the role of bacteria in the supergene environment of the

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IntroductionALTHOUGH traditional genetic models of supergene enrich-ment have followed an inorganic geochemical approach toore-forming processes (Brimhall et al., 1985), it is reasonableto ask what role organic processes may have played. Titley(1975) postulated a link between microorganisms and therapid rates of copper leaching in porphyry copper districts inthe southwestern Pacific region where high precipitation anderosion rates occur. Acidophilic iron- and sulfur-oxidizingbacteria (thiobacilli: e.g., Acidithiobacillus thiooxidans, a sul-fur oxidizer, Acidithiobacillus ferrooxidans, an iron and sulfuroxidizer, and Leptospirillum ferrooxidans, an iron oxidizer),

common in acidic environments containing pyritic minewaste (Edwards et al., 1999), play a fundamental role in theformation of acid mine drainage (Colmer and Hinkle, 1947;Jambor and Blowes, 1984; Sand et. al., 1995; Schippers et. al.,1996; Nordstrom and Southam, 1997; Gehrke et. al., 1998;Schippers and Sand, 1999) and presumably contribute to su-pergene weathering. These aerobic bacteria are lithoau-totrophic, i.e., they get their energy from the oxidation of in-organic constituents and utilize this energy to fix carbondioxide into cell biomass (growth). In acid mine drainage sys-tems, thiobacilli are responsible for the generation of sulfuricacid and the concurrent release of heavy metals (Lizama andSuzuki, 1989; Norris and Kelly, 1982) from a wide range ofsulfide minerals (Lawrence et al., 1997). When they are pre-sent under acidic conditions (pH <4; Kirby et al., 1999), the

The Role of Bacteria in the Supergene Environment of the Morenci Porphyry Copper Deposit, Greenlee County, Arizona

M. STEPHEN ENDERS,†

Newmont Mining Corporation, 1700 Lincoln Street, Denver, Colorado 80203

CHRIS KNICKERBOCKER,Department of Biological Sciences, Northern Arizona University, Flagstaff, Arizona 86011-5640

SPENCER R. TITLEY,Department of Geosciences, University of Arizona, Tucson, Arizona 85721

AND GORDON SOUTHAM

Department of Earth Sciences, University of Western Ontario, London, Ontario, Canada N6A 5B7

AbstractGeological and microbiological examination of water and mineral samples collected primarily along the 5200

bench of the Metcalf pit within the Morenci copper deposit revealed that acidophilic iron oxidizing bacteria(thiobacilli) and sulfate-reducing bacteria have contributed to leaching and, in part, to the enrichment of cop-per in the supergene environment, respectively. The 5200 bench traverses a classic, but tilted, enrichment pro-file consisting of a 200-m-thick zone of leached capping (0.06% Cu, 0.06% S) that overlies a partially leached,180-m-thick enriched blanket (0.42% Cu, 0.39% S). The modern climate in southwestern Arizona is semiaridand exhibits a biannual wet-dry cycle where active weathering zones, associated with fracture-flow regimesalong the 5200 bench, provided a natural laboratory to study supergene processes. Samples from these weath-ering zones contained viable thiobacilli that thrived during wet periods of the year reaching populations >107

bacteria/ml within an environment where the pH was diluted to near-neutral conditions. The population ofthiobacilli decreased during the dry periods, presumably due to low water activity. During dry periods, evapo-rative concentration of sulfuric acid also promoted sulfide mineral dissolution and the formation of sulfate salts(e.g., chalcanthite). During the subsequent wet phase, these salts dissolved, contributing to the soluble, mobilefraction of copper important to supergene enrichment. Acidification of the sulfide zone was also promoted bythe formation of authigenic iron hydroxides such as goethite and jarosite, similar to acid mine drainage systems.Order of magnitude estimates of contemporary bacterial iron oxidation of a 100-m3 block in the active weath-ering environment reveals that an oxidized cap could be produced in as little as 9 × 102 to 5 × 103 yr. While itis interesting to think that leaching at Morenci could form on a time scale of several thousand years, it is in-correct to assume that an entire 100-m3 block could be active at one time (i.e., leaching occurs in discrete frac-tures). The most active fracture in the Metcalf pit, Morenci, which produces a few kilograms of bacteria/yr,could leach between 0.14 and 0.87 t of Cu annually. Small populations of viable sulfate-reducing bacteria(~103/g) occurred within this partially leached zone and at the ground-water interface (i.e., at the redox bound-ary). Even though the enriched blanket possesses viable sulfate-reducing bacteria, inorganic geochemicalprocesses dominate supergene enrichment in this system. Comparing the annual carbon fixation occurring dur-ing biooxidation versus leaching, bacterial sulfate reduction could only fix between 0.2 and 1.1 percent of thecopper being leached.

† Corresponding author: e-mail, [email protected]

©2006 Society of Economic Geologists, Inc.Economic Geology, v. 101, pp. 59–70

oxidation of pyrite can be 105 times faster than the abiotic rate(Singer and Stumm, 1970).

The opposing bacterial process is carried out by anaerobic,dissimilatory sulfate-reducing bacteria that oxidize low mole-cular weight organic compounds utilizing sulfate as their ter-minal electron acceptor, forming and releasing hydrogen sul-fide as a byproduct of metabolism (Donald and Southam,1999). Alpers and Brimhall (1989) proposed that sulfate-re-ducing bacteria activity might explain a thin zone of massivechalcocite at the top of the enrichment blanket at La Escon-dida, Chile. Lichtner and Biino (1992) also questioned thepotential role of sulfate-reducing bacteria in the direct pre-cipitation of pyrite, bornite, and chalcocite in enrichmentblankets. Sillitoe et al. (1996) proposed bacteria as mediatorsof copper sulfide enrichment based on a scanning electronmicroscope study of chalcocite from copper deposits in north-ern Chile. In a related study, Bawden et al. (2003) used sulfurisotopes to demonstrate the involvement of sulfate-reducingbacteria in the enrichment of Zn at the Mike gold deposit,Carlin Trend, Nevada. Dissimilatory sulfate-reducing bacte-ria occur naturally in acid mine drainage sites (Fortin et al.,1995, 1996; Fortin and Beveridge, 1997; Donald andSoutham, 1999) and are responsible for precipitation of dis-solved heavy metals in natural environments (Labrenz et al.,2000; Druschel et al., 2002) or have been used in engineeredsystems to clean up acid mine drainage sites by precipitatingmetal sulfides (Dvorak et al., 1992; Hammack et al., 1994;Benner et al., 2000).

Thiobacilli and sulfate-reducing bacteria are naturally oc-curring and will be enriched to populations exceeding 105

cells/ml (or /g for soils and sediments) under favorable growthconditions (Southam and Beveridge, 1992), where they canhave a profound affect on geochemical processes. However,the question remains, what role have they played in the for-mation of supergene copper deposits? An A. ferrooxidansspp., recovered from Morenci, is capable of oxidizing pyriteunder circumneutral pH laboratory conditions, causing acidi-fication of the bulk fluid phase (Mielke et al., 2003). Also,Melchiorre and Enders’ (2003) oxygen isotope thermometrydata (20°–34°C) suggest that bacterial growth could occur inMorenci under incubatorlike conditions, which would pro-mote near-optimum mesophilic microbial activity (Southamand Beveridge, 1992; Fortin et al., 1995; Nordstrom andSoutham, 1997). The goal of the present study was to exam-ine the role of thiobacilli and sulfate-reducing bacteria inweathering and in the supergene enrichment of copper, re-spectively, at Morenci, Arizona.

Geologic SettingThe Morenci district (Fig. 1) hosts a porphyry copper de-

posit that was formed by magmatic-hydrothermal processesabout 55 m.y. ago (McCandless and Ruiz, 1993). Widespreadhypogene mineralization as pyrite + chalcopyrite ± sphaleritewas deposited in a stockwork of veins and fractures in Pre-cambrian granite, Paleozoic sedimentary rocks, andLaramide-age felsic intrusive rocks over a 19-km2 area. Theresulting stockwork of veins and veinlets has a fracture den-sity ranging from about 0.10 to over 1.0/cm (length/area). Hy-pogene mineralization was accompanied by pervasive quartz-sericite-pyrite alteration that resulted in a deposit with

average contents of 3 wt percent pyrite and 0.16 wt percentcopper as chalcopyrite.

Supergene mineralization at Morenci was formed by thecoupled processes of erosion and chemical weathering duringat least three periods of enrichment and four periods of leach-ing (Enders, 2000). These processes began about 55 Ma andended with the onset of middle Tertiary volcanism. Super-gene enrichment resumed at about 20 Ma when middle tolate Tertiary extension reexposed the deposit (Moolick andDurek, 1966; Langton, 1973; Cook, 1994; Enders, 2000). Ox-idation and enrichment produced a classic supergene profile(Titley and Marozas, 1995) and a three-fold average increasein copper grade. This classic profile contains a copper-poor,limonitic leached cap that overlies an enriched blanket ofhigher grade chalcocite ± covellite mineralization (0.42% Cu)as replacements and coatings of the primary chalcopyrite andpyrite. During subsequent enrichment cycles, copper fromthe enriched blankets was leached and transported to a lowerwater table to make a thicker and higher grade enrichmentblanket, leaving hematite after chalcocite in the former blan-ket. In areas with low pyrite to chalcocite ratios (<2–1) earlierenriched blankets were oxidized inplace to form brochantite,malachite, azurite, chrysocolla, tenorite, and other copper ox-ides (Melchiorre and Enders, 2003).

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FIG. 1. Location map showing the Clifton-Morenci area in a regional con-text. The inset map shows the location of the principal subdivisions of theMorenci district in outline and with mine coordinates (in feet) for scale.

Metcalf geology

Mining in the Metcalf area has exposed a classic, but tilted,enrichment profile (Fig. 2) that is well displayed on the 5200bench along the south side of the Metcalf pit. Supergene min-eralization and argillic alteration are superimposed onstrongly fractured and quartz-sericite-pyrite–altered, Laramide-age monzonite porphyry and older granite porphyry, and Pro-terozoic granite. The War Eagle fault zone has both offset theenriched blanket and enhanced permeability along the zone,resulting in a sharp transition from strong leached capping inthe hanging wall to enriched sulfides in the footwall.

The leached cap contains abundant iron oxides, including astockwork of quartz-hematite + goethite ± jarosite veins andveinlets, left behind as a result of nearly complete leaching ofthe precursor pyrite and chalcocite mineralization. Boxworksof hematite after chalcocite, and quartz after pyrite are com-mon throughout the leached capping along the 5200 bench.

The hematite boxworks have been interpreted to representthe destruction of a preexisting enriched blanket at Metcalfduring the Miocene to the Pliocene (Moolick and Durek,1966; Langton, 1973; Cook, 1994; Titley and Marozas, 1995;Enders, 2000).

The enriched blanket contains disseminated pyrite andchalcocite mineralization in the matrix of the rock and astockwork of 0.1- to 10-mm-wide veins and veinlets of chal-cocite, djurelite, and covellite that both coat and totallyreplace pyrite and chalcopyrite. In addition, very thin molyb-denite veinlets crosscut the earlier quartz-sericite-pyrite-chal-cocite veins. Late-stage, coarse-grained quartz-pyrite veinsare also common throughout this part of the deposit.

Partial leaching in the upper 90 m of the enriched blankethas resulted in a rock mass that contains a mixture ofhematite, goethite, and jarosite that partially, to completelyreplace chalcocite and pyrite in places. Partial leaching is ev-ident as relatively narrow, oxidized fractures that penetrate

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FIG. 2. Cross section 15,600 N looking north across the Metcalf area demonstrating the supergene mineral profile, pitlimit as of August 1998, the water table as of April 1995 (dashed line), and sample locations along the 5200 bench. Inset isthe plan map of the Metcalf 5200 bench (labeled with the sample sites), showing its position in the Metcalf pit along thesouthern pit wall. Sample locations for this study are shown at the toe of the bench. Sample locations are labeled in the planmap and cross section as follows: 1 = MET-5200-LC01, 2 = MET-5200-LC02, 3 = MET-5200-LC03, 4 = MET-5200-PL01,5 = MET-5200-EB01, 6 = MET-5200-EB02, 7 = MET-5200-EB03, 8 = MET-5200-EB04, 9 = MET-5200-EB05, 10 = MET-5200-EB06, and 11 = MET-5200-PL02. Bench faces are shaded in dark gray and pit floors are shaded in white. The refer-ence line for cross section 15,600 N is located just south of the pit wall. Elevations are in feet.

approximately 180 to 270 m into the enriched blanket ex-posed on the east side of the Metcalf pit wall. This style ofleaching is consistent with a drop in the water table, as a re-sult of downcutting of the Gila River during the Pleistocene(Enders, 2000).

Recent mining has exposed all of these zones and causedthe water table to further drop in the Metcalf pit area. Seepsand springs along faults and fractures above the current watertable are accompanied by strong oxidation of chalcocite andpyrite in the enriched blanket. Oxidation has formed jarosite± goethite after pyrite, chalcanthite after chalcocite, and leftgoethite and hematite behind in the leached fractures andiron hydroxides plus copper sulfates on the adjacent pit floorand walls (Enders, 2000).

Physiography

The Morenci district is located on the southeastern edge ofthe Transition zone physiographic province between the Col-orado Plateau to the north and the Basin and Range to thesouth (Fig. 1). Elevations average about 1,700 m above sealevel (asl) and range from a high of about 2,100 m in the northto a low of about 1,035 m in the south. The district is locatedin the Gila River drainage basin and is partially surrounded bythree perennial streams: the Gila River, Eagle Creek, and theSan Francisco River. Prior to mining, a southerly flowing in-termittent stream (Chase Creek) occupied an axial positionthrough the center of the district. As a result, there are strongtopographic and hydrologic gradients affecting the supergeneenvironment at Morenci.

Hydrology

The hydrogeologic system in the Morenci sub-basin is char-acterized almost entirely by fracture flow in consolidatedbedrock. Ground water originates as infiltration from rainfalland snowmelt in the higher elevations of the basin and flowstoward the lower elevations where it appears as springs andprovides underflow to perennial streams. Ground-water ele-vations tend to follow topography and are highest in thenorthern part of the district and lowest near the perennialstreams on the eastern, southern, and western boundaries ofthe district. In the Metcalf pit area, ground-water depthsrange from about 80 to 260 m below the premine topography.As a result, ground water has a near-neutral pH and is domi-nated by calcium and sulfate, with lower concentrations ofsodium, potassium, chloride, magnesium, carbonate, and bi-carbonate (Aquifer Protection permit application, PhelpsDodge Morenci district, Arizona Department of Environ-mental Quality, Dames and Moore Inc., March 28, 1996).

Climate

The climate of the Morenci district is semiarid and typicalof the Mexican Highland portion of the Basin and Rangewhere a wide range of conditions exist that are directly linkedto variations in altitude and geomorphology. Average dailyminimum and maximum temperatures for the region are –5o

to 15oC in January and 17o to 35oC in July (Remick, 1989).Annual precipitation at Morenci has averaged 33 cm over a43-yr period (Phelps Dodge Morenci, Inc., 1996, monthlyand annual precipitation records for the period January 1953-March 1996), while the lake evaporation rate in the Duncan

valley is about 15 to 17 cm/yr (Anderson et. al., 1992). Mostof the precipitation occurs in July, August, and Septemberduring the annual monsoon season and again in Decemberthrough February from winter storms (Fig. 3). As a result,there are strong, seasonal, warm to cold, and wet to dry cyclesaffecting the supergene environment at Morenci.

Methods

Sampling

A wide variety of materials (natural waters, ground waterfrom monitoring wells, process solutions and solids, seeps,springs, water-saturated weathered products and rocks) weresampled to characterize background populations of A. fer-rooxidans in the natural and mining environment before fo-cusing in on key exposures in the Metcalf pit (Fig. 2). Theseeps, springs, and rocks sampled in the Metcalf pit occurredalong fractures in the vadose zone across an actively weather-ing sulfide-bearing enriched zone.

Sampling was carried out in two stages between November1997 and November 1999. The first stage determined thepresence and levels of A. ferrooxidans in background watersof the Morenci area outside the Metcalf pit, in waters andmineral samples within the deposit across the supergene pro-file exposed on the 5200 bench, and at different times duringthe year. The second stage examined the distribution of sul-fate-reducing bacteria in the same sample locations and innew areas as mining progressed. Samples for microbiologicalstudies were collected in sterile 50-ml polypropylene tubes.Samples for sulfate-reducing bacteria were processed onsitedue to the oxygen stress associated with sampling, whereasthe samples for thiobacilli were stored at 4°C until processed.Water samples for chemical analysis were collected by fillingclean, dry 1,000-ml polypropylene bottles (minimal head-space) and placed on ice for shipment to the lab.

Enumeration of thiobacilli and sulfate-reducing bacteria

During the orientation survey, selective media, designed toenumerate iron or sulfur oxidizing bacteria (Southam andBeveridge, 1992) were employed to characterize the thio-bacilli in the Morenci district. The positive cultures from this

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FIG. 3. Morenci daily precipitation record from October 1, 1997, throughAugust 31, 1999. Dashed lines with dates mark the four sampling events formicroorganisms during this period.

survey were able to oxidize both iron and elemental sulfur,putting them in the A. ferrooxidans group. Samples possess-ing A. ferrooxidans were serially diluted and grown in quin-tuplicate in a selective growth medium which consisted of 9Kbuffer [0.4 g (NH4)2SO4, 0.1 g K2HPO4, 0.4 g MgSO4·7H2Oper L] , pH 2.3 (Silverman and Lundgren, 1959) containing33.3 g/l FeSO4·7H2O. Positive tubes demonstrating growth(e.g., iron oxidation) are scored for each dilution and the mostprobable number score is assigned using statistical tables pro-vided by Cochran (1950), which take into account the dilutionand the number of tubes possessing positive growth, and hasa resolution down to 0.2 bacteria/g. The most probable num-ber method produces a cell count, and the data are reportedas viable bacteria/g or /ml. Data from mixed samples of solidsin water are reported as viable bacteria/gram. The A. ferroox-idans cultures were grown at 25°C for 6 weeks to ensure thatthe endpoint of growth had been reached.

Dissimilatory sulfate-reducing bacteria were serially di-luted in anaerobic saline (8-ml saline plus 1 ml of a reducingagent, described below) to protect them from the toxic effectsof oxygen and enumerated using the most probable numbermethod and the following selective culture medium: 10 g/lBacto® Tryptone, 2 g/l MgSO4·7H2O, 0.5 g/l FeSO4·7H2O,and 6 ml/l 60 percent(aq) sodium lactate, pH 7.5 (Fortin et al.,1995). A reducing agent supplement (RAS, 7.5 g/l ascorbicacid and 7.5 g/l thioglycollic acid, pH 7.5) was added to themedium at a concentration of 10 percent (vol/vol). The sul-fate-reducing bacteria cultures were grown at 25°C for 3weeks to ensure that the endpoint of growth was reached.

Microscopic examination and energy dispersive X-ray spectroscopy (EDS)

Rock samples were examined using conventionally polishedthin sections and reflected light microscopy.

The samples of weathered mineral products collected forscanning electron microscopy (SEM) transmission electronmicroscopy (TEM) were fixed with 1 percent (vol/vol) glu-taraldehyde, washed once using distilled water, embedded in2 percent low-melt agarose, dehydrated using a 100 percentacetone dehydration series and embedded in Epon 812 resin(Graham and Beveridge, 1990). Ultra-thin sections (70 nm)were cut using a Reichert-Jung® Ultracut E ultramicrotome,placed on Formvar carbon-coated 200-mesh nickel grids andviewed unstained in a JEOL®-1200EX TEM. Samples forSEM-EDS were applied directly to an aluminum SEM stuband allowed to dry under vacuum in the SEM chamber. EDSwas performed using a Quantum Kevex® 3300 light elementdetector attached to a LEO®-435VP SEM and quantifiedusing a Kevex® software program to determine the relativesample composition.

Water chemistry

Five samples of seep and spring waters were shipped toMcKenzie Laboratories or Bolin Laboratory in Phoenix, Ari-zona, for analysis. The samples were filtered (0.1 µm) and an-alyzed for pH, total dissolved solids (TDS), major cations andanions, alkalinity, and the following dissolved metals (Sb, As,Be, Cd, Cr, Pb, Se, Al, Ba, Ca, Cu, Fe, Mg, Mn, Ni, K, Na,Zn, Hg, Ag, Tl). Sampling and analytical procedures followedappropriate EPA test methods. Laboratory pH measurements

(Denver Instruments Basic® pH meter) were made directlyon liquid samples. In order to estimate the likely pH of waterafter a rain event in dry areas, 1 ml of distilled water wasadded to 1 g of solid sample (weathered products or sedi-ment), vortexed for 1 min, and allowed to stand for 1 min (seeSoutham and Beveridge, 1993).

Acid and/or base accounting and rock geochemistry

A total of 14 rock samples from the Metcalf 5200 bench andpit were submitted to SVL Analytical Inc. in Kellogg, Idaho,for geochemical analysis. The samples were analyzed for totalsulfur and soluble sulfur forms, and the acid generating andacid neutralization potentials of the samples were measuredby SVL, using standard accepted industry and EPA analyticalprocedures as summarized in Jennings and Dollhopf (1995).In addition, splits of those 14 samples plus four additionalsamples were analyzed by the Phelps Dodge Analytical Ser-vices laboratory in Morenci for total copper, acid-soluble cop-per, molybdenum, ferric-soluble copper, iron, and total sulfur.The acid-soluble method provides an estimate of the amountof oxide copper minerals (i.e., chrysocolla, malachite, andazurite) leachable in a dilute solution containing H2SO4. Theferric-soluble method provides an estimate of the amount ofcopper sulfide minerals (i.e., chalcocite, covellite) leachablein a dilute solution containing H2SO4 and Fe2 (SO4)3.

Results

Orientation studies

Samples of the Morenci water supply and upgradientground-water monitor wells around the district showed near-neutral pH and no viable A. ferrooxidans (data not shown;Table 1). However, viable A. ferrooxidans populations, up to10 bacteria/ml, were cultured from circumneutral pH watercollected from some of the downgradient water wells in theChase Creek stockpile and Tailings Dam areas. Bacterial pop-ulations in acidic (pH 1.8), solvent extraction/electrowinning(SX/EW) process fed solutions were greater than backgroundlevels and ranged between 102 to approximately 103

thiobacilli/ml. Background bacterial populations in seeps,springs, and weathered mineral products from five of themost accessible sites in the three pits contained up to 105

thiobacilli/g and were associated with acidic conditions, whichranged from pH 2.1 to 5.4.

Populations of A. ferrooxidans and sulfate-reducing bacteria at Metcalf

Populations of viable A. ferrooxidans ranged from less thandetection (0.2 bacteria/g) up to 3.5 × 107 bacteria/g (Table 2,Fig. 4). A. ferrooxidans was not detected at any of the sites inleached capping or in the partially leached zone at the top ofthe enriched blanket. However, populations increased almostsix orders of magnitude in the enriched blanket and were stillpresent in the partially leached fracture zone. Interestingly,no viable A. ferrooxidans were recovered from MET-5200-EB04, a site that contained pyrite in the groundmass butnone in veins or on open fracture surfaces (data not shown)where water and bacteria could not access this material.

Seasonal effects: Sample pH and A. ferrooxidans popula-tions showed a significant seasonal effect (Figs. 3–5). During


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the wet season, the sample pH values were near neutral toslightly acidic; however these values became more acidic dur-ing the dry season (cf. Figs. 3, 5). This seasonal effect wasmore pronounced in the enriched blanket than in the leachedcapping and upper, partially leached zone. A similar patternoccurred in the A. ferrooxidans populations, where the bacte-ria thrived during wet periods, but the populations dropped

off several orders of magnitude during the dry seasons (cf.Figs. 3, 4).

TEM observations of unstained ultra-thin sections revealedthe presence of abundant A. ferrooxidans cells in samplesfrom the actively weathering sites in the Metcalf pit (e.g.,MET-5200 SP01; Fig. 6). These bacteria are considered tohave been healthy, because they are not mineralized and are

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TABLE 1. Background Bacterial Populations and Characteristics of Natural Waters (creek, seeps, springs), Process Solutions, and Mineral Samples

Sample no.1 Sample description pH A. ferrooxidans (viable bacteria/g or /ml) SRB (viable bacteria/g or /ml)

Creeks, seeps, wellsMD-SW03 Upgradient 7.7 n.d n.a.MD-GW02 Upgradient 7.1 n.d. n.a.MD-SW04 Chase creek 7.0 2.6 × 100 n.a.MD-SW05 Tailings dam 7.3 1.0 × 101 n.a.

Process solutionsSX/EW-LS01 Copper rich 1.8 1.1 × 103 n.a.SX/EW-LS02 Copper depleted 1.4 1.3 × 102 n.a.

Mineral samplesTD-S06 Dry sand 2.7 2.3 × 102 n.a.TD-S07 Moist fines 2.6 1.3 × 103 n.a.TD-SW03 Pond water 2.5 1.7 × 106 n.a.TD-S08 Pond sediment 2.8 5.4 × 106 n.a.

MOR-3600-SW01 Spring water 3.1 4.9 × 102 n.a.MOR-3600-S01 Brown mud 2.5 4.9 × 105 n.a.MOR-3600-S02 Gray mud 4.5 n.a. 1.3 × 10–1

MOR-3600-S03 Sediment 1.9 n.a. n.d.

NWX-4350-SW02 Standing water 5.4 n.a. n.d.

MET-5200-EB05 Seep/rock scraping 2.1 2.2 × 100 n.a.MET-5200-EB06 Seep/rock scraping 2.8 8.2 × 103 n.a.MET-5000-WP01 Seep/rock scraping 4.3 1.7 × 105 n.a.MET-4900-SP02 Spring/Fe(OH)3(s) 2.8 7.0 × 104 n.a.

Notes: n.a. = not analyzed, n.d. = not detected, SRB = sulfate-reducing bacteria, upgradient = upstream or up ground-water gradient; EB = enriched blan-ket, GW = ground water, MD = Morenci district, MET = Metcalf pit, MOR = Morenci pit, MPN = most probable number, NWX = North West Extractionpit, SX/EW = solvent extraction/electrowinning, S = sediment, SW = surface water, TD = Tailings dam; WP = weathered products,

1 Sample number corresponds to mine elevation in ft

TABLE 2. A Comparison of Average pH and Populations of A. ferrooxidans and Sulfate-Reducing Bacteria in Samples Collected from Morenci

Sample 1 Sample description pH A. ferrooxidans (viable bacteria/ml) SRB (viable bacteria/ml)

MET-5200-EB02 Weathered products 5.0 6.5 × 104 n.dMET-5200-EB03 Weathered products 2.3 2.3 × 102 3.5 × 102

MET-5200-EB05 Weathered products 4.0 9.3 × 104 5.4 × 102


MET-5200-EB06 Water 2.4 3.5 × 107 2.8 × 102

MET-4900-SP04 Water 2.4 2.4 × 107 2.6 × 103

MET-4900-WP02 Weathered products 2.5 n.a. 1.8 × 102

MET-4750-SP03 Water 2.4 3.3 × 106 2.9 × 101

MET-4750-S04 Weathered products 2.6 n.a. 2.0 × 102



NWX-4200-EB09 Weathered products 4.3 1.1 × 106 2.6 × 102

Notes: n.a. = not analyzed, n.d. = not detected, SRB = sulfate-reducing bacteria; EB = enriched blanket, MET = Metcalf pit, NWX = Northwest Exten-sion pit, S = sediment, SP = spring water


typical of acidophilic iron-oxidizing bacteria in acid minedrainage systems (Southam and Beveridge, 1992). SEM-EDSanalysis of the fibrous minerals in the TEM image indicatedthese are jarosite [KFe3(SO4)2(OH)6] (data not shown).

Sulfate-reducing bacteria populations: Sulfate-reducingbacteria were not found at the ground-water interface locatedat the bottom of the Morenci or Northwest Extension pits(Table 1). However, they were found throughout the enrichedblanket and in the partially leached zone and in close proxim-ity to (i.e., in the same samples) the A. ferrooxidans popula-tions, averaging ~500 bacteria/g (Table 2). Only one of thesesamples did not contain detectable sulfate-reducing bacteria.Interestingly, the site (MET-4900-SP04) with the highestnumber of A. ferrooxidans also had the highest sulfate-reduc-ing bacteria population.

Relationship to site conditions and geochemistry

Water chemistry: The geochemistry of the five seeps andsprings in the mine area associated with the sample sites istypical of acid mine drainage systems (Table 3). Average pHvalues were acidic and ranged from 2.6 to 4.6, they contain

high concentrations of copper and sulfate, have no detectablealkalinity, have elevated trace metal contents, and are all as-sociated with significant populations of A. ferrooxidans (Ta-bles 1, 2). For example, the seep at MET-5200-EB06 had thehighest copper content (960 mg/l), the highest sulfate content(4,300 mg/l), and consistently high bacterial populations (Fig.5). Overall, copper concentrations were higher than iron, butlocally, the metal ratios were highly variable. These waterswere significantly different than the typical ground water inmonitoring wells around the district.

Rock geochemistry: Laboratory tests on sulfur speciationand acid-base accounting showed that the wall rocks in thedeposit have high acid-generating potential and no acid neu-tralization capacity (Table 4). Except for the leached capping,where sulfides have been previously weathered or trapped


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FIG. 4. Seasonal variability of A. ferrooxidans along the Metcalf 5200bench. The bars below the reference line indicate less than detection limit(<0.13 bacteria/ml) for those sites during the respective sampling event.Sample locations correspond to those in Figure 2 and with the results shownin Table 1.

FIG. 5. Seasonal variability of sediment sample pH along the Metcalf 5200bench. Sample locations correspond to those in Figure 2.

FIG. 6. A representative unstained, ultra-thin section TEM micrograph ofa sample of mineralized biofilm from the seep at sample site MET-5200-EB05. Although the thiobacilli are in an intensively mineralizing environ-ment, they are not coated with iron hydroxides. These unmineralized bacte-ria are presumably alive (supported by the viable bacterial counts; see Fig. 4)and contribute to the development of the weathering profile via the contin-ual regeneration of ferric iron catalyst. The secondary jarosite(KFe3(SO4)2(OH)6) minerals coat the clay-sized silicates produced by acidweathering of the host rock. Bar equals 0.5 µm.

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TABLE 3. Geochemistry of Seep and Spring Waters in the Morenci District

MET-5200-EB06 MET-4700-EB08 Analysis (mg/l) (seep)1 MET-4900-SP041 (seep)1 MOR-3600-SW011 NWX-4350-SW021

Specific Conductance (µmhos/cm) 4200 3100 6000 1400 1600pH 2.6 3.2 2.8 4.6 4.1Total dissolved solids 6300 4000 3520 1300 1100Chloride <25 <50 <50 <5 11Fluoride <0.1 1.1 9.6 1.9 4.2Nitrogen, from ammonia 1.1 5.8 n.a 0.11 29Nitrogen from nitrate-nitrite <2.5 <5 n.a. <0.5 67Phosphorous 0.17 <0.050 n.a. <0.050 <0.050Sulfate 4300 2700 2150 840 550Alkalinity, bicarbonate (as CaCO3) <20 <20 <2 <20 <20Alkalinity, carbonate (as CaCO3) <20 <20 <0.5 <20 <20Alkalinity, hydroxide (as CaCO3) <20 <20 <0.5 <20 <20Alkalinity, total (as CaCO3) <20 <20 <2 <20 <20Metals

Antimony 0.0096 0.018 0.006 0.007 <0.005Arsenic 0.011 0.0098 <0.005 <0.005 <0.005Beryllium 0.013 0.026 0.054 0.012 0.018Cadmium 0.028 2.6 0.49 <0.0005 0.12Chromium 0.023 <0.005 <0.05 <0.005 <0.005Lead <0.005 0.011 0.011 <0.0005 0.17Selenium <0.005 <0.005 <0.005 <0.0005 <0.005Aluminum 370 140 74.9 0.48 7.8Barium 0.04 <.01 <0.10 0.02 0.02Calcium 21 150 270 150 55Copper 960 87 92.62 0.46 100Iron 260 420 91.44 66 15Magnesium 11 55 69 40 23Manganese 0.35 45 43.4 9.1 10Nickel 0.18 0.15 0.21 <0.05 0.06Potassium 10 20 21 21 66Sodium 30 39 33 70 41Zinc 4.7 62 159 0.8 18Mercury <0.0002 <0 .0002 0.0038 <0.0002 <0.0002Silver <0.005 <0.005 <0.40 <0.0005 <0.005Thallium <0.002 0.0059 <0.001 <0.002 0.0037

Notes: n.a. = not analyzed; EB = enriched blanket, MET = Metcalf pit, MOR = Morenci pit, NWX = North West Extraction pit, PL = partially leached,SP = spring water, SW = surface water


TABLE 4. Acid-Base Accounting Data for Selected Morenci Samples

Acid generating Acid neutralizing Nonextractable potential potential Acid-base

Sample no. Total S (%) HCl soluble S (%) S (%) Sulfide S (%) (CaCO3/ 1,000 t) (CaCO3/ 1,000 t) account

(CaCO3/ 1,000 t)MET-5200-LC01 0.24 0.22 0.02 <0.01 <0.3 1.07 1.07MET-5200-LC02 0.06 0.04 0.02 0.00 <0.3 <0.5 0.00MET-5200-LC03 11.70 1.40 0.67 9.63 301.00 2.01 –299.00MET-5200-PL01 0.18 0.01 0.01 0.17 5.31 <0.5 –5.31MET-5200-EB01 9.66 0.59 0.76 8.31 260.00 2.76 –257.00MET-5200-EB02 1.82 1.41 0.01 0.40 12.50 <0.5 –12.50MET-5200-EB03 0.24 0.10 0.02 0.12 3.75 <0.5 –3.75MET-5200-EB04 0.95 0.01 0.01 0.94 29.40 <0.5 –29.40MET-5200-EB05 2.34 0.27 0.48 1.59 49.70 <0.5 –49.70MET-5200-EB06 0.94 0.46 0.03 0.45 14.1 <0.5 –14.1MET-5200-PL02 0.15 0.12 0.03 0.00 <0.3 <0.5 0.00MET-5000-WP01 0.89 0.38 0.05 0.46 14.40 <0.5 –14.40MET-4900-SP02 1.43 0.19 0.08 1.16 36.30 <0.5 –36.30

Notes: EB = enriched blanket, LC = leached capping, MET = Metcalf pit, PL = partially leached, SP = spring water, SW = surface water, WP = weath-ered products

above the water table, most of the sulfur in the enriched blan-ket of the Metcalf pit (except MET-5200-EB04) is present aspyrite and chalcocite and available for biooxidation. Coppercontents of the samples varied widely from <0.01 wt percentCu in the leached capping to over 10 wt percent Cu in veinsand averaged about 0.52 percent Cu overall in the enrichedblanket. With the exception of site MET-5200-EB4, all sam-ples in the enriched blanket contained pyrite and chalcocitein veins and along fracture surfaces. All of the samples arestrongly altered granite or monzonite porphyry. These rockscontained pervasive quartz and sericite replacing the originalfeldspars and ranging from 5 to 40 percent of the matrix ofthe rock and 100 percent of the vein and fracture selvagesand, therefore, had no buffering capacity for acidic solutions.

Discussion

Role of bacteria in leaching

Active weathering of copper in porphyry copper deposits,consisting of low-grade ore (i.e., 0.05–0.35% Cu as chalcopy-rite and pyrite), occurs under acidic pH conditions (Tables3–5). Because biological oxidation of pyrite (3 × 10–7 mol L–1

s–1 by reactions (1) and (2)) can be up to five orders of mag-nitude greater than the abiotic rate (3 × 10–12 mol L–1 s–1 byreaction (3); Singer and Stumm, 1970), thiobacilli have beenimplicated in this process (Titley, 1975).

Fe2+ + 1⁄4O2 + H+ thiobacilli → Fe3+ + 1⁄2H2O, (1)14Fe3+ + FeS2 + 8H2O → 15Fe2+ + 2SO4

2– + 16H+, (2)

and

FeS2 + 7/2O2 + H2O → Fe2+ + 2SO42– + 2H+. (3)

In Morenci, acidophilic iron-oxidizing bacteria (Tables 1, 2,Fig. 5) enhance metal sulfide dissolution by catalyzing theabiotic oxidation of metal sulfides (Table 5; reaction (2);Singer and Stumm, 1970). The regeneration of Fe2+ for thethiobacilli establishes the propagation cycle between the iron-oxidizing bacteria, dissolution of sulfide minerals, and the for-mation of metal-rich sulfuric acid leachate (Fortin et al.,1995). Not only is the Fe3+ available to leach pyrite, it is alsoavailable to leach chalcopyrite, chalcocite, and covellite asdemonstrated in the ferric-soluble leach tests of the wall-rocksamples (Table 5). The excess H2SO4 in reaction (2) can pro-mote chalcocite and covellite dissolution, further enhancingCu mobility as demonstrated in the acid-soluble leach tests ofthe wall-rock samples (Table 5). The recovery of viablethiobacilli from every hydraulically active fracture demon-strates that these weathering reactions are active throughoutthe year at Morenci.

Although the optimal condition for growth ofAcidithiobacillus spp. is at pH <3 (Colmer et al, 1950; Amaroet al., 1991; Hallmann et al., 1993), the colonization of sulfideminerals and resulting chemolithotrophy is possible undercircumneutral pH conditions (Southam and Beveridge, 1992;Mielke et al., 2003). At Morenci, seasonal (i.e., wet) cyclespromoted the rapid growth of thiobacilli across a wide zone ofactive weathering in populations exceeding 106 bacteria/g atpH values that were extremely acidic (pH <3 up to pH 7.2; cf.Figs. 4, 5, Table 3). The decrease in the populations duringthe dry season is presumably due to low water activity, whichis deleterious to A. ferrooxidans (Southam and Beveridge,1992) because it is unable to produce a cyst or endosporeresting stage (Kelly and Harrison, 1984).


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TABLE 5. Copper Leaching Potential for Selected Wall-Rock Samples

Sample no.1 Total Cu (%) Acid-soluble Cu % Acid-soluble Ferric-soluble Cu % Ferric- soluble

MET-5200-LC01 0.01 0.01 100 0.01 100MET-5200-LC02 0.01 0.01 100 0.01 100MET-5200-LC03 10.92 1.98 18 4.34 40MET-5200-PL01 0.09 0.03 33 0.05 56MET-5200-PL02 0.05 0.04 80 0.04 80MET-5200-EB01 10.59 1.50 14 4.07 38MET-5200-EB02 0.46 0.07 15 0.24 52MET-5200-EB03 0.10 0.04 40 0.05 50MET-5200-EB042 0.32 0.20 63 0.22 69MET-5200-EB05 3.08 0.28 9 1.20 39MET-5200-EB06 0.34 0.01 3 0.01 3

MET-5000-WP01 0.19 0.02 11 0.05 26

MET-4900-WP02 0.21 0.03 14 0.13 62MET-4900-SP023,4 0.57 0.07 12 0.17 30

MET-4700-EB08 0.36 0.01 3 0.01 3

MOR-3600-S043 0.25 0.02 8 0.05 20

NWX-4200-EB09 0.36 0.02 6 0.03 8

Notes: EB = enriched blanket, GW = ground water, MET = Metcalf pit, MOR = Morenci pit, NWX = North West Extraction pit, PL = partially leached,S = solid, SP = spring water, SW = surface water, WP = weathered products

1 Sample number corresponds to mine elevation in ft2 Disseminated sulfide mineralization, i.e., within rock matrix3 Enriched blanket4 Solid collected from spring outflow

Acid attack of the sericite-bearing wall rocks during theseprocesses results in the formation of jarosite, alunite, andkaolinite in the actively weathering zone (Bladh, 1982; Titleyand Marozas, 1995). The kaolinite tends to retain moisture,which promotes bacterial growth. Thus the moist mixture ofchalcanthite, jarosite, ferrihydrite, and kaolinite that occursalong the actively weathering fractures in the Metcalf 5200bench can be thought of as a biogeochemical wedge of leach-ing. During dry periods, the evaporative concentration of thebiogenic sulfuric acid and ferric sulfate promotes mineral dis-solution and the formation of soluble salts such as chalcan-thite. These salts, then, constitute the mobile fraction of min-erals that are ready to be leached and transported during thenext hydrologic pulse. Once the sulfides have been oxidized,all of the copper and some of the kaolinite appear to beflushed from the fractures leaving a quartz + goethite ±hematite secondary mineral complex.

The bacterial populations highlighted in Figure 5 corre-spond to two hydrologic (i.e., growth) cycles that occur an-nually at Morenci. Using the data from MET-5200-EB6, thebiomass produced during these two growth cycles (i.e., aproxy for annual bacterial growth), equaled 3.7 × 107 cells/gof fracture material. Using the mass of Escherichia coli (3 ×10–13 g dry wt: Neidhardt et al., 1990), which is similar in sizeto Acidithiobacillus spp. (Kelly and Harrison, 1984), thisseep produced 1.1 × 10–5 g of bacteria (dry wt) or 4.5 × 10–6

g organic carbon (40 wt % C as CH2O) per gram of fracturematerial. When applied to the 3.8 vol percent fractures re-ported by Enders (2000), which equals 3.8 × 1010 cm3 offracture material in a 100-m3 block, an active 100-m3 super-gene block could produce 4.3 × 10-1 t of organic carbon/yr(similar to Melchiorre and Enders, 2003). Since the utiliza-tion of ferrous iron is an energetically poor reaction, bacteriamust oxidize 18.5 mol of iron for each mol of carbon fixed(Silverman and Lundgren, 1959). However, this value as-sumes 100 percent metabolic efficiency, and bacteria (all en-ergy systems) are not 100 percent metabolically efficient.For A. ferrooxidans, metabolic efficiency ranges between 3.2percent (Temple and Colmer, 1951) and 20.5 percent (Sil-verman and Lundgren, 1959). Also, 14 moles of ferric iron isrequired to oxidize 1 mol of pyrite (reaction 2). When theseenergy-reaction requirements are applied to the 100-m3

block of hypogene ore, containing 1.3 × 109 mol pyrite (3%)and 1.0 × 108 mol chalcopyrite (0.46%), it would only take 9× 102 to 5 × 103 yr to oxidize all of the metal sulfides pre-sent in this block, assuming a process that is entirely meta-bolically driven. While it is interesting to think that thisleaching could occur on a time scale of several thousandyears, this is clearly too fast since the leached capping atMorenci took several million years to form (Enders, 2000). Itis unlikely that an entire 100-m3 block could be active at onetime when the actual process occurs in discrete fractures andis cyclical, reflecting episodic (overall) downward movementin the position of the redox boundary as a result of tectonic,physiographic, and climatic changes. Using average erosionrates (0.12 mm/yr) during the main stage of leaching and en-richment at Morenci (18–2 m.y.: Enders, 2000), it would takeabout 0.8 m.y. to erode that same 100-m3 block, thus allow-ing for leaching over a much longer time than that estimatedfor microbial oxidation.

Using MET-5200-EB6 as a model fracture (1 cm wide, ex-tending up and down 3 benches, ~ 50 m in height), 570 g oforganic carbon would be produced by bacterial growth in thisfracture each year, and ~300 to 2,000 mol of pyrite(4,300–27,000 mol of Fe2+) would be oxidized annually, pro-ducing 4,300 to 27,000 mol of Fe3+ by reaction (1). Since 4mol of Fe3+ is required to oxidize each mol of chalcocite, mi-crobially driven reactions in this fracture could leach between0.14 and 0.87 t of Cu/yr. It is important to note that this cal-culation represents the potential activity of a single fractureand that viable thiobacilli were found across this active weath-ering horizon; therefore, the impact of biooxidation would begreater than that described. Where there is sufficient pyriteremaining, further dissolution of pyrite and chalcocite in theenriched blanket left behind a mixture of transportedhematite and goethite or a classic and distinctive hematiteboxwork (Anderson, 1982; Titley and Marozas, 1995).

The leached capping is typically devoid of copper and con-tains a mixture of the iron oxide minerals hematite, goethite,and jarosite, which comprise the classic limonite assemblage(Blanchard, 1968; Anderson, 1982). In the regions where allof the reduced iron and sulfur minerals have been oxidizedthere is no available energy for thiobacilli. Under these con-ditions, no thiobacilli are detected (Fig. 5). Where thisprocess is incomplete, a zone of partial leaching is left behindbeneath the leached capping (Enders, 2000). In Morenci, ac-tive thiobacilli were recovered as long as there were metalsulfides and seasonal inputs of ground water. However, sur-face water (i.e., rain or snow melt) did not provide enoughwater to support the growth of thiobacilli on any exposed sul-fides along the benches. Rather, the importance of groundwater in biooxidation is demonstrated by the presence of sul-fide zones that are stranded above the water table, as in theMorenci district, or left to desiccate in dry climatic conditionssuch as in northern Chile since the mid-Miocene (Alpers andBrimhall, 1988). In these environments A. ferrooxidans dieand the abiotic geochemical reactions will only proceedslowly.

Role of bacteria in enrichment

The presence of viable sulfate-reducing bacteria in the ac-tively weathering environment at Morenci suggests that sul-fate-reducing bacteria activity contributes to the enrichmentand/or precipitation of secondary copper sulfide mineralsthrough sulfate reduction. Although this mechanism has beenspeculated to occur in some supergene Cu environments(Alpers and Brimhall, 1989; Lichtner and Biino, 1992) and isconsidered to be responsible for the supergene enrichment ofZnS at the Mike gold deposit (Bawden et al., 2003), the dis-covery of viable sulfate-reducing bacteria at Morenci is evi-dence that sulfate-reducing bacteria contribute to supergeneenrichment in this contemporary system and suggest thatthey may have contributed to the historical enrichment of Cu.

The basic biochemical and geochemical reactions mediatedby dissimilatory sulfate-reducing bacteria (Tuttle et al., 1969;Trudinger et al., 1985) in Morenci are:

2CH2O + SO42– → H2S + 2HCO–

3 (4)and

Cu2+ + H2S → CuS(s) + 2H+. (5)

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CuS-So concentrates containing 33 percent Cu have beenproduced from acid mine drainage from the abandoned RioTinto copper mine in Nevada in bench-scale bioreactor ex-periments (Hammack et al., 1994). However, this level of sul-fate-reducing bacteria activity requires abundant organic car-bon and sulfate, which is not limited in these environments.In the Morenci supergene system, the source of organic car-bon presumably results from the fixation of CO2 within theoverlying weathering profile by thiobacilli, similar to that ob-served in acid mine drainage systems (Fortin et al., 1995,Fortin and Beveridge, 1997). During the wet season thethiobacilli flourish, in part due the meteoric input of dissolvedoxygen, fixing carbon into the supergene system. During thedry season, a lot of the thiobacilli die, releasing organic car-bon. Subsequent wet seasons provide more favorable redoxconditions for bacterial sulfate reduction and promote the be-ginning of another cycle of carbon fixation. The highest sul-fate-reducing bacteria population was found associated withthe highest population of thiobacilli (Table 2), where biogeniccopper sulfides (i.e., covellite) are produced across the broadweathering and enrichment profiles at Morenci. However, in-organic geochemical processes dominate supergene enrich-ment in this system. Using the annual carbon fixation occur-ring in MET-5200-EB6 fracture and reactions (4) and (5),bacterial sulfate reduction could only fix 0.2 to 1.1 percent ofthe copper leached from MET-5200-EB6. Therefore, inor-ganic enrichment processes (Enders, 2000) are primarily re-sponsible for forming the enriched blanket at Morenci.

ConclusionsGeological and microbiological examination of water and

mineral samples collected primarily along the 5200 bench ofthe Metcalf pit within the Morenci copper deposit revealedthat acidophilic iron-oxidizing bacteria (thiobacilli) are re-sponsible for significant leaching, and sulfate-reducing bac-teria can be minimally responsible for the enrichment ofcopper in the supergene environment. Theoretically, biooxi-dation of sulfides in a 100-m3 block of ore at Morenci couldhave been completed in a few thousand years. In contrast,abiotic oxidation may be too slow (by a factor of 105), suchthat a supergene zone would be completely eroded beforethorough leaching and enrichment could take place. There-fore, biooxidation may be essential for the formation of su-pergene Cu deposits.

AcknowledgmentsThe authors would like to thank Phelps Dodge Morenci

Inc. for their generous and enthusiastic support of this pro-ject. This paper is an abridgement of part of the first author’sPh.D. dissertation for the Department of Geosciences, Uni-versity of Arizona. We thank the anonymous reviewers whohelped focus our manuscript.April 18, 2001; January 3, 2006

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