Bioleaching of Phosphate Minerals Using Aspergillus Niger: Recovery of Copper and Rare Earth Elements

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Article Bioleaching of Phosphate Minerals Using Aspergillus niger: Recovery of Copper and Rare Earth Elements

Laura Castro *, Maria Luisa Blázquez, Felisa González and Jesús Angel Muñoz

Department of Chemical and Materials Engineering, Complutense University of Madrid, 28040 Madrid, Spain; [email protected] (M.L.B.); [email protected] (F.G.); [email protected] (J.A.M.) * Correspondence: [email protected]; Tel.: +34-91-394-4354

Received: 11 June 2020; Accepted: 15 July 2020; Published: 20 July 2020

Abstract: Rare earth elements (REE) are essential in high-technology and environmental applications, where their importance and demand have grown enormously over the past decades. Many lanthanide and actinide minerals in nature are phosphates. Minerals like monazite occur in small concentrations in common rocks that resist weathering. Turquoise is a hydrous phosphate of copper and aluminum scarcely studied as copper ore. Phosphate-solubilizing microorganisms are able to transform insoluble phosphate into a more soluble form which directly and/or indirectly contributes to their metabolism. In this study, bioleaching of heavy metals from phosphate minerals by using the fungus Aspergillus niger was investigated. Bioleaching experiments were examined in batch cultures with diﬀerent mineral phosphates: aluminum phosphate (commercial), turquoise, and monazite (natural minerals). The experiments were performed at 1% pulp density and the phosphorous leaching yield was aluminum phosphate > turquoise > monazite. Bioleaching experiments with turquoise showed that A. niger was able to reach 8.81 mg/l of copper in the aqueous phase. Furthermore, the fungus dissolved the aluminum cerium phosphate hydroxide in monazite, reaching up to 1.37 mg/L of REE when the fungus was grown with the mineral as the sole phosphorous source. Furthermore, A. niger is involved in the formation of secondary minerals, such as copper and REE oxalates.

Keywords: Aspergillus niger; mineral phosphate; bioleaching; rare earth elements

1. Introduction Rare Earth Elements (REE) attract enormous interest because of their importance to current and high technological manufacturing industries and the expectation of an increase in requirements throughout the world on a monopolistic market of REE due to geopolitical controls [1]. The distribution of rare earths by their end use is estimated as follows: catalysts, 60%; ceramics and glass, 15%; metallurgical applications and alloys, 10%; polishing, 10%; others, 5% [2]. The production of rare earths increased with renewed mining activities in the United States of America as well as new and or increased production in Australia, Burma (Myanmar), and Burundi. Mine production in China increased 14% in 2018 compared with the quota in 2017. In 2018, the USA Department of the Interior and other executive branch agencies published a list of 35 critical minerals, including rare earths [3]. Likewise, the European Commission carries out a critical assessment on non-energy and non-agricultural raw materials that, in 2017, included heavy rare earth elements, light rare earth elements, and platinum group metals [4]. Despite their name, these elements are not rare and occur abundantly in the Earth’s crust, however, not at high enough concentrations to be considered economically recoverable. The elements vary in abundance from cerium, the 25th most abundant element of common elements at 60 parts per million,

Metals 2020, 10, 978; doi:10.3390/met10070978 www.mdpi.com/journal/metals Metals 2020, 10, 978 2 of 13 to thulium and lutetium, the least abundant rare earth elements in the Earth at about 0.5 ppm [5]. The three main REE ores that are currently mined for production are bastnaesite (REE-FCO3), monazite (light REE-PO4), and xenotime (heavy REE-PO4), together representing approximately 95% of known REE minerals. Conventional methods of REE extraction from ores involve the use of high temperatures and harsh chemicals (either concentrated sulfuric acid or concentrated NaOH), resulting in high energy usage and the production of toxic waste streams [6]. Biohydrometallurgical methods are generally considered as an eco-friendly alternative with low cost and low energy requirements. Bioleaching allows the recovery of some valuable heavy metals and reduces the toxicity of the waste materials using microorganisms [7,8]. Recently, some research work has been developed, especially with monazite, using microorganisms able to dissolve phosphorus from inorganic rocks, named as phosphate-solubilizing microorganisms (PSMs). PSMs have been previously used in agriculture to enhance crop production, but there are few studies related to the recovery of valuable elements from phosphate minerals. Several microorganisms, such as the bacteria Enterobacter aerogenes, Pantoea agglomerans, and Pseudomonas putida, have demonstrated to grow in the presence of natural rare earth phosphate minerals, releasing phosphorous, iron, thorium, and REE. Numerous organic acids were produced by the microorganisms, but microbial processes were also playing a role in solubilization of the monazite ore [9]. Azospirillum brasilense, Azospirillum lipoferum, Pseudomonas rhizosphaerae, and Mesorhizobium cicero, but particularly Acetobacter aceti, were able to solubilize cerium and lanthanum from monazite, although the efficiency of the process was low [10]. Fungal strains able to solubilize phosphate minerals have been also used to leach monazite releasing rare earth elements to the aqueous phase, such as Aspergillus niger ATCC 1015, Aspergillus terreus strain ML3-1, and a Paecilomyces spp. strain WE3-F [11]. The fungus Penicillum sp. solubilized a total 1 concentration of 12.32 mg L− rare earth elements after 8 days. Although monazite also contains radioactive thorium, bioleaching by these fungi preferentially solubilized rare earth elements over thorium that remained in the solid residual [9]. The most important mechanism of metal leaching by heterotrophic fungus is the production of organic acids, however, phosphatase activity and the microbial attachment could be involved in the dissolution of monazite [12]. Aspergillus species have shown potential for rare earth metals bioleaching of various waste materials, such as fly ash [13], spent catalysts [14], and electrical waste [15]. Some of these processes reached leaching recoveries similar to the yields offered by chemical leaching. Copper has been extensively used in industry due to its excellent ductility and electric and thermal conductivity. Copper bioleaching has commercial application in the metal extraction from low grade and secondary mineral resources, mainly sulfides [16,17]. Nevertheless, copper phosphate bioleaching has been scarcely investigated [18]. Several hundred phosphate minerals are known and the most important mineral is apatite. Apatites and poorly crystalline or amorphous phosphorite sediments are mined and treated with acids to obtain fertilizers. Nevertheless, phosphates have been scarcely mined to extract valuable metals. Many of the f-block elements are extracted from phosphate minerals, notably monazite [19]. Turquoise mineral was considered in this study as a source of copper. Some copper companies do not consider turquoise valuable enough for sale; instead, the mineral is crushed and processed for its metal content. The main objective of the present work is to study the solubilization of phosphorous and the recovery of valuable elements from mineral phosphates (aluminum phosphate, turquoise, and monazite) using the fungus A. niger. The release of copper from turquoise and rare earth elements (Ce, La, and Nd) from monazite was monitored. In addition, the pH and the redox potential (EhAg/AgCl) were followed during the process, evidencing that the mineral dissolution is caused by the fungal growth and the production of organic acids that acidified the medium and complexed the metals. The samples were examined by X-ray diffraction (XRD) and Scanning Electron Microscopy (SEM), showing that A. niger metabolites are involved in the formation of secondary minerals, such as copper and REE-oxalates. Metals 2020, 10, 978 3 of 13

2. Materials and Methods

2.1. Minerals Diﬀerent phosphate minerals, both commercial and natural, were used in the bioleaching tests (Table1).

Table 1. Mineral phosphates tested in the bioleaching experiments using A. niger.

Mineral Formula Type Origin

Berlinite AlPO4 Commercial Fluka Turquoise CuAl (PO ) (OH) 5H O Natural Mirandilla, Badajoz (Spain) 6 4 4 3· 2 Monazite (Ce, La, Nd, Th)PO4 Natural Seis Lagos (Brazil)

The elemental composition of the natural phosphate minerals was determined by X-ray ﬂuorescence chemical analysis. The composition of the turquoise was (wt. %): Si, 22.4; Al, 12.9; P, 7.3; Cu, 3.5; Fe, 2.0; O, 48.9. The main elements in the monazite were (wt. %): Al, 16.7; Si, 16.5; Fe, 4.95; S, 4.59; P, 1.42; Ce, 1.23; Nd, 0.69; La, 0.62; Th, 0.54; O, 46.8.

2.2. Leaching Fungal Strain and Bioleaching Experiments The fungus Aspergillus niger CECT2807 was provided by the Spanish Type Culture Collection. The bioleaching experiments were carried out in 250 mL Erlenmeyer flasks in triplicate. Each flask containing 1% mineral (w/v) was autoclaved for 30 min at 121 ◦C. No mineralogical changes were observed in X-ray diffractograms before and after autoclaving. A volume of 100 mL of sterile potato dextrose broth (potato starch (from infusion) 4 g/L, dextrose 20 g/L, Difco, Carlsbad, CA, USA) at initial pH 5.3 was added to each flask. Flasks were inoculated with the fungus A. niger, and then, cultures were incubated aerobically on an orbital shaker (New Brunswick Scientific Innova 44R) at 150 rpm and 30 ◦C. Non-inoculated flasks were used as controls. An amount of 5 mL was periodically withdrawn during the experiments for further analysis (phosphorous and metal concentrations, pH, and EAg/AgCl. In addition, A. niger was grown on non-sterile monazite and sterile potato dextrose broth to observe the influence of indigenous biota on the bioleaching process. The fungus was also grown with monazite as the sole phosphate source on modified minimal medium containing (g/L): NaNO3, 6.0; KC1, 0.52; MgSO 7H O, 0.52; glucose, 10. All the reagents were provided by Fisher Chemicals 4· 2 (percent purity 99%). ≥ 2.3. Analytical Methods

pH and redox potential (EhAg/AgCl) of the samples were measured using a pHmeter Crison Basic 20 (sensitivity: 98%) (Hach Lange., Barcelona, Spain). The concentration of phosphorous was determined using an ICP-OES equipment (Perkin Elmer Optima 2100 DV, Perkin Elmer Inc., Waltham, MA, USA). In addition, the concentration of copper during turquoise experiments and cerium, lanthanum, and neodymium concentrations in monazite experiments were measured by ICP-OES. Each measurement was performed in triplicate with RSD <3%. The morphology of fungi and the different phosphate minerals was observed using Scanning Electron Microscopy (SEM, JEOL JSM-6330 F, JEOL, Boerne, TX, USA). Samples with fungus were prepared by filtering through 0.2 µm pore-size filters and successively dehydrated with acetone/water mixtures of 30%, 50%, and 70% acetone, respectively, and stored overnight at 4 ◦C in 90% acetone for cell dehydration. After critical-point drying and coating with gold, samples were observed under the microscope FE-SEM at 15–20 kV. Mineral residues in culture flasks were characterized using powder X-ray diffraction (XRD) on a Philips X’pert-MPD equipment (Malvern Panalytical, Eindhoven, The Netherlands) with a Cu anode operating at a wavelength of 1.5406 Å as the radiance source. Samples were placed on off-axis quartz Metals 2020, 10, 978 4 of 13 plates (18 mm diameter 0.5 mm DP cavity). The scanning range was from 10 to 90 2θ, with an × ◦ ◦ angular interval of 0.05◦ and 4 s counting time. The crystalline phases were identified using standard cards from the International Centre for Diffraction Data (ICDD, Newtown Square, Pennsylvania) PowderMetals 2020 Di, 10ffraction, x FOR PEER File REVIEW database. 4 of 13

3.3. ResultsResults

3.1.3.1. PhosphatePhosphate MineralMineral SolubilizationSolubilization PhosphorousPhosphorous is is an essentialan essential macronutrient macronutrient for all livingfor all organisms. living organisms. This element This has element a fundamental has a rolefundamental in biochemical role reactionsin biochemical involving reactions genetic materialinvolving (DNA, genetic RNA) material and energy (DNA, transfer RNA) (ATP), and energy and in thetransfer formation (ATP), of celland membranesin the formation (phospholipids) of cell me thatmbranes provide (phospholipids) structural support that forprovide organisms structural [20]. Consequently,support for organisms phosphorous [20]. isConsequently, a limiting nutrient phosphorous for microorganisms is a limiting in nutrient many environments for microorganisms due to the in deﬁcitmany ofenvironments organophosphates due to or the the lowdeficit solubility of organo of inorganicphosphates mineral or the phosphates low solubility (usually of presentinorganic in soilsmineral and phosphates rocks). Microorganisms (usually present have toin producesoils and enzymes rocks). Microorganisms named phosphatases have or to generate produce metabolic enzymes productsnamed phosphatases such as organic or generate acids to dissolvemetabolic the products insoluble such phosphorous as organic immobilizedacids to dissolve in the the crystalline insoluble structurephosphorous of some immobilized minerals [in21 ].the crystalline structure of some minerals [21]. CalciumCalcium phosphates phosphates can can be be solubilized solubilized by by a wide a wide range range of microorganisms.of microorganisms. Nevertheless, Nevertheless, there there are otherare other valuable valuable phosphate phosphate minerals minerals which arewhich more are refractory. more refractory. The fungus TheAspergillus fungus Aspergillus niger CECT2807 niger wasCECT2807 grown inwas the grown presence in the of aluminumpresence of phosphate, aluminum turquoise, phosphate, and turquoise, monazite and and monazite phosphorous and concentrationphosphorous wasconcentration measured periodically.was measured The periodical initial concentrationly. The initial of phosphorousconcentration in of the phosphorous medium was in 27.63the medium mg/L. Aluminum was 27.63 mg/L. phosphate Aluminum was abiotically phosphate dissolved, was abiotically but A.niger dissolved,enhanced but A. the niger phosphorous enhanced solubilizationthe phosphorous rate. Whensolubilization the fungus rate. strain When was grownthe fungus with turquoise,strain was phosphorous grown with concentration turquoise, decreasedphosphorous to 5.10 concentration mg/L after 7decreased days, and then,to 5.10 the mg concentration/L after 7 increased,days, and reaching then, the 47.97 concentration mg/L after 14increased, days. A. reaching niger also 47.97 grew mg/L in presence after 14 of days. monazite, A. niger as also shown grew in in Figure presence1. After of monazite, 7 days, phosphorous as shown in concentrationFigure 1. After decreased 7 days, ph uposphorous to 0.70 mg concentration/L, and then, decreased increased up to 8.810.70 mgmg/L,/L. and then, increased up to 8.81 mg/L.

160

140

120 Control AlPO4 100 -1 A. niger AlPO4 80 Control turquoise

[P]/mg·l 60 A. niger turquoise 40 Control monazite 20 A. niger monazite 0 051015 Time / days

FigureFigure 1.1. EvolutionEvolution ofof phosphorousphosphorous concentrationconcentration fromfrom didifferentﬀerent mineralmineral phosphatesphosphates (aluminum(aluminum phosphate,phosphate, turquoise turquoise and and monazite) monazite) at at 1 1 g g/L/L in in abiotic abiotic and and biotic biotic ( A.(A. niger niger)) tests. tests.

A.A. nigernigerwas was ableable toto transformtransform remarkablyremarkably twotwo veryvery insolubleinsoluble minerals,minerals, turquoiseturquoise andand monazite,monazite, withwith an an acidiﬁcation acidification of of the the medium medium (Figure (Figure2a). 2a). The Th protone proton substitution substitution reaction reaction is responsible is responsible for the for mineralthe mineral phosphate phosphate solubilization solubilization through through the biological the biological production production of organic of acids,organic which acids, could which be representedcould be represented by the following by the fo generalllowing Equation general [Equation22]: [22]:

[Mn+][PO43−] + [HA] = [H+] [PO43−] + [Mn+] [A−] (1)

where [HA] is the corresponding organic acid and Mn+ is a metallic cation. The stoichiometry is not included in this equation because it depends on the mineral phosphate and the type of organic acid generated by the microorganism. The efficiency of solubilization depends on the mineral phosphate solubility, the amount of protons generated by the organic acid, and its pKa. After 7 days, the pH values in the inoculated flasks with AlPO4 and turquoise experienced a sharp increase associated with the release of phosphorous in the aqueous phase due to proton

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n+ 3 + 3 n+ [M ][PO4 −] + [HA] = [H ] [PO4 −] + [M ] [A−] (1) where [HA] is the corresponding organic acid and Mn+ is a metallic cation. The stoichiometry is not included in this equation because it depends on the mineral phosphate and the type of organic acid generated by the microorganism. The efficiency of solubilization depends on the mineral phosphate solubility, the amount of protons generated by the organic acid, and its pKa. After 7 days, the pH values in the inoculated flasks with AlPO4 and turquoise experienced aMetals sharp 2020 increase, 10, x FOR associated PEER REVIEW with the release of phosphorous in the aqueous phase due to proton5 of 13 consumption. In addition, turquoise is a hydrated basic phosphate that raises the OH- concentration duringconsumption. its solubilization. In addition, turquoise is a hydrated basic phosphate that raises the OH- concentration duringThe its redox solubilization. potential was also monitored during the bioleaching processes (Figure2b). The increase in theThe redox redox potential potential observed was also at the monitored initial stage, during in parallel the bioleaching to the decrease processes in the (Figure pH value 2b). in The the flasks,increase was in attributedthe redox potential to the solubilization observed at of the mineral initial phosphatesstage, in parallel by the to fungus the decrease via direct in the oxidation pH value of glucosein the flasks, to gluconic was attributed acid and other to the organic solubilization acids. of mineral phosphates by the fungus via direct oxidation of glucose to gluconic acid and other organic acids.

9 350

8 300 7 250 6 200

5 mV / pH 4 150 Ag/AgCl 3 E 100 2 1 50 0 0 0 5 10 15 0 5 10 15 Time / days Time / days a b pH control AlPO4 pH A. niger AlPO4 E control AlPO4 E A. niger AlPO4 pH control turquoise pH A. niger turquoise E control turquoise E A. niger turquoise pH control monazite pH A. niger monazite E control monazite E A. niger monazite

Figure 2. Evolution of (a) pH and (b) EAg/AgCl in abiotic and biotic (A. niger) tests for different mineral Figure 2. Evolution of (a) pH and (b)E in abiotic and biotic (A. niger) tests for diﬀerent mineral phosphates (aluminum phosphate, turquoise,Ag/AgCl and monazite) at 1 g/L. phosphates (aluminum phosphate, turquoise, and monazite) at 1 g/L. Microbial bioleaching of insoluble metal phosphate minerals is caused by protonation of the Microbial bioleaching of insoluble metal phosphate minerals is caused by protonation of the anion anion of the metal compound. In fungi, two mechanisms seem to be the most relevant for of the metal compound. In fungi, two mechanisms seem to be the most relevant for solubilization solubilization of phosphate minerals: (1) the H+-translocating ATPase of the plasma membrane that of phosphate minerals: (1) the H+-translocating ATPase of the plasma membrane that is a source is a source of protons, and (2) the production of organic acids, which can be effective complexing of protons, and (2) the production of organic acids, which can be eﬀective complexing agents for agents for metal cations [21]. In addition, the excretion of metabolites, siderophores, and CO2 from metal cations [21]. In addition, the excretion of metabolites, siderophores, and CO2 from microbial microbial respiration also contributes to the dissolution process [23]. The importance of all these respiration also contributes to the dissolution process [23]. The importance of all these processes varies processes varies depending on the microorganism and the growth conditions. Previous research has depending on the microorganism and the growth conditions. Previous research has evidenced that evidenced that A. niger produces citric acid when it is grown with metal phosphates [24]. Organic A. niger produces citric acid when it is grown with metal phosphates [24]. Organic acids in the soil 3− acids in the soil promote the chelation of cations and favor anions release,3 such as PO4 , leading to promote the chelation of cations and favor anions release, such as PO4 −, leading to the solubilization the solubilization of phosphate compounds. of phosphate compounds.

3.2. Turquoise BioleachingBioleaching usingusing AspergillusAspergillus nigerniger There areare nono previousprevious studiesstudies onon turquoiseturquoise bioleachingbioleaching toto recoverrecover copper.copper. The evolutionevolution of copper concentration in the aqueous phase during thethe dissolution of turquoise by fungal action is shown inin Figure Figure3. Copper3. Copper concentration concentration in the in aqueous the aqueous phase wasphase 8.81 was mg 8.81/L after mg/L 14 days.after However,14 days. theHowever, maximum the concentrationmaximum concentration was 13.02 mg was/L, 13.02 reached mg after/L, reached 9 days. after At that 9 days. moment, At that the pHmoment, increased the andpH increased the redox and potential the redox decreased potential as a decreased consequence as a of consequence copper precipitation of copper (Figure precipitation2). (Figure 2).

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-1 10

8 Control

[Cu]/ mg·L 6 A. niger 4

0 051015 Time /days

Figure 3.3. EvolutionEvolution of of copper copper concentration concentration from from turquoise turquoise (1 g(1/L) g/L) in abiotic in abiotic and bioticand biotic (A. niger (A.) niger tests.) tests. Organic acids, citric, oxalic and gluconic acid, are generated by the A. niger, whether a mineral phosphateOrganic is presentacids, citric, or not oxalic [25]. and The gluconic solubilization acid, are of insoluble generated metal by the phosphates A. niger, whether release freea mineral metal cationsphosphate and /isor present soluble or metal not complexes,[25]. The solubilization increasing their of insoluble availability. metal Attending phosphates to their release high free solubility metal andcations the bindingand/or constants,soluble metal gluconate complexes, ([CuL+ ],increasi 7.1 10ng7; [CuLtheir ],availability. 2.5 1014) and Attending citrate ([CuL] to their-, 1 high1018; × 2 × × [CuHL],solubility 2 and10 22the; [CuH bindingL]+, constants, 2 1028) would gluconate be involved ([CuL+ in], 7.1 the high× 107 copper; [CuL2 concentration], 2.5 × 1014) and in solution. citrate × 2 × ([CuL]The-, 1decrease × 1018; [CuHL], in copper 2 × concentration1022; [CuH2L]+, could 2 × 10 be28) causedwould bybe involved the precipitation in the high of organiccopper metabolitesconcentration [26 in] solution. or by mineralogical changes (dissolution and subsequent reprecipitation) in the mineralThe substrate decrease during in copper biological concentration solubilization could [27]. be caused by the precipitation of organic metabolitesRaw turquoise [26] or andby mineralogical the mineral residues changes obtained (dissolution after and fungal subsequent growth were reprecipitation) analyzed by in XRD the diffractionmineral substrate to compare during mineral biological phases solubilization (Figure4). The [27]. turquoise diffractogram confirmed the composition obtainedRaw byturquoise X-ray fluorescence and the mineral and its residues high crystallinity. obtained after In addition, fungal growth these XRD were patterns analyzed evidenced by XRD significantdiffraction changesto compare in mineralogy mineral phases in the inoculated(Figure 4). samples. The turquoise Turquoise diffractogram (CuAl (PO )confirmed(OH) 5H theO) 6 4 4 8· 2 wascomposition completely obtained transformed by X-ray by A. fluorescence niger CECT2807 and 14 its days high after crystallinity. inoculation. In The addition, diffractogram these ofXRD the mineralpatterns residue evidenced obtained significant showed thechanges presence in ofmine copperralogy phosphate in the andinoculated copper oxalate, samples. evidencing Turquoise the copper(CuAl6(PO reprecipitation4)4(OH)8·5H2 byO) thewas biological completely action. transformed by A. niger CECT2807 14 days after inoculation.The residues The fromdiffractogram the inoculated of the flasks mineral containing residue turquoise obtained were showed observed the by presence FE-SEM of (Figure copper5). Turquoisephosphate particles and copper were oxalate, transformed evidencing and very the densecopper mycelium reprecipitation was formed by the by biologicalA. niger, whereaction. copper phosphate and copper oxalate crystals were precipitated and adsorbed. Fungi play an important role in the formation of organic and inorganic secondary minerals through precipitation and deposition of crystalline compounds on and within cells walls, such as oxalates and phosphates. Previous studies evidenced that fungi can generate metal oxalates with different chemical elements and minerals [28]. The fungus Beauveria caledonica can solubilize minerals containing heavy metals such as cadmium, copper, lead or zinc and form oxalates in the local microenvironment and in association with the mycelium. This fungus accumulated high amounts of copper as copper oxalate in the presence of copper phosphate [29]. A. niger was able to solubilize metal compounds (ZnO, Zn3(PO4)2 and Co3(PO4)2) and precipitate metal oxalates [26]. The precipitation of metal oxalates may provide a detoxification mechanism, whereby fungi can survive at high concentrations of hazardous metals.

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Figure 4. X-ray diffraction (XRD) pattern of the turquoise (a) before and (b) after 14 days of bioleaching using A. niger (● potassium aluminum silicate hydroxide, ■ silicon oxide, ♦ copper aluminum phosphate hydroxide hydrate, copper phosphate, ▲ copper oxalate). Figure 4. X-ray diffraction (XRD) pattern of the turquoise (a) before and (b) after 14 days of Figure 4. X-ray diﬀraction (XRD) pattern of the turquoise (a) before and (b) after 14 days of bioleaching Figure 4. X-ray Thebioleachingdiffraction residues (XRD) using frompattern A. the ofniger the inoculated turquoise(● potassium (a) flasksbefore aluminum and containing (b) after silicate14 turquoisedays ofhydroxide, were ■observed silicon oxide, by FE-SEM ♦ copper (Figure bioleaching usingusing A. nigerA. niger(● potassium( potassium aluminum aluminum silicate hydroxide, silicate ■ hydroxide,silicon oxide, ♦ siliconcopper oxide, copper aluminum phosphate 5). Turquoise particles• were transformed and very dense mycelium▲ was formed by A. niger, where hydroxidealuminum hydrate,phosphate copperhydroxide phosphate, hydrate,▲ N copper copper oxalate). phosphate, copper oxalate). aluminumcopper phosphate phosphate hydroxide hydrate, and copper copper phosphate,oxalate crys coppertals oxalate). were precipitated and adsorbed. The residues fromThe the residues inoculated from flasks containingthe inoculated turquoise flasks were observed containing by FE-SEM turquoise (Figure were observed by FE-SEM (Figure 5). Turquoise 5).particles Turquoise were transformed particles and were very densetransformed mycelium wasand formed very bydense A. niger mycelium, where was formed by A. niger, where copper phosphate and copper oxalate crystals were precipitated and adsorbed. copper phosphate and copper oxalate crystals were precipitated and adsorbed.

Figure 5. Scanning Electron Microscopy (SEM) images of (a) A.niger grown with turquoise after 14 days; (b) detail ofFigure precipitates 5. Scanningon fungal biomass. Electron Microscopy (SEM) images of (a) A.niger grown with turquoise after 14 Figure 5. Scanning Electron Microscopy (SEM) images of (a) A.niger grown with turquoise after 14 days; Fungi play an days;important (b) detailrole in ofthe precipitates formation of onorganic fungal and biomass. inorganic secondary minerals (b) detail of precipitates on fungal biomass. through precipitation and deposition of crystalline compounds on and within cells walls, such as oxalates and phosphates.Fungi play an important role in the formation of organic and inorganic secondary minerals Previous3.3. studies MonaziteFigure evidenced 5. BioleachingScanning that fungi Electron can gene Microsrate metalcopy oxalates (SEM) with images different of (chemicala) A.niger grown with turquoise after 14 elements and throughmineralsdays; [28]. precipitation ( bThe) detail fungus of Beauveria precipitates and caledonicadeposition on canfungal solubilize of biomass.crystallin minerals e containingcompounds heavy on and within cells walls, such as Aspergillus niger CECT2807 was grown on monazite from the Morro dos Seis Lagos (Brazil). metals such asoxalates cadmium, and copper, phosphates. lead or zinc and form oxalates in the local microenvironment and The Morro dos Seis Lagos deposit is formed by siderite as the main mineral and is covered by a in association with PreviousFungithe mycelium. play studies anThis important fungus evidenced accumulated role that in high thefungi amounts formation can ofgene copper ofrate organicas coppermetal and oxalates inorganic with secondary different chemicalminerals oxalate in the presence of copper phosphate [29]. A. niger was able to solubilize metal compounds thickelements ferruginous and minerals lateritic [28]. crust. The fungus This site Beauveria hosts one caledonica of the largestcan solubilize Nb deposits minerals in the containing world. Itheavy also (ZnO, Zn3(POthrough4)2 and Co precipitation3(PO4)2) and precipitate and deposition metal oxalates of [26]. crystallin The precipitatione compounds of metal on and within cells walls, such as contains iron and manganese and important resources of REE [30]. The evolution of REE and thorium metalsoxalates such and as phosphates. cadmium, copper, lead or zinc and form oxalates in the local microenvironment and concentrations in the aqueous phase during the bioleaching experiments with A. niger are shown in in associationPrevious studieswith the evidenced mycelium. that This fungi fungus can accumulatedgenerate metal high oxalates amounts with of differentcopper as chemical copper Figure6. It is known that REE-phosphates have particularly low solubilities in water, in the order of oxalateelements in and the mineralspresence [28]. of copper The fungus phosphate Beauveria [29]. caledonica A. niger wascan solubilizeable to solubilize minerals metal containing compounds heavy 10 13 M (10 11 g/L) [31]. After three days of bioleaching, the REE concentration reach a maximum (ZnO,metals− Znsuch3(PO− as4 )cadmium,2 and Co 3copper,(PO4)2) andlead orprecipitate zinc and metalform oxalates oxalates in [26]. the localThe microenvironmentprecipitation of metal and (0.97 mg/L) corresponding to the production of higher concentrations of citric and gluconic acids by in association with the mycelium. This fungus accumulated high amounts of copper as copper oxalate in the presence of copper phosphate [29]. A. niger was able to solubilize metal compounds (ZnO, Zn3(PO4)2 and Co3(PO4)2) and precipitate metal oxalates [26]. The precipitation of metal

Metals 2020, 10, x FOR PEER REVIEW 8 of 13 oxalates may provide a detoxification mechanism, whereby fungi can survive at high concentrations of hazardous metals.

3.3. Monazite Bioleaching Aspergillus niger CECT2807 was grown on monazite from the Morro dos Seis Lagos (Brazil). The Morro dos Seis Lagos deposit is formed by siderite as the main mineral and is covered by a thick ferruginous lateritic crust. This site hosts one of the largest Nb deposits in the world. It also contains iron and manganese and important resources of REE [30]. The evolution of REE and thorium concentrations in the aqueous phase during the bioleaching experiments with A. niger are shown in Metals 2020, 10, 978 8 of 13 Figure 6. It is known that REE-phosphates have particularly low solubilities in water, in the order of 10−13 M (10−11 g/L) [31]. After three days of bioleaching, the REE concentration reach a maximum (0.97 mg/L)A. niger corresponding(Figure6a) [ 11 to]. the A signiﬁcant production REE of losshigher was concentrations observed after fourof citric days and that gluconic may be causedacids by by A. the nigerremoval (Figure of REE 6a) [11]. from A the significant aqueous phase REE loss due was to processes observed such after as four re-precipitation days that may (e.g., be as caused REE-oxalates) by the removalor adhesion of REE to microbial from the cells aqueous [32,33]. Then,phase REE due concentration to processes began such to as increase, re-precipitation probably because (e.g., as the REE-oxalates)fungus needed or the adhesion phosphorous to microbial from the cells rock for[32,33]. its microbial Then, REE growth concentration and phosphorous began concentrationto increase, probablychanged because from 0.70 the mg fungus/L at day needed 9 to 8.80 the mg phosphor/L at dayous 14 (Figurefrom the1). rock On the for other its microbial hand, fungus growth biomass and phosphorouswas able to adsorbconcentration thorium changed generated from during 0.70 mg/L the bioprocess. at day 9 to Other 8.80 mg/L works at evidenced day 14 (Figure that thorium 1). On theis adsorbedother hand, onto fungus the external biomass cell was wall able by to coordination adsorb thorium with generated the nitrogen during of the the chitin bioprocess. present Other in the worksfungal evidenced cell wall [ 34that,35 ].thorium is adsorbed onto the external cell wall by coordination with the nitrogen of the chitin present in the fungal cell wall [34,35].

1.2 0.9 0.8 1 0.7 -1

0.8 -1 0.6 0.5 0.6 0.4 [Th]/ mg·L

[REE]/ mg·L 0.4 0.3 0.2 0.2 0.1 0 0 051015051015 Time / days Time/ days

ab Control A. niger Control A. niger Figure 6. Evolution of REE and thorium concentration released from monazite (1 g/L) in abiotic and Figure 6. Evolution of REE (a) and thorium concentration released (b) from monazite (1 g/L) in abiotic biotic (A. niger) tests. and biotic (A. niger) tests.

MonaziteMonazite is is thethe mainmain source source of thorium,of thorium, which whic is ah radioactiveis a radioactive element. elem Consequently,ent. Consequently, the monazite the monaziteprocessing processing consumes consumes more energy more and energy chemicals and ch thanemicals other than REE other minerals REE because minerals of thebecause requirement of the requirementof additional of separationadditional ofseparati radioactiveon of radioactive thorium. This thorium. processing This processing generates significantgenerates significant amounts of amountshazardous of hazardous wastes, leading wastes, to radioecologicalleading to radioecological issues [36]. Consequently,issues [36]. Consequently, the bioleaching the ofbioleaching REE while ofthorium REE while is adsorbed thorium on is theadsorbed fungus on would the fungus represent would an important represent advantagean important for theadvantage development for the of developmentthe bioprocess. of the bioprocess. MonaziteMonazite was was characterized characterized using using XRD XRD and and the the main main phases phases identified identified were were aluminum aluminum silicate silicate hydroxide,hydroxide, silicon oxide,oxide, aluminum aluminum cerium cerium phosphate phosphate hydroxide, hydroxide, sodium sodium cerium cerium carbonate carbonate fluoride, fluoride,and potassium and potassium hydronium hydronium iron sulphate iron hydroxide sulphate hydroxide (Figure7a). (Figure The REE 7a). ore The minerals REE inore the minerals Morro dosin theSeis Morro Lagos dos deposit Seis Lagos include deposit monazite include and monazite its alteration and products, its alteration such products, as florencite, such a as REE florencite, aluminum a REE aluminum phosphate mineral, and rhabdophane ((Ce,La)PO4·(H2O)) [30]. XRD also confirmed phosphate mineral, and rhabdophane ((Ce,La)PO4 (H2O)) [30]. XRD also confirmed the presence of the presence of REE fluoride carbonates showing ·peaks corresponding to CeCO3F. The mineral REE fluoride carbonates showing peaks corresponding to CeCO3F. The mineral residue after fungal residuebioleaching after wasfungal analyzed bioleaching and the was results analyzed evidenced and the that results the aluminum evidenced cerium that the phosphate aluminum hydroxide cerium phosphatewas transformed hydroxide by A. was niger transformed. Besides the by mineral A. niger phases. Besides in the the substrate, mineral peaksphases corresponding in the substrate, to Ce peaksand La corresponding oxalates were to observed Ce and La in samplesoxalates afterwere twoobserved weeks in of samples incubation after (Figure two weeks7b). of incubation (FigureMonazite 7b). samples treated with the fungus A. niger CECT2807 were examined by FE-SEM (Figure8). A. niger developed a dense mycelium around monazite particles (Figure8a,b). In addition, REE oxalates were precipitated by the metabolites generated during the fungal growth and adsorbed onto the mycelium (Figure8b). The backscattered electron image showed bright areas on the filamentous structure of the fungus corresponding to REE, giving evidence of the REE oxalate precipitation and REE adsorption (Figure8c). Metals 2020, 10, x FOR PEER REVIEW 9 of 13 Metals 2020, 10, x FOR PEER REVIEW 7 of 13

Metals 2020, 10, 978 9 of 13

Metals 2020, 10, x FOR PEER REVIEW 9 of 13

Figure 7. XRD pattern of the monazite (a) before and (b) after 14 days of bioleaching using A. niger (● aluminum silicate hydroxide, ■ silicon oxide, x aluminum cerium phosphate hydroxide, ▲ sodium cerium carbonate fluoride, ♦ potassium hydronium iron sulphate hydroxide, cerium oxalate).

Monazite samples treated with the fungus A. niger CECT2807 were examined by FE-SEM (Figure 8). A. niger developed a dense mycelium around monazite particles (Figure 8a,b). In addition, REE oxalates were precipitated by the metabolites generated during the fungal growth and FigureFigure 7.7. XRDXRD pattern of the monazite Figure ( (aa )) 4.before before X-ray and and diffraction (b (b) )after after 14 14(XRD) days days ofpattern of bioleaching bioleaching of the using usingturquoise A.A. niger niger (a ()● before and (b) after 14 days of adsorbed onto the mycelium (Figure■ 8b). The backscattered electron image showed bright areas▲ on (aluminumaluminum silicate silicate hydroxide, bioleachingsilicon silicon oxide, oxide, usingx aluminum xA. aluminum niger cerium(● potassium cerium phosphate phosphate aluminum hydroxide, hydroxide, silicateN sodium hydroxide, ■ silicon oxide, ♦ copper the filamentous• structure of the fungus corresponding to REE, giving evidence of the REE oxalate ceriumsodium carbonate cerium carbonate ﬂuoride, fluoride,potassium ♦ hydroniumpotassium hydronium iron sulphate iron hydroxide, sulphate hydroxide,cerium oxalate). cerium▲ aluminum phosphate hydroxide hydrate, copper phosphate, copper oxalate). precipitationoxalate). and REE adsorption (Figure 8c). The residues from the inoculated flasks containing turquoise were observed by FE-SEM (Figure Monazite samples treated5). withTurquoise the fungus particles A. wereniger transformedCECT2807 wereand veryexamined dense myceliumby FE-SEM was formed by A. niger, where (Figure 8). A. niger developedcopper a dense phosphate mycelium and copperaround oxalate monazite crys talsparticles were precipitated(Figure 8a,b). and In adsorbed. addition, REE oxalates were precipitated by the metabolites generated during the fungal growth and adsorbed onto the mycelium (Figure 8b). The backscattered electron image showed bright areas on the filamentous structure of the fungus corresponding to REE, giving evidence of the REE oxalate precipitation and REE adsorption (Figure 8c).

Figure 5. Scanning Electron Microscopy (SEM) images of (a) A.niger grown with turquoise after 14 days; (b) detail of precipitates on fungal biomass.

Fungi play an important role in the formation of organic and inorganic secondary minerals through precipitation and deposition of crystalline compounds on and within cells walls, such as oxalates and phosphates. Previous studies evidenced that fungi can generate metal oxalates with different chemical Figure 8. SEM images of (a)elements monazite andparticles minerals after [28].24 h Theunder fungus fungal Beauveria bioleaching, caledonica (b) mycelium can solubilize minerals containing heavy Figure 8. SEM images of (a) monazite particles after 24 h under fungal bioleaching, (b) mycelium developed after 14 days of fungalmetals growth.such as ( ccadmium,) Backscattered copper, electron lead SEMor zinc micrograph and form of oxalates A. niger in the local microenvironment and developed after 14 days of fungal growth. (c) Backscattered electron SEM micrograph of A. niger mycelium. Arrows indicatedin bright association areas on thewith mycelium the mycelium. due to REE This deposition. fungus (daccumulated) SEM image ofhigh amounts of copper as copper mycelium. Arrows indicated bright areas on the mycelium due to REE deposition. (d) SEM image of mycelium encapsulating monazite mineral after 14 days of fungal growth in minimal medium. mycelium encapsulating monaziteoxalate mineral in the after presence 14 days of of copper fungal growthphosphate in minimal [29]. A. medium. niger was able to solubilize metal compounds (ZnO, Zn3(PO4)2 and Co3(PO4)2) and precipitate metal oxalates [26]. The precipitation of metal

The biotransformation of REE-bearing phosphates and the precipitation of lanthanide oxalates could follow the following equation [32]: Figure 8. SEM images of (a) monazite particles after 24 h under fungal bioleaching, (b) mycelium + 2 2 2LnPO +9H O + 2H + 3(C O ) − Ln (C O ) 9H O + 2HPO (2) developed after 14 4days of2 fungal growth. 2(c) 4Backscattered→ 2 2electron4 3· SEM2 micrograph4 of A. niger mycelium. Arrows indicated bright areas on the mycelium due to REE deposition. (d) SEM image of mycelium encapsulating monazite mineral after 14 days of fungal growth in minimal medium.

Metals 2020, 10, x FOR PEER REVIEW 10 of 13

The biotransformation of REE-bearing phosphates and the precipitation of lanthanide oxalates could follow the following equation [32]: Metals 2020, 10, 978 10 of 13 2LnPO4 +9H2O + 2H+ + 3(C2O4)2− → Ln2(C2O4)3·9H2O + 2HPO42 (2)

The formation of REE oxalates limits the long-termlong-term dissolution of these elements. Nevertheless, recent studies have demonstrateddemonstrated that biogenic REE-oxalates can be transformedtransformed through thermal decomposition into REE-oxides that may be precursorsprecursors forfor otherother valuablevaluable REEREE materialsmaterials [[37].37]. In orderorder to to optimize optimize the the solubilization solubilization of REE of fromREE monazitefrom monazite by A. niger by, diA.ff nigererent, growthdifferent conditions growth wereconditions studied. were The studied. bioleaching The ofbioleaching REE by A. of niger REEgrown by A. on niger sterilized grown monazite on sterilized with dextrosemonazite potato with brothdextrose was potato compared broth with was thecompared yield obtained with the when yield the obtained fungus when was grown the fungus with awas minimal grown medium with a (withoutminimal phosphorous).medium (without In addition, phosphorous). the REE release In addition, by the indigenous the REE microorganisms release by the was indigenous monitored inmicroorganisms experiments performed was monitored on non-inoculated in experiments flasks with performed non-sterilized on monazite.non-inoculated Furthermore, flasksA. nigerwith wasnon-sterilized inoculated mona on non-sterilizedzite. Furthermore, monazite A. niger with was the inoculated aim of evaluating on non-sterilized a possible monazite syntrophic with eff theect betweenaim of evaluating the microbial a possible populations syntrophic (Figure effect9). between the microbial populations (Figure 9). When A.A. niger nigerwas was grown grown on dextrose on dextrose potato brothpotato with brot sterilizedh with monazite,sterilized the monazite, REE concentration the REE reachedconcentration a maximum reached of 0.97a maximum mg/L after of 30.97 days. mg/L Furthermore, after 3 days. a significant Furthermore, increase a significant in REE concentration increase in wasREE observedconcentration after was 10 days observed (Figures after6a and10 days9 A. (Figure niger). A.6a nigerand FigureCECT2807 9 A. canniger grow). A. niger using CECT2807 monazite ascan sole grow phosphate using monazite source, as reaching sole phosphate 1.37 mg/ Lsource, of REE reaching (Figure 91.37 Minimal mg/L of medium). REE (Figure Consequently, 9 Minimal themedium). minimal Consequently, medium favors the the minimal fungal dissolutionmedium favors of REE the and fungal seems dissolution to avoid the of notable REE and precipitation seems to ofavoid oxalates the notable observed precipitation when A. niger of wasoxalates grown observed in a rich when medium. A. niger Nevertheless, was grown a slowerin a rich growth medium. was observedNevertheless, when a the slower fungus growth grew inwas minimal observed medium when and the monazite fungus particlesgrew in remained minimal unreacted medium afterand 14monazite days (Figure particles8d). REEremained concentration unreacted in theafter non-inoculated 14 days (Figure flask reached8d). REE a maximum concentration of 0.78 in mg the/L (Figurenon-inoculated9, unsterilized flask monazite).reached a Concentrations maximum of of REE0.78 inmg/L the leachate (Figure when 9, unsterilizedA. niger was monazite). inoculated ontoConcentrations non-sterilized of REE monazite in the wasleachate 0.83 mgwhen/L (FigureA. niger9 ,wasA. nigerinoculatedunsterilized onto non-sterilized monazite). Among monazite all REEwas 0.83 leached, mg/L concentrations (Figure 9, A. niger of cerium unsterilized were the monazite). greatest, Among followed all by REE Nd leached, and La. concentrations When the native of consortiacerium were and A.the niger greatest,were combined,followed by a syntrophic Nd and La. eff ectWhen between the native both populations consortia and leading A. niger to a higher were amountcombined, of REEsa syntrophic would have effect been between expected. both Although populations the resultsleading were to a onlyhigher slightly amount improved, of REEs the would REE precipitationhave been expected. was significantly Although avoided. the results were only slightly improved, the REE precipitation was significantly avoided.

A. niger Unsterilized A. niger 1.6 A. niger Minimal Medium monazite Unsterilized monazite 1.4

1.2

-1 1

0.8 [Nd] [La] 0.6 [REE]/ mg·l [REE]/ [Ce] 0.4

0.2

0 01234681014 01234681014 01234681014 01234681014 Time / days Time/ days Time/ days Time/ days

Figure 9. Total concentration of rare earth elements, cerium (Ce), lanthanum (La), and neodymium Figure 9. Total concentration of rare earth elements, cerium (Ce), lanthanum (La), and neodymium (Nd) bioleached during the experiments under different conditions. (Nd) bioleached during the experiments under diﬀerent conditions.

Previous works have determined remarkable alterationsalterations in the natural microbialmicrobial populationspopulations during bioleaching of monazite ores. The The existence of native Firmicutes on the monazite seems to have significantlysignificantly contributed to the increase in REEREE leachingleaching observedobserved whenwhen usingusing non-sterilizednon-sterilized monazite [[38].38]. The leachingleaching yields yields and and rates rates in bioleachingin bioleaching are usuallyare usually lower lower in comparison in comparison to chemical to chemical leaching usingleaching strong using (in)organic strong (in)organic acids or acids chelants. or chelants. Sulfuric Sulfuric acid is acid often is preferred often preferred to other to inorganicother inorganic acids becauseacids because it dissolves it dissolves fewer fewer impurities, impurities, such such as Caas Ca and and Sr Sr [6 ].[6]. However, However, bioleaching bioleaching ooffersffers some advantages over chemical leaching in the case of certain low-grade REE ores; especially, the secondary sources. Bioleaching of waste phosphors and cracking catalysts using biogenic gluconic acid presented Metals 2020, 10, 978 11 of 13 higher efficiency than chemical leaching using pure organic acids at even higher concentrations, due to the presence of other metabolites in the leaching agent [39]. Techno-economic and life cycle analysis indicated that a bioleaching plant using agricultural or food wastes is an economical and environmental viable alternative for future REE recovery [40]. The use of waste products as alternative substrates for microbial metabolism may be more profitable and cleaner for several impact categories, with the same leaching efficiency but remarkably reduced processing costs and impacts.

4. Conclusions This work has shown that valuable metals from mineral phosphates were mobilized using A. niger CECT2807. The phosphorous leaching rates were aluminum phosphate > turquoise > monazite. According to our results, an important amount of copper was recovered from turquoise. A. niger leached also REE from monazite, obtaining the best results when the fungus was cultivated on minimal medium with the mineral as the sole phosphorous source. The XRD results evidenced that A. niger was able to leach the copper aluminum phosphate hydroxide hydrate (turquoise) and the aluminum cerium phosphate hydroxide in monazite. Nevertheless, part of the leached copper and REE were precipitated as secondary minerals such as phosphates and oxalates. Bioleaching using phosphate solubilizing microorganisms could be an alternative to conventional methods for valuable metals extraction.

Author Contributions: Conceptualization, M.L.B.; Funding acquisition, J.A.M.; Investigation, L.C.; Methodology, J.A.M.; Project administration, F.G.; Resources, M.L.B.; Validation, F.G.; Writing—review and editing, L.C. and J.A.M. All authors have read and agreed to the published version of the manuscript. Funding: This work was supported by the Complutense University of Madrid (project PR87/19-22648). Conﬂicts of Interest: The authors declare no conﬂict of interest.

References

1. Goodenough, K.M.; Wall, F.; Merriman, D. The rare earth elements: Demand, global resources, and challenges for resourcing future generations. Nat. Resour. Res. 2018, 27, 201–216. [CrossRef] 2. Charalampides, G.; Vatalis, K.I.; Apostoplos, B.; Ploutarch-Nikolas, B. Rare earth elements: Industrial applications and economic dependency of Europe. Procedia Econ. 2015, 24, 126–135. [CrossRef] 3. U.S. Department of the Interior and U.S. Geological Survey. Critical Mineral Resources of the United States—Economic and Environmental Geology and Prospects for Future Supply. In Professional Paper 1802; U.S. Department of the Interior and U.S. Geological Survey: Reston, VA, USA, 2017; p. 797. 4. European Commission. Communication from the Commission to the European Parliament, the Council, the European Economic and Social Committee and the Committee of the Regions on the 2017 list of Critical Raw Materials for the EU. 2017. Available online: https://ec.europa.eu/transparency/regdoc/rep/1/2017/EN/ COM-2017-490-F1-EN-MAIN-PART-1.PDF (accessed on 17 July 2020). 5. Balaram, V. Rare earth elements: A review of applications, occurrence, exploration, analysis, recycling, and environmental impact. Geosci. Front. 2019, 10, 1285–1303. [CrossRef] 6. Peelman, S.; Sun, Z.H.I.; Sietsma, J.; Yang, Y. Chapter 21-Leaching of rare earth elements: Review of past and present technologies A2-Lima, Ismar Borges De. In Rare Earths Industry; Filho, W.L., Ed.; Elsevier: Boston, MA, USA, 2016; pp. 319–334. 7. Watling, H. Microbiological Advances in Biohydrometallurgy. Minerals 2016, 6, 49. [CrossRef] 8. Zhuang, W.-Q.; Fitts, J.P.; Ajo-Franklin, C.M.; Maes, S.; Alvarez-Cohen, L.; Hennebel, T. Recovery of critical metals using biometallurgy. Current Opin. Biotech. 2015, 33, 327–335. [CrossRef][PubMed] 9. Corbett, M.K.; Eksteen, J.J.; Niu, X.-Z.; Croue, J.-P.; Watkin, E.L.J. Interactions of phosphate solubilising microorganisms with natural rare-earth phosphate minerals: A study utilizing Western Australian monazite. Bioprocess Biosyst. Eng. 2017, 40, 929–942. [CrossRef] 10. Shin, D.; Kim, J.; Kim, B.S.; Jeong, J.; Lee, J.C. Use of phosphate solubilizing bacteria to leach rare earth elements from monazite-bearing ore. Minerals 2015, 5, 189–202. [CrossRef] Metals 2020, 10, 978 12 of 13

11. Brisson, V.L.; Zhuang, W.Q.; Alvarez-Cohen, L. Bioleaching of rare earth elements from monazite sand. Biotechnol. Bioeng. 2016, 113, 339–348. [CrossRef][PubMed] 12. Fathollahzadeh, H.; Eksteen, J.J.; Kaksonen, A.H.; Watkin, E.L.J. Role of microorganisms in bioleaching of rare earth elements from primary and secondary resources. Appl. Microbiol. Biotechnol. 2018, 103, 1043–1057. [CrossRef] 13. Xu, T.-J.; Ramanathan, T.; Ting, Y.-P. Bioleaching of incineration fly ash by Aspergillus niger-precipitation of metallic salt crystals and morphological alteration of the fungus. Biotechnol. Rep. (Amst.) 2014, 3, 8–14. [CrossRef] 14. Mouna, H.M.; Baral, S.S. A bio-hydrometallurgical approach towards leaching of lanthanum from the spent fluid catalytic cracking catalyst using Aspergillus niger. Hydrometallurgy 2019, 184, 175–182. 15. Faraji, F.; Golmohammadzadeh, R.; Rashchi, F.; Alimardani, N. Fungal bioleaching of WPCBs using Aspergillus niger: Observation, optimization and kinetics. J. Environ. Manag. 2018, 217, 775–787. [CrossRef] [PubMed] 16. Gentina, J.C.; Acevedo, F. Copper bioleaching in Chile. Minerals 2016, 6, 23. [CrossRef] 17. Yin, S.; Wang, L.; Kabwe, E.; Chen, X.; Yan, R.; An, K.; Zhang, L.; Wu, A. Copper bioleaching in China: Review and Prospect. Minerals 2018, 8, 32. [CrossRef] 18. Delvasto, P.; Valverde, A.; Ballester, A.; Muñoz, J.A.; González, F.; Blázquez, M.L.; Igual, J.M.; García-Balboa, C. Diversity and activity of phosphate bioleaching bacteria from a high-phosphorus iron ore. Hydrometallurgy 2008, 92, 124–129. [CrossRef] 19. Attfield, J.P. Phosphates. In Encyclopedia of Materials: Science and Technology; Buschow, K.H.J., Cahn, R.W., Flemings, M.C., Ilschner, B., Kramer, E.J., Mahajan, S., Veyssière, P., Eds.; Elsevier: Oxford, UK, 2001; pp. 6896–6901. 20. Ruttenberg, K.C. 8.13-The global phosphorus cycle. In Treatise on Geochemistry; Holland, H.D., Turekian, K.K., Eds.; Pergamon: Oxford, UK, 2003; pp. 585–643. 21. Alori, E.T.; Glick, B.R.; Babalola, O.O. Microbial phosphorus solubilization and its potential for use in sustainable agriculture. Front. Microbiol. 2017, 8, 971. [CrossRef] 22. Ivanova, R.P.; Bojinova, D.Y.; Gruncharov, I.N.; Damgaliev, D.L. The solubilization of rock phosphate by organic acids. Phosphorus Sulfur 2006, 181, 2541–2554. [CrossRef] 23. Welch, S.A.; Taunton, A.E.; Banfield, J.F. Effect of microorganisms and microbial metabolites on apatite dissolution. Geomicrobiol. J. 2002, 19, 343–367. [CrossRef] 24. Show, P.L.; Oladele, K.O.; Siew, Q.Y.; Aziz Zakry, F.A.; Lan, J.C.-W.; Ling, T.C. Overview of citric acid production from Aspergillus niger. Front. Life Sci. 2015, 8, 271–283. [CrossRef] 25. Sayer, J.A.; Gadd, G.M. Binding of cobalt and zinc by organic acids and culture filtrates of Aspergillus niger grown in the absence or presence of insoluble cobalt or zinc phosphate. Mycol. Res. 2001, 105, 1261–1267. [CrossRef] 26. Sayer, J.A.; Gadd, G.M. Solubilization and transformation of insoluble inorganic metal compounds to insoluble metal oxalates by Aspergillus niger. Mycol. Res. 1997, 101, 653–661. [CrossRef] 27. Illmer, P.; Schinner, F. Solubilization of inorganic calcium phosphates—Solubilization mechanisms. Soil Biol. Biochem. 1995, 27, 257–263. [CrossRef] 28. Gadd, G.M.; Bahri-Esfahani, J.; Li, Q.; Rhee, Y.J.; Wei, Z.; Fomina, M.; Liang, X. Oxalate production by fungi: Significance in geomycology, biodeterioration and bioremediation. Fungal Biol. Rev. 2014, 28, 36–55. [CrossRef] 29. Fomina, M.; Hillier, S.; Charnock, J.M.; Melville, K.; Alexander, I.J.; Gadd, G.M. Role of oxalic acid overexcretion in transformations of toxic metal minerals by Beauveria caledonica. Appl. Environ. Microbiol. 2005, 71, 371–381. [CrossRef][PubMed] 30. Takehara, L.; Silveira, F.V.; Santos, R.V. Chapter 4-Potentiality of rare earth elements in Brazil. In Rare Earths Industry; Borges De Lima, I., Leal Filho, W., Eds.; Elsevier: Boston, MA, USA, 2016; pp. 57–72. 31. Firsching, F.H.; Brune, S.N. Solubility products of the trivalent rare-earth phosphates. J. Chem. Eng. Data 1991, 36, 93–95. [CrossRef] 32. Kang, X.; Csetenyi, L.; Gadd, G.M. Monazite transformation into Ce- and La-containing oxalates by Aspergillus niger. Environ. Microbiol. 2020, 22, 1635–1648. [CrossRef] 33. Xu, S.; Zhang, S.; Chen, K.; Han, J.; Liu, H.; Wu, K. Biosorption of La3+ and Ce3+ by Agrobacterium sp. HN1. J. Rare Earth 2011, 29, 265–270. [CrossRef] Metals 2020, 10, 978 13 of 13

34. Tsezos, M.; Volesky, B. The mechanism of thorium biosorption by Rhizopus arrhizus. Biotechnol. Bioeng. 1982, 24, 955–969. [CrossRef] 35. Bhainsa, K.C.; D’Souza, S.F. Thorium biosorption by Aspergillus fumigatus, a filamentous fungal biomass. J. Hazard. Mater. 2009, 165, 670–676. [CrossRef] 36. Mohd Salehuddin, A.H.J.; Ismail, A.F.; Che Zainul Bahri, C.N.A.; Aziman, E.S. Economic analysis of thorium extraction from monazite. Nucl. Eng. Technol. 2019, 51, 631–640. [CrossRef] 37. Kang, X.; Csetenyi, L.; Gadd, G.M. Biotransformation of lanthanum by Aspergillus niger. Appl. Microbiol. Biotechnol. 2019, 103, 981–993. [CrossRef][PubMed] 38. Corbett, M.K.; Eksteen, J.J.; Niu, X.-Z.; Watkin, E.L.J. Syntrophic effect of indigenous and inoculated microorganisms in the leaching of rare earth elements from Western Australian monazite. Res. Microbiol. 2018, 169, 558–568. [CrossRef][PubMed] 39. Reed, D.W.; Fujita, Y.; Daubaras, D.L.; Jiao, Y.; Thompson, V.S. Bioleaching of rare earth elements from waste phosphors and cracking catalysts. Hydrometallurgy 2016, 166, 34–40. [CrossRef] 40. Jin, H.; Reed, D.W.; Thompson, V.S.; Fujita, Y.; Jiao, Y.; Crain-Zamora, M.; Fisher, J.; Scalzone, K.; Griffel, M.; Hartley, D.; et al. Sustainable bioleaching of rare earth elements from industrial waste materials using agricultural wastes. ACS Sustain. Chem. Eng. 2019, 7, 15311–15319. [CrossRef]

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