Crocetti G R, Hugenholtz P, Bond P L, Schuler A, Keller J, Jenkins D, Blackall L L
Department of Microbiology and Parasitology, Advanced Wastewater Management Centre, The University of Queensland, St. Lucia, 4072 Queensland, Australia.
Appl Environ Microbiol. 2000 Mar;66(3):1175-82. doi: 10.1128/AEM.66.3.1175-1182.2000.
Laboratory-scale sequencing batch reactors (SBRs) as models for activated sludge processes were used to study enhanced biological phosphorus removal (EBPR) from wastewater. Enrichment for polyphosphate-accumulating organisms (PAOs) was achieved essentially by increasing the phosphorus concentration in the influent to the SBRs. Fluorescence in situ hybridization (FISH) using domain-, division-, and subdivision-level probes was used to assess the proportions of microorganisms in the sludges. The A sludge, a high-performance P-removing sludge containing 15.1% P in the biomass, was comprised of large clusters of polyphosphate-containing coccobacilli. By FISH, >80% of the A sludge bacteria were beta-2 Proteobacteria arranged in clusters of coccobacilli, strongly suggesting that this group contains a PAO responsible for EBPR. The second dominant group in the A sludge was the Actinobacteria. Clone libraries of PCR-amplified bacterial 16S rRNA genes from three high-performance P-removing sludges were prepared, and clones belonging to the beta-2 Proteobacteria were fully sequenced. A distinctive group of clones (sharing >/=98% sequence identity) related to Rhodocyclus spp. (94 to 97% identity) and Propionibacter pelophilus (95 to 96% identity) was identified as the most likely candidate PAOs. Three probes specific for the highly related candidate PAO group were designed from the sequence data. All three probes specifically bound to the morphologically distinctive clusters of PAOs in the A sludge, exactly coinciding with the beta-2 Proteobacteria probe. Sequential FISH and polyphosphate staining of EBPR sludges clearly demonstrated that PAO probe-binding cells contained polyphosphate. Subsequent PAO probe analyses of a number of sludges with various P removal capacities indicated a strong positive correlation between P removal from the wastewater as determined by sludge P content and number of PAO probe-binding cells. We conclude therefore that an important group of PAOs in EBPR sludges are bacteria closely related to Rhodocyclus and Propionibacter.
实验室规模的序批式反应器（SBR）作为活性污泥法的模型，用于研究废水中强化生物除磷（EBPR）。通过提高进入SBR的进水磷浓度，基本实现了聚磷菌（PAO）的富集。使用域、纲和亚纲水平的探针进行荧光原位杂交（FISH），以评估污泥中微生物的比例。A污泥是一种高效除磷污泥，生物量中含磷量为15.1%，由大量含聚磷酸盐的球杆菌簇组成。通过FISH分析，A污泥中超过80%的细菌是排列成球杆菌簇的β-2变形菌，强烈表明该菌群包含负责EBPR的PAO。A污泥中的第二优势菌群是放线菌。制备了来自三种高效除磷污泥的PCR扩增细菌16S rRNA基因的克隆文库，并对属于β-2变形菌的克隆进行了全序列测定。一组与红环菌属（序列同一性为94%至97%）和嗜丙酸栖菌（序列同一性为95%至96%）相关的独特克隆群（序列同一性≥98%）被确定为最有可能的候选PAO。根据序列数据设计了三种针对高度相关候选PAO菌群的特异性探针。所有这三种探针都能特异性地与A污泥中形态独特的PAO簇结合，与β-2变形菌探针完全吻合。EBPR污泥的连续FISH分析和聚磷酸盐染色清楚地表明，与PAO探针结合的细胞含有聚磷酸盐。随后对一些具有不同除磷能力的污泥进行PAO探针分析，结果表明，根据污泥磷含量测定的废水中磷的去除量与PAO探针结合细胞的数量之间存在很强的正相关。因此，我们得出结论，EBPR污泥中一组重要的PAO是与红环菌和丙酸杆菌密切相关的细菌。
