| Along with the abundant production and use of antibiotics,its residues and ecological effects in environmental media have caused wide public concern.Although the trace levels of residual antibiotics in the environment is not enough to kill microorganisms,the long-term exposure in this circumstance may still affect for the function systems of microbial,adversely affecting its physiological and biochemical properties,even inducing the generation of resistance genes.In this research,the erythromycin with high detective rate was chosen as a representative of the antibiotics,and phenanthrene-degrading strain Sphingomonas sp GY2 B which was screened by our lab in the earlier stage with high efficiency of phenanthrene,was chosen for the research object.The study was conducted on the microscopic morphology,cell activity,key enzyme gene and protein expression differences of PAHs-degrading strain under antibiotics stress,providing the theoretical basis for correctly understanding and estimating the antibiotic environmental risk.Main findings are as follows:(1)The erythromycin showed toxicity to Sphingomonas sp GY2 B with concentration dependence.When erythromycin concentration is higher than 0.25 mg/L,the GY2 B growth and degradation of phenanthrene are inhibited.Stress effects of erythromycin showed adverse influence on cell morphology and internal structure.Moreover,the higher concentration of erythromycin was,the more serious of the cell morphological changed.Under the low concentrations of erythromycin stress,the cytoplasmic density decreased,but the complete structure of cells as well as the genetic material and inclusions can still guarantee the function of the strain,realizing to the degradation and removal of phenanthrene.However,high levels of erythromycin stress will make the degrading strain with almost no capacity.(2)Erythromycin stress effects would change the cytoactive of degrading bacteria GY2 B and presented the concentration of correlation.Flow cytometry analysis showed that when erythromycin concentration higher than 1 mg/L,the death cells increased and the activity of cells decreases,the cell integrity of GY2 B bacteria was destroyed.When erythromycin concentration higher than 25 mg/L,the cytoplasm membrane permeability would increased and membrane potential would decreased,then cells would occur the polarization phenomenon,thus promoting cell apoptosis.(3)Erythromycin stress could significantly change the degrading bacteria GY2 B protein expression.Erythromycin stress effects could make the proteins which related to degradation down-regulated,including polycyclic aromatic hydrocarbons dioxygenase ferredoxin reductase,2,3-catechol dioxygenase,polycyclic aromatic hydrocarbons dioxygenase small subunit,glutathione transferase,Lys R family transcription regulators,etc.,Among them,polycyclic aromatic hydrocarbons dioxygenase ferredoxin reductase had mainly influences on the aromatic ring open epoxidation,catechol 2,3-dioxygenase was associated with process of oxidizing catechol to carbon dioxide and water in phenanthrene degradation,LysR family transcription regulators could activate the target gene.With the increase of erythromycin stress concentrations,the differential protein types connected with degradation reduced,the amount of protein expressions cut.When reaching to 100 mg/L erythromycin,the differential protein productions associated with degradation even could not detected,the degrading bacteria does not exist proteins related to PAHs degradation.Erythromycin stress would change cell integrity,membrane permeability and functional protein expression of degradating microbial in the environment,thus affecting the physiological function of the microbial,but the influences presented the concentration of correlation obviously.Therefore,when evaluating the environmental risk of antibiotics,the concentration levels should be analyzed to avoid overstating their risks. |
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