The entomopathogenic fungi Beauveria bassiana and Metarhizium robertsii are natural pathogens of the Colorado potato beetle (CPB) and hold promise for biocontrol. However, the insect's immune responses to fungal infections remain poorly understood. In this study, we performed transcriptomic analyses of haemocytes and fat body tissue from CPB larvae topically infected with B. bassiana and M. robertsii at the stage of pathogen encapsulation. Quantitative PCR validation of gene expression, along with complementary physiological and biochemical assays, was also conducted. Reference gene annotations were expanded using both the classical bioinformatic tool InterProScan and the AI-based tool ProteInfer. For the first time, pathogen-specific differences in CPB tissue responses to fungal infections were revealed. Gene Ontology enrichment and expression profiles of immune-related genes indicated a stronger antifungal response to M. robertsii, whereas infection with B. bassiana was associated with enhanced protection against self-damage. Pathogen-specific responses were linked to the expression of genes encoding certain receptors, serine proteases, serpins, Toll signalling components, antimicrobial peptides, chitin deacetylases, chitin synthases, laccases, as well as to the production of phenoloxidases and reactive oxygen species. The most diverse and pathogen-specific gene expression changes occurred in haemocytes rather than in the fat body. We propose that the observed differences in immune responses are mainly driven by the fungal enzymatic machinery, secondary metabolites and pigments. The differentially expressed genes identified in this study provide novel insights into insect-pathogen interactions and represent promising molecular targets for advancing both fundamental research on insect immunity and the applied development of innovative biocontrol strategies.

Behavior-modifying effects of methanolic seed extract of Garcinia kola (Clusiaceae) against Colorado potato beetle [Leptinotarsa decemlineata (Say)

Biocontrol potential of native Beauveria bassiana isolates from Türkiye against Leptinotarsa decemlineata (Say) (Coleoptera: Chrysomelidae)

The rice striped stem borer (Chilo suppressalis) and Colorado potato beetle (Leptinotarsa decemlineata) are notorious pests that have developed widespread resistance to conventional insecticides. RNA interference (RNAi) represents a breakthrough in pest management contingent on identifying effective target genes. This study isolated and characterized transcription factor E93 from both pests. Spatiotemporal expression profiles revealed a similar pattern: low levels in early instars, a progressive increase in later instars, and peak expression in adults, particularly within the ovaries. Functional analysis via CRISPR/Cas9 and RNAi across both species elicited analogous defects: abnormal pupation, aberrant eclosion, and impaired egg development. Furthermore, feeding on potato plants treated with dsLdE93 in pot trials significantly reduced the level of L. decemlineata pupation, emergence, and viable egg production. This study identifies E93, a transcription factor pivotal for development in both C. suppressalis and L. decemlineata, and validates it as a potential novel target for RNAi-mediated pest control.

Abstract BACKGROUND Foliar application of RNA interference (RNAi)‐based products offers a promising strategy for protecting crops against insect pests. This approach involves the direct spraying of exogenous double‐stranded RNA (dsRNA) to silence specific target genes in pests. However, its practical efficacy is constrained by the environmental degradation of the dsRNA, rapid degradation by gut nucleases, inefficient cellular internalization and limited endosomal escape. The integration of nanotechnology with RNAi has emerged as a promising frontier in sustainable pest control, offering improved dsRNA stability, enhanced cellular delivery, and the potential to reduce conventional pesticide use. In this study, we explored the use of a positively charged covalent organic framework (COF)‐based nanocarrier functionalized with polyethylenimine (PEI) and polyethylene glycol (PEG) for dsRNA delivery. RESULTS The resulting COF‐PEI‐PEG@dsRNA complexes effectively resisted degradation by nuclease‐rich midgut extracts. Moreover, COF‐PEI‐PEG facilitated efficient dsRNA delivery into Drosophila S2 and Lepidopteran Sf9 cells, resulting in significant improvement in target gene knockdown. In vivo , COF‐PEI‐PEG@dsRNA significantly enhanced RNAi efficacy in Leptinotarsa decemlineata and Drosophila suzukii , although no improvement was observed in Spodoptera exigua . CONCLUSION Collectively, our findings highlight the potential of COF‐PEI‐PEG as effective dsRNA delivery platforms, offering a novel and versatile tool to enhance RNAi‐based insect management and fundamental entomological research. © 2026 Society of Chemical Industry.

The damage caused by herbivores is generally measured as the amount of leaf tissue consumed, without accounting for the fate of the leftover tissue. As a result, the plant defense mechanisms that promote resistance to herbivore feeding by photosynthetically acclimating the rest of the plant to the feeding spot leaf area have not been well exploited. Plant-insect interactions are now becoming better defined with the development of visualization methods that permit spatial whole-leaf assessment of photosynthetic efficiency after herbivore attack. The purpose of our study was to evaluate the spatial heterogeneity of photosystem II (PSII) function at the whole-leaf level before and after herbivory by the Colorado potato beetles. Twenty minutes after Colorado potato beetle (Leptinotarsa decemlineata) feeding, the maximum efficiency of PSII photochemistry (Fv/Fm) decreased significantly, suggesting photoinhibition due to reduced efficiency of the oxygen-evolving complex (OEC). The decreased quantum yield of PSII photochemistry (ΦPSII) after feeding, at the neighboring area of the feeding spot and at the rest of the leaf area, was attributed to the reduced efficiency of the open PSII reaction centers (Fv'/Fm'), since there was no change in the fraction of open PSII reaction centers (qp). Nevertheless, plant defense elicitation was activated by the photoprotective mechanism of non-photochemical quenching (NPQ) that reduced the singlet oxygen (1O2) formation in potato plants in the neighboring area of the feeding spot and at the rest of the leaf area. In addition, the increased production of hydrogen peroxide (H2O2) triggered by this increase suggests that it acted as a signaling molecule in the biotic stress defense response.

Plants in the family Solanaceae produce steroidal glycoalkaloids (SGAs), specialized metabolites that are toxic to consumers while providing protection against herbivores. The Colorado potato beetle (CPB) is a major foliar pest of potatoes, and the wild species Solanum chacoense PI458310 displays resistance through the accumulation of the specific SGAs leptine I/II, the C-23-acetoxylated derivatives of α-chaconine/α-solanine. Although these leptines and their presumed C-23-hydroxylated precursors, leptinine I/II, have been recognized for decades, their biosynthetic routes have remained unresolved. In this study, we show that leptinines and leptines do not originate from direct modificfation of α-chaconine/α-solanine, but instead arise from earlier transformations of spirosolane-type precursors. We identified Sc23DOX, a spirosolane C-23 hydroxylase, and Sc23ACT, a 23-O-acetyltransferase, and demonstrated in vitro that these enzymes convert α/β-solamarine into 23-hydroxy and 23-O-acetyl intermediates, which are subsequently processed into leptinines and leptines by the ring-rearrangement enzymes Dioxygenase for Potato Solanidane synthesis (DPS), Reductase for Potato Glycoalkaloid biosynthesis 1 (RPG1), and RPG2. Notably, these C-23 hydroxylation and O-acetylation reactions parallel the initial steps of α-tomatine detoxification during fruit ripening in tomato (Solanum lycopersicum). Moreover, when Sc23DOX and Sc23ACT were expressed heterologously in cultivated potato, the plants produced leptinines and leptines de novo. Genomic and transcriptomic analyses further revealed that many cultivated potatoes (Solanum tuberosum) retain apparently functional 23DOX and 23ACT orthologues, and that expression of 23DOX alone can restore leptine biosynthesis in certain cultivars. Collectively, these findings clarify the leptinine/leptine biosynthetic pathway, illustrate the co-option of a detoxification-related module into a defense system, and provide a genetic basis for breeding or engineering CPB-resistant potatoes.

Vulnerability of potato crops to diseases and pest infestation can affect its quality and lead to significant yield losses. Timely detection of such diseases can help take effective decisions. For this purpose, a deep learning-based object detection framework is designed in this study to identify and classify major potato diseases and pests under real-world field conditions. A total of 2,688 field images were collected from two research farms in Punjab, Pakistan, across multiple growth stages in various seasonal conditions. Excluding 285 symptoms-free images from the earliest collection led to 2,403 images which were annotated into four biotic-stress classes: blight disease (n = 630), leaf spot disease (n = 370), leafroll virus (viral symptom complex; n = 888), and Colorado potato beetle (larvae/adults; n = 515), indicating class imbalance. Several state-of-the-art models were used including YOLOv8 variants (n/s/m), YOLOv7, YOLOv5, and Faster R-CNN, and the results are discussed in relation to recent potato disease classification studies involving cropped leaf images. Stratified splitting (70% training, 20% validation, 10% testing) was applied to preserve class distribution across all subsets. YOLOv8-medium achieve the best performance with mean average precision (mAP)@0.5 of 98% on the held-out test images. Results for stable 5-fold cross-validation show a mean mAP@0.5 of 97.8%, which offers a balance between accuracy and inference time. Model robustness was evaluated using 5-fold cross-validation and repeated training with different random seeds, showing a low variance of ±0.4% mAP. Results demonstrate promising outcomes under the real-world field conditions, while, broader cross-region and cross-season validation is intended for the future.

A major challenge in crop improvement is enhancing resistance to diverse biotic stresses. Because terpenoids play key roles in chemical defense, an envisioned strategy is to introduce new terpene metabolic pathways into crops through engineering. Microbial-type terpene synthase-like (MTPSL) genes are widespread in nonseed plants but absent in seed plants. Here, we engineered terpene metabolism in Nicotiana benthamiana using MTPSL genes, enabling production of sesquiterpenes absent in flowering plants and enhanced resistance to pest insects and fungal pathogens. Two liverwort MTPSL genes, RlMTPSL3 and RlMTPSL4, which produce sesquiterpenes absent from flowering plants, were selected for metabolic engineering. In N. benthamiana, both genes generated sesquiterpenes consistent with their in vitro activities, and co-expression yielded combined profiles. Co-expression of RlMTPSL3 and RlMTPSL4, individually or together, with 3-hydroxy-3-methylglutaryl-CoA reductase, the rate-limiting enzyme in sesquiterpene pathway, substantially increased sesquiterpene production. Bioassays of engineered tissues with two defoliating herbivores beet armyworm (Spodoptera exigua) and Colorado potato beetle (Leptinotarsa decemlineata) showed growth suppression and up to 30% mortality. The gut microbiome of beet armyworm feeding on engineered tissues showed differences from those feeding on control tissues, suggesting a potential mechanism underlying reduced pest insect performance. Engineered sesquiterpenes were recovered from larval frass, indicating stability through digestion. Transformed leaves emitted elevated sesquiterpenes as volatiles that repelled beet armyworm. In addition, extracts of engineered tissues inhibited the growth of Fusarium oxysporum, a fungal pathogen, by ∼50%. Together, these results demonstrate that MTPSL-based engineering can introduce new sesquiterpenes into flowering plants, providing a promising strategy for broad-spectrum crop protection.

Diapause in the Colorado potato beetle is characterized by sex-specific gene expression.

Population dynamics are controlled by life history and moderated by the environment. These factors determine the timing and magnitude of population abundance and are used to predict crop pest populations. Forecasts of pest populations typically focus on heat accumulation during the growing season when the pests are active or emerging from the soil if they have a diapause period. Weather conditions before and during diapause can impact population dynamics as well but tend to be understudied. This study used 16 years of Colorado Potato Beetle (Leptinotarsa decimlineata) abundance data combined with landscape and daily weather data to understand the importance of seasonal weather and landscape on predicting population abundance. Boosted-tree multiclass models predicted abundance at each life stage and across all models. Cumulative degree days, a measure of developmental time, were overwhelmingly important. To understand how the top non-temporal variables could interact with time, we used general additive mixed models to examine interactions between time and the top six ranked variables for each life stage, then generated predicted abundances across the growing season for the 10th, 50th, and 90th percentiles of each variable, keeping all other variables constant. This revealed that while Colorado Potato Beetle abundance was most strongly affected by heat accumulation, other weather factors like precipitation and air saturation, as well as soil temperature during diapause, can also influence abundance trends. The only landscape variable consistently ranked in the top six was potato acreage.

Dillapiol is a naturally occurring methylenedioxyphenyl compound with insecticide-synergizing activity comparable to piperonyl butoxide (PBO). This study identified structurally related molecules with practical potential for managing insecticide-resistant insects. Six new dillapiol analogs, containing ester- or ether-linked side chains, were synthesized and evaluated as pyrethrum synergists against the Colorado potato beetle (CPB) Leptinotarsa decemlineata (Say) (Coleoptera: Chrysomelidae). Their activity was assessed through bioassays and by quantifying inhibition of Phase I and II detoxification enzymes in vitro and in vivo. All six compounds displayed higher synergistic activity by ingestion than by topical exposure, and each structural class included at least one compound with a synergism ratio greater than 20. In the resistant CPB strain (RS-CPB), two ester compounds inhibited P450 monooxygenase activity in vitro as effectively as PBO, while dillapiol and one ether analog reduced P450 activity in vivo. Notably, all six analogs reduced glutathione S-transferase (GST) activity; the most active was an ether analog with an in vitro IC50 of 0.23 (±0.04) mM. Dillapiol also significantly reduced GST activity in vivo. These analogs demonstrated PBO-equivalent P450 inhibition combined with unique GST inhibition and show promise as alternative synergists for managing insecticide-resistant insects.

While potato (Solanum tuberosum L.) is the world's most important non-cereal food crop, its production faces increasing pressure from climate change, pests, and pathogens. Genetic diversity among its wild relatives in the Solanum section Petota has long contributed to improving cultivated lines. This mini review contributes by examining how genomic advances have transformed our ability to utilize this genetic diversity, from domestication to the first doubled monoploid reference genome and the recent developments in polyploid genome assembly and super-pangenome construction. Analyses of these resources reveal that wild species exhibit extensive allelic diversity for stress tolerance, with examples including frost resistance from S. commersonii and S. acaule Bitt., late blight resistance from S. demissum Lindl. and others, and Colorado potato beetle resistance from S. okadae Hawkes. We conclude that integrating genetic material from potato wild relatives with improved variety germplasm provides an essential pathway for developing climate-resilient potato varieties. Consistent and increasing pangenome characterization will be essential for identifying novel genes conferring pathogen resistance, stress tolerance, and agronomic traits, particularly given the complex polyploid nature and extensive phenotypic diversity of the potato.

Leptinotarsa decemlineata (Say) is among the most destructive insect pests of potato Solanum tuberosum L. worldwide. While conventional chemical tactics have repeatedly failed to control this pest due to the rapid development of resistance, elicitor-based enhancement of plant defenses using jasmonic acid (JA) or salicylic acid (SA) pathways offers a promising complementary approach. Hence, this study evaluated the effects of 3 commercially available plant defense elicitors: Blush 2X (JA type), Actigard 50WG (SA type), and Regalia (JA and SA type), on the growth, development, and performance of L. decemlineata larvae across 2 phenological stages of “Russet Burbank” potatoes, with and without Myzus persicae (Sulzer) co-infestation. Defoliation, larval development, weight, and excretion rates were assessed through a series of greenhouse bioassays. Blush 2X significantly reduced defoliation and delayed larval development, supporting the role of JA signaling in mediating antifeedant defenses. In contrast, high-rate Actigard 50WG increased defoliation and excretion while reducing larval weight, suggesting altered sugar allocation and a potential diuretic effect. Aphid presence induced signal interference, modifying elicitor efficacy in a plant stage-dependent manner. These findings highlight the potential of elicitor treatments to enhance integrated pest management strategies for L. decemlineata, while underscoring the importance of plant phenology, signal cross-talk, and ecological context in optimizing their deployment.

The paper focuses on the advancement of experimental and analytical methodologies for examining the mechanical properties of living larvae of Leptinotarsa decemlineata (Colo-rado potato beetle) to facilitate their eradication through acoustic resonance. The signifi-cance of this research is underscored by the necessity to substitute chemical pest control methods, which are detrimental to human health and the environment, with ecological acous-tic technologies. This research is conducted within the domains of applied mechanics and biomechanics, building upon prior investigations into the acoustic eradication of detrimental insects.The examination of contemporary scientific literature reveals that low-frequency acous-tic and vibrational phenomena exert both detrimental and beneficial effects on biological tis-sues. Research conducted by V. S. Didkovskyi, V. T. Hrynchenko, G. I. Sokol, and others vali-dates that resonance phenomena in biological structures can induce structural disruption when the external frequency aligns with the organism's natural frequency. The authors sug-gest using controlled acoustic waves to kill pest larvae by taking advantage of their biome-chanical resonance.To implement this concept, the authors devised a novel methodology for assessing the mechanical properties of living larvae, encompassing stiffness, mass, and Young’s modulus. The larvae were represented as cylindrical elastic entities with a uniform cross-section. A new measuring tool was created and patented (Patent of Ukraine No. 153662, 2023) for test-ing deformation in a controlled way with known loads. The setup has a micrometer, a ma-nometer, and a loading mechanism that can be changed to measure deformation in living bio-logical specimens very accurately. The experimental results indicated stiffness values between 0.3248 N/m and 1.1621 N/m, Young’s modulus ranging from 36.62 Pa to 79.07 Pa, and natural resonance frequen-cies from 5.23 Hz to 8.54 Hz. These results show that it is possible to get destructive reso-nance with acoustic emitters set to these frequencies.The developed methodologies and instrumentation establish a basis for the design of ef-fective acoustic devices for pest management. The use of these kinds of systems will make it possible to get rid of pests in agriculture in a way that is safe for the environment and doesn't use chemicals. This will greatly lower ecological risks and make sure that crops are protected in a way that is sustainable.

Smart sensing of Leptinotarsa decemlineata infestations using a portable optical array

Deep learning driven edge inference for pest detection in potato crops using the AgriScout robot

The initiation of pupal ecdysis behavior by ecdysis-triggering hormone (ETH) has been established in two Lepidopterans and two Dipterans. In Coleoptera Leptinotarsa decemlineata, RNA interference (RNAi) targeting eth impairs pupation. However, whether the RNAi phenotypes result from defects in organ formation or disruption of pupal ecdysis behavior has not been established. Here, we characterized ETH physiological functions during pupation in a potato defoliating ladybird, Henosepilachna vigintioctomaculata. Hveth levels were high just before or right after each molting, it was highly expressed in the trachea-rich tissues, such as dissected epidermis, fat body, and head. RNAi against Hveth impaired pupation of more than 80% of the resultant larvae. They were partially enveloped by the old larval cuticle until death. HE stained slices displayed that new cuticles were well formed under the old ones at the head, pleuron, and pronotum of prothorax. Moreover, adult organs such as compound eyes, mouthparts, and legs were well developed in both the control and the Hveth RNAi groups. Conversely, the freezing sections revealed that no clear separation between the old and the newly formed cuticles in the Hveth RNAi head, in contrast to the control head that showed a visible gap. In addition, the pupal cuticle under the old larval integument was not pigmented. Our findings suggest that ETH initiates pupal ecdysis behavior to shed off old larval exoskeleton and activates pigmentation of pupal cuticle during pupation in H. vigintioctomaculata.

Fungal lectins, with their specific glycan-binding activity, represent promising biopesticide candidates. This study investigates the potential of six fungal lectins to control Colorado potato beetle (CPB) larvae (Leptinotarsa decemlineata). In vitro feeding trials with recombinant lectins were performed to evaluate their individual toxicity. Marasmius oreades agglutinin (MOA) caused significantly higher mortality than the other lectins, whereas Coprinopsis cinerea lectin 2 (CCL2) and Aleuria aurantia lectin (AAL) completely inhibited weight gain. Functional analysis of MOA mutants revealed that both carbohydrate-binding and proteolytic domains are essential for toxicity. MOA binds to midgut glycoproteins in CPB larvae, disrupting midgut epithelium. Histology and ultramicroscopy showed that MOA causes loss of apicobasal polarity and detachment of basal lamina. No acute toxicity of lectins against adult European honeybees (Apis mellifera) was found; however, significant sublethal effects on larval development were observed. MOA shows promise as a selective bioinsecticide for sustainable CPB control with minimal nontarget effects.

The Colorado potato beetle (CPB), Leptinotarsa decemlineata Say (Coleoptera: Chrysomelidae), is as important pest of potato, Solanum tuberosum, in the U.S. CPB management is heavily dependent on broad-spectrum insecticides, thereby stimulating an interest to identify alternative solutions. In this study, we evaluated the efficacy of ledprona, a dsRNA interference-based bioinsecticide, following different application timings and frequencies to reduce CPB populations, minimize defoliation, and protect potato yields across diverse conditions in the northern U.S. Ledprona applied three to five times, starting when overwintered adults were present to a week after initial egg hatch, consistently provided the greatest suppression of first-generation larvae, and significantly reduced potato defoliation and yield loss. Applications initiated when larvae were abundant and most eggs had hatched, followed by one or two applications showed reduced effectiveness. While adult CPB control was inconsistent, ledprona effectively suppressed first-generation larval populations and defoliation, demonstrating its value as an effective novel bioinsecticide. Optimal timing and frequency of ledprona applications are critical for effective CPB control in potato. Our findings indicated that the phenology of the CPB infestation will need to be monitored closely early in the season to enable proper timing of the first ledprona application. We offer practical insights for how best to integrate dsRNA-based insecticides into season-long CPB management programs. © 2025 Society of Chemical Industry.