Nano-enabled plant fortification: green-synthesized SiO₂ and emamectin benzoate nanoparticles synergistically boost maize defense and agronomic performance against Spodoptera frugiperda infestation

Abstract

Zea mays L., a globally vital C₄ cereal, is increasingly threatened by fall armyworm (FAW), Spodoptera frugiperda (J. E. Smith), a destructive and insecticide-resistant pest. This study developed a nano-enabled strategy integrating green-synthesized SiO₂ nanoparticles (GS-SiNPs) for plant fortification with nano-formulated emamectin benzoate (EMB-NPs) for enhanced insecticidal activity. Laboratory bioassays on 4^th-instar FAW larvae evaluated acute toxicity (LC₅₀ and LC₉₀) and detoxification enzyme activity. A field experiment in Egypt, autumn 2024 used a randomized complete block design to test two foliar sprays on maize in ten treatments with four replicates. Larval counts, leaf damage, anatomy, photosynthesis, leaf area (LA) plant⁻¹, Si content, and yield were assessed. Laboratorially, LC₉₀ (ppm) values were 93.6 (EMB-NPs), and 122.7 (EMB bulk), with GS-SiNPs exhibiting the steepest (5.18). GS-SiNPs with EMB bulk or EMB-NPs exhibited LC₅₀ values of 102.0 and 71.8 ppm, respectively, indicating a synergistic effect of both mixtures. EMB bulk + GS-SiNPs and EMB-NPs + GS-SiNPs suppressed larval detoxification enzymes. Field results revealed 100% initial larval mortality. The ½EMB-NPs + GS-SiNPs reduced leaf damage by 64.2% after the 1^st spray, while ¾EMB-NPs + GS-SiNPs achieved 86.4% after the 2^nd spray. This treatment also induced significant anatomical modification, increasing blade, midvein, and vascular bundle thickness. It enhanced photosynthesis, leaf Si, and LA plant⁻¹, and boosted yield by 54.5% vis-à-vis control. Combining GS-SiNPs with EMB-NPs, particularly ¾EMB-NPs + GS-SiNPs, enhanced EMB bioefficacy and suppressed FAW detoxification while improving maize’s physio-anatomical resilience. This nano-enabled sustainable strategy offers a dose-efficient and eco-friendly approach for FAW management and maize productivity.

Background

Maize (Zea mays L.) is a globally cultivated C4 staple cereal crop with high yield potential, broad agro-climatic adaptability, and pivotal contributions to food security, nutrition, and industry^1,2. Its grains are rich in starch, protein, fiber, fat, essential minerals, and vitamins, and possess superior antioxidant and nutritional properties compared to other cereals³. Globally, maize was harvested from ~ 208.2 million hectares (M ha) in 2023, yielding ~ 1241.6 million tons; M t (5.96 t ha⁻¹), while in Africa, ~ 44.1 M ha produced ~ 95.0 Mt (2.15 t ha⁻¹)⁴. In Egypt, 2022 summer maize cultivation covered ~ 2.0 million feddans (1.75 white and 0.25 yellow), yielding ~ 6.43 Mt at ~ 3.24 and 3.04 t feddan⁻¹ (one feddan = 4200 m²) in old and new agricultural lands, respectively⁵.

The fall armyworm (FAW), Spodoptera frugiperda (Lepidoptera: Noctuidae; J. E. Smith), is a highly destructive polyphagous lepidopteran pest^6,7,8. Historically restricted to the Americas, it has emerged in recent decades as a formidable invasive species, now posing substantial threats to agricultural systems and food security in Africa and Asia^9,10,11. The FAW is regarded as the most economically important pest of maize due to its capability to cause severe yield losses^{12,13,14,15,16}. Owing to its heightened resistance to multiple synthetic insecticides and transgenic maize hybrids, FAW has become a formidable agricultural pest, urging effective and sustainable control management approaches^{17,18,19,20,21}. To ensure control effectiveness, insecticide selection should be guided by field-evaluated resistance patterns. In field populations of FAW, emamectin benzoate (EMB) treatments did not result in resistance-conferring alterations in the glutamate-gated chloride channel (GluCl) gene^22,23,24,25. Emamectin benzoate, a semi-synthetic macrolide, is a derivative of avermectin produced by the actinomycete Streptomyces avermitilis via natural fermentation. Its insecticidal activity results from its high-affinity and selective binding to GluCl channels interrupts nerve impulses and paralyzes insects. GluCl channels are crucial for regulating Cl⁻ flux across insect neuronal membranes^26,27.

Taken together, the above information underscores that effective and sustainable control management of the FAW remains a complex ongoing challenge. Overcoming this challenge requires the development of control materials and innovative application technologies to control this highly destructive pest¹³. Nanotechnology is poised to play a transformative role in plant protection by facilitating the development of environmentally sustainable, targeted, and controlled-release pest management strategies^28,29. These advancements help resolve shortcomings of conventional pesticides, such as off-target effects, environmental persistence, and rapid degradation^30,31. The formidable challenge posed by S. frugiperda (J. E. Smith) in maize necessitates synergistic and multi-pronged control strategies³². While EMB, as an organic insecticide, is a highly effective insecticide against FAW, its short residual activity and potential for off-target effects limit its long-term efficacy^33,34. The integration of green-synthesized SiO₂ nanoparticles (GS-SiNPs) with EMB nanoparticles (EMB-NPs)³² provides an innovative dual function, simultaneously serving as a plant-fortifying micronutrient and a nanoscale carrier for enhanced insecticidal bioefficacy and delivery^35,36. As a microelement, silicon (Si) deposition in plant cell walls physically strengthens maize against larval feeding³⁷, while also inducing systemic resistance by activating defense-related genes and antioxidant systems under biotic stress³⁸. In a nano-formulation, nanosilicon (SiNPs) can encapsulate EMB, facilitating controlled and sustained release of the active ingredient, thereby prolonging its insecticidal activity^39,40 and reducing application frequency. This integrated approach not only improves the efficacy and environmental profile of EMB but also leverages a plant’s intrinsic defense mechanisms, representing a significant advancement in sustainable pest management against S. frugiperda (J. E. Smith)^41,42.

The insecticidal efficacy of EMB bulk⁴³, EMB-NPs⁴⁴ or GS-SiNPs⁴⁵ against S. frugiperda (J. E. Smith) has been individually documented. However, a critical knowledge gap remains regarding the synergistic potential of co-applying these nanomaterials to overcome the limitations of single-component treatments, such as the short residual activity of biopesticides or the variable efficacy of physical fortification. Therefore, this study was designed to address this gap by developing a novel, integrated strategy that couples GS-SiNPs with EMB-NPs, under both laboratory and field conditions. Unlike prior studies, this approach is designed to function as a dual-action defense system: utilizing GS-SiNPs to structurally fortify maize anatomical barriers and suppress larval detoxification enzymes, thereby maximizing the insecticidal impact of EMB-NPs. We hypothesized that the EMB-NPs and GS-SiNPs co-treatment would provide a dual-function strategy: (i) direct effectively insecticidal action against FAW through EMB, and (ii) enhanced plant fortification through Si-mediated improvements in anatomical integrity, morpho-physiological resilience, and grain productivity. Mechanistically, EMB-NPs may provide sustained pest suppression through controlled release, while GS-SiNPs enhance cell wall integrity, and antioxidative defenses together creating a synergistic plant-pest management interface. Accordingly, our objectives were to (i) evaluate EMB (bulk and nano) alone and in combination with GS-SiNPs for FAW control and detoxification enzyme modulation, (ii) determine their integrated effects on photosynthetic function, leaf anatomical structure, morpho-physiological attributes, and yield, and (iii) elucidate the extent to which GS-SiNPs enhance EMB performance and persistence.

Materials and methods

Chemicals and analytical reagents

Emamectin benzoate (EMB; Speedo® 5.7% WDG, Shoura Chemicals) was applied as the insecticidal treatment in this study. Sodium metasilicate pentahydrate (Na₂SiO₃·5H₂O, ≥ 98.8%) and ammonium hydroxide solution were purchased from Advent Chembio Pvt. Ltd. (Mumbai, India). All solvents used were of analytical grade and dried over molecular sieves before use. Grinding media consisting of stainless-steel balls with diameters of 3 mm, 5 mm, and 10 mm were employed for different stages of the ball milling process. All chemicals were used without further purification unless otherwise specified. The reference standard for quantifying total protein was bovine albumin powder (98%, VWR chemicals Avantor®, Belgium). Folin-ciocalteu’s phenol reagent (FCR) (Sigma-Aldrich) was used to form a colored complex. To assess esterase activity, 1-Naphthyl acetate (99.5% was obtained from Alpha Chemika, India) was utilized as substrate, sodium dodecyl sulfate (SDS 99%, AVI-Chem Laboratories), and fast Blue B salt 95% powder (Sigma-Aldrich) as a colorimetric reagent. GST activity was measured react 1-chloro 2,4-dinitrobenezene (CDNB 97%, Sigma-Aldrich) as a substrate with reduced glutathione (GSH, Sigma-Aldrich).

Preparation of green-synthesized SiO₂ nanoparticles (GS-SiNPs) and emamectin benzoate nanoparticles (EMB-NPs)

Preparation of Pelargonium odoratissimum (Linnaeus) leaf extract

The air-dried powder leaves of P. odoratissimum L. (20 g) were taken and then submerged in 400 mL of deionized water (dH₂O). The ultrasonic-assisted solvent extraction (UASE) method⁴⁶ was used for the extraction process by placing the Erlenmeyer flask in a Probe Sonicator homogenizer (Benchmark Scientific, USA, 150 W, 25 kHz) for 30 min at ambient temperature (35 ± 2 ºC). Whatman filter paper No. 1 was used after muslin cloth to filter the solvent (dH₂O) and powder layer. The filtrate solution of P. odoratissimum L. leaf extract was stored in a refrigerator until use. The selection of P. odoratissimum L. leaf extract for the biosynthesis of GS-SiNPs is due to its rich profile of bioactive phytochemicals, particularly phenolics and flavonoids. These secondary metabolites contain hydroxyl and carboxyl functional groups that facilitate the reduction of silica precursors, such as sodium metasilicate pentahydrate, to silica nanoparticles. Furthermore, these biomolecules serve a critical role as natural capping and stabilizing agents, forming a coating on the nanoparticle surface that prevents agglomeration and enhances stability without the need for toxic chemical surfactants.

Green synthesis of SiO₂ nanoparticles

Sodium metasilicate pentahydrate, as a precursor, was used for synthesizing nanosilica using the green synthesis route. 20 mL of precursor solution (0.5 mol L⁻¹) was added dropwise to 100 mL of freshly P. odoratissimum L. aqueous leaf extract and the mixture was being continuous stirring at 50 °C for 1 h until it was reduced into a jelly-like precipitation was formed. The pH of the nanoparticles mixture was adjusted by adding an ammonium hydroxide solution (0.1 mol. L⁻¹) to maintain the pH value of 11.50. Afterwards, the precipitate was washed several times with dH₂O, centrifuged (Sigma 2k15, USA) at 4000 rpm for 10 min at 4 °C and collected the precipitate in a crucible, and dried overnight in an oven at 100 °C then calcined in a muffle furnace at 600 °C for 2 h. Elevating the temperature in the muffle furnace was mandatory to remove any organic residuals. Finally, a white powder was obtained and stored carefully in a sterile air-tight container for further characterization.

Ball milling synthesis procedure for emamectin benzoate nanoparticles (EMB-NPs)

Equipment and setup

The synthesis of EMB was carried out using a high-energy planetary ball mill (Retsch PM 400, Retsch GmbH, Haan, Germany) equipped with a 500 mL stainless steel grinding jar and an automated vacuum chamber system. The milling apparatus featured programmable time and speed control with automatic start-stop functionality and vacuum maintenance throughout the synthesis process. The vacuum chamber was evacuated to 10⁻² mbar and backfilled with high-purity nitrogen gas to maintain an inert atmosphere⁴⁷. Temperature control is achieved through integrated thermocouples within the milling jar, ensuring that the temperature does not exceed 65 °C during operation. The grinding media consists of stainless-steel balls with three different diameters: 3 mm, 5 mm, and 10 mm. These varying ball sizes are strategically employed across different stages of the milling process to achieve progressive particle size reduction and enhanced mechanical activation.

Three-stage processing approach

The ball milling procedure was conducted in three distinct stages with systematically varying parameters. The 1^st stage employs 10 mm stainless steel balls at a ball-to-powder weight ratio of 8:1, with the planetary ball mill operating at 300 rpm for 2 h total duration. This initial processing stage utilizes programmed cycles of 20 min of milling followed by 10 min of pause to prevent overheating and allow for controlled particle size reduction. The 2^nd stage utilizes 5 mm balls at an increased ratio of 12:1, with the rotation speed elevated to 450 rpm for 4 h total duration. The milling cycles during this intermediate processing phase consist of 30 min of milling followed by a 15-min pause, providing enhanced particle reduction and improved mixing efficiency. This stage represents the most intensive phase of the process in terms of total duration. The final stage employs the smallest 3 mm balls at the highest ratio of 15:1, operating at the maximum rotation speed of 500 rpm for 2 h. The milling cycles in this stage feature 45 min of continuous operation followed by 10-min cooling periods. This final processing stage achieves the finest particle refinement and maximum mechanical activation of the material^48,49.

Process control and monitoring

Throughout the entire 8-h process, the vacuum system maintains consistent pressure below 10⁻² mbar with automatic nitrogen backfilling between stages. Intermediate sampling is performed at the end of each stage under controlled vacuum conditions to monitor progress. The automated pause cycles allow for periodic scraping of adhered material from the jar walls using appropriate tools, ensuring uniform mixing throughout each synthesis stage.

The progressive processing principle underlying this multi-stage approach allows for systematic particle size reduction, enhanced mechanical activation, improved uniformity, controlled energy input, and optimized processing efficiency. The combination of decreasing ball sizes with increasing speeds and ball-to-powder ratios creates a controlled environment for achieving desired material properties through purely mechanical means^50,51,52.

Characterization of the prepared nanoparticles

The GS-SiNPs and EMB-NPs were characterized using advanced spectroscopy techniques. The size and shape of the green synthesized GS-SiNPs and EMB-NPs were determined using high-resolution transmission electron microscopy (JEM-2100, JEOL, Japan). A particle size analyzer and zeta potential (PSS Nicomp ZPW388-V2.14, Inc., Santa Barbara, Calif., USA) were utilized to determine the particle size distribution (PSD) and identify the stability of the GS-SiNPs and EMB-NPs obtained.

Laboratory experiment

Rearing of Spodoptera frugiperda (J. E. Smith)

The S. frugiperda (J. E. Smith) larvae were collected from maize plantations in the Fayoum Governorate, Egypt, to establish a laboratory strain in order to allow the insect to adapt to laboratory conditions. The larvae were reared up to the third generation (G3) without exposing them to any pesticides. The collected larvae were transported to the Plant Protection Laboratory, Faculty of Agriculture, Fayoum University, and reared in an incubator under controlled conditions (20 ± 5 °C, 60 ± 5% relative humidity (ReH), and a 14 h light/10 h dark). The larvae were reared in a 32-cell artificial diet tray system with detachable four-cell lids (RT32W white trays and TRPV4 lids; Frontier Agricultural Sciences, Newark, DE, USA) and supplied daily with fresh Ricinus communis L. leaves as a feed source. The diet tray system was cleaned daily, and the bottom of each tray was lined with a thin layer of fine sawdust to facilitate moisture absorption and promote successful pupation. The pupae were then transferred to clean plastic containers (10 cm in diameter and 20 cm in height). The containers were covered with muslin cloth to ensure adequate aeration until adult emergence. Emerged adults were maintained in aerated containers provided with paper rolls moistened with a 10% sucrose solution as an energy-rich dietary supplement. Filter papers were provided as egg-laying substrates, and eggs were carefully collected daily to maintain colony propagation⁵³.

Insecticidal activity via larval bioassay

The acute toxicity of EMB bulk, EMB-NPs, and GS-SiNPs against 4^th-instar S. frugiperda (J. E. Smith) larvae was evaluated via the standardized leaf-dip bioassay. Three concentrations of EMB bulk (25, 50, and 75 mg L⁻¹), EMB-NPs (25, 50, and 75 mg L⁻¹), and GS-SiNPs (200, 250, and 300 mg L⁻¹) were prepared. The tested EMB bulk + GS-SiNPs and EMB-NPs + GS-SiNPs mixtures were assessed for determination of their joint toxicity on 4^th-instar FAW larvae at concentrations corresponding to their respective lethal concentration for 25% (LC₂₅) values. Each treatment consisted of four replicates, with ten larvae per replicate. Fresh Ricinus communis leaves were immersed for 30 s in either the tested solutions or dH₂O as a control, air-dried at room temperature until completely dry, and then offered to larvae^53,54. Mortality was recorded 24 h post-exposure treatment. Corrected mortality rates were calculated using Abbott’s⁵⁵ formula, and median lethal concentrations (LC₅₀) were calculated through Finney’s Probit analysis⁵⁶. Co-toxicity factor (CF) was calculated based on the LC₂₅ values according to Mansour et al.⁵⁷ to express the joint activity between combinations. Three groups of findings differentiated by this factor. A positive factor of ≥ 20 indicates a synergism, a negative factor of ≤  − 20 indicates an antagonism, and intermediate values of >  − 20 to < 20 represent an additive effect, as following equation: CF = [(O – E)/E] × 100; where O and E are observed and expected mortality percentages, respectively.

Larval sample preparation for total protein quantification and detoxifying enzyme bioassays

The 4^th-instar larvae of S. frugiperda (J. E. Smith) were exposed to LC₂₅, with four independent replicates, 24 h post-exposure treatment. Each replicate comprised 1000 mg of surviving larvae. The selection of 4^th-instar larvae for biochemical analyses was based on specific physio-toxicological considerations. This developmental stage coincides with peak metabolic activity and intense feeding behavior⁵⁸, ensuring measurable baseline activities of key detoxification and digestive enzymes such as glutathione S-transferase (G-S-T) and total esterases (TE), as well as total protein content (TPC), which are essential for assessing physiological disruption. Moreover, 4^th-instar larvae possess sufficient biomass to provide adequate enzyme extract volumes for parallel spectrophotometric assays, thereby reducing sampling errors associated with the lower protein yields of earlier instars. From a toxicological standpoint, 4^th-instar represents a relatively more tolerant phenotype than younger larvae; consequently, pronounced enzymatic inhibition at this stage constitutes a robust indicator of the nano-formulation’s efficacy against field-relevant, damaging larvae⁵⁹. To assess the TPC and the activities of G-S-T and TE, whole larvae were homogenized in ice-cold 0.1 M NaH₂PO₄ buffer (pH 7.0). The homogenates were centrifuged at 5000 rpm for 20 min in a refrigerated centrifuge (4 °C), and the resulting supernatants were transferred to sterile Eppendorf tubes and kept frozen at − 20 °C pending enzymatic assays.

Larval detoxifying enzyme activity was evaluated after exposure to EMB bulk, EMB-NPs, or GS-SiNPs, and their mixtures (EMB bulk + GS-SiNPs and EMB-NPs + GS-SiNPs) at the LC₂₅ level. The TPC of 4^th-instar larvae was quantified following the Lowry method⁶⁰, using bovine serum albumin as the reference standard. The TE enzyme activity was assayed according to Gomori⁶¹, while G-S-T enzyme activity was determined following the protocol described by Scharf et al.⁶².

Field-based agronomic experiment

Site description, experimental setup and design, and crop management

An on-farm agronomic experiment was carried out at the Experimental Station Farm (29° 19′ 30.4″ N, 30° 51′ 44.8″ E), Faculty of Agriculture, Fayoum University, Egypt, during the 2024 autumn season. The experimental soil was classified as clay texture. At 0.0–0.4 m depth, bulk density (g cm⁻³), organic matter (%), available N (%), available P (mg kg⁻¹), available K⁺ (mg kg⁻¹), and CaCO₃ (%) were 1.38, 1.15, 0.04, 5.7, 60.95, and 4.79, respectively, as determined following Klute and Dirksen⁶³ and Page et al.⁶⁴. The available Si content was 32.6 mg kg⁻¹⁶⁵. The climatic data for the maize growing season (Table 1) were obtained from the nearest official meteorological station, located approximately 5 km from the experimental site. These conditions reflect a typical hot-arid summer with gradual autumnal moderation. The maximum air temperature was consistently high from June to August (~ 39–40 °C), while maximum temperatures declined in September and October. Th ReH gradually increased from early to late season, whereas solar radiation increased steadily, peaking in September–October, indicating severe atmospheric dryness and high irradiance during the grain-filling stage. Precipitation was negligible throughout the season, rendering regular irrigation is essential for sustaining crop growth.

Table 1 Mean monthly air temperature, average relative humidity (ReH), solar radiation (SR), and precipitation at Fayoum, Egypt, during the 2024 maize growing summer season.

The experiment was laid out in a randomized complete block design (RCBD) consisting of ten treatments as detailed in Table 2. Each tested treatment was replicated four times, resulting in a total of 40 experimental plots. Each experimental plot area measured 12.6 m², consisting of six planting ridges (3 m in length × 0.7 m in width). Healthy grains of maize (Zea mays L., white single-cross hybrid Hytech 2031) were obtained from Misr Hytech Seed International Company, Cairo, Egypt. This hybrid is known for strong stalks, a stay-green habit, tolerance to lodging and late wilt, dual suitability for grain and silage, and a maturity period of 115–120 days after planting (DAP). At a 3–5 cm soil depth, two maize grains were manually sown in hills spaced 0.25 m apart along one side of the planting ridge on 25 June and harvested on 20 October during the 2024 autumn growing season. At 21 DAP, maize seedlings were hand-thinned to one vigorous and morphologically uniform plant per hill to optimize stand density, minimize inter-plant competition, and equalize growth conditions.

Table 2 Description of field experimental treatments applied for controlling the invasive S. frugiperda (J. E. Smith), in Zea mays L. in this study during the 2024 autumn growing season.

The entire recommended basal phosphorus fertilizer rate (74.4 kg P₂O₅ ha⁻¹) was incorporated into the upper soil horizon (0–0.2 m) prior to planting. Potassium fertilizer, at the recommended rate of 57.6 kg K₂O ha⁻¹, was applied in two equal splits, topdressed post-thinning (21 DAP) and again at 35 DAP. Nitrogen (N) fertilizer was applied at a total rate of 288 kg N ha⁻¹ in three topdressings. The initial dose (48 kg N ha⁻¹) was incorporated into the root zone during sowing, while the 2^nd and 3^rd doses (120 kg N ha⁻¹ each) were manually topdressed post-thinning and at 35 DAP, respectively. All other standard agronomic practices for maize cultivation, apart from targeted pest management, i.e., S. frugiperda (J. E. Smith), were conducted according to local commercial recommendations under the Delta Nile clay soil conditions in Egypt.

Field application protocol of emamectin benzoate (EMB) and green-synthesized SiO ₂ nanoparticles (GS-SiNPs) in maize field

The commercial insecticide EMB was applied at a concentration of 400 mg L⁻¹ (equivalent to 192 g ha⁻¹) based on the official recommendations of the Egyptian Agricultural Ministry for S. frugiperda (J. E. Smith) management in maize⁶⁶. However, the GS-SiNPs concentration of 370.8 mg L⁻¹ (equivalent to 177.98 g ha⁻¹) was selected based on its lethal concentration required to cause 90% mortality (LC₉₀) reported under these study conditions. Before initiating spray treatments, the baseline larval infestation incidence was determined to standardize comparisons across all experimental treatments. This was calculated by dividing the number of larva-harbored maize plants by the total number of examined maize plants, expressed as a percentage. The ten treatments were applied at 07:00 a.m. via two complementary delivery methods: (i) foliar spraying with a 20 L hand-operated knapsack sprayer and (ii) direct application into the leaf whorls of each maize plant (3 mL each) with a 50 mL plastic syringe. Applications were applied twice, at 14 DAP, when the mean infestation averaged 38.6% (33.3–42.9%), and again at 33 DAP, with treatments delivered in 480 L ha⁻¹ of dH₂O as the diluent. These application timings were synchronized with specific maize development stages according to the Biologische Bundesanstalt, Bundessortenamt und Chemische Industrie (BBCH) scale in Germany⁶⁷: BBCH 13–14 (≈ 3–4 leaves unfolded) and BBCH 18–19 (≈ 8–10 leaves fully unfolded, extending to the onset of stem elongation). To enhance wettability and ensure efficacious penetration of the spray into maize tissues, each application was supplemented with Tween®-20 (0.02%, v/v) as a non-ionic surfactant.

Sampling, larval infestation/crop measurements, and calculations

Larval counts and canopy damage assessment

To assess treatment efficacy, the numbers of live and dead larvae plant⁻¹ were recorded in each experimental plot both before and after treatment application. In each plot, ten maize plants (40 plants per treatment) exhibiting visible S. frugiperda (J. E. Smith) feeding symptoms were selected for observation. Pre-treatment larval counts were conducted 1 day before by carefully opening the leaf whorl of each sampled plant, whereas post-treatment counts were performed at 1, 5, and 14 days post-application. The extent of canopy damage severity was then calculated as described by Toepfer et al.⁶⁸.

Determination of photosynthetic parameters and leaf area

At 70 DAP, which corresponds to the BBCH 60/61 stage (beginning of tassel emergence and anthesis), five plants were randomly chosen in each plot to measure Soil Plant Analysis Development (SPAD)-chlorophyll content index (SPAD-CCI) using a non-destructive SPAD₅₀₂ chlorophyll meter (Konica Minolta, Inc., Tokyo, Japan). Leaf photosynthetic performance index on an absorption basis (PPI_ABS) was assessed using a Portable Handy PEA chlorophyll fluorimeter (Hansatech, Kings Lynn, UK). Measurements were taken from the middle portions of the youngest fully expanded leaves at 10:00 a.m. After a 20-min dark adaptation, minimum (F_o) and maximum (F_m) fluorescence were measured to calculate variable fluorescence (Fᵥ = F_m – F_o), maximum quantum yield of photosystem II (PSII) [F_v/F_m = (F_m − F_o)/F_m], and potential PSII activity (F_v/F_o) following Maxwell and Johnson⁶⁹. The PPI_ABS, reflecting electron flux from PSII to intersystem electron acceptors, was determined following Clark et al.⁷⁰. The five previously selected maize plants were harvested and immediately transported to the laboratory to count the leaf number plant⁻¹ and to determine the leaf area (LA) plant⁻¹⁷¹, calculated as follows:

$${\text{LA}}\;{\text{plant}}^{ - 1} \left( {{\text{dm}}^{2} } \right) = \frac{{\left( {{\text{maximum}}\;{\text{width}} \times {\text{maximum}}\;{\text{length}}\;{\text{of}}\;{\text{the}}\;{\text{fourth}}\;{\text{leaf}}\;{\text{from}}\;{\text{the}}\;{\text{plant}}\;{\text{tip}} \times 0.75 \times {\text{leaf}}\;{\text{number}}\;{\text{plant}}^{ - 1} } \right)}}{100}$$

Leaf anatomical structure

Flag leaf samples were fixed in FAA (50 ml 95% ethanol + 10 ml formaldehyde + 5 ml glacial acetic acid + 35 ml dH₂O) for 2 d^72,73. Thereafter, the samples were washed in 50% ethanol, dehydrated through a tert-butanol series, and embedded in paraffin wax. Transverse sections, 20 μm thick, were cut by a rotary microtome (Zhejiang Jinhua Kedi, China), mounted with Haupt’s adhesive, stained with crystal violet-erythrosin⁷⁴, cleared in carbol xylene, and sealed in Canada balsam. The slides were examined under an upright light microscope (AxioPlan, Zeiss, Jena, Germany), and images were analyzed with CaseViewer 2.3 (3DHISTECH Ltd.).

Determination of leaf silicon (Si) content

For leaf Si determination, maize leaf tissue samples were oven-dried at 70 °C, ground, and screened through a 60-mesh sieve. A 0.2 g representative sample was then placed into a 100 mL polyethylene vessel, to which 10 mL of 1 M HCl and 20 mL of 2.3 M HF were added, following the procedure of Novozamsky et al.⁷⁵ as adapted by Taber et al.⁷⁶. The bottles were sealed and agitated on a rotary shaker at 180 rpm for 15 h. The resulting extract was filtered through Whatman #41 ashless filter paper into a 50 mL polyethylene tube. An aliquot of this filtered extract was transferred into a 10 mL polycarbonate test tube for analysis using inductively coupled argon plasma optical emission spectroscopy (ICAP-OES, Thermo-Jarrell Ash model IRIS 1000 dual-view, Franklin, MA, USA). Plasma power was adjusted to 1150 W, and the detection wavelength for Si detection was 250.690 nm. A standard calibration curve was established using 0, 10, and 50 mg Si L⁻¹ standards, and a reagent blank was included to correct for background contamination.

Grain yield and its related attributes

At 117 DAP, aligned with the BBCH 88/89 developmental stage (physiological maturity), grain yield-associated parameters were assessed on 10 randomly sampled plants from the plot. Cob length and diameter (cm) were measured. The shelling percentage for each treatment was calculated as the ratio of grain weight from 10 cobs to their pre-shelling weight prior to shelling. The 100-grain weight (100-GW in g) was determined by weighing 100 grains randomly bulked from each plot after oven-drying at 80 °C to a constant weight. To estimate grain yield (t ha⁻¹), the maize grain harvested from all plants within the central four ridges (8.4 m² per plot), together with the grain weight of the ten previously sampled plants, was measured. The values were then converted to tons per hectare on a 15.5% seed moisture basis.

Data processing and statistical analysis

Mortality data were corrected using Abbott’s⁵⁵ formula, and the lethal dose-probability (Ldp) line was estimated through Probit analysis following Finney’s⁵⁶ method. Normality of the data was first assessed using the Shapiro–Wilk test (p > 0.05) to verify the assumptions underlying analysis of variance (ANOVA)⁷⁷. To meet the assumptions of ANOVA, percentage data were transformed prior to analysis using the arcsine square root transformation (arcsine √x/100). A randomized complete block design (RCBD) with four replicates was used in the current laboratory and field experiments. Post-hoc mean comparisons were conducted using Duncan’s multiple range test at p ≤ 0.05 probability level, implemented through INFOSTAT software version 2020 (Córdoba University, Córdoba, Argentina).

Results

Characterization of green-synthesized SiO₂ (GS-SiNPs) and emamectin benzoate (EMB-NPs) nanoparticles

The obtained image (Fig. 1A) showed that the GS-SiNPs have particle sizes within the nanometer range. High-resolution transmission electron microscopy (HR-TEM) was used to verify the morphology and size distribution of the nanoparticles. The HR-TEM micrographs (Fig. 1B) revealed that GS-SiNPs occurred in multiple size classes and exhibited a predominantly spherical morphology. The particles were composed of finer subunits, with an average size of 32.18 nm (Fig. 1D). Furthermore, the stability of the GS-SiNPs was confirmed by zeta potential analysis, which recorded a value of − 29.7 mV. The selected area electron diffraction (SAED) pattern (Fig. 1C) showed well-defined diffraction rings generated by the superposition of diffraction spots from many randomly oriented crystallites, confirming the crystalline nature of the prepared material.

Fig. 1

Zeta sizer analysis (A); zeta potential (B); high-resolution transmission electron microscopy (HR-TEM) image (C); selected-area electron diffraction (SAED) pattern (D); and particle-size distribution histogram (E) of the green-synthesized SiO₂ nanoparticles (GS-SiNPs) prepared in this study.

Figure 2A–C displays the shape, size, and size distribution of EMB NPs. The HR-TEM image reveals a good distribution of spherical EMB-NPs without any aggregation. Moreover, image analysis revealed that the average size of EMB-NPs was approximately 45.58 nm. This finding was corroborated by zeta potential measurements, which showed a surface charge of + 24.6 mV.

Fig. 2

Zeta sizer analysis (A), zeta potential (B), high-resolution transmission electron microscopy (HR-TEM) image (C), and particle-size distribution histogram (D) of nano-emamectin benzoate (EMB-NPs) synthesized using high-energy planetary ball milling in this study.

Laboratory experiment

Acute toxicity

The acute toxicity of the tested substances and their mixtures was evaluated based on their LC₅₀ values, as presented in Table 3. After 24 h of exposure, the LC₅₀ values against 4^th-instar FAW larvae for GS-SiNPs, EMB-NPs, and EMB bulk, individually, were recorded as 209.7, 20.7, and 35.9 ppm, respectively. The corresponding LC₉₀ values were 370.8, 93.6, and 122.7 ppm, respectively. All evaluated substances exhibited larvicidal efficacy against FAW. Moreover, GS-SiNPs showed the steepest slope (5.18), followed by EMB bulk (2.39) and EMB-NPs (1.95), indicating that a small increase in GS-SiNPs concentration results in a substantial rise in larval mortality. Whilst the LC₅₀ of EMB bulk + GS-SiNPs and EMB-NPs + GS-SiNPs mixtures were 102.0 and 71.8 ppm, respectively. Effect of combined for the two mixtures showed synergistic activity against S. frugiperda (J. E. Smith) larvae, where the co-toxicity factor recorded 34.8 and 37.5, respectively. Consequently, the efficacy of GS-SiNPs + EMB bulk or NPs combinations against FAW can make them promising insecticides for field application.

Table 3 Lethal concentration and probit slope of S. frugiperda (J. E. Smith) larvae with green synthesized silicon nanoparticles (GS-SiNPs), emamectin benzoate (EMB) nanoparticles (EMB-NPs), and EMB bulk formulation.

Enzymatic activity

Table 4 presents the TPC and activities of key detoxifying enzymes, namely G-S-T and TE, in S. frugiperda (J. E. Smith) larvae treated with EMB bulk, EMB-NPs, and GS-SiNPs, either individually or in binary mixtures with GS-SiNPs. The TPCs in S. frugiperda (J. E. Smith) larvae treated with EMB bulk and EMB-NPs were 3.39 and 3.19 mg g⁻¹ body weight (BW), respectively. When combined with GS-SiNPs, these values were enhanced to 5.75 and 3.67 mg g⁻¹ BW, respectively. S. frugiperda’s larvae treated with GS-SiNPs alone also exhibited significantly higher protein levels than the control (sprayed only with dH₂O). The highest protein content was recorded in S. frugiperda (J. E. Smith) larvae treated with the EMB bulk + GS-SiNPs mixture.

Table 4 Total protein content (TPC) and detoxification enzyme activities in S. frugiperda (J. E. Smith) larvae treated with emamectin benzoate (EMB) in bulk and nano forms and/or green synthesized silicon nanoparticles (GS-SiNPs)-based treatments under laboratory conditions.

The G-S-T activity in S. frugiperda (J. E. Smith) larvae treated with EMB bulk + GS-SiNPs or EMB-NPs + GS-SiNPs mixture was significantly lower than in those receiving their individual counterparts. The G-S-T activities were 0.393 and 0.453 mmol min⁻¹ mg⁻¹ protein for EMB bulk + GS-SiNPs and EMB-NPs + GS-SiNPs, respectively. The S. frugiperda (J. E. Smith) larvae treated with EMB bulk showed higher G-S-T activity than those treated with EMB-NPs. The GS-SiNPs treatment alone induced the highest G-S-T activity (0.630 mmol min⁻¹ mg⁻¹ protein), followed closely by EMB bulk (0.587 mmol min⁻¹ mg⁻¹ protein) and EMB-NPs (0.460 mmol min⁻¹ mg⁻¹ protein). The combination of EMB and GS-SiNPs led to a significant decline in G-S-T activity. The lowest activities were recorded in the EMB bulk + GS-SiNPs (0.393 mmol min⁻¹ mg⁻¹ protein) and EMB-NPs + GS-SiNPs (0.453 mmol min⁻¹ mg⁻¹ protein) mixtures (Table 4).

Among all treatments, the highest TE activity was observed in S. frugiperda (J. E. Smith) larvae treated with EMB bulk (0.371 µmol min⁻¹ mg⁻¹ protein), followed closely by those treated with EMB-NPs (0.329 µmol min⁻¹ mg⁻¹ protein). Conversely, TE activities in S. frugiperda (J. E. Smith) larvae exposed to EMB bulk + GS-SiNPs (0.244 µmol min⁻¹ mg⁻¹ protein) and EMB-NPs + GS-SiNPs (0.293 µmol min⁻¹ mg⁻¹ protein) were significantly lower than in their individual counterparts. The lowest TE activity was recorded in S. frugiperda (J. E. Smith) larvae treated solely with GS-SiNPs (0.244) (Table 4).

Field experiments

Number of Spodoptera frugiperda (J. E. Smith) larvae per maize plant

Results presented in Table 5 revealed that, for the 1^st spray interval, pre-spray (1 DBT) mean S. frugiperda (J. E. Smith) larvae counts ranged from 1.90 to 3.10 larvae plant⁻¹, with no statistically significant differences (p > 0.05) among plots. All tested treatments exhibited total larval mortality (100% reduction) 24 h post-spray (1 DAT), whereas S. frugiperda (J. E. Smith) larvae remained abundantly in the control plots (3.90). Surviving S. frugiperda (J. E. Smith) larvae in the control and dead larvae in treated plots mostly belonged to the early (1^st or 2^nd) instar. By 5 DAT, larval resurgence occurred in all treatments, with the mean number of surviving larvae ranging from 1.90 larvae plant⁻¹ in the ½EMB-NPs + GS-SiNPs to 4.80 larvae plant⁻¹ in the full-dose EMB bulk treatment. Differences remained statistically insignificant (p > 0.05) among treatments or between treated and control plots at this interval. At 14 DAT, larval counts escalated further across treatments, reaching up to 4.30 larvae plant⁻¹ in the full-dose EMB bulk treatment. Intriguingly, the control plants displayed a relatively lower mean of 1.90 larvae plant⁻¹, with no significant differences (p > 0.05) among treatments at this stage.

Table 5 Mean number of S. frugiperda (J. E. Smith) larvae per maize plant treated with emamectin benzoate (EMB) in bulk and nano forms and/or green synthesized silicon nanoparticles (GS-SiNPs)-based treatments under field conditions during the 2024 autumn growing season.

For the 2^nd spray interval, pre-spray (1 DBT) larval counts ranged from 2.00 to 2.60 plant⁻¹ in treated plots and reached 3.50 larvae plant⁻¹ in the control, with no statistically significant differences (p > 0.05) observed (Table 5). One day post-spray (1 DAT), all tested treatments resulted in 100% larval mortality, while the control plots maintained an average of 3.70 larvae plant⁻¹, encompassing developmental stages from newly hatched to sixth instars. By 5 DAT, S. frugiperda (J. E. Smith) larval counts remained at zero in plots treated with GS-SiNPs alone or in combination with ¾EMB bulk, whereas only minor infestations (0.10–0.20 larvae plant⁻¹) persisted in other treatments. Statistically significant differences (p ≤ 0.05) were evident between all treated plots and the control at both 1 and 5 DAT. At 14 DAT, larval numbers slightly increased, ranging from 0.10 to 0.80 larvae plant⁻¹ across treatments, except for the ½EMB-NPs + GS-SiNPs combination, which maintained 100% larval mortality (0.00 larvae plant⁻¹). Significant differences (p ≤ 0.05) were still evident between treated plots and the control. No significant differences (p > 0.05) were detected among GS-SiNPs, full-dose EMB-NPs, or their blends, nor among EMB bulk, ¾EMB bulk, ¾EMB-NPs, and ½EMB-NPs treatments.

Leaf damage scores per maize plant under Spodoptera frugiperda (J. E. Smith) infestation

As presented in Table 6, before the 1^st spray (1 DBT), leaf damage scores were statistically similar among all treatments, ranging from 0.63 to 0.74. One day after the 1^st spray (1 DAT), the leaf damage score declined in all treated plots due to S. frugiperda (J. E. Smith) larval mortality, while the control exhibited increase slightly to 0.81. Although reductions were observed in all treatments, these differences were not statistically significant (p < 0.05) compared with the control. At 5 DAT, leaf damage score declined markedly in all treated plots relative to the control. The most pronounced reduction was recorded with the ½EMB-NPs + GS-SiNPs treatment, corresponding to 64.2% of the control. Significant differences (p ≤ 0.05) were detected between all treated plots and the control; however, no significant variation was found among the treatments themselves. At 14 DAT, a slight increase in damage occurred in most treatments, yet all remained significantly lower than those of the control. The highest and lowest leaf damage scores were observed with the ¾EMB bulk (1.16), and EMB-NPs had the (0.63), representing 34.5% and 64.4% reductions, respectively, relative to the control value (1.77).

Table 6 Leaf damage scores of Zea mays L. treated with emamectin benzoate (EMB) in bulk and nano forms and/or green synthesized silicon nanoparticles (GS-SiNPs)-based treatments under field conditions during the 2024 autumn growing season.

Following the 2^nd spray, leaf damage severity exhibited a subsequent reduction at 24 h post-spray, ranging from 65.9% (EMB-NPs) to 36.3% (¾EMB bulk), relative to the control (Table 6). At 5 DAT, all applied treatments showed a pronounced reduction in leaf damage score compared with the control, reaching as low as 78.8% of the control value in the EMB-NPs, ¾EMB bulk + GS-SiNPs, and ¾EMB-NPs + GS-SiNPs treatments. At 14 DAT, leaf damage scores remained relatively low across all treatments, varying from 0.29 (¾EMB-NPs + GS-SiNPs) to 0.80 (¾EMB bulk), representing 86.4% and 62.4% reductions, respectively, relative to the control (2.13) (Fig. 3). Collectively, the ¾EMB-NPs + GS-SiNPs combination consistently produced the lowest leaf damage score, followed sequentially by the EMB-NPs, ¾EMB bulk + GS-SiNPs, and ½EMB-NPs + GS-SiNPs, whereas the ¾EMB bulk recorded the highest leaf damage score among the treated plots. Statistically, no significant disparity (p > 0.05) was observed among the EMB-NPs, ¾EMB bulk + GS-SiNPs, and ½EMB-NPs + GS-SiNPs treatments.

Fig. 3

Leaf morphology of Zea mays L. as influenced by ten green-synthesized SiO₂ nanoparticles with either bulk or nano-emamectin benzoate (T1-T10 as described in Table 2) during the 2024 season. T1: Control (only dH₂O); T2: GS-SiNPs, T3: full dose of EMB bulk; T4: ¾ full dose of EMB bulk; T5: full dose of EMB-NPs; T6: ¾ full dose of EMB-NPs; T7: ½ full dose of EMB-NPs; T8: ¾ full dose of EMB bulk + GS-SiNPs; T9: ½ full dose of EMB-NPs + GS-SiNPs; T10: ¾ full dose of EMB-NPs + GS-SiNPs. Bars represent 30 cm.

Leaf anatomical structure

Leaf anatomical characteristics of Zea mays displayed highly significant (p ≤ 0.01) differences among treatments, with all combined applications of EMB and GS-SiNPs outperforming individual treatments (Table 7 and Fig. 4A–J). The co-treatment of ¾EMB-NPs + GS-SiNPs achieved the greatest blade thickness (262.3 µm), midvein thickness (663.0 µm), and main vascular bundle (VB) diameter (220.2 µm), reflecting respective increases of 41.8%, 63.4%, and 20.5%, respectively, relative to the control. Similarly, the ½EMB-NPs + GS-SiNPs treatment produced the highest metaxylem (Mx) vessel diameter of main VB (82.01 µm with a 24.3% increase), while the ¾EMB bulk + GS-SiNPs combination maximized cuticle (17.67 µm with a 117.9% increase) and epidermis (33.0 µm with a 52.8% increase) thicknesses. Notably, several treatments, including GS-SiNPs alone, ¾EMB bulk + GS-SiNPs, and ¾EMB-NPs + GS-SiNPs formed statistically parallel groups for most parameters.

Table 7 Leaf anatomy structure of Zea mays L. treated with emamectin benzoate (EMB) in bulk and nano forms and/or green synthesized silicon nanoparticles (GS-SiNPs)-based treatments under field conditions during the 2024 autumn growing season.

Fig. 4

Transverse sections in leaf’s blade of Zea mays L. as influenced by ten green-synthesized SiO₂ nanoparticles with either bulk or nano-emamectin benzoate (T1-T10 as described in Table 2) during the 2024 season. (A) Control (only dH₂O); (B) GS-SiNPs, (C) full dose of EMB bulk; (D) ¾ full dose of EMB bulk; (E) full dose of EMB-NPs; (F) ¾ full dose of EMB-NPs; (G) ½ full dose of EMB-NPs; (H) ¾ full dose of EMB bulk + GS-SiNPs; (I) ½ full dose of EMB-NPs + GS-SiNPs; (J) ¾ full dose of EMB-NPs + GS-SiNPs. m = mesophyll; mvb = main vascular bundle; xv: xylem vessel. Bare represents 500 µm.

Photosynthetic traits, leaf area (LA) plant⁻¹, and leaf silicon (Si) content

Based on the results presented in Table 8, all treatments significantly (p ≤ 0.01) enhanced maize photosynthetic traits, LA plant⁻¹, and leaf Si content compared to the untreated, infested control. A pronounced synergistic effect was observed, particularly with the combined treatment of ¾EMB-NPs + GS-SiNPs consistently recorded the highest F_v/F_m (0.818), F_v/F_o (4.62), PPI_ABS (4.67), LA plant⁻¹ (7.68 dm²), and a leaf Si content of 24.5 mg g⁻¹ DW, representing respective increases of 2.3%, 22.9%, 43.3%, 32.0%, and 44.1%, respectively, relative to the control. The greatest enhancement in SPAD-CCI (48.6) was achieved by the GS-SiNPs treatment alone, resulting in a 14.9% increase compared with the control. Statistical parallelism was evident among ¾EMB bulk + GS-SiNPs, ½EMB-NPs + GS-SiNPs, and ¾EMB-NPs + GS-SiNPs for most photosynthetic traits and leaf Si content.

Table 8 Photosynthetic traits, leaf area plant⁻¹, and leaf silicon (Si) content of Zea mays L. treated with emamectin benzoate (EMB in bulk and nano forms) and/or green synthesized silicon nanoparticles (GS-SiNPs)-based treatments against invasive S. frugiperda (J. E. Smith) larvae during the 2024 autumn growing season.

Grain yield and its related attributes

Analysis of maize yield components (Table 9) revealed that, while cob length was not significantly (p = 0.0833) altered across treatments, all other yield-related attributes, i.e., cob diameter (Fig. 5), 100-GW, shelling percentage, and grain yield, were significantly (p ≤ 0.01) impacted. The combined application of ¾EMB-NPs + GS-SiNPs and EMB-NPs treatments resulted in the highest grain yield (6.83 and 6.67 t ha⁻¹), corresponding to 54.5% and 50.9% increases, respectively, over the control (4.42 t ha⁻¹). The highest cob diameter (4.12 cm) and shelling percentage (78.70%) were obtained under the ½EMB-NPs + GS-SiNPs treatment, which was statistically comparable to several other treatments. The latter represented 11.1% and 10.8% increases, respectively, relative to the control. Conversely, the greatest 100-GW values were observed with GS-SiNPs (46.40 g), followed by ¾EMB-NPs + GS-SiNPs (42.30), reflecting 25.1% and 14.0% increases over the control treatment.

Table 9 Grain yield and its attributes of Zea mays L. treated with emamectin benzoate (EMB in bulk and nano forms) and/or green synthesized silicon nanoparticles (GS-SiNPs)-based treatments against invasive S. frugiperda (J. E. Smith) larvae during the 2024 autumn growing season.

Fig. 5

Cob morphology of Zea mays L. as influenced by ten green-synthesized SiO₂ nanoparticles with either bulk or nano-emamectin benzoate (T1-T10 as described in Table 2) during the 2024 season. T1: Control (only dH2O); T2: GS-SiNPs, T3: full dose of EMB bulk; T4: ¾ full dose of EMB bulk; T5: full dose of EMB-NPs; T6: ¾ full dose of EMB-NPs; T7: ½ full dose of EMB-NPs; T8: ¾ full dose of EMB bulk + GS-SiNPs; T9: ½ full dose of EMB-NPs + GS-SiNPs; T10: ¾ full dose of EMB-NPs + GS-SiNPs. Bar represents 20 cm.

Relations

The 28-variable mean variables were analyzed to compute Pearson’s correlation coefficients and their significances (Fig. 6). The analysis included FAW larval counts plant⁻¹, leaf damage, leaf photosynthetic and anatomical traits, LA plant⁻¹, leaf Si content, yield and its related attributes, all of which were influenced by the EMB bulk or EMB-NPs, and/or GS-SiNPs-based treatments under field conditions. Positive correlations (p ≤ 0.05) were detected among selected physiological responses (F_v/F_o, F_v/F_m, PPI_ABS, leaf Si content, and SPAD-CCI), leaf anatomical (blade and midvein thickness, Mx vessel diameter of main VB, epidermal thickness, and cuticle thickness), and yield-related attributes (cob length and diameter, and 100-GW), and grain yield. In contrast, number of FAW larvae plant⁻¹ at 1 DAT in the 1^st spray and at 5 DAT in the 2^nd spray (NL1.1DAT and NL2.5DAT, respectively), as well as leaf damage recorded at 1, 5, and 14 DAT in the 1^st spray (LDS1.1DAT, LDS1.5DAT, and LDS1.14DAT, respectively) and at 1 and 5 DAT in the 2^nd spray (LDS2.1DAT and LDS2.5DAT, respectively), were negatively correlated (p ≤ 0.05) with the aforementioned photosynthetic and anatomical, and yield traits. Moreover, significant positive correlations (p ≤ 0.05) were observed among NL1.1DAT, NL2.5DAT, LDS1.1DAT, LDS1.5DAT, LDS1.14DAT, LDS2.1DAT, and LDS2.5DAT themselves.

Fig. 6

Correlogram of the correlation matrix for all studied traits related to the control of Spodoptera frugiperda (Lepidoptera: Noctuidae; J. E. Smith) in Zea mays L. The color hue of each circle represents the correlation direction and magnitude, whereas the circle size reflects the strength and significance of the relationship. An asterisk (*) denotes a significant correlation (p ≤ 0.05). NL1.1DAT, NL1.5DAT, and NL1.14DAT: number of FAW larval plant⁻¹ at 1, 5, and 14 DAT, respectively, in the 1^st spray, NL2.1DAT, NL2.5DAT, and NL2.14DAT: number of FAW larval plant⁻¹ at 1, 5, and 14 DAT, respectively, in the 2^nd spray, LDS1.1DAT, LDS1.5DAT, and LDS1.14DAT: leaf damage score at 1, 5, and 14 DAT, respectively, in the 1^st spray, LDS2.1DAT, LDS2.5DAT, and LDS2.14DAT: leaf damage score at 1, 5, and 14 DAT, respectively, in the 2^nd spray, BL-Th: blade thickness, Mid-Th: midvein thickness, M-VB-D: main vascular bundle diameter, MxV-Di-VD: metaxylem vessel diameter of main vascular bundle, Cut-Th: cuticle thickness, Epi-Th: epidermis thickness, F_v/F_m: maximum quantum yield of photosystem II, F_v/F_o: potential photosystem II activity, PPI_ABS: photosynthetic performance index on an absorption basis, SPAD-CCI: Soil Plant Analysis Development (SPAD)-chlorophyll content index, LA/P: leaf area plant⁻¹, LSiC: leaf silicon content, Cob-L: cob length, Cob-D: cob diameter, 100-GW: 100-grain weight, and GY: grain yield.

A heatmap with hierarchical analysis was performed and visualized to reveal the interactive connection between the EMB bulk or EMB-NPs, and/or GS-SiNPs-based treatments against invasive S. frugiperda (J. E. Smith) larvae (Fig. 7). The studied treatments (X-axis) were separated into four main groups using the hierarchical cluster analysis. Control (T1; only dH₂O) treatment was separated in the 1^st main group, while the 2^nd group included GS-SiNPs (T2), ¾EMB bulk + GS-SiNPs (T8), ½EMB-NPs + GS-SiNPs (T9), and ¾EMB-NPs + GS-SiNPs (T10). The 3^rd group included ¾EMB-NPs (T6) and ½EMB-NPs (T7), while the 4^th group included EMB bulk (T3), ¾EMB bulk (T4), and EMB-NPs (T5). Moreover, the Y-axis (selected parameters) were divided into three main groups (1, 2, and 3). Group 1 included F_v/F_o, F_v/F_m, PPI_ABS, and SPAD-CCI, thickness of blade, cuticle, and midvein, cob length and diameter, 100-GW, and grain yield. The group 2 included NL1.5DAT, NL1.14DAT, Mx vessel diameter of main VB, leaf Si content, epidermal thickness, shelling percentage, while group 3 included other remain parameters. These results highlighted that application of the EMB bulk or EMB-NPs, and/or GS-SiNPs-based treatments effectively improved leaf photosynthetic and anatomical traits, including the lignified tissues and affected the number of FAW larvae plant⁻¹, and grain yield under field conditions.

Fig. 7

Heatmap illustrating hierarchical clustering across all measured parameters and the ten tested treatment groups (T1-T10 as described in Table 2); the color scale denotes the corresponding Z-score values for each parameter. T1: Control (only dH2O); T2: GS-SiNPs, T3: full dose of EMB bulk; T4: ¾ full dose of EMB bulk; T5: full dose of EMB-NPs; T6: ¾ full dose of EMB-NPs; T7: ½ full dose of EMB-NPs; T8: ¾ full dose of EMB bulk + GS-SiNPs; T9: ½ full dose of EMB-NPs + GS-SiNPs; T10: ¾ full dose of EMB-NPs + GS-SiNPs. NL1.1DAT, NL1.5DAT, and NL1.14DAT: number of FAW larval plant⁻¹ at 1, 5, and 14 DAT, respectively, in the 1^st spray, NL2.1DAT, NL2.5DAT, and NL2.14DAT: number of FAW larval plant⁻¹ at 1, 5, and 14 DAT, respectively, in the 2^nd spray, LDS1.1DAT, LDS1.5DAT, and LDS1.14DAT: leaf damage score at 1, 5, and 14 DAT, respectively, in the 1^st spray, LDS2.1DAT, LDS2.5DAT, and LDS2.14DAT: leaf damage score at 1, 5, and 14 DAT, respectively, in the 2^nd spray, BL-Th: blade thickness, Mid-Th: midvein thickness, M-VB-D: main vascular bundle diameter, MxV-Di-VD: metaxylem vessel diameter of main vascular bundle, Cut-Th: cuticle thickness, Epi-Th: epidermis thickness, F_v/F_m: maximum quantum yield of photosystem II, F_v/F_o: potential photosystem II activity, PPI_ABS: photosynthetic performance index on an absorption basis, SPAD-CCI: Soil Plant Analysis Development (SPAD)-chlorophyll content index, LA/P: leaf area plant⁻¹, LSiC: leaf silicon content, Cob-L: cob length, Cob-D: cob diameter, 100-GW: 100-grain weight, and GY: grain yield.

Principal component analysis (PCA)-biplot was performed to explore the high variations resulted by the foliar application of the EMB bulk or EMB-NPs, and/or GS-SiNPs-based treatments on the studied maize variables. The PCA-dimensions, i.e., Dim 1 and Dim 2, respectively, explored 64.5% and 11.5% variability of data, respectively (Fig. 8). The elevated variability between the different treatments indicated the high variability in plant defense performance against S. frugiperda (J. E. Smith) larvae by application of the EMB bulk or EMB-NPs, and/or GS-SiNPs-based treatments. The ¾EMB-NPs + GS-SiNPs-treated maize plants under field conditions showed highly difference as compared to the control (T1; only dH₂O)-treated plants, while the difference was not high between T2, T9, and T10. Therefore, application of EMB bulk or EMB-NPs, and/or GS-SiNPs-based treatments has a distinct role in decreasing the number of S. frugiperda (J. E. Smith) larvae and improving the leaf photosynthetic and anatomical traits, and maize yield under field conditions.

Fig. 8

Biplot graph of studied traits and the ten tested treatment groups (T1-T10 as described in Table 2), illustrating the first two principal component analysis (PCA) dimensions (Dim1 and Dim2). T1; Control (only dH₂O), T2; GS-SiNPs, T3; EMB bulk, T4; ¾EMB bulk, T5; EMB-NPs, T6; ¾EMB-NPs, T7; ½EMB-NPs, T8; ¾EMB bulk + GS-SiNPs, T9; ½EMB-NPs + GS-SiNPs, T10; ¾EMB-NPs + GS-SiNPs. Each black dot refers to a treatment number. NL1.1DAT, NL1.5DAT, and NL1.14DAT: number of FAW larval plant⁻¹ at 1, 5, and 14 DAT, respectively, in the 1^st spray, NL2.1DAT, NL2.5DAT, and NL2.14DAT: number of FAW larval plant⁻¹ at 1, 5, and 14 DAT, respectively, in the 2^nd spray, LDS1.1DAT, LDS1.5DAT, and LDS1.14DAT: leaf damage score at 1, 5, and 14 DAT, respectively, in the 1^st spray, LDS2.1DAT, LDS2.5DAT, and LDS2.14DAT: leaf damage score at 1, 5, and 14 DAT, respectively, in the 2^nd spray, BL-Th: blade thickness, Mid-Th: midvein thickness, M-VB-D: main vascular bundle diameter, MxV-Di-VD: metaxylem vessel diameter of main vascular bundle, Cut-Th: cuticle thickness, Epi-Th: epidermis thickness, F_v/F_m: maximum quantum yield of photosystem II, F_v/F_o: potential photosystem II activity, PPI_ABS: photosynthetic performance index on an absorption basis, SPAD-CCI: Soil Plant Analysis Development (SPAD)-chlorophyll content index, LA/P: leaf area plant⁻¹, LSiC: leaf silicon content, Cob-L: cob length, Cob-D: cob diameter, 100-GW: 100-grain weight, and GY: grain yield.

Discussion

The S. frugiperda (J. E. Smith) represents one of the most destructive lepidopteran pests worldwide, owing to its ability to infest more than 350 plant species, predominantly maize^20,21. Due to its high fecundity, strong migratory potential, and rapid adaptation to several insecticide classes, conventional chemical control has become increasingly ineffective^34,78. The present study provides mechanistic insights that combining GS-SiNPs with EMB or its nanoform (EMB-NPs) represents an effective eco-nanotechnological tactic against S. frugiperda (J. E. Smith). Combined laboratory-field evaluations revealed a dual-mode suppression mechanism, direct larvicidal biochemical toxicity, and Si-mediated strengthening of maize’s physio-structural resilience, resulting in superior crop protection and productivity.

The small particle size observed in the GS-SiNPs is likely influenced by the reducing and stabilizing constituents in the P. odoratissimum L. leaf extract. It is important to clearly distinguish the green synthesis approach employed in this study from methods based on the extraction of biogenic silica from Si-accumulating biomass, such as rice straw or wheat residues. While such sources yield amorphous silica, the P. odoratissimum L. extract here functions as a bioactive reducing and capping matrix for the sodium metasilicate precursor⁷⁹. A key advantage of P. odoratissimum L. lies in its high abundance of specific phenolic and flavonoid compounds, which promote rapid silicate ion reduction and immediate nanoparticle stabilization. This phytochemical capping effectively suppresses agglomeration⁸⁰, resulting in highly stable, spherical GS-SiNPs with uniform morphology and offering a more controlled and reproducible synthesis route than the inherently variable extraction of silica from crop residues. The acute toxicity bioassay revealed a pronounced larvicidal potency gradient, as LC₅₀ value indicated that EMB-NPs was about tenfold higher larvicidal toxicity than GS-SiNPs and outperformed EMB bulk about 1.7-fold. Therefore, EMB-NPs demonstrated its excellent efficacy against S. frugiperda (J. E. Smith) larvae⁸¹. Furthermore, the larvicidal activity of EMB bulk or EMB-NPs enhanced when tiny quantities of it combined with GS-SiNPs, which contributed to decreasing the use of pesticides³². This enhanced potency can be attributed to nanostructural superiority, greater surface-to-volume ratio, optimized cellular penetration, and controlled release kinetics, which intensify interaction with the insect neurophysiological system⁸². EMB exerts its toxic action by affinity for GluCl channels, triggering unregulated Cl⁻ influx, leading to neuromuscular paralysis²⁴. In nanoparticulate form, the nanoscale size facilitates efficient epidermal penetration and uniform diffusion through the leaf tissue, maintaining persistent larval exposure and higher contact toxicity^39,83.

The relatively sharp slope of the GS-SiNPs dose-toxicity curve indicates a high concentration lethality sensitivity, implying that minimal nanoparticle concentration increments substantially enhance lethality⁸⁴. This finding concurs with the established biophysical action of silica nanoparticles, cuticular physical abrasion and lipid layer perturbation, resulting in dehydration and cuticular dysfunction^85,86. Such functionality disruption may also facilitate EMB diffusion through larval integuments, explaining the synergistic improvement observed when the GS-SiNPs + EMB-NPs are co-applied^41,82. The current study’s findings also proved that the synergistic effect of GS-SiNPs and EMB was induced by the inhibition of detoxifying enzymes in FAW. Thus, taken GS-SiNPs, EMB-NPs and EMB-bulk, as well as their combinations safe, and efficient insecticidal strategy against FAW larvae. The GS-SiNPs induce cuticular failure through a nano-abrasive mechanism governed by their geometry, surface reactivity, and phytochemical capping⁸⁷. Particle accumulation at epicuticular folds promotes localized scraping of the wax-lipid barrier, followed by chitin-protein micro-fracturing. These structural defects enhance lipid extraction and water efflux, ultimately compromising integumental integrity and permeability⁸⁸.

The synergistic interaction observed between GS-SiNPs and EMB-NPs suggests a dual mode of action targeting both the physical integrity and metabolic defenses of S. frugiperda (J. E. Smith). Structurally, GS-SiNPs abrade the insect midgut epithelium and cuticle; this functionality disruption increases permeability and facilitates rapid EMB diffusion through larval integuments into the hemolymph^41,82,89. Consequently, this physical damage constitutes a standalone stressor that potentiates EMB bioavailability, accelerating and homogenizing its neurotoxic action^36,90. This structural compromise is further amplified by metabolic interference, as our bioassays (Table 4) reveal that co-application significantly suppresses G-S-T and TE, which were otherwise elevated in single treatments. We propose that this synergistic suppression stems from a physiological trade-off induced by the nanoparticles. The abrasive damage caused by GS-SiNPs to the larval midgut likely forces a reallocation of metabolic energy toward urgent cellular repair and osmoregulation, thereby limiting the resources available for synthesizing energetically costly detoxification enzymes. Furthermore, the suppression may result from direct catalytic interference, where the GS-SiNPs’s high surface area and reactivity⁸² generate oxidative stress or facilitate protein corona formation that structurally denatures or sterically hinders the active sites of G-S-T and TE. Consequently, the GS-SiNPs do not merely deliver the insecticide but actively dismantle the pest’s biochemical capacity to metabolize it, rendering the larvae hypersensitive to the EMB payload³².

The enzymatic activity profile offers a biochemical synergy between GS-SiNPs and EMB. Larvae exposed to EMB (bulk or nano) alone exhibited enhanced G-S-T and TE activities, indicating an induced detoxification response to xenobiotic stress⁹¹. In contrast, co-treatment with GS-SiNPs significantly suppressed these enzymatic activities^92,93. This inhibition is plausibly attributed to the strong binding affinity of Si-based nanoparticles to sulfhydryl or carboxyl functional groups within catalytic sites, inducing conformational perturbations that compromise catalytic function^41,94. Moreover, silica nanoparticle-induced oxidative stress may disturb redox homeostasis and deplete intracellular reduced glutathione, thus limiting G-S-T-mediated detoxification⁸⁸. Consequently, the suppressed detoxification capacity enhances EMB bioavailability and toxic persistence in larval tissues, thereby amplifying lethality⁹⁵. Comparable biochemical responses have been reported for abamectin- or chlorpyrifos-loaded mesoporous SiO₂ nanoparticles, corroborating their synergistic pesticidal efficiency³⁵.

The field experiment validated the laboratory results, demonstrating that all tested formulations achieved 100% larval lethality within 24 h post-spraying, while control remained infested. The transient resurgence observed after 5 and 14 days coincides with with EMB’s limited environmental stability under open-field conditions, attributed to photodegradation and rapid microbial breakdown^96,97. Notably, maize plants treated with GS-SiNPs, alone or in mixtures, maintained suppressed larval populations beyond EMB’s degradation period, indicating Si’s residual protective role⁴¹. Silicon deposition within leaf epidermal tissues forms a silicified bilayer, phytolith barriers that reinforce leaf rigidity, decrease palatability, and erode larval mandibles^42,98. Moreover, Si primes maize defensive metabolism through jasmonic-acid-dependent signaling, stimulating phenolics, lignin, and defense-related proteins that suppress herbivory^38,42. The persistent reduction in larval counts at 14 d post-spraying therefore denotes both residual Si fortification and possible induced systemic resistance effects rather than direct chemical toxicity^8,99.

The pronounced difference in larval suppression sustainability between the 1^st and 2^nd sprays highlights the cumulative nature of the nano-fortification strategy. Following the 1^st application (14 DAP) at the seedling stage, the rapid expansion of leaf surface area likely led to a dilution effect of the applied nanoparticles, rendering the physical barrier less permanent and allowing for larval resurgence by 14 days post-treatment⁹⁸. Conversely, the 2^nd application (33 DAP) appeared to benefit from the residual Si deposited by the 1^st spray, creating a cumulative hardening effect⁴¹. This hypothesis is substantiated by the significant elevation in leaf Si content (up to 24.5 mg g⁻¹ DW; Table 8) observed in the treated plants. We postulate that this progressive Si accumulation formed a dense, abrasive physical barrier on the leaf epidermis that was impenetrable to early-instar larvae during the re-infestation waves³⁸, thereby maintaining population suppression significantly longer than the initial application.

Microscopic observations corroborate the defensive/protective function of Si integration. The co-treatment of ¾EMB-NPs + GS-SiNPs significantly increased blade and midvein thickness, vascular bundle diameter, and cuticle and epidermal layers, morphological adaptations that restrict larval penetration and and reinforce structural sturdiness^8,100. Silicon deposits as amorphous silica (SiO₂) within epidermal cell walls and vascular tissues, strengthening mechanical fortitude and minimizing tissue deformation under chewing and piercing-sucking^42,101,102. Concurrently, thicker cuticle and epidermis reduce transpirational water loss, maintaining cellular turgidity and optimizing gas exchange during larva-induced stress¹⁰³.

These leaf anatomical modifications resulted in enhanced photosynthetic functionality. The enhanced F_v/F_m, F_v/F_o, and PPI_ABS ratios indicate improved photochemical efficiency and electron transport capacity, likely due to the preserved ultrastructural integrity of chloroplast under Si-mediated protection^104,105. Silicon is known to stabilize thylakoid membranes and alleviate photooxidative damage through modulating reactive oxygen species (ROS) detoxification¹⁰⁶. Furthermore, increased SPAD-CCI and LA plant⁻¹ confirm enhanced chlorophyll retention and sustained an active photosynthetic canopy, essential for compensating biomass losses caused by S. frugiperda’s larvae feeding. The notable increase in leaf Si under mixture treatments implies efficient absorption and deposition, correlating nanoparticle internalization to improve structural–functional integrity^100,107.

The resulting physiological improvements culminated in significant gains in cob diameter, grain weight, shelling efficiency, and total grain yield. Specifically, the ¾EMB-NPs + GS-SiNPs treatment yielded a 54.5% yield advantage over the control, matching the efficacy of lower EMB-NPs doses when co-applied with GS-SiNPs. This parity demonstrates that coupling GS-SiNPs allows dose minimization without efficacy loss, an economically and environmentally advantageous outcome^32,36. Silicon’s role in reinforcing sink capacity and photosynthate partitioning likely accounts for the higher kernel weight and shelling percentage^37,108. Simultaneously, sustained pest suppression through combining biochemical and structural barriers minimized metabolic diversion for tissue repair, ensuring prioritized resource allocation toward yield formation⁹².

Overall, the results integrated a multi-tiered mechanism wherein GS-SiNPs and EMB-NPs act cooperatively across biochemical, physio-anatomical, and ecological levels. Biochemically, GS-SiNPs compromise larval detoxification enzymes like G-S-T and TE, thereby magnifying EMB-induced neurotoxicity. Physiologically, Si deposition within maize tissues reinforces photosynthetic efficiency and alleviates oxidative stress by chloroplast stabilization and enhancing antioxidant defense system. Anatomically, reinforced cuticular layers, epidermal thickness, and vascular integrity, collectively as mechanical barriers, hinder larval invasion and feeding activity. Ecologically, these combined effects maintain suppressed pest populations and reduce canopy damage across consecutive spray cycles¹⁰⁹, ultimately restoring crop productivity. This mechanistic sequence emphasizes a synergistic interplay whereby GS-SiNPs act concurrently as nanocarriers facilitating EMB delivery and as a benificial micronutrient that trigger plant-mediated resistance^41,82,110. This dual-mode defense integrates bottom-up (plant-induced)⁸⁴ and top-down (insecticidal)¹¹¹ control of S. frugiperda (J. E. Smith), establishing an emergent trend in nano-enabled integrated pest management (Fig. 9).

Fig. 9

Multi-tiered mechanism of green-synthesized SiO₂ nanoparticles (GS-SiNPs) integrating direct larvicidal action with emamectin benzoate (EMB) and indirect plant-mediated resistance for enhanced pest management and crop productivity. G-S-T = glutathione S-transferase.

Broader implications, future perspectives, and key limitations

The observed synergistic between EMB-NPs and GS-SiNPs offers implications extend beyond conventional control management of S. frugiperda (J. E. Smith). It exemplifies how nanotechnology can harmonize pest control efficacy with environmental safety via targeted delivery, lower dosages, and induction of endogenous plant defenses. To further substantiate the environmental safety of the applied nanomaterials, the assessment was conducted in alignment with established international regulatory frameworks. Specifically, toxicity evaluations followed the OECD Guidelines for the Testing of Chemicals¹¹², Material Safety Data Sheet (MSDS) standards for nanomaterial hazards, and emerging pesticide ecotoxicity risk assessment methodologies¹¹³. Collectively, these frameworks provide a standardized basis for confirming that non-target toxicity remains within acceptable limits and support that GS-SiNPs and EMB-NPs pose a lower environmental risk than conventional high-dose chemical pesticide applications. However, advancing this strategy toward large-scale adoption requires multi-seasonal field studies to validate long-term stability of these formulations, Si-pesticide interactions in soil–plant systems, and potential effects on non-target organisms. Coupling nanoparticle toxicodynamics with transcriptomic and metabolomic analyses may elucidate molecular interplay between Si signaling and larva detoxification processes. Such mechanistic insight will be crucial for optimizing nanopesticide formulations compatible with sustainable agriculture practices. Despite promising outcomes, mechanistic interpretations regarding enzyme inhibition and Si-mediated defense enhancement in this study were based on biochemical and anatomical indicators rather than validated via direct molecular or transcriptomic analysis. Additionally, nanoparticle stability, persistence, and potential trophic transfer within the soil–plant-insect continuum were not assessed, leaving gaps regarding long-term ecological safety and environmental behavior. Finally, although synergistic effects between GS-SiNPs and EMB-NPs were evident, the optimal nanoparticle ratios, application regimes, and potential non-target organisms remain to be systematically evaluated through multi-location studies and comprehensive omics analyses.

Furthermore, with respect to the environmental safety and potential phytotoxicity of the applied nanomaterials, the observed photosynthetic performance provides a reliable proxy for overall plant health. The significant enhancement in F_v/F_m and PPI_ABS under the ¾EMB-NPs + GS-SiNPs treatment (Table 8) indicates the absence of oxidative stress typically associated with phytotoxic bioaccumulation¹⁰⁶. In contrast to synthetic bulk compounds, which may induce ROS accumulation due to slow metabolic turnover, GS-SiNPs have been reported to function as nano-scavengers or elicitors that enhance the plant’s total antioxidant capacity¹¹⁴. Previous studies have demonstrated that mesoporous silica nanoparticles degrade into non-toxic Si(OH)₄, a bioavailable form readily assimilated by plants to reinforce cell wall structure without adverse bioaccumulation or soil persistence¹¹⁵. Accordingly, the observed improvements in maize morpho-physiological traits confirm that the applied nano-formulations are cytocompatible and contribute to mitigating the oxidative pressure, posing minimal ecological risk relative to conventional bulk pesticide applications.

Conclusion

In conclusion, the integration of GS-SiNPs with EMB-NPs establishes a potent and environmentally adaptive strategy for suppression of S. frugiperda (J. E. Smith) in maize fields. This functional dual-action strategy merges enhanced insecticidal toxicity and reinforced plant defense system. The synergistic interaction impairs larval detoxification pathways, reinforcement of photosynthetic and structural integrity, and sustains field-level pest suppression, ultimately restoring grain yield. These findings highlight the practical potential of nano-enabled formulations as sustainable alternatives to conventional chemical control. This approach effectively minimizes required insecticide loads while simultaneously maximizing control efficacy. Therefore, field implementation should prioritize optimizing nanoformulation ratios and spray intervals. This optimization will harmonize insecticidal bioefficacy with inherent plant health benefits, advancing the integration of GS-SiNPs and EMB-NPs into future nanobiotechnological integrated pest management program frameworks. Based on field evaluation of ten treatments, the co-application of ¾EMB-NPs (144.0 g ha⁻¹) with GS-SiNPs (177.98 g ha⁻¹) was identified as the optimal treatment. This combination achieved S. frugiperda (J. E. Smith) larval lethality comparable to full-dose applications while increasing grain yield by 54.5% via Si-mediated enhancement. Accordingly, this reduced-dose nano-enabled formulation is recommended for maize production, offering an effective balance between agronomic performance, reduced chemical input, and economic sustainability under open-field conditions.