| 其他摘要 | Yutangba, in Enshi City, Hubei Province, is one of the typical high-Se areas in China，and it has given rise to lots of environmental problems, due to selenium enrichment in local biotope. Using Se-laden carbonaceous mudstones as studying subjects, and three Se(IV)-reducing strains, Bacillus licheniformis SeRB-1, Bacillus sp. SeRB-2, and Arthrobacter sp. SeRB-3, screened form the mudstones, as researching tools, we preliminary studied the high Se(IV)-resistant cabalicity of strains, kinetics of selenite bioreduction and biomineralization under ion stress. Based on the above studies, major conclusions are drawn as follows. (1) The carbonaceous mudstones are super high-Se samples, within selenium highly enriched, and the average concentration is as high as 3000 mg/kg, with the highest value to 21830 mg/kg. The enrichment of bioactive elements, V, Mo, Cr, Zn, Cu and so on, tells that there are biological activity enrolling in the formation of the mudstones. While, the rich organic matter and nitrogen composition, and the unconsolidated structures create survival conditions for bacteria. (2) It shows that the doubling time got longer, growth rate turned smaller, and percentages of growth inhibition became larger, with the increasing of Se(IV) concentration, from the Se(IV)-resistant experiments. It also tells that the Se(IV)-reducing tolerance concentration is as high as 800 mM, and the median lethal concentration is high to 100～ 150 mM, indicating the strains belonging to high Se(IV)-reducing tolerance bacteria. Of which, strain SeRB-1 is better, with faster growing rate and short lag phase. (3) It illustrates that selenite reduction is mainly concentrated in exponential phase and earlier stationary phase. The reduction is generally divided into two stages. Stage I: From inoculation to earlier logarithmic growth phase, in this phase, cells grew slowly and the reducing ratio was small. Stage II: From the end of earlier logarithmic growth phase to time over, in later logarithmic phase, cells grew most quickly and the reducing ratio was largest,especially from 12～24 h. After that, in the stationary phase, reducing ratio turned smaller again. Finally, the average Se(IV)-reducing kinetics of the three strains at the concentrations from 1～10 mM were obtained, through average reducing efficiency calculating at each concentration and combining Michaelis-Menten equation. The kinetic equations of strains SeRB-1, SeRB-2 and SeRB-3 are Ci/V=76.14+13.72Ci, Ci/V=61.49+13.21Ci, and Ci/V=58.96+13.51Ci, respectively. According to which, Michaelis constants Km of enzyme reaction and maximum growth rate Vmax were calculated. The Km values are 5.55 mM, 4.22 mM and 4.65 mM and Vmax values are 0.073 μM/(L?h), 0.076 μ M/(L?h) and 0.074 μ M/(L?h), for SeRB-1, SeRB-2 and SeRB-3, respectively. Of which, strain SeRB-2 has a larger Vmax and longer reducing time. (4) The Se(IV)-reducing efficiency is dreasing with selenite concentrations increasing from 1 mM to 10 mM, and the maxnium reducing efficiency (or RE) was reached at Se(IV)=1 mM. Of which, the RE value of strain SeRB-2 is high to 90%, with 70% and 60% followed for SeRB-1 and SeRB-3, respectively. It indicates these bacteria have the application potential in bioremediation of selenium-contaminated sites, and strain SeRB-2 is more advantage. More Satisfied results would be obtained when selenite concentration is no more than 1mM. (5) There are also biominerals formation, during selenite bioreduction under Cu+2 and Fe+2 stress, but the rate is much slower than selenite bioreduction. Transmission electron microscopy revealed the process of biomineral formation at different periods. In earlier period, small granules(10～20 nm) were present mainly on the membranes, a few in the cytoplasm, and they are regular arranged around the cell envelopes. In the middle period, cells lysis appeared, and partial cell walls were completely decomposed, only leaving larger mineral particles exposed outside. Cytoplasm appeared vacuoles, and larger particles filled the vacancies. These particles, about 100～20 |
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