摘要
高氯酸盐(ClO4-)污染修复是地下水污染修复系统研究的热点之一。本文以室内研究为主，通过添加不同电子供体实现ClO4-的降解，并引入氮素混合污染，将ClO4-降解过程中关键功能酶编码基因pcrA、 cld，脱氮过程中功能酶编码基因NirS及表征整个细菌群落的16SrRNA基因的表达量联立，与污染物浓度变化之间进行相关性耦合分析，从基因水平阐释ClO4-降解过程中生物作用的详细机理，研究内容及结果包括：
（1）不同电子供体存在条件下的ClO4-降解研究结果表明，将10mg/LClO4-降至检出限以下，添加醋酸盐作为电子供体的AD体系降解速率(100小时)>未添加电子供体的ND体系(240小时)>添加氢气作为电子供体的HD体系(71天)。
（2）在pH值和DO等外界条件适合ClO4-降解的条件下，pcrA基因代表的高氯酸盐还原酶的活性是制约ClO4-降解的关键性因素。ND体系中16SrRNA基因与污染物浓度变化相关性最高(p=0.979)；AD体系中为cld基因(p=0.157)；HD体系中为pcrA与cld之和(p=0.732)。功能基因相关性分析结果表明不同电子供体存在条件下，ClO4-的降解与反应体系内不同生物群落相关。
（3）若ClO4-和NO3-N同时存在，在外源添加足够的醋酸盐作为电子供体条件下， NO3-N与ClO4-浓度比为1：1和5：1的范围内，硝酸盐的存在不会完全抑制ClO4-的降解，当NO3-N降解完全时，可以促进ClO4-的降解。
（4）当反应体系内存在氨氮时，氨氮可以作为氮源促进ClO4-的降解。若反应体系中仅有一种形式的氮素污染存在，即使提高初始氮浓度也不会对ClO4-的降解产生明显抑制作用，但若两种形式的氮素同时存在时则可以加速ClO4-降解的进行。在有氮存在的混合降解体系内，pcrA和cld基因与ClO4-的浓度变化之间的相关性不是很高，证明该功能基因对复杂环境中特定生物群落的表征有一定局限性。
（5）对添加不同电子供体降解体系前后菌种多样性变化分析可知ClO4-降解特征菌如Azospira和Dechloromonas等的存在及丰度的多少对ClO4-的降解具有决定性的作用，并可以作为表征环境中ClO4-降解可能性的依据之一。
Perchlorate contamination in the environment has been considered as a hotspot issuein the recent years. Included in this laboratory research were completely reducingperchlorate throμgh adding different electron donors, and introducing nitrogen as co-contaminant. The correlation between the change of perchlorate concentration andfunctional genes which encoded key enzyme during perchlorate reduction as pcrA andcld, denitrification process as NirS, or total bacteria as16SrRNA were assessed at thelevel of genes to further interpretation the mechanisms of perchloratebiodegradation.The main contents and results of this study were as follows:
(1) The efficiency of degradation of10mg/L perchlorate in three systems withdifferent electron donors were shown as: the rate of perchlorate reduction with acetateas electron donor (AD)(completely degradation in100hours) was more rapid thanperchlorate reduction without any electron donor (ND)(completely degradation in240hours) and ND was faster than with hydrogen as electron donor (HD)(completelydegradation in71days).
(2)Under the conditions of chemical indexes such as pH and DO which are suitablefor perchlorate bioremediation, the key factor that controlled the perchlorate reductionwas the expression of pcrA gene which encoded the α-subunit of perchlorate reductase.Pearson correlation index results showed that the expression of16SrRNA gene wasthe most apparently related to the change of ClO4-concentration in ND system(p=0.979); whereas cld to ClO4-in the AD system (p=0.157) and pcrA plus cld gene toClO4-in HD systems (p=0.732). It proved that ClO4-degradation under differentelectron donors were regulated by different microbial community in the reductionsystems, respectively.
(3)When NO3-N was present as co-contaminant with ClO4-, under the condition thatadding sufficient acetate as electron donor, NO3-did not completely inhibit the ClO4-reduction in the concentration ratio of NO3-N to ClO4-were1and5. NO3-promotedClO4-reduction once the NO3-was completely removed.
(4)Ammonium can be used as an extra nitrogen source for promoting the perchlorate reduction bacteria growth in the medium and accelerate the ClO4-bioremediation.When there existed only one kind of nitrogen contaminant in thesystem, ClO4-reduction did not be inhibited by the single nitrogen contaminant evenrisen the initial concentration of nitrogen. However, simultaneously existence of twokinds of nitrogen co-contaminants obviously improved the degradation efficiency ofClO4-.The correlation between the functional gene pcrA、cld and the concentrationof ClO4-were not so apparently, due to the limitations of using functional gene asmaker to represent specific microbial community in the mixed contamination systems.
(5) The phylogenic analysis of the changes in microbial communities indicated thatthe typical perchlorate reduction bacteria such as Azospira and Dechloromonas wereprevalently existent in ND, HD and AD respectively. The composition and theabundance of specific microbial community played an important role in perchloratebioremediation, and can be used as a symbol of perchlorate reduction in environment.
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