Modern medicine, veterinary, food, and agriculture sectors are struggling with microbial diseases due to the increasing resistance of microorganisms to available antimicrobials agents (Food and Agriculture Organization of the United Nations (FAO), 2017; Xie et al., 2018; Hashempour-Baltork et al., 2019). Antibiotic resistance results from the excessive and reckless use of antibiotics in agricultural food production, healthcare, and environmental protection sectors (Xie et al., 2018; Caniça et al., 2019; Dadgostar, 2019; Larsson and Flach, 2022). A wide range of pathogenic bacteria cause chronic infections by forming complex multicellular structures known as biofilms. Moreover, some studies indicate that conventional antibiotics may induce phenotypic changes in bacterial cells which subsequently trigger biofilm formation (Olivares et al., 2020). Biofilms provide a stable protective environment for the dissemination of microorganisms due to a self-synthesized structure which is an organic and highly hydrated matrix composed of exopolysaccharides, proteins, and nucleic acids (Hall-Stoodley and Stoodley, 2005; Olivares et al., 2020). There are several hypotheses of biofilm recalcitrant to antibiotics, including persistent cells, adaptive responses, and lower penetration of antimicrobial agents (Sahoo et al., 2021). In response to the growing threat of bacterial pathogens, it is necessary to develop approaches that are wide-ranging and effective in controlling bacterial outbreaks. An alternative to conventional chemicals is the formulation of agents that may prevent biofilm formation and also act on individual bacterial cells.

The growing population and increasing demand for food, require solutions to increase crop yields and ensure safety in food production, distribution, and storage (Fung et al., 2018; Wypij et al., 2023). Nowadays, microbial diseases of plants are becoming an increasing problem as they reduce crop yields and significantly affect the food industry on both a global and regional scale (Charkowski, 2018; Charkowski et al., 2020). Among the most notable bacterial pathogens of plants are Pseudomonas syringae, Ralstonia solanacearum, Agrobacterium tumefaciens, Xanthomonas spp., Erwinia amylovora, and Pectobacterium carotovorum (Mansfield et al., 2012; Iwu and Okoh, 2019). The bacterial diseases of crop plants including black rot (Schaad et al., 1980; Maji and Nath, 2015), soft rot (Pérombelon and Kelman, 1980; Pérombelon, 2002), bacterial speck (Shenge et al., 2007; Butsenko et al., 2020), and fire blight (Johnson and Stockwell, 1998; Vanneste, 2000; Peil et al., 2021). While, fungal pathogens, for example, Fusarium oxysporum, Botrytis cinerea, Colletotrichum acutatum, Puccinia sp., Sclerotinia sclerotiorum, etc., have a broad host range and cause pre-and post-harvest diseases. Among them, causal agents of grey and white mold as well as black leg and dry rot are worthy of note (Amselem et al., 2011; Dean et al., 2012; Gabal et al., 2019). Plant pathogens spread rapidly and are increasingly aggressive and harmful leading to yield losses in cereals (maize, wheat, rice), vegetables (potatoes, tomatoes, brassicas), and fruits (Sundin et al., 2016; Nawaz et al., 2020; Hampf et al., 2021). Indeed, foodborne illnesses caused by bacteria, are another concern with ensuring human food security (Gandhi and Chikindas, 2007; Scallan Walter et al., 2021). Bacterial pathogens such as Listeria sp., or the the members of Enterobacteriaceae family are proliferated via the fecal-oral routes by ingestion of contaminated water or food. Virulent strains of these bacteria cause a broad-spectrum of disorders like nausea, watery or bloody diarrhea, vomiting, and inflammatory changes (Allocati et al., 2013). Thus, intracellular pathogen Listeria monocytogenes is responsible for listeriosis, a very severe and deadly illness (Chlebicz and Śliżewska, 2018). Symptoms of the infection in healthy adult patients may include lack of appetite, stomachache, nausea, or diarrhea. Nonetheless, the infection is particularly dangerous for newborns, the elderly, immunocompromised patients, and poses a risk of miscarriage for pregnant women (Zhu et al., 2017).

Silver nanoparticles (AgNPs) have pronounced antimicrobial activity, even at low concentrations (Salleh et al., 2020). Some studies indicate, that AgNPs reveal long-term antibiofilm activities by multi-site action due to their unique properties, which include nano-size and high surface area (Martinez-Gutierrez et al., 2013; Ali et al., 2015; Soliman et al., 2022). Basically, biological methods of nanoparticle synthesis have been recognized as a substitute for physical and chemical ones (Rai et al., 2021b; Al-Rajhi et al., 2022). In addition, biologically synthesized AgNPs are capped with molecules of natural origin, which enables them to interact easily with bactericides along with bacterial cells and therefore, improves their antimicrobial efficiency (Wypij et al., 2020). Among the biological synthesis methods of nanoparticles (NPs) are those mediated by plants, algae, bacteria or fungi (Wypij et al., 2021; Al-Rajhi et al., 2022; Salem et al., 2022, 2023). Among microorganisms, fungi reveal the strong potential to secrete metabolites involved in the synthesis of metallic nanoparticles (MNPs) (Rai et al., 2021a; Shaheen et al., 2021). Mycosynthesis is an environmentally friendly, inexpensive, and simple method, which can be used for the efficient production of AgNPs (Lahiri et al., 2021; Salem et al., 2023). Current research efforts focus on the optimization of mycosynthesis of AgNPs with desired physio-chemical and biological properties, which may be used to eradicate bacterial pathogens (Zhao et al., 2018).

Hence, the present study was designed to synthesize AgNPs from Fusarium culmorum strain JTW1 in an easy, eco-friendly, efficient, and inexpensive way and to estimate the potential use of AgNPs in the biomedicine, agriculture, and food production industry. The mycosynthesized AgNPs were characterized using UV–vis spectrophotometry (UV–Vis), Transmission Electron Microscopy (TEM), X-ray diffraction (XRD) spectroscopy, Energy-dispersive X-ray spectroscopy (EDX), Fourier-Transform Infrared Spectroscopy (FTIR), Nanoparticle Tracking Analysis (NTA), Dynamic Light Scattering (DLS) and Zeta potential measurement. The comprehensive antibacterial activity of AgNPs, including estimation of minimal inhibitory and biocidal concentrations (MIC and MBC, respectively), efficacy of combined antibiotics and AgNPs as well as antibiofilm activity against a wide range of Gram-positive and Gram-negative pathogenic bacteria of humans and plants were investigated. Mycosynthesized AgNPs were tested against a set of fungal and oomycete plant pathogens, including Alternaria alternata IOR 1783, Botrytis cinerea IOR 1873, Colletotrichum acutatum IOR 2153, Fusarium oxysporum IOR 342, Fusarium solani IOR 825, Phoma lingam IOR 2284, Sclerotinia sclerotiorum IOR 2242, Phytophthora cactorum IOR 1925, Phytophthora cryptogea IOR 2080, Phytophthora megasperma IOR 404 and Phytophthora plurivora IOR 2303.

We synthesized AgNPs by mycelial extract of Fusarium culmorum isolate JTW1 after treatment with 1 mM aqueous solution of silver nitrate, and visually characterized by the color change, from light-yellow to dark-brown. The color change provides evidence of the formation of AgNPs due to the surface plasmon resonance (SPR) exhibited by the synthesized AgNPs (Smitha et al., 2008). Synthesis of myco-AgNPs was confirmed spectroscopically by observation of a characteristic absorbance peak at 430 nm, which is characteristic of AgNPs. According to the literature, the absorption peaks between 380–450 nm are related to the surface plasmon resonance (SPR) of silver nanoparticles (Desai et al., 2012; Mistry et al., 2021). Our UV–Vis results corroborate with those presented by Huang et al. (2018) and Osorio-Echavarría et al. (2021) who showed that AgNPs synthesized from fungi releveled strong peaks at 430 nm.

Mycosynthesized silver nanoparticles are well known for their bioactivities including antimicrobial activity against both Gram-positive and Gram-negative bacteria (Majeed et al., 2018). The bioactivity of AgNPs is highly dependent on the size, shape, stability, and capping molecules of nanoparticles as well as target cells (e.g., bacterial strain; Li et al., 2012). Therefore, comprehensive characterization of the physical and chemical properties of mycosynthesized AgNPs is important for their potential application.

In the present study, various analytical techniques were used to characterize silver nanoparticles, namely TEM, EDX, XRD, FTIR, DLS, NTA, and Zeta potential measurement. TEM analysis showed small (average size 16 nm) and spherical nanoparticles from F. culmorum. Similarly, Bawaskar et al. (2010) reported the synthesis of spherical and small (average size 11 nm) AgNPs by F. culmorum extract after challenging with 1 mM AgNO₃ at ambient temperature. Our results are also in accordance with the findings of Azmath et al. (2016), who reported the fabrication of AgNPs from Colletotrichum sp. ALF2-6 with a size ranging from 5 to 60 nm, but with highly variable shapes. However, TEM micrographs showed a clearly smaller size of AgNPs from F. culmorum strain JTW1 than NTA and DLS analyses (188.4 and 169.9 nm, respectively) used for further characterization. In the case of the latter analyses the larger size of nanoparticles is due to the presence of capping molecules from fungal extract on the surface of AgNPs (Joshi et al., 2013). Moreover, smaller nanoparticles may not be detected in the DLS technique, as they can be covered by bigger ones (Tomaszewska et al., 2013). In turn, EDX spectrum confirmed elemental composition of AgNPs and the peaks observed at 3, 22, and 24 keV were typical for the absorption of AgNPs due to surface plasmon resonance (Soliman et al., 2022). The X-ray diffraction pattern confirmed the reduction of Ag ⁺ ions and the formation of AgNPs with a face-centered cubic (fcc) structure. However, several unassigned peaks were also recorded in the XRD analysis. It is suggested that these additional peaks arise due to the crystallization of the bioorganic phase on the surface of the AgNPs (Elumalai et al., 2017; Wypij et al., 2022). The presence of biomolecules on the surface of synthesized AgNPs was confirmed by FT-IR measurements which revealed characteristic peaks. The band recorded at 3448.75 cm⁻¹ was assigned to the stretching vibrations of primary and secondary amines and indicated the presence of free OH and NH groups. The peaks at 2923.34 and 2852.38 cm⁻¹ showed the presence of CH stretching (Ghaseminezhad et al., 2012; Wypij et al., 2021). A band at 1632.52 cm^− 1 corresponded to the bending vibrations of the amide I and amide II bands of proteins (Elamawi et al., 2018). The bands observed at 1384.60 and 1352.28 cm⁻¹ can be assigned to the C–N stretching vibrations of aromatic amines (Jain et al., 2011; Salem, 2022) while the peak at 1085.60 cm⁻¹ can be associated with the C–N stretching vibrations of the aliphatic amines (Vigneshwaran et al., 2007). The presence of different functional groups associated with amines, alkanes, and a carboxylic acid, on the surface of AgNPs suggest that various biomolecules from fungal extracts can be involved in nanoparticle formation and stabilization (Ammar and El-Desouky, 2016; Akther et al., 2019). Besides, the biological capping of AgNPs prevent their aggregation and may affect their biological activity, including antimicrobial activity (Hamouda et al., 2019). The high negative value of zeta potential (−38.43 mV) confirmed the repulsion among the particles and indicates their high stability. Elamawi et al. (2018) reported negatively charged (−19.7 mV) and well-dispersed silver nanoparticles synthesized from Trichoderma longibrachiatum. However, their zeta potential was much lower suggesting lower stability.

AgNPs synthesized from F. culmorum strain JTW1 demonstrated remarkable antimicrobial activity against human pathogens (Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Staphylococcus aureus), food-borne pathogens (Listeria monocytogenes, Salmonella enterica, Salmonella infantis) and plant pathogens (Agrobacterium tumefaciens, Pectobacterium carotovorum, Pseudomonas syringae, and Xanthomonas campestris). Interestingly, among human pathogens, the most sensitive to mycosynthesized AgNPs was P. aeruginosa, followed by E. coli, K. pneumoniae, and food-borne pathogens, including Gram-positive L. monocytogenes. It is particularly significant that AgNPs from F. culmorum JTW1 inhibited the growth of members of Gram-negative bacteria that are on the list of multidrug-resistant taxa highlighted by the World Health Organization (WHO) (2021).

AgNPs have been found to be effective in the management of plant diseases caused by bacteria, such as Xanthomonas oryzae, X. phaseoli, Ralstonia solanacearum, Clavibacter michiganensis and many more (Tariq et al., 2022). In the present study, P. carotovorum was found to be the least sensitive to mycosynthesized AgNPs (MIC and MBC values were 128 and 512 μg mL⁻¹, respectively) when compared to P. syringae, X. campestris, and A. tumefaciens.

It is suggested that effective action of silver nanoparticles against bacteria may be through the progressive release of lipopolysaccharides and proteins as well as the further formation of irregularly shaped pits at the outer membrane of Gram-negative bacteria (Amro et al., 2000; Fayaz et al., 2010). Salvioni et al. (2017) demonstrated that antimicrobial activity of negatively charged AgNPs depended on the concentration and was associated with their incorporation into the membrane of Gram-negative bacteria (E. coli) and penetration into the cells. In addition, intracellular leakage and coagulation of nanoparticles on the bacterial surface were also observed. In turn, treatment of Gram-positive S. aureus with AgNPs resulted in the appearance of blisters on the surface of cells and cell lysis (Salvioni et al., 2017). Other studies indicated that AgNPs or released Ag⁺ ions after penetration into a cell may generate free radicals (Qing et al., 2018; Tian et al., 2018) and interact with biomolecules and intracellular structures (Li et al., 2010; Adeyemi et al., 2020; Gul et al., 2021). Yuan et al. (2017) reported that AgNPs caused lower lactate dehydrogenase activity (LDH) and decreased levels of adenosine triphosphate (ATP) in P. aeruginosa and S. aureus. Moreover, ROS generated after AgNPs treatment led to inhibition of metabolic activity and the growth of bacterial cells. It is claimed that due to the different mechanisms of AgNPs action, which affect various aspects of bacterial growth and metabolism, the development of microbial resistance to nanoparticles is limited (Wang L. et al., 2017).

In the present study, tested antibiotics demonstrated antibacterial activity against Gram-negative and Gram-positive bacteria which was increased in combination with AgNPs from F. culmorum strain JTW1. The enhanced antimicrobial activity against E. coli, K. pneumoniae, and P. aeruginosa was observed for a combination of AgNPs and streptomycin and against P. aeruginosa with ampicillin. In the case of S. aureus, increased activity of ampicillin and kanamycin was observed when combined with AgNPs. The increased antimicrobial effect may be due to the binding of antibiotics to AgNPs. The molecules of antibiotics react easily with nanoparticles due to many active groups such as hydroxyl and amido groups (Wypij et al., 2022). Hence, AgNPs may be covalently bound to streptomycin due to the presence of four-terminal –NH₂ groups in antibiotic structure. In addition, this association might be enhanced by the electrostatic interactions between negatively charged AgNPs and positively charged streptomycin molecules (Wang C. et al., 2017). Moreover, biologically synthesized nanoparticles showed more stable adsorption of ampicillin when compared to chemically synthesized ones due to the presence of an organic layer on their surface, which acts as a stabilizer (Rogowska et al., 2017). It is claimed that AgNPs in combination with antibiotics can promote cell wall disruption and damage, and facilitate the transport of antibiotics into the cell by increasing membrane permeability. Other mechanisms of AgNPs action include inhibition of enzymes responsible for the hydrolysis of antibiotics due to the disruption of their structure and release of Ag⁺ ions (Panáček et al., 2016). Therefore, it is assumed that the combination of AgNPs and antibiotics enhanced their inhibitory activity against pathogenic bacteria. This allows the use of lower doses of antibacterial agents and reduces the risk of resistance to antimicrobial agents (Hemeg, 2017).

AgNPs are reported to hinder the biofilm formation in many bacteria (Markowska et al., 2013; Ansari et al., 2021; Hetta et al., 2021). Crystal violet staining and fluorescein diacetate assay was used for the visualization of adherent biofilm bacteria and measurement of biofilm activity, respectively. Interestingly, AgNPs from Fusarium culmorum strain JTW1, at low concentration (0.25 μg mL⁻¹), efficiently inhibited the formation and hydrolytic activity of P. aeruginosa biofilm. Similar results were reported by Akther et al. (2020) who studied the efficiency of AgNPs synthesized from Rhizopus arrhizus BRS-07 in reducing biofilm formation by P. aeruginosa and found that the formation of this structure was inhibited by 7–93% in a dose-dependent manner (5–25 μg mL⁻¹). Furthermore, AgNPs synthesized from F. culmorum strain JTW1 at a higher concentration of 64 μg mL⁻¹ also efficiently (by 67–96%) reduced the formation of biofilms in E. coli strains, P. aeruginosa, and K. pneumoniae. These nanoparticles were found to be more active against biofilm than those reported by Ramachandran and Sangeetha (2017) which completely inhibited the formation of Escherichia coli (ETEC12), Klebsiella pneumoniae (SKP7) and Pseudomonas aeruginosa (ETPS11) biofilms after treatment with AgNPs at a concentration of 100 μg mL⁻¹. Ansari et al. (2014) found the highest antibiofilm efficiency of commercially available water-soluble AgNPs (5–10 nm) against Escherichia coli and Klebsiella spp. at the concentration of 50 μg mL⁻¹. The inhibition of biofilm formation by AgNPs is a consequence of bacterial growth arrest, prevention of exopolysaccharide formation (Ansari et al., 2014), and disruption of bacterial quorum sensing signaling (Akther et al., 2020). In our study, although AgNPs at the lower concentrations had no impact on biofilm formation by E. coli ATCC 8739, K. pneumoniae, and S. aureus ATCC 25923, they significantly decreased the activity of hydrolases.

Food-borne pathogens, such as Listeria monocytogenes and Salmonella enterica are known to form biofilms, which ultimately leads to serious problems in the food industry (Harrell et al., 2021; Muthulakshmi et al., 2022). Our findings confirmed that both the aforementioned bacteria showed sensitivity to mycofabricated AgNPs. However, the development of biofilm in L. monocytogenes was more prevented than S. infantis. Our results are in line with those reported by Chlumsky et al. (2020) who showed higher activity of AgNPs in reducing biofilm formation and a decrease in metabolic activity against L. monocytogenes, S. aureus than S. infantis S59. The in vitro studies on the AgNPs from Terminalia catappa leaf extract against L. monocytogenes revealed the inhibition of biofilm formation and reduction in the virulence factors such as protease production, at concentrations of 50 and 100 μg mL⁻¹ (Muthulakshmi et al., 2022). Similarly, AgNPs from F. culmorum strain JTW1 remarkably prevented biofilm formation ability in bacterial plant pathogens, especially in A. tumefaciens and P. carotovorum. Moreover, a significant reduction in the hydrolase enzymatic activity of these bacteria was recorded. Recently, Olfati et al. (2021) also reported the inhibitory effect of AgNPs synthesized from Calendula officinalis extract against the biofilm formation of P. carotovorum. Consequently, the obtained results clearly reveal that mycosynthesized AgNPs not only effectively inhibited the activity of enzymes but also the growth of the bacteria. Therefore, it can be hypothesized that the ability to inhibit the hydrolytic enzyme activity of bacteria may occur due to an interaction of AgNPs with proteins, causing an alteration in structure and interfering with enzymatic functions (Wigginton et al., 2010).

Silver nanoparticles from F. culmorum strain JTW1 showed strong inhibitory activity against the growth and spore germination of Botrytis cinerea, Sclerotinia sclerotiorum, and Phoma lingam, but not against remaining fungal strains. Similarly, antifungal activity of AgNPs obtained from strains of the genus Trichoderma to plant pathogens was shown in other studies reported by Guilger et al. (2017) and Tomah et al. (2020). For instance, AgNPs synthesized from Trichoderma sp. at a concentration of 200 μg mL⁻¹ reduced the growth of mycelia of Sclerotinia sclerotiorum, formation of new sclerotia and their germination (Tomah et al., 2020). The authors further reported the changes in fungal cells by induction of pore formation in the cell wall and further accumulation of AgNPs inside the fungal cells (Tomah et al., 2020). Similarly, Guilger et al. (2017) found that AgNPs at a concentration of 0.31 × 10¹² NPs/mL inhibited mycelial growth and the formation of new sclerotia of Sclerotinia sclerotiorum. However, in our study, inhibition of sclerotia germination was achieved at a much lower concentration of myco-AgNPs (32 μg mL⁻¹) showing their higher effectiveness in elimination of S. sclerotiorum. In another study, Malandrakis et al. (2019) tested the sensitivity of a number of fungal pathogens to commercially available silver nanoparticles (size <100 nm) and reported the highest susceptibility of Botrytis cinerea, followed by Verticillium dahliae, Colletotrichum gloeosporioides, Monilia fructicola, Alternaria alternata, Fusarium oxysporum f. sp. Radicis-Lycopersici and Fusarium solani (the latter was unaffected in tested concentration range). The higher inhibitory effect of AgNPs was observed against spores germination (EC₅₀ values in the range 5.08–390.20 μg mL⁻¹) than mycelial growth (EC₅₀ value were > 306 μg mL⁻¹) of tested pathogens (Malandrakis et al., 2019). Our findings provide evidence of the differential response of fungal pathogens to AgNPs. The highest inhibition of mycelial growth was recorded in P. lingam (9.54 mm), followed by B. cinerea (4.93 mm) and S. sclerotiorum (3.41 mm) whereas, in the case of spore germination, maximum inhibition was found in S. sclerotiorum (32 μg mL⁻¹), followed by B. cinerea (64 μg mL⁻¹) and P. lingam (256 μg mL⁻¹). Nevertheless, our findings of antimicrobial potential of AgNPs against a wide range of fungal crop pathogens indicate their potential for use in agricultural crop management to protect against diseases such as gray or white mold, blackleg, canker, and dry rot.

In this study, AgNPs were effectively synthesized from Fusarium culmorum strain JTW1. They were comprehensively characterized using UV–vis, TEM, XRD, NTA, Zeta potential measurements, and FTIR which revealed small size, spherical shape, stability, crystalline nature of AgNPs, and that mycosynthesized AgNPs were capped with biomolecules. Moreover, AgNPs were found to have strong antimicrobial potential against bacterial pathogens of humans and plants. AgNPs also enhanced antibacterial activity of streptomycin against Gram-negative bacteria, namely E. coli strains ATCC 8739 and ATCC 25922, K. pneumoniae ATCC 700603, P. aeruginosa ATCC 10145, ampicillin against P. aeruginosa, and ampicillin and kanamycin against Gram-positive S. aureus ATCC 25923 and S. aureus ATCC 6538, respectively. Furthermore, AgNPs significantly reduced bacterial biofilm formation and inhibited activity of hydrolytic enzymes synthesized by biofilm cells. In addition, AgNPs showed great potential to inhibit fungal spore germination, which is crucial in the fungal spread in the environment, and growth of fungal mycelia. Therefore, these mycosynthesized silver nanoparticles in view of their unique properties, have a high potential as an antimicrobial agent in medicine and promising solution to control or reduce pathogens in agriculture and the food industry.

The original contributions presented in the study are included in the article/Supplementary material, further inquiries can be directed to the corresponding authors.

PG and MR conceived research and edited and reviewed it. PG and JT-W designed research. JT-W and MW conducted experiments and acquired the funds. JT-W analyzed data and wrote the manuscript. All authors have read and agreed to the published version of the manuscript.

This research was funded by Excellence Initiative – Research University, Nicolaus Copernicus University in Torun, Grant4NCUStudents (38/2021/Grants4NCUStudents and 02/2021/Grants4NCUStudents). The ACP was funded from EF-IDUB of Nicolaus Copernicus University.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.