Bright Side of Fusarium oxysporum: Secondary Metabolites Bioactivities and Industrial Relevance in Biotechnology and Nanotechnology

Fungi have been assured to be one of the wealthiest pools of bio-metabolites with remarkable potential for discovering new drugs. The pathogenic fungi, Fusarium oxysporum affects many valuable trees and crops all over the world, producing wilt. This fungus is a source of different enzymes that have variable industrial and biotechnological applications. Additionally, it is widely employed for the synthesis of different types of metal nanoparticles with various biotechnological, pharmaceutical, industrial, and medicinal applications. Moreover, it possesses a mysterious capacity to produce a wide array of metabolites with a broad spectrum of bioactivities such as alkaloids, jasmonates, anthranilates, cyclic peptides, cyclic depsipeptides, xanthones, quinones, and terpenoids. Therefore, this review will cover the previously reported data on F. oxysporum, especially its metabolites and their bioactivities, as well as industrial relevance in biotechnology and nanotechnology in the period from 1967 to 2021. In this work, 180 metabolites have been listed and 203 references have been cited.


Nanotechnological Applications
Nanotechnology holds promise in the medicine, agriculture, and pharmaceutical industries [112]. Natural nanostructures have gained more attention due to the wide spectrum of bioactivities and fewer animals, humans, and environmental toxicity. The microbial synthesis of nanoparticles is an approach of green chemistry that combines both nanotechnology and microbial biotechnology [113]. Metals nanoparticles are increasingly used in various biotechnological, pharmaceutical, and medicinal applications, including drug delivery, gene transfer, insect-pests management in agriculture, and bioelectronics devices fabrication, as well as antibacterial agents towards many pathogenic bacteria, including the MDR (multidrug-resistant) strains [114][115][116].

Metal Nanoparticles
Several studies reported the synthesis and characterization of metal nanoparticles (NPs) using F. oxysporum, as well as their bioactivities. Additionally, some studies dealt with optimizing the conditions for the synthesis of NPs by F. oxysporum, including temperature, media, pH, salt concentration, light intensity, the volume of filtrate, and biomass quantity [44,47,50,51,54,55]. Marcato et al., synthesized AgNPs (silver nanoparticles) using F. oxysporum. The incorporation of these NPs in cotton cloth was found to exhibit a bactericidal effect towards S. aureus, leading to its sterilization [50]. Ishida et al., synthesized AgNPs using F. oxysporum aqueous extract that showed significant antifungal potential towards Cryptococcus and Candida (MIC values ≤ 1.68 µg/mL) [51]. Moreover, it was found that the biosynthesized AgNPs by two F. oxysporum isolates exhibited higher antibacterial potential towards human-pathogenic bacteria; E. coli, Proteus vulgaris, S. aureus, and K. pneumonia than the used antibiotics. These AgNPs could be favorable antibacterial agents, especially towards MDR bacteria [44]. Ahmed et al., synthesized AgNPs using F. oxysporum, which inhibited some MDR species of Staphylococcus and Enterobacteriaceae (conc. 50% v/v), as well as Candida krusei and C. albicans, suggesting that they might be potential alternatives to antibiotics [46]. The in-silico and in-vitro studies demonstrated the immense antibacterial potential of F. oxysporum's AgNPs against P. aeruginosa and E. coli [45]. The AgNPs synthesized using nitrate reductase purified from F. oxysporum IRAN-31C showed potent antimicrobial potential towards a wide array of human pathogenic bacteria and fungi in the disk diffusion method [117]. A study by Ballottin et al., revealed that the cotton fibers impregnated with biogenic AgNPs synthesized from F. oxysporum filtrate solution possessed potent antimicrobial potential even after repeated mechanical washing cycles. This might highlight the potential use of biogenic AgNPs as an antiseptic in textiles for medical applications [118].
Moreover, a study by Hamedi et al., revealed that the existence of ammonium lowered the productivity of AgNPs using F. oxysporum cell-free filtrate and prohibited the nitrate reductase enzyme secretion [119]. Longhi et al., reported that the combination of AgNPs synthesized using F. oxysporum with FLC (fluconazole) reduced the MIC of FLC around 16 to 64 times towards planktonic cells of C. albicans and induced a significant dose-dependent inhibition of both initial and mature biofilms of FLC-resistant C. albicans. Therefore, these AgNPs could represent a new strategy for treating FLC-resistant C. albicans infections [49]. Additionally, the combination of simvastatin with these AgNPs demonstrated antibacterial activity towards E. coli-producing ESBL (extended-spectrum β-lactamase) and MRSA (methicillin-resistant S. aureus). This could be a great future alternative in bacterial infection control, where smaller doses of these AgNPs are required with the same antibacterial activity [120]. Besides, its combination with polymyxin B showed a 16-fold reduction of the MIC of polymyxin B and decreased carbapenem-resistant Acinetobacter baumannii viability with additive and synergic effects, as well as significantly reduced cytotoxicity towards mammalian Vero cells, indicating its pharmacological safety [121]. The AgNPs synthesized with F. oxysporum f.sp. pisi were found to have moderate adulticidal potential on Culex quinquefasciatus (vector of filariasis) (LC 50 0.4, LC 99 4.8, and LC 90 4 µL/cm 2 ) after 24 h exposure [122]. The synthesized AgNPs using F. oxysporum aqueous extract had anticancer potential towards MCF7 (IC 50 14 µg/mL) that was characterized using CLSM (confocal laser scanning microscopic) technique [123]. Bawskar et al. stated that the biosynthesized AgNPs using F. oxysporum possessed more potent antibacterial potential towards E. coli and S. aureus than chemo-synthesized AgNPs that may be due to the protein capping and their mode of entry into the bacterial cell, which encouraged biosynthetic method over the chemosynthetic one in AgNPs synthesis [124]. Two types of AgNPs, phyto-synthesized and myco-synthesized NPs were biosynthesized by AgNO 3 reduction with Azadirachta indica extract and F. oxysporum cell filtrate, respectively that possessed lower cytotoxic potential on C26 and HaCaT cell lines as compared with citrate coated AgNPs [125]. Santos et al. proved that F. oxysporum-biosynthesized AgNPs without pluronic F68 (stabilizing agent) had high antibacterial potential towards E. coli, P. aeruginosa, and S. aureus. On the contrary, chemo-synthesized AgNPs exhibited synergism in antibacterial activity in the presence of pluronic F68 [126]. Streptococcus agalactiae is an important cause of invasive diseases, mainly in newborns, pregnant women, and elderly individuals [127]. The combination of F. oxysporum-produced AgNPs (AgNPbio) and eugenol led to a remarkable synergistic effect and significant reduction of the MIC values of both eugenol and AgNPbio towards planktonic cells of S. agalactiae [127]. Thakker et al., reported the synthesis of GNPs (gold nanoparticles) using F. oxysporum f. sp. cubense JT1 that showed antibacterial potential versus Pseudomonas sp. [128]. Moreover, the conjugated GNPs with tetracycline demonstrated powerful antibacterial activity against Gram-negative and -positive bacteria in comparison to tetracycline and free GNPs. Therefore, tetracycline conjugation with these GNPs enhanced the antibacterial potential, which may have significant therapeutic applications [129]. Yahyaei and Pourali studied the conjugation of GNPs with chemotherapeutic agents such as paclitaxel, tamoxifen, and capecitabine. Moreover, the cytotoxic effect of conjugated GNPs was assessed towards MCF7 and AGS cell lines, using MTT assay. Unlike the paclitaxel conjugated GNPs, the tamoxifen and capecitabine conjugated GNPs revealed no toxic effects due to their low half-lives and deactivation [130]. Further, Syed and Ahmad reported the synthesis of stable extracellular platinum nanoparticles, using F. oxysporum [131]. CdSe (cadmium/selenium) quantum dots are often used in industry as fluorescent materials. Kumar et al., and Yamaguchi et al., reported the synthesis of highly luminescent CdSe quantum dots by F. oxysporum [132,133]. In 2013, Syed and Ahmad synthesized highly fluorescent CdTe quantum dots using F. oxysporum at ambient conditions by the reaction with a mixture of TeCl 4 and CdCl 2 . These nanoparticles exhibited antibacterial potential towards Gram-negative and -positive bacteria [53]. Riddin et al., analyzed the biosynthesized platinum (Pt) nanoparticles by F. oxysporum f. sp. lycopersici at both intercellular and extracellular levels. It was found that only the extracellular nanoparticle production was proved to be statistically significant with a yield of 4.85 mg/L [134].

Metal Sulfide Nanoparticles
In addition, Q-state CdS NPs were biosynthesized by the reaction of aqueous CdSO 4 solution with F. oxysporum [135]. The chemically-synthesized CdSQDs inhibited E. coli cell proliferation in a dose-dependent manner, unlike the biogenic CdSQDs synthesized by F. oxysporum f. sp. lycopersici, which showed an antibacterial potential only at high concentration. Additionally, only the biogenic CdSQDs showed no inhibition on seed germination after incubation of biogenic and chemical CdSQDs with Lactuca sativa seeds [43]. Bi 2 S 3 (bismuth sulfide) NPs have significantly varied applications, including photodiode arrays, photovoltaic materials, and bio-imaging. Uddin et al., synthesized a highly fluorescent, natural protein capped Bi 2 S 3 NPs by subjecting F. oxysporum to bismuth nitrate penta-hydrate, along with sodium sulfite under ambient conditions of pressure, temperature, and pH. It was found that they were fundamentally much more fluorescent than fluorophores (toxic fluorescent chemical compounds), which are largely utilized in immunohistochemistry, imaging, and biochemistry [48].

Metal Oxide Nanoparticles
It was reported that F. oxysporum might have vast commercial implications in low-cost, room-temperature, ecofriendly syntheses of technologically significant oxide nanomaterials from available potentially cheap naturally raw materials [136]. F. oxysporum rapidly biotransformed the naturally occurring amorphous biosilica in rice husk into crystalline silica NPs. This could lead to an economically viable and energy-conserving green approach toward the large-scale synthesis of oxide nanomaterials [136]. Moreover, the mesophilic F. oxysporum bioleached Fly-ash at ambient conditions produced highly stable, crystalline, fluorescent, water-soluble, and protein-capped silica nanoparticles [52]. It was found that F. oxysporum enriched zirconia in zircon sand by a process of selective extracellular bioleaching of silica nanoparticles. It was proposed that the fungal enzymes specifically hydrolyzed the silicates in the sand to form silicic acid, which on condensation by certain other fungal enzymes resulted in silica nanoparticles synthesis at room temperature [136]. A water dispersible and thermo-stable Ag/Ag 2 O NPs were produced from silver oxide micro-powder using F. oxysporum. These Ag/Ag 2 O NPs may become a potential candidate for enzyme-free glucose determination and exhibited catalytic potency for MB (methylene blue) degradation in presence of NaBH 4 (reducing agent). Additionally, they showed an excellent antimicrobial potential against A. niger and B. subtilis [137].

Biotechnological and Industrial Relevance of F. oxysporum
F. oxysporum is a wealthy source of enzymes with significant biotechnological and industrial potential. In various studies, F. oxysporum demonstrated a remarkably high enzymatic performance and the ability to degrade different biomasses. Herein, the reported enzymes from F. oxysporum and their industrial and biotechnological applications are highlighted.

Glycoside Hydrolases
Cellulases are accountable for cellulose hydrolysis, including β-1,4-endoglucanase, cellobiohydrolase, and β-glucosidases (BGL), which catalyze the hydrolysis of aryl-and alkyl-β-glucosides, as well as oligosaccharides and diglucosides [1,3,5]. Cellulases' preparations have been added to the ruminant animals' diets to stimulate feed processing and fiber digestion to increase the extent and rate of digestion [138]. Zhao et al., purified extracellular BGL from F. oxysporum that had high acid stability (pH 3) and cellobiose hydrolytic activity relative to Celluclast ® (commercial cellulase). Its supplementation also released more reducing sugars (330 mg/g substrate) from cellulose, in comparison to Novozymes (commercial BGL, 267 mg/g substrate) under simulated gastric conditions. Thus, it could be a good source for a new commercial BGL for improving the feed and food quality in the animal feed industry and could be used in combination with Celluclast for industrial applications that required degradation of cellulose at acidic pH [37].
Fucose is a low abundant deoxy-hexose sugar, usually attached to the non-reducing ends of oligolipids, oligosaccharides, and other glycoconjugates (e.g., immunoglobulins, glycoproteins, blood group substances, and mucins). Besides, it is a component of marine algal polysaccharides, human milk oligosaccharides, and plant gums [139]. FUC (α-Lfucosidase) a glycoside hydrolase, catalyzes the breakdown of the terminal α-L-fucosidic bonds. It has remarkable roles in various bioprocesses, and it is used as a marker for hepatocellular carcinoma detection and structural analyses of complex natural products [140]. F. oxysporum produced FUC in large amounts through induction by L-fucose. This enzyme hydrolyzed p-nitrophenyl α-L-fucoside (synthetic substrate) like marine gastropod and mammalian enzymes, thus it could replace these enzymes. It had beneficial use as an analytical tool for the structural elucidation of complex carbohydrates and oligosaccharides [41]. Additionally, Yano et al., purified a novel FUC from F. oxysporum culture broth. Besides nitrophenyl compounds, this enzyme had a novel substrate specificity. It could hydrolyze porcine mucin and blood group substances [42].
α-D-Galactopyranosidase (GPase) is a glycoside hydrolase that hydrolyzes the αgalactopyranosyl linkages at non-reducing ends of sugar chains. They are utilized for various applications, including eliminating non-digestible oligosaccharides such as stachyose in legume products and soybean, improving the digestibility of animal feed, and increasing the yield and quality of sucrose in sugar refineries [141]. FoAP1 and FoAP2 are two bifunctional enzymes that were isolated and characterized from the culture supernatant of F. oxysporum 12S, possessing GPase (α-D-galactopyranosidase)/APase (β-L-arabinopyranosidase) activities in a ratio 1.7 and 0.2, respectively using PNP-α-D-Galp (para-nitrophenyl α-D-galactopyranoside) and PNP-β-L-Arap (para-nitrophenyl α-l-arabinopyranoside) as substrates [38]. A novel GPase, FoGP1 was purified from F. oxysporum, exhibiting degrading activity with terminal α-1,3-galactosyl linkages in gum Arabic side chains. Therefore, it might be used for improving gum Arabic physical properties, which is an industrially important polysaccharide used as a coating agent and an emulsion stabilizer [34].
Xylan is one of the most abundant carbohydrates on earth. Its complete degradation is accomplished by the action of various enzymes such as β-D-xylosidases and endoβ-1,4-xylanases. Alconada and Martínez characterized extracellular β-xylosidase and endo-1,4-β-xylanase from F. oxysporum f. sp. melonis, growing in a medium containing oat spelt xylan. The latter had a high affinity towards oat spelt xylan [142]. Additionally, FoXyn10a, new GH10 xylanase was purified and structurally characterized from F. oxysporum [143]. Anasontzis et al., reported that the constitutive homologous overexpression of the endo-xylanase in F. oxysporum increased ethanol production during CBP of lignocellulosics [144]. Xyn11a is an endo-1,4-β-xylanase gene from F. oxysporum, belonging to the fungal glycosyl hydrolase family 11 (GH-11) that was cloned and expressed in Pichia pastoris. Recombinant P. pastoris possessed efficient xylanase secreting potential and a high level of enzymatic activity under methanol induction [145]. Gómez et al., identified Xyl2, an endo-β-1,4-xylanase from F. oxysporum. It was highly active at alkaline pH and its immobilization on certain supports significantly increased its thermal stability. Together, these properties rendered Xyl2, an attractive biocatalyst for the sustainable industrial degradation of xylan [146]. Najjarzadeh et al., compared xylanase production by different inducers, such as lactose, sophorose, xylooligosaccharides, and cellooligosaccharides in F. oxysporum f. sp. lycopersici. It was found that xylooligo-saccharides were more effective than other substrates at the induction of β-xylosidases and endoxylanases. Moreover, xylotetraose, xylohexaose, and xylobiose were the best inducers of endoxylanase, extracellular β-xylosidase, and cell-bound β-xylosidase, respectively [147].

Nitrilases
Microbial nitrilases are biocatalysts of remarkable organic importance in terms of nitrile conversion. Nitrile compounds include aromatic and simple aliphatic metabolites, cyano-lipids, and cyano-glucosides which serve as key intermediates and compounds in various biochemical pathways [148]. Processes involving enzymatic conversion of nitrile substrates to higher value amides and carboxylic acid groups are preferred over the chemical synthesis for their production of fewer harmful reaction by-products and greater reaction specificity [149]. Industrial application of nitrile-converting enzymes includes acrylamide and nicotinamide production [35]. The newly isolated nitrilase from F. oxysporum f. sp. lycopercisi ED-3 strain had a wide substrate specificity toward ortho-substituted heterocyclic, aliphatic, and aromatic nitriles and had optimal activity at temperature 50 • C and pH 7.0 [35].

Nitric Oxide Reductases
Denitrification is a substantial process in the nitrogen cycle, which involves the reduction of NO −2 and/or NO 3− to either N 2 O or N 2 . This process can be performed by many bacteria, as well as fungi through a series of consecutive metallo-enzyme-catalyzed chemical reactions [150]. It is a reversible process of nitrogen fixation, where it carries back the fixed N 2 to the atmosphere. It was found that the main source of global N 2 O emissions is the microbial activities of denitrification and nitrification [151]. Therefore, controlling microbial denitrification is most important for N 2 O emission reduction [152]. It was reported that F. oxysporum exhibited a distinct denitrifying potential that resulted in the anaerobic evolution of N 2 O from NO −2 and NO 3− [153]. Further, a nitric oxide reductase (NOR), cytochrome P450nor, belonging to P450 superfamily was purified from F. oxysporum, which exhibited remarkable NO reduction potential [36,150,152]. It showed a unique bio-function compared with other usual P450s. While the usual P450s are involved in metabolizing various biological substances through mono-oxygenation reaction using O 2 , P450nor catalyzed the NO reduction but not the mono-oxygenation [150,154]. Shoun and Tanimoto described a heme-thiolate protein or P450 from F. oxysporum that was involved in the NO (nitrogen monoxide) reduction to N 2 O (dinitrogen oxide) [153]. In contrast to other bacterial cytochrome bc-containing NO reductases, it did not need a flavoprotein for electron transfer from NADH to the heme, but it utilized NADH directly for the reduction process [155]. Additionally, Daiber et al., identified P450nor (cytochrome P450 NADH-NO), a heme-thiolate protein that catalyzed the reduction of two NO molecules to N 2 O. P450nor was observed to have a remarkable role in protecting the fungus from NO inhibition of mitochondria [156].

Cutinases
Esters having a chain of fewer than 10 carbon atoms are used as flavor compounds in the pharmaceutical, cosmetic, and food industries [157]. Natural synthesis of flavor compounds takes place either by enzymatic bioconversion or by microorganisms, the former path was found to be an easier and more suitable method [157]. The high demand of various industries for fatty acid ester leads to possible growth of the market to $2.44 billion by 2022, from $1.83 billion in 2014 [33,158]. Enzyme immobilization increases their stability and allows their easy separation from the reaction and reuse to overcome the drawbacks of utilizing enzymes such as low operational or storage stability, and/or heat and organic solvent sensitivity [159]. The imFocut5a a CLEAs (cross-linked enzyme aggregates) was produced from a crude F. oxysporum cutinase preparation. This immobilized cutinase possessed a remarkable thermo-stability and was able to synthesize butyl butyrate (pineapple flavor) at a high yield of bioconversion (99%) through the trans-esterification of vinyl butyrate with butanol. This bioconversion presented an eco-friendly and sustainable production of natural flavor compounds, underpinning its industrial potential for use in food bioprocesses [33]. FoCut5a, a cutinase was purified from F. oxysporum and expressed either in the periplasm or cytoplasm of E. coli BL21. It could hydrolyze PET (polyethylene terephthalate) and synthetic polymers. Therefore, it could be used in industrial applications as a biocatalyst for the eco-friendly treatment of synthetic polymers [160].

Fructosyl Amino Acid Oxidases
Glycation is the non-enzymatic glycosylation of proteins due to the condensation of reducing sugars such as glucose with the proteins (α-or ε-amino groups) to form a Schiff's base [161]. Glycation leads to browning of foods during long-term storage, which represents a problem in the food industry [39]. Glycation of hemoglobin, blood proteins, and albumin was found to be enhanced in diabetic patients. Glycated proteins, particularly glycated hemoglobin A1c, are important markers for assessing the effectiveness of anti-diabetic agents. Fructosyl amino acid oxidase (FAOD) based assays have become an attractive alternative to conventional detection methods for measuring glycated proteins [161]. Sakai et al., purified FLO (fructosyl lysine oxidase) from F. oxysporum S-lF4 that acted against fructosyl poly L-lysine. FLOD could be used for measuring glycated proteins such as glycated albumin in the serum [39].

Lipoxygenase
Lipoxygenase (LOX) is a dioxygenase that catalyzes the hydro-peroxidation of polyunsaturated fatty acids such as arachidonic and linoleic acids [162]. It is expressed in epithelial, tumor, and immune cells that have various physiological functions such as skin disorders, inflammation, and tumorigenesis [163]. Bisakowski et al., extracted and purified LOX from F. oxysporum that shared many of the characteristics with LOXs reported from other sources such as substrate specificity, pH, enzyme inhibition, activation, and other kinetic studies [40].

Laccases
Laccases are belonging to oxidoreductases that catalyze the O 2 reduction to H 2 O with simultaneous organic substrates oxidation. They oxidize phenolic substrates but are also able to oxidize bigger or non-phenolic substrates by LMS (laccase-mediator system), where a small phenolic compound acts as a mediator [164]. Additionally, they have been reported as potential lignin-degrading enzymes and as solubilizing agents [165,166]. They have attracted great interest because of their wide applications in diverse biotechnological and industrial fields such as textile dye decolorization, pulp bleaching, organic synthesis, bioremediation, and detoxification of environmental pollutants, delignification, or biofuel production [1,3,5].
Kwiatos et al., expressed F. oxysporum Gr2 laccase in Saccharomyces cerevisiae and engineered it for getting higher effect against 2,6-dimethoxyphenol and higher expression levels. The resulted laccase had a promising potential for different industrial uses such as solubilization of brown coal, which is a clean coal technology, aiming at converting lignite to its cleaner form [164]. In 2018, Kwiatos et al., reported that F. oxysporum LOCK-1134 isolated from brown coal, efficiently bio-solubilized lignite, producing liquefied products that had over 99% less Hg and 50% less sulfur than the crude coal. Additionally, its laccase was expressed in Pichia pastoris. The resulted novel laccase improved the biodegradation process in presence of LMS. It released fulvic and humic acids from liquefied coal. The latter are environmentally friendly fertilizers that possessed a stimulating influence on crop growth [165].

Aromatic Carboxylic Acid Decarboxylases
The non-oxidative aromatic carboxylic acid decarboxylases catalyze the reversible decarboxylation of phenolic carboxylic acids. Therefore, they are useful biocatalysts for preparing high-value phenolic compounds by the decarboxylation of phenolic carboxylic acids derived from lignin, which opens up a new prospect for high-value utilization of the world second most abundant organic substance [167,168]. Song et al., characterized 2,3-DHBD_Fo, a 2,3-dihydroxybenzoic acid decarboxylase from F. oxysporum that possessed a relatively high catalytic decarboxylation efficiency for DHBA (2,3-dihydroxybenzoic acid) and catechol, hence it had a different substrate spectrum from other benzoic acid decarboxylases [167].

Keratinases
Keratins are complex proteins that formed of β-sheets and α-helix structures. They are commonly found in agro-industrial residues such as swine hair and chicken feathers. Keratinous wastes are treated in non-eco-friendly ways, including landfills and incinerators [169,170]. F. oxysporum isolated from chicken feathers showed potential for keratinase production that had the highest degradation percentage (59.20% w/w) in swine hair [169].

Phospholipase B
Phospholipase B (PLB) hydrolyzes the phospholipid acyl groups to produce fatty acids and phosphoglycerates [171]. PLB is utilized to produce beneficial phospholipid derivatives, reduce food's cholesterol content, and refine vegetable oils, especially in terms of crude oil degumming [172]. Su et al., characterized a putative lipase from F. oxysporum NCBI-EGU84973.1 that was expressed in P. pastoris and classified as a PLB. It had phospholipids hydrolyzing potential greater than its lipase capacity where it hydrolyzed the fatty acyl ester bond at the sn-1 and -2 positions of the phospholipids and reduced the oil phosphorus contents. This proved the potential industrial use of this PLB in oil degumming applications [172].
3.1.11. Triosephosphate Isomerase TPI (triosephosphate isomerase) is a glycolysis enzyme that catalyzes the reversible isomerization between DHAP (dihydroxyactetone-3-phosphate) and GAP (glyceraldehyde-3-phosphate) [173]. Therefore, TPI is essential for pathogenic organisms to get the energy needed for survival and infection. Hernández-Ochoa et al., isolated, cloned, and overexpressed Tpi gene from F. oxysporum isolated from a wild species collected from a bean crop. They purified FoxTPI recombinant protein that had the TPIs classical topology conserved in other organisms [14].

Applications of F. oxysporum
Biodiesel (biofuel) is obtained from renewable sources such as animal fat or vegetable oil by trans-esterification of triglycerides to give fatty acid alkyl esters [174]. It is used as a full or partial substitute for petrol diesel in combustion engines [175]. Its production attracts attention worldwide due to the environmental benefits such as biodegradation that reduced the emission of sulfur and aromatic hydrocarbons during fuel combustion and decreased emission of CO 2 , CO, and particulate materials. The accumulated lipids in microorganisms such as algae, fungi, and bacteria are mainly triacylglycerols (TAG) that are utilized as metabolites for biodiesel production [32]. F. oxysporum NRC2017 isolated from Egyptian soil had remarkable lipid producing capacity (55.2%). It showed the highest lipid accumulation 98.3 mg/g in the presence of baggase and its fatty acids were found to be suitable for biodiesel production based on GC analysis [32].
On earth, the most abundant source of biomass is lignocellulosic material that includes agricultural residues, grasses, wood, or any non-food-plant sources. Its microbial fermentation produces ethanol and other solvents that represent an alternative path for wastes treatment and production of fuel additives and chemical feedstocks. F. oxysporum was found to have the potential for converting D-xylose, as well as cellulose to ethanol in a one-step process, indicating its capacity for ethanol production [176].
Bioethanol production is a harsh operational process that needs potent biocatalysts. CBP (consolidated bioprocessing) is an economical and efficient method of manufacturing bioethanol from lignocellulose. CBP integrates the fermentation and hydrolysis steps into a single process, leading to a significant reduction in the steps of the biorefining process. Ali et al., reported that F. oxysporum had a high potential for CBP of lignocellulose to bioethanol and it could be a commercially competitive CBP agent [177]. It was observed a significant inter-strain divergence regarding the capacity of different F. oxysporum strains to produce alcohol from wheat straw [178]. Nait M'Barek et al., assessed the potential of F. oxysporum for bioethanol production from non-valorized OMW (olive mill waste) using CBP. It showed maximum bioethanol yield and production of 0.84 g/g and 2.47 g/L, respectively, indicating its importance as a bio-agent for single-pot local bio-refinery [179]. Moreover, F. oxysporum BN converted imidazolium-based ionic liquid (IL)-pretreated rice straw to bioethanol via CBP with 64.2% of the theoretical yield of 0.125 g ethanol/g rice straw [164]. It secreted a novel IL-tolerant cellulase that can direct the conversion of ILpretreated lignocellulose residue to ethanol, which had a significant potential to bring a breakthrough in commercial ethanol production by the reduction of the overall cost [180].

Anthranilates
Anthranilates are derivatives of anthranilic acid that constitute an important part of several bio-metabolites and serve as a scaffold for developing remarkable pharmaceuticals for the management of the pathogenesis and pathophysiology of diverse disorders. They possessed impressive bioactivates such as antiviral, antimicrobial, insecticidal, antiinflammatory, anti-diabetic, and anticancer [181]. Compounds 1-21 are anthranilic acid derivatives that had been purified and characterized only from F. oxysporum f. sp. dianthi extracts using HPLC-, pyrolysis-, and HR-MS [56]. It was reported that the anthranilic acid derivatives are originated from 2 that is formed from benzoate and anthranilate [182]. Subsequently, it undergoes hydroxylation at C-2 to produce dianthalexin, hydroxylation at C-4 to yield 3 or 5, and methylation to 9 or 8. Moreover, 20 and 19 are produced from 5 and 3 [56,183] (Scheme 1, Figure 1). Moreover, F. oxysporum BN converted imidazolium-based ionic liquid (IL)-pretreated rice straw to bioethanol via CBP with 64.2% of the theoretical yield of 0.125 g ethanol/g rice straw [164]. It secreted a novel IL-tolerant cellulase that can direct the conversion of ILpretreated lignocellulose residue to ethanol, which had a significant potential to bring a breakthrough in commercial ethanol production by the reduction of the overall cost [180].

Anthranilates
Anthranilates are derivatives of anthranilic acid that constitute an important part of several bio-metabolites and serve as a scaffold for developing remarkable pharmaceuticals for the management of the pathogenesis and pathophysiology of diverse disorders. They possessed impressive bioactivates such as antiviral, antimicrobial, insecticidal, antiinflammatory, anti-diabetic, and anticancer [181]. Compounds 1-21 are anthranilic acid derivatives that had been purified and characterized only from F. oxysporum f. sp. dianthi extracts using HPLC-, pyrolysis-, and HR-MS [56]. It was reported that the anthranilic acid derivatives are originated from 2 that is formed from benzoate and anthranilate [182]. Subsequently, it undergoes hydroxylation at C-2′ to produce dianthalexin, hydroxylation at C-4 to yield 3 or 5, and methylation to 9 or 8. Moreover, 20 and 19 are produced from 5 and 3 [56,183] (Scheme 1, Figure 1). Scheme 1. Possible biosynthetic pathway for the formation of anthranilic acid derivatives [56,182,183].

Jasmonates
Jasmonates are lipid-based metabolites, possessing jasmonic acid (3-oxo-2-(pent-2enyl)cyclopentane acetic acid) framework that are found in fungi, bacteria, and plants [186,187]. In plants, they function as growth regulators and play major roles in the defense of plants against insects and diseases [187]. In addition, hydroxylated jasmonic acids, unsaturated or saturated, and cisor trans-configured elongated side-chain derivatives were reported from fungi. They can also form conjugates with amino acids such as isoleucine. In a study by Miersch et al., jasmonates derivatives 31-52 were purified from F. oxysporum f. sp. matthiolae using RP-18 Lichrolut, DEAE-Sephadex-A25, and 100-C18 Eurospher and characterized by GC-MS and HPLC [61] (Figures 3 and 4).

Cyclic Peptides and Depsipeptides
F. oxysporum yielded bioactive cyclic depsipeptides such as enniatins (ENs) and beauvericin (BEA, 99) (Figures 9 and 10). ENs are characterized by an alternating sequence of three D-α-hydroxyisovaleric acids and three N-methyl-L-amino acids in their structure. ENs H (96), I (97), and MK1688 (98), and BEA (99) were purified from F. oxysporum KFCC- Furthermore, it prohibited migration of MDA-MB-231 and PC-3M cells (conc. ranging from 3.0 to 4.0 µ M and 2.0 to 2.5, respectively) in the WHA (wound healing assay). NIH ImageJ software and WHA suggested that 99 was able to inhibit PC-3M and MDA-MB-231 migration at sub-lethal concentrations. Moreover, it possessed potent antiangiogenic potential at sub-lethal concentrations, as indicated by complete inhibition of HU-VEC-2 network formation at 3.0 µ M below IC25 (5.0 µ M) and IC50 (7.5 µ M) [64]. Cyclosporine A (100) was isolated from mycelia extract of non-pathogenic F. oxysporum S6 using reversed-phase silica gel and HPLC. It prohibited the growth and suppressed sclerotia formation of the phytopathogenic fungus Sclerotinia sclerotiorum with MIC 0.1 µ g/disc that made it suitable to be utilized as a bio-fungicide. Moreover, a remarkable increase in the number of surviving soybean plants was noted when F. oxysporum and S. sclerotiorum were inoculated together, in comparison to plants inoculated with S. sclerotiorum alone in the greenhouse assay. Hence, F. oxysporum could be a good biocontrol agent for S. sclero- onistic activity [86]. It is noteworthy to mention that some F. oxysporum strains could repress the growth of Pythium ultimum in cucumber [193] and affected S. sclerotiorum sclerotia germination [194].

Glucosylceramides
Glucosylceramides (GCs) are neutral glycosphingolipids, having glucose in 1-O-βglycosidic linkage with a ceramide [195]. Bernardino et al, isolated and purified the GCs, 102-104 from F. oxysporum. These GCs were assessed for their potential in inducing resistance in Nicotiana tabacum cv Xanthi plants against TMV (Tobacco mosaic virus) ( Figure  11). Spraying tobacco plants with GCs before virus infection reduced the incidence of necrotic lesions caused by TMV. After GCs treatment, the infected plants with the virus exhibited a reduction in HR (hypersensitive response) lesions, indicating GCs antiviral effect. The results revealed that GCs stimulated the early accumulation of H2O2 and superoxide radicals, which act as a plant immunity elicitor to combat diseases influencing the plants [87].  [64]. Cyclosporine A (100) was isolated from mycelia extract of non-pathogenic F. oxysporum S6 using reversedphase silica gel and HPLC. It prohibited the growth and suppressed sclerotia formation of the phytopathogenic fungus Sclerotinia sclerotiorum with MIC 0.1 µg/disc that made it suitable to be utilized as a bio-fungicide. Moreover, a remarkable increase in the number of surviving soybean plants was noted when F. oxysporum and S. sclerotiorum were inoculated together, in comparison to plants inoculated with S. sclerotiorum alone in the greenhouse assay. Hence, F. oxysporum could be a good biocontrol agent for S. sclerotiorum in soybean because of its metabolite 100 that was responsible for the in vitro antagonistic activity [86]. It is noteworthy to mention that some F. oxysporum strains could repress the growth of Pythium ultimum in cucumber [193] and affected S. sclerotiorum sclerotia germination [194].

Xanthone Derivatives
Bikaverin (119), intensively colored pigment was reported firstly from F. vasinfectum and F. lycopersici [85,96]. It belongs to the NRPKs (non-reducing polyketides) group that is produced by type I PKS [196,197]. By genetic engineering together with HPLC-HRMS and NMR tools, Arndt et al., identified the biosynthetic way for 119 and characterized its intermediates [198] (Scheme 3). Compounds 119 and 125 were isolated from F. oxysporum CECIS associated with Cylindropuntia echinocarpus (Figure 13). They were assessed for their cytotoxic activity towards a panel of four sentinel cancer cell lines by the MTT assay. Only 119 was cytotoxic towards NCI-H460, MIA Pa Ca-2, MCF-7, and SF-268 (IC 50 0.26-0.43 µM), compared with doxorubicin (IC 50 0.01-0.07 µM). It is noteworthy that 125 that lacks the C-6-OH group did not have cytotoxic activity even at concentrations of 4.0 and 2.0 µg/mL [64]. Further, 119 isolated from F. oxysporum f. sp. lycopersici as a purple-colored compound, exhibited a protective effect on oxidative stress and attenuated H 2 O 2 -induced neurotoxicity on human neuroblastoma SH-SY5Y cells. Pretreatment of neurons with 119 attenuated the H 2 O 2 (100 µM)-induced oxidative stress through improving the cell viability, antioxidant status, mitochondrial membrane integrity, and regulation of gene expression [95]. Therefore, it could be utilized as an alternative to some of the toxic synthetic antioxidants and a preventive agent against neurodegeneration [95]. Carmen et al., reported that bikaverin-contaminated products had no negative effect on human health [199]. Kundu et al., also purified 119 from F. oxysporum f. sp. ciceris ITCC-3636 EtOAc extract that had a weak anti-nemic potential towards M. incognita (LC 50 392.9 µg/mL) [88]. Additionally, Son et al., reported that 119 isolated from F. oxysporum EF119 showed antimicrobial activities against various phyto-pathogenic oomycetes and fungi. It suppressed the development of tomato late blight by 71% at conc. 300 µg/mL. Therefore, it may be used as a bio-control agent towards P. infestans-caused tomato late blight [66]. is produced by type I PKS [196,197]. By genetic engineering together with HPLC-HRMS and NMR tools, Arndt et al, identified the biosynthetic way for 119 and characterized its intermediates [198] (Scheme 3). Compounds 119 and 125 were isolated from F. oxysporum CECIS associated with Cylindropuntia echinocarpus (Figure 13). They were assessed for their cytotoxic activity towards a panel of four sentinel cancer cell lines by the MTT assay.  [95]. Therefore, it could be utilized as an alternative to some of the toxic synthetic antioxidants and a preventive agent against neurodegeneration [95]. Carmen et al, reported that bikaverin-contaminated products had no negative effect on human health [199]. Kundu et al, also purified 119 from F. oxysporum f. sp. ciceris ITCC-3636 EtOAc extract that had a weak anti-nemic potential towards M. incognita (LC50 392.9 µg/mL) [88]. Additionally, Son et al, reported that 119 isolated from F. oxysporum EF119 showed antimicrobial activities against various phyto-pathogenic oomycetes and fungi. It suppressed the development of tomato late blight by 71% at conc. 300 µ g/mL. Therefore, it may be used as a bio-control agent towards P. infestans-caused tomato late blight [66].

Scheme 3.
Putative biosynthetic pathway of 119 and its intermediates. The bold arrows represent the preferred pathway and dashed lines represented other possible reaction steps [196][197][198].

Pyran and Furan Derivatives
Chlamydosporol (158) a pyran lactone derivative was isolated from marine-mudflatderived F. oxysporum and assessed for its antibacterial potential towards MRSA and

Conclusions and Future Research Directions
Currently, more focus has been directed to fungi as they are a wealthy platform for the biosynthesis of a huge number of structural diverse metabolites. F. oxysporum is a species with great physiological and morphological variations and its wide-ranging existence in ecological activities worldwide indicates its profoundly diversified and significant role in nature. It can produce various bio-metabolites that may directly and indirectly be utilized as therapeutic agents for various health problems. In this work, 180 metabolites were reported from F. oxysporum in the period from 1967 to 2021. Alkaloids quinones, and jasmonates, and anthranilate derivatives represented the major metabolites that were isolated from this fungus ( Figure 19). Although, this big number of reported metabolites, few of them are evaluated for their bioactivities. The assessed activities of these metabolites were antimicrobial, cytotoxicity, nematicidal, antiviral, leishmanicidal, antiviral, and antioxidant. Additionally, there is a lack of pharmacological studies that focus on exploring the possible mechanisms of the active metabolites. In addition, the untested metabolites should be further explored for their possible bioactivities. Co-cultivation experiments should be employed to elicit the production of these metabolites. The discovery of the underlying biosynthetic pathways of these bio-metabolites is needed, which would allow the rational engineering or refactoring of these pathways for industrial purposes. Further, research for identifying the responsible biosynthetic genes for these metabolites may open the opportunity to explore the genetic potential of F. oxysporum for discovering novel metabolites by metabolic engineering that could result in more affordable and novel pharmaceutics and food additives. Moreover, studies on the structure-activity relationships and/or derivatization of these fungus metabolites should be carried out.
Although, the reported data revealed that F. oxysporum is widely employed for the synthesis of different types of metal nanoparticles that could have various biotechnological, agronomical, pharmaceutical, industrial, and medicinal applications. Many of these biosynthesized NPs possessed favorable antimicrobial potential, especially towards MDR microbes that can be potential alternatives to antibiotics. Further, it was found that the combination of NPs synthesized using F. oxysporum with antibiotics produced additive and synergic effects that could represent a new strategy for treating some antibiotics resistant strains and lower the doses of the used antibiotics. F. oxysporum might have vast commercial implications in low-cost, room-temperature, ecofriendly syntheses of technologically significant oxide nanomaterials from naturally available potentially cheap raw materials. However, the NPs synthesized from F. oxysporum are limited to metals and fewer metal oxides and sulfides. Therefore, future research should focus on developing protocols for implementing the biosynthesis of NPs of other metals, metal oxides, nitrides, and carbides. Research on the toxic effect of these NPs, as well as their effects on animals and human health and accumulation in the environment, is needed.
Additionally, in-vivo studies and clinical trials are needed to elaborate the exact mechanism responsible for their observed bioactivities. There is also a need for evaluating these NPs for their effectiveness towards various diseases, which can open in the future a new avenue in the biomedical field. More research is required for optimizing various reaction conditions to achieve better control over the shape, size, stability, and monodispersity of these NPs. F. oxysporum is considered as an efficient enzyme producer. Its enzymes have attracted great interest because of their possible applications in diverse biotechnological and industrial fields such as pharmaceutical, cosmetic, and food industries, organic synthesis, bioremediation, and detoxification of environmental pollutants, delignification, denitrification, or biofuel production. Additionally, they are involved in eco-friendly bioconversion processes of various substrates to highly valuable products that could be preferred more over the chemical synthesis. Research that focuses on engineering enzymes in such a way for maximum stability and activity under appropriate conditions is desirable. Recombinant DNA technology and engineering of proteins are required to improve the industrial production of these enzymes. Additionally, some F. oxysporum strains can be utilized as bio-control agents because of their ability to prohibit the growth of several fungal plant pathogens.

Conflicts of Interest:
The authors declare no conflict of interest.