**4. Toxicity of** *Fusarium* **Nanoparticles**

Nanoparticles are unique materials as they have property combinations compared with conventional materials [114]. There is a wide range of applications of NPs such as in human health appliances, industrial, medical and biomedical fields, engineering, electronics, and environmental applications [115]. Among all nanomaterials, AgNPs are the most widely used in medicine, medicinal devices, pharmacology, biotechnology, electronics, engineering, energy, magnetic fields, and also in environmental remediation [116]. Their highly effective antibacterial activity has found applications in industrial sectors including textiles, food, consumer products, medicine, etc. [117].

The unique physical, chemical and biological (e.g., antimicrobial, anticancer, antiparasitic) properties of nanoparticles differ largely from corresponding bulk materials and make them a high-demand material in different sectors. However, the widespread and increased use of nanoparticles may pose a risk to both the environment and living organisms by increasing the level of toxicity [118]. To date, several studies have used different model cell lines to exhibit the cytotoxicity of nanoparticles from *Fusarium* species, mainly AgNPs, and their underlying molecular mechanisms [31,60,65,119]. Biogenic nanoparticles are capped with natural molecules like proteins [120–122]. This capping is defined as corona. This nanoparticle corona significantly affects the biological response [123]. Based on the surface affinity and exchange rate, the corona can be divided into two forms: hard corona and soft corona. The soft corona proteins are 'vehicles' for the silver ions, whereas the hard coronas are rigid for the trespass into the cellular system [124]. The functional groups of the corona play a key role in the formation of the nanoparticle–protein corona [125]. These functional groups along with the protein charges also regulate the cytotoxic properties of the nanoparticle corona [123,125]. The surface charge of nanoparticles plays an important role in their bactericidal activity against both Gram-positive and Gram-negative bacteria, as exemplified by AgNPs [126]. It was confirmed using transmission electron microscopy (TEM) that AgNPs can penetrate cellular compartments such as endosomes, lysosomes, and mitochondria [127]. Several reports indicate that the proteins contained in the nanoparticle corona interact with the cells and not the nanoparticles themselves [128,129]. Thus, the corona formation and composition have important implications for both toxicity [130] and internalization [131].

To sum up, particle size [132], particle shape [133], particle surface properties [134], biological fluid properties, and composition affect the corona structure and thus the adverse effects on human health and the environment [131,135].

#### *4.1. Effect of Size and Shape of Nanoparticles on Cytotoxicity*

It is claimed that apart from the size, the shape of nanoparticles affects their toxicity to cells. Mohamed et al. [31] investigated the cytotoxicity of the two different shapes of zinc oxide nanoparticles (ZnONPs) biosynthesized from *Fusarium keratoplasticum* (A1-3) and *Aspergillus niger* (G3-1). These nanoparticles displayed a similar size (10–42 and 8–38 nm) but different shapes, namely hexagonal and nanorods, respectively. It was reported that a safe dose of these nanoparticles for applications in animal cells should be lower than 20.1 and 57.6 ppm, respectively. Therefore, the rod ZnONPs were more applicable to safety at high concentrations in contrast to hexagonal ZnONPs [31]. Soleimani and co-authors [136] studied biological activity of different shapes (cube, sphere, rice and rod) of AgNPs synthe-

sized using chitosan in acetic acid solution and 0.2 M of AgNO<sup>3</sup> (cubes) and *F. oxysporum,* starch and 0.08 M of AgNO<sup>3</sup> at pH 6.8 (spheric) or 1.0 M of AgNO<sup>3</sup> at pH 3.0 (nanorice), and finally, *F. oxysporum*, starch and 1.2 M AgNO<sup>3</sup> at pH 3.0 and 30 ◦C for 3 days (blunt ends rods) or with 10-days incubation (sharp-ends rods). They found that silver nanostructures with different shapes are not inherently toxic to human cells at concentrations lower than 10 µg ml−<sup>1</sup> , whereas for higher concentrations cell viability decreased in a shape and dose-dependent manner. Nanocubes, nanorice and sharp-nanorods were found to be more toxic than spheres and blunt-nanorods. The former significantly decreased cell viability at concentrations 25 µg mL−<sup>1</sup> or higher, and the latter at concentrations of 50 and 75 µg mL−<sup>1</sup> . Moreover, they showed that cubic nanoparticles inhibited the growth of all tested bacteria (*Bacillus subtilis, Staphylococcus aureus, Escherichia coli,* and *Pseudomonas aeruginosa*) at the lowest-used concentrations (10 ppm). The toxic effect of nanostructures against bacterial cells increased in order spheres, blunt-rods, nanorice, sharp-rods, and nanocubes [136]. It was concluded that due to stronger vertex in sharp ends of silver nanostructures, some geometries of AgNPs, especially cubic structures, have more interaction with the bacterial cell membrane, resulting in stronger biological activity [136]
