internal modification of RNA in eukaryotes, serving as a key post-transcriptional regulator of gene expression that is dynamically modulated by methyltransferases (writers) and demethylases (erasers). However, while the functions of m 6 A methylases have been partially elucidated in insects, the identity of m 6 A erasers in arthropods and their chemical catalytic mechanisms, as well as biological functions, remains largely enigmatic. Here, we uncovered 2499 putative methylase genes and 1148 putative demethylase genes in 266 insect genomes, and demonstrated that ALKBH4 functions as an m 6 A demethylase in the whitefly, Bemisia tabaci , catalyzing the oxidative reversal of mRNA m 6 A modifications both in vitro and in vivo. Furthermore, we established that ALKBH4, in coordination with other core components of the m 6 A pathway, fulfills an essential function in regulating the transcript stability of Imaginal Disk Growth Factor 1 (IDGF1) during whitefly development. Collectively, our findings expand the evolutionary scope of the eukaryotic m 6 A modification system, and reveal a conserved yet insect-specific epitranscriptomic regulatory mechanism governing fundamental physiological processes and adaptive phenotypes. Significance statement The addition of a methyl group to the N6-position of adenosine (m 6 A) is a highly abundant chemical modification of RNA. However, the functional role of m 6 A in insects and the key enzymes that regulate its levels remains poorly understood. In this study, we explored putative methylase genes and demethylase genes in hundreds insect genomes, and identified an m 6 A RNA demethylase, ALKBH4, in the whitefly, Bemisia tabaci . We demonstrate that ALKBH4 oxidatively reverses mRNA methylation in vivo and in vitro, in combination with other components of the m 6 A pathway, plays an important role in whitefly development. These findings provide new insight into m 6 A methylation system of insect.

Foliar application of nanocoated inoculates encapsulating Klebsiella variicola W12 enhanced nitrogen fixation under nitrogen-depleted conditions, representing a sustainable strategy to increase crop productivity.