Microorganisms as Nano-factories for the Synthesis of Metal Nanoparticles

Nanoparticles applications have revolutionized different areas of the research. These include medicine, surgery, drug delivery, wastewater treatment, agriculture, cancer therapy, etc. The use of nanoparticles is increasing day by day due to their promising characteristics. With the excessive use of the nanoparticles, their accumulation in the organisms and different environments have been reported. A very high increase in the accumulation and toxicity of nanoparticles has been reported in the last decade. Therefore, the nanoparticle research has now been shifted to find new techniques and methods to minimize the toxic effects of nanoparticles. In this context, the requirement of a safe design approach and the generation of fewer toxic nanoparticles are required. One of the eco-friendly approaches for safer nanoparticles synthesis is the use of living organisms for nanoparticles production. Microbes especially, bacteria, fungi, and yeasts, are considered safe, secure, and efficient systems for nanoparticle biosynthesis. This review is an attempt to understand the potential of microbes for the biosynthesis of nanoparticles. A R T I C L E H I S T O R Y Received: November 14, 2019 Revised: January 14, 2020 Accepted: February 12, 2020 DOI: 10.2174/2665980801999200507090343


INTRODUCTION
Nanoparticles have been used as promising candidates in diverse areas, including medicine. There are various strategies for the synthesis of nanoparticles. The most common strategy to produce nanoparticles involves the utilization of toxic organic solvents and reducing agents. These reagents are used to prevent agglomeration and defects in the nanoparticles [1]. The use of chemical methods is hazardous due to the use of toxic chemicals, such as hydrazine, dimethyl sulfoxide, acetonitrile, methanol, acetone, toluene, and potassium bitartrate [2,3]. These chemicals are responsible for genotoxicity, carcinogenicity, and cytotoxicity [4,5]. The use of these reagents is not environmentally-friendly. Therefore, various green matrices, such as DNA, bacteria, peptide, amino acid, protein, natural products, and polysaccharide were used for biomimetic synthesis [6][7][8][9][10][11][12]. With the increasing toxicity imposed by the nanoparticles on the living organisms. The necessity for the discovery of nanoparticle toxicity reduction methods is observed. In this connection, few strategies have been proposed. One of them is the use of eco-friendly approaches, such as biogenic nanoparticles. The use of biological organisms for the production of nanoparticles has emerged as a promising solution to the traditional nanoparticles production methods [13]. Fig. (1) presents different microorganisms, which can be used for the biogenic nanoparticles production.

BIOGENIC NANOPARTICLES
Biogenic nanoparticles are produced using living organisms. These nanoparticles are produced from bacteria, fungi, yeast, etc. They are alternative to the toxic nanoparticles, which are artificially synthesized. Biogenic nanoparticles have greater acceptance in the medical applications because of their advantages over the chemically produced nanoparticles [14]. Fig. (2) presents various advantages offered by biogenic nanoparticles. They offer biocompatibility, which gives them added stability and improved interaction with the pathogens involved [15]. Other advantages of using biogenic nanoparticles include low cost, reduced toxicity, and high efficiency [16]. Table 1 represents a few methods used in the biogenic nanoparticles synthesis along with their advantages   Fig. (2). Advantages of using biosynthesized nanoparticles. and disadvantages. In a study, the importance of biogenic silver nanoparticles was demonstrated. As compared to the synthetic silver nanoparticles, the silver nanoparticles produced from Aspergillus tubingensis were less harmful to the soil microbiota [17]. In the study, varying concentrations of Aspergillus tubingensis produced silver nanoparticles were used on rice seeds, soil microorganisms, and zebrafish. The study demonstrated better performance of the Aspergillus tubingensis produced biogenic silver nanoparticles over synthetic nanoparticles. A recent study demonstrated that magnetic nanoparticles could be synthesized in human stem cells from the nano-degradation products [18]. The study was performed by internalization in the mesenchyme stem cells. The nanoparticles were degraded during chondrogenesis, and stem cells were remagnetized. This remagnetization was the direct demonstration of magnetic nanoparticle's biosynthesis.

MICROBIAL SYNTHESIS OF NANOPARTICLES
Microorganisms, such as actinomycetes, bacteria, fungi, and yeast received attention since the past decade to produce nanoparticles [19]. Microorganisms are known to precipitate the nanoparticles by their metabolic activities [20]. Microorganism produces inorganic nanoparticles with immense applications in the biomedical fields [21]. Different microorganisms have been used for the synthesis of silver, gold, copper, and zinc nanoparticles [22][23][24][25]. Fig. (3) presents various applications involved in the use of biogenic nanoparticles. Table 2 represents microorganisms, along with the types of nanoparticles produced.

Nanoparticles Biosynthesis in Bacteria
Bacteria are a popular target for nanoparticles production due to their easier genetic manipulation and higher growth rates [26]. Recently, bacterium Desulfovibrio vulgaris has been used to produce nanoparticles useful in pharmaceutical compounds removal [27]. A gram-negative bacteria Acinetobacter calcoaceticus produced monodispersed cuboidal nanoparticles [28]. Another bacteria Shewanella algae stored the nanoparticles in periplasmic space [29]. Filamentous cyanobacteria such as Calothrix and Plectonema boryamum were known to produce nanoparticles with the size 3.2 and 5, respectively [29,30]. In a very recent report, biogenic copper nanoparticles were synthesized from Escherichia sp. for textile effluent treatment and azo dye degradation [31].
In a recent study, copper nanoparticles synthesized from Escherichia sp. were used for the treatment of textile effluents and azo dye photocatalysis [31].

Nanoparticles Biosynthesis in Fungi
Nanoparticle biosynthesis from fungi is another simple method which has been recently used. Fungi have enormous potential for nanoparticle production as compared to the bacteria. They have higher productivity and higher metal ions tolerance [13]. The biomass treatments and downstream processing are relatively straightforward in fungi when compared to bacteria. Also, they have metal ions bioaccumulation capacity, which results in cost-effective and efficient nanoparticle production. In this context, fungi Cladosporium oxysporum was used for the production of gold nanoparti-cles. The study investigated the effect of salt concentration, pH, and reaction time on yield and particle size [32]. The biosynthesized gold nanoparticles exhibited efficient catalytic activity in textile dye degradation. In another study, the effect of incubation time and reaction temperature were observed for the production of extracellular gold nanoparticles in the culture filtrate of Trichoderma viride and Hypocrea lixii [33]. White rot fungus, Schizophyllum radiatum isolated from the forest, had the potential to produce silver nanoparticles with the size range 10-40 nm [34]. The produced silver nanoparticles exhibited significant antibacterial activity against a variety of bacterial strains. Fungi Neurospora crassa produced nanoparticles in various shapes such as round, crystalline and quasi-spherical [35]. Another fungus Fusarium oxysporum produced variously shaped nanoparticles in size range of 5-140 nm [36,37].

Nanoparticle Biosynthesis in Algae
Algae are considered as promising candidate research due to their different properties. One of these properties is their ability to accumulate heavy metal ions [56,57]. After accumulation, metal ions can be transformed into useful materials [58]. Various properties mark the algae as nanofactories. These properties include lower doubling time, simple cell disruptions, quick harvesting, easy scalability at a high scale and low-cost production [59][60][61][62][63]. Various micro, as well as macroalgae, has been known to produce nanoparticles [64][65][66].
In microalgae, various strains have been used for the production of nanoparticles. Chlorella vulgaris produced up to 2 µm and 60 nm gold nanoparticles [67]. In this strain, a protein of 28kDa was identified, which was responsible for the shape direction of the gold nanoparticles. Klebsormidium Flaccidum produced gold nanoparticles in the range of 10-20 nm with the Sol-gel methods [68]. The study evidenced the gold reduction in the thylakoids. Tetraselmis suecica produced spherical gold nanoparticles in the range 51-120 nm [69]. Chlorella pyrenoidusa produced spherical and icosahedral gold nanoparticles in the range of 25-30 nm using the NADH-dependent enzyme [70]. Tetraselmis kochinensis produced triangular and spherical gold nanoparticles in the range of 5-35 nm using reducing enzymes in the cytoplasmic membrane and cell wall [71].
Algae can produce secondary metabolites and proteins, which can be used to produce potential metal nanoparticles [72][73][74][75]. The preparation of algae mediated nanoparticles synthesis involves algal extract preparation. The extract is then mixed with the metal precursor solution [76]. This procedure can be used to form stable nanoparticles in terms of different shapes and sizes [58,76,77]. Algae mediated nanoparticle synthesis can be intracellular [78] and extracellular [58,79]. Spirulina subsalsa and Lyngbya majuscula were used for the production of gold nanoparticles [80]. Spherical nanoparticles were produced from the alga Prasiola crispa [81].
Discussed here are a few very recent studies where algae were used to synthesize nanoparticles. In a study, silver nanoparticle was synthesized using algae. The study investigated the effect of silver nanoparticle formation on the life cycle of algae [82]. Silver nanoparticles were synthesized using green algae Botryococcus braunii, red algae Portieria hornemannii and Gelidium corneum [83][84][85]. A comparison was made for the two red algae for silver nanoparticles biosynthesis [86]. A macro algae polysaccharide was used for the silver nanoparticle's green synthesis [87]. An algae obtained from the Mediterranean sea was used for the biogenic synthesis of iron oxide nanoparticles [88]. A brown algae Cystoseira baccata was used for the gold nanoparticle's green synthesis [72].

MECHANISMS FOR BIOGENIC NANOPARTI-CLES PRODUCTION
A variety of factors are responsible for the reduction of metals ions to biosynthesize the nanoparticles in the microorganisms. One of them is the presence of an organic functional group on the cell wall, which is responsible for the induction of biomineralization of the metal ions. Another critical factor is the environmental conditions, which include pH, temperature, medium composition, and metal salt concentration [89]. These environmental conditions are known to affect nanoparticle morphology, size, and composition [90]. The efficiency of nanoparticle biosynthesis can be enhanced by the optimization of the factors. In such an effort, growth kinetics was optimized for the biosynthesis of silver nanoparticles using Morganella psychrotolerans to observe the effects on the nanoparticle's morphology [91]. In the study, the temperature played an important role where spherical nanoparticles were produced at 20°C, mixed spherical, hexagonal and triangular nanoplates at 25°C, mixed nanoplates and spherical at 15°C and significant enhancement of nanoplates number at 4°C. The biosynthesis of silver nanoparticles produced by Arthrobacter sp. demonstrated that factors such as pH, temperature, and metal ions concentrations could modulate nanoparticle synthesis [92]. The study demonstrated that a reduction in the silver nitrate concentration caused the synthesis of face-centered cubic nanoparticles, and increased silver nitrate concentration resulted in the aggregation of silver nanoparticles. The study concluded that metal ion concentration and medium pH could have a direct influence on nanoparticle synthesis.
In a study conducted for the biosynthesis of gold and silver nanoparticles demonstrated that low molecular weight secretory proteins which were present in the supernatant, which was responsible for nanoparticles synthesis [93]. Several studies reported the nanoparticle biosynthesis via bacterial extracellular polymeric substances (EPS) [94][95][96]. These substances act as effective capping and bio-reductant agents. In a similar study, EPS secreted from a marine isolate was used to produce silver nanoparticles [94].
A bacterium Bacillus cereus was isolated from a site contaminated with heavy metals. The bacterium synthesized extracellular silver nanoparticles with surface plasmon resonance properties [99]. These properties could be used in various applications. In an experiment, radiation-resistant strain Deinococcus radiodurans biosynthesized extracellular silver nanoparticles by silver chloride solution reduction [103]. The biosynthesized silver nanoparticles demonstrated antibacterial, anti-biofilm and anti-cancerous activities. In a study, palladium and platinum nanoparticles were synthesized, which demonstrated methyl orange dye decomposition [106]. In a different study, Ochrobactrum sp. produced tellurium nanoparticles, which demonstrated the potential of this bacterium for the conversion of tellurite oxyanions to the useful nanopar-ticles [107]. The toxic gold ions were converted to the gold nanoparticles by Bacillus subtilis [108]. The produced gold nanoparticles were used as a biocatalyst for methylene blue degradation and can be used for degradation of other dyes toxic to the environment. Silver nanoparticles produced by Bacillus brevis demonstrated antibacterial activities against drug-resistant strains of Staphylococcus Aureus and Salmonella typhi [100].

Mechanisms of Nanoparticles Biosynthesis in Yeast Cells
Yeast cells adapt themselves in the metal toxicity conditions by detoxification mechanisms such as chelation, precipitation and intracellular sequestration. In a study, Yarrowia lipolytica yeast cells were used for the biosynthesis of silver nanoparticles. The study demonstrated that brown pigment in the yeast cells was responsible for the biomineralization of metal ions and pigment derived nanoparticles exhibited antibiofilm activity against pathogen Salmonella paratyphi [40]. Extracellular eco-friendly silver nanoparticles were synthesized by Candida utilis NCIM 3469 [41]. The produced nanoparticles were circular, with a size range of 20-80 nm and had antibacterial activity against pathogenic strains such as Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa. A genetically modified Pichia pastoris was used for silver nanoparticles biosynthesis [45]. The yeast overexpressed cytochrome b5 reductase enzyme, which was responsible for metal ions reduction to the nanoparticles. A yeast strain, Candida Lusitania isolated from termite gut, produced silver nanoparticles in the size range 2-10 nm [46]. The biosynthesized nanoparticles exhibited antiproliferative activity to Klebsiella pneumoniae and S. aureus. In another study, Saccharomyces cerevisiae was used for the biosynthesis of palladium nanoparticles, which exhibited photocatalytic textile azo dye degradation [54].

Mechanisms for Intracellular Nanoparticle Biosynthesis
In the mechanism of intracellular biomineralization of silver ions, it was considered that the enzymes present on the cell wall of the bacteria were responsible for the production of silver nuclei. In a study, silver nanoparticles were produced by the reduction of silver ions by Streptomyces sp. LK-3 [112]. The study reported that the extracellular production of stable silver nanoparticles was conducted by NADHreductase by an electron transfer reaction. The produced nanoparticles exhibited strong antiparasitic and acaricidal activities. In a recent study, Streptacidiphilus durhamensis was used for the synthesis of silver nanoparticles [113]. The produced nanoparticles demonstrated antimicrobial activity against Proteus mirabilis, Staphylococcus aureus and Pseudomonas aeruginosa. In general, the nanoparticles synthesized from the microorganisms have higher antimicrobial activities due to the stabilization and capping of the nanoparticles as compared to the traditional nanoparticles synthesized.

Mechanisms for Extracellular Nanoparticle Biosynthesis
Most of the studies reported have demonstrated the extracellular secretion of the nanomaterials. The advantages of extracellular nanoparticle synthesis are that it is devoid of impurities such as intracellular proteins. Moreover, treatment of detergent is not involved, and ultrasound is not required. Understanding the mechanism of nanoparticle biosynthesis in fungi is indispensable for various applications. In this context, the effect of temperature, pH and isolate selection on nanoparticle morphology was observed in the nanoparticle production using Fusarium oxysporum [119]. Aluminum oxide nanoparticles were synthesized using Colletotrichum sp., and the synthesized nanoparticles were functionalized by essential oils [120]. The study results demonstrated that functionalized oil could be used as an antimicrobial agent for food-borne pathogens. Recently, antimicrobial and anticancer nanoparticles have been synthesized using edible mushroom Pleurotus ostreatus and two fungi Scopulariopsis brumptii and Penicillium citreonigrum [121].

BIOGENIC NANOPARTICLES CYTOTOXICITY
There are various routes through which organisms come in contact with the nanoparticles. Humans can be exposed to nanoparticles through inhalation, skin contact, and gastrointestinal absorption [122]. After entering the human body, nanoparticles can be engulfed by the macrophages, which may lead to the development of inflammation [123,124]. On ingestion, it reaches the liver and accumulates there [125]. Nanoparticle accumulation results in the glutathione level alteration increased reactive oxygen species (ROS) and mitochondrial potential reduction [126]. Nanoparticles are also known to induce oxidative stress in the respiratory system [127].
Studies have reported that silver nanoparticles can disturb the plasma membrane integrity, which leads to leakage [128]. The normal function of the mitochondrial membrane can be impaired with the nanoparticle interaction [129]. In a study, nanoparticles caused the mitochondrial membrane lipids peroxidation after accumulation around the mitochondria [130]. Reactive Oxygen Production mediated apoptosis was observed in the cells exposed to metal nanoparticles [131]. DNA damage is caused after nanoparticles enter the nucleus [132]. Nanoparticles exposure can disturb regular cell metabolism leading to apoptosis and cell death [133]. Apoptosis mediated cell death is governed by combined molecular mechanisms, such as DNA fragmentation and caspase pathway [134].
All the toxicity, as mentioned above, were reported for artificially synthesized nanoparticles. Biosynthesized nanoparticles also cause some toxicity. In a bacteria mediated nanoparticle synthesis, the silver nanoparticles completely inhibited the growth of brine shrimp at up to 100 µg/mL [135]. In plants, iron nanoparticles caused the alteration in the root cell walls structure in Arabidopis thaliana [136]. The structural changes were mediated by the loosening of the cells. The toxicity of the nanoparticles depends on the shape, size, and structure of the nanoparticle [137]. The dose of the nanoparticles also plays an important factor in the toxicity. Higher dose results in more toxicity as compared to the lower doses [138]. The dispersion of nanoparticles is another important factor affecting toxicity. The well-dispersed nanoparticles cause more toxicity as compared to the agglomerated nanoparticles [139]. Toxicity evaluation of the biogenic silver nanoparticles produced from Althaea officinalis was performed. The study evaluated the toxicity imposed by the two types of silver nanoparticles prepared from the infusion of roots and extracts [140]. The study results demonstrated that the nanoparticles prepared from the extract were more toxic as compared to the nanoparticles prepared from the root infusion.
A recent study evaluated the toxicity imposed by the biogenic synthesized nanoparticles in the zebrafish and the human endothelial cells [141]. The results from the study demonstrated reactive oxygen species formation in human endothelial cells, and the induction of apoptosis was also observed. Induction of the apoptosis was supported by the increased expression of the apoptotic biomarkers. The in vivo trials on the zebrafish resulted in the imbalanced heart rate, mortality, and cell death in the embryo. The study also confirmed the morphological changes in the tail and yolk sac of the zebrafish. The biogenic functionalized nanoparticles cause less toxicity as compared to the chemically functionalized nanoparticles. Iron oxide nanoparticle's neurotoxicity was observed on retention in mitochondria or lysosomes [142]. In the study, three types of iron oxide nanoparticles were synthesized, and cytotoxicity in Parkinson`s disease cellular model was analyzed. The study confirmed significant toxicity on SH-SY5Y cells imposed by lysosometargeted nanoparticles. Inhibition of AMPK caused the increased neurotoxicity in lysosome as well as mitochondriatargeted nanoparticles . The neurotoxicity also caused the mitochondrial membrane potential alteration.

CONCLUSION
Biogenic nanoparticles are a new concept that came into existence after the immense harmful effects of artificially synthesized nanoparticles were observed. Biogenic nanoparticles can be produced from microbes, plants, and animals. It was observed that microbes could be considered as a useful and efficient system for the biosynthesis of nanoparticles. These microbes, especially bacteria, fungi, yeast, and algae, have the potential to produce different types of nanoparti-cles. They have been reported to produce spherical, hexagonal, cuboidal, and nanoplates. Also, the potential to produce nanoparticles utilizing various metal ions such as gold, silver, platinum, palladium, etc. When compared with the artificially designed nanoparticles, the biosynthesized nanoparticles exhibited less toxicity. The studies have reported the various mechanisms involved in the reduction of metal ions to produce nanoparticles by the microbes. The biogenic nanoparticles are produced both intracellularly as well as extracellularly. Although the biogenic nanoparticles are considered less toxic, a few recent studies have demonstrated their toxic effects on the organisms and organ systems.

CONSENT FOR PUBLICATION
Not applicable.