Long-Term Biogeochemical Stabilization of Carbonate-Bearing Pb–Zn–Cu Mine Tailings by Amendment-Assisted Phytostabilization

Legacy mine tailings remain persistent sources of metal exposure because they are not inert waste deposits but reactive geochemical systems. Their low organic matter, poor structure, limited nutrient availability, and elevated metal concentrations restrict plant and microbial establishment while leaving tailings surfaces vulnerable to wind dispersion, runoff, and pore-water transport. (1, 2) In Pb–Zn–Cu tailings, risk is governed not only by total metal concentrations but also by the distribution of metals among pore water, readily labile pools, reactive carbonate- and Fe/Mn oxide-associated phases, organic/sulfide-associated pools, and more stable residual fractions. (3−5) This distinction is particularly important for carbonate-bearing tailings, where circumneutral to alkaline pH can suppress immediate metal solubility, but longer-term mobility may still be influenced by carbonate weathering, dissolved organic carbon, ionic strength, and redox-sensitive mineral phases. (6−8)

Amendment-assisted phytostabilization offers a practical alternative to excavation or engineered capping for large mine-waste areas, but its success is often evaluated too narrowly. Recent restoration approaches increasingly emphasize nature-based strategies that combine contaminant stabilization with development of ecological functions, including carbon cycling and microbial development. (9, 10) Field-based phytomanagement studies have demonstrated that combined amendment and native plant strategies can improve restoration outcomes in metal-contaminated mine residuals. (11, 12) Vegetation can reduce surface erosion, improve surface stability, and initiate organic carbon deposition; however, plant survival or biomass alone does not demonstrate geochemical stabilization or development of broader biological and biogeochemical functions. (13, 14) A more rigorous assessment requires evidence that amendment–plant systems reduce metal lability, promote retention in less mobile solid-phase pools, and support the development of biological and biogeochemical functions. Biosolids (BS) can improve tailings fertility, organic matter content, microbial activity, and metal retention through organic matter inputs and sorption/complexation processes. (15) However, their long-term effectiveness may be limited by decomposition of labile organic matter, redistribution of nutrients, and potential contributions of dissolved organic ligands, salts, and metal inputs that influence pore-water chemistry and metal mobility. (16, 17) Biochar (BC) and arbuscular mycorrhizal fungi (AMF) may complement biosolids by increasing sorptive capacity, improving nutrient retention, promoting aggregation, and supporting root- and mycorrhizosphere-mediated carbon inputs and microbial development. (18, 19) Recent studies have shown that integrated amendment mixtures containing biochar, organic amendments, and microbial inocula can improve plant establishment, modify tailings chemistry, and influence microbial communities in mine rehabilitation settings. (18, 20)

Despite these potential benefits, the time-resolved and depth-dependent coupling between biological development and geochemical stabilization in amended tailings remains insufficiently explored. Previous studies have examined improvements in vegetation establishment, metal bioavailability, microbial indicators, or microbial community structure during tailings and mine-soil rehabilitation. (21−25) Long-term rehabilitation studies have further demonstrated progressive changes in tailings chemistry, organic matter characteristics, and microbial communities following revegetation. (26) However, the persistence and extension of coupled biological and geochemical responses into initially unamended tailings layers remain poorly resolved, particularly when considering long-term changes in pore-water chemistry alongside solid-phase metal partitioning, organic carbon content, and microbial characteristics. This limitation is important because sustained phytostabilization requires not only reduced metal mobility but also development of functional tailings properties beyond the amended surface layer. Biological amendments such as AMF may contribute to belowground development through root-associated processes and microbial activity, potentially facilitating carbon accumulation and microbial development beyond the amended surface layer. The contribution of these processes to sustained geochemical stabilization in initially unamended deeper tailings, however, remains uncertain.

Here, we evaluated the long-term biogeochemical stabilization of carbonate-bearing Pb–Zn–Cu mine tailings under amendment-assisted revegetation with purple prairie clover (Dalea purpurea), a native prairie legume species. Biosolids, biochar, and arbuscular mycorrhizal fungi were applied only to the upper tailings layer, while the underlying tailings remained unamended, providing a depth-resolved framework for evaluating treatment-associated responses beyond the zone of direct amendment incorporation. Over seven years, tailings pore-water was monitored annually for pH, electrical conductivity, dissolved organic carbon, dissolved nitrogen, and dissolved Pb, Zn, and Cu. At the end of the seventh growth season, tailings cores were analyzed for organic carbon, phospholipid fatty acid (PLFA)-based microbial indicators, aqua-regia-extractable metals, sequentially extracted Pb, Zn, Cu, Fe, and Mn fractions, and mineralogy by X-ray diffraction (XRD). We hypothesized that long-term amendment-assisted revegetation would: (i) alter pore-water chemistry and dissolved metal concentrations; (ii) promote organic carbon accumulation and microbial development within and below the amended layer; and (iii) shift Pb, Zn, and Cu among operationally defined fractions in a metal- and depth-dependent manner. Together, this framework evaluates phytostabilization as a coupled aqueous, solid-phase, and biological process rather than vegetation establishment alone.

For annual tailings pore-water (TPW) collection, one Rhizon MOM soil moisture sampler (10 cm porous membrane length, 2.5 mm diameter, 0.15 μm pore size; Royal Eijkelkamp, The Netherlands) was installed in each pot. Samplers were inserted diagonally into the tailings profile, with the porous membrane section extending into the lower unamended tailings layer, as shown in Figure 1. Prior to each extraction, pots were watered to approximately 70% of field capacity and allowed to equilibrate for 24 h under greenhouse conditions. TPW was collected annually at the end of each growing season over the seven-year growth period by applying suction to the sampler tubing. A 50 mL TPW sample was collected from each pot.

Immediately after collection, pH and electrical conductivity (EC) were measured on fresh samples using calibrated pH and EC electrodes (Orion 370, Thermo Fisher Scientific, Inc.; VWR 61161-362, respectively). Dissolved organic carbon (DOC) and dissolved nitrogen (DN) were measured on fresh, nonacidified aliquots using a TOC analyzer (Shimadzu TOC-5000A). A 15 mL aliquot of each TPW sample was acidified to 2% with trace-metal-grade nitric acid for elemental analysis. Dissolved Pb, Zn, and Cu concentrations were determined by inductively coupled plasma optical emission spectroscopy (ICP-OES; PerkinElmer Avio 200).

TPW pH remained narrowly circumneutral to slightly alkaline, generally around 7.4–7.6. MT and BS+BC had the highest overall pH, whereas BS, BS+AMF, and BS+BC+AMF had lower values (Table S4). This modest pH depression is consistent with the lower pH of the biosolids amendment and may also reflect organic acid production during decomposition and rhizosphere development. (16, 54) However, persistent circumneutral pH is consistent with strong carbonate buffering in the dolomite-rich tailings matrix. (55)

EC, DOC, and DN showed a clear amendment-associated pulse. During the first three years, amended-treatment means were 4.89–6.24 mS cm^–1 for EC, 41.4–51.5 mg L^–1 for DOC, and 39.8–71.2 mg L^–1 for DN, compared with MT means of 2.33–2.54 mS cm^–1, 2.39–2.77 mg L^–1, and 1.49–1.93 mg L^–1, respectively. By year seven, amended-treatment means declined to 3.20–3.88 mS cm^–1 for EC, 20.37–30.46 mg L^–1 for DOC, and 4.93–17.27 mg L^–1 for DN. These trends are consistent with an initially salt-, nutrient-, and DOC-rich TPW environment following amendment application, followed by a decline in readily soluble constituents over time that may reflect processes such as depletion, retention, redistribution, and biological assimilation.

The early elevation of soluble constituents is consistent with the dual role of biosolids as a fertility amendment and a source of labile nutrients, salts, DOC, and metals. (16, 17) However, the later TPW patterns suggest that DOC dynamics were not controlled solely by depletion of biosolids-derived soluble organic matter. Although DOC declined substantially from early year values, DOC remained relatively higher in some combined-amendment treatments than in BS alone by the end of the experiment, consistent with continued contributions from vegetation and microbial processing. Organic carbon amendments and vegetation establishment may influence dissolved organic matter composition, microbial associations, mineral–organic interactions, and aggregation during biogeochemical development in the residuals. (56−58) Thus, the subsequent decline in EC, DOC, and DN may reflect a transition from an initially amendment-dominated soluble phase toward a TPW environment increasingly influenced by biological and mineralogical processes, rather than simple loss of amendment-derived constituents. (59)

The seven-year results show that amendment-assisted revegetation was associated with a transition from early amendment-associated mobilization toward greater biological development and metal attenuation. Although biosolids initially increased EC, DOC, DN, and dissolved metals, these responses attenuated over time, particularly in treatments containing BC and/or AMF. Concurrent increases in OC, PLFA-based microbial indicators, and fungal:bacterial ratios in the middle and lower unamended layers, together with metal-specific repartitioning of Pb, Zn, and Cu, indicate that long-term stabilization was associated with coupled aqueous, solid-phase, and biological changes extending beyond the directly amended surface layer (Figures 2–4 and S2). Successful phytostabilization should therefore be evaluated in terms of both contaminant stabilization and substrate development, because surface vegetation alone may overstate restoration progress if deeper tailings remain carbon-poor, biologically inactive, and geochemically labile. Deeper root penetration, AMF-associated PLFA, microbial proliferation, and changes in pore-water chemistry were consistent with biological and geochemical influence extending into underlying unamended tailings, including carbon accumulation, microbial habitat development, and potential C and nutrient cycling. However, the relative contributions of roots, AMF, microbial processes, and pore-water-mediated transport cannot be resolved from the present data.

From a management perspective, the results support complementary amendment strategies that combine fertility, sorption, biological development, and metal retention. Biosolids supported revegetation, but BS alone maintained higher dissolved Zn and Cu by year seven, whereas treatments containing BC and/or AMF showed stronger long-term metal attenuation and biological development. Restoration performance should therefore consider pore-water chemistry, solid-phase lability, OC accumulation, and microbial indicators in addition to vegetation establishment and total metal concentrations. Because the biosolids also contained elevated Pb, Zn, and Cu, their metal loading should be considered as a potential secondary contamination source under environmental conditions that promote remobilization, such as changes in pH or dissolved organic matter. Field-scale economic feasibility will further depend on the local and sustained availability of amendment materials, transport distances, application rates, and site-preparation requirements, and the cost, availability, and scalability of AMF inoculation; these factors require site-specific evaluation before large-scale implementation.

Field-scale restoration should also consider diverse native plant communities rather than relying on the single species evaluated here. Multispecies communities may provide complementary rooting patterns, carbon inputs, and nutrient-acquisition strategies that influence microbial development, ecosystem development, and succession, consistent with field restoration studies showing benefits of AMF-associated plant–microbe interactions for native plant establishment and community development. (74, 75) N-fixing legumes, such as the purple prairie clover used herein, can contribute to nitrogen availability in nutrient-limited tailings, while AMF-mediated belowground interactions may influence nutrient acquisition and interactions among co-occurring plant species. (76, 77) However, the greenhouse pot design does not reproduce field-scale hydrologic variability, preferential flow, seasonal climatic conditions, or spatial root and plant-community development, and the study did not directly quantify leaching fluxes or field-scale metal transport. Accordingly, these responses represent stabilization under the experimental conditions rather than direct evidence of reduced metal export at field scale. Field studies are needed to determine whether they persist under variable hydrologic and climatic conditions and translate into reduced metal transport and sustained ecosystem development.