Phosphate-solubilizing bacteria (PSB) can alleviate available phosphorus deficiency without causing environmental pollution, unlike chemical phosphate fertilizers. However, the phosphate solubilization mechanisms of PSB are still unclear. Transcriptome sequencing was used to analyze the expression patterns of differential expressed genes (DEGs) of the phosphate-solubilizing bacterium W134 under the conditions of soluble phosphorus (group A), insoluble phosphorus (group B), and lacking phosphorus (group C). Nine DEGs in three different groups were detected by quantitative real-time polymerase chain reaction (qRT-PCR). Then, high performance liquid chromatography (HPLC) was applied to detect the concentrations and composition of organic acids. Compared with group A, Gene Ontology (GO) annotation showed that the cluster of W134 DEGs in groups B and C were basically the same. Besides, the results of enrichment Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway indicated that genes in the Citrate cycle (TCA cycle) pathway closely related to organic acid production were significantly upregulated. The qRT-PCR results were almost consistent with the expression trends of the transcriptome data. The HPLC results showed that the formic acid, ascorbic acid, acetic acid, citric acid, and succinic acid concentrations were significantly increased in group B and C (p < 0.05), while the contents of lactic acid and malic acid were significantly increased in group B (p < 0.05). The above results provided further validation that the upregulated genes should be related to W134 secretion of organic acids. Our study revealed several potential candidate genes and tried to explain phosphate solubilization mechanisms. This provides a new insight for calcareous reclaimed soil, and it will reduce the need of chemical phosphate fertilizers to promote environmentally friendly agriculture.

Coal mining has caused significant soil nitrogen loss in mining areas, limiting reclamation and reuse in agriculture. This article studies the effects of organic fertilizer, inorganic fertilizer, and the combined application of Pseudomonas fluorescens with the ability of nitrogen fixation on soil nitrogen accumulation and composition in the reclamation area of the Tunlan Coal Mine from 2016 to 2022 under the conditions of equal nitrogen application, providing a scientific basis for microbial fertilization and the rapid increase in nitrogen content in the reclaimed soil of mining areas. The results showed that as the reclamation time increased, the nitrogen content and the composition and structure of the soil treated with fertilization rapidly evolved toward normal farmland soil. The soil nitrogen content increased most rapidly in the presence of added P. fluorescens + organic fertilizer (MB). Compared to other treatments (inorganic fertilizer (CF), organic fertilizer (M), and P. fluorescens + inorganic fertilizer (CFB)), MB increased total nitrogen (TN) to normal farmland soil levels 1–3 years earlier. The comprehensive scores of MB and CFB on the two principal components increased by 1.58 and 0.79 compared to those of M and CF treatments, respectively. This indicates that the combination of P. fluorescens and organic fertilizer improves soil nitrogen accumulation more effectively than the combination of P. fluorescens and inorganic fertilizer. In addition, the application of P. fluorescens increases the content of unknown nitrogen (UN) in acid-hydrolysable nitrogen (AHN) and decreases the content of amino acid nitrogen (AAN) and ammonia nitrogen (AN). However, there was no significant effect on the content of ammonium nitrogen (NH₄⁺-N) and nitrate nitrogen (NO₃⁻-N) in soil-mineralized nitrogen (SMN). When combined with inorganic fertilizer, the contribution of SMN to TN increased by 14.78%, while when combined with organic fertilizer, the contribution of AHN to TN increased by 44.77%. In summary, the use of P. fluorescens is beneficial for nitrogen recovery in the reclaimed soil of coal-mining areas. The optimal fertilization method under the experimental conditions is the combination of P. fluorescens and organic fertilizer.

The initial variations in soil bacteria at the very beginning of reclamation still remains unclear. This study investigates the impact on bacterial communities of eight different treatments, including uncultivated land, unfertilized cultivation, chemical fertilizer, chemical fertilizer + bacterial fertilizer, manure, manure + bacterial fertilizer, manure + chemical fertilizer, and manure + chemical fertilizer + bacterial fertilizer, during the short-term reclamation of coal-mining soils. The results showed that total nitrogen, available phosphorus, soil organic carbon, microbial biomass carbon, and alkaline phosphatase activity were significantly increased in all fertilization treatments compared to uncultivated land (p < 0.05). All fertilization treatments other than chemical fertilizer harbored significantly higher activities of urease, catalase, and invertase than unfertilized cultivation (p < 0.05). The bacterial communities structures in manure-amended treatments significantly differed in uncultivated land and unfertilized cultivation and were phylotypically shifted from oligotrophic to Actinobacteria-dominant copiotrophic traits, accompanied with phenotypic succession of the enriching characteristics of Gram-positive, biofilms formation, and stress tolerance. The co-occurrence network in manure-amended treatments harbored a simple co-occurrence network, indicating more productive soils than in no-manure treatments. Manure amendment, total nitrogen, microbial biomass carbon, invertase, catalase, and soil moisture were the key driving factors. Our study underscores the bacterial initialization characteristics promoted by manure at the very beginning of coal-mining reclamation.

Fertilization is an effective measure to rapidly improve soil quality in reclaimed mining areas. However, the combined effects of fertilization regimes and reclamation age on phosphorus (P) fraction transformation and the pqqC-harboring microbial community in reclaimed soils remain unclear. In this study, we investigated the dynamics of inorganic P fractions and the pqqC-harboring bacterial community under different fertilization treatments (no fertilizer: CK; chemical fertilizer: CF; organic manure: M) and reclamation ages (1, 5, and 10 years) in a coal mining reclamation area of Shanxi Province, using long-term field experiments combined with high-throughput sequencing. Results showed that compared with the CF and CK treatments, the M treatment significantly increased soil organic matter (SOM), available P (AP), and total nitrogen (TN) content, and promoted the conversion of moderately labile P (NaOH-Pi) to labile P fractions (H₂O-Pi, NaHCO₃-Pi). Meanwhile, the pqqC gene abundance increased with reclamation age, with the M treatment maintaining the highest levels in all fertilization regimes. Co-occurrence network analysis of core species revealed that the number of connections gradually decreased and the network structure simplified with increasing reclamation age. Correspondingly, the microbial community transitioned from an initial stage characterized by rapid response and intense competition to a stable phase. Specifically, Pseudomonas spp. played a key role in P mobilization. Structural equation modeling (SEM) further demonstrated that reclamation age directly promoted the pqqC gene abundance and AP content, whereas fertilization indirectly influenced P transformation by regulating microbial diversity. Our findings reveal that reclamation age and fertilization synergistically shape the inorganic P profile and the associated bacterial community, providing insights for developing targeted P management strategies in reclaimed lands.

Coal mining has caused a significant loss of nitrogen content of soil in mining areas. This article studies the effects of organic fertilizer, inorganic fertilizer, and combined application of Pseudomonas fluorescens with the ability of nitrogen fixation on soil nitrogen accumulation and composition in the reclamation area of Tunlan Coal Mine from 2016 to 2022 under the condition of equal nitrogen application, providing scientific basis for microbial fertilization and rapid increase of nitrogen content in the reclaimed soil of mining areas. The results showed that as the reclamation time increased, the nitrogen content and its composition structure of the soil treated with fertilization rapidly evolved towards normal farmland soil. The soil nitrogen content increased the fastest in the treatment of Pseudomonas fluorescens + organic fertilizer (MB), compared with inorganic fertilizer (CF), organic fertilizer (M), and Pseudomonas fluorescens + inorganic fertilizer (CFB) treatments, the treatment of Pseudomonas fluorescens + organic fertilizer (MB) can bring the total nitrogen (TN) content of the soil to normal farmland soil levels 1-3 years earlier. The comprehensive scores of Pseudomonas fluorescens + organic fertilizer (MB) and Pseudomonas fluorescens + inorganic fertilizer (CFB) on the two principal components were increased 1.58 and 0.79 than those of organic fertilizer (M) and inorganic fertilizer (CF) treatments, respectively. This further indicates that the combination of Pseudomonas fluorescens and organic fertilizer has a better effect on improving soil nitrogen accumulation than the combination of Pseudomonas fluorescens and inorganic fertilizer. At the same time, the application of Pseudomonas fluorescens will increase the content of unknown nitrogen (UN) in acid-hydrolysable nitrogen (AHN), and decrease the content of amino acid nitrogen (AAN) and ammonia nitrogen (AN), relatively. However, there was no significant effect on the content of ammonium nitrogen (NH4+-N) and nitrate nitrogen (NO3--N) in soil inorganic nitrogen (SMN). When combined with inorganic fertilizer, the contribution of soil mineralized nitrogen (SMN) to total nitrogen (TN) increased by 14.78%, while when combined with organic fertilizer, the contribution of acid-hydrolysable nitrogen (AHN) to total nitrogen (TN) increased by 44.77%. In summary, the use of Pseudomonas fluorescens is beneficial for nitrogen recovery in the reclaimed soil of coal mining areas. The optimal fertilization method under the experimental conditions is the combination of Pseudomonas fluorescens and organic fertilizer.