The most important task of modern nanotechnology is the development of reliable and efficient techniques allowing one to produce monodisperse nanoparticles with needed parameters. Nanoparticle production methods can be divided into physical, chemical and biological (or bio-assisted) ^[1][2][1,2], as well as combined methods putting together biological materials and physical influences, such as microwave radiation ^[3][4][3,4]. The chemical and physical methods traditionally used to produce nanoparticles allow large quantities of particles to be synthesized in a short time, but they are often expensive and difficult to perform; the issue of their environmental safety is a big problem as well. Therefore, in recent years, there has been an increasing interest in green nanotechnology and the biological synthesis of nanoparticles ^[5][6][7][8][5,6,7,8]. The introduction of green synthesis techniques can reduce the negative impact of nanotechnology on the environment by using less toxic reagents and reducing the risks of secondary pollution. Furthermore, nanoparticles produced via biosynthesis may have higher stability and biocompatibility and lower toxicity owing to their coating with biogenic surfactants or capping agents ^[9][10][11][9,10,11].

The ability to form nanoparticles has been found in all groups of organisms. Numerous studies have shown that plants ^{[12][13][14][15]}[12,13,14,15], animals ^[10][16][17][10,16,17], bacteria ^[16][18][16,18], fungi ^{[11][19][20][21]}[11,19,20,21], actinomycetes ^[22][23][22,23], algae ^{[14][16][24][25]}[14,16,24,25], lichens [26] and viruses [10] can be successfully used to produce nanoparticles. Along with living cultures, their biomass, cell fractions, extracts, metabolites and spent media can also be used in green nanosynthesis ^{[6][11][12][14][27][28]}[6,11,12,14,27,28], as well as various plant and animal food products ^{[29][30][31][32]}[29,30,31,32] and organic industrial wastes ^[33][34][35][33,34,35]. Fungal cultures of different taxonomic groups are very promising biological objects for the green synthesis of nanoparticles ^[11][20][36][11,20,36]. The advantage of fungi in comparison to other organisms is their ability to produce a wide range of active protein molecules, convert ions of heavy metals and other trace elements into less toxic forms under the action of their enzymes and accumulate them in large quantities both within their mycelium and extracellularly. Therefore, fungi-mediated nanosynthesis has been increasingly studied in the past two decades. In fungal cultures, the ability to form a wide range of nanoparticles of different chemical compositions was found, including metals, metalloids, metal oxides and sulfides and other compounds, as well as composite nanoparticles ^[19][36][37][19,36,37].

All biological methods of nanosynthesis can be divided into two main types. In the first one, living cultures are used directly to serve as “nanofactories“, forming nanoparticles in vivo and accumulating them in their cells, on their cell surface or in the medium. Fungal biomass separated from the culture liquid can also be used. The disadvantage of this method is the need to separate the obtained nanoparticles from the bio-object’s cells; moreover, the process of biosynthesis by growing cultures may take a long time. The level of the precursor may also be a limiting factor, because high concentrations of metals and other compounds used to biosynthesize nanoparticles inhibit the growth processes. Another option for green nanoparticle synthesis is the use of various substances derived from bio-objects, such as culture liquids, intracellular extracts, protein fractions or individual fungal metabolites. The use of such techniques greatly facilitates the process of nanosynthesis, as there is no need to destroy the producing organism’s cells and separate nanoparticles from them.

The physico-chemical properties of nanoparticles are closely related not only to their chemical composition and crystal structure but also to their size and morphology, including particle physical shape, surface topography and the presence of pores and cavities ^[38][39][38,39]. It is also important that the synthesized nanoparticles are homogeneous in size and shape and resistant to aggregation in suspensions. The properties of biogenic nanoparticles have been found to depend on the species and strain of the microorganism, the extracts and metabolites used, the precursor compound and its concentration, media composition, stirring rate, incubation time, temperature, pH and other conditions, the varying of which can control nanoparticle formation ^[40][41][42][40,41,42]. The characteristics of the resulting nanoparticles determine their future applications. In this regard, an important challenge in nanobiotechnology is to develop methods allowing one to produce nanoparticles with better control over their size, shape and other properties. However, not enough attention has been paid to the study of the influence of the synthesis conditions on the properties of biogenic nanoparticles. In particular, there are still few comparative studies on the mycosynthesis of nanoparticles with different characteristics using the same fungal species but under different conditions.

2. Fungi-Mediated Synthesis of Nanoparticles

2.1. Mycosynthesis of Silver Nanoparticles

Owing to their unique features, silver nanoparticles (AgNPs) have many applications in various fields of medicine and engineering. The area of their applications includes electronic components, biomedical devices, textile engineering, cosmetics, agricultural engineering and many others ^[43][44][43,44]. AgNPs have been found to have a wide range of biological activities, including antibacterial, antifungal, antiviral, antitumor, hepatoprotective and hypotensive properties, which is why they are actively used for therapeutic purposes ^[45][46][45,46]. To date, the biological synthesis of AgNPs in fungi has been the most extensively studied of all elements. The ability to produce nanosilver has been detected in more than 120 species of fungi from different taxa, including Ascomycota (Alternaria [47], Aspergillus ^[48][49][48,49], Beauveria [50], Bionectria [51], Botryodiplodia [52], Chrysosporium [53], Cladosporium [54], Colletotrichum [55], Epicoccum [56], Fusarium ^[57][58][57,58], Geotricum [59], Guignardia [60], Helvella [61], Hormoconis [62], Humicola [63], Macrophomina [64], Neurospora [65], Paecilomyces [66], Penicillium ^{[67][68][69][70][71][72]}[67,68,69,70,71,72], Pestalotia [73], Phoma ^[74][75][74,75], Picoa [76], Saccharomyces [77], Sclerotinia [78], Scopulariopsis [79], Talaromyces [80], Tirmania [81], Trichoderma ^{[42][48][82][83][84]}[42,48,82,83,84], Verticillium [85], Yarrowia [86]), Mucoromycota (Rhizopus [87]) and Basidiomycota (Agaricus ^{[41][88][89][90][91]}[41,88,89,90,91], Auricularia [92], Bjerkandera [93], Boletus [94], Calocybe [95], Coriolus [94], Cryptococcus [96], Flammulina ^[97][98][97,98], Fomes [99], Fomitopsis [100], Ganoderma ^{[41][82][101][102]}[41,82,101,102], Grifola [41], Hypsizygus [103], Inonotus [104], Lactarius [105], Laxitextum [106], Lentinus ^[41][107][41,107], Microporus [108], Phaenerochaete [109], Phellinus [88], Piriformospora [110], Pleurotus ^{[41][111][112][113][114]}[41,111,112,113,114], Pycnoporus [115], Rhodotorula [116], Schizophylluum [117], Trametes [118], Tricholoma [119], Volvariella [120]). Basidiomycetes are of particular interest as promising bio-objects for nanoparticle fabrication. Most of the basidiomycetes studied for mycosynthesis belong to edible and medicinal mushrooms, many of which are grown in artificial culture. These fungi produce a wide range of biologically active molecules, which not only can act as capping and stabilizing agents but also have anticancer, anti-inflammatory, antioxidant and antimicrobial activities themselves, allowing the production of nanoparticles with complex biomedical properties. The number of research papers on nanosilver mycosynthesis includes many dozens and is constantly growing. In recent years, there have been several reviews detailing the production of biogenic nanoparticles of this element using fungal cultures ^{[45][121][122]}[45,121,122]. Therefore, below reswearchers will focus on some of the most recent publications in the past five years (Table 1). cultural liquid [139]. Spherical, pentagonal and hexagonal nanoparticles (5–30 nm) were obtained with Trichoderma hamatum mycelial extract [149]. The authors optimized the conditions for the synthesis of AuNPs with the smallest size using T. hamatum. Nanoparticles biosynthesized using T. harzianum mycelial biomass had a nanometric size distribution below 30 nanometers and a spherical shape [150]. AuNPs of variable shapes with considerable antibacterial, antioxidant and antimitotic activities were obtained with an Alternaria spp. extract [135]. Gold nanospheres (10–100 nm) with antibacterial and antifungal properties were obtained using Phoma sp. mycelial biomass [148]. Cubic AuNPs with strong antimicrobial, cytotoxic and antioxidant activity were synthesized using a Morchella esculenta fruit body extract [147]. Molnár et al. studied AuNP mycosynthesis by 29 thermophilic fungi and compared the results of three different approaches for the synthesis of gold nanoparticles using the extracellular fraction, the autolysate or the intracellular fraction of the fungi [152]. They observed the formation of nanoparticles with different sizes (ranging between 6 nm and 40 nm) and size distributions depending on the fungal strain and experimental conditions. Vetchinkina et al. studied AuNP mycosynthesis by A. bisporus and Agaricus arvensis cultures ^[41][153][41,153]. The use of live cultures, culture liquids and mycelial extracts resulted in the formation of nanoparticles of different sizes and shapes. Nanospheres were formed with living cultures and culture liquids, while irregularly spherical particles in the case of A. bisporus and various shapes with A. arvensis were formed using intracellular mycelial extracts. An extract from the A. bisporus fruit body was prepared and utilized as a reducing and stabilizing agent toward a green synthesis of AuNPs [134]. The different parameters such as the precursor concentration, precursor:extract ratio, pH, temperature, reaction mode and reaction time were optimized for the mycosynthesis of AuNPs. The synthesized gold nanospheres (10–50 nm) significantly inhibited the growth of clinically important pathogenic Gram-positive and Gram-negative bacteria and pathogenic fungi. AuNPs with a dye-degrading activity obtained by Dheyab et al. using an A. bisporus fruit body extract were oval, spherical, drum-like, hexagonal and triangular (average size of 53 nm) [133]. An A. bisporus mushroom extract was also used to synthesize gold nanospheres through a hydrothermal process (at a pressure of 15 psi and a temperature of 121°C for 15 min) [132]. The optimal conditions for the maximum nanoparticle concentration and stability were selected. Face-centered cubic nanocrystals with dye-reducing properties were synthesized using phenolic compounds isolated from Ganoderma applanatum [141]. Anticancer AuNPs biofabricated using a G. lucidum fruit body extract exhibited shapes such as spherical, oval and irregular, and their size ranged between 1 and 100 nm [142]. AuNPs synthesized using G. lucidum living cultures, as well as cultural liquid, were spherical, while the use of G. lucidum mycelial extract resulted in spherical, hexagonal, tetragonal and triangular particle formation [41]. The same results were obtained for Grifola frondosa and Pleurotus ostreatus cultures as well [41]. Chaturvedi et al. combined AuNP synthesis with the use of a P. sajor-caju fruit body extract followed by microwave irritation to further enhance the effects of fabricated gold nanospheres [113]. Vetchinkina et al. studied AuNP mycosynthesis by the L. edodes culture [41]. Living cultures formed nanospheres of 5–50 nm; smaller nanospheres were formed by the incubation of culture liquid with HAuCl₄, and spherical, hexagonal, tetragonal and triangular particles of various sizes were formed with mycelial extract. Nanoparticles different in shape and size were synthesized using enzymes isolated and purified from the L. edodes mycelium. Spherical nanoparticles (2–20 nm) were obtained using intracellular Mn-peroxidase, and particles forming with the use of intracellular laccases and tyrosinases were bigger and irregularly spherical, triangular and tetrahedral in shape. When AuNPs were made with extracts from different morphogenetic stages of L. edodes and G. lucidum, their size, shape and degree of aggregation differed between the morphological structures involved [128]. The cytotoxicity of the AuNPs was negligible in a broad concentration range. Other researchers used an L. edodes fruit body extract to produce AuNPs of various shapes [145]. Basu et al. obtained variously shaped gold nanoparticles using a Tricholoma crassum mycelial extract [151]. They showed that particle size could be altered by changing synthesis parameters such as temperature and substrate and precursor concentrations. A mixture of triangular, spherical and irregular shapes with an average size of 74.32 nm was fabricated using a Flammulina velutipes fruit body extract [138]. A chaga (Inonotus obliquus) medicinal mushroom extract induced the formation of mostly spherical AuNPs with a size below 20 nm [143]. These AuNPs are promising dual-modal (chemo-photothermal) therapeutic candidates for anticancer applications. The production of AuNPs by a Coprinus comatus fruit body extract and the effect of UV irradiation at different times on nanoparticle size were investigated [137]. Gold nanospheres were also obtained using fruit body extracts of Cantharellus sp. (average particle size of 60.6 nm) [136] and Laetiporus versisporus (average particle size of 10 nm) [144].

2.3. Mycosynthesis of Platinum Nanoparticles

Platinum nanoparticles (PtNPs) are of great interest in various fields of engineering and biomedicine owing to their unique physico-chemical (catalytic, magnetic and optical) and biological (antimicrobial, antioxidant, anticancer) properties ^{[154][155][156]}[154,155,156]. The mycosynthesis of PtNPs is much less studied, as compared to that of silver and gold. To date, the ability to form nanoparticles of this noble metal has been detected in several Ascomycota species (Table 3). . The extract containing intracellular components of fungal strains was obtained from a mechanically disrupted mycelium, while for the extract containing extracellular components of fungal strains, the biomass was extracted without disruption. The second method produced smaller particles. A number of researchers have studied the effect of various additional external physical influences on the fungi-mediated nanoparticle formation and developed combined methods of myconanosynthesis to improve AgNP characteristics. For example, UV radiation enhanced the characteristics of AgNPs obtained with an Agaricus bisporus pilei extract [89]. Microwave irritation enhanced the properties of AgNPs synthesized with the use of a Pleurotus sajor-caju fruit body extract [113]. AgNPs were synthesized from Pleurotus florida fruit body extracts using different electro-magnetic radiations, microwaves, visible light and UV rays [111]. Microwave irradiation led to the synthesis of monodisperse AgNPs of 10 nm size within 150 s of exposure, whereas visible light and UV radiation led to the synthesis of polydisperse AgNPs with inconsistent dimensions. Numerous studies have shown that mycosynthesized AgNPs have antibacterial, antifungal, anticancer, antioxidant, larvicidal and other properties, and the same nanoparticles can exhibit a wide range of biological activities. For example, silver nanospheres obtained by using Flammulina velutipes had bactericidal, fungicidal, anti-Alzheimer, anticancer, antioxidant and anti-diabetic activities, as well as good biocompatibility against human red blood cells [97]. Silver nanospheres produced using Aspergillus niger and Trichoderma longibrachiatum xylanases exhibited antibacterial, antifungal, antioxidant, anticoagulant, thrombolytic and dye-degrading activities [48]. All these properties offer great prospects for biomedical applications of mycogenic AgNPs.

2.2. Mycosynthesis of Gold Nanoparticles

Gold nanoparticles (AuNPs) have attracted attention owing to their unique optical, electronic, thermal, chemical and biological properties. They have been used in chemical and biological sensing, bio-imaging, nonlinear optics, catalysis, targeted drug delivery, gene delivery and as antimicrobial and antioxidant agents, as well as in cancer, Alzheimer’s, cardiovascular and infectious disease therapy ^{[129][130][131]}[129,130,131]. In the past two decades, the biological synthesis of gold nanoparticles by fungi has been studied almost as extensively as that of silver. The ability to form AuNPs has been found in dozens of micro- and macromycete species. The table shows the AuNP mycosynthesis data published in the past five years (Table 2). As can be seen from the table, AgNPs are commonly spherical in shape; irregular, oval, cubic, triangular, polygonal and other shapes are less common. Extracts from fruit bodies and mycelium are the most frequently used biological material for AgNP mycosynthesis, while culture liquids, biomass, living cultures and fungal metabolites of different purity (including enzymes, polysaccharides and phenolic compounds) are less commonly used. A number of researchers have screened fungal cultures to find the most promising ones for AgNP biofabrication. For example, Qu et al. studied 10 Trichoderma species and found that AgNPs obtained using different species differed in the degree of antimicrobial activity [42]. Other researchers found that among nine different fungi isolated from metal-rich sites, a strain of Penicillium janthinellum exhibited maximum metal tolerance capacity and AgNP-synthesizing ability [69]. The shape, size, homogeneity and stability of nanoparticles are influenced by the process conditions, the optimization of which can improve the quality of the obtained particles. For example, Mohanta et al. used various ratios of a Ganoderma sessiliforme mushroom extract to AgNO₃ for AgNP synthesis [101]. At a 0.5:10 ratio, nanoparticles formed very slowly; at a 1.5:10 ratio, the reaction was very rapid but nanoparticles formed large aggregates. The 1:10 ratio was optimal and allowed the authors to obtain nanoparticles with an average size of 45 nm with antimicrobial and antioxidant activity. In another study, the utilization of a Ganoderma lucidum fruit body extract was scrutinized under different operational conditions including the AgNO₃:extract ratio, reaction time and temperature to establish an effective myconanosynthesis method with a high yield rate and nanoparticle stabilization [125]. Vetchinkina et al. studied the effect of the Lentinus edodes culture age and stage of ontogenesis on the biogenic AgNP synthesis using culture liquids of different ages [41] and extracts obtained from the different morphological structures of L. edodes [128]. Parametric optimization, including the concentration of AgNO₃, fungal biomass, ratio of cell filtrate to AgNO₃, pH, reaction time and presence of light, was performed for the rapid synthesis of silver nanoparticles by Penicillium polonicum [72]. For Trichoderma harzianum and Ganoderma sessile, different methods of mycelial extraction for silver mycosynthesis were compared [82] As with silver, the ability to biosynthesize gold nanoparticles has been studied mainly in two groups of fungi—ascomycetes and basidiomycetes. Mycogenic AuNPs most often have a spherical shape, but triangular, hexagonal, cubic, irregular and other shapes were also found. Needle- and flower-like nanostructures with a spindle shape were obtained using Fusarium solani biomass extract [140]. Spherical and hexagonal particles 22–30 nm in size were mycosynthesized with the use of Fusarium oxysporum PtNP biosynthesis has been best studied in F. oxysporum. Riddin et al. showed that the mycelial biomass of F. oxysporum is capable of producing nanoparticles of various shapes (hexagons, pentagons, circles, squares, rectangles) and sizes (10–100 nm) and determined the optimal conditions (pH, temperature and concentration of the precursor compound H₂PtCl₆) for maximum nanoparticle yield [158]. Nanoparticles were formed both extracellularly and intracellularly as well as on the hyphae surface, but only the extracellular production of nanoparticles proved to be statistically significant. In further studies [161], a hydrogenase with Pt(IV)-reductase activity was isolated from this strain of F. oxysporum. It was shown that the bioreduction of platinum salt by hydrogenase takes place by a passive process and not an active one as previously understood. PtNPs formed by cell-free mycelial extract and purified hydrogenase differed in size and shape. The particles formed with the extract were irregular in shape, with an average nanoparticle size of 30–40 nm. Circular, triangular, pentagonal and hexagonal nanoparticles, often appearing as nanoplates, with a mean size range of 40–60 nm, were formed using the enzyme. It was found that the oxidation state of the platinum salt also plays an important role in nanoparticle formation [160]. When PtCl₂ was used as a precursor, large (100–180 nm) nanoparticles of predominantly rectangular and triangular shape forming aggregates were biosynthesized with F. oxysporum hydrogenase. Bioreduction of H₂PtCl₆ produced spherical monodisperse nanoparticles varying in size with the mean nanoparticle size between 100 and 140 nm. Syed and Ahmad were able to produce spherical PtNPs with a diameter of 15–30 nm using F. oxysporum mycelial biomass [161]. The particles were formed extracellularly and were capped by natural proteins secreted by the fungus and therefore did not require the addition of stabilizing agents. Gupta and Chundawat obtained face-centered cubic nanoparticles with an average size of 25 nm with antimicrobial and photocatalytic activity using F. oxysporum filtrate [162]. The use of Penicillium chrysogenum culture filtrate made it possible to obtain highly dispersed non-aggregating platinum nanospheres (5–40 nm) [164]. Another ascomycete in which the ability to synthesize platinum nanoparticles has been found is Neurospora crassa [163]. Incubation of mycelial biomass with H₂PtCl₆ produced extracellular PtNPs (4–35 nm in diameter) and spherical nanoaggregates (20–110 nm in diameter). Using a mycelial extract from the same fungi, round single-crystal nanoagglomerates with diameters of 17 to 76 nm were obtained, containing individual single crystals of approximately 2–3 nm in diameter. Nanoplatinum was also obtained using the culture filtrate of the phytopathogenic fungus Alternaria alternata [157]. The particles were found to be irregular in shape presenting an overall quasi-spherical, rectangular, tetrahedral and hexagonal as well as polygonal morphology. Their size varied in the range of 50–315 nm with an average size of 135 nm. Borse et al. investigated the production of platinum nanospheres using cell-free extract of Saccharomyces boulardii yeast biomass and the effect of parameters such as the concentration of H₂PtCl₆, temperature, pH, reaction time and cell concentration [165]. A cell mass concentration of 500 mg/ mLin, the presence of 0.5 mM chloroplatinic acid at 35 °C, pH 7 and 200 rpm for 36 h showed maximum PtNP synthesis. Under these conditions, platinum nanospheres (80–150 nm) with anticancer activity were formed. It was also shown that nanoparticles were formed intracellularly when whole yeast cells were incubated with H₂PtCl₆.