Ecological Toxicology Studies of Herbicide Fomesafen on Soil Enzyme, Microorganism and Earthworm(Eisenia Fetida)
|School||Shandong Agricultural University|
|Keywords||Fomesafen Soil enzyme Soil microorganism Earthworm DGGE T-RFLP Antioxidant enzymes DNA damage|
Fomesafen is a diphenyl ether herbicide that is mainly used in soybean and peanut fields to control the broad-leaved weeds. The improper use, long-term use and large amounts use of fomesafen, long residual herbicide, caused not only carryover injury to sensitive crops but also potential damaged the soil ecosystem. To evaluate the safety of fomesafen on the soil environment, studies were conducted concerning its influences on soil enzymes and soil microorganisms by conventional physiological and biochemical methods and combining with the denaturing gradient gel electrophoresis (DGGE), terminal restriction fragment length polymorphism (T-RFLP) technologies. In addition, the oxidative stress and DNA damage in the earthworm Eisenia fetida induced by low doses of fomesafen were also investigated. The main results were summarized as follows:1. The fomesafen was determined in culture soil by high-performance liquid chromatography (HPLC), which showed that the dissipation kinetics of fomesafen in soil was described using a first order kinetics model, the half-time was38.9d. Followed that, the soil microbe quantity in the soil samples treated with different doses of fomesafen were measured by the plate counting method. Results showed that the number of bacterial and actinomyces in the10,100and500μg/kg fomesafen treated soil were higher than those in the control groups, but the fungi was slightly inhibited in the40days of incubation. With the incubation time prolonged, the number of bacteria, actinomyces and fungi were gradually returned the control levels at60d. In the whole, low doses (≤500μg/kg) of fomesafen has less effect on the soil microorganism quantity2. Effect of low doses of fomesafen on soil enzyme activities under the laboratory simulation conditions was investigated. Five enzymes, i.e., Acid phosphatase, alkaline phosphatase, urease, dehydrogenase and β-glucosidase, were selected. The results found that the activities of acid phosphatase, alkaline phosphatase and P-glucosidase were significantly stimulated by the fomesafen in the30days of incubation. The activity of soil urease presents the effect of inhibit-stimulate-return to a control level. In the60days of incubation, the dehydrogenase activities of soils treated with fomesafen were higher that those of the controls, and showed a statistical dose-dependent relationship. Because of the sensitive to fomesafen, the soil urease and dehydrogenase can be used as one of the indicator to evaluate its soil ecological effect.3. The potential influences of fomesafen on soil bacterial and fungal community in brown soil were assessed by16S rDNA-PCR-DGGE. During the60days of incubation, fomesafen could stimulate the bacteria populations, especially for Emticicia, Luteibacter, Bacillus and Pontibacter. These mean that low doses (≤500μg/kg) of fomesafen had no toxicity on soil bacteria. But the Jacarrd indices calculated from DGGE indicated that the similarity gradually reduced with the increased of fomesafen concentration in soil compared to the controls. These showed that fomesafen could change the soil bacteria community structure in some extent. Soil fungi community was also affected by low doses of fomesafen. In the20days of incubation, soil fungi were stimulated by minimum concentration (10μg/kg) of fomesafen, but the inhibited effect was found at the higher concentrations (100and500μg/kg). After30days of incubation, DGGE profiles showed that Trichosporon、Ascobolus and uncultured fungi were simulated under the stresses of fomesafen.4. T-RFLP was used to assess the impact on soil bacterial community by low doses of fomesafen in this study. The soil bacterial was significantly affected by fomesafen during10days of incubation. Some soil bacterial floras were inhibited, but this inhibition effect gradually disappeared after days of10. Another some soil bacterial floras were always stimulated by fomesafen during the whole cultural period (60d). It was found that these findings were similar to results obtained through DGGE and plate counting method. The biodiversity of soil bacterial community (Shannon index, H’) reduced during the30days of cultural period and then recovered to that in the control. The Simpson index was increased under the stress of fomesafen, which showed that fomesafen could stimulate the growth of some bacteria. The relative content of TRF138flora was always higher than that in the control. This showed that some strains could utilize fomesafen as the energy sources for growth. And this may be also providing the foundation for isolating fomesafen degrading bacterium.5. Oxidative stress and DNA damage in the earthworm Eisenia fetida induced by low doses (≤500μg/kg) of fomesafen were investigated in the OECD soil. Compared to the control, the SOD activity increased on the3rd and7th days but decreased on the14th day due to treatment with100and500μg/kg of fomesafen. The activities of CAT and POD increased significantly on the3rd,7th and14th days of exposure. In addition, the ROS level was significantly enhanced throughout the entire experimental period. When exposure period was prolonged21and28days, the activities of SOD, CAT, and POD returned to the control levels. Slight increase of ROS could not effect the changes of three enzymes activity on the exposure days of21and28. Compared to the controls, MDA contents had no obvious changes, in addition to the seventh day had significance increase. Based on the results of the present study, low doses of fomesafen (≤500μg/kg) could not lead to oxidative stress and peroxidation in E. fetida after exposure of21days. The slightly DNA damage in earthworm coelomocytes was observed after treatment with low doses fomesafen in14days of exposure. With the extension of exposure time, the DNA damage in earthworm coelomocytes gradually disappeared under the defense mechanism of earthworm.