Diversity and functional attributes of microorganisms from stockpiled soils of coal mines in Mpumalanga Province, South Africa

Abstract

Coal mining in South Africa is renowned for large-scale removal of topsoil and subsoil through opencast mining. Such processes lead to an enormous amount of land degradation, and thus limit the land when mining operation ceases. The removal and stockpiling of topsoil lead to adverse effects not only on the physicochemical properties of the soil but also to the microbiological properties of the soil which greatly limit the ability of the soil to sustain plant development. Microbial properties are useful indicators of soil quality and could possibly serve as assessment criteria for successful rehabilitation of ecologically disturbed areas. The purpose of this study was to investigate the impact of soil stockpiling activities on diversity and the functional properties of microorganisms from opencast coal mines in Mpumalanga Province, South African. Samples were randomly collected from the stockpile soils of three opencast coal mines and adjacent unmined land (control) was sampled at depths of 0-20 cm ("topsoil") and >20 cm ("subsoil") in summer, winter and spring seasons. Physicochemical properties, β-glucosidase and urease activities in soils were determined using standard methods, while bacterial (16S rRNA gene) and fungal (Internal transcribed spacer 2 region) diversity were determined using culture-based methods and Polymerase Chain Reaction-denaturing gradient gel electrophoresis (PCR-DGGE). The pure bacterial and fungal isolates obtained from culture-based methods were further evaluated for their soil fertility attributes potentials to play roles in soil nutrient cycling as well as in plant growth enhancement such as phosphate solubilisation, atmospheric nitrogen fixation, and indoleacetic acid (IAA) production. The pattern of differences in the physicochemical properties of soils between unmined and stockpiled soils was not drastic across the three seasons, neither did the nutrient composition and soil physical properties clearly suggest that stockpiled soils were in poorer physicochemical condition compared to soil samples from control sites. The β-glucosidase and urease activities in stockpiled soils were mostly higher (p<0.05) than in unmined soils, and varied significantly (P<0.05) between seasons in some sites. PCR-DGGE patterns and Shannon-Wiener indices obtained revealed higher microbial diversity in unmined soils than in stockpiles soils across all seasons. Taxonomic analyses of sequences obtained (both PCR-DGGE bands and pure isolates) revealed that phyla Firmicutes (bacteria) and Ascomycota (fungi) were dominant. PCR-DGGE further revealed that Phialocephala humicola, Mortierella sp and Phoma sp were unique to Mine C. Several potential plant growth promoting microorganisms were obtained. Most of the isolates from both control and stockpiled soils had the potential to fix atmospheric nitrogen. None of the bacterial isolates from stockpiled soils produced IAA. The bacterial isolates from control soils were more efficient in phosphate solubilisation than those obtained in stockpiled soils. In general, the fungal isolates obtained from both control and stockpiled soils were more efficient in phosphate solubilisation and IAA production than bacterial isolates. The results suggest that microbial diversity, bacterial IAA production and phosphate solubilisation, and enzyme activities in soil stockpiles are affected by stockpiling operations. This may have negative implications for nutrient cycling and soil health during post-mining rehabilitation.