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Microbial community analysis of enhanced biological phosphorus removal process and identification of polyphosphate accumulating organisms by molecular approach

强化生物除磷工艺的微生物群落分析及聚磷积累微生物的分子鉴定

基本信息

批准号：
14350283
负责人：
MINO Takashi
金额：
$ 9.6万
依托单位：
The University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2003
项目状态：
已结题

项目摘要

Fundamental research about enhanced biological phosphorus removal (EBPR) process is now making great progress because of the new microbial community analysis methods such as molecular techniques. This research was carried to reveal the relationship between microbial community and function of EBPR sludge using 16S-rDNA approach.In this research, at first, laboratory scale reactors were operated using acetate, glutamate, propionate, glucose or peptone as a main carbon source respectively. The expression of phosphorus removal activity was monitored by batch experiments and the microbial community change was monitored by Denaturing gradient gel electrophoresis (DGGE) using 16S-rDNA fragments. By comparing phosphorus removal activity and microbial community change, polyphosphate accumulating organisms were screened. In addition, by sequencing the whole length of 16S-rDNA of target bacteria by using cloning method and developing gene probes which can specifically detect the target bacteria and combining with the polyphosphate staining method and fluorescent in situ hybridization, capability of accumulating polyphosphate by screened bacteria was investigated.As a result, it was demonstratedd that gram-positive bacteria with high GC content accumulated polyphosphate in the reactor fed with mainly peptone and this gram positive bacteria was considered as the second putative polyphosphate accumulating organisms (PAOs) after Rhodocyclus related bacteria, which has been already considered as PAOs candidates. In addition, as another bacteria which accumulated polyphosphate but was not Rhodocyclus related bacteria nor gram positive bacteria was detected, it was clearly shown that still unknown PAOs should exist. In this way, it was shown that the EBPR is achieved by the combination of several PAOs

随着分子生物学技术等新的微生物群落分析方法的出现，强化生物除磷（EBPR）工艺的基础研究取得了很大进展。本研究以乙酸盐、谷氨酸盐、丙酸盐、葡萄糖和蛋白胨为主要碳源，采用16 S-rDNA技术研究了EBPR污泥微生物群落结构与污泥功能的关系。通过批式实验监测除磷活性的表达，利用变性梯度凝胶电泳（DGGE）技术检测微生物群落的变化。通过比较除磷活性和微生物群落变化，筛选聚磷菌。此外，通过克隆法对目的菌16 S-rDNA进行全序列测定，并研制出能特异性检测目的菌的基因探针，结合多聚磷酸盐染色法和荧光原位杂交法，对筛选出的细菌积累多聚磷酸盐的能力进行了研究，结果表明，结果表明，在以蛋白胨为主的反应器中，GC含量较高的革兰氏阳性菌对聚磷酸盐有一定的积累作用，该菌被认为是第二大聚磷酸盐积累菌。红环菌属（Rhodocyclus）相关细菌之后，其已经被认为是PAO候选物。此外，由于检测到另一种积累多磷酸盐的细菌，但不是红环菌属相关细菌，也不是革兰氏阳性细菌，因此清楚地表明应该存在仍然未知的PAO。通过这种方式，表明EBPR是通过几个PAO的组合来实现的