Plant diseases are commonly managed using crop protection chemicals, including fungicides, antibiotics, and insecticides. Excessive use of these chemicals adversely affects soil health, the environment, and human health. Utilizing crop protection chemicals at the nanoscale level addresses these issues. Nanotechnology is a multidisciplinary field with broad applications in the fields of science and technology. Silver (Ag) nanoparticles were the first to be investigated for plant disease management. Studies by Park et al. (2016), Lamsal et al. (2011), and Kim et al. (2008) examined the early use of silver nanoparticles in controlling powdery mildew disease in various crops. However, chemically synthesized nanoparticles can increase particle reactivity and toxicity, which may lead to negative effects on plant and human health and the environment through the breakdown of chemical groups and the formation of by-products (Meena et al., 2021).

Many biocontrol agents are widely used for the management of fungal and bacterial diseases, as these agents produce antifungal and antibacterial metabolites, which, in turn, control plant diseases (Vinodkumar et al., 2018; Basavarajappa et al., 2023). However, most biocontrol agents are non-specific and adversely affect beneficial microbes present in the soil. Beneficial endophytic and antagonistic microorganisms have been widely explored in the management of crop diseases (Johnson et al., 2014; Gill et al., 2016). The promising endophytic and antagonistic fungi or bacteria produce antimicrobial metabolites having either antifungal or antibacterial properties (Vinodkumar et al., 2018). Green synthesis of nanoparticles of antimicrobial compounds derived from potential endophytic and antagonistic microbes offers a sustainable approach for managing plant diseases because they are non-toxic, environmentally safe, effective at low dosage, target multiple fungal or bacterial pathogens, and safe for the beneficial organisms in the crop niche.

Tomato (Lycopersicon esculentum Mill.) is a widely grown vegetable that is rich in vitamins A, E, and C, as well as calcium, niacin, and organic acids, and has a high water content (Aslam et al., 2017). Due to their short growth period and high profitability, tomatoes are commercially used in the production of various food products (Karthika et al., 2020). As a result, there is an increasing demand for improving production techniques, fruit quality, yield, storage methods, and effective disease and pest management practices. Tomatoes are highly susceptible to various oomycetes, fungal, bacterial, and viral diseases, resulting in a substantial yield reduction and decreased nutritional value. Common fungal diseases affecting tomato cultivation include damping off, Phytophthora root rot, Fusarium wilt, and anthracnose (Singh et al., 2017). Among bacterial diseases, wilt and spot are particularly destructive, accounting for approximately 90% of the yield losses (Huang et al., 2013). Additionally, tomato spotted wilt virus and tomato leaf curl New Delhi virus pose a high risk to tomatoes, with yield losses ranging from 70 to 95% (Ong et al., 2020). The above diseases are managed by the regular use of fungicides or antibiotics. Insecticides are used to control vectors that transmit viral diseases. Moreover, the development of fungicide and antibiotic resistance adds further challenges to management. Therefore, nanopesticides offer an advanced solution for controlling various diseases in crop plants, as the quantity required is low.

Among metal nanoparticles, zinc nanomaterials are cost-effective, less phytotoxic, environmentally safe, and exhibit antimicrobial activity against phytopathogenic fungi (Yehia and Ahmed, 2013; Zabrieski et al., 2015; De La Rosa-García et al., 2018; Dos Santos et al., 2019; Kumawat et al., 2025), bacteria (Almoudi et al., 2018; Khan and Siddiqui, 2018; Rashid et al., 2024), viruses (Cai et al., 2019), and algae (Qureshi et al., 2018). Moreover, zinc is a crucial micronutrient for plant growth, carbohydrate metabolism, and the regulation of gene expression linked to biotic and abiotic stresses (Sabir et al., 2014). Therefore, zinc oxide nanoparticles have gained growing interest in agriculture because they are recognized as safe by the United States Food and Drug Administration (USFDA; FDA, 2015).

Green synthesis of nanoparticles can be performed using derivatives of plants as well as microorganisms such as fungi, bacteria, actinobacteria, yeasts, molds, and algae. Biomolecules found in plants or microorganisms, such as proteins, enzymes, phenolic compounds, amines, alkaloids, and pigments, serve as reducing agents in the synthesis of nanoparticles (Nadaroglu et al., 2017). In recent years, endophytes have gained importance in sustainable agriculture because of their unique ability to colonize plant tissues without causing disease. Besides being non-pathogenic, endophytes promote plant health and protect plants from biotic and abiotic stresses through the production of bioactive metabolites, improvement of nutrient availability, and modulation of the plant immune responses (Johnson et al., 2018, 2019; Khan et al., 2025). Endophytic bacteria are widely used for nanoparticle synthesis because of their ability to reduce metal toxicity (Korbekandi et al., 2009). They transform metals into nanoparticles through the activity of cellular enzymes and secondary metabolites (Joshi et al., 2017; Meena et al., 2021). This study highlights the simultaneous broad-spectrum antimicrobial activity of WDE and Gs-ZnO-NPs of the promising endophytic and antagonistic Ba-8SE-IF1 against tomato pathogens, including P. aphanidermatum (Edson), P. nicotianae (Breda de Haan), C. gloeosporioides (Penzig.), F. oxysporum f. sp. lycopersici (Sacc.), R. solanacearum (Smith), X. campestris (Dowson), TSWV, and ToLCNDV. GC-MS/MS analysis of the WDE of Ba-8SE-IF1 identified 26 major organic compounds with antioomycetes, antifungal, antibacterial, and antiviral properties. Gs-ZnO-NPs of Ba-8SE-IF1-WDM significantly inhibited the above pathogens, and the endophyte promotes the growth of tomato seedlings.

The present study focused on the extraction and identification of antimicrobial compounds from WDE of a promising endophytic and antagonistic bacterium, B. amyloliquefaciens 8SE-IF1; green synthesis of zinc oxide nanoparticles using WDM of Ba-8SE-IF1; and evaluation of antimicrobial activity of Ba-8SE-IF1-WDE and its Gs-ZnO-NPs against different phytopathogenic oomycetes, fungi, bacteria, and viruses infecting tomato plants. WDE of Ba-8SE-IF1 exhibited significant broad-spectrum antimicrobial activities against P. aphanidermatum, P. nicotianae, F. oxysporum f. sp. lycopersici, C. gloeosporioides, R. solanacearum, and X. campestris and also resulted in remission of symptoms caused by TSWV and ToLCNDV. The inhibition of these phytopathogens indicates that the WDE of Ba-8SE-IF1 contains multiple antimicrobial compounds. GC-MS/MS analysis revealed that WDE contains at least 26 major organic compounds and has antifungal, antibacterial, antiviral, or simultaneous antimicrobial properties. The identified antimicrobial compounds have functional groups such as phenols, carboxylic acids, alcohols, carbonyl groups of aldehydes, and aliphatic hydrocarbons. Moreover, five compounds, viz., phenol 3,5-bis (1,1-dimethyl-ethyl), hexadecane, 1-tetradecene, 2,6,10,14-tetramethyl hexadecane, 2,6,11, and 15-tetramethyl hexadecane, have antifungal, antibacterial, antiviral, and antimicrobial properties. These abilities enabled endophytic bacteria to significantly inhibit the growth of phytopathogenic oomycetes, fungi, and bacteria, as well as reduce the symptoms produced by TSWV and ToLCNDV. This is the premier study that reports simultaneous inhibition of a multitude of phytopathogens belonging to different kingdoms by WDE of an endophytic and antagonistic bacterium in crop plants. However, the antimicrobial properties of culture filtrates of antagonistic bacteria have been reported against either phytopathogenic fungi or bacteria. A 40% culture filtrate of Bacillus sp. B44 inhibited the growth of F. oxysporum f. sp. lycopersici to 70% (Jangir et al., 2018), a 50% culture filtrate of B. licheniformis suppressed the growth of R. solani, C. gloeosporioides, and P. capsici (Jeong et al., 2017), and the culture filtrate of Pseudomonas aeruginosa Os_12 impeded F. oxysporum f. sp. pisi (Gupta et al., 2022). The culture filtrates of Pseudomonas kilonensis Ba35 and Serratia liquefaciens Ou55 inhibited Agrobacterium tumifaciens (Etminani et al., 2024), and volatile compounds from Bacillus strain D13 reduced the growth of X. oryzae pv. oryzae (Xie et al., 2018). The fatty acids from B. amyloliquefaciens VB7 had antiviral activity against the tobacco streak virus in cotton (Vinodkumar et al., 2018).

GC-MS/MS analysis also revealed the presence of major functional groups, including phenols, carboxylic acids, alcohols, aldehydes (carbonyl compounds), and aliphatic hydrocarbons (alkanes) in the Gs-ZnO-NPs of WDM of the promising endophytic bacterial strain. Phenols and carboxylic acid compounds are known to possess antifungal properties by directly interacting with fungal cell membranes. These compounds integrate into the lipid bilayer, increasing membrane fluidity and leading to disorganization and eventual cell lysis (Avis and Bélanger, 2001). The phenolic compound, phenol 2,4-bis (1,1-dimethylethyl), exhibited antifungal activity against Alternaria solani and Botrytis cinerea by disrupting membranes, inhibiting lipid peroxidation, and inducing cell death (Gao et al., 2017). Hexadecanoic acid is fungistatic and targets the fungal cell wall and interferes with ergosterol biosynthesis in F. oxysporum, A. solani, and C. lagenarium (Liu et al., 2008). The aliphatic hydrocarbons primarily exhibit antifungal activity by inhibiting spore germination (Prakash and Arora, 2021) and causing mycelial disruption and distortion (Muhialdin et al., 2020). Similarly, alcohols and aldehydes exhibit antibacterial activity by inducing potassium ion (K⁺) leakage from bacterial cells, resulting in membrane disruption and structural damage (Togashi et al., 2007). Oleic and hexadecanoic acids disrupt the membranes of Pseudomonas syringae, R. solanacearum, and X. campestris, resulting in increased permeability, oxidative stress, and metabolic disruption (Sohn et al., 2013; Idris, 2022), and interrupt the entry and movement of tobacco mosaic virus by altering the lipid makeup of the host cell membrane (Zhao et al., 2017). Pentadecenoic, heptadecenoic, and octadecenoic acids produced by B. amyloliquefaciens VB7 synergistically suppress tobacco streak virus replication in cotton (Vinodkumar et al., 2018). However, the antiviral mechanisms of these compounds remain unclear.

The green synthesis of ZnO nanoparticles was confirmed by the immediate color change of the reaction mixtures after the incubation period. A yellowish-brown color was observed, while green-synthesizing nanoparticles of Ba-8SE-IF1-WDM were observed. Similarly, white color changes were observed in the green synthesis of zinc nanoparticles of Pseudomonas fluorescens (Vinay et al., 2018). The color changes observed in aqueous solutions are attributed to the plasmon resonance phenomenon. This process is involved in the reduction of zinc metallic salts into nanoparticles and is facilitated by the oxidation of aldehyde groups in biomolecules into carboxylic acids (Shameli et al., 2012). The peak of UV-Vis absorption observed at 387 nm for the Gs-ZnO-NPs of Ba-8SE-IF1-WDM confirms the formation of biologically active monodisperse ZnO-NPs in the solution. The observed absorption peak is due to the surface plasmon resonance directly linked to the particular size, shape, and composition of the solution used in the nanoparticle synthesis. Comparable absorption spectra were observed for green-synthesized zinc oxide nanoparticles of Cinnamomum camphora leaf extracts at 368–374 nm (Zhu et al., 2021) and Bacillus cereus RNT6 at 382 nm (Ahmed et al., 2021).

The stability of the Gs-ZnO-NPs of Ba-8SE-IF1-WDM was assessed based on its zeta potential. It resulted in −33.1 mV, which demonstrated the stability of the antimicrobial metabolites in the Gs-NPs. A high magnitude of either positive or negative zeta potential indicates strong electrostatic repulsion between the Gs-NPs, which prevents their aggregation and ensures long-term stability. Moreover, this property ensures the consistent biological activity of the Gs-NPs in inhibiting phytopathogens and the symptoms produced by them, in addition to growth promotion in crop plants. Singh et al. (2019) reported a zeta potential of −17.1 mV for ZnO-NPs of the leaf extract of Punica granatum L.

The FTIR spectrum of the GS-ZnO-NPs confirms the presence of O-H stretching in alcohols and phenolic compounds, in addition to carbonyl (C=O) groups in the antimicrobial compounds present in the water-diffusible metabolites of Ba-8SE-IF1. Similarly, the FTIR spectrum shows the presence of O-H and N-H stretching of aliphatic primary amines, broad O-H stretching, and amide I b and C-N stretching of amines in the biologically active Gs-ZnO-NPs. These properties of other Gs-NPs were reported by Fouda et al. (2018, 2020), Mohamed et al. (2019), and Mohamed et al. (2021). X-ray diffraction of Gs-ZnO-NPs of Ba-8SE-IF1-WDM reinstated its crystalline structure, whereas the sharp peak confirmed its purity and crystallinity.

FE-SEM analysis revealed that the Gs-ZnO-NPs exhibited an oval to spherical shape with a size of 59.94 nm. The spherical form of nanoparticles indicates the minimization of the total surface energy, isotropic growth conditions (even distribution of reducing agents), and uniform encapsulation of biomolecules, which in turn enhances their biological properties and performance. Comparable results have been reported in the shape of other green-synthesized ZnO nanoparticles, i.e., spherical particles of 21–35 nm (Ahmed et al., 2021), irregular particles of ~100 nm (Mohamed et al., 2019), and hexagonal particles with an average size of 50–90 nm (Gupta et al., 2018).

The Gs-ZnO-NPs exhibited substantially high antimicrobial activity against all the tested phytopathogens, underscoring their role in enhancing different biological functions. These Gs-NPs interact electrostatically with microbial membranes to facilitate their cellular uptake. They generate redox-active electrons to produce reactive oxygen species (ROS), which in turn damage key cellular components such as proteins, lipids, enzymes, and nucleic acids of phytopathogenic microbes (Sánchez-López et al., 2020). Additionally, NPs interfere with protein function, disrupt membrane integrity, alter cell morphology, and induce cytoplasmic leakage (Díez-Pascual, 2018; Kalia et al., 2020). The Gs-ZnO-NPs have a lower risk of resistance development due to their multiple mechanisms of action against plant pathogens, as they contain at least 26 major antimicrobial compounds. This contrasts with commercial crop protection chemicals, which typically have a single mode of action, making it easier for pathogens to adapt and develop resistance over time. There are very limited reports on the antimicrobial activity of Gs-ZnO-NPs from beneficial microbes against multiple phytopathogens. However, plant-based Gs-ZnO-NPs were developed and reported to inhibit plant pathogens, viz., Alternaria mali, Botryosphaeria dothidea, and Diplodia seriata (Ahmad et al., 2020); Alternaria alternata (Zhu et al., 2021); Aspergillus niger (Kumawat et al., 2025); X. oryzae pv. oryzae (Ogunyemi et al., 2019); R. solanacearum, Erwinia carotovora, and Clavibacter michiganensis (Rashid et al., 2024); and tobacco mosaic virus (Abdelkhalek and Al-Askar, 2020).

Tomato seedlings inoculated with R. solanacearum showed a high bacterial wilt incidence of 60.3%, whereas Gs-ZnO-NPs of Ba-8SE-IF1-WDM-treated plants showed an enhanced tolerance to R. solanacearum, resulting in a lower incidence of 9.9%. The observed reduction in disease incidence is attributed to the antibacterial properties of the Gs-ZnO-NPs of Ba-8SE-IF1-WDM. 1-hexadecanol, 4,6-dimethyl dodecane, 5-methyl tetradecane, 2,6,10,15-tetramethyl heptadecane, heneicosane, and n-tetradecane present in Gs-NPs are reported to inhibit bacterial proliferation by disrupting tolerance mechanisms, impairing transmembrane proton translocation, and suppressing glycolytic activity (Applerot et al., 2009; Reddy et al., 2007). Interestingly, Gs-ZnO-NPs did not show any inhibitory effects on commonly used biocontrol agents such as P. fluorescens and Trichoderma spp. (data not shown).

The commercial fungicides (metalaxyl for oomycetes and carbendazim for fungi at 100 ppm) exhibited antifungal activity against F. oxysporum f. sp. lycopersici (47.7%), followed by C. gloeosporioides (36.6%), P. nicotianae (32.2%), and P. aphanidermatum (19.44%) in the poisoned food assay. Streptocycline (100 ppm) showed antibacterial activity against R. solanacearum and X. campestris, with inhibition zones of 13.25 mm and 11.47 mm, respectively, in the agar well method. Under in vivo conditions, streptocycline (100 ppm)-treated plants showed green wilt symptoms, with a disease incidence of 20.8%. However, WDE and Gs-NPs outperformed the commercial fungicides and antibiotics used for the management of fungal and bacterial diseases in tomatoes.

Furthermore, the biometric parameters were significantly improved in tomato plants treated with Gs-ZnO-NPs of Ba-8SE-IF1-WDM. There was a 35 to 100% increase in the different biometric parameters of vegetative growth compared to the control plants. It also showed at least a 25–90% improvement in the vegetative parameters compared to either metalaxyl- or carbendazim-treated plants (data not shown). Zinc plays a crucial role as a micronutrient in plant growth and development, functioning as a cofactor for numerous enzymes involved in protein synthesis, auxin metabolism, and cell division. However, zinc applied through conventional fertilizers is often immobilized in the soil, limiting its availability to plants (Sharma et al., 2022). The nanoscale formulation of the Gs-ZnO-NPs of WDM enhances zinc solubility and bioavailability, thus enabling its efficient uptake and translocation within plant tissues. This improved zinc nutrition is probably responsible for the observed stimulation of vegetative growth.

To summarize, this study clearly demonstrates that WDE of Ba-8SE-IF1 and its Gs-ZnO-NPs exhibited synchronized antioomycete, antifungal, antibacterial, and antiviral activities against an array of phytopathogens in addition to growth-promoting effects in tomato plants. WDE and its Gs-ZnO-NPs have at least 26 major compounds that are responsible for their antimicrobial properties. Thus, they are highly promising candidates for the development of nanopesticide formulations. The experimental results should be validated under multi-location field trials in different agroecological units. Additional studies are needed to understand the complex interactions between green-synthesized nanoparticles and tomato plants under diverse agroecological conditions. In addition, the mechanism underlying the plant growth-promoting potential of green-synthesized nanoparticles remains to be elucidated. Such investigations are very crucial in assessing the long-term efficacy of Gs-NPs to facilitate the development of sustainable nanotechnology in crop production and protection systems.