Multi-mechanism synergistic remediation of phosphate-solubilizing bacteria under tetracycline and lead co-contamination stress: phosphate precipitation and organic acid complexation

Anjie Zhou¹Jiangyan Wu²Qi Tang²Huihui Chen³Yuan Liu²Haoming Chen^2*

• ¹Southeast University, Nanjing, China
• ²Nanjing University of Science and Technology, Nanjing, China
• ³Chinese Academy of Sciences, Beijing, China

Tetracycline (TC) and lead (Pb) contamination have become globally urgent environmental challenges. They are widely distributed in soil, water bodies, and other environments, posing severe threats to ecosystems and human health. Microbial remediation, as a cost-effective and environmentally friendly pollution control approach, has garnered increasing attention in recent years. This study isolated and screened a highly efficient phosphate-solubilizing bacteria strain CZ-M3 (Microbacterium sp.) from a chemical factory contaminated environment. The strain achieved a phosphate solubilization capacity of 125.46 mg/L in PVK medium, significantly outperforming the control strain CZ-B5 (69.1% increase). Stress tolerance experiments demonstrated that strain CZ-M3 maintained robust activity under TC (≤ 200 mg/L) and Pb2+ (≤1000 mg/L) stress, achieving 72-hour removal rates of 57.36% for 200 mg/L TC and 28.5% for 1000 mg/L Pb²⁺, thus selected as the core functional strain for remediating TC-Pb co-contamination. Notably, under co-contamination conditions, TC stress was found to stimulate Pb immobilization by CZ-M3. First, the strain releases PO₄³⁻ by secreting various organic acids, forming stable Pb-phosphate precipitates [e.g., Pb₅ (PO₄)₃Cl] with Pb²⁺, which was confirmed by X-ray diffraction (XRD) and attenuated total reflectance infrared spectroscopy (ATR-IR) results. Second, three-dimensional excitation-emission matrix (3D-EEM) spectroscopy revealed that combined stress induced enhanced secretion of extracellular polymeric substances (EPS) (humic acid-like substances), whose abundant functional groups (e.g., carboxyl and hydroxyl groups) effectively complexed and adsorbed contaminants. Most critically, organic acid secretion profiling found that high-concentration TC stress inhibited the tricarboxylic acid (TCA) cycle, causing a significant increase in tartaric acid content (significantly higher than the control group). Tartaric acid formed stable complexes with Pb²⁺ via its functional groups and promoted phosphate precipitation. This study provides a solid theoretical basis and a valuable microbial resources for the bioremediation of heavy metal-antibiotic co-contaminated environments.