| Nanosilver has become one of the most commonly used nanomaterials in consumer products due to its broad spectrum of effective antibacterial properties.The widespread production,transportation and use of nanosilver lead to its inevitable entry into the soil environment,which had a negative impact on microorganisms.The biological nitrogen fixation in soil was an important nitrogen input process in natural ecosystems,but how the nitrogen-fixing microflora associated with this process responded to the stimulation of nanosilver was currently unknown,so it is particularly important to known about the changes of soil nitrogen-fixing bacteria and biological nitrogen fixation under nanosilver exposure,which not only can further determine the changes of nitrogen-fixing microbial community structure,but also provide a theoretical basis for the evaluation of ecotoxicity of nanosilver.The effects of the highest dose of nanosilver on the soil nitrogen-fixing bacteria community structure were studied.The types of nitrogen-fixing bacteria,the a diversity,the community composition and abundance of nitrogen-fixing bacteria,and the structure of nitrogen-fixing bacteria were investigated.The results showed that the species of nitrogen-fixing bacteria decreased under the exposure of nanosilver,and its diversity and abundance decreased,and the community composition changed significantly.The short-term exposure of nAg10(7 d)had the greatest impact on the structure of nitrogen-fixing bacteria.The long-term exposure of nAg10(90 d)and short-term exposure of nAg50(7 d)had little effect on the structure of nitrogen-fixing bacteria community,but the change of nitrogen-fixing bacteria community structure was the smallest when nAg50 was exposed for a long time(90d);the effects of nanosilver on soil nitrogen-fixing bacteria were related to its size and dosage,as well as exposure time,the long-term retention of nanosilver in the soil affects its toxic effects.The toxic effects of two different particle sizes of nanosilver on soil nitrogen-fixing model strain Azotobacter vinelandii were investigated.The cell activity,apoptosis,cell morphology,nitrogenase activity,gene expression,and oxidative stress and Ag uptake of Azotobacter vinelandii cells were investigated by nAg10 and nAg50.The results showed that the exposure of nanosilver inhibited the activity of A.vinelandii cells,accelerated the apoptosis process of A.vinelandii cells,and increased the mortality rate.After 12 hours of exposure to nanosilver,the structure of A.vinelandii cells was severely damaged,and the nitrogenase in the body was destroyed.The activity was significantly reduced,the expression of nitrogen-fixing gene nifH was down-regulated,and reactive oxygen species(ROS)were detected in the cells.ESR further proved that nanosilver appeared to produce ·OH.In addition,Ag was detected in the cell membrane and cytoplasm of A.vinelandii after the nanosilver exposure.The destruction of cell membrane by nanosilver and the generation of oxidative stress are the main reasons for the toxic effect of nanosilver.The toxicity of nAg10 was stronger than that of nAg50.The effects of nanosilver on soil nitrogen fixation were investigated by the changes of soil pH,autotrophic nitrogen-fixing bacteria and nitrogenase activity under the exposure of two different particle sizes of nanosilver.The results showed that the soil pH did not change under the exposure of nanosilver,and remained at about 6.54:the number of soil nitrogen-fixing bacteria and soil nitrogenase activity were affected to different extents after exposure to nanosilver.The short-term exposure of nanosilver(7d?28d)had a great influence on the numbers of soil nitrogen-fixing bacteria,and the effect of long-term exposure(60d?90d)was less.The toxicity of nanosilver showed dose and size effect.The effect of nanosilver on soil nitrogen fixation was extended with exposure time and the doses of nanosilver,this effect was less related to the size of nanosilver;the inhibition of nanosilver was related to its dose and exposure time. |
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