Contribution of the Soil Macro- and Microstructure to Organic Matter Stabilisation in Natural and Post-Mining/Industrial Soils under Temperate Climatic Conditions
1. Introduction
Soil ecosystems contain more carbon (C) than the terrestrial biomass and atmospheric resources combined. Soil C is bound in the soil organic matter (SOM), thereby reducing the emission of carbon dioxide (CO2) into the atmosphere [1,2]. The SOM also supplies the materials and energy required to support plant metabolism, improve plant growth, and retain soil water and is a main source of nutrients [3]. For this reason, SOM is essential to the soil environment, sustaining ecosystem health and functions and, ultimately, human life [4]. Even an insignificant decrease in SOM could have considerable negative consequences for the entire environment [2].
The environmental services that soil provides depend to a great extent on its SOM content and quality, in addition to the SOM-related soil features, such as structure and porosity [4]. The particles forming the SOM are bound within the soil structure, specifically in microaggregates (∅ = 50–250 µm) and macroaggregates (>250 µm) [5]. The small pores resulting from the arrangement of the primary soil particles are known as textural (intra-aggregate) pores, while the larger (inter-aggregate) pores, known as structural pores, result from biological activity, climatic conditions and management practices [6]. The soil structure controls several processes, such as the retention of water, gas exchange, nutrient dynamics and root penetration. It is also a habitat populated by soil organisms, controlling their diversity and activity [6].
To interpret soil processes and the nature and arrangement of soil features, micromorphological techniques based on thin sections made from undisturbed soil samples have often been used. These allow the structure and porosity to be assessed and the soil constituents to be identified, including the SOM, its quality, and its degree of decomposition [4,7,8]. Microscopic observations can be complemented with quantifications of the micromorphological constituents based on scanned colour images processed using image analysis computer software [9,10].
Soil contaminated with trace metals is widespread as a result of industrial activities, such as the mining, extraction, and processing of mineral ores [11,12,13]. These activities lead to the destruction of the whole soil profile, resulting in the substantial loss of SOM. Soil recovery in such areas mainly involves the development of surface horizons and SOM storage [14]. According to Slukovskaya et al. [15], the restoration of ecosystems on brownfield sites takes at least 30–35 years. In the first recovery phase, which takes about 5–7 years, an organic accumulative layer forms, fixed by the root systems of perennial grasses. Where the soil pollution is heavy, this stage can take an extended amount of time due to the need to reduce the toxicity of the soil. However, after several years, the recovered soils of mining/industrial areas could potentially become important C reservoirs.
Studies on the original structure of SOM and its relationship to soil porosity and aggregation at detailed scales have already been undertaken, investigating different aspects and the effects of different climatic conditions. Such studies have illuminated the impact of the addition of organic products on aggregate stability [16] and the soil pore structure effect on the rate of SOM decomposition [1,10]. Forms of organic substances in soils have been studied at the microscopic level in a riparian zone [17], while Virto et al. [18] investigated the effect of soil mineral composition on aggregation, with a focus on SOM, calcium carbonate (CaCO3) and clay contributions to semi-arid Mediterranean soils, emphasising the role of SOM in aggregate stabilisation. The soil quality in restored mining areas was the subject of a study by Bosch-Serra et al. [13], who linked soil biological activity, aggregation, and porosity with chemical properties. A micromorphological investigation allowed the authors to confirm that the development of pore structure in restored mining soils, accompanied by high micro- and mesofaunal activity, led to pedogenesis mainly through the formation of pedogenic structures. The relationships between soil chemical and micromorphological properties and SOM transformation have been investigated in soils derived from gypsum under Mediterranean climatic conditions [7]. The authors focused on problems associated with plant root penetration and water infiltration due to microcrystalline gypsum pore infillings.
Having reviewed the literature, to our knowledge, few studies have investigated SOM at the microscale in terms of its relationship with the soil physicochemical properties of two contrasting soils—natural soils derived from gypsum and old, self-restored soils from mining and industrial areas. Soils derived from gypsum occupy large areas under arid or semi-arid climatic conditions, where the presence of precipitated gypsum in the soil profile often inhibits plant growth and worsens the soil properties and fertility [7,19]. In temperate areas, although such soils occupy small areas and are usually shallow, they are environmentally important because they contain high amounts of SOM [20] and exhibit high biological activity [21]. For these reasons, we chose gypsum soils to compare with self-restored mining/industrial soils treating them as a pattern for mining/industrial soils. The significance of this research leads in emphasising the environmental role of degraded soils, especially those left for years without reclamation, which may help in making appropriate decisions regarding their sustainable management. In the literature, mining/industrial soils are studied mainly due to the risk to human health and environmental quality resulting from their contamination [11,13,16]. Their importance in the context of SOM storage and possible role in climate mitigation is much less considered [3,22].
We hypothesised that both groups of soils (natural gypsum and post-mining/industrial) were important C sinks, storing high amounts of SOM. However, we thought that excessive amounts of trace metals in the post-mining/industrial soils would have negatively affected their structural development, which would thus affect SOM stabilisation compared to the case in natural gypsum soils. In our work, we aimed at: (i) checking to what degree physical properties related to macro- and microstructure affect the soil organic matter properties and stability and (ii) comparing natural soils to those of mining areas to ascertain their carbon sequestration effects. In order to test the hypothesis and accomplish the goals, we compared the SOM decomposition state, structure and porosity in natural soils derived from gypsum and in self-restored soils from former mining and industrial areas. We examined thin sections of the soils in order to determine their microstructures and degree of SOM transformation at the microscopic level and quantified the results using image analysis.
2. Materials and Methods
2.1. Study Areas, Soil Bedrock, and Sampling
The studies were carried out in four areas located in southern Poland: (1) the Nida Basin, which is the main area where gypsum rock occurs in Poland. These deposits formed in the Miocene as a result of salt precipitating from warm seawater. In the Pleistocene, the Nida Basin was covered by the Cracow glaciation, which left behind mainly sand, clay, and loess, which occur as admixtures to the gypsum parent materials in the soils of the region. The gypsum rock in the area usually forms gentle hills [20,23]; (2) the area around Bukowno, where the mining and processing of zinc-lead (Zn-Pb) ores have been performed since the 15th century, leaving heaps of tailings that have never been reclaimed in about 150 to 400 years; (3) close to Jaworzno, an area of recent Zn-Pb mining, the tailings heaps left behind composed of waste rock. The Zn-Pb ore deposits in the Bukowno and Jaworzno areas occur in Jurassic limestone and dolomite, with admixtures of Pleistocene sand; and (4) in the district of Krakow (Nowa Huta), where iron (Fe) processing has been taking place for 50 years, leaving heaps of metallurgical slag on which soils with loess and sand admixtures have formed [24] (Figure 1).
All the waste materials were characterised by properties unfavourable to ecosystem development, such as sensitivity to erosion, poor water retention, nutrient deficiency, and very high—or at least excessive—contents of trace metals. For these reasons, pedogenesis at these sites has been occurring very slowly, and full recovery of the soils is taking a long time. In the Nida Basin area, the soils are derived naturally from gypsum, while in the Bukowno, Jaworzno, and Nowa Huta areas, the soils are self-restored mining/industrial soils. Both groups represent shallow soils in which the accumulation of SOM is the main soil-forming process, and all sites are fully covered with grasses belonging to the Molinio–Arrhenatheretea class (main species listed in Appendix A, Table A1).
The climate of the study area is of continental type with a warm summer and rainfalls throughout the whole year, with an average annual precipitation about 600 mm and a temperature about 6 °C, classified as: Dfb according to the climate classification of Köpper Geiger.
Sampling was performed at four natural soil sites in the Nida Basin area (Gacki (Gac), Busko-Lagiewniki (Bus-L), Busko (Bus), and Chotelek (Chot)) and at four sites hosting post-mining/industrial soils (Jaworzno (Jaw), Bukowno-Warpie (Buk-W), Bukowno (Buk), and Nowa Huta (NH)) (Figure 1). As noted above, the soils in the post-mining and industrial areas differed in age—with the Buk-W and Buk soils formed on very old heaps about 400 and 150 years old, respectively—and were referred to as old brownfield soils, while the Jaw and NH soils were formed on younger heaps—about 50 years old—were referred to as brownfield soils.
At each site, three sampling points were designated, totalling 24 sampling points. Undisturbed and disturbed soil samples were taken out from the same randomly selected sampling points at the site after digging a pit. For the analyses, we collected soil samples from the top layers 0–15 cm deep.
According to the IUSS Working Group WRB (2022) soil classification, the soils derived from gypsum belonged to Gypsiric Chernic Phaeozems (Siltic or Arenic), while the mining and industrial soils were Spolic Technosols (Humic, Siltic, Toxic).
2.2. Field Analyses
The macrostructure (i.e., the size, shape, and resistance of the soil aggregates) was determined for each sample based on the Guidelines for Soil Description [25]. The colour of the soil was established using a Munsell chart [26]. Based on soil colour, the values of the A-horizon development index (ADI) were calculated according to Equation (1)
2.3. Laboratory Analysis of Disturbed Soils Samples
Soil samples were air-dried and sieved through a 2 mm mesh sieve. On the fine soil particles, we determined the texture with the Bouyoucos hydrometer–sieve method, classified in accordance with International Union of Soil Sciences Working Group World Reference Base [29] recommendations. Values of pH were measured using a potentiometer CPI-551 Elmetron (Elmetron, Zabrze, Poland) in a suspension of soil and deionised water at a ratio of 1:2.5 [30]. The total nitrogen (N) content was determined with the use of the Kjeldahl method [30] on a FOSS Kjeltec TM 8100 apparatus (Tecator, Höganäs, Sweden). The CO2 volumetric method was used to determine the CaCO3 content on a Scheibler apparatus (WPL Gliwice, Poland) [30]. The organic carbon content was measured after removal of the carbonates by hydrochloric acid (HCl), and dry combustion using a Vario MACROcube analyser equipped with a CO2 detector (Elementar Analysensysteme GmbH, Langenselbold, Germany), with sulphanilic acid (C6H7NO3S) as the reference material and with a detection limit of 0.001%. The SOM content was calculated by multiplying the organic C content by the Van Bemmelen factor (equal to 1.724) [30]. The exchangeable Ca2+, magnesium (Mg2+), sodium (Na+) and potassium (K+) cations were extracted using 1 mol·dm−3 ammonium acetate (NH4OAc) [30]. The determined cation contents were summed and shown as the total content of basic cations (BCs). The concentrations of cadmium (Cd), Pb, and Zn were determined after digestion of the soil in a mixture (1:3 v/v) of concentrated perchloric and nitric acids (HClO4 and HNO3, respectively) using the wet method in a closed microwave oven (Multiwave 3000, Anton Paar, PerkinElmer Inc., Waltham, MA, USA). The concentrations of the exchangeable cations (Ca2+, Mg2+, Na+ and K+) and the trace metals (Zn, Pb and Cd) were assessed using an atomic emission spectrometer-ICP-OES Optima 7300 DV (PerkinElmer Inc., Waltham, MA, USA) and a multi-element ICP-IV Merck standard solution. The data quality was verified using internal standards and the certified reference material CRM023-050—Trace Metals—Sandy Loam 7 (RT Corporation). The dehydrogenase activity (indicator of overall soil microbial activity) was determined after incubating the samples for 24 h at 37 °C using a 2,3,5-triphenyltetrazolium chloride solution as the substrate. The intensity of the colouration was measured using a Shimadzu UV-1800 spectrophotometer (Kyoto, Japan) at 450 nm [31].
2.4. Laboratory Analysis of Undisturbed Soils Samples
Undisturbed aggregates were collected in order to establish the aggregates’ stability, which was determined via their water resistances using the sieve method. Air-dried aggregates were placed on a 0.25 mm sieve, which was slowly immersed in distilled water, and then mechanically raised for 3 min in the sieving apparatus (Eijkelkamp Agrisearch Equipment, Giesbeek, The Netherlands). The sieving process was repeated using a dispersing solution containing 2 g·dm−3 sodium hydroxide (NaOH) until total breakdown of the aggregates was achieved. Both fractions: the unstable (sieved in distilled water) and stable (sieved in dispersing solution) were dried at 110 °C and then weighed. For the calculation of the water resistance index (WRI) the following formula was used:
where MDS = weight of the soil obtained from the dispersing solution (stable fraction) and MH2O = weight of the soil obtained from the water (unstable fraction).
The dry soil bulk density (BD) was established by weighing it in 100-cm3 cylinders filled with soil with undisturbed structure [32]. The total porosity (TPor) was calculated from the BD, given the particle density (PD) according to the formula:
For the micromorphological study, thin sections were prepared from the undisturbed soil samples collected in Kubiena boxes. The samples were consolidated with Araldite® epoxy resin cut into slices; the slices were glued to a microscope slide and then thinned to 30 µm and polished. To obtain digital images suitable for the micromorphometric analysis of the images, all the thin sections were scanned using a high-resolution Epson scanner. The files were converted to TIFF format and transferred to a computer. Areas of 3.5 × 5.0 cm were processed using the 2400 dpi option. The digital images were assessed using Aphelion software v.4.6.0 (ADCIS S.A. and AAI, Inc., Saint Contest, France). We selected the organic and inorganic parts in these images. The organic parts were then divided into well- and slightly decomposed or undecomposed organic substances. Pores and minerals were isolated from the inorganic parts. For the identification and quantification of selected objects, the Otsu method was used [33], which is based on minimising the inter-class variance. This fully automated method is especially useful for images with a bimodal distribution of pixel values, representing dark and bright objects. Detection of the non-decomposed organic substance and pores was performed using manual thresholding (Figure A1). All the image processing and measurements of the images were performed using Aphelion software.
The slightly decomposed organic matter (OM) comprised recognisable tissue remains, but with evidence of decomposition caused by microfloral or microfaunal activity. The decomposed OM occurred mainly in the form of organic aggregates (also containing small mineral particles). The identifiable aggregates were divided into two classes based on the mean equivalent diameter (diameter of an equivalent circle having the same area)—microaggregates (Agg mic), with a diameter of 50–250 μm, and macroaggregates (Agg mac) >250 μm. The areas of slightly decomposed OM (UndOM) were similarly divided into “mic” and “mac” size ranges. The size and shape of selected porous areas (Por) were determined, with two pore size categories—the medium pores (<150 μm) referred to as Por mic and the coarse pores (>150 μm) as Por mac. The aggregate, UndOM, and pore properties (area and percentage of the image surface) were determined together with their shape factors—circularity and elongation. In the digital images, for circularity, a value of 1 represented a circle, with a trend towards 0 for long, thin objects or those very irregular edges. Elongation was the absolute value of the difference in moments of inertia in the main directions divided by the sum of these moments, with 0 representing a circle and 1 an elongated ellipse. The means of these properties were calculated for each size class and each sample.
Observations of the soil thin-sections were also performed using a Nikon Eclipse E400 POL microscope under plane-polarised light and cross-polarised light. The microscope observations focused on small features, such as textural pores, enchytraeid droppings and separate plant tissues that were barely or not visible in the scans. The micromorphological descriptions of the thin sections were based on nomenclature adopted from Stoops [34].
2.5. Statistical Analyses
Using one-way analysis of variance (ANOVA), the differences in individual parameters between the soil properties were studied. In order to estimate the least significant differences between the mean values of homogeneous groups, we applied the post hoc test by means of the Bonferroni correction (at p 35] and Canoco 5 [36] software.
