Toxic copper compounds are one of the most spread and ecologically hazardous contaminants of the environment. They are released into the environment in the places of its deposits and as a result of human industrial and household activities. Environmental pollution with copper compounds has a detrimental effect on ecosystems. The most powerful sources of copper pollution are mining sites, as well as wastewater of industrial enterprises. Nowadays, the development of the effective methods of ecosystems purification is relevant for the preservation of the environment. The development of microbial methods of toxic copper detoxification is promising. These methods require the isolation and investigation of microorganisms that are resistant to copper in high concentrations and are able to interact with it (accumulate, reduce, precipitate, etc.). Screening ecosystems for the presence of such microorganisms and understanding their molecular and physiological mechanisms of resistance and detoxification of hazardous copper compounds are of priority importance for environmental protection. These knowledges are necessary for the development of environmental protection biotechnologies for the purification of copper-containing wastewater and copper-contaminated soils.
The position of the thermodynamic prognosis about the possibility of growth and interaction of microorganisms with soluble copper(II) compounds at extremely high, one-molar (63546 mg/L) concentrations was confirmed experimentally for the first time. The natural ecosystems of five geographical regions of the globe (Ukraine, Antarctica, the Arctic, Israel, and South America) were screened for the presence of copper-resistant microorganisms. Their wide distribution was shown and the quantitative parameters of resistance to toxic copper compounds were determined. It was determined that there are microorganisms in the studied samples of soil, clay and sand, resistant to copper in extremely high concentrations (1000 – 15500 mg/L) in a significant amount (from n×102 to n×104 CFU/g) during cultivation in the agar nutrient medium. Nine new copper-resistant bacterial strains (Pseudomonas lactis UKR1, P. panacis UKR 2, P. veronii UKR 3 and UKR4, Staphylococcus succinus Cop98, Pantoea agglomerans Cop101, Bacillus mycoides Cop102, B. megaterium Cop99, B. velezensis Cop41) and 1 yeast strain (Rhodotorula mucilaginosa UKR5) were isolated capable to grow in the presence of 1 M or 63,546 mg/L Cu2+ in a liquid nutrient medium.
For the first time, the genomes of the four strains of Pseudomonas lactis UKR1, P. panacis UKR 2, P. veronii UKR 3 and UKR4 resistant to 1 mol/L Cu2+ were sequenced. The ability of Pseudomonas lactis strain UKR1 to interact with copper compounds via all thermodynamically acceptable pathways (immobilization (accumulation in cells, reduction precipitation to insoluble compounds and precipitation without valency changing) as well mobilization) was shown. Genomic sequences of highly resistant strains of Pseudomonas lactis UKR1, Pseudomonas panacis UKR2, Pseudomonas veronii UKR3 and Pseudomonas veronii UKR4 have been deposited in the DDBJ/ENA/GenBank database with bioproject accession number PRJNA565195 and genome accession numbers – VWXW00000000, VWXV00000000, VWXU00000000, VWXT00000000, respectively. Initial screening of the four genomes for genes encoding copper resistance mechanisms was shown the presence A, B, D proteins, copper-exporting ATPase copA3, copper chaperone copZ, as well as the two-component regulatory system cusRS. The ability of the unadapted to Cu2+ strict anaerobic hydrogen-synthesizing strain Clostridium butyricum 92 to adapt and immobilize Cu2+ with the efficiency of 88,0–99.2% in the concentration range of 50–200 mg/L Cu2+ has been proven. However, the efficiency of Cu2+ immobilization by the highly resistant to copper P. lactis strain UKR1 was 18.0–76.8% at the initial concentration of 200 mg/L Cu2+. For the first time, the possibility of regulation of types of the microorganisms interaction with copper compounds was established on the example of P. lactis strain UKR1. The high effectiveness of the copper immobilization via reduction to insolvable Cu2O↓ as well as precipitation of Cu2+ without valence changing by unadapted diversified hydrogen-synthesizing and methanogenic microbial communities has been proven. The high efficiency of the toxic metals ions accumulation by plants due to their stereochemical analogy with macroelements has been theoretically substantiated and experimentally confirmed. The effectiveness of using tobacco Nicotiana tabacum L. variety Djubek for the removal of specified metals from contaminated soils at a high initial concentration of each metal – 500 mg/kg soil has been proven in the vegetation conditions. The copper-resistant and unadapted microorganisms can be used for the development of industrial biotechnologies.
